SCIENCE OF PAIN

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 SCIENCE OF PAIN

Editors
Dr Allan I. Basbaum
University of California, San Francisco, CA, USA

Dr M. Catherine Bushnell
McGill University, Montreal, Quebec, Canada

AMSTERDAM BOSTON HEIDELBERG LONDON NEW YORK OXFORD

PARIS SAN DIEGO SAN FRANCISCO SINGAPORE SYDNEY TOKYO
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PSYCHOPHYSICS OF PAIN

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Contents
Introduction ix

1 The Adequate Stimulus 1

R D Treede, Johannes Gutenberg-University, Mainz, Germany
2 Pain Theories 5
F Cervero, McGill University, Montreal, QC, Canada
3 Anatomy of Nociceptors 11
S Mense, Institut fur Anatomie und Zellbiologie, Universitat Heidelberg, Heidelberg, Germany
4 Molecular Biology of the Nociceptor/Transduction 43
M S Gold, University of Pittsburgh, Pittsburgh PA, USA
M J Caterina, Johns Hopkins School of Medicine, Baltimore, MD, USA
5 Zoster-Associated Pain and Nociceptors 75
H Maija, Helsinki University Hospital, Helsinki, Finland
6 Ectopic Generators 83
M Devor, Hebrew University of Jerusalem, Jerusalem, Israel
7 Sodium Channels 89
John N Wood, University College London, London, UK
8 Physiology of Nociceptors 97
M Ringkamp and R A Meyer, Johns Hopkins University, Baltimore, MD, USA
9 Itch 115
E Carstens, University of California, Davis, CA, USA
10 Thermal Sensation (Cold and Heat) through Thermosensitive TRP Channel Activation 127
Makoto Tominaga, National Institutes of Natural Sciences, Okazaki, Japan
11 The Development of Nociceptive Systems 133
G J Hathway and M F Fitzgerald, University College London, London, UK
12 Appropriate/Inappropriate Developed Pain Paths 147
Jens Schouenborg, Lund University, Lund, Sweden
13 Pain Control: A Child-Centered Approach 155
Patricia A McGrath, The University of Toronto, Toronto, ON, Canada
14 Assaying Pain-Related Genes: Preclinical and Clinical Correlates 165
V E Scott, R Davis-Taber, and P Honore, Global Pharmaceutical Research and
Development, Abbott Park, IL, USA
15 Evolutionary Aspects of Pain 175
E T Walters, University of Texas at Houston, Medical School, Houston, TX, USA

v
vi Contents

16 Redheads and Pain 185

J S Mogil, McGill University, Montreal, QC, Canada
17 Autonomic Nervous System and Pain 193
Wilfrid Janig, Physiologisches Institut, Christian-Albrechts-Universitat zu Kiel, Germany
18 Sympathetic Blocks for Pain 227
A Sharma, Columbia University, New York, NY, USA
J N Campbell and S N Raja, Johns Hopkins University, Baltimore, MD, USA
19 Sprouting in Dorsal Root Ganglia 237
E M McLachlan, Prince of Wales Medical Research Institute, Randwick, NSW, Australia
20 Vagal Afferent Neurons and Pain 245
W Janig, Christian-Albrechts-Universitat zu Kiel, Kiel, Germany
21 Sex, Gender, and Pain 253
R B Fillingim, University of Florida College of Dentistry, Community Dentistry and
Behavioral Science Gainesville, FL, USA
22 Neurotrophins and Pain 259
Lorne M Mendell, State University of New York, Stony Brook, NY, USA
23 Morphological and Neurochemical Organization of the Spinal Dorsal Horn 279
A Ribeiro-da-Silva, McGill University, Montreal, QC, Canada
Y De Koninck, Centre de recherche Universite Laval Robert-Giffard, Quebec, QC, Canada
24 Spinal Cord Physiology of Nociception 311
A R Light, University of Utah, Salt Lake City, UT, USA
S Lee, Korea Institute of Science and Technology, Seoul, Korea
25 What is a Wide-Dynamic-Range Cell? 331
D Le Bars, INSERM U-713, Paris, France
S W Cadden, University of Dundee, Dundee, UK
26 Spinal Cord Mechanisms of Hyperalgesia and Allodynia 339
T J Coderre, McGill University, Montreal, QC, Canada
27 Glycine Receptors 381
H U Zeilhofer, University of Zurich, Zurich, Switzerland
28 Pain Following Spinal Cord Injury 387
R P Yezierski, Comprehensive Center for Pain Research and The McKnight Brain Institute,
University of Florida, Gainesville, FL, USA
29 Long-Term Potentiation in Pain Pathways 401
J Sandkuhler, Medical University of Vienna, Vienna, Austria
30 Immune System, Pain and Analgesia 407
H L Rittner, H Machelska, and C Stein, Charite Universitatsmedizin Berlin,
Campus Benjamin Franklin, Berlin, Germany
31 Mechanisms of Glial Activation after Nerve Injury 429
L R Watkins, E D Milligan, and S F Maier, University of Colorado at Boulder, Boulder, CO, USA
32 Trigeminal Mechanisms of Nociception: Peripheral and Brainstem Organization 435
D A Bereiter, University of Minnesota, Minneapolis, MN, USA
K M Hargreaves, University of Texas Health Science Center, San Antonio, TX, USA
J W Hu, University of Toronto, Toronto, ON, Canada
33 Migraine A Disorder Involving Trigeminal Brainstem Mechanisms 461
P J Goadsby, University of California, San Francisco, CA, USA
34 Tooth Pain 469
M R Byers, University of Washington, Seattle WA, USA
 Contents vii

35 Ascending Pathways: Anatomy and Physiology 477

D Lima, Universidade do Porto, Porto, Portugal
36 Dorsal Columns and Visceral Pain 527
W D Willis Jr. and K N Westlund, University of Texas Medical Branch, Galveston, TX, USA
37 Visceral Pain 543
G F Gebhart and K Bielefeldt, University of Pittsburgh, Pittsburgh, PA, USA
38 Irritable Bowel Syndrome 571
S Bradesi and E A Mayer, University of California, Los Angeles, CA, USA
I Schwetz, Medical University, Graz, Austria
39 Pain in Childbirth 579
U Wesselmann, The Johns Hopkins University School of Medicine, Baltimore, MD, USA
40 Urothelium as a Pain Organ 585
L A Birder, University of Pittsburgh School of Medicine, Pittsburgh, PA, USA
41 The Brainstem and Nociceptive Modulation 593
M M Heinricher, Oregon Health & Science University, Portland, OR, USA
S L Ingram, Washington State University, Vancouver, WA, USA
42 Emotional and Behavioral Significance of the Pain Signal and the Role of the Midbrain
Periaqueductal Gray (PAG) 627
K Keay and R Bandler, University of Sydney, Sydney, NSW, Australia
43 The Thalamus and Nociceptive Processing 635
J O Dostrovsky, University of Toronto, Toronto, ON, Canada
A D Craig, Barrow Neurological Institute, Phoenix, AZ, USA
44 Psychophysics of Sensations Evoked by Stimulation of the Human Central Nervous System 655
S Ohara, C A Bagley, H C Lawson, and F A Lenz, Johns Hopkins Hospital, Baltimore, MD, USA
45 Nociceptive Processing in the Cerebral Cortex 669
R D Treede, Ruprecht-Karls-University Heidelberg, Heidelberg, Germany
A V Apkarian, Northwestern University, Chicago, IL, USA
46 Phantom Limb Pain 699
H Flor, Central Institute of Mental Health, Mannheim, Germany
47 Human Insular Recording and Stimulation 707
F Mauguiere, Lyon I University and INSERM U879, Bron, France
M Frot, INSERM U879, Bron France
J Isnard, Lyon I University and INSERM U879, Bron, France
48 The Rostral Agranular Insular Cortex 717
L Jasmin, Neurosurgery and Gene Therapeutics Research Institute, Los Angeles, CA, USA
P T Ohara, University of California, San Francisco, CA, USA
49 Descending Control Mechanisms 723
K Ren and R Dubner, University of Maryland, Baltimore, MD, USA
50 Diffuse Noxious Inhibitory Controls (DNIC) 763
D Le Bars, INSERM U-713, Paris, France
J C Willer, INSERM U-731, Paris, France
51 Fibromyalgia 775
R Staud, University of Florida, Gainesville, FL, USA
52 Pain Perception Nociception during Sleep 783
G J Lavigne, Universite de Montreal, Montreal, QC, Canada
K Okura, Tokushima Graduate School, Tokushima, Japan
M T Smith, John Hopkins Medical School, Baltimore, MD, USA
viii Contents

53 Pharmacological Modulation of Pain 795

A Dray, AstraZeneca Research and Development, Montreal, PQ, Canada
54 Forebrain Opiates 821
J-K Zubieta, University of Michigan, Ann Arbor, MI, USA
55 Neuropathic Pain: Basic Mechanisms (Animal) 833
M H Ossipov and F Porreca, University of Arizona, Tucson, AZ, USA
56 Animal Models and Neuropathic Pain 857
I Decosterd and T Berta, University of Lausanne, Lausanne, Switzerland
57 Neuropathic Pain: Clinical 865
R Baron, Christian-Albrechts-Universitat Kiel, Kiel, Germany
58 Neurogenic Inflammation in Complex Regional Pain Syndrome (CRPS) 901
F Birklein, University of Mainz, Mainz, Germany
M Schmelz, University of Heidelberg, Mannheim, Germany
59 Complex Regional Pain Syndromes 909
R Baron, Christian-Albrechts-Universitat Kiel, Kiel, Germany
60 Poststroke Pain 919
T S Jensen and N B Finnerup, Aarhus University Hospital, Aarhus, Denmark
61 Psychophysics of Pain 927
R H Gracely, University of Michigan Health System, VAMC, Ann Arbor, MI, USA
E Eliav, UMDNJ-New Jersey Dental School, Newark, NJ, USA
62 Consciousness and Pain 961
M Devor, Hebrew University of Jerusalem, Jerusalem, Israel
63 Assessing Pain in Animals 969
S W G Derbyshire, University of Birmingham, Birmingham, UK
64 Psychological Modulation of Pain 975
D D Price, A Hirsh, and M E Robinson, University of Florida, Gainesville, FL, USA
65 The Placebo Effect 1003
F Benedetti, University of Turin Medical School, Turin, Italy
66 Hypnotic Analgesia 1009
P Rainville, Universite de Montreal, Montreal, QC, Canada
I Marc, Universite Laval, Quebec City, QC, Canada

Index 1017
Introduction
There is no coming to consciousness without pain. Carl Gustav Jung

It has been argued that pain, unlike audition, vision, somatosensation, and olfaction, is not a primary sense,
but instead is more of an emotional experience. Most researchers of pain, however, consider pain to be a
complex perception evoked by noxious stimuli. Pain is probably far more complicated than the other
perceptual modalities described in this series. For example, in the setting of tissue or nerve injury, where
pain is persistent, the stimulus that evokes pain can change. In fact, under these conditions innocuous stimuli
can readily evoke the perception of pain.
But even these unusual characteristics do not capture the features that make pain among the most complex
of perceptions. The International Association for the Study of Pain defines pain as An unpleasant sensory and
emotional experience associated with actual or potential tissue damage, or described in terms of such damage.
In other words, although there is a very discrete anatomical and physiological basis for the detection and
transmission of messages that are interpreted as painful, what makes the experience of pain so special is that
there is always a profound emotional quality to the experience.
Pain in general, and pain research in particular, is especially exciting as it brings together elements of so
many disciplines. This volume is comprehensive. It includes a wealth of information on the molecular biology,
anatomy, physiology, and biochemical bases of pain, both in the normal and injury setting. But this volume also
addresses the critical cognitive component of the pain experience, including some of the most provocative
cerebral imaging studies that for the first time are providing insights into the gestalt of brain activity that occurs
when pain is experienced. There are chapters on the pharmacological basis of the placebo, on the utility of
hypnosis for the treatment of pain, and even an essay on consciousness and pain.
This is not a how to treat clinical textbook. Nevertheless, the editors are advocates of the new mantra
in the field, namely that chronic pain is not a symptom of disease, but rather is a disease entity itself, a
disease of nervous system function. Therefore, in addition to covering the fundamentals of acute pain
processing, from the nociceptor to cortical activation, we also cover, in depth, the changes that occur in the
setting of injury, including molecular, structural, and biochemical alterations in the properties of nocicep-
tors and central nervous system pathways. Some of the particularly intractable clinical pain conditions are
discussed. These chapters not only provide insights into pathophysiology but also clues to pain
management.
Of course, a variety of compendia have recently appeared, and many also provide comprehensive reviews of
the field. With this in mind, the editors have made a concerted effort to produce a final product that is different.
Too often the excitement that epitomizes the field of pain research is buried within, or indeed omitted from, the
typical edited book. Some textbooks include the proverbial box that highlights an interesting topic, but these
are generally very limited. We wanted to bring these topics to the forefront. Our approach is to include, in
association with each major chapter, at least one or two cameos that illustrate fascinating and provocative areas
of basic and clinical neuroscience that intersect the study of pain.
A few years ago we knew almost nothing about the cortical mechanisms that underlie the pain experience.
Today some scientists, albeit the minority, believe that cortical imaging can provide an objective measure of the
pain experience. A few years ago, the tetrodotoxin-resistant subtype of voltage-gated sodium channel, NaV1.8,

ix
x Introduction

was considered the Holy Grail for the next breakthrough in pain management. How fast things change. The
discovery that a loss of function mutation of NaV1.7 underlies a condition of congenital insensitivity to pain and
that a gain of function mutation underlies the excruciatingly painful condition of erythromelalgia has
dramatically altered the focus, not only of the science community but also of the pharmaceutical industry.
The pace of discovery in pain research is indeed remarkable. We hope that this volume conveys the excitement
inherent in this discovery process and, most importantly, that it stimulates the next generation of basic and
clinical scientists to unravel the mystery of the pain experience.

Allan I. Basbaum and M. Catherine Bushnell

 1 The Adequate Stimulus
R D Treede, Johannes Gutenberg-University, Mainz, Germany
2009 Elsevier Inc. All rights reserved.

1.1 Noxious Stimulus 1

1.2 Nociceptive Stimulus 3
References 3

Glossary
nociception The processes of encoding and nociceptor A primary afferent nerve fiber that is
processing of noxious stimuli by the nervous capable of encoding noxious stimuli. All non-noci-
system. ceptive afferents (e.g., tactile receptors, warm
nociceptive stimulus An actually or potentially receptors) do respond to noxious stimuli (mechan-
tissue damaging event that is encoded by ical or thermal, respectively), because these stimuli
primary nociceptive afferents. Although actual or are way above their respective thresholds. But only
potential tissue damage is the common denomi- nociceptors are capable of encoding the relevant
nator of those stimuli that may cause pain, there properties of those stimuli (e.g., sharpness, heat
are some types of tissue damage that are not intensity in the painful range).
detected by any afferents, and thus do not cause noxious stimulus An actual or potential tissue
pain. Therefore, not all noxious stimuli are damaging event. This was found to be the common
adequate stimuli of nociceptive afferents. The denominator of those stimuli that may cause pain.
adequate stimuli of nociceptors are termed But there are some types of tissue damage that are
nociceptive stimuli, which is a subset of noxious not detected by any afferents, and thus do not
stimuli. cause pain. See nociceptive stimulus.

The term adequate stimulus (Kandel, E. R. et al., 2000) Noxe) or environmental phenomena that threaten to
is used in sensory physiology to describe the class of cause such damage. Hence, the adequate stimulus to
environmental phenomena that requires the least elicit pain is traditionally called a noxious stimulus. It
amount of energy in order to elicit a percept mediated may be defined as follows.
by a particular sensory system, for example, a visual
percept can be elicited by a punch to the eye, but light
from a television screen needs much less energy for 1.1 Noxious Stimulus
this purpose. The implication from such observations
in subjective sensory physiology is that the receptive A noxious stimulus is an actual or potential tissue
organs of each sensory system are specialized to detect damaging event. Noxious stimuli may belong to ther-
a corresponding class of environmental phenomena mal, mechanical, or chemical modalities of energy
(i.e., photoreceptors in the eye are specialized to detect supply. Therefore, the sensory organs of the nocicep-
photons). tive system, free nerve endings of thinly myelinated A
It was difficult to transfer this concept to the per- fibers, and unmyelinated C fibers in the skin and most
ception of pain and to the nociceptive system. Many other tissues, have a characteristic property: most of
different stimuli may cause pain (e.g., pin prick, burn them are polymodal, that is, they respond to more than
injury, freeze injury, and inflammation), none of which one modality of stimulus energy. Some nociceptive
needs particularly low amounts of energy. Sherrington nerve endings, particularly those of A fibers, are spe-
is credited with identifying the common denominator cialized to be most sensitive to one particular stimulus
of those stimuli as being tissue damage (in Greek: vo modality (e.g., either heat or pin prick). The differential

1
2 The Adequate Stimulus

sensitivity spectra of nociceptive afferents suggest that efficacy of its central synapses. This process is called
pain quality may be encoded by a similar population central sensitization and may be considered to be one
code in the nociceptive system as taste quality in the of the phylogenetically oldest mechanisms of learning
gustatory system (Treede, R. D. et al., 1998). and memory (Woolf, C. J. and Walters, E. T., 1991).
Polymodality is not the result of a primitive non- Primary nociceptive afferents may also enhance their
specific responsivity to tissue damage. Instead, it is responsiveness to their adequate stimuli, a process
mediated by specialized signal transduction pathways, called peripheral sensitization (Raja, S. N. et al., 1999).
some of which have been elucidated at the molecular Both peripheral and central sensitization contribute to
level ( Julius, D. and Basbaum, A. I., 2001). One member the warning function of the nociceptive system.
of the gene family of transient receptor potential chan- As a warning system, the nociceptive system has
nels (TRPV1), for example, is specifically activated by gaps in its sensitivity: there are some types of tissue
moderate heat stimuli, low tissue pH, and a class of damage that are not detected by any afferents, and thus
irritant substances called vanilloids. For most inflam- do not cause pain or any protective behavior. This is a
matory mediators, specific receptor molecules are well-known phenomenon in internal organs such as the
present in the membranes of nociceptive nerve endings. liver or the brain, where a malignant tumor may cause
The topic of this volume of the Handbook of the Senses extensive damage without being noticed by the patient.
is pain. The taxonomy of the International Association There is an even more common phenomenon of tissue
for the Study of Pain (IASP) defines pain as, An damage that goes unnoticed by the nociceptive system:
unpleasant sensory and emotional experience asso- damage by ultraviolet radiation (Figure 1). The pain of
ciated with actual or potential tissue damage, or sunburn always comes too late, after the skin has
described in terms of such damage (Merskey, H.
et al., 1979). This definition implicitly refers to the
adequate stimulus as identified by Sherrington, in its
extended form including potential tissue damage rather
than calling for outright tissue damage. There are two
good reasons for this extension. First, a system that only
responds after tissue damage has occurred, cannot sub-
serve a warning function for the organism. The sensory
system that mediates pain sensation (the nociceptive
system), however, provides even primitive organisms
with an array of protective reflexes that can flexibly
respond to threatening environmental challenges. Such
a system needs to be sensitive enough to signal
impending tissue damage before it occurs. Second,
although primary nociceptive afferents, the sensory
organs of the nociceptive system, encode different
intensities of manifest tissue damage as graded action
potential discharge rates (Raja, S. N. et al., 1999),
the nociceptor activation thresholds were found to be
clearly lower than the intensity needed to damage the
skin. In humans as well as many animal species, heating
the skin to above 40  C, cooling to below 30  C, or
punctate pressure around 15 bars activates nocicep-
tors, but none of these stimuli damages the skin. In fact,
for most people these stimuli are not even painful.
The relatively low peripheral nociceptor activation
thresholds allow the nociceptive system to subserve its Figure 1 The concept of the adequate stimulus to activate
warning function in a highly flexible and plastic way. the nociceptive system and to elicit pain can be illustrated
by this scene. Ultraviolet radiation causes tissue damage,
The peripheral input can be shut down by descending
but this noxious stimulus is not detected by the nociceptive
inhibition, if there is an a priori reason to ignore the nerve endings. Only the inflammatory response of the skin
warning signal. The system can also enhance its leads to adequate activation of nociceptive nerve endings
response to the warning signal by enhancing the via chemical mediators, which serve as nociceptive stimuli.
 The Adequate Stimulus 3

already been damaged. This pain does not signal the by the visual system and auditory stimuli are
initial damage but the bodys response by an inflam- encoded by the auditory system. However, a full
matory reaction. The ensuing peripheral and central appreciation of our senses goes beyond these aspects
sensitization of the nociceptive system lead to pro- of sensory physiology (Bieri, P., 1995; Metzinger, T.,
nounced hyperalgesia of the injured skin to heat and 2000): seeing comprises more than vision, hearing
mechanical stimuli, which then serves to protect the comprises more than audition, and pain comprises
skin from further damage. These observations have led more than nociception. This is reflected in the defi-
to the introduction of a new term, nociceptive stimulus nition of pain as given above that does not relate to
(Cervero, F. and Merskey, H., 1996). any objective observable measure, but refers to the
subjective experience of the person in pain.

1.2 Nociceptive Stimulus

A nociceptive stimulus is an actual or potential tissue References

damaging event that is encoded by primary nocicep-
tive afferents. In summary, the adequate stimulus to Bieri, P. 1995. Pain a Case Study for the MindBody Problem.
In: Pain and the Brain: From Nociception to Cognition
activate the receptive organs of the nociceptive sys- (eds. B. Bromm and J. E. Desmedt), pp. 99110. Raven
tem consists of either actual or potential tissue Press.
damage (noxious stimulus). However, not all noxious Cervero, F. and Merskey, H. 1996. What is a noxious stimulus?
Pain Forum 5, 157161.
stimuli are detected by the nociceptive system. Julius, D. and Basbaum, A. I. 2001. Molecular mechanisms of
Therefore, the adequate stimulus for this system in nociception. Nature 413, 203210.
the strict sense is that subset of noxious stimuli that Kandel, E. R, Schwartz, J. H, and Jessell, T. M 2000. Principles
of Neural Science. 4th edn., p. 414. McGraw-Hill.
can be encoded by the nociceptive system (nocicep- Merskey, H., Albe-Fessard, D., Bonica, J. J., Carmon, A.,
tive stimuli). It is not unusual for a sensory system to Dubner, R., Kerr, F. W. L., Lindblom, U., Mumford, J. M.,
encode only a part of the range of environmental Nathan, P. W., Noordenbos, W., Pagni, C. A., Renaer, M. J.,
Sternbach, R. A., and Sunderland, S. 1979. Pain terms: a list
phenomena that its receptive organs are specialized with definitions and notes on usage. Recommended by the
for: visual stimuli consist of a restricted range of IASP subcommittee on taxonomy. Pain 6, 249252.
wavelengths of electromagnetic waves, and auditory Metzinger, T. 2000. The Subjectivity of Subjective Experience: A
Representationalist Analysis of the First-Person Perspective.
stimuli consist of a restricted frequency range of In: Neural Correlates of Consciousness: Empirical and
pressure waves in the air. Likewise, nociceptive sti- Conceptual Questions (ed. T. Metzinger), pp. 285306. MIT
muli consist of a restricted range of actually or Press.
Raja, S. N., Meyer, R. A., Ringkamp, M., and Campbell, J. N.
potentially tissue-damaging events. Linguistically, 1999. Peripheral Neural Mechanisms of Nociception.
the term nociceptive stimulus may appear to be a In: Textbook of Pain (eds. P. D. Wall and R. Melzack),
little odd, as it also implies reception rather than just 4th edn, pp. 1157. Churchill Livingstone.
Treede, R. D., Meyer, R. A., and Campbell, J. N. 1998.
stimulation, but for exactly that reason it fits well into Myelinated mechanically insensitive afferents from monkey
the broader system of terms in sensory physiology. hairy skin: heat response properties. J. Neurophysiol.
According to the concept of the adequate 80, 10821093.
Woolf, C. J. and Walters, E. T. 1991. Common patterns of
stimulus, nociceptive stimuli are encoded by the plasticity contributing to nociceptive sensitization in
nociceptive system, just as visual stimuli are encoded mammals and aplysia. Trends Neurosci. 14, 7478.
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 2 Pain Theories
F Cervero, McGill University, Montreal, QC, Canada
2009 Elsevier Inc. All rights reserved.

References 9

From the beginning of scientific enquiry there have necessarily have time-locked responses to painful
been two opposing views on the biological meaning stimuli. If pain is a passion of the soul then we need
of pain. One view proposes that pain is a sense similar distributed networks and interactive parallel proces-
to vision or hearing, a component of the sensory sing and not a pain pathway. This view has been
repertoire of most animals that warns us of impend- articulated throughout the last 100 years as the
ing damage, gives accurate information to the brain pattern theory of pain, denying the existence of
about injuries, and helps us to heal. The inclusion of dedicated sensory elements for pain processing and
pain in The Senses: A Comprehensive Reference, alongside attributing the perception of pain to interactions
vision, hearing, or olfaction shows that this view is between patterns of impulses in nonspecific neuronal
persuasive. But there has always been an alternative networks. The influential gate theory of pain
interpretation of pain that denies it being a sense like (Melzack, R. and Wall, P. D., 1965) is the best con-
vision or hearing and attaches to both pain and its temporary example of such an interpretation.
opposite pleasure fundamental roles in shaping the The specificity theory of pain was a natural devel-
emotions and behaviors of the individual. Pain is seen opment of the Doctrine of Specific Nerve Energies
as a trigger of emotional states, a behavioral drive, put forward by the German physiologist Johannes
and a highly effective learning tool. Aristotle, who Muller in the nineteenth century. The basic proposal
was the originator of this view, made it very clear: of Mullers doctrine is that each sensory modality is
there are only five senses vision, hearing, touch, the result of the activation of a specific neural system
taste, and smell. Pain and pleasure are not senses but in the brain. If we are able to perceive touch it is
passions of the soul. because we have a subset of cells in our peripheral
Whether or not pain is a sense like the others is and CNS that respond to touch; if an animal can
not just an academic exercise. The experimental perceive infrared light or an electromagnetic field
paradigms used to study the nervous system can be then it must have a subset of neurons capable of
fundamentally different depending on the theoretical sensing and processing these stimuli. As he put it:
approach to the object of the study. If pain is regarded Sensation is not the conduction of a quality or
as a sense, like vision or hearing, then we will look for state of external bodies to consciousness, but the
sensors that are activated selectively by painful conduction of a quality or state of our nerves to
stimuli and for sensory pathways in the brain and consciousness, excited by an external cause
spinal cord that carry pain information much in the (Muller, J., 18351840). At the end of the nineteenth
same way as we identify photoreceptors in the retina century, von Frey M. (1895) extended Mullers doc-
and a visual pathway to the cortex. This approach has trine to pain sensation, thus reinforcing a strict
generated an interpretation of pain mechanisms sensory interpretation of pain. He proposed that the
known as the specificity theory, which maintains fine nerve endings of unmyelinated afferents were
that there are elements of the peripheral and central the pain receptors in the periphery and that there was
nervous system (CNS) specifically and exclusively a specific pain pathway taking their signals to the
dedicated to the processing of pain-related brain. This influential proposal is responsible for
information. the well-known model of pain mechanisms often
However, if pain is not a sense like vision or found in textbooks whereby a pain receptor in the
hearing, then we do not need to look for a specific periphery is activated by a noxious stimulus and
neural machinery for its processing but for patterns of sends impulses to the spinal cord and from there to
activation, spatial or temporal, in neurons that do not the thalamus and cortex via a crossed spinothalamic

5
6 Pain Theories

pathway. In other words, a relatively simple and animal models of inflammatory and pathological
straight forward pain pathway (Figure 1(a)). pain (see Cervero, F., 2005 for a recent discussion
In contrast, the pattern theory of pain represents a on the gate theory).
development of the Aristotelian concept of pain as a The considerable amount of new information about
passion of the soul and was articulated in modern pain mechanisms gathered in the last 40 years has
form by Goldscheider A. (1898) at the turn of the relegated the specificity versus pattern argument to a
nineteenth century. The basic proposal is that pain is secondary role. It is now well established that there are
the result of intense stimulation of peripheral recep- specialized sensors in the skin, muscles, and viscera of
tors, regardless of modality and tissue origin. For the most animals that are activated exclusively by stimuli
pattern theory the neural substrate of pain perception that cause injury and whose excitation leads to the
consists of sequences of impulses in peripheral and sensation of pain (Belmonte, C. and Cervero, F.,
central neurons that lead to pain sensations when 1996). It is also known that pain cannot be evoked,
certain spatial and temporal patterns of activity are under normal circumstances, by changing the patterns
produced. The emphasis of this interpretation is on of activation of tactile sensory receptors (Ochoa, J. and
patterns of neural activity evoked in the brain by a Torebjork, E., 1983; 1989). There are also substantial
painful stimulus. In the 1960s, the gate theory of pain data showing that there are neurons in the spinal cord
proposed a specific neural mechanism located at the and brain driven mainly or exclusively by nociceptive
first afferent relay in the spinal cord to illustrate how stimuli (Hunt, S. P. and Mantyh, P. W., 2001).
patterns of impulses in sensory receptors could lead However, there is also significant experimental evi-
to the modulation of pain sensation (Figure 1(b)). dence in favor of plasticity in the nociceptive sensory
The gate theory emphasized the dynamic and plastic channel and of the existence of dynamic processes that
components of pain sensations and drew attention to can alter profoundly the functional properties of per-
pain modulation as opposed to an interpretation of ipheral nociceptors and of nociceptive central neurons
pain exclusively as an alarm system. It also focused (Treede, R. D. et al., 1992; Hunt, S. P. and Mantyh, P.
the attention of many researches on the clinical W., 2001; Julius, D. and Basbaum, A. I., 2001). It has also
aspects of pain, away from physiological pain, been demonstrated that following a peripheral injury
which contributed a surge of studies on the effects or inflammation, the activation of tactile afferents from
of neuropathic lesions and on the development of uninjured skin can evoke pain sensations (see Cervero,

(a) (b)

Central
control

Gate control system

L +
+
Action
Input SG T system

+
S

Figure 1 (a) Descartes drawing of a pathway-oriented pain mechanism. The fire activates pain nerves in the foot of the
child and these signals are transmitted to the brain where they are reflected into motor nerves that draw the foot away from
the fire. (b) Schematic diagram of the gate theory of pain mechanisms. The diagram shows the presynaptic interaction model
between large (L) and small (S) afferent fibers and the key role of substantia gelatinosa (SG) neurons controlling the activity of
transmission (T) cells. (a) From Descartes, R. 1664. Lhomme. Chez Jacques Le Gras. (b) Reprinted with permission from
AAAS from Melzack, R. and Wall, P.D. 1965. Pain mechanisms: a new theory. Science 150, 971979.
 Pain Theories 7

F. and Laird, J. M. A., 1996). Clearly, a point has been 100

reached where neither a strict specificity nor a pattern Hyperalgesia
interpretation can account for all that it is known about 75

Pain sensation
pain mechanisms.
We now realize that there are many different Injury
50
forms of pain (acute, traumatic, inflammatory, and Normal
Allodynia pain
neuropathic) and that pain is a dynamic process that
25
cannot be explained with a single theory or a unique
mechanism. One of the most striking expressions of
the dynamic nature of pain sensation is its lack of 0
Innocuous Noxious
adaptation. A continuous and uniform visual or audi-
Stimulus intensity
tory stimulus leads to sensory adaptation; we simply
stop feeling this stimulus after a few seconds or Figure 2 Diagram illustrating the changes in pain
minutes. However, the sensation of pain not only sensation induced by injury. The normal relationship
between stimulus intensity and the magnitude of pain
does not adapt to a continuous noxious stimulus but sensation is represented by the curve at the right-hand side
gets progressively worse so that, after a few minutes of the figure. Pain sensation is only evoked by stimulus
of persistent stimulation with a relatively mild pain- intensities in the noxious range (the vertical dotted line
ful stimulus, the sensation becomes unbearable. This indicates the pain threshold). Injury provokes a leftward shift
change in pain sensitivity generates a state of pain in the curve relating stimulus intensity to pain sensation.
Under these conditions, innocuous stimuli evoke pain
amplification or hyperalgesia that is normally (allodynia). Reproduced From Cervero, F. and Laird, J. M. A.
triggered and maintained by a persistent noxious 1996. Mechanisms of touch-evoked pain (allodynia): a new
stimulus but that can, under pathological circum- model. Pain 68, 1323, used with permission.
stances, appear without an obvious cause so that the
normal relationship between injury and pain is lost.
Hyperalgesia and allodynia are the main symptoms to the alterations in central processing that are, in
of many chronic pain states and is the property of turn, responsible for secondary hyperalgesia (Treede,
pain sensation that makes it particularly unpleasant R. D. et al., 1992).
and often unbearable. Secondary hyperalgesia is defined as an increased
In psychophysical terms a hyperalgesic state is sensitivity to pain occurring in areas adjacent or even
represented by a leftward shift that occurs, following remote to the site of injury. For instance, following an
a peripheral injury, in the curve that relates stimulus injury to the hand an area of hyperalgesia may
intensity to pain sensation (Cervero, F. and Laird, J. develop covering the entire arm or an inflammation
M. A., 1996) (Figure 2). This shift causes the lower of the gastrointestinal tract or the bladder may pro-
portion of the pain curve to fall in the innocuous duce an area of hyperalgesia in the abdominal or
stimulus intensity range (allodynia or pain produced pelvic regions. Secondary hyperalgesia is the result
by an innocuous stimulus) whereas the top portion of an alteration in the processing by the CNS of
shows an increased pain sensation to noxious stimuli impulses from low-threshold mechanoreceptors,
(hyperalgesia proper or increased pain sensitivity to a such that, these impulses are able to activate noci-
noxious stimulus). Allodynia and hyperalgesia pro- ceptive neurons and evoke pain. This central
vide protective mechanisms to the organism, alteration is initially triggered and later maintained
preventing the individual from stimulating an injured by the enhanced afferent discharges from the primary
area and in so doing helping the healing process. hyperalgesic area (Treede, R. D. et al., 1992).
There are two forms of hyperalgesia: primary and Different forms of pain are mediated by different
secondary. Primary hyperalgesia is an increased pain mechanisms which participate in various ways in the
sensitivity that occurs at the site of injury and it is generation of pain and hyperalgesia (Cervero, F. and
the consequence of nociceptor sensitization, that is, Laird, J. M. A., 1991; Klein, T. et al., 2005). Generally
the increased firing of peripheral nociceptors at the speaking we can identify three different forms of pain
site of injury whose excitability has been increased by taking into account the relationship between noxious
a number of locally released sensitizing agents. These stimulus and pain sensation: nociceptive, inflamma-
sensitized nociceptors send enhanced afferent dis- tory, and neuropathic (also called phase 1, 2, and 3
charges to the CNS thus evoking increased pain pains by Cervero, F. and Laird, J. M. A., 1991)
from the primary hyperalgesic area and contributing (Figure 3). Nociceptive pain refers to the processing
8 Pain Theories

CNS Pain
Phase 1

Brief Brief
injury

Phase 2

Persisting
+
Inflammation

Phase 3

Nerve +
Abnormal
or CNS
damage

Figure 3 Diagram showing the three models of pain processing for the three main types of pain (nociceptive, inflammatory,
and neuropathic or phases 1, 2, and 3). See text for further explanation. Reproduced from Cervero, F. and Laird, J. M. A. 1991
One pain or many pains? A new look at pain mechanisms. News Physiol. Sci. 6, 268273, used with permission.

of brief noxious stimuli; inflammatory pain is the discharges in peripheral nociceptors and pain per-
consequence of prolonged noxious stimulation lead- ception is lost.
ing to tissue damage; and neuropathic pain is the Neuropathic pain syndromes are the consequence
consequence of neurological damage, including per- of damage to peripheral nerves or to the CNS itself
ipheral neuropathies and central pain states. and produce pain sensations well outside the range of
Nociceptive pain is a protective sensation needed the sensations produced by the normal nociceptive
for the survival and well-being of the individual. The system, even after serious peripheral injury or
mechanisms subserving the processing of brief inflammation. These include spontaneous pain,
noxious stimuli can be viewed as a fairly simple greatly reduced pain thresholds, and mechanical allo-
pathway that carries impulses in peripheral nocicep- dynia. Neuropathic pain states are characterized by
tors centrally toward the thalamus and cortex and an almost complete lack of correlation between
leads to brief pain perception. In contrast, injury and peripheral noxious stimuli and pain sensation and
tissue damage evoke an inflammatory reaction as part are produced by neurological lesions that cause
of the healing process and generate a pain state abnormal impulse activity generated in nerve
different from nociceptive pain as the response prop- sprouts, neuromas, or in dorsal root ganglion cells,
erties of the various components of the nociceptive ephaptic coupling between adjacent nerve fibers and
system change. These changes include nociceptor abnormal responses of peripheral nociceptors and
sensitization and recruitment of populations of CNS neurons. Nociceptive and inflammatory pains
previously unresponsive receptors. In turn, CNS are symptoms of peripheral injury, whereas neuro-
neurons show an amplification of their excitability pathic pain is a symptom of neurological disease.
expressed as increases in receptive field size and Whereas the specificity theory can explain fairly
greater spontaneous and evoked firing. All of these well the simpler forms of pain, such as a pin prick or
changes indicate that the CNS has moved to a new, the acute pain of a minor burn, complex pain experi-
more excitable state as a result of the noxious input ences, including hyperalgesic states require
generated by tissue injury and inflammation. Under substantial peripheral and central plasticity such
these conditions, an immediate correlation between that low-intensity stimuli evoke pain (i.e., secondary
 Pain Theories 9

CNS
Nociceptor
sensitization A/C

Synaptic
strengthening
by incoming
A afferent volleys

LT
Secondary
hyperalgesia A/C

Activation of
nociceptive N
neurons by LT
afferents

Figure 4 Diagram representing the basic mechanisms of primary and secondary hyperalgesia and the three key processes
implicated in their generation. Primary hyperalgesia is produced by the stimulation of nociceptors connected to A- and
C-afferent fibers which activate nociceptive CNS pathways (N). Secondary hyperalgesia is produced by stimulation of tactile
receptors connected to A-afferents which normally activate low-threshold (LT) pathways but that as a consequence of the
amplification of the nociceptive input from the injured area can now access nociceptive neurons (N). See text for further
explanation.

hyperalgesia or touch-evoked allodynia). It is very the result of sensitization of peripheral nociceptors

likely that both specific and nonspecific nociceptive leading to decreases in their activation threshold or
systems participate in the generation and mainte- be mediated by a purely central mechanism whereby
nance of different pain states. low-threshold mechanoreceptors access nociceptive
We can currently identify three key processes in neurons. The emphasis has moved from theoretical
the neurobiological approach to the mechanisms of interpretations based on a specific pain channel or on
pain and hyperalgesia: (1) the process of nociceptor patterns of impulses to the in depth analysis of the
activation and sensitization, responsible for the initial functional processes that may cause the different pain
signaling of injury and the peripheral changes in the and hyperalgesic states. Neither the specificity nor
nociceptive system induced by a noxious stimulus; the pattern theories have been able to provide work-
(2) the process of central amplification of nociceptive able models for all forms of pain sensation. They still
signals, known as central sensitization, generated by offer a useful theoretical framework for some of our
synaptic strengthening of connections between CNS observations but the weight of work has shifted to
neurons and responsible for the enhanced excitability studying those processes that will help the develop-
that accompanies persistent pain states; and (3) the ment of effective pain relief therapies.
process whereby activity in low-threshold sensory
receptors from undamaged peripheral areas can
access the nociceptive system and evoke pain sensa- References
tions and hyperalgesic states (e.g., touch-evoked pain,
tactile allodynia) (Figure 4). Belmonte, C. and Cervero, F. 1996. Neurobiology of
The analysis of these three processes supersedes Nociceptors. Oxford University Press.
Cervero, F. 2005. The Gate Theory Then and Now. In: The Paths
interpretations of pain based on a specificity or pat- of Pain (eds. H. Mershey, J. De Loeser, and R. Dubner),
tern approach as all three contain elements that pp. 3348. IASP Press.
require the existence of groups of neurons dedicated Cervero, F. and Laird, J. M. A. 1991. One pain or many pains?
A new look at pain mechanisms. News Physiol. Sci.
to the processing of nociceptive signals as well as the 6, 268273.
expression of functional changes induced by afferent Cervero, F. and Laird, J. M. A. 1996. Mechanisms of touch-
activity. Several of these changes can lead to major evoked pain (allodynia): a new model. Pain 68, 1323.
Descartes, R. 1664. Lhomme. Chez Jacques Le Gras.
alterations of pain sensitivity through quite separate Goldschaider, A. 1898. Uber den Schmerz: Gesammelte
mechanisms. For instance, touch-evoked pain can be Abhandlungen. Ambros.
10 Pain Theories

Hunt, S. P. and Mantyh, P. W. 2001. The molecular dynamics of Ochoa, J. and Torebjork, E. 1983. Sensations evoked by
pain control. Nature Rev. Neurosci. 2, 8391. intraneural microstimulation of single mechanoreceptor units
Julius, D. and Basbaum, A. I. 2001. Molecular mechanisms of innervating the human hand. J. Physiol. 342, 633654.
nociception. Nature 413, 203210. Ochoa, J. and Torebjork, E. 1989. Sensations evoked by
Klein, T., Magerl, W., Rolke, R., and Treede, R. D. 2005. intraneural microstimulation of C nociceptor fibres in human
Human surrogate models of neuropathic pain. Pain skin nerves. J. Physiol. 415, 583599.
115, 227233. Treede, R. D., Meyer, R. A., Raja, S. N., and Campbell, J. N.
Melzack, R. and Wall, P. D. 1965. Pain mechanisms: a new 1992. Peripheral and central mechanisms of cutaneous
theory. Science 150, 971979. hyperalgesia. Prog. Neurobiol. 38, 397421.
Muller, J. 18351840. Handbuch der Physiologie des Menschen von Frey, M. 1895. Beitrage zur sinnesphysiologie der haut. Ber.
fur Vorlesungen. J. Holscher. Sachs. Ges. Wiss. 47, 166184.
 3 Anatomy of Nociceptors
S Mense, Institut fur Anatomie und Zellbiologie, Universitat Heidelberg, Heidelberg, Germany
2009 Elsevier Inc. All rights reserved.

3.1 Introduction 12
3.2 General Morphological Features of Nociceptors 14
3.2.1 Light Microscopic Structure 14
3.2.2 Ultrastructure 15
3.3 Nociceptors in Various Tissues 17
3.3.1 Skin 17
3.3.1.1 Light microscopy 18
3.3.1.2 Electron microscopy 19
3.3.2 Cornea 20
3.3.2.1 Light microscopy 20
3.3.2.2 Electron microscopy 21
3.3.3 Deep Somatic Tissues 22
3.3.3.1 Muscle pain compared to cutaneous pain 22
3.3.3.2 Nociceptors in muscle and tendon 22
3.3.3.3 Nociceptors in joints and ligaments 25
3.3.3.4 Tooth pulp 27
3.3.3.5 Dura mater encephali 32
3.3.4 Visceral Organs 33
3.3.4.1 Visceral pain 33
3.3.4.2 Testis 34
3.4 Neuropeptide Content of Nociceptors 34
3.4.1 General Remarks 34
3.4.2 Neuropeptides in Afferent Units of Different Tissues 35
3.4.3 Neuropeptides in Nociceptors and Other Types of Free Nerve Ending 36
3.5 Efferent Function of Nociceptors 37
3.5.1 Release of Neuropeptides from the Nociceptive Ending 37
3.5.2 The Axon Reflex 37
3.5.3 Neurogenic Inflammation 37
3.6 Receptor Molecules in the Membrane of Nociceptors 38
References 38

Glossary
anterograde transport Intra-axonal transport in antidromically (against the normal direction of pro-
the distal direction (away from the soma) of sub- pagation) by action potentials. The released
stances synthesized in (or injected into) spinal or neuropeptides influence the microcirculation in the
cranial ganglion cells. The transport is brought vicinity of the ending.
about by molecules (kinesin for anterograde, and deep somatic tissues All subcutaneous tissues
dynein for retrograde, transport) that bind sub- that are not viscera, namely tendon, fascia, mus-
stances and vesicles and move along the cles, ligaments, and joints.
microtubules in the axoplasm. epitendineum (peritendineum externum) Dense
axon reflex Release of neuropeptides from a connective tissue with blood vessels, lymphatic
branch of a nociceptor when the branch is invaded vessels, and nerve fibers surrounding a tendon.

11
12 Anatomy of Nociceptors

group IIV fibers Nomenclature for afferent fibers skeletal muscle, it is often supplied by a group III
that originate in deep somatic tissues. Group IV fiber.
fibers correspond to C-fibers from the skin, and primary afferent unit The first sensory neuron in
group III to A-fibers. Group I fibers comprise the the body periphery. It includes the receptive
primary endings from muscle spindles (Ia-fibers) ending, the afferent fiber, the soma in the spinal
and Ib-fibers from Golgi tendon organs. Group II or cranial ganglion, and the central process in the
fibers and cutaneous A-fibers are largely identical. dorsal root and the central nervous
The denominations using roman numerals are system (CNS).
based on the diameter of the nerve fibers, those receptor matrix An arrangement of cell organelles
using arabic letters on the conduction velocity. (mitochondria, vesicles, and axonal reticulum, i.e.,
noceffector This term was coined by Kruger L. a network of fluid-filled vacuoles) embedded in a
(1988) for nociceptors in the tooth; it emphasizes granulated axonal cytoplasm.
the efferent function of the receptor. Efferent func- Ruffini corpuscle A flat, encapsulated mechan-
tion means that the receptor is involved in the oreceptor supplied by several myelinated nerve
maintenance of the tissue under normal and fibers. The fibers form a dense arborization within a
pathological circumstances by releasing sub- capsule of connective tissue.
stances stored in the ending. One aspect of such a sensory terminal tree The preterminal axon and
noceffector role in the skin would be regulation of its branches. The term is part of the concept that
Langerhans cell type expression and proliferation both group III and IV endings do not have just one
of keratinocytes. Under pathophysiological cir- sensory site, but possess several branches with
cumstances, the effector role of a nociceptor is many receptive loci that together enhance the
reflected in the axon reflex (see above) and neuro- sensitivity of the free nerve ending as a sense
genic inflammation. organ.
paciniform corpuscle A rapidly adapting varicosity An expansion of the preterminal axon of
mechanoreceptor with a morphology similar to that a slowly conducting sensory fiber (A- or C-fiber).
of a Pacinian corpuscle. It consists of a layered The varicosity contains mitochondria, vesicles, and
capsule that surrounds a central receptive core. other cell organelles. It is discussed as a receptive
The capsule is derived from the perineurium; the site. The varicosities of a free nerve ending are
central core contains the receptive portion of the connected by thin stretches of axon; this arrange-
axon with Schwann cells around it. Compared to ment causes the beaded appearance of a free
the Pacinian corpuscle, the paciniform corpuscle is nerve ending and the preterminal axon under the
smaller and has fewer laminae in its capsule. In light microscope.

3.1 Introduction nociceptor is an ending that by its discharge behavior

is capable of distinguishing between an innocuous
Originally, pain was assumed to be an emotion like and a noxious stimulus. An important point for the
pleasure and fear. In the nineteenth century, this understanding of the function of nociceptors is that
concept was replaced by the view that pain is due their stimulation threshold is just below tissue-dama-
to activation of a set of specialized nerve endings. ging intensity. The function of nociceptors is not to
Von Frey M. (1894) was the first to link pain to fine signal existing tissue damage, but to inform the cen-
nerve terminals in the skin. The functional term tral nervous system (CNS) when stimuli approach
nociceptor is derived from the Latin word noxius for tissue-threatening intensities.
damaging or harmful. It denotes a sensory ending Nociceptive endings are present in almost all tis-
that detects actual or potential tissue damage. When sues and organs of the organism. However, the fact
activated, it may cause pain in humans and pain- that lesions of the brain and parenchyma of the lung,
related reflexes in animals. Another definition of a liver, and cartilage are not painful suggests that in
 Anatomy of Nociceptors 13

these tissues nociceptors are not present. Conversely, (a) 3000 (b)
by stimulation of cornea, dura mater, and tooth pulp,

(m2)
pain is the predominant or only sensation that can be
6000
elicited. These observations led to the assumption

Cross-sectional area (m2)

that these tissues are equipped exclusively with 5000
nociceptors.
The available clinical and experimental evidence 4000
1 2 (m s1)
indicates that small-diameter afferent fibers have to
be activated in order to elicit pain. These fibers 3000
conduct action potentials at a relatively slow velocity
(below 30 m s1 in the cat); histologically they com- 2000

prise either thin myelinated (A- or group III) fibers

1000
or nonmyelinated (C- or group IV) fibers. However,
there are also nociceptors supplied by faster-conduct-
ing thick myelinated A/-fibers: in the rat, 10 20 30 CV(m s1)
Group: lV lll
approximately 20% of all A/-fibers are nociceptive
(for review, see Djouhri, L. and Lawson, S. N., 2004). Figure 1 Relationship between cross-sectional area of
The nomenclature with roman numerals (group IIV dorsal root ganglion (DRG) cells and conduction velocity
(CV) of the peripheral axon. (a) Data from 12 unmyelinated
fibers) was developed by Lloyd D. P. C. (1943) for
(C- or group IV) fibers and 10 thin myelinated (A- or group
muscle afferent fibers. It is now being used for afferent III) fibers. (b) Data from the unmyelinated units at an
fibers from muscle, joint, tendons, and fascia (the so- expanded scale of the abscissa. There was no significant
called deep somatic tissues). correlation between soma size and CV within group III
Generally, there is a negative relationship fibers, and for group IV units the correlation was negative.
(Modified from Hoheisel, U. and Mense, S., 1987, with
between mechanical threshold and conduction velo- permission).
city (CV) of sensory afferents: the higher the CV, the
lower the mechanical threshold (Burgess, R. P. and
Perl, E. R., 1967). In the dorsal root ganglion (DRG),
the correlation between soma diameter and axonal neuropeptides may be helpful, since for instance
CV is weaker than generally thought. In Figure 1 CGRP is absent from postganglionic sympathetic
(Hoheisel, U. and Mense, S., 1987), this correlation is fibers ( Ju, G. et al., 1987). However, postganglionic
shown for intracellularly stained DRG cells of the parasympathetic fibers have also been shown to con-
cat. For group III units (those having CVs between tain neuropeptides. Therefore, if neuropeptide-
2.5 and 30 ms1), there was no significant correlation immunoreactive (ir) free nerve endings are being
between soma size and CV, and for group IV units studied, one has to assume that a certain (probably
(conducting at <2.5 ms1), the correlation was nega- small) proportion of the endings originate from auto-
tive. Thus, from the soma size in the DRG, the nomic efferent fibers. Of course, the best way of
diameter or CV of group III and IV afferent fibers eliminating this problem is to cut the autonomic
cannot be inferred. Generally, the majority of noci- supply to the studied body region.
ceptive free nerve endings will originate from small- Not all small-diameter, slowly conducting affer-
diameter DRG cells, but there are also relatively ent fibers are nociceptive. In cutaneous nerves, there
large somata that supply nociceptive terminals. are thermoreceptors and low-threshold mechanore-
Conversely, not all the small DRG cells supply noci- ceptors that have unmyelinated afferent fibers, and
ceptive endings. also in skeletal muscle low-threshold mechanorecep-
In morphological studies on the endings of tors with group IV afferents can be found (Light, A. R.
unmyelinated fibers, the first hurdle to be taken is and Perl, E. R., 2003; Hoheisel, U. et al., 2005). The
the distinction between efferent and afferent units, presence of certain neuropeptides such as CGRP does
because postganglionic sympathetic fibers may look not distinguish between high- and low-threshold
identical to unmyelinated afferent ones, except mechanosensitive (HTM and LTM) group IV units,
that the former have a larger mean diameter because the neuropeptide was found in both functional
(Heppelmann, B. et al., 1988). The presence of types (Hoheisel, U. et al., 1994).
14 Anatomy of Nociceptors

The development of myelinated nociceptors has 3.2 General Morphological Features

been reported to depend upon the presence of trkA of Nociceptors
(tyrosin kinase A) and trkC receptors in the DRG,
3.2.1 Light Microscopic Structure
but not upon trkB receptors (Ichikawa, H. et al., 2004).
The trk receptors bind neurotrophins (NTs) selec- Morphologically, a nociceptor is a free nerve ending
tively; the ligand for trkA is nerve growth factor that is usually connected to the CNS through thin
(NGF), that for trkB, brain-derived neurotrophic myelinated or unmyelinated nerve fibers. (As stated
factor (BDNF), and that for trkC, NT-3. above, there are also nociceptors with thick myeli-
Even though several groups have studied and nated fibers.) C- or group IV fibers are assumed to
described the structure of putative nociceptors, terminate exclusively in free nerve endings while
clear morphological (light or electron microscopic) group III fibers supply both free nerve endings and
differences between the various functional types of other types of receptors (for instance in muscle pacini-
nociceptor are not apparent. The functional differ- form corpuscles; Barker, D., 1967; Stacey, M. J., 1969).
ences are probably due to the equipment of the A frequent location of free nerve endings is the wall of
endings with different sets of receptor molecules arterioles and the surrounding connective tissue; the
(Cesare, P. and McNaughton, P., 1997). capillaries proper do not have a rich supply with these
In neurophysiological experiments on cats and endings (Stacey, M. J., 1969; Reinert, A. et al., 1998).
rats, an unexpected finding was that many unmyeli- The marked sensitivity of free nerve endings to che-
nated afferent units had two receptive fields (RFs), mical stimuli, particularly to those associated with
i.e., the same fiber could be activated by applying disturbances of the microcirculation, may be related
stimuli to more than one location. In the deep tissues to their location in or close to the wall of blood vessels.
of the cat tail, afferent units were found that had one The term free nerve ending or unencapsulated end-
RF in deep tissues (muscle, joint, and periosteum) ing is derived from the fact that in the light microscope
and another one in the skin distal to the deep RF this type of ending lacks a visible (corpuscular) recep-
(Mense, S. et al., 1981). Sometimes, the distance tive structure (Stacey, M. J., 1969). Typically, such an
between the RFs could be several centimeters. The ending consists of several branches or terminals that
anatomical basis of this feature may be branching of altogether form the receptor in the morphological
the afferent fiber close to its area of termination. sense. An afferent fiber together with its receptive end-
Anatomical and neurophysiological studies (Devor, ing is an afferent unit. In the fluorescence microscope,
M. et al., 1984; Pierau, E.-K. et al., 1984) have shown the receptive ending and the preterminal axon look like
that afferent units with long branched axons are a string of beads with relatively wide diameter (so-
rare (a few percent of all afferent fibers), but they called varicosities) connected by very thin stretches of
may be functionally relevant, since they are likely to axon. The diameter of a branch of a free nerve ending is
reduce the spatial resolution of the nociceptive 0.51.0 mm, i.e., at the limits of the resolution of the light
system, and thus could contribute to the diffuse microscope. Figure 2 shows a free nerve ending in a rat
nature of deep pain.
The data that are most important to this chapter
were obtained in studies employing a combination of Sp-ir terminal axons
and receptive endings
electrophysiological and neuroanatomical techni-
ques, because otherwise one does not know if a
given free nerve ending in a histological section has
a nociceptive function. However, many purely mor-
phological investigations on free nerve endings
Muscle fiber
yielded results that are relevant to this chapter,
even though the evidence for a nociceptive function
is indirect. Therefore, many of these studies are
included below. 30 m
The main emphasis of this chapter is on nocicep-
Figure 2 Histological section from rat gastrocnemius
tors or free nerve endings, respectively, in skin,
muscle showing two nerve fibers with varicosities visualized
muscle/joint, tooth pulp, and testis, because large with fluorescent antibodies to substance P (SP; U. Hoheisel
sets of data are available from these tissues. and S. Mense, unpublished data).
 Anatomy of Nociceptors 15

skeletal muscle after labeling with fluorescent antibo- channels than the IB4-negative fibers (Wu, Z.-Z. and
dies to SP. Pan, H.-L., 2004). Another difference between these
Most of the unmyelinated afferents enter the two types of putative nociceptors is that the IB4-
spinal cord via the dorsal root. However, there are positive endings terminate freely in the tissue
also unmyelinated afferent fibers in the ventral root: without close spatial relationship to blood vessels,
approximately 30% of all ventral root fibers are whereas the peptidergic fibers are often associated
unmyelinated and 50% of these fibers survive in with blood vessels (Silverman, J. D. and Kruger, L.,
the distal stump after cutting the ventral root. 1988; Figure 3). At present, however, the assumption
Therefore, they were assumed to be afferent with prevails that the separation between these two
their soma located in the DRG (Applebaum, M. L. postulated types is probably not that sharp and that
et al., 1976). IB4-positive endings may also express TRPV1
In immunohistochemical studies, sensory free receptors.
nerve endings have been described as either peptider- In experiments employing a combination of electro-
gic or lectin-positive, the latter being largely peptide physiological and immunohistochemical techniques to
free. The lectin-positive endings possess membrane- study functionally identified DRG cells, Lawson S. N.
associated glycoconjugates that bind the plant lectin et al. (1997) reported that cells terminating in cutaneous
Griffonia simplicifolia isolectin B4 (IB4). They are nociceptive endings showed a strong tendency to
assumed to be identical to the fluoride-resistant acid express SP, particularly if they had a slow CV or
phosphatase (FRAP) endings. Many of the IB4-posi- small somata in the DRG.
tive endings express the purinergic P2X3receptor. In
contrast, the peptidergic endings are equipped with
the vanilloid receptor TRPV1 (transient receptor 3.2.2 Ultrastructure
potential subtype V1). These cytochemical differences In electron microscopic studies, often uncertainties
led to the hypothesis that the IB4-positive and -nega- remain as to the identity of the ending. Even when in
tive endings represent two functionally distinct classes combined electrophysiologicalmorphological investi-
of nociceptors. (Snider, W. E. and McMahon, S. B., gations an ending is first functionally characterized and
1998; Stucky, C. L. and Lewin, G. R., 1999). Recent then morphologically reconstructed, it is sometimes
evidence indicates that the peptidergic endings difficult to make sure that a given ending in a histolo-
express trkA receptors, are dependent on NGF in gical section is the one that was studied functionally.
their development, and express predominantly tetro- The published findings demonstrated that free
dotoxin (TTX)- sensitive Na channels, whereas the nerve endings are not free in the strict sense, because
nonpeptidergic IB4-positive endings depend on glial the majority are ensheathed by Schwann cells (there-
cellderived neurotrophic factor (GDNF) in their fore, the term free nerve ending is a misnomer).
development and express more TTX-resistant Na Exceptions to this rule are endings in the epidermis

(a) (b)

V
V

Figure 3 Testicular whole-mount preparation of the rat epididymal tunica vasculosa stained with Griffonia simplicifolia A-B4
lectin (a) and antibodies to calcitonin gene-related peptide (CGRP; (b)). Slender lectin-stained axonal bundles and terminals
distribute freely throughout the tissue (a). The lectin-labeled axons bear little relation to blood vessels (V) whose endothelial basal
lamina is also lectin-stained. In contrast, CGRP immunoreactivity is evident as heavily labeled coarse nerve bundles from which
one can trace fine granular axons running closely adherent to blood vessels (V). (From Silverman, J. D. and Kruger, L., 1988, with
permission.)
16 Anatomy of Nociceptors

that lose their Schwann cell covering, when they pass

the basal lamina of the epidermal layer (see below).
Usually, also the receptive terminals are ensheathed
by Schwann cells, and only small areas of the axonal
membrane remain uncovered by Schwann cell pro-
cesses. These areas are separated from the interstitial
fluid only by the basal lamina that spans the exposed
axonal membrane areas (Andres, K. H. et al., 1985;
Heppelmann, B. et al., 1990a; 1990b; Messlinger, K.,
1997). The varicosities in the exposed membrane
areas are supplied with mitochondria and vesicles
and show other structural specializations character- A
istic of receptive structures, and together with the
end bulb they are assumed to be the site where
external stimuli act. A reconstruction of such an end-
SC
ing is shown in Figure 4.
The arrangement of cell organelles (mitochon-
dria, vesicles, and axonal reticulum, i.e., a network
of fluid-filled vacuoles) embedded in a granulated
axonal cytoplasm was called receptor matrix by
Andres K. H. and von During M. (1973). Often,
nociceptive endings exhibit granular or dense-core
vesicles containing neuropeptides. However, vesicles
containing catecholamines can have the same
appearance, so that dense-core vesicles are not an
unambiguous sign of a nociceptive ending. The func-
tion of the round clear vesicles in the peripheral
ending is still obscure. They may contain the same
transmitters as the central synaptic terminal (e.g.,
glutamate). Recent reports have shown that gluta-
mate is an effective stimulant for nociceptive
endings in the body periphery (Svensson, P. et al.,
2003). Therefore, it is conceivable that nociceptors
enhance their own excitation by releasing glutamate
when they are activated. z
In a recent survey, Kruger L. et al. (2003a) list
some features that were considered characteristic y x
for nociceptive terminals in the body periphery: 1 m

(1) An axonal reticulum that may be derived from Figure 4 Reconstruction of an end branch of an articular
the smooth endoplasmic reticulum. group III ending. Left: Contours of the cross-sectioned
(2) Vesicle aggregates embedded in a granular axo- nerve fiber at a distance of 1 mm each (computer-aided
reconstruction). Right: Hand drawing based on the stack of
nal matrix that may be identical to the receptor sections showing the surface of the end branch. The bare
matrix described by Andres K. H. and von axon areas (A) are densely dotted; SC, Schwann cell. (From
During M. (1973). Messlinger, K. et al., 1995, with permission.)
(3) Exposed membrane areas that are not ensheathed
by Schwann cells and directly contact the basal
lamina. branches constitute a sensory tree as shown in
Figure 5 (Messlinger, K., 1996).
The receptive terminal as a whole consists of several It is important to note that the above features
branches equipped with many receptive sites that cannot be found at the synaptic terminal of the
have the appearance of varicosities. Together the same neuron, i.e., the central and distal terminals of
 Anatomy of Nociceptors 17

One of the characteristic functional properties of a

nociceptor is its relatively high mechanical stimula-
tion threshold. The high mechanical threshold is
surprising, considering the fact that a free nerve end-
ing is a fragile structure with a semifluid membrane.
One factor that may be responsible for the high
mechanical threshold is a special mechanosensitive
ion channel, the transient receptor potential subunit
V4 (TRPV4; Liedtke, W., 2005).

3.3 Nociceptors in Various Tissues

A complete overview of all tissues is not attempted.

Only those organs/tissues for which comprehensive
sets of data are available are addressed below.
CF VV LV

PN 3.3.1 Skin
The cutaneous nociceptors are generally subdivided
into several types according to the fibers supplying
them, namely A/- or C-fibers. Among the recep-
tors supplied by myelinated fibers are (data
from monkey, after Djouhri, L. and Lawson, S. N.,
Figure 5 So-called sensory terminal tree of a group III 2004):
fiber (schematic representation) as a possible
morphological basis for sensitization of polymodal
nociceptors. The summation of local receptive potentials
Type I A mechano-heat receptors (AMH, sup-
plied by A- and A-fibers). They have a relatively
created in perivascular (chemoreceptive) branches at
venous vessels (VV) or lymphatic vessels (LV) in
low mechanical, but a high thermal, threshold. They
combination with the local potentials of mechanoreceptive are called moderate pressure receptors by some.
branches in collagenous tissue (CF) may lead to a
suprathreshold excitation at the basis of the sensory
Type II A mechano-heat nociceptors (AMHs,
supplied by A-fibers). They have high mechanical
terminal tree within the perineurium (PN). (From and low thermal thresholds.
Messlinger, K., 1996, with permission.)
Among the receptors supplied by C-fibers, which are
a sensory neuron exhibit morphological differences. by far more frequent, are
Small clear vesicles are present at both terminals, but
at the peripheral terminal they show diversities in C-fiber mechano-heat nociceptors (CMHs),
which have little sensitivity to algesic agents
size, shape, and electron lucency, which are not pre-
sent at the central terminal. The differences in vesicle Cold nociceptors
content between the peripheral and the central term- Polymodal nociceptors that respond to all
noxious stimuli including algesic chemical stimuli
inal are difficult to explain, because if the vesicles are
synthesized in the soma, as is generally assumed, a In a cutaneous nerve of the cat, approximately 50% of
complicated sorting process is required. This is not all unmyelinated afferent fibers were found to have a
impossible, because at least for channel proteins an high mechanical threshold in the noxious range
exclusive transport to the peripheral terminal has (Bessou, P. and Perl, E. R., 1969). These units also
been reported (Garcia-Anoveros, J. et al., 2001). responded to noxious heat and irritant chemicals and
Another explanation would be that the small clear therefore were considered to be polymodal nociceptors.
vesicles in the distal terminal are synthesized in the The measured size of the RF of a single unmye-
ending itself, but at present this appears unlikely. As linated fiber supplying a polymodal nociceptor
the origin and the contents of the small clear vesicles depends on the thickness of the probe used for
in the peripheral terminal are unknown, the function mechanical stimulation: Thick probes yielded areas
of these vesicles is likewise obscure. of 632 mm2 (mean 18 mm2) in the rabbit, whereas
18 Anatomy of Nociceptors

with small probes mean areas of 7.5 mm2 were mea- extensive innervation. Sebaceous glands and arrector
sured, which consisted of several mechanosensitive pili muscles were not supplied with CGRP-ir fibers.
spots that could be activated independently from To find out if the effect of capsaicin treatment of
each other (Kenins, P., 1988). skin pain (e.g., erythralgia) is due to a desensitization
or destruction of nociceptive nerve fibers, Simone and
coworkers (Simone, D. A. et al., 1998) performed a light
3.3.1.1 Light microscopy microscopic study of capsaicin effects in skin biopsies
The nociceptive endings of the skin can extend in the of human skin. In the epidermal layer of intact skin,
epidermis to just beneath the surface. This explains mainly fibers ir for PGP 9.5 (protein gene product 9.5,
why superficial abrasions of the skin can cause pain a general marker for nerve fibers) were found, whereas
but no bleeding, because there are no blood vessels in in the dermis a dense innervation of CGRP- and SP-
the epidermis. ir fibers was present. After topical capsaicin treatment,
A study of the distribution of CGRP-ir fibers in the the CGRP- and PGP-ir fibers were almost comple-
rat glabrous skin (Kruger, L. et al., 1989) revealed a tely gone, but the SP-ir fibers were less affected.
dense network in the subpapillary layer largely con- Simultaneously, heat pain and pain produced by
fined to dermal blood vessels and sweat glands. The sharp objects were markedly reduced. This finding is
intraepidermal fibers branched extensively in the stra- surprising, because most SP-ir fibers are assumed to be
tum spinosum, but some extended nearly to the nociceptive (Lawson, S. N. et al., 1997).
stratum corneum (Figures 6 and 7). These fibers Recently, unmyelinated fibers stained for PGP 9.5
were considered to fulfill a noceffector role rather in the skin of human biopsy material have been studied
than a nociceptive one. Noceffector means that these using confocal microscopy (Kennedy, W. R., 2004). Of
fibers have an efferent function and are involved in the interest in the context of the present chapter are fibers
maintenance of the tissue under normal and patholo- that penetrated the basal lamina and branched in the
gical circumstances (Kruger, L., 1988). One aspect of epidermis, because they can be assumed to be free
such a noceffector role in the skin would be regulation nerve endings. The PGP 9.5-positive fibers in the
of Langerhans cell type expression and proliferation of epidermis had an appearance similar to that described
keratinocytes (Kruger, L. and Halata, Z., 1996). In in other reports: From a subepidermal plexus single
hairy skin of the rat, a similar arrangement of fibers coursed in the epidermis, where they could be
CGRP-ir fibers was found with the additional feature easily counted (Figure 8(a)). In skin biopsies of patients
that the lower portion of each hair follicle exhibited an with diabetic neuropathy, the density of the epidermal

(a) (b)

Figure 6 Digital glabrous skin. (a) A dermal (d) cutaneous nerve gives rise to an extensive calcitonin gene-related peptide-
immunoreactive (CGRP-ir) subpapillary nerve plexus (p) from which individual axons can be traced into epidermal pegs (e)
and dermal papillae (arrow). Some branches appear to end at varying depths within the stratum spinosum revealing terminal
expansions (arrowhead) (63). (b) Branched CGRP-ir axons in palmar glabrous skin extending from the dermis (d) nearly to
the stratum corneum (s.cor) (320). (From Kruger, L. et al., 1989, with permission.)
 Anatomy of Nociceptors 19

first identified functionally in electrophysiological

recordings and then examined morphologically.
Mechano-nociceptors had RFs consisting of several
sensitive spots with relatively insensitive areas in
between. After functional identification, the location
of the mechanosensitive spots was marked with fine
steel pins and the tissue processed for light and elec-
tron microscopy. The study focused on afferent units
conducting at 2030 m s1, i.e., on myelinated
mechano-nociceptors. Some of the thin myelinated
fibers approached the papillary layer of the corium
10 (dermis), lost their myelin sheath in this area but
Figure 7 A dermal calcitonin gene-related peptide- retained their Schwann cell layer, and penetrated
immunoreactive (CGRP-ir) axon terminating as a free nerve the epidermal basal layer. The penetration site was
ending with a bulbous enlargement (arrow) (320). (From considered to be the receptive portion of the ending;
Kruger, L. et al., 1989, with permission.) here the axons exhibited mitochondria, clear round
and dense-core vesicles. Most of the axons that pene-
branches was much less, and some fibers showed swel- trated into the epidermal layer still had a Schwann
lings that were interpreted as an indication of de- or cell covering (Figure 9), but in some cases the
regeneration (Figure 8(b)). Schwann cell sheath was lost and the basal lamina of
the axon fused with that of keratinocytes (Cauna, N.,
3.3.1.2 Electron microscopy 1966).
In a combined electrophysiologicalelectron micro- Nociceptive endings originating from A-fibers
scopic study on mechanical nociceptors in the cat skin have been reported to terminate in the basal layer
(Kruger, L. et al., 1981), single receptive endings were of the epidermis, i.e., they do not penetrate into the

(a)

(b)

Figure 8 (a) Normal human epidermal and papillary dermis innervation. Nerve fibers (green and yellow) course in bundles
through the dermis and branch into the papillary dermis to form the subepidermal neural plexus. Fibers from this plexus
penetrate the epidermaldermal basement membrane (red) to enter the epidermis (blue). Note that the basement membrane
surrounding capillaries also appears red and some nonneuronal fibroblasts appear green. Epidermal nerve fibers are abundant
and uniformly distributed. (b) Diabetic skin. Two tufts (arrows) arise from the subepidermal neural plexus (arrowhead). Nerve
fibers comprising the right tuft have many swellings. Irregular distribution characterized by tufting and clustering of nerve fibers
and the presence of swelling is distinctive of neuropathy. (From Kennedy, W. R., 2004, with permission.)
20 Anatomy of Nociceptors

Recordings from single corneal sensory fibers

demonstrated that the RFs of LTM endings were
K relatively large with a size of 50200 mm2 in the cat
(Tower, S., 1940). Most of the slowly conducting (A-
or C-) fibers of the cornea were found to be polymodal
a
v
and responded to mechanical, thermal, and chemical
m SC stimuli. They had a smaller RF size of 25 mm2.
Acidic solutions were used to test these units, and
BL the endings responded readily to a pH of 5 to 4.5
with a discharge that was nearly proportional to the
proton concentration. Capsaicin a specific ligand for
the TRPV1 receptor was also excitatory for slowly
12 conducting corneal afferents (Belmonte, C. et al., 1991).
Figure 9 Cat hairy skin. An intradermal axon (a) In contrast to most other body regions, the cornea
containing mitochondria (m) and clear vesicles (v) is not supplied by blood vessels. Therefore, after a
(arrowheads) enclosed by a Schwann cell (SC). The lesion has occurred, blood cells have no rapid access
basal lamina (BL) of the keratinocyte (K) is not shared by to the damaged area. In this case, lesion-induced sen-
the Schwann cell, which lacks a basal lamina only within
sitization of corneal nociceptors can occur through
the epidermis (40 000). (From Kruger, L. et al., 1981,
with permission.) activation of tissue cyclooxygenase and lipoxygen-
ase-generating prostaglandins (PGs) and other
sensitizing agents.
upper epidermal layer (Kruger, L. and Halata, Z., An interesting feature of corneal sensory endings is
1996). At present, these endings cannot be morpho- that the continuous shedding of epithelial cells on the
logically distinguished from cutaneous cold receptors surface of the cornea changes the morphology of the
that have the same location (Hensel, H. et al., 1974). nerve terminals. This results in a rearrangement of the
Du J. et al. (2003) reported that nociceptive arborization pattern of superficial terminals within 1
unmyelinated endings in the rat digital nerves exhibit week. Also after a relatively mild injury, a remodeling
N-methyl-D-aspartate (NMDA) receptor molecules of the corneal sensory ending takes place, and this
type 1 (NMDAR1) and can be excited by glutamate. remodeling is associated with an increased expression
These data support those of Svensson P. et al. (2003). of the growth-associated protein (GAP)-43 in the
The NMDAR1 were assumed to contribute to the corneal afferents (Belmonte, C. and Gallar, J., 1996).
sensitization of nociceptors in inflamed tissue, This protein is known to be involved in the develop-
because they showed an increased expression 7 days ment and regeneration of neurons; it is also present in
after induction of an experimental inflammation. growth cones of regenerating nerve fibers.

3.3.2.1 Light microscopy

Light microscopic investigations showed that corneal
3.3.2 Cornea
afferents were either thin myelinated or unmyelinated.
Hundred years ago, von Frey assumed that from the As in the skin, they formed a dense plexus in the stroma
cornea only pain can be evoked, but later several from which branches ascended through the basal
reports were published indicating that also sensation epithelial layer and terminated in bulb-like structures
of touch, warmth, and cold can be elicited. The just underneath the corneal surface. In the electrophy-
corneal epithelium is one of the tissues with the siological study mentioned above (MacIver, M. G. and
highest innervation density of the body and in this Tanelian, D. L., 1993) the fibers differed in their mor-
regard is comparable with the tooth pulp. phology: A-fibers looked like leashes and polymodal
In an electrophysiological study on fine nerve C-fiber terminals like stranded endings under
fibers of the cornea in vitro, A-fibers were described epifluorescence.
that were sensitive to mechanical and heat stimuli. As in free nerve endings of other tissues, neuro-
These units were situated in the basal epithelial peptides such as SP and CGRP are present in corneal
layer, whereas polymodal C-fiber endings were terminals. Interestingly, in the cornea SP has a
found in the superficial layers (MacIver, M. G. and trophic effect on the corneal epithelium. The healing
Tanelian, D. L., 1993). of a corneal wound is promoted by external
 Anatomy of Nociceptors 21

administration of SP. Conversely, the healing process (a) BC

is delayed if SP is missing in the corneal terminals,
e.g., after injection of capsaicin into the retrobulbar
space to deplete SP-containing fibers. Generally, SP
appears to promote mitosis in epithelial cells of the
cornea, while CGRP inhibits it. After corneal irrita-
tion or damage, signs of neurogenic inflammation
occur such as edema, conjunctivial vasodilatation, BM
M
and photophobia. These symptoms are attributed to
the release of neuropeptides from the sensory corneal
terminals (e.g., SP and CGRP).
1 m

3.3.2.2 Electron microscopy

Electron microscopic studies showed that thin fibers (b)
entering the epithelium lost their Schwann cell cov-
ering, often branched and exhibited varicosities that
contained mitochondria, neurofilaments and round BC
clear vesicles (Figures 10 and 11; Belmonte, C. and
Gallar, J., 1996; Muller, L. J. et al., 1996). Terminals
containing granular vesicles were numerous only at
the limbus of the cornea. These endings disappeared
after sympathectomy and, therefore, were considered
to be adrenergic terminals. Sensory endings (i.e.,
those having their somata in the trigeminal ganglion)
were often ir to SP and CGRP. Hoyes A. D. and 5 m
Barber P. (1976) distinguished between two types of
sensory free nerve endings that originated from (c)
unmyelinated corneal fibers:

M
BC

1 m

Figure 11 Nerve fibers in the epithelium of the human

cornea. (a) Cross section of a single nerve fiber at the level of
26 an axonal expansion (a bead), showing numerous
mitochondria (M), glycogen particles (large arrow), and
Figure 10 Flat mount preparation of cornea. The focal vesicles (small arrow) (16576). (b) Frontal section of a
plane is through the base of the corneal epithelium, which is nerve filament that bifurcates at the points indicated by the
heavily innervated by varicose calcitonin gene-related arrowheads. The single fiber running in the upper part of the
peptide-immunoreactive (CGRP-ir) axons coursing radially picture has a bead filled with mitochondria (large arrow)
inward from the limbus. Branches can be traced in more (3000). (c) Frontal section of a single fiber, showing two
superficial focal planes ascending perpendicularly between beads filled with numerous dark mitochondria (M),
epithelial cells to approach the epithelial surface (200). glycogen particles (small arrows), and vesicles (large
(From Silverman, J. D. and Kruger, L., 1988, with arrow) (12544). BC, basal cell; BM, Bowmans membrane.
permission.) (From Muller, L. J. et al., 1996, with permission.)
22 Anatomy of Nociceptors

(1) Units that contained many mitochondria and Besides capsaicin, ATP (binding to the purinergic
were considered to be mechano- or thermo- P2X3 receptor), NGF, and protons (acting through
receptors. TRPV1 and acid-sensing ion channels (ASICs)) are
(2) Units exhibiting clear and dense-core vesicles. effective stimulants for muscle nociceptors
These units were assumed to be nociceptive (Hoheisel, U. et al., 2004; 2005). Similar to nociceptive
afferents; they were concentrated in the central afferents from the skin, afferent fibers from nocicep-
parts of the cornea where no blood vessels are tors in rat muscle are equipped with TTX-resistant
present. The majority of this fiber type had an (TTX-r) Na channels (Steffens, H. et al., 2003).
intraepithelial location.
3.3.3.2.(i) Light microscopy In a quantitative eva-
Attempts to correlate morphology with function in luation of neuropeptide-ir free nerve endings and
corneal receptive endings did not yield unequivocal preterminal axons (both characterized by varicosities)
results. Therefore, in their review, Belmonte C. and in the gastrocnemiussoleus muscle of the rat, most
Gallar J. (1996) stated that no morphological specia- endings were found around small blood vessels (arter-
lization has been found in corneal nerve terminals ioles or venules), whereas capillaries and the muscle
associated with the various functional classes of cor- cells were not supplied by these endings. Most numer-
neal afferents. ous were the CGRP-ir endings followed by endings
with SP-immunoreactivity (IR), VIP-IR, NGF-IR,
and GAP-43-IR (Reinert, A. et al., 1998). Many end-
ings exhibited IR for more than one peptide, e.g., for
3.3.3 Deep Somatic Tissues
SP and CGRP or SP and VIP (Figure 12).
3.3.3.1 Muscle pain compared After 12 days of an experimental myositis, the inner-
to cutaneous pain vation density of the muscle with neuropeptide-ir free
Muscle pain is less well localized than cutaneous pain. nerve endings was significantly increased. The effect
One reason for this difference may be the fact that was particularly marked for endings with SP-IR, GAP-
muscle pain but not cutaneous pain is often 43-IR, and NGF-IR (Reinert, A. et al., 1998; Figure 13
referred to sites distant from the lesion. Another pos- (b and c)). The density of the SP-ir fibers doubled in
sible reason is the lower innervation density of muscle inflamed muscle. Of course the question arose if this
tissue. Direct quantitative comparisons between the increase was due to inflammation-induced sprouting of
innervation density of muscle and skin have not been the nerve fibers or due to an increase in the neuropep-
published so far and are difficult to make, because tide content of the individual fiber so that a higher
muscle tissue is 3D and skin largely 2D. proportion of fibers were above the detection threshold
Free nerve endings in muscle are partly nocicep- in the fluorescence microscope. The finding that the
tive and partly nonnociceptive. The latter ones have a density of NGF-ir and GAP-43-ir fibers increased
low mechanical threshold and are assumed to mediate together with that of SP-ir endings was interpreted
pressure and tension sensations from muscle tissue. that sprouting had occurred, because NGF and GAP-
Another important function for these nonnociceptive 43 are strongly expressed in growth cones.
endings in muscle is to control the adjustment of Free nerve endings in the calcaneal tendon of the
circulation and respiration to the requirements of rat showed IR to the same neuropeptides, but the
muscle work. They are activated by physiological distribution of the fibers was different from muscle.
degrees of exercise and send their information to the In the epitendineum, the fibers formed a much den-
circulatory and respiratory centers in the medulla ser network than in muscle, but the collagen fiber
(McCloskey, D. I. and Mitchell, J. H., 1972). bundles in the center of the tendon were largely free
from free nerve endings (Reinert, A. and Mense, S.
3.3.3.2 Nociceptors in muscle and tendon unpublished data). This finding may explain why
Several studies on the response properties of unmye- chronic mechanical stress to the epitendineum can
linated afferent fibers from cat and dog muscle be very painful, whereas ruptures of collagen bundles
demonstrated that a subpopulation of these afferents within the tendon may occur without pain.
are excited by stimuli (strong mechanical and algesic
chemical) that are known to elicit pain (Kumazawa, T. 3.3.3.2.(ii) Electron microscopy
and Mizumura, K., 1977; Mense, S., 1977; Mense, S. 3.3.3.2.(ii).(a) Muscle The first comprehensive
and Meyer, H., 1985). report on the morphology of free nerve endings
 Anatomy of Nociceptors 23

(a) (b) i

(c) (d)

(e) (f)

(g) (h)

Figure 12 (a and b) Sections of intact (a) and persistently inflamed (b) muscle. Scale bar 100 mm. The insert (i) in (b) shows
lymphoid cells between vacuoles at a higher magnification. Scale bar 50 mm. (cf ) Neuropeptide-ir nerve fibers in muscle.
(c) Varicose SP-ir fibers (arrows) in the adventitia of an artery (arrowheads). (d) The fibers shown in (c) double-labeled for
calcitonin gene-related peptide (CGRP). (e) Perivascular SP-ir fiber that is also ir for vasoactive intestinal polypeptide (VIP; (f )).
(g) Perivascular fiber in inflamed muscle exhibiting immunoreactivity for NGF (arrow, same magnification as in (h)). (h) Growth-
associated protein 43 (GAP 43)-ir fiber (arrow) in the wall of an artery (arrowheads). Scale bar 25 mm (ch). (From Reinert, A.
et al., 1998, with permission.)
24 Anatomy of Nociceptors

(a) SP-ir fibers (b) GAP 43-ir fibers

Fibers per cm2 (Mean + S.E.M.)

20
*
12
Intact ** 15

Acute infla. (10 h)

10 10
Fibers per cm2 (Mean + S.E.M.)

Persistent
infla. (12 days) 5
** *
8
0

6
(c) NGF-ir fibers

Fibers per cm2 (Mean + S.E.M.)

25
4 **
20
* 15
2
10

0 5
Muscle Conn. Arteries Arterioles Capillaries Veins All
fibres tissue (Venules) 0

Figure 13 Innervation density of substance P-immunoreactive (SP-ir), growth-associated protein 43 (GAP 43)-ir, and nerve
growth factor (NGF)-ir fibers in intact and inflamed muscle. Acute inflammation (yellow bars), duration 10 h; persistent
inflammation (red bars), duration 12 days. (a) Distribution of SP-ir fibers in the components of muscle tissue. Note that no
fibers were found close to muscle fibers and capillaries. Asterisks mark the level of significance compared to intact muscle.

p < 0.05; p < 0.01; p < 0.001, U test. (Modified after Reinert, A. et al., 1998.)

in skeletal muscle of the cat was published by Stacey precapillary segment of a blood vessel in the soleus
M. J. (1969). He used the silver impregnation techni- muscle. The ending contained clear and dense-core
que in sympathectomized animals and focused on vesicles and had exposed membrane areas. The authors
endings supplied by group III and IV fibers. The suggested that those terminals (mainly originating from
majority of the latter had a diameter of 0.35 mm; the group III fibers) that had a close association with con-
unmyelinated afferents outnumbered the myelinated nective tissue may have a mechanoreceptive function
ones by a factor of 2. A prominent location of free mediating stretch or pressure, whereas endings that
nerve endings supplied by group IV fibers was the lacked this feature but had a spatial relation to mast
adventitia of arterioles and venules. Myelinated cells were nociceptors.
(group II and III) afferents generated not only free
nerve endings but also paciniform corpuscles, 3.3.3.2.(ii).(b) Tendon The electron microscopic
whereas unmyelinated fibers terminated exclusively reconstruction of free endings in the calcaneal tendon
in free nerve endings. of the cat yielded various morphological types of free
In another study on sympathectomized cats, von nerve endings connected to group III and IV afferent
During M. and Andres K. H. (1990) reported that the fibers (Andres, K. H. et al., 1985). Based on morpho-
predominant location of group III and IV endings was logical criteria or the location in the tissue, the
in the perimysium surrounding larger or smaller bun- authors distinguished five types of free nerve endings
dles of muscle fibers. Other locations were the supplied by group III fibers: Type 1 terminated in
adventitia of arterioles, venules, and lymphatic vessels venous vessels (VV) (called the lanceolate type); it
(LV), and finally the endoneurium of nerve fiber bun- was special in that it had a flattened profile in cross
dles. The latter terminals were assumed to belong to sections and possessed exposed receptive areas on its
nervi nervorum. In muscle, the terminals of group III edges. Type 2 ended in the wall of LV. Types 3 and 4
fibers were generally larger than those of group IV supplied the connective tissue around blood vessels,
fibers, and they contained more mitochondria and a and one of these types had contacts to collagen fiber
more distinct receptor matrix. Figure 14 shows a group bundles. The collagen bundles were assumed to
IV terminal in the endomysium around the transfer mechanical forces to the receptive ending
 Anatomy of Nociceptors 25

(a) Schaible, H.-G. and Schmidt, R. F., 1985; Schaible,

H.-G. and Grubb, B. D., 1993; Schaible, H.-G., 2006).
c
In inflamed joint, initially mechanoinsensitive afferents
(silent nociceptors) are sensitized and may become
enm mechanosensitive (Schaible, H.-G. and Schmidt, R. F.,
1988).
pc
3.3.3.3.(ii) Ultrastructure of articular free nerve
endings In a thorough investigation of articular end-
10 m
ings, Heppelmann B. et al. (1990a) studied free nerve
endings from the knee joint of the cat in sympathecto-
mized animals. Figure 17 shows the reconstruction of
(b) group III and IV nerve endings from the joint capsule of
pc the cats knee joint. As in other tissues, free nerve end-
ings in joint were ensheathed by Schwann cells, and
only some sites were not covered. Usually, these
SC
exposed areas were located in the varicosities of the
ending (the beads) and the end bulb, here the axon,
often directly abutted the basal lamina. The exposed
membrane areas were assumed to be the receptive sites
of the fibers; they exhibited structures that are often
1 m associated with receptive sites: aggregates of mito-
chondria, glycogen particles, vesicles in the
Figure 14 Nerve terminal of a group IV fiber in the soleus axoplasm, and an electron-dense filamentous struc-
muscle of the cat. Rectangle in (a) represents the area of the
ture of the axoplasm (the receptor matrix). The end
receptor shown in (b). The terminal axon contains numerous
dense-core and clear vesicles. pc, precapillary segment; c, bulb, i.e., the most distal beaded end of the fiber
capillary; SC, Schwann cell. Arrows indicate exposed (Figure 18 shows the end bulb of one branch of the
axonal membrane areas. (From von During, M. and Andres, group III unit reconstructed in Figure 17), had gen-
K. H., 1990, with permission.) erally the same structure as the axonal beads, except
that the mitochondria showed a special arrangement:
and, therefore, the endings were viewed as mechan- the organelles were not oriented in parallel as in the
oreceptors. The fifth type of group III ending axonal beads but pointed with their axis in all possi-
innervated the endoneural connective tissues of ble directions.
small nerve fiber bundles (Figure 15). In the cats knee joint capsule, free nerve endings
In the same study, two types of free nerve endings generated by group III fibers were smaller in axon
of group IV fibers were described; both were located diameter and had shorter branches (up to 200 mm)
in the connective tissue of blood vessels, and some of than those supplied by group IV fibers (more than
the fibers contained granulated vesicles. The term- 300 mm). The main difference between group III and
inals showed penicillate formations (Figure 16). IV endings was that the latter never had a neurofila-
ment core in their axoplasm, which was present in all
axons and axonal beads of group III fibers (but not in
3.3.3.3 Nociceptors in joints the end bulbs). The functional significance of the
and ligaments neurofilament core is obscure; one possible function
3.3.3.3.(i) Pain from joints In humans, experimen- is mechanical stabilization of the group III endings
tal joint pain has been evoked by administration of that usually have a more unfavorable quantitative
noxious mechanical and chemical stimuli to the articular relation between the fluid axoplasm and the more
capsule and ligaments. No pain is elicited by stimulation rigid axolemma.
of cartilage, because cartilage is not innervated. Joint The authors of these and similar data
inflammation is characterized by hyperalgesia and (Heppelmann, B. et al., 1990a; 1990b; Messlinger, K.,
persistent pain at rest. The discharge properties of 1996) put forward the concept that both group III and
joint nociceptors largely reflect these subjective phe- IV fibers form sensory terminal trees that consist of
nomena of joint pain (Guilbaud, G. et al., 1985; the preterminal axon and its branches (Figure 5). The
26 Anatomy of Nociceptors

av a2 T lll VV
a1 sm

a3
200 m

SC

VV

T lll PTgc
cfb
T lll EN
pn

SC

SC
av

VV

Figure 15 Group III fiber terminals in cat calcaneal tendon. Reconstruction of a series of sections over a length of 1 mm
showing three different terminals of group III afferents. A fascicle of arterial vessels (av) and venous vessels (VV) with some
arterio-venous anastomoses is accompanied by small nerve fiber bundles of which the myelinated axons (a1, a2, a3) are
reconstructed. The schematic drawings show segments of three different terminals at a higher magnification (T III VV, T III
PTgc, and T III EN) with their typical relations to the surrounding tissue structures. Axon membrane segments without any
Schwann cell (SC) covering are marked by higher contrast. The terminal of the a1 axon is the T III VV type with two
lanceolate terminals in the adventitia of the vessel. The arrows indicate the free edges of the terminal. Smooth muscle cell
(sm). The terminal of the a2 axon is type T III PTgc in the connective tissue of the vesselnervefiber fascicle. The terminals
are oriented to collagen fiber bundles (cfb). The terminal of the a3 axon is the T III EN type. The terminals ramify within the
endoneural space. Another small myelinated axon passes the terminals. Perineural sheath (pn). The length of the receptive
areas of each myelinated axon is indicated in the reconstruction by arrows. (From Andres, K. H. et al., 1985, with
permission.)

terminal tree is not covered by myelin (in group III axonal beads may not only serve as receptive sites
endings) and lacks a perineurial sheath (in both group but also as amplifiers of the receptor potential.
III and IV endings); it is assumed to form the sensory The group IV endings terminating in the medial
ending proper. collateral ligament (Figure 19) exhibited the same fea-
Considerations on the space constant of free nerve tures as the terminals in the joint capsule: They had
endings prompted Heppelmann B. et al. (1990a; 1990b) exposed membrane areas and contained mitochondria,
to hypothesize that the distance from the end bulb as glycogen particles, and vesicles (Heppelmann, B. et al.,
the main receptive region to the regenerative region 1990b; Messlinger, K., 1996).
where the action potential originates may be dan- Group III and IV fibers of the joint terminate as
gerously great for a safe transformation of the receptor free nerve endings in the fibrous capsule, adipose
potential into an action potential. Therefore, the tissue, ligaments, menisci, and periosteum. Staining
 Anatomy of Nociceptors 27

1
VV

A
LV Fig . 7
SC SC

T IV A

pn

Fig . 2
B Fig . 3

pn

SC
SC
Fig . 4
T IV

A A

Fig . 8
m z z
y x y x

Figure 16 Group IV fiber terminals (T IV) in cat calcaneal Group lll Group lV
tendon. Schematic drawing of group IV fibers and their Figure 17 Sensory endings in the knee joint capsule of
sensory regions in the surrounding connective tissue of the cat. Reconstructions based on ultrathin sections at 1-
venous (VV) and lymphatic (LV) vessels. Perineural sheath mm intervals. Areas of bare axolemma without Schwann cell
(pn). The various parts of the scheme are not to scale. The covering are marked by dense stippling. A, sensory axon;
terminal bundles end in penicillate formations. (From SC, Schwann cell; arrows, thin segments between axonal
Andres, K. H. et al., 1985, with permission.) beads (6000). x, y, z bars, 1 mm. Left: group III fiber, length
of reconstructed fiber 70 mm. Right: group IV fiber. The
axonal beads are shorter and the thin segments longer than
for nerve fibers and neuropeptides also demonstrated in group III fibers. Areas of bare axolemma are interrupted
endings in the synovial layer. Interestingly, a few by axon segments with complete Schwann cell wrapping.
(From Heppelmann, B. et al., 1990b, with permission.)
group IV endings terminated in the perineurium
(PN) of the articular nerve. Assuming that these end-
ings are nociceptors, they might be responsible for 3.3.3.4 Tooth pulp
neuropathic pain originating in the joint. 3.3.3.4.(i) Toothache In psychophysiological stu-
The major neuropeptides in small-diameter joint dies, electrical stimulation of the tooth pulp has been
afferents were SP, CGRP, and somatostatin (SOM). reported to elicit pain exclusively (Andersson, S. A.
Neurokinin (NK)-A, galanin, enkephalins, and et al., 1973). Therefore, the free nerve endings in the
neuropeptide Y have also been localized to joint pulp were assumed to be either nociceptors or sym-
afferents (Schaible, H.-G. and Grubb, B. D., 1993; pathetic postganglionic fibers. Rats show aversive
Schaible, H.-G., 2006). behavior when capsaicin is applied to the exposed
28 Anatomy of Nociceptors

(1) The great majority (5679.6% in the cat mandib-

ular canine tooth; Holland, G. R. and Robinson,
P. P., 1983) of the fibers supplying the tooth pulp
are unmyelinated with a mean diameter of 0.4 mm
in man (Koling, A., 1985); they often branch and
form a dense subodontoblastic plexus (the per-
ipheral plexus of Raschkow, see Byers, M. R.,
1984, for review) in the apex of the tooth.
Between the plexus and the odontoblast layer,
there is the cell-free zone of Weil. Figure 20
shows an overview of the afferent fibers in the
Figure 18 Axonal end bulb of an articular group III fiber in
root and coronal pulp of a rat molar. The data
the cat. In addition to mitochondria and glycogen particles, were obtained in experiments in which the
the end bulb contains several clear vesicles (arrow) and trigeminal ganglion was injected with an antero-
larger dense-core vesicles (arrowhead). In contrast to more gradely transported marker (WGA-HRP) to
proximal axonal beads, microtubules and a neurofilament visualize afferent fibers only (Ibuki, T. et al.,
core are missing in the end bulb. (From Heppelmann, B.
et al., 1990a, with permission.)
1996). In the subodontoblastic region, axo-axonal
appositions and appositions between axons and
odontoblasts have been described by some
dentine of the incisors (Chidiac, J. J. et al., 2002; Rifai, authors, but others could not support this finding.
K. et al., 2004). The latter observation indicates that The other pulpal afferents conduct in the A-
nociceptive afferents of the tooth pulp are equipped range; A-fibers are sparse.
with the TRPV1 receptor for which capsaicin is a (2) Many axons are incompletely ensheathed by
specific ligand. Schwann cells (Figure 21); they sometimes
Functional studies of the receptive properties of leave the Schwann cell sheath completely to
single unmyelinated tooth pulp afferents are scarce. terminate as naked axons in the extracellular
The published data show that slowly conducting space. In those fibers that do have a sheath, the
afferent units from cat are sensitive to stimuli that Schwann cells often lack a basal lamina
are known to elicit pain in humans (hot and cold, (Figure 22; Byers, M. R. et al., 1982). The majority
local administration of bradykinin (BK), and mechan- of the afferent fibers form beaded free nerve end-
ical stimulation) ( Jyvasjarvi, E. and Kniffki, K. D., ings in odontoblast layer, predentin, and in
1992). Many of the recorded units responded to dentinal tubules, many of which are innervated.
more than one stimulus and thus behaved like poly- The intradentinal endings extend only 100
modal nociceptors. In another study, the sensitivity 200 mm into the dentin, and sometimes more
to capsaicin of A-, A-, and C-fibers was compared than one axon accompanies an odontoblast pro-
in cat tooth pulp afferents. Not only C-fiber endings cess (Figure 23). Gunji T. (1982) put forward the
were excited but also A-units, albeit at a smaller concept that the odontoblast processes and the
proportion. None of the A-endings responded to axons in the dentinal tubules form a mechanor-
the stimulant (Ikeda, H. et al., 1997). eceptive complex. In contrast to the intradentinal
No studies combining electrophysiological and axons, the odontoblast processes reach the den-
morphological techniques were conducted on teeth. tinenamel border and could be deformed by
Therefore, in morphological investigations a possible mechanical stimuli. The mechanical movements
nociceptive function of the free nerve endings can be or deformations of the processes may then excite
discussed, but is not sure. the sensory axons that do not penetrate that far
into the dentinal tubules.
3.3.3.4.(ii) Light and electron microscopy Data are (3) The varicosities of the free nerve endings exhibited
available from many species including man. The mor- specializations known from nociceptors in other
phology of the free nerve endings is similar to that tissue such as mitochondria, small clear and large
described above for other tissues in more detail. In the dense-core vesicles, smooth endoplasmic reticu-
following paragraph, possible differences between lum, and few microtubules and neurofilaments
nerve fibers in mature and continuously growing (Figure 24; Byers, M. R., 1984; Ibuki, T. et al.,
teeth (e.g., mice incisors) are not considered. 1996). Ibuki T. et al. (1996) distinguished between
 Anatomy of Nociceptors 29

(a)

VV

C
N

10 m

(b) (c)

F2 F2 F1
F1

Figure 19 Group IV fiber endings at a venous vessel (VV) in the medial collateral ligament of the cat. (a) Reconstruction
based on serial semi- and ultrathin sections. Besides a small peripheral nerve (N), two terminal nerve fiber bundles without
perineurial sheath that enclose two group IV fibers (left) and one single fiber (right) run along the VV. (b and c) Cross sections of
the nerve fiber bundle with two group IV fibers as marked in Figure 3(a) at high magnification. Drawings from original electron
micrographs. Scale bar 1 mm. (b) Cross section showing a bulge of one of the nerve fibers (Fl) with bare areas of the
axolemma (arrows), mitochondria, glycogen particles, and vesicles. The other nerve fiber (F2) is completely ensheathed by
the Schwann cell. C, cross section 40 mm distal from that of Figure 3(b) showing the final thickening of the nerve fiber Fl with
bare area of axolemma (arrow). The second nerve fiber (F2) continues. (From Heppelmann, B. et al., 1990b, with permission.)

two types of varicosity: one contained many mito- endings have a lower probability of being excited,
chondria, while the other was characterized by the and therefore need less mitochondria.
presence of large dense-core and small clear vesi- (4) Besides a nociceptive function many of the small-
cles. Generally, a rich supply with mitochondria caliber tooth afferents may have a trophic influ-
and glycogen particles is interpreted as an indica- ence on blood vessels in the pulp or on
tion of high energy demands and, therefore is likely odontoblasts. They may be necessary for repair
to occur in endings with a low mechanical thresh- or normal growth, as in continuously growing
old and high firing probability, while nociceptive teeth. The finding that the anterograde marker
30 Anatomy of Nociceptors

(a) (b)

D
BV

O
S

BV
0.2 mm

50 m

Figure 20 Innervation of root pulp (a) and coronal pulp (b) in a rat tooth. The fibers were labeled anterogradely from the
trigeminal ganglion. (a) Bundles of labeled fibers (arrows) run along the blood vessels (BV) of the root pulp. Some fibers
surround the blood vessels (arrowheads). (b) The fibers form a subodontoblast plexus (S) and penetrate the dentin (arrows). D,
dentin; O, odontoblast. (From Ibuki, T. et al., 1996, with permission.)

(a) D (b)
S

S
BV

5 m 2 m

Figure 21 (a) Fibers labeled anterogradely with horseradish peroxidase in the subodontoblastic region of the tooth. BV,
blood vessel; D, dentin; O, odontoblast. (b) High magnification of the area marked by an arrow in (a). Horseradish peroxidase
reaction product is present in the myelinated A-axon (large arrow) and unmyelinated axons (small arrow). The unmyelinated
axon has partially lost its Schwann cell sheath (arrowheads). S, Schwann cell. (From Ibuki, T. et al., 1996, with permission.)

(WGA-HRP) injected into the trigeminal gang- was assumed to reflect a release process that
lion by Ibuki T. et al. (1996) was found not only normally supplies the odontoblasts with trophic
within the axons but also outside between the substances. This hypothesis resembles the nocef-
axons and the odontoblast processes (Figure 24) fector concept of Kruger.
 Anatomy of Nociceptors 31

5 6
P
A

P A

SC

A A
SC

Figure 22 Nerve fibers in the cell-free zone of the tooth pulp. Unmyelinated axons (A) were identified in this zone when
surrounded by a Schwann cell (SC), but no basal lamina accompanied the Schwann cell. Other cell processes (P) could be
axons or fibroblasts. Scale bars, 0.5 mm (42300). (From Byers, M. R. et al., 1982, with permission.)

(a)

A
A A
1.0 m

(b)

A
A

Figure 23 Axons in dentinal tubules. (a) A cluster of adjacent dentinal tubules at the predentindentin border. Each tubule
contains an axon (A) and an odontoblast process () (10350). (b) Two adjacent axons in the same dentinal tubule. (From
Byers, M. R. et al., 1982, with permission.)

3.3.3.4.(iii) Neuropeptide content and equipment were thought to have a noceffector function
with receptor molecules Sensory dental axons (Kruger, L., 1988).
contain many neuropeptides (SP, CGRP, VIP, enke- SP-ir fibers predominated in the center of the
phalin), the most interesting one being SP because of tooth pulp. The expression of SP in carious human
its relation to a nociceptive function. A particularly teeth has been found to be increased, and the increase
rich innervation with CGRP-ir fibers was found in was larger in symptomatic than in asymptomatic
the subodontoblast layer and the dentinal tubules teeth. This finding parallels the results obtained
(Kruger, L. and Halata, Z., 1996). These endings from inflamed muscle (see above) and suggests that
32 Anatomy of Nociceptors

development, differentiation, or regeneration of the

tooth pulp innervation (Foster, E. et al., 1995).
However, not all free nerve endings can be con-
LV sidered to be nociceptive; in the tooth pulp many
endings appear to supply blood vessels (Zhang, J. Q.
et al., 1998). The vascular innervation with neuro-
peptide-ir nerve endings is largely restricted to
SV arterioles; capillaries and lymphatics were mostly
devoid of an innervation. This distribution pattern
resembles that of skeletal muscle. Substances
involved in the vascular and cellular regulation of
the tooth pulp in the rat are SP, NK-A, and CGRP,
which are thought to be released from nerve endings.
This function was inferred from the finding that
pulpal hard tissue cells, blood vessels, and fibroblasts
were equipped with NK-1 and NK-2 receptors (for
OP
SP and NK-A, respectively) as well as CGRP1
D receptors (Fristad, I. et al., 2003).

3.3.3.5 Dura mater encephali

3.3.3.5.(i) Headache Generally, those types of
headache that are associated with vascular changes
in the meninges (e.g., migraine) are being attributed
0.5 m to the activation of para- or perivascular nociceptive
endings by agents released from the nerve terminals.
Figure 24 Anterogradely labeled sensory varicosity
Among these substances are SP, CGRP, and NK-1.
adjacent to an odontoblast process (OP) in dentin. The
varicosity is full of large dense-core vesicles (LV) and small Observations supporting this assumption are that
clear vesicles (SV). Some vesicles (arrowheads) contain the when tested in awake subjects meninges are most
horseradish peroxidase reaction product used for painful close to blood vessels, and that migraine pain
anterograde labeling. The reaction product is also present in is most intense during the phase of vasodilatation and
the extracellular space (arrow). D, dentin. (From Ibuki, T.
increase of vascular permeability.
et al., 1996, with permission.)

3.3.3.5.(ii) Light and electron microscopy In

their light and electron microscopic study on recep-
SP plays an important role in the mediation of tooth- tive endings in the dura mater of the rat, Andres K. H.
ache (Rodd, H. D. and Boissonade, F. M., 2000). In et al. (1987) described terminals of myelinated and
one patient with hereditary sensory and autonomic unmyelinated fibers. The tissue around postcapillary
neuropathy, CGRP- and SP-ir fibers were comple- blood vessels and the superior sagittal sinus exhibited
tely missing, which could explain the sensory deficit a particularly high innervation density of endings.
associated with this condition (Rodd, H. D. et al., Contrary to former belief, the dura also contained
1998). The lack of CGRP- and SP-ir fibers was LV (Figure 25). These vessels were innervated
accompanied by an upregulation of neuropeptide Y. mainly by endings from unmyelinated fibers.
The nerve endings in the tooth pulp have been Some of the myelinated fibers terminated in
shown to be equipped with receptor molecules that Ruffini-like nerve endings, while most of the myeli-
are thought to be associated with a nociceptive func- nated and all of the unmyelinated fibers terminated
tion, e.g., P2X3, TRPV1 (Renton, T. et al., 2003), and in free nerve endings. The Ruffini-like receptors
the vanilloid receptor-like protein, VRL1. In rat were located close to collagen fiber bundles of the
molar tooth pulp, fibers ir to trkB (the receptor for wall of the sagittal sinus and of other blood and LV.
BDNF and NT-4) were found. These fibers included They were viewed as stretch receptors.
both unmyelinated and myelinated units. This and Among the free nerve endings formed by unmye-
other findings were interpreted as indicating that linated fibers, one type was located very close (at a
BDNF and/or NT-4 are involved in the distance of <1 mm) to the endothelial cells of the
 Anatomy of Nociceptors 33

EV F P
B
MV
NFB
NFB
MA SS
LV

MA CO

SAS

LV
CO
NFB

Figure 25 Vascular bed of the dura underneath the

parietal bone (B) and dorsal to the sagittal sinus (SS).
Arrows, entrance of the superior cerebral veins into the
sagittal sinus; MA, branches of meningeal artery; MV,
branches of meningeal vein; EV, emissary vein; LV,
lymphatic vessels; NFB, nerve fiber bundles exhibiting a
terminal plexus; CO, cortex; SAS, subarachnoidal space.
(From Andres, K. H. et al., 1987, with permission.)

postcapillary venules. It had the typical appearance of

free nerve endings in other tissues with exposed mem-
brane areas, mitochondria, vesicles, and a receptor
matrix. The endings that were located close to the
wall of blood vessels would be in a good position to
monitor the composition of the interstitial fluid in the
vicinity of the vessels (Figure 26). They could well
perform nociceptive functions, for instance in cases of
headache when sensitizing and pain-producing sub-
stances are released from the blood plasma and
afferent nerve fibers.
Many of the nerve fibers accompanying blood ves-
sels contained CGRP or SP (Figure 27; Hanesch, U., F
1996). Like in other tissues, SP-ir fibers were less
numerous than CGRP-ir ones. While SP-IR was
located inside the axoplasm, the SP receptor NK-1
was found on the outer membrane of the Schwann
cells surrounding the axons. Figure 26 Nerve terminations in the dura at the wall of a
venous vessel. Axon profiles with vesicles and receptor
matrix are indicated (arrows); the other axons exhibit
3.3.4 Visceral Organs cytoskeletal elements (45000). Scale bar 1 mm. The gap
between the terminal and the postcapillary venule with
3.3.4.1 Visceral pain endothelial fenestration is <1 mm. F, process of fibrocyte; P,
The mechanisms of visceral pain are still poorly pericyte. (From Andres, K. H. et al., 1987, with permission.)
understood. Apparently, some inner organs are
equipped with specific nociceptors (HTM and poly-
modal ones; Janig W. (1996), for review), but their in the pattern of discharge in populations of rela-
existence has not been proven for all viscera. It is tively unspecific visceral afferents. The clinical
conceivable that the pain from some viscera is observation that a small cut into the gut is not painful,
mediated not by specific nociceptors but is encoded but spasm or ischemia of a larger area causes strong
34 Anatomy of Nociceptors

(a) (b)

MMA
MMA

Figure 27 Neuropeptide-ir nerve fibers in the dura mater of the rat. (a) CGRP-ir fibers accompany and cross the middle
meningeal artery (MMA). Small bundles of fibers branch off the main bundle and terminate in the connective tissue thereby
forming tree-like endings (arrowheads). Some fibers end at the artery (arrows). (b) SP-ir fibers accompanying the MMA. Single
beaded nerve fibers terminate at the vessel wall (arrows) or in the connective tissue between blood vessels (arrowheads).
(From Hanesch, U., 1996, with permission.)

pain, is generally attributed to the low innervation this arrangement is special for testicular nociceptors,
density of the inner organs. or if it had been overlooked in other tissues.
In the distal tip of the axon, the axolemma exhibited
a greater number of exposed areas that directly con-
3.3.4.2 Testis tacted the basal lamina. The authors stress the presence
Electrophysiological studies by Kumazawa and cow- of clusters of small spherical vesicles, aggregates of
orkers (Kumazawa, T. and Mizumura, K., 1980; mitochondria, and axonal reticulum in a granular matrix
Kumazawa, T. et al., 1987) have shown that in the in this part of the axon that apparently forms the recep-
tissue layers covering the seminiferous tubules in the tive terminal (Figure 30). Interspersed between the
rat, endings with unmyelinated afferent fibers are pre- spherical vesicles were so-called granular vesicles that
sent that behave like polymodal nociceptors in that differed somewhat from the dense-core vesicles of end-
they can be activated by mechanical stimuli, pain- ings containing catecholamines and, therefore, could be
producing substances, and heat. The innervation den- distinguished from sympathetic nerve endings.
sity of the tissue is low, and the receptive endings are The functional interpretation of the ultrastructure
located directly underneath the tunica albuginea. of the terminal poses some problems: Even though it
These properties made it possible to relate the is characterized by special features (vesicle clusters,
mechanosensitive spots found in electrophysiological mitochondria, and exposed axolemma), the axonal
experiments to structures in histological specimen. zones close to the terminal likewise exhibited some
Kruger L. and coworkers (2003b) used this of these specializations. Therefore, it is difficult to
approach to elucidate the ultrastructure of nociceptive decide if only the ultimate terminal is the receptive
endings of the testis. In the electrophysiological site where the electrical receptor potential originates
experiments, the location of the receptive ending was or if also other parts of the preterminal branches can
marked with insect pins. The endings were recon- be viewed as receptive, as in the sensory tree concept.
structed from serial electron microscopic sections An important question in this context is where the
(Figure 28), and in general they showed the features generator portion of the nociceptor axon is located
described above for putative nociceptors in other tis- and if it exhibits structural specializations different
sues. The preterminal axon had expansions from those of the receptive terminal.
(varicosities) with axonal constrictions in between.
Approaching the terminal proper, the axons often
lost their microtubules and neurofilaments. Instead, a
network of axonal reticulum (probably endoplasmic 3.4 Neuropeptide Content
reticulum) appeared that was embedded in a granular of Nociceptors
matrix (similar to the receptor matrix of Andres, K. H.
3.4.1 General Remarks
and von During, M., 1973). This arrangement of an
axonal reticulum in a granular matrix (Figure 29) has In many cells, neurotransmitters coexist with
not been described by other authors, but it is unclear if neuropeptides, or one neuron contains more than
 Anatomy of Nociceptors 35

A
B d

C
e

D
b
E
F

h G
H
g
i

l
k
J
K

Figure 28 Three-dimensional reconstruction of a pin-marked receptive field (RF) identified from a physiologically
characterized nociceptor polymodal fiber discharge in an in vitro canine testicular preparation based on a series of semithin
(1 mm) sections from which ultrathin sections were obtained in each of the sectors, labeled AK. In the segments indicated in
lower case, axons are shown in red and Schwann cell basal lamina boundaries in white. Scale bar 1 mm. (From Kruger, L.
et al., 2003b, with permission.)

one neuropeptide. It is well known that a large

proportion of SP-expressing DRG neurons also
contain glutamate. Another example is that almost
all DRG cells that are ir for CGRP also exhibit IR
for SP and other neuropeptides (Ju, G. et al., 1987).
S It is generally assumed that glutamate acts as the
main neurotransmitter in nociceptive afferents,
m
whereas the neuropeptides function as neuromodu-
lators that with few exceptions enhance the
central nervous effects of peripheral noxious sti-
muli (Hokfelt, T. et al., 1980; Kow, L.-M. and Pfaff,
ar D. W., 1988). The peptides are synthesized in the
somata of the DRG or in ganglion cells of cranial
nerves. They are transported to both the central
and the peripheral terminal of the primary afferent
unit.
Figure 29 Electron micrograph (from sector E of Figure 28)
displaying a putative receptive terminal containing a
characteristic preterminal array of axonal reticulum (ar),
microtubules (arrows), glycogen granules, and spherical 3.4.2 Neuropeptides in Afferent Units
vesicles (asterisks) embedded in a granular matrix and of Different Tissues
clustered below a surface ensheathed by dark Schwann cell
processes (S) and containing vesicles. The bare upper No neuropeptide has been found that can be consid-
axolemma is in direct contact with the basal lamina. Scale ered specific for afferent fibers from a particular
bar 0.5 mm. (From Kruger, L. et al., 2003, with permission.) tissue. DRG cells projecting in a cutaneous nerve
36 Anatomy of Nociceptors

(a) (b) are likely to be involved in painful alterations of

muscle tissue.

gm 3.4.3 Neuropeptides in Nociceptors

and Other Types of Free Nerve Ending
Whether or not a particular neuropeptide or combi-
nation of neuropeptides is associated with a
particular type of receptor is obscure. There is evi-
dence from studies on DRG cells (Lawson, S. N. et al.,
1997; Djouhri, L. and Lawson, S. N., 2004) indicating
that SP-like IR (SP-LI) and to a lesser extent, also
CGRP-LI was present predominantly in nocicep-
tive units. Arguments supporting a nociceptive
function of SP are that noxious stimulation is fol-
lowed by a release of SP in the dorsal horn, and that
the mRNA for SP and other tachykinins is upregu-
lated under these circumstances. On the other hand,
there is also evidence speaking against such a relation
between the nociceptive function and the presence of
SP. In a study by Leah J. D. et al. (1985), the great
majority (10 out of 12) of individually identified
nociceptive DRG cells of the cat did not exhibit
SP-LI. Virtually all primary afferent units with SP-
Figure 30 Electron micrograph of the final traced LI also exhibited CGRP-LI (Garry, M. G. et al.,
portion of the marked receptive field (RF) indicated in 1989); both peptides are presumably released
sector K of Figure 28 containing the final vesicle-rich together when the fiber is active. In the spinal cord,
axonal profiles, including several dense-core vesicles CGRP has been reported to prolong the action of SP
(arrowheads), all within a granular matrix (gm) exposed in
b to the basal lamina. Arrow, additional unidentified
by inhibiting its degradation and to facilitate synaptic
profile. Scale bar 1 mm. (From Kruger, L. et al., 2003b, transmission in general by enhancing the calcium
with permission.) influx into afferent fibers.
Data from experiments in which single DRG
cells with receptive endings in muscle were first
have been shown to contain SP, CGRP, and SOM functionally identified and then injected with a
and thus present a peptide pattern similar to that of dye, showed that at least some cell bodies
muscle nerves, whereas visceral afferent units lack IR whose peripheral processes terminated in muscle
to SOM (Molander, C. et al., 1987; OBrien, C. et al., HTM (presumably nociceptive) receptors exhibited
1989). In comparison to skin nerves, muscle nerves CGRP-LI (Hoheisel, U. et al., 1994). However, IR to
have been reported to contain less SP. This finding SP, CGRP, and other neuropeptides was not only
has been teleologically explained by assuming that present in nociceptive units but also in other types of
the vasodilatation and plasma extravasation caused muscle receptors (e.g., in some muscle spindles and
by the release of SP and CGRP from afferent fibers other LTM units). For instance, SP-LI has been
would be deleterious for skeletal muscle, since it is found in LTM receptors, e.g., in nerve fibers from
surrounded by a tight fascia. Because of the fascia, an Krause corpuscles in the dogs tongue. Most of the
SP-induced muscle edema would result in a high cutaneous and deep receptor types studied so far
increase in interstitial pressure and could cause showed CGRP-LI in some of their cell bodies in
muscle necrosis. On the other hand, there can be no the DRG (Hoheisel, U. and Mense, S., 1987). The
doubt that a considerable proportion of free nerve only condition for the presence of CGRP-LI seemed
endings in muscle exhibit IR to SP and CGRP to be that the afferent unit had a small soma size and/
(Reinert, A. et al., 1998). Therefore, SP and CGRP or a slow CV.
 Anatomy of Nociceptors 37

3.5 Efferent Function of Nociceptors The area of wheal and flare after a localized
damage to the skin for instance around a needle
3.5.1 Release of Neuropeptides from the
prick could be an indicator of the extent of the
Nociceptive Ending
excited nociceptive ending. Of course, this is true
Neuropeptides (e.g., SP, CGRP, VIP, and SOM) are only if a single ending is stimulated and if the dis-
stored in vesicles in the varicosities of the peripheral tance covered by diffusion of the neuropeptides is
terminal. Whenever a nociceptor is excited, it constant.
releases neuropeptides into the interstitial tissue. SP The size of the RFs of cutaneous polymodal noci-
then releases histamine from mast cells, and together ceptors was found to be <2 mm2 in cat (Bessou, P. and
with CGRP these agents cause vasodilatation and an Perl, E. R., 1969) and 632 mm2 in rabbit (Kenins, P.,
increase in vascular permeability of the blood vessels 1988). These figures are greater than the reported
around the active ending. The result is a shift of length of the branches of a nociceptor ending (a few
blood plasma from the intravascular to the interstitial hundred micrometers). The difference may be due to
space. Outside the blood vessel, BK is cleaved from the fact that nociceptors can be excited from a certain
the plasma protein kallidin, and serotonin (5-HT) is distance, particularly when coarse probes are used. For
set free from platelets and PGs (particularly PG E2) a muscle nociceptor, the size of the RF can be deter-
from endothelial and other tissue cells. All these mined only with limitations, particularly if it is located
substances sensitize nociceptors. Thus, the main tis- deep within the muscle. The reported sizes of super-
sue alteration induced by a nondestructive noxious ficially located fields or the projections of deep RFs on
mechanical stimulus is a localized region of vasodi- the muscle surface range from spotlike to several
latation, edema, and sensitized nociceptors. square centimeters in the gastrocnemius muscle of
the cat and dog (Kumazawa, T. and Mizumura, K.,
1977; Mense, S. and Meyer, H., 1985).

3.5.2 The Axon Reflex

3.5.3 Neurogenic Inflammation
The release of neuropeptides from an activated
nociceptor is an essential aspect of its function. The release of SP, CGRP, NK-A, and other agents
from nociceptors is the central factor in the cascade
A nociceptor is not a passive sensor for tissue-
of events that lead to neurogenic inflammation in
threatening stimuli; it actively influences the micro-
the periphery (Lembeck, F. and Holzer, P., 1979).
circulation and chemical composition of the
Neurogenic inflammation is characterized by tissue
interstitial space around it. The morphological basis
edema and infiltration by immune cells after antidro-
of the axon reflex is that the branches of a single
mic activation of nociceptive nerve endings, i.e.,
nociceptive ending can extend over a relatively
when action potentials are generated somewhere
large area (several hundred micrometers). If a nox-
along the course of primary afferent units (spinal
ious stimulus activates only parts of the ending, the
nerve or dorsal root). The action potentials propagate
action potentials originating in the regenerative to both the CNS (causing pain) and the peripheral
region of the ending can invade antidromically ending (causing release of neuropeptides and neuro-
(against the normal direction of propagation) those genic inflammation). The published data indicate
branches of the ending that were not excited by the that vasodilatation can be elicited by antidromic sti-
stimulus. These antidromic action potentials release mulation of both A- and C-fibers, but increase in
neuropeptides from the unstimulated branches. The vascular permeability and plasma extravasation by
whole process is called axon reflex. It is assumed to stimulation of C-fibers only. Neuropathies and radi-
be the reason for the wheal and flare around a cuta- culopathies and other pathological conditions that
neous lesion. The vascular permeability is increased are associated with antidromic activity in sensory
mainly by SP (and by the NK-A and -B; Gamse, R. nerve fibers are examples of such events.
and Saria, A., 1985), whereas CGRP is assumed to act Neurogenic inflammation is likely to increase the
primarily as a vasodilatator. There is evidence show- dysesthesias and pain of patients suffering from
ing that CGRP enhances the plasma extravasation neuropathies.
induced by SP and NK-A and -B, but reduces the Neuropeptides also influence immune cells (for a
vasodilatory action of SP by desensitizing muscle review, see Morley, J. E. et al., 1987) and synoviocytes.
arterioles to the peptide (Ohlen, A. et al., 1988). These actions may be of particular importance for the
38 Anatomy of Nociceptors

development and maintenance of chronic arthritis and concentrations of 5-HT and PGE2 are released,
other inflammatory disorders of deep somatic tissues. which are sufficient for sensitizing, but not for acti-
vating, nociceptors.
Novel stimulants for nociceptors are ATP, cap-
3.6 Receptor Molecules in the saicin, low pH, and NGF (Caterina, M. J. and
Membrane of Nociceptors David, J., 1999). ATP binds to purinergic receptors
(in nociceptive endings, the P2X3 receptor; Ding,
Classic stimulants for nociceptors are endogenous Y. et al., 2000) and opens a cation channel. Animal
pain-producing (algesic) substances such as BK, experiments have shown that nociceptors respond
5-HT, and high concentrations of potassium ions. to ATP in concentrations that are present in tissue
Injections of hypertonic NaCl solutions (4.56.0%) cells. This means that every time a cell is damaged
have been used for many decades to elicit pain from (by trauma or a pathological process), it releases
deep somatic tissues. Similar to potassium, the stimu- ATP in amounts that can cause pain. For this
lating effects of hypertonic NaCl are considered to be reason, ATP has been considered a general signal
unspecific (i.e., not mediated by a receptorligand substance for tissue trauma (Cook, S. P. and
interaction). McCleskey, E. W., 2002).
The effects of BK on nerve endings are mediated Capsaicin, the active ingredient of chilli peppers,
by G protein-coupled B1 and B2 receptors. Binding is a natural stimulant for the vanilloid receptor VR1,
of the ligands to this and other receptors of this now called TRPV1 (Caterina, M. J. and Julius, D.,
type results in changes of the state of activation of 2001). The receptor is also sensitive to an increase in
intracellular second messenger systems such as phos- H concentration and to heat with a threshold of
pholipase C (PLC), cyclic adenosine monophosphate approximately 39  C. The sensitivity of this receptor
(cAMP), cyclic guanosin monophosphate (cGMP), to protons is important under conditions in which the
and protein kinases (PK). One possible sequel of pH of the tissue is lowered (exhaustive muscle work,
the activation of these messengers is the phosphor- ischemia, and inflammation). TRPV2 is present
ylation of ion channels. This increases the opening mainly in myelinated heat nociceptors that have a
time or opening probability of the channels. thermal threshold above 52  C, and TRPV4 is being
Collectively, the processes lead either to direct discussed as the main receptor of mechano-nocicep-
excitation of the ending or to a changed reaction of tors (Liedtke, W., 2005).
the nerve ending to external stimuli (sensitization or A special feature of nociceptive afferent fibers are
desensitization). TTX-r sodium channels. TTX is a neurotoxin that
In intact tissue, the discharges evoked in nerve blocks TTX-sensitive sodium channel and thus inhi-
endings by BK are mainly due to the activation of bits conduction in nerve fibers that possess this type
the BK receptor B2, whereas under pathological con- of Na channel (mostly large-diameter fibers).
ditions the receptor B1 is the predominant one (for a Nociceptive fibers are equipped with TTX-r chan-
review of receptor molecules mediating the effects of nels; therefore, TTX has no blocking action. Two
pain-producing substances, see Kumazawa, T., 1996). TTX-r Na channels that are important for nocicep-
It is often overlooked that the above substances, tion are the voltage-gated sodium channels (Nav) 1.9
particularly BK, excite not only nociceptors but also and 1.8. Nav 1.9 has been found exclusively in noci-
nonnociceptive endings with thin myelinated and ceptive primary afferent neurons, whereas Nav 1.8 is
unmyelinated afferent fibers. Therefore, BK cannot present in both nociceptive and nonnociceptive ones
be considered a specific excitant of nociceptors. (for review, Djouhri, L. and Lawson, S. N., 2004).
However, muscle receptors with group I and II affer-
ents (e.g., muscle spindles and tendon organs) are not
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Kumazawa, T. and Mizumura, K. 1980. Chemical responses visceral convergence in cat dorsal root ganglion neurones
of polymodal receptors of the scrotal contents in dogs. demonstrated by double-labelling with fluorescent tracers.
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 Anatomy of Nociceptors 41

Reinert, A., Kaske, A., and Mense, S. 1998. Inflammation- Steffens, H., Eek, B., Trudrung, P., and Mense, S. 2003.
induced increase in the density of neuropeptide- Tetrodotoxin block of A-fibre afferents from skin and
immunoreactive nerve endings in rat skeletal muscle. Exp. muscle A tool to study pure C-fibre effects in the spinal
Brain Res. 121, 174180. cord. Pflugers Arch. 445, 607613.
Renton, T., Yiangou, Y., Baecker, P. A., Ford, A. P., and Stucky, C. L. and Lewin, G. R. 1999. Isolectin B(4)-positive
Anand, P. 2003. Capsaicin receptor VR1 and ATP and negative nociceptors are functionally distinct.
purinoceptor P2X3 in painful and nonpainful human tooth J. Neurosci. 19, 64976505.
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Rifai, K., Chidiac, J. J., Hawwa, N., Baliki, M., Jabbur, S. J., and Nielsen, T., Arendt-Nielsen, L., and Sessle, B. J. 2003.
Saade, N. E. 2004. Occlusion of dentinal tubules and Glutamate-evoked pain and mechanical allodynia in the
selective block of pulp innervation prevent the nociceptive human masseter muscle. Pain 101, 221227.
behaviour induced in rats by intradental application of Tower, S. 1940. Unit for sensory reception in cornea. With notes
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Rodd, H. D. and Boissonade, F. M. 2000. Substance P 3, 486500.
expression in human tooth pulp in relation to caries and pain Wu, Z.-Z. and Pan, H.-L. 2004. Tetrodotoxin-sensitive and
experience. Eur. J. Oral Sci. 108, 467474. resistant Na channel currents in subsets of small sensory
Rodd, H. D., Loescher, A. R., and Boissonade, F. M. 1998. neurons of rats. Brain Res. 1029, 251258.
Immunocytochemical and electron-microscopic features of Zhang, J. Q., Nagata, K., and Iijima, T. 1998. Scanning electron
tooth pulp innervation in hereditary sensory and autonomic microscopy and immunohistochemical observations of the
neuropathy. Arch. Oral Biol. 43, 445454. vascular nerve plexuses in the dental pulp of rat incisor.
Schaible, H.-G. 2005. Basic Mechanisms of Deep Somatic Anat. Rec. 251, 214220.
Pain. In: Textbook of Pain (eds. S. B. McMahon and
M. Koltzenburg), pp. 621633. Churchill Livingstone.
Schaible, H.-G. and Grubb, B. D. 1993. Afferent and spinal
mechanisms of joint pain. Pain 55, 4554.
Schaible, H.-G. and Schmidt, R. F. 1985. Effects of an Further Reading
experimental arthritis on the sensory properties of fine
articular afferent units. J. Neurophysiol. 54, 11091122.
Schaible, H.-G. and Schmidt, R. F. 1988. Time course of Heppelmann, B, Messlinger, K., Neiss, W. F., and Schmidt, R. F.
mechanosensitivity changes in articular afferents during a 1994. Mitochondria in fine afferent nerve fibres of the knee
developing experimental arthritis. J. Neurophysiol. joint in the cat: a quantitative electron-microscopical
60, 21802195. examination. Cell Tiss. Res. 275, 493501.
Silverman, J. D. and Kruger, L. 1988. Lectin and neuropeptide Heppelmann, B., Pfeffer, H.-G., Schaible, H.-G., and
labeling of separate populations of dorsal root ganglion Schmidt, R. F. 1986. Effects of acetylsalicylic acid and
neurons and associated nociceptor thin axons in rat testis indomethacin on single groups III and IV sensory units from
and cornea whole-mount preparations. Somatosens. Res. acutely inflamed joints. Pain 26, 337351.
5, 259267. Messlinger, K., Pawlak, M., Steinbach, H., Trost, B., and
Simone, D. A., Nolano, M., Johnson, T., Wendelschafer- Schmidt, R. F. 1995. A new combination of methods for
Crabb, G., and Kennedy, W. R. 1998. Intradermal injection localization, identification, and three-dimensional
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subsequent reinnervation of epidermal nerve fibers: characterized by electrophysiology. Cell Tiss. Res.
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cat. J. Anat. 105, 231254.
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 4 Molecular Biology of the Nociceptor/Transduction
M S Gold, University of Pittsburgh, Pittsburgh PA, USA
M J Caterina, Johns Hopkins School of Medicine, Baltimore, MD, USA
2009 Elsevier Inc. All rights reserved.

4.1 Introduction 44
4.1.1 Structure Dictates Function: The Anatomy of the Nociceptor 45
4.1.2 Nociceptor Inputs and Outputs: More than Meets the Eye 46
4.1.2.1 The onset of nociceptor excitation: hyperpolarization versus depolarization 46
4.1.2.2 Efferent function 47
4.1.2.3 Spike integration within the cell body 48
4.1.2.4 Slow axonal transport 48
4.2 The Molecular Toolbox of Sensory Neurons 49
4.2.1 Ion Channels Involved in the Transduction of Extracellular Signals 49
4.2.1.1 P2X adenosine triphosphate-gated ion channels 49
4.2.1.2 Proton-gated ion channels 50
4.2.1.3 Transient receptor potential ion channels 51
4.2.2 Ion Channels Influencing Passive Membrane Properties: Membrane Resistance 53
4.2.3 Ion Channels Underlying Active Membrane Processes 54
4.2.3.1 Action potential upstroke 54
4.2.3.2 Action potential downstroke 54
4.2.3.3 After potential 55
4.2.3.4 Action potential threshold 56
4.2.3.5 High-threshold voltage-gated Ca2 current 57
4.2.4 Membrane Receptors that Indirectly Alter Nociceptor Function 57
4.2.4.1 G-protein-coupled receptors 57
4.2.4.2 Neurotrophin receptors 58
4.3 Additive and Emergent Properties of Nociceptor Membrane Proteins 59
4.3.1 Introduction 59
4.3.2 Shared Modalities and Polymodality 59
4.3.3 Functional Interactions between Nociceptor Proteins 60
4.4 Injury-Induced Plasticity in Transduction 61
4.4.1 Introduction 61
4.4.2 Inflammation-Induced Changes in Nociceptor Sensory function Inflammatory Pain 61
4.4.3 Changes in Nociceptor Function Following Traumatic Nerve Injury Neuropathic Pain 62
4.4.3.1 Nerve injury-induced changes in transduction 62
4.4.3.2 Changes in membrane stability 63
4.4.3.3 Mechanisms underlying nerve injury-induced changes in transduction 64
4.5 Conclusion 64
References 65

Glossary
action potential A characteristic pattern of neu- afterpotential The change in membrane potential
ronal depolarization and hyperpolarization that that follows the action potential.
mediates transmission of information along the cable properties Passive electrophysiological
length of the neuron and triggers communication properties of a neuron that influence the movement
with synaptically connected neurons. of charge, and therefore the spread of a change in

43
44 Molecular Biology of the Nociceptor/Transduction

membrane potential, along a neuron. These prop- neurotrophins A diverse family of soluble signal-
erties include internal resistance (influenced by the ing molecules that promote the differentiation,
diameter of the neural process), membrane resis- survival, and/or function of neurons.
tance (influenced by the number of open ion nociceptor Specialized sensory neuron whose
channels in the membrane), and membrane capa- activation signals the presence of noxious, or
citance (determined by the total membrane surface potentially damaging stimuli. These neurons are
area). selectively activated by noxious stimuli, under nor-
generator potential Membrane depolarization mal circumstances, or by molecules released once
resulting from the transduction of environmental tissue damage has already occurred.
stimuli (i.e., thermal, mechanical, or chemical). This neuropathic pain Spontaneous pain or hyper-
transduction results in a change in membrane sensitivity to environmental stimuli arising from
conductance that underlies the change in mem- damage to neural tissue.
brane potential. receptive field The area in or on the body within
G-protein-coupled receptors A diverse family of which adequate stimuli are capable of generating
membrane proteins that trigger changes in cell activity in a given neuron.
function by signaling through intracellular guanine polymodality The ability of a molecule or neuron
nucleotide-binding proteins. to be activated by more than one type (modality) of
hyperalgesia A state in which the sensitivity of stimulus (e.g., mechanical and chemical).
nociceptive signaling pathways to environmental tranduction A process in which environmental sti-
stimuli is enhanced. muli evoke conformational changes in the structure of
inflammatory pain Spontaneous pain or hyper- specific proteins located on nociceptor terminals that
sensitivity to environmental stimuli arising from directly or indirectly (i.e., via cellular signaling cas-
inflammation in areas innervated by nociceptive cades) trigger the opening or closing of ion channels.
neurons. transient receptor potential A historically based
length constant A measure of the distance over name for a family of related iron channels that are
which a current injected at a point source will prevalent in nociceptive neurons and other sensory
spread within a neuron. neuron subtypes.

4.1 Introduction generator potential reaches a critical threshold,

action potential firing ensues. In order for these
The perception of pain is usually triggered when action potentials to trigger a perception of pain,
mechanical, chemical, or thermal stimuli of high however, they must be propagated to the central
intensity or a particular quality are detected by a terminal of the sensory neuron and subsequently
specialized subpopulation of sensory neurons evoke neurotransmitter release in sufficient quanti-
referred to as nociceptors. Although the term noci- ties to activate postsynaptic spinal cord neurons.
ceptor has probably outlived its utility, we will use it Thus, there are multiple critical steps in the afferent
here as a means of distinguishing afferents capable transmission of nociceptive information, each of
of responding to tissue damage from those that which involves a unique compilation of specialized
normally only encode innocuous stimuli. proteins. We will begin this chapter by describing
Nociceptor activation typically begins with trans- several important but sometimes overlooked anato-
duction, a process in which environmental stimuli mical and physiological features of nociceptive
evoke conformational changes in the structure of neurons. We will then introduce some of the major
specific proteins located on nociceptor terminals protein classes that contribute to transduction and,
that directly or indirectly (i.e., via cellular signaling to a more limited extent, the ensuing neurophysio-
cascades) trigger the opening or closing of ion chan- logical processes that result in the transmission of
nels. The resulting changes in ionic flow across the nociceptive information to the spinal cord.
plasma membrane produce a change in membrane Subsequently, we will provide a glimpse at how
potential, referred to as a generator potential. If the such variables as molecular heterogeneity,
 Molecular Biology of the Nociceptor/Transduction 45

intermolecular interactions, posttranslational mod- processes merges to form a T junction, leaving the
ifications, and ion channel distribution can vastly cell body attached to peripheral and central processes
expand the sensory versatility of the nociceptive via a single short length of axon (Fitzgerald, M., 2005)
neurons. Finally, we will describe in greater detail (Figure 1). Both central and peripheral processes
a few of the changes that underlie nociceptor signal- terminate in a branching pattern referred to as a
ing plasticity in the context of inflammation and terminal arbor (Figure 1). The extent of the periph-
nerve injury. Although neurotransmission within eral arbor depends on the afferent type and site of
the spinal cord dorsal horn represents another innervation. For example, afferents innervating the
important function of nociceptors, we will defer a fingertips may innervate a square millimeter or less
description of this process to other chapters of this of skin, and therefore appear to have relatively few
volume. terminal branches (Darian-Smith, I. et al., 1980;
Andrew, D. and Greenspan, J. D., 1999). In contrast,
afferents innervating a visceral structure such as the
4.1.1 Structure Dictates Function: colon may have an extensive terminal arborization
The Anatomy of the Nociceptor that extends over many centimeters of tissue
Sensory neurons are referred to as pseudounipolar (Berthoud, H. R. et al., 2004). Afferents responsible
neurons because they start out during development for the transduction of low-threshold mechanical
as bipolar neurons with a central process extending stimuli such as brush or vibration terminate in spe-
into the spinal or trigeminal dorsal horn and a per- cialized structures such as Rufinni endings or Merkel
ipheral process extending out to peripheral targets. discs that are critical for mediating the appropriate
During development, the initial segment of these two response to these stimuli (Mearow, K. M. and

Schematic of cell body axon and

Normal Pathological
terminals in rough proportion
Cell body/ganglion: Cell body/ganglion:
1. Import of neurotrophin To spinal 1. Excitatory transducer, spike initiator
receptor signalling complexes cord synthesis
2. K+ channel, inhibitory GPCR synthesis
2. Ca2+ binding, store handling 3. Changes in Ca2+ binding proteins, store
3. Synthesis of transducers and handling
neuropeptides 4. Changes in gene transcription/translation
4. Transport towards spinal cord 5. Intraganglionic transmitter release and
and/or periphery cross-talk
6. Local immune cells, cytokines
7. Sympathetic sprouting
Up to
1m
Axon: Axon:
1. Ion channels spaced Cut axon
appropriately for action potential 1. Proximal accumulation of transducers,
channels
propagation
2. neurotrophin and neuropeptide transport
2. Ortho- and anterograde 3. Local immune cells, cytokines
transport Neighboring Uninjured axon
1. channel density
2. neurotrophin transport
Peripheral Terminal:
1. Multiple sites of transduction
2. Proximity of transducers to Peripheral terminal:
spike initiators 1. Excitatory transducer and spike initiator
3. Modality specificity number
4. Polymodality of transducers 2. Change in transducer subtype composition
5. Efferent function peripheral 3. Transducer phosphorylation
4. K+ channel, inhibitory GPCR number
release of SP, CGRP 5. Changes in modality specificity
6. Anatomical changes (terminal
sprouting/branching)
7. pH, inflammatory mediators

Figure 1 Schematic diagram of three major peripheral compactments of a nociceptive neuron: the cell body (top),
axon (middle), and peripheral terminal arbor (bottom). Blowups at left: functional and anatomical properties of these
compartments under normal conditions. Blowups at right: characteristic changes associated with nerve injury or
inflammation.
46 Molecular Biology of the Nociceptor/Transduction

Diamond, J., 1988; Johnson, K. O., 2001). In contrast, structures. As suggested above, the structure of
nociceptors are said to have free nerve endings, the terminal arbor varies with target of innovation.
because peripheral terminals of these afferents do There is also evidence that subpopulations of
not appear to be associated with any specific cell nociceptive afferents have distinct terminal arbor
type (Kruger, L. et al., 2003a). Further description of patterns within the same structure. For example, the
this anatomical arrangment can be found in the chap- terminal arbor morphology of cutaneous C fibers
ter by Mense in this volume (Chapter Anatomy of varies according to whether the C fiber is peptidergic
Nociceptors). (i.e., expresses substance P or calcitonin gene-related
There are at least three important implications of peptide (CGRP)) or expresses the mas-related gene
the observation that peripheral terminals of noci- (mrg), MrgD (Zylka, M. J. et al., 2005). Furthermore,
ceptors are free. First, while recent evidence in the cornea, the terminal arbor of C-fiber afferents
suggests that transduction may occur in cells sur- is distinct from that of A fibers (MacIver, M. B. and
rounding nociceptive terminals (i.e., urothelial cells Tanelian, D. L., 1993). At an ultrastructural level,
(Birder, L. A., 2005) or keratinocytes (Guler, A. D. nociceptive terminals appear to have beaded endings
et al., 2002; Peier, A. M. et al., 2002b; Khodorova, A. with clusters of organelles such as mitochondria at
et al., 2003; Wu, G. et al., 2004a) (see below)), the fact sites of swelling (Kruger, L. et al., 2003b). While the
that nociceptive terminals are generally not asso- functional implications of this beaded structure
ciated with specialized cells suggests that these remain to be determined, it has been suggested that
neurons themselves possess the machinery neces- swellings may be sites for transduction and/or the
sary for transduction. Consistent with this peripheral release of transmitter.
suggestion, the isolated cell bodies of putative noci- Finally, it is clear that active membrane proper-
ceptors in vitro have been show to respond to ties, and therefore the nature of the afferent signal,
thermal (Cesare, P. and McNaughton, P., 1996; are influenced by an interaction between the dis-
Reichling, D. B. and Levine, J. D., 1997; Reid, G. tribution, density, and biophysical properties of ion
and Flonta, M., 2001; Thut, P. D. et al., 2003), channels and transducers present in afferent term-
mechanical (McCarter, G. C. et al., 1999), and var- inals and the morphological features of the terminal
ious chemical (Baccaglini, P. I. and Hogan, P. G., arbor. Cable theory describes the impact of leak
1983) stimuli. Second, if transduction machinery is channels, terminal branch points, and irregularities
present in nociceptive terminals, transduction may in membrane shape such as those associated with
occur whenever and wherever a transducer is prop- the beaded pattern described above. As a first pass,
erly positioned in the cell membrane. The result is all of these features, in addition to the relatively
that transduction can and probably does occur at large surface-to-volume ratio associated with
multiple sites in a peripheral arbor (Gover, T. D. fine terminals suggest that the length constant, a
et al., 2003; Zhao, J. et al., 2006). This has important measure of the distance over which a current
implications for spike initiation and integration of injected at a point source will spread within the
stimuli applied throughout a receptive field terminal, is quite small in nociceptive terminals.
(Darian-Smith, I. et al., 1980). It also has important This implies that there are likely to be multiple
implications for the generation of ectopic activity in sites of spike initiation throughout the terminal
the presence of injury as there is evidence that the arbor. Importantly, it is likely that the cable proper-
cut ends of axons may rapidly develop mechanical, ties, and therefore length constant, of nociceptive
thermal, and chemical sensitivity (Michaelis, M. terminals are dynamically regulated. This notion
et al., 1999) and that spontaneous activity may arise is based on the observation that potassium
from within sensory ganglia following nerve injury leak currents, which determine membrane resis-
(Kajander, K. C. et al., 1992). Third, because nerve tance, may be regulated by a number of factors
endings are not only free of an association with a such as transmitters and second messengers
specific cell type, but free of even perineurial (Maingret, F. et al., 2000). Furthermore, evidence
encapsulation (Devor, M., 1999), these endings are from dorsal horn neurons (Mantyh, P. W. et al.,
not only accessible to the chemical milieu of the 1995) suggests that terminal morphology may be
tissue, but may be accessible to therapeutic rapidly altered. Importantly, a change in a length
interventions. constant may influence active membrane properties
Both light and electron micrographic analyses of such as action potential threshold and spike
nociceptive terminals reveal complex anatomical accommodation.
 Molecular Biology of the Nociceptor/Transduction 47

4.1.2 Nociceptor Inputs and Outputs: More et al., 2002). Second, the presence of a significant
than Meets the Eye depolarizing conductance in nociceptive afferents
suggests that the resting membrane potential will be
The major focus of this chapter is on molecules
relatively depolarized. This has been observed
involved in the initiation of fast orthograde electrical
repeatedly in studies on dissociated sensory neurons,
signaling by nociceptors to the central nervous sys-
where the resting membrane potential of putative
tem. However, several physiological features of
nociceptive afferents is more depolarized than
nociceptors fall outside this conventional framework
that of putative non-nociceptive afferents (see
but are important within the larger context of sensory
Gold, M. S. et al., 1996a). There is evidence that
transduction. Therefore, before we dissect the
intact nociceptive terminals in situ may be depolar-
mechanisms of nociceptor stimulus transduction,
ized as well (Brock, J. A. et al., 2001). As mentioned
action potential generation, and action potential pro-
above, resting membrane potential may influence not
pagation in any detail, it is worth considering a few of
only transduction, given the voltage dependence of
these features.
several transducers (see below), but also spike
initiation.
4.1.2.1 The onset of nociceptor excitation:
hyperpolarization versus depolarization 4.1.2.2 Efferent function
Sensory transduction in the mammalian nervous It is clear that nociceptor afferent neurotransmission
system can arise from either membrane hyper-polar- contributes to the perception of pain associated with
ization or membrane depolarization. The visual tissue injury (Treede, R. D. et al., 1992). However, it is
system utilizes the former while the somatosensory also clear there are tissues distributed throughout the
system utilizes the latter. Consequently, in the absence body that are heavily innervated by nociceptive
of stimuli, the resting membrane potential of nocicep- afferents, but that will never give rise to the sensation
tive afferents is relatively hyperpolarized, with little, if of pain in many of us. For example, even though
any spontaneous activity (Rose, R. D. et al., 1986; pericardial tissue receives nociceptive innervation
Ritter, A. M. and Mendell, L. M., 1992). This resting thought to be responsible for pain associated with
membrane potential appears to reflect a relatively cardiac ischemia (Immke, D. C. and McCleskey, E. W.,
large K conductance through both voltage-insensi- 2001), the majority of us will never experience dis-
tive (Baumann, T. K. et al., 2004) and voltage-gated K comfort arising from this structure. What might be
channels (Kirchhoff, C. et al., 1992). However, this K the function(s) of nociceptors in these locations? One
conductance appears to be working against a relatively potential answer to this question stems from the
high membrane permeability to depolarizing cations ability of many nociceptors to release bioactive sub-
(i.e., Na and Ca2), at least in cutaneous afferents stances with the potential to exert trophic and/or
(Kirchhoff, C. et al., 1992). Data from studies of the inflammatory effects on targets of innervation. For
isolated sensory neuron cell body in vitro suggest that a example, CGRP, which can be released by many
significant depolarizing conductance may be present nociceptive neurons, has been shown to regulate
in a relatively large percentage of putative nociceptive cellular activity underlying bone formation and
afferents (Viana, F. et al., 2002). resorption, two processes critical to the maintenance
There are at least two implications of a significant of healthy bone (Lerner, U. H., 2002). Such seemingly
depolarizing conductance in nociceptive afferents. paradoxical efferent nociceptor activity is particu-
First, it suggests that a viable mode of transduction larly important in tissues subjected to injury. Under
may involve the closing of a K channel. In the these circumstances, the release of transmitters from
absence of a depolarizing conductance, closing a K the peripheral nociceptor terminals has been shown
channel in a membrane in which K conductance is to contribute to local hyperemia and plasma extra-
the only major conductance responsible for the vasation (hallmarks of the so-called neurogenic
establishment of the resting membrane potential, inflammation), to the orchestration of tissue repair,
will result in little, if any change in membrane poten- and to the initiation and maintenance of pain and
tial. Consistent with the suggestion that there is a hyperalgesia (Willis, W. D., 1999). Much of the focus
significant depolarizing conductance in putative in this area has been on nociceptor release of CGRP
nociceptive afferents, there is evidence that some and another peptide transmitter, substance P
forms of cold transduction reflect the closing of a (Figure 1). However, it is now clear that glutamate
K channel (Reid, G. and Flonta, M., 2001; Viana, F. is also released into the periphery, where it appears
48 Molecular Biology of the Nociceptor/Transduction

able to act in an autocrine fashion to further activate Wall, P. D., 1990). While the exact mechanisms
and/or sensitize nociceptive afferents (deGroot, J. underlying such cross talk have yet to be completely
et al., 2000). Appreciation of these efferent capabilities worked out (Amir, R. and Devor, M., 1996), there is
has led to the suggestion that a better name for evidence that the sensory neuron cell body is capable
nociceptive sensory neurons might be noceffector of releasing transmitter (Huang, L. Y. and Neher, E.,
afferents (Kruger, L., 1988). 1996) and that peripheral nerve stimulation results in
Three mechanisms have been described that may the release of transmitter within the ganglia
contribute to the injury-induced efferent release of (Matsuka, Y. et al., 2001).
neurotransmitter. One is the axon reflex, thought to More relevant to the issue of transduction, how-
be mediated by action potential spread from one branch ever, is the observation that afferent activity is capable
in a terminal arbor to other branches (Lewis, T., of driving changes in gene transcription (Figure 1).
1935). A second, relatively rare, mechanism is axonal For example, there is evidence that electrical activity
coupling, in which axons traveling in the same bundle in sensory neurons is capable of changing Na chan-
may, as a result of injury, come into such close nel expression (Klein, J. P. et al., 2003). Interestingly,
proximity that activity in one axon is able to initiate the pattern of activity appears to influence the pattern
activity in a second axon (Meyer, R. A. et al., 1985). of gene transcription (Fields, R. D. et al., 2005). The
A third mechanism actually involves the initiation of result may be as profound as to determine whether or
action potentials in the central terminals of nociceptive not sprouting axons are able to coalesce into a nerve.
afferents within the spinal cord dorsal horn that are One mediator that appears to be critical for the inte-
propagated antidromically out into the periphery gration of activity within the cell body is Ca2, acting
(Sluka, K. A. et al., 1995). While the exact mechanism through a number of Ca2-dependent molecules
of this activation is still debated, there is evidence that in (Eshete, F. and Fields, R. D., 2001). Thus, the trans-
the presence of inflammation there is a change in duction of activity within the sensory neuron cell
presynaptic gamma-aminobutyric acid (GABA) ergic body involves the influx of Ca2, which then initiates
input, such that primary afferent-driven GABA release a number of cellular processes including gene
from interneurons is capable of action potential genera- transcription.
tion at central terminals (Sluka, K. A. et al., 1995). A
fourth mechanism, often overlooked in this discussion, 4.1.2.4 Slow axonal transport
involves the possible direct coupling between calcium- Another general mechanism critical to nociceptive
permeable transducers and transmitter release. Such a signaling involves axonal transport to the cell body,
mechanism appears to contribute to the inflammation rather than action potential propagation. This is a
associated with capsaicin-induced activation of its remarkable process, given the tremendous length of
receptor transient receptor potential vanilloid 1 sensory nerve axons (up to 1 m), relative to their soma
(TRPV1) (see below). size (1830 mm). Among the primary functions of this
form of signaling, which may take hours to days, are
4.1.2.3 Spike integration within the cell the maintenance of nociceptor survival during devel-
body opment, the maintenance of nociceptive phenotype
There is evidence indicating that once the T junction and function in the adult, and orchestration of the
is formed within the sensory ganglia, action poten- afferent response to tissue injury (Koltzenburg, M.,
tials need not invade the sensory neuron cell body 1999). The best-studied example of signaling via slow
(Amir, R. and Devor, M., 2003). This observation axonal transport is that of neurotrophic factors bound
begs the question of why action potentials invade to their receptors (Figure 1) (Zweifel, L. S. et al., 2005).
the cell body at all. One answer to this question However, slow axonal transport can also lead to signal-
appears to be that the invading action potential pro- ing in the distant sensory neuron cell body following
vides a mechanism for the recruitment of additional ligand-dependent internalization of G-protein-
activity. The term cross-excitation has been used to coupled receptors (GPCRs) (e.g., those for neuroki-
describe the observation that activity in one axon can nins, opioids, and extracellular proteases) (Tseng, L. F.
induce activity in neighboring axons (Devor, M. and and Narita, M., 1995; Vergnolle, N. et al., 2001). In
Wall, P. D., 1990). Interestingly, the prevalence of addition, transcription factors, second messenger
cross-excitation increases in the presence of tissue molecules, cell surface glycoproteins, and foreign par-
injury, providing a mechanism for peripherally ticles such as plant lectins, toxins, and viruses can all be
mediated secondary hyperalgesia (Devor, M. and transported retrogradely from peripheral sensory
 Molecular Biology of the Nociceptor/Transduction 49

neuron terminals toward the cell body (Wang, H. F. 4.2.1 Ion Channels Involved in
et al., 1998; Vulchanova, L. et al., 2001; Tarpley, J. W. the Transduction of Extracellular Signals
et al., 2004; Yeomans, D. C. et al., 2005; Mata, M. et al.,
4.2.1.1 P2X adenosine triphosphate-
2006). In what has been a boon to researchers, there are
gated ion channels
some molecules that are selectively taken up by sub-
Extracellular adenosine triphosphate (ATP), released
populations of sensory neurons, and have therefore upon cell damage, has been recognized as a nocicep-
been used to characterize innervation patterns (see tive stimulus ever since the demonstration that
Wang, H. F. et al., 1998). Sensory neurons also appear application of this molecule to a blister base produces
to be able to take up inflammatory mediators such as pain in human psychophysical studies (Bleehan, T.
TNF (Schafers, M. et al., 2002). Exactly how sensory and Keele, C. A., 1977). At least one mechanism by
neurons use the information associated with the which ATP activates nociceptors is via activation of
presence or absence of various molecules transported ATP-gated ion channels of the P2X family. These
back to the cell body remains to be determined. channels are comprised of a heterotrimer of P2X
However, the internalization and transport of mole- subunits, each with two transmembrane domains
cules present at sensory neuron terminals appears to and a large extracellular loop that contains the ATP
be a mechanism for sensory neurons to monitor binding site. Seven P2X subtypes have been cloned in
tissue integrity and adjust to changes in the local mammals. They differ with respect to their sensitiv-
environment. ity to ATP or ATP analogs, as well as the kinetics of
their ATP-evoked currents. For example, homo-
meric P2X3 is responsive to ,-methylene ATP,
and exhibits a rapidly desensitizing current response,
4.2 The Molecular Toolbox of whereas homomeric P2X2 exhibits slowly desensitiz-
Sensory Neurons ing currents that are ,-methylene ATP insensitive.
When coassembled, these channels form an ,-
Sensory neurons express a multitude of cell surface methylene sensitive, slowly desensitizing heteromul-
proteins that serve anatomically as markers of neu- timeric channel. P2X channels are typically
ronal subtypes and functionally as mediators of the nonselective cation channels that exhibit consider-
fundamental signaling and transmission functions able permeability to both Na and Ca2 ions. In
outlined in the previous section. Three classes of several cases, however, (P2X2, P2X4, and P2X7)
sensory neuron cell surface proteins have been stu- protracted stimulation of these channels can lead to
died in the greatest detail: ion channels, metabotropic an apparent dilation of the channel pore, such that
GPCRs, and receptors for neurotrophins or cyto- much larger organic cations such as N-methyl
kines. Together, these proteins endow sensory D-glucamine are able to permeate the channel. The
afferents with responsiveness to a qualitatively and significance of this dynamic ion selectivity remains
quantitatively diverse collection of environmental unclear (North, R. A., 2002).
variables and dictate the passive and active properties Although mRNAs encoding P2X1 through P2X6
essential for converting a generator potential into a have all been detected in sensory neurons, P2X2 and
signal that reaches the spinal cord. As detailed below, P2X3 appear to be expressed most prominently in
certain membrane proteins found in nociceptors (e.g., these cells. P2X2 is expressed in neurons with a wide
P2X3, TRPM8, mrgA, and NaV1.8) are expressed range of cell body sizes. In contrast, at least in skin,
within subsets of sensory neurons at levels much P2X3 expression appears to be confined to a sub-
higher than anywhere else in the body. Such enriched population of small-diameter sensory neurons that
expression has provided reagents that can be used bind isolectin B4 (IB4) and that project to inner
for sensory neuron- or nociceptor-specific gene dis- lamina II of the spinal cord dorsal horn (North, R.
ruption or transgene expression in the laboratory A., 2002). It is important to note, however, that the
setting. More importantly, it provides at least the precise relationship between P2X3 expression and
theoretical opportunity to develop antinociceptive IB4 binding may vary between cutaneous affer-
drugs that have minimal effects on other cell types. ents with their cell bodies in dorsal root ganglia and
A few of the more prominent classes of cell surface other nociceptor populations such as those in the
proteins expressed in sensory neurons are described trigeminal ganglion (Ambalavanar, R. et al., 2005).
below. Genetic disruption of both P2X3 and P2X2 results
50 Molecular Biology of the Nociceptor/Transduction

in elimination of all transient ATP-evoked currents 4.2.1.2 Proton-gated ion channels

and all but a small persistent ATP-evoked current in Reductions in tissue pH, as might occur during ische-
cultured sensory neurons (Cockayne, D. A. et al., mia or inflammation, can potently activate
2005). nociceptors and sensitize them to heat and other
P2X channels also contribute to pain sensation in stimuli. In vitro, low pH activates cationic channels
vivo. For example, although application of P2X3 in sensory neurons that can exhibit transient, sus-
antagonists (Jarvis, M. F. et al., 2002; McGaraughty, S. tained, or biphasic kinetics (Bevan, S. and Yeats, J.,
et al., 2003) or antisense oligodeoxynucleotide- 1991). At least some of these responses can be attrib-
mediated knockdown of P2X3 (Barclay, J. et al., uted to the acid-sensing ion channel (ASIC) family.
2002; Honore, P. et al., 2002) have no apparent effect These multimeric ion channels are comprised of
on acute nociceptive responses to mechanical or subunits that topologically resemble the P2X chan-
thermal stimuli, both interventions have been nels, with two transmembrane domains each and an
shown to reduce thermal and/or mechanical hyper- extracellular ligand (H) binding domain. Five ASIC
algesia associated with complete Freunds adjuvant subtypes have been cloned, of which four (ASIC14)
(CFA) administration into the paw and several mod- are expressed in peripheral sensory neurons
els of neuropathic pain, as well as reducing the extent (Waldmann, R. et al., 1999; Waldmann, R., 2001).
of the first and second phases of paw licking follow- Splice variants of several subtypes add to the overall
ing intraplantar formalin injection. Gene knockout of complexity, and one such variant, ASIC1, appears
P2X3 or P2X2/3 receptors in mice has also been to be selectively enriched in sensory neurons
reported to diminish formalin-evoked nocifensive (Chen, C. C. et al., 1998). Upon activation by low
behavior. However, these knockout animals exhibit a pH (from <7 to <4, depending on the subtype)
perplexing mixture of pain-related phenotypes, ASICs typically mediate a prominent and rapidly
desensitizing Na current. However, some subtypes
including paradoxically enhanced behavioral avoid-
(e.g., ASIC3), exhibit a biphasic current consisting of
ance of heat and cold stimuli (Cockayne, D. A. et al.,
an initial desensitizing sodium current, followed by a
2000; Souslova, V. et al., 2000; Cockayne, D. A. et al.,
more sustained current that includes Ca2 ions
2005; Shimizu, I. et al., 2005). In visceral structures,
(Waldmann, R. et al., 1999; Waldmann, R., 2001).
epithelial cells appear capable of the storage and
Several lines of evidence support the participation
Ca2-dependent release of ATP in response to
of ASIC channels in acid responsiveness of sensory
mechanical and chemical stimuli. P2X3 and P2X2/3
neurons. First, the kinetics of sensory neuron
receptors on primary afferent terminals enable visceral
responses to a pH 6 stimulus are altered in neurons
afferents to respond to this released ATP, perhaps con- isolated from ASIC1, ASIC2, or ASIC3 knockout
tributing to visceral pain sensation (Cockayne, D. A. mice or in wild-type neurons exposed to APETx2,
et al., 2000; Vlaskovska, M. et al., 2001; Andersson, K. E., an ASIC3-selective antagonist from sea anemone
2002; Birder, L. A., 2005). A similar mechanism may (Price, M. P. et al., 2001; Benson, C. J. et al., 2002;
underlie ATP-mediated stimulus transduction in cuta- Xie, J. et al., 2002; Diochot, S. et al., 2004). Second, pH
neous tissue in the presence of inflammation (Wu, G. 5-evoked C-fiber responses are reduced in ex vivo
et al., 2004a). P2X receptors contribute to nociceptive skinnerve explants derived from ASIC3 knockout
neurotransmission in a number of other ways, as well. mice (Price, M. P. et al., 2001). Third, mice lacking
In the spinal cord, for example, activation of presynap- ASIC3 exhibit defects in the ipsilateral mechanical
tic P2X3-containing channels and postsynaptic P2X hyperalgesia that typically develops 4 h after injec-
channels of other subtypes, modulates neurotransmit- tion of acid to the gastrocnemius muscle (Price, M. P.
ter release and excitability, respectively (Gu, J. G. and et al., 2001) as well as defects in the bilateral hyper-
MacDermott, A. B., 1997; Nakatsuka, T. and Gu, J. G., algesia that develops after two intramuscular acid
2001; Nakatsuka, T. et al., 2003; Tsuzuki, K. et al., 2003). injections, 5 days apart (Sluka, K. A. et al., 2003).
In addition, although it appears not to be expressed Interestingly, no changes in behavioral responses to
within neurons themselves, P2X7 is necessary for nor- acute subcutaneous acid injection have been reported
mal inflammation, nerve injury-induced hyperalgesia in these knockout studies. However, ASIC3 may be
(Chessell, I. P. et al., 2005), and CFA-induced hyper- particularly important for sensing reductions in mus-
algesia in rats (DellAntonio, G. et al., 2002a; 2002b), cle pH during ischemia (Immke and McCleskey 2001).
probably owing to its role in cytokine release from In addition to acid detection, ASICs have also
inflammatory cells (Labasi, J. M. et al., 2002). been implicated in mechanosensation. This
 Molecular Biology of the Nociceptor/Transduction 51

hypothesis was initially based upon the recognition these animals exhibit diminished responsiveness not
that ASICs bear structural and pharmacological simi- only to subsequent capsaicin challenge, but also to
larities (i.e., amiloride sensitivity) to degenerins, other painful stimuli ( Jancso, G. et al., 1977). The
cationic channels identified in genetic screens for highly enriched expression of a vanilloid receptor
mechanosensory defects in the nematode, in nociceptive neurons appears to underlie these
Cenorhabditis elegans (OHagan, R. and Chalfie, M., incredibly selective effects of capsaicin and related
2005). The immunolocalization of ASIC2 and 3 to vanilloid compounds (Szallasi, A. and Blumberg, P.
sensory afferent terminals (Price, M. P. et al., 2000; M., 1999). This conclusion was corroborated by the
Garcia-Anoveros, J. et al., 2001; Price, M. P. et al., molecular cloning of a capsaicin-gated ion channel
2001) and the systematic examination of mechano- named TRPV1 (Caterina, M. J. et al., 1997).
sensation in ASIC knockout mice have lent support TRPV1 belongs to the TRP family, which in
to this notion. Mice lacking ASIC1 were recently mammals contains approximately 30 members. TRP
reported to exhibit no detectable changes in mechan- channel subunits have six transmembrane domains
oreceptor properties in the skinnerve preparation. and intracellular N- and C-termini, and apparently
However, there was an enhancement of mechanically form tetrameric channels on the cell surface.
evoked firing rate in gastrointestinal afferents in A remarkable feature of TRPV1 is that, in addition
these mice, accompanied by a reduction in gastric to vanilloid compounds, it can be activated by a
emptying rate (Page, A. J. et al., 2004). Studies of number of other stimuli, including protons (pH < 5)
ASIC2 KO mice have yielded somewhat conflicting (Tominaga, M. et al., 1998), certain endogenous
results. In one study, these mice were reported to arachidonic acid derivatives (anandamide, N-arachi-
exhibit reduced mechanically evoked firing rate of donyl dopamine, and 12-HPETE) (Zygmunt, P. M.
rapidly adapting A mechanoreceptors (RAM), et al., 1999; Hwang, S. W. et al., 2000; Huang, S. M.
without a change in RAM threshold (Price, M. P. et al., 2002) and even noxious heat (>42  C)
et al., 2000). In another study, however, no change (Caterina, M. J. et al., 1997). Nonvanilloid exogenous
in RAM response characteristics was observed (Roza, chemicals capable of activating TRPV1 include etha-
C. et al., 2004). ASIC3 KO mice were reported to nol (Trevisani, M. et al., 2002), camphor (Xu, H. et al.,
exhibit decreased sensitivity of A mechanoreceptors 2005), and 2-aminoethoxydiphenyl borate (2-APB)
but increased sensitivity of RAM, together with (Hu, H. Z. et al., 2004).
increased behavioral withdrawal from mechanical Sensory neurons derived from TRPV1 knockout
stimulation after carageenan (Price, M. P. et al., 2001). mice not only are devoid of vanilloid responsiveness
In contrast to the situation for acid detection, it has but also exhibit deficits in current responses evoked
been relatively difficult to correlate mechanically by pH 5 or moderate heat (4550  C) stimuli. All in
evoked currents with the presence of particular ASIC vivo vanilloid-evoked responses are correspondingly
subunits. Indeed a recent in vitro study of mechanically absent in TRPV1 knockout mice. These animals also
evoked currents revealed no differences between wild exhibit modestly reduced (but not absent) behavioral
type and ASIC2 knockout, ASIC3 knockout, or responses to acute noxious heat. A far more striking
ASIC2/3 double knockout mice. Instead, the authors nociceptive phenotype is apparent in these mice fol-
of this study observed that the cationic dye, ruthenium lowing cutaneous inflammation by mustard oil,
red, could inhibit mechanosensitive currents, suggest- carageenan, or complete Freunds adjuvant or follow-
ing the possible participation of transient receptor ing surgical incision of the skin. Under these
potential (TRP) channels (see below) (Drew, L. J. conditions, almost none of the thermal hyperalgesia
et al., 2004). typically exhibited by wild-type mice is observed in
mice lacking TRPV1, although mechanical hyper-
4.2.1.3 Transient receptor potential ion algesia is unaffected (Caterina, M. J. et al., 2000;
channels Davis, J. B. et al., 2000; Pogatzki-Zahn, E. M. et al.,
One early clue that nociceptive neurons might 2005). More recent pharmacological studies have
express signaling proteins distinct from those of further supported a role for TRPV1 in inflammatory
other neuronal subtypes was the observation that thermal hyperalgesia (Pomonis, J. D. et al., 2003;
exposure of neonatal rats to capsaicin, the main pun- Gavva, N. R. et al., 2004; Honore, P. et al., 2005).
gent ingredient in hot peppers, produces a lifelong However, the degree to which this channel may or
depletion of small-diameter sensory neurons, with no may not participate in neuropathic pain states is
such effects on larger-diameter neurons. As adults, much less clear. Although some pharmacological
52 Molecular Biology of the Nociceptor/Transduction

studies using TRPV1 antagonists with significant et al., 2002b), where TRPV3-mediated current
potential for off-target effects have suggested a pos- responses can be readily generated (Chung, M. K.
sible role (Pomonis, J. D. et al., 2003; Walker, K. M. et al., 2004a; Moqrich, A. et al., 2005). These findings
et al., 2003), mice lacking TRPV1 exhibit apparently have led to the notion that TRPV3 may signal the
normal thermal hyperalgesia following partial presence of cutaneous warmth via keratinocytesen-
ligation of the sciatic nerve (Caterina, M. J. et al., sory neuron communication. Support for a role of
2000). The recent availability of more selective TRPV3 in thermosensation has come from the obser-
TRPV1 antagonists promises to resolve this issue vation that in mice lacking this protein, heat-evoked
(Rami, H. K. and Gunthorpe, M. J., 2004). One parti- nociception is blunted in the hot plate and tail
cularly promising recent finding was that a TRPV1- immersion assays. In addition, on a thermal gradient,
selective antagonist was shown to partially attenuate selection of preferred floor temperatures is slowed,
the mechanical hyperalgesia associated with experi- but not absent, in these mice (Moqrich, A. et al., 2005).
mental bone cancer pain (Ghilardi, J. R. et al., 2005). TRPV4 can be gated by warm temperatures
Following the identification of TRPV1, a host of (>32  C), hypo-osmolarity, cytochrome P450 meta-
other TRP channels were found to be expressed in bolites of arachidonic acid, or the synthetic phorbol
sensory ganglia. Like TRPV1, many of these channels ester, 4-phorboldibutyryl didecanoate (Nilius, B.
are polymodal, exhibiting responsiveness to a multiple et al., 2004). Like TRPV3, TRPV4 is expressed
physical and chemical stimuli ( Jordt, S. E. et al., 2003; more prominently in keratinocytes than in sensory
Patapoutian, A. et al., 2003). Whereas TRPV1 appears afferents (Guler, A. D. et al., 2002), although there is
to be the only member of this family gated by capsai- evidence for low-level TRPV4 protein expression in
cin, TRPV2, TRPV3, and TRPV4, like TRPV1, can a population of medium- to large-diameter sensory
be activated by elevated ambient temperatures. afferents (Liedtke, W. and Friedman, J. M., 2003).
TRPV2 is expressed most prominently in a subpopu- Mice lacking TRPV4 exhibit reduced sensitivity to
lation of neurons with medium- to large-diameter cell noxious mechanical or hypertonic stimuli, prolonged
bodies. Based on their size, expression of CGRP and tail immersion latencies at modestly hot tempera-
neurofilament proteins, and projection pattern within tures, and a shift in thermal selection behavior
the spinal cord, many of these neurons are likely to toward floor temperatures warmer than those
correspond to A neurons. However, some presumably selected by wild-type mice (Liedtke, W. and
A neurons that project within the dorsal columns also Friedman, J. M., 2003; Suzuki, M. et al., 2003;
appear to express TRPV2. Heterologously expressed Alessandri-Haber, N. et al., 2005; Lee, H. et al., 2005b).
TRPV2 can be activated by temperatures >52  C, a Just as heat can activate TRPV1TRPV4, cold
thermal response pattern reminiscent of those exhib- temperatures have been reported to activate at least
ited by Type II A fibers in peripheral nerve recording two other TRP channels. TRPM8 was identified on the
studies and by a subpopulation of medium-diameter, basis of its responsiveness to the cold-mimetic agent,
capsaicin-insensitive dorsal root ganglion (DRG) neu- menthol. Examination of its thermosensory responsive-
rons in isolated cultures (Caterina, M. J. et al., 1999). ness revealed, however, that it could alternatively be
TRPV2 can alternatively be activated by hypo-osmo- activated by modest reductions in ambient temperature
larity (Iwata, Y. et al., 2003), cell stretch (Muraki, K. (<22  C) (McKemy, D. D. et al., 2002; Peier, A. M. et al.,
et al., 2003), or 2-APB (Hu, H. Z. et al., 2004). 2002a). Another cold-mimetic agent, icilin, can also
TRPV3 is activated by warm temperatures (>34 activate TRPM8, but only when intracellular Ca2
37  C), although its thermal responsiveness extends levels are elevated (McKemy, D. D. et al., 2002;
well into the noxious range (Peier, A. M. et al., 2002b; Chuang, H. H. et al., 2004). TRPM8 is expressed in a
Smith, G. D. et al., 2002; Xu, H. et al., 2002). This subset of small-diameter neurons that lack expression
channel can, alternatively, be activated by certain of TRPV1, substance P or Isolectin B4 (Peier, A. M.
exogenous chemical substances (camphor, 2-APB) et al., 2002a).
(Chung, M. K. et al., 2004b; Hu, H. Z. et al., 2004; Another icilin-sensitive TRP channel that has been
Moqrich, A. et al., 2005). The expression pattern of implicated in cold sensation is TRPA1. This channel is
TRPV3 remains somewhat controversial. Although expressed in a subpopulation of small-diameter sen-
there are reports of TRPV3 mRNA or protein sory neurons and is commonly coexpressed with
expression in sensory neurons (Smith, G. D. et al., TRPV1. In some studies, recombinant TRPA1 has
2002; Xu, H. et al., 2002), its expression in these cells been reported to be responsive to intense cooling
is far less clear than in skin keratinocytes (Peier, A. M. (<18  C) (Story, G. M. et al., 2003). Consistent with
 Molecular Biology of the Nociceptor/Transduction 53

this finding, mice lacking TRPA1 were recently have no intrinsic voltage sensitivity and are active at
reported to exhibit reduced withdrawal behavior on resting membrane potentials thereby contributing to
a cold plate or in response to acetone-mediated cool- resting input resistance. TWIK related potassium
ing of the hind paw (Kwan, K. Y. et al., 2006). However, channel (TREK-1) was the first of these K channels
the cold sensitivity of TRPA1 is not universally to be identified in sensory neurons. TWIK stands for
accepted, since other investigators have failed to tandem of P domains in a weak inward rectifier
detect such responses and have reported normal potassium channel (Patel, A. J. and Honore, E.,
cold-evoked behavioral responses in a distinct line of 2001). TREK-1 was cloned in 1996 based on homol-
TRPA1 null mice (Jordt, S. E. et al., 2004). Another ogy to TWIK. While TREK-1 was originally shown
point of contention revolves around the potential con- to be activated by arachidonic acid and inhibited by
tributions of TRPA1 to mechanosensation, which is cAMP, it was subsequently demonstrated that
supported by immunolocalization studies (Nagata, K. TREK-1 was activated by membrane stretch, osmotic
et al., 2005) and by analysis of one knockout mouse line welling as well as molecules that caused membrane
(Kwan, K. Y. et al., 2006) but not another (Bautista, D. crenation. Two more mechanosensitive two-pore
M. et al., 2006). The basis for these apparent discre- potassium channels, TREK-2 and TRAAK were sub-
pancies remains unclear. However, there is uniform sequently described (Maingret, F. et al., 1999; Bang,
consensus that TRPA1 is a mediator of nociceptive H. et al., 2000), both of which are also activated by
responses to certain irritant chemicals, including mus- membrane stretch and crenators and both of which
tard oil (allyl isothiocyanate), cinnamaldehyde, and are present in sensory ganglia (Kang, D. and Kim, D.,
acrolein and that it serves as a downstream target 2006). Nevertheless, it remains to be determined
following the activation of bradykinin receptors whether any of these channels underlies the mechan-
(Bandell, M. et al., 2004; Jordt, S. E. et al., 2004; osensitivity of primary afferent neurons (Patel, A. J.
Bautista, D. M. et al., 2006; Kwan, K. Y. et al., 2006). and Honore, E., 2001).
Indeed, both lines of TRPA1 null mice were found to A second class of ion channel that may also
be defective in nocifensive behavioral responses to contribute to resting input resistance is a member of
mustard oil injection, mustard oil-evoked allodynia, the voltage-gated Na channel family. One member of
and bradykinin-evoked pain-like behavior (Bautista, this family, NaV1.9 appears to encode a voltage-gated
D. M. et al., 2006; Kwan, K. Y. et al., 2006). channel with unique gating properties (Cummins, T.
Thus far, the examination of TRP channels within R. et al., 1999). The channel has a very low threshold for
sensory neurons has been confined largely to those activation, activates very slowly upon membrane depo-
that either exhibit preferentially high expression in larization, and undergoes very little inactivation with
these cells or have been identified as being respon- small membrane depolarizations. These gating proper-
sive to chemical stimuli with recognizable sensory ties have two important consequences. First, it is likely
effects. However, other TRP channels are known to that this channel contributes little, if anything to the
be expressed quite broadly in neuronal and non- rapid upstroke of the action potential. Second, the
neuronal cells, where they participate in numerous channel should, in theory, contribute a depolarizing
functions that are important to sensory neuron biol- drive at resting membrane potentials (Herzog, R. I.
ogy (e.g., store-operated Ca2 entry, Mg2 uptake, et al., 2001). The problem with this theory is that
mechanosensation, axon guidance) (Ramsey, I. S. biophysical analysis of persistent Na currents in sen-
et al., 2006). Therefore, studies aimed at the analysis sory neurons (putative NaV1.9-mediated currents)
of those channels in sensory ganglia may yield (Priest, B. T. et al., 2005) indicate that this current is
further important insights. subject to a profound slow inactivation (Cummins, T. R.
et al., 1999). This voltage and time-dependent process
is associated with a channel transition from a closed
state, from which it could be activated with
4.2.2 Ion Channels Influencing Passive
membrane depolarization, to an inactivated state,
Membrane Properties: Membrane
from which the channel is no longer available for
Resistance
activation. The extent of slow inactivation of
While a number of channels contribute to the estab- NaV1.9 is such that there is virtually no detectable
lishment of input resistance, two general classes have persistent current evoked from a typical resting
been described at a molecular level. One class membrane potential (i.e., 60 mV). There is
includes the two-pore K channels. These channels evidence suggesting that the voltage dependence of
54 Molecular Biology of the Nociceptor/Transduction

slow inactivation is influenced by recording condi- MacDermott, A. B., 1997; Brock, J. A. et al., 1998;
tions, such that the use of fluoride-based electrode Khasar, S. G. et al., 1998), while NaV1.6 is present
solutions results is a hyperpolarizing shift in the along the axons (Rios, J. C. et al., 2003). Furthermore,
voltage dependence of this process (Coste, B. et al., there is evidence that channel expression is devel-
2004). Consequently, the extent of inactivation opmentally regulated. For example, in the rat,
may be overestimated when the whole-cell patch expression levels of NaV1.3 and 1.5 are high during
configuration is used, if stability of the internal milieu development, and then reduced to almost undetect-
is as important for the stability of NaV1.9 as it is for able levels after birth (Waxman, S. G. et al., 1994;
other ion channels (Kepplinger, K. J. et al., 2000). Renganathan, M. et al., 2002). Importantly, the pat-
Either way, what is clear is that even a few NaV1.9 tern of both - and -subunit expression is altered
channels active at rest could have a profound influ- both within and between neurons following injury,
ence on neuronal excitability (Herzog, R. I. et al., changes that are thought to play a significant role in
2001). pain and hyperalgesia associated with tissue injury.
Voltage-gated Na currents in sensory neurons
are generally described in terms of their sensitivity
4.2.3 Ion Channels Underlying Active
to tetrodotoxin (TTX) where currents are TTX-
Membrane Processes
sensitive (NaV1.1, 1.2, 1.3, 1.6, and 1.7), TTX-insen-
4.2.3.1 Action potential upstroke sitive (NaV1.5), or TTX-resistant (NaV1.8 and 1.9).
Voltage-gated Na channels are critical for the The TTX-resistant currents are further subdivided
upstroke, or rapid depolarization, phase of the neu- into inactivating (NaV1.8) and persistent current
ronal action potential. Typical voltage-gated Na (NaV1.9). Based on data from biophysical character-
channels consist of an -subunit and two -subunits ization of these subunits and toxin sensitivity,
(Catterall, W. A., 1992). The -subunit contains NaV1.8 appears to be largely responsible for spike
everything necessary for a functional voltage-gated initiation in the terminals and cell body of nocicep-
channel, including the ion pore, voltage sensor, and tive afferents (Gold, M. S., 2000a), while the TTX-
inactivation gate. -subunits appear to both influence sensitive channels, NaV1.6 and NaV1.7, are likely to
channel gating as well as play a major role in channel underlying action potential conduction. Recent evi-
trafficking (Isom, L. L., 2001). Recent evidence sug- dence from the NaV1.7 null mutant mouse suggests
gests that one of the -subunits may also influence that NaV1.7 may also play a significant role in
channel inactivation (Grieco, T. M. et al., 2005). Nine action potential generation in nociceptive afferent
-subunits have been cloned and clearly demon- terminals (Nassar, M. A. et al., 2004). A small cohort
strated to underlie voltage-gated Na currents of individuals was also recently identified who do
(Catterall, W. A. et al., 2005a). A tenth -subunit has not experience pain in response to noxious stimuli,
been identified that is distantly related to the other even though they appear to be aware of stimuli that
nine and has been suggested to function as a Na should produce pain, exhibiting few, if any detect-
transporter rather than a voltage-gated channel able somatosensory abnormalities (Cox, J. J. et al.,
(Goldin, A. L. et al., 2000). Four -subunits have 2006). All of these individuals possess loss of func-
been cloned (Goldin, A. L. et al., 2000; Yu, F. H. tion mutations in NaV1.7, indicating that this
et al., 2003). There is evidence that eight of the nine channel is necessary for the stimulus evoked percep-
-subunits and all four -subunits are present in tion of pain.
sensory neurons (Black, J. A. et al., 1996; Shah, B. S.
et al., 2000; Renganathan, M. et al., 2002; Takahashi, N. 4.2.3.2 Action potential downstroke
et al., 2003). -subunits are differentially distributed The falling phase of the action potential is the result of
both between and within sensory neurons. For exam- two processes. One is the inactivation of voltage-gated
ple, NaV1.7, 1.8, and 1.9 are preferentially expressed Na channels and the second is the activation of K
in putative nociceptive afferents (Fang, X. et al., 2002; channels. One class of channels that plays a major role
Djouhri, L. et al., 2003a; 2003b), while NaV1.1 is in controlling the rate of membrane repolarization are
preferentially expressed in non-nociceptive afferents the sustained (i.e., delayed rectifier type) voltage-gated
(Black, J. A. et al., 1996). Within nociceptive afferents, K channels (Werz, M. A. and MacDonald, R. L., 1983).
NaV1.8 appears to be normally localized to the cell There appear to be at least three types of sustained
body and terminal arbor (Ritter, A. M. and Mendell, voltage-gated K currents present in sensory neurons
L. M., 1992; Jeftinija, S., 1994; Gu, J. G. and based on biophysical and pharmacological properties
 Molecular Biology of the Nociceptor/Transduction 55

(Gold, M. S. et al., 1996c). While the K channel sub- feature of nociceptive afferents is that the somatic
units underlying these currents have yet to be action potential is associated with a relatively long
conclusively identified, immunohistochemical data AHP (McLachlan, E. M. et al., 1993; Villiere, V. and
have implicated several possible candidates (Rasband, McLachlan, E. M., 1996; Djouhri, L. et al., 2003a). A
M. N. et al., 1998). number of distinct currents have been suggested to
Another class of channel that contributes to mem- influence the magnitude and duration of the AHP.
brane repolarization is the large conductance Ca2- Furthermore, because of differences between cur-
modulated K channel, (i.e., BK or Maxi-K channel). rents with respect to biophysical properties of
These channels are encoded by a single -subunit channel gating, changes in intracellular Ca2 con-
(slo, which has several splice variants), whose gating centrations, etc., the relative contribution of different
properties, Ca2 sensitivity, and pharmacological ion channels to the AHP changes with time. Because
sensitivity is influenced by the presence of one of of this feature, investigators have often divided the
four distinct -subunits (Wallner, M. et al., 1998). AHP into fast (early), intermediate, and slow (late)
While these channels have intrinsic voltage sensitiv- AHP. Channels mediating the fast AHP are generally
ity, the voltage dependence of activation is such that those that contributed to the downstroke of the
in the absence of Ca2, it is unlikely that these action potential (i.e., sustained voltage-gated K
channels would be sufficiently activated to contribute and BK currents). These channels generally deacti-
to the downstroke of the action potential (Wallner, vate (i.e., close) rapidly and therefore do not
M. et al., 1998). This suggests that there is sufficient generally contribute to the AHP for more than sev-
Ca2 entry via voltage-gated Ca2 channels during eral of milliseconds (Fowler, J. C. et al., 1985a).
the action potential (Blair, N. T. and Bean, B. P., The hyperpolarization associated with the fast
2002), to enable activation of BK channels. The nat- AHP may result in the activation of three additional
ure of the association between BK channels and types of currents that have opposing influences on
voltage-gated Ca2 channels in afferent terminals membrane potential. First, there may be the activa-
has yet to be described. However, the timing of BK tion of inward rectifying K currents (Scroggs, R. S.
current activation in the cell body suggests that there et al., 1994). These currents appear to activate and
is a relatively tight association between the two. inactivate rapidly in sensory neurons and therefore
appear to primarily influence the fast AHP. Second,
4.2.3.3 After potential there may be the activation of low-threshold inacti-
The after potential is the change in membrane poten- vating or A-type voltage-gated K currents
tial that follows an action potential. This is generally (Cardenas, C. G. et al., 1995). These channels are
a membrane hyperpolarization (more negative than largely inactivated at rest. However, there may be
resting membrane potential), but may also be a depo- enough membrane hyperpolarization associated with
larizing after potential (DAP). Two different the fast AHP to enable recovery from inactivation of
mechanisms have been described that contribute to a number of A-type K channels. Because these
the DAP. One mechanism involves a high density of channels have a low threshold for activation, once
low threshold, or T-type voltage-gated Ca2 chan- recovered from inactivation, the channels will open
nels (White, G. et al., 1989). The biophysical and therefore prolong the decay of the AHP. Because
properties of these channels are such that if present A-type currents in sensory neurons may inactivate
at a high enough density, they are able to depolarize slowly (i.e., with time constants of decay will over
the membrane following an action potential, provid- 100 ms (Gold, M. S. et al., 1996c)), these channels can
ing enough depolarizing drive to evoke a burst of and do contribute significantly to the intermediate
action potentials. The second current suggested to AHP (Harriott, A. et al., 2006). As such, these channels
contribute to a DAP is a Ca2-dependent Cl cur- play a significant role in regulating interspike inter-
rent (Mayer, M., 1985). As the name implies, val (Connor, J. A. and Stevens, C. F., 1971;
activation of this current requires sufficient Ca2 Yoshimura, N. and de Groat, W. C., 1999).
entry during the action potential. It also requires Importantly, persistent inflammation and nerve
the intracellular Cl concentration is such that the injury have been shown to result in a decrease in
equilibrium concentration for Cl is depolarized. these currents, which is associated with a concomi-
In the majority of nociceptive sensory neurons, tant increase in membrane excitability (Yoshimura, N.
the somatic action potential is followed by an after and de Groat, W. C., 1999; Moore, B. A. et al., 2002;
hyperpolarization (AHP). In fact, a distinguishing Stewart, T. et al., 2003; Dang, K. et al., 2004; Harriott, A.
56 Molecular Biology of the Nociceptor/Transduction

et al., 2006). Third, there may be the activation of experiment suggesting that others are expressed as
hyperpolarization-activated cationic current (Ih). well (Gold, M. S., unpublished observation). It
These currents, carried by family channels called remains to be determined which if any of the SK
hyperpolarization-activated cyclic nucleotide-gated channels mediates the slow AHP given several strik-
(HCN) channels, are nonselective cationic currents ing differences between the SK channels and the
(Robinson, R. B. and Siegelbaum, S. A., 2003). slow AHP in terms of pharmacological sensitivity,
Consequently, activation of these channels results in Ca2 dependence and gating properties (Cordoba-
a depolarizing current, which, if of sufficient magni- Rodriguez, R. et al., 1999).
tude, may contribute to spike initiation. A high
density of these channels can result in membrane 4.2.3.4 Action potential threshold
oscillations and spontaneous activity. Indeed, there The action potential threshold is where the various
is evidence that these currents are upregulated in the influences of all the ion channels in the membrane
presence of injury, a change that has been suggested come into play. The magnitude and nature of leak
to contribute to spontaneous afferent activity conductance determines membrane resistance and
observed following nerve injury (Chaplan, S. R. the impact a generator current will have on the
et al., 2003). Furthermore, there is evidence that membrane potential. The biophysical properties of
blocking these channels attenuates mechanical voltage-gated K currents will determine the magni-
hypersensitivity observed following nerve injury tude of the K conductance associated with
(Lee, D. H. et al., 2005a). These channels are also membrane depolarization as well as the impact of
modulated by cyclic nucleotides (Ingram, S. L. and resting membrane potential on that value. Similarly,
Williams, J. T., 1994; Robinson, R. B. and the biophysical properties of voltage-gated Na cur-
Siegelbaum, S. A., 2003), which suggests that they rents will determine the voltage at which these
may contribute to the increased excitability observed channels begin to open, the rate at which they open
in the presence of inflammation as well, given that and the rate at which they inactivate in the face of
many inflammatory mediators induce an increase in slow membrane depolarizations. Given the fact that
cAMP concentrations (Levine, J. D. et al., 1993). there is considerable variation in the biophysical
In a subpopulation of sensory neurons, a slow properties of Na channels depending on which -
AHP has also been described (Fowler, J. C. et al., and -subunits are present, the relative density of
1985a; 1985b; Weinreich, D. and Wonderlin, W. F., these subunits will have a significant impact on action
1987). This AHP apparently reflects the activation potential threshold (Matzner, O. and Devor, M.,
of a Ca2-dependent K channel that takes several 1992).
hundred milliseconds to activate, but once activated Another class of ion channels that may contribute to
takes several seconds to close (Cordoba-Rodriguez, the action potential threshold are the low threshold
R. et al., 1999). Consequently, activation of these (i.e., T-type) voltage-gated Ca channels. As with
channels has a profound influence of spike accom- voltage-gated Na channels, T-type channels consist
modation. The slow AHP has been well described in of an -subunit that contains the channel-pore,
nodose ganglion neurons, where it appears to be a voltage-sensing, and inactivation gates, and ancillary
target for inflammatory mediators that are able to subunits that influence channel gating and trafficking
block it in a cAMP dependent manner. The result is (Catterall, W. A. et al., 2005b). Three -subunit genes
a dramatic increase in excitability (Weinreich, D. have been cloned that encode for T-type currents.
and Wonderlin, W. F., 1987). While there is evi- T-type currents appear to be present in a low density
dence that a similar channel may be present in DRG in putative nociceptive afferents (Scroggs, R. S. and
neurons, it does not appear to play a major role Fox, A. P., 1992). While the activation rate of these
in inflammatory mediator-induced sensitization of channels does not appear to be sufficient to enable
nociceptive afferents (Gold, M. S. et al., 1996d). these channels to contribute to the action potential
Four Ca2-dependent but voltage-independent upstroke, the low threshold for activation enables
channels have been cloned and are referred to as these currents to amplify the effects of small membrane
SK14, in reference to their small single-channel depolarizations. Consequently, an increase in the den-
conductance (Bond, C. T. et al., 2005). There is sity of these channels can have a significant influence
immunohistochemical evidence for the presence of on membrane excitability. In support of this suggestion,
these SK1 in sensory neurons (Boettger, M. K. et al., these channels have recently been implicated in epi-
2002) and we have data from a single-cell PCR leptogenisis (Czapinski, P. et al., 2005). More recent
 Molecular Biology of the Nociceptor/Transduction 57

evidence suggests that they may contribute to the terminus, signal by catalyzing the dissociation of
sensitization of nociceptive afferents in the presence guanosine diphosphate (GDP) from the -subunit
of tissue injury (Barton, M. E. et al., 2005). of heterotrimeric G proteins associated with the
intracellular face of the plasma membrane. As a con-
4.2.3.5 High-threshold voltage-gated sequence of its dissociation, GDP is replaced with
Ca 2 current guanosine triphosphate (GTP), causing the hetero-
The structure of high-threshold voltage-gated Ca2 trimer to dissociate into a GTP-bound -subunit and
channels is analogous to that of voltage-gated Na a free  dimer. Both the GTP-bound -subunit and
channels and T-type Ca2 channels, with a single the free  dimer have specific cellular targets
-subunit that contains the ion pore, voltage-sensor, through which they are able to activate or inhibit
etc., and several different ancillary subunits that signaling cascades and/or directly modify the activ-
influence channel gating (i.e., the -subunit) and/ ity of specific ion channels (Gilman, A. G., 1987;
or channel trafficking (i.e., the 2-subunit complex) Pierce, K. L. et al., 2002). This versatile signal trans-
(Catterall, W. A. et al., 2005b). Seven -subunits duction strategy is used by at least 1000 different
encoding high-threshold channels have been cloned, GPCRs expressed throughout the body, and dozens
resulting in four currents that are distinguishable on of these are expressed by nociceptors and other
the basis of unique pharmacological sensitivity. sensory neurons. A few notable examples include
Molecular biological and pharmacological analyses B1 and B2 receptors for bradykinin (Steranka, L. R.
indicate that all four current types are likely present et al., 1988), P2Y2 receptors for ATP (Tominaga, M.
in nociceptive afferents (Kostyuk, P. G. et al., 1981; et al., 2001), PAR2 protease-activated receptors
Scroggs, R. S. and Fox, A. P., 1992; Kim, D. S. et al., (Vergnolle, N. et al., 2001), EP1 EP3C and EP4
2001; Yusaf, S. P. et al., 2001; Wu, Z. Z. et al., 2004b). receptors for prostaglandin E2 (Ferreira, S. H. et al.,
While these channels are critical for transmitter 1978; Sugimoto, Y. et al., 1994; Oida, H. et al., 1995),
release, their role in direct regulation of excitability mu, kappa, and delta opioid receptors (Dado, R. J.
is less clear. On the one hand, they enable inward et al., 1993; Arvidsson, U. et al., 1995), several subtypes
current that contributes to membrane depolarization of metabotropic glutamate receptor (Bhave, G. et al.,
and prolongation of the action potential. On the 2001; Yang, D. and Gereau, R. W. T., 2002), and at
other hand, they enable activation of K and Cl least seven subtypes of 5HT receptor (Nicholson, R.
currents that may be inhibitory. Thus, the site of et al., 2003). One recently discovered family of appar-
channel activation is critical. Of note, in studies ently pronociceptive GPCRs are those of the so-called
involving the isolated sensory neuron cell body, mrg or sensory neuron specific (SNS) family (Dong, X.
there is evidence to suggest that inflammatory med- et al., 2001; Lembo, P. M. et al., 2002; Zylka, M. J.
iators decrease the density of high-threshold et al., 2003). The expression pattern of this GPCR
voltage-gated currents (Borgland, S. L. et al., 2002), family exhibits remarkable species-specific heteroge-
and in the presence of nerve injury, there is a neity, as there appear to be at least 50 members
decrease in the density of high-threshold voltage- in mice, but only four in rat and only a few distantly
gated currents as well (Baccei, M. L. and Kocsis, J. related homolog in humans (Zylka, M. J. et al., 2003).
D., 2000; Hogan, Q. H. et al., 2000; Kim, D. S. et al., GPCRs expressed in sensory ganglia can be
2001). Authors of both the studies suggested that broadly classified into those that enhance nociceptor
these changes may contribute to pain associated excitability and those that diminish it. Sensitizing
with tissue injury and all are certainly correct on receptors tend to couple to either Gq (e.g., B2,
this point. P2Y2, PAR2, and EP1) or Gs (e.g., EP3C, EP4).
The activated -subunit of Gq activates phospholi-
pase C (PLC), an enzyme that catalyzes cleavage of
4.2.4 Membrane Receptors that Indirectly
the membrane phospholipid phosphatidylinositol
Alter Nociceptor Function
bisphosphate to inositol 1,3,5-trisphosphate (IP3)
4.2.4.1 G-protein-coupled receptors and diacylglycerol (DAG). IP3 evokes the release of
Many of the cell surface proteins that define the Ca2 ions from intracellular stores that, together with
functional and histological identity of sensory neu- DAG, go on to trigger the plasma membrane locali-
rons are the so-called GPCRs. These proteins, which zation and activation of protein kinase C (PKC)
possess seven transmembrane domains, an extracel- (Gilman, A. G., 1987). Multiple PKC isoforms (, ,
lular amino terminus, and an intracellular carboxyl ", and ) have been shown to facilitate nociceptor
58 Molecular Biology of the Nociceptor/Transduction

sensitization through their phosphorylation of ion functions is the nerve growth factor (NGF) family
channels and other target proteins (Cesare, P. et al., (NGF, BDNF, neurotrophin-3, and neurotrophin-4).
1999; Khasar, S. G. et al., 1999; Olah, Z. et al., 2002; The receptors for this family of nociceptors fall into
Wang, Y. et al., 2004). Ca2 ions can also act in many two categories: a low-affinity receptor p75, which
other ways, such as by binding to proteins such as appears to bind all members of the family, and homo-
calmodulin that can allosterically regulate ion chan- dimeric high-affinity receptors that are specific for
nels and intracellular enzymes in a Ca2-dependent each of the family members (TrkA, TrkB, or TrkC)
manner (Numazaki, M. et al., 2003). The GTP-bound (Huang, E. J. and Reichardt, L. F., 2001). Interestingly,
Gs subunit activates the enzyme adenylyl cyclase, one p75 monomer and one Trk dimer can apparently
which catalyzes the generation of cyclic adenosine bind the same neurotrophin dimer simultaneously
monophosphate (cAMP) from ATP (Gilman, A. G., (He, X. L. and Garcia, K. C., 2004). During develop-
1987). cAMP, in turn, activates cAMP-dependent ment, all sensory neurons destined to become
protein kinase (protein kinase A, PKA), which can nociceptors are dependent on NGF for survival.
also phosphorylate and augment the responsiveness However, with maturation, only a subset of nocicep-
of certain ligand-, heat-, or voltage-gated channels tive afferents continues to express the high-affinity
(Gold, M. S. et al., 1998; Bhave, G. and Gereau, R. W. NGF receptor, TrkA. In these neurons, continued
T., 2004). Another intracellular enzyme recently access to NGF is no longer necessary for survival,
implicated in nociception is the cAMP-activated but remains essential for the maintenance of the affer-
guanine nucleotide exchange protein, Epac, which ent phenotype, which includes peptide expression and
couples intracellular cAMP generation to the PLC- the specific composition of ion channels and transdu-
dependent sensitization of TTX-resistant sodium cers. Afferents have continued access to NGF
channels (Hucho, T. B. et al., 2005). Interestingly, produced by target tissue, but the amount of NGF
although all of the actions of pronociceptive available is critical: a decrease in access to NGF
GPCRs on excitability are indirect, through their appears to result in the loss of nociceptive phenotype,
actions on downstream signaling mechanisms, activa- while increased access to NGF results in an increase in
tion of some (e.g., bradykinin receptors) is sufficient the expression of nociceptive markers (Koltzenburg,
to trigger action potential firing, whereas activation M., 1999). BDNF and neurotrophin-3 can also alter
of others (e.g., prostaglandin receptors) primarily nociceptor function (Shu, X. Q. and Mendell, M. L.,
enhances neuronal responsiveness to other exogenous 1999).
stimuli (Birrell, G. J. et al., 1991; 1993; Kumazawa, T. Transduction of the signal arising from the bind-
et al., 1996). This distinction may reflect differences ing of NGF family members to their receptors
in signal amplification or differences in the proximity consists of both slow processes requiring internaliza-
of a given GPCR, its target enzymes, and a given tion of the neurotrophinreceptor complex and rapid
depolarizing ion channel. signaling processes that occur at the nociceptor term-
In contrast to the pronociceptive receptors, inal. In both cases, NGF binding drives the
GPCRs that couple to Gi or Go proteins (e.g., m- dimerization of the TrkA receptor, resulting in
opioid receptor, type II metabotropic glutamate TrkA autophosphorylation and the recruitment of a
receptors) tend to suppress neuronal activity. This complex of molecules that signal through distinct
outcome results from inhibition of adenylyl cyclase enzymatic pathways. These include, but are not lim-
(to diminish PKA phosphorylation of target pro- ited to, pathways involving phospho-inositol 3 kinase
teins), inhibition of voltage-gated Ca2 channels, (PI3K), ras-mitogen-associated protein kinase
and/or activation of hyperpolarizing G-protein- (MAPK), and PLC . The activated complex with
coupled inwardly rectifying K channels (GIRKs of NGF-bound TrkA at its core is internalized into a
the Kv4 family) (McFadzean, I., 1988; Bhave, G. and specialized endosomal vesicle and transported along
Gereau, R. W. T., 2004). microtubules to the cell body, where signaling from
this complex can trigger changes in transcription,
translation, and protein trafficking (Zweifel, L. S.
4.2.4.2 Neurotrophin receptors et al., 2005). For many years, this slow retrograde
As described in detail elsewhere in this chapter, neu- process was viewed as the predominant mode of
rotrophins are also important regulators of sensory neurotrophin signaling in sensory neurons. More
neuron development, survival, and excitability. One recently, however, it has become evident that
major class of neurotrophic factors that carry out these NGFTrkA activation of PI3K, MAPK, and PLC
 Molecular Biology of the Nociceptor/Transduction 59

produces rapid effects at the nociceptor terminal However, a full mechanistic understanding of noci-
by altering ion channel sensitivity and plasma ceptive signal transduction, action potential
membrane localization within seconds, chiefly via generation, and spike propagation will require that
receptor phosphorylation (Shu, X. and Mendell, pain biologists move beyond a mere itemization of
M. L., 2001; Bonnington, J. K. and McNaughton, the contents of this toolbox to an appreciation of the
P. A., 2003; Zhang, X. et al., 2005). These events will additive and emergent properties that can arise from
be described in greater detail in the final section of the polymodal nature of some of these molecules,
this chapter. The p75 receptor is associated with their posttranslational modulation, their interac-
the activation of a distinct set of second messenger tions, and their spatial distribution. In this section,
pathways, including one involving the activation of we attempt to illustrate these principles using a few
sphingomyelinase and the liberation of ceramide specific examples, some well established, and others
(Zhang, Y. H. et al., 2002; Zhang, Y. H. and Nicol, more speculative in nature.
G. D., 2004).
Nociceptive afferents expressing TrkA constitute
4.3.2 Shared Modalities and
a subpopulation of nociceptive afferents. Other sub-
Polymodality
populations of nociceptive afferents express
receptors for other neurotrophic factors. For exam- One source of signaling complexity and heterogene-
ple, one subpopulation of nociceptive afferents that ity among sensory neurons is the existence of
do not express the neuropeptides substance P or multiple transducers for a given stimulus modality.
CGRP, lose their NGF dependence during early For example, as described above, at least six TRP
postnatal life and begin to express a receptor for channels expressed in sensory neurons can be gated
glial cell-derived neurotrophic factor (GDNF) by heat or cold. Although each channel exhibits a
(Molliver, D. C. et al., 1997). GDNF is a member of characteristic temperature range over which it is
a second family of neurotrophins that includes neu- thermally responsive, in many cases these ranges
turin, artemin, and persephin. As with NGF in TrkA- overlap, theoretically allowing continuous recogni-
expressing neurons, the amount of GDNF to which tion of the thermal environment. At the same time,
these nonpeptidergic nociceptive afferents have the largely nonoverlapping expression patterns of
access influences their phenotypic properties (see TRPM8, TRPV1, and TRPV2 lend some credence
Fjell, J. et al., 1999a). GDNF signaling involves a to the labeled-line hypothesis, which states that
specific GDNF family receptor (GFR), which acts distinct sensory modalities (innocuous cold, pain,
in conjunction with another receptor tyrosine kinase, itch, etc.) are conveyed by anatomically distinct
ret, that is shared by all members of this family. subsets of sensory afferents with specific central
Interestingly, GDNF signaling involves second mes- projection patterns (Craig, A. D., 2002). However, it
senger pathways similar to those underling the should be noted that this segregation of thermotrans-
actions of NGF, including PLC-, MAPK, and duction mechanisms is not absolute. For example,
PI3K (Airaksinen, M. S. and Saarma, M., 2002). some polymodal nociceptors exhibit paradoxical
Thus, the phenotype of subpopulations of nocicep- dual responsiveness to both intense heat and
tive afferents is regulated by different receptor intense cold (Dodt, E. and Zotterman, Y., 1952).
systems that appear to ultimately use similar trans- Even at the molecular level, TRPV1 and TRPA1
duction cascades. are coexpressed in a significant fraction of small-
diameter sensory neurons. Though the implications
for thermosensation remain controversial, such coex-
pression appears to be critical for full responsiveness
4.3 Additive and Emergent to bradykinin, perhaps owing to the potentiation of
Properties of Nociceptor Membrane TRPA1 by Ca2 ions flowing through TRPV1
Proteins (Bautista, D. M. et al., 2006). Thus, while the labeled
line hypothesis may adequately address some aspects
4.3.1 Introduction
of thermosensation, the overall situation may be
It should be evident from the preceding section that more complex.
sensory afferents have, at their disposal, an exten- Mechanotransduction represents an even more
sive repertoire of cell surface proteins with diverse extreme example of likely complementarity and
stimulus specificities and signaling capabilities. redundancy among transducing molecules. Members
60 Molecular Biology of the Nociceptor/Transduction

of both the ASIC and the TRP families have TRPM8, itself (Rohacs, T. et al., 2005). Similarly,
been implicated in this process, but only modest def- TRPV1 can activate MAPK signaling pathways
icits in mechanotransduction have been reported in (Zhuang, Z. Y. et al., 2004). An additional interaction
animals lacking any single putative mechanosensory mechanism that has been reported in other types of
protein (Price, M. P. et al., 2000; Liedtke, W. and sensory neurons, but could also conceivably occur in
Friedman, J. M., 2003; Suzuki, M. et al., 2003; Page, nociceptors, is the stimulation of Ca2-activated K
A. J. et al., 2004; Kwan, K. Y. et al., 2006). One channels by the Ca2 ions that flow through ligand-
interpretation of these findings is that mechano- gated ion channels (Yuhas, W. A. and Fuchs, P.A.,
transduction involves complexes of intracellular, 1999; Isaacson, J. S. and Murphy, G. J., 2001). This
transmembrane, and extracellular molecules, and arrangement has the potential to shape the ampli-
that multiple interchangeable ion channel subunits tude, duration, and even the direction of generator
can contribute to the ion permeation pathways of potential.
these transduction complexes. Active membrane properties are also strongly
Another source of complexity in sensory nerve dependent upon the interaction, at multiple levels,
signal transduction is responsiveness to multiple between the ion channels and the other membrane
modalities within a single transduction molecule. proteins underlying transduction and those regulat-
One of the best-studied examples of this phenom- ing action potential initiation and propagation. For
enon is the capsaicin receptor, TRPV1. This example, even small changes in resting membrane
channel protein can be activated by a diverse list potential or generator potential will dramatically
of chemical or thermal stimuli (Caterina, M. J. and influence the availability of different voltage-gated
Julius, D., 2001). In addition, there is evidence that ion channels. Similarly, the second messenger path-
this channel or its splice variants may participate in ways activated by GPCRs and neurotrophin
osmotic or mechanical sensitivity in osmosensory receptors can impact many facets of action potential
regions of the brain and in non-neuronal cells of generation and propagation, as described in the final
the urinary bladder (Birder, L. A., 2005; Naeini, R. section of this chapter. The spatial distribution of
S. et al., 2006). Such polymodal responsiveness pro- ion channels underlying each of the steps in afferent
vides ample opportunities for convergent stimuli to signaling is another critical determinant of whether
combine additively or even supraadditively to pro- or not there will be successful transmission of sen-
duce greater nociceptor activation than any single sory information. The ion channels underlying spike
stimulus. For example, even a modest reduction in initiation must be close enough to the ion channels
pH (to 6.4) can drop the apparent thermal threshold underlying stimulus transduction that the generator
for TRPV1 activation to near body temperature, potential pushes the membrane potential above the
possibly leading to spontaneous pain in the context action potential threshold (Figure 1). Similarly, the
of inflammation or ischemia (Tominaga, M. et al., ion channels underlying action potential propaga-
1998). tion must be close enough to the site of spike
initiation, that the action potential is faithfully pro-
pagated toward the central nervous system. In
4.3.3 Functional Interactions between
addition, action potential threshold may be rela-
Nociceptor Proteins
tively high in nociceptors that utilize the high-
Interactions between different proteins on the surface threshold voltage-gated Na channel NaV1.8 for
of the nociceptor terminal can also have profound spike initiation, whereas action potential burst dura-
effects on nociceptors transduction and therefore tion may be relatively short in nociceptors in which
function. Sometimes, these interactions are direct, as Ca2-dependent K channels are present in high
in the case of ion channel or GPCR heteromultimer- densities.
ization. In other cases, second messenger molecules One implication of the precise nature of these
and intracellular enzymes facilitate functional inter- spatial relationships between of ion channels within
actions among membrane proteins. Recently, for sensory neurons is that cellular processes underlying
example, it was demonstrated that TRPM8 can acti- the trafficking and anchoring of ion channels within
vate Ca2-dependent phospholipases that, in turn, sensory neurons must be tightly regulated. While
cleave the phosphatidyl inositol bisphosphate that there is much to be learned about the distribution
inhibits or activates a host of channels, including and anchoring of ion channels in sensory neurons,
inwardly rectifying potassium channels, as well as the observation that it is possible to induce modality
 Molecular Biology of the Nociceptor/Transduction 61

specific sensitization in polymodal nociceptors, 4.4.2 Inflammation-Induced Changes

raises the possibility that ion channels and/or trans- in Nociceptor Sensory function
ducers are differentially distributed throughout the Inflammatory Pain
terminal arbor. Adaptor/scaffold proteins can also
One of the best examples of injury-induced changes
shape nociceptor responsiveness by bridging ion
in the transduction properties of nociceptive neurons
channels or GPCRs with intracellular enzymes or
is the enhanced responsiveness to heat that occurs
with proteins that regulate intracellular transport. following tissue inflammation. As indicated in
For example, PICK1, a PDZ domain-containing Section 4.2 of this chapter, the thermal hyperalgesia
protein, appears to regulate the subcellular traffick- associated with several distinct inflammatory insults
ing of ASIC channel subunits (Duggan, A. et al., (CFA, carageenan, and mustard oil) or specific
2002), while annexin has been demonstrated to reg- inflammatory mediators (bradykinin, NGF, ATP,
ulate surface expression and function of NaV1.8 proteases, and prostaglandin E2) depends strongly
(Okuse, K. et al., 2002). on the presence of TRPV1. Consistent with this
In summary, signaling heterogeneity, conver- observation, TRPV1 ion channel activity can be sen-
gence of function, proteinprotein interactions, and sitized by these various agents in dissociated DRG
cytosolic signaling, as well as spatiotemporal organi- neurons or heterologous expression systems
zation of ion channels allow an already complex set (Lopshire, J. C. and Nicol, G. D., 1998; Shu,X. and
of membrane proteins to vastly expand the sensory Mendell, M. L., 2001; Vellani, V. et al., 2001;
and coding capacity of nociceptive neurons. Bonnington, J. K. and McNaughton, P. A., 2003).
One lesson emerging from these studies, however,
is that TRPV1 sensitization by inflammatory media-
tors can proceed via a number of alternative
4.4 Injury-Induced Plasticity in mechanisms.
Transduction The most direct mechanism by which inflamma-
4.4.1 Introduction tory mediators can sensitize TRPV1 is by evoking
the generation or liberation of TRPV1 coagonists,
Research into the underlying mechanisms of pain such as protons (Caterina, M. J. et al., 1997) or arachi-
associated with tissue injury has revealed that while donic acid metabolites (Hwang, S. W. et al., 2000;
there are a number of key players that are involved Huang, S. M. et al., 2002). Convergent interactions
with a wide range of injury types the nature of the of TRPV1 with multiple stimulators result in channel
contribution of these players varies as a function of activity levels greater than those evoked by one
the type of injury. For example, acute, phosphoryla- agonist alone. Another extensively studied TRPV1
tion-dependent modulation of the voltage-gated Na sensitization mechanism involves direct phosphory-
channel NaV1.8 (Fitzgerald, E. M. et al., 1999), results lation of this channel by serine/threonine kinases
in an increase in current that contributes to an (PKC, PKA, Ca2/calmodulin-dependent protein
inflammation-induced increase in nociceptors excit- kinase II) (Bhave, G. and Gereau, R. W. T., 2004) or
ability (England, S. et al., 1996; Gold, M. S. et al., the src tyrosine kinase (Zhang, X. et al., 2005).
1996b). In contrast, following traumatic nerve injury, Although there is some overlap in their recognition
redistribution of NaV1.8 to the axons of uninjured sequences, each kinase apparently targets a different
afferents appears to be necessary for the expression of collection of phosphorylation sites, resulting in the
mechanical hypersensitivity associated with nerve generation of distinct phospho-TRPV1 species. In
injury (Gold, M. S. et al., 2003). There are many the case of ser/thr kinases, phosphorylation appears
such examples, a number of which have been alluded to enhance TRPV1 sensitivity to its chemical and
to in the preceding discussion. However, while it is thermal agonists and/or reverse TRPV1 desensitiza-
beyond the scope of the present chapter to detail the tion (Bhave, G. and Gereau, R. W. T., 2004). In
array of injury-induced changes in mechanisms contrast, TRPV1 phosphorylation by src kinase,
underlying stimulus transduction, it is informative which is activated by NGFtrkA signaling, leads
to consider several specific examples of modulatory within 10 min to a translocation of TRPV1 from an
events that occur in the context of inflammation and intracellular compartment to the plasma membrane,
nerve injury. A few of these mechanisms are illu- resulting in an increase in the overall number of
strated schematically in Figure 1. TRPV1 channels on the cell surface (Zhang, X.
62 Molecular Biology of the Nociceptor/Transduction

et al., 2005). The MAPK, p38, has also been shown to both injured afferents and their uninjured neighbors
participate in NGF-stimulated facilitation of (Gold, M. S., 2000b). Most compelling are the obser-
TRPV1 signaling during inflammation. Although vations that ongoing neuropathic pain may be
the relevant target of p38 phosphorylation is not blocked with the administration of local anesthetics
known, the end result is an apparent enhancement (Gracely, R. H. et al., 1992). However, there is also a
of TRPV1 translation and/or trafficking to periph- very tight correlation between the development of
eral nociceptor terminals that requires 12 days (Ji, R. nociceptive behaviors and the emergence of ongoing
R. et al., 2002). afferent activity in animal models of neuropathic
A third general mechanism by which TRPV1 can pain (Liu. C. N. et al., 2000). Because of this associa-
be modulated in the context of inflammation is by tion between ongoing afferent activity and ongoing
alterations in membrane phospholipid content. For pain, considerable effort has been dedicated to the
example, it has been proposed that phosphatidylinosi- identification of mechanisms that may contribute to
tol bisphosphate exerts a tonic inhibition on TRPV1 this activity. Central to this search is the question of
activity and that its hydrolysis by PLC relieves that whether the activity is driven by stimulus transduc-
inhibition (Chuang, H. H. et al., 2001; Prescott, E. D. tion, or whether there are changes in the relative
and Julius, D., 2003). The effects of PIP2 on TRPV1 contribution of ion channels resulting in membrane
may be complex, however, given that this phospholi- potential instability, the emergence or amplification
pid is also required for resensitization of TRPV1 of membrane potential oscillations that are capable of
following agonist-dependent desensitization (Liu, B. driving action potential generation. There is evi-
et al., 2005). More recently, work from several labora- dence for both sources of activity.
tories has provided evidence that phosphorylation of
PIP2 to PIP3 by PI3 kinase to form PIP3 represents 4.4.3.1 Nerve injury-induced changes in
yet another mechanism for TRPV1 sensitization transduction
(Bonnington, J. K. and McNaughton, P. A., 2003; Several nerve injury-induced changes in transduction
Zhuang, Z. Y. et al., 2004; Zhang, X. et al., 2005). have been identified. These can be loosely grouped into
Although the precise basis for this effect remains those involving aberrant expression of transducers nor-
unclear, one intriguing suggestion is that it is due to mally present in peripheral nerve, aberrant coupling
the effects of PIP3 on src activity, thus potentially between the transducers and cellular pathways and the
linking lipid-based and protein kinase-based sensitiza- emergence of inappropriate sources of stimulation. The
tion events (Zhuang, Z. Y. et al., 2004). Much remains list of tranducers aberrantly expressed following nerve
to be learned about the relative importance of and injury has grown as fast as transducers have been iden-
interactions between these different processes. tified. Aberrant expression comes in at least three forms.
However, it is clear that nociceptors have developed One is the expression of transducers in the wrong place
multiple means of responding to tissue inflammation within a neuron. The most dramatic example of this is
by becoming hypersensitive. the emergence of mechanical and thermal sensitivity at
the cut end of a peripheral axon (Michaelis, M. et al.,
1999). More recently, it have been demonstrated that
compression of a nerve is sufficient to alter the distribu-
4.4.3 Changes in Nociceptor Function
tion of tranducers in the membrane (Ma, C. et al., 2006).
Following Traumatic Nerve Injury
Importantly, aberrant expression patterns are not
Neuropathic Pain
restricted to the point of compression or injury as a
Pain and hyperalgesia arising from direct damage to cut axon will result in increased mechanical sensitivity
sensory neurons come under the heading of neuro- of the sensory neuron soma (Howe, J. F. et al., 1977).
pathic pain. One of the most dramatic and common Two consequences of these changes are the emergence
features of neuropathic pain is the presence of of mechanical sensitivity at sites that are normally
ongoing pain (Backonja, M. M. and Stacey, B., mechanically insensitive and mechanical allodynia, a
2004). This feature is distinct from inflammatory situation in which a mechanical stimulus, such as that
pain, where it is often possible to relieve pain if it is associated with movement, that is normally not painful,
possible to eliminate stimuli that impact the inflamed becomes a source of pain. Worse still, mechanical
tissue. There is compelling evidence to suggest that stimuli associated with physiological functions, such as
ongoing pain associated with peripheral nerve injury movement of tissue associated with blood flow, may also
is the result of aberrant and/or ongoing activity in become a source of stimuli for these transducers. A
 Molecular Biology of the Nociceptor/Transduction 63

second form of aberrant expression is the emergence of The emergence of inappropriate sources of stimu-
transducers in the wrong neurons. For example, as lation may involve sources that are both expected
indicated above, TRPV1 is primarily expressed in noci- and those that are much less so. Following nerve
ceptive afferents. However, following nerve injury, this injury, there are a number of changes that occur in
transducer is expressed in what were previously low- both the injured axon and the surrounding tissue that
threshold non-nociceptive afferents (Rashid, M. H. et al., facilitate restoration of the normal innervation pat-
2003). A third form of aberrant expression is a change in tern (see Murinson, B. B. et al., 2005). Injured nerves
the levels of expression of transducers normally present extend processes, which if scar tissue forms at the
in a neuron. There is evidence for both decreases in same time, may not be able to access former nerve
inhibitory transducers and increases in excitatory trans- tracks. The result may be the formation of a neuroma.
ducers. For example, nerve injury is associated with a Thus, an expected source of stimulation would be
decrease in the expression of opioid receptors (Kohno, mechanical stimulation of the neuroma associated
T. et al., 2005), which are the transducers for the inhibi- with traction of the surrounding scar tissue.
tory signal of both endogenous and exogenous opioids. Interestingly, nerve injury may also be associated
The result is a decrease in the efficacy of mechanisms with the sprouting of sympathic postganglionic term-
normally involved in attenuating pronociceptive pro- inals into the sensory ganglia (McLachlan, E. M. et al.,
cesses. An example of an increase in an excitatory 1993). This striking observation provided anatomical
transducer is an increase in the expression of the ATP evidence of sympathic/afferent coupling within the
receptor, P2X3 (Fukuoka, T. et al., 2002). The combina- sensory ganglia. While many details about the
tion of these changes in expression levels is a net mechanisms underlying this sprouting have yet to
be determined, access to NGF appears to be a reg-
increase in afferent excitability.
ulating factor (Ramer, M. S. and Bisby, M. A., 1999).
Evidence in support of aberrant coupling between
transducers and signaling pathways following nerve
4.4.3.2 Changes in membrane stability
injury is less compelling but should be mentioned.
As emphasized above, the distribution, expression
The pathway that has received the greatest attention
levels and biophysical properties of ion channels in
is the adrenergic system, largely as a result of the
a neural membrane are critical to the maintenance of
observation that pain associated with nerve injury
a given level of membrane excitability. Change in
may be sympathetically dependent. What most pain
any of these parameters may have profound effects.
scientists agree on, is that in normal tissue, activation
Not surprisingly, nerve injury is associated with
of the sympathetic nervous system is not painful. In
changes in all three, with clear examples of changes
fact, primary afferent neurons express adrenergic
in distribution (Gold, M. S. et al., 2003), expression
receptors that are critical for the antinociceptive levels (Waxman, S. G. et al., 1994; Dib-Hajj, S. et al.,
actions of spinally administered adrenergic agonists. 1996; Decosterd, I. et al., 2002), and biophysical prop-
However, following nerve injury, adrenergic agonists erties (Cummins, T. R. and Waxman, S. G., 1997) of
may become excitatory (Sato, J. and Perl, E. R., 1991; ion channels in both injured neurons and their unin-
Devor, M. et al., 1994; Chen, Y. et al., 1996; Liu, X. et al., jured neighbors. One of the more striking
1999; Moon, D. E. et al., 1999). Mechanisms of this ramifications of changes in all three of these proper-
excitation are still debated, but there is evidence to ties is the emergence and/or amplification of
support at least two mechanisms. One is that there is a membrane potential oscillations which, it has been
change in adrenergic receptor expression and/or cou- argued, are sufficient to drive ongoing activity fol-
pling such that activation of adrenergic receptors on lowing nerve injury (Amir, R. et al., 1999). Two
primary afferent becomes excitatory (Davis, K. D. et al., critical players have been identified. One is a vol-
1991; Birder, L. A. and Perl, E. R., 1999). A second is tage-gated Na channel with appropriate biophysical
that there is a change in the signaling pathway under- properties to sustain the upstroke of the oscillation.
lying the actions of adrenergic agonists at The second is a K leak current that drives the
autoreceptors on the sympathetic postganglionic neu- downstroke of the oscillation (Amir, R. et al., 1999).
ron terminals: in the presence of elevated levels of While the K channels have yet to be identified,
intracellular Ca2, norepinephrine becomes capable several different Na channels have been implicated.
of driving the release of arachidonic acid metabolites One possibility is a TTX-resistant channel such as
such as prostaglandin E2 (Gonzales, R. et al., 1989; NaV1.8 or NaV1.9. This would be consistent with the
1991), which then act directly on the primary afferent. observation that the upstroke of the oscillation in
64 Molecular Biology of the Nociceptor/Transduction

trigeminal neurons is TTX-resistant (Puil, E. et al., peripheral nerve (Coward, K. et al., 2001). Second,
1989) as well as the biophysical properties of the there appears to be no deficit in the nerve injury-
current implicated in studies of isolated DRG neu- induced hypersensitivity in an NaV1.7 null mutant
rons (Liu, C. N. et al., 2002). However, spontaneous mouse (Nassar, M. A. et al., 2005). Thus, while the
activity in injured afferents is most robust in sensory specific channels underlying nerve injury-induced
neurons with a large cell body diameter giving rise to increases in afferent excitability have yet to be con-
myelinated axons (Liu, C. N. et al., 2000); neither clusively identified, it is clear that ion channels alone
TTX-resistant channel is present in this subpopula- may become a generator of ongoing activity in injured
tion of sensory neurons and both are dramatically neurons.
downregulated in injured neurons (Dib-Hajj, S.
et al., 1996; Decosterd, I. et al., 2002). Another channel 4.4.3.3 Mechanisms underlying nerve
that has received a lot of attention is NaV1.3. This injury-induced changes in transduction
channel is upregulated in the presence of injury While many of the mechanistic details underlying
(Waxman, S. G. et al., 1994; Black, J. A. et al., 1999) nerve injury-induced changes in transducer and/or
and appears to have biophysical properties consistent ion channels have yet to be identified, it is clear that
with ongoing high-frequency activity (Cummins, T. changes in access to trophic factors plays a major role
R. and Waxman, S. G., 1997; Cummins, T. R. et al., in the process (Waxman, S. G. et al., 1999; Sah, D. W.
2001). There is evidence for the accumulation of et al., 2003). Following nerve injury, different groups of
TTX-sensitive Na channels at sites of nerve injury afferents may either be deprived of access to neuro-
(Devor, M. et al., 1993). Furthermore, there is evi- trophic factors or bathed in excess levels with those
dence that at least some of the oscillatory activity in that are injured being deprived and those that remain
sensory neurons is sensitive to TTX (Amir, R. et al., being bathed in excess. The suggestion that access to
1999). However, antisense knockdown of this channel neurotrophic factors plays a critical role in driving
has no impact on nociceptive behavior associated nerve injury-induced changes is supported by the
with nerve injury (Lindia, J. A. et al., 2005). observation that exogenous application of factors
Recent evidence from a cohort of individuals who such as NGF or GDNF to injured neurons restores
suffer from a paroxysmal pain disorder raise the pos- ion channel expression levels and may eliminate
sibility that NaV1.7 may contribute to nerve injury- ongoing activity. Exogenous NGF restores expression
induced increases in afferent excitability. Sufferers of levels of NaV1.8 and NaV1.9 and reduces expression
this disorder appear to share gain of function muta- levels of NaV1.3 (Black, J. A. et al., 1997; Fjell, J. et al.,
tions in NaV1.7, decreasing the rate and/or extent of 1999b). GDNF has also been shown to differentially
channel inactivation (Fertleman, C. R. et al., 2006). regulate expression levels of NaV1.3 (Boucher, T. J.
While the link between neuropathic pain and parox- et al., 2000; Leffler, A. et al., 2002). Similarly, the per-
ysmal pain disorder is tenuous, manifestations of this ipheral administration of NGF results in the
disorder share similarities with signs and symptoms sensitization of nociceptive afferents and increases in
neuropathic pain, most notably a sensitivity to the the expression of many proteins that are increased in
membrane stabilizer carbamazepine. A different set uninjured afferents following nerve injury (e.g.,
of NaV1.7 mutations appears to underlie primary TRPV1) (Ji, R. R. et al., 2002; Rashid, M. H. et al.,
erythermalgia (Yang, Y. et al., 2004), a rare pain dis- 2003). Attenuation of neurotrophin signaling through
order associated with pain and redness in the feet and antibodies or antagonist has also been shown to reverse
hands. Mutations in NaV1.7 associated with eryther- inflammation-induced changes in afferent excitability
malgia contribute to an increase in the excitability of (McMahon, S. B., 1996; Koltzenburg, M. et al., 1999).
primary afferent neurons (Rush, A. M. et al., 2006), but
they also appear to result in a decrease in the excit-
ability of sympathetic postganglionic neurons, and it is 4.5 Conclusion
likely the combination of these changes in excitability
that appear to mediate the unique symptoms of the Adequate treatment of chronic pain in the absence of
disorder. At least two lines of evidence argue against a serious side effects remains an elusive goal for patients,
role for NaV1.7 in pain associated with traumatic clinicians, and basic researchers. What should be clear
nerve injury. First, in patients with peripheral nerve from the preceding discussion is that the identification
injury NaV1.7 protein appears to decrease in the DRG of effective therapeutic interventions has been ham-
neurons and remains undetectable in injured pered by the realization that the system is far more
 Molecular Biology of the Nociceptor/Transduction 65

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 5 Zoster-Associated Pain and Nociceptors
H Maija, Helsinki University Hospital, Helsinki, Finland
2009 Elsevier Inc. All rights reserved.

5.1 Epidemiology and Course of Herpes Zoster and Postherpetic Neuralgia 75

5.2 Neuropathology of Herpes Zoster and Postherpetic Neuralgia 77
5.3 Mechanisms of Acute Zoster Pain 77
5.4 Pathophysiology of Postherpetic Neuralgia 78
References 79

Glossary
allodynia Painful response to normally pain-free postherpetic neuralgia (PHN) Prolonged pain
stimulus on the skin. after herpes zoster.
herpes zoster (HZ) Usually painful reactivation of varicella-zoster virus (VZV) The causative agent
varicella-zoster virus. of herpes zoster.

5.1 Epidemiology and Course of paresthesias and itching. Patients with HZ affecting
Herpes Zoster and Postherpetic the extremities can have complex regional painlike
Neuralgia symptoms (color changes, skin temperature changes,
edema, weakness, and extradermatomal involve-
Postherpetic neuralgia (PHN) is a painful aftermath ment), and the risk of PHN is high in these cases
of herpes zoster (HZ), an acute inflammation of (Berry, J. D. et al., 2004). Most patients experience
spinal nerves due to reactivation of varicella-zoster pain during HZ, with only about 10% remaining
virus (VZV) residing dormant in sensory ganglia. HZ pain-free. Pain precedes rash in 75% of cases
is the most common neurological disorder with a (Haanpaa, M. et al., 1999a; Chidiac, C. et al., 2001),
cumulative lifetime incidence of 1020% in general lasting usually for some days, but in some cases weeks
population, and as high as 50% of a cohort surviving to and even months before the eruption of rash (Gilden,
85 years of age (Dworkin, R. H. and Schmader, K. E., D. H. et al., 1991). Zoster pain tends to resolve spon-
2001). Surveys from two United Kingdom general- taneously with time, but a minority of patients
practice populations reported annual incidence rates continue to have pain after the healing of rash.
of 3.44.8 per 1000 persons (McGregor, R. M., 1957; Retrospective epidemiological studies based on
Hope-Simpson, R. E., 1965). Two population-based help-seeking behavior reported that about 10% of
studies of epidemiology of HZ have been published the patients have pain after the healing of rash, 5%
from Rochester, Minnesota (Ragozzino, M. W. et al., of the patients have pain at 3 months, and 2% have
1982; Donahue, J. G. et al., 1995) and reported the pain at 1 year (Burgoon, C. F. et al., 1957; Hope-
annual age-adjusted incidences of 1.3 and 2.1 per Simpson, R. E., 1965; Ragozzino, M. W. et al., 1982).
1000. About 50% of the HZ cases are thoracic, A prospective long-term follow-up study from
whereas trigeminal, cervical, and lumbar locations Iceland reported that 19% of the patients have pain
represent each 1015% of the cases and sacral derma- at 1 month, 7% at 3 months, and 3% at 1 year
tomes about 5% of the cases. The distribution of HZ (Helgason, S. et al., 2000). A prospective observational
reflects the initial virus burden to sensory ganglia community study reported pain in 30% of the
(Hope-Simpson, R. E., 1965). patients at 6 weeks, 27% at 12 weeks, 16% at 6
In HZ, viral replication first causes ganglionitis months, and 9% at 1 year (Scott, F. T. et al., 2003).
followed by infection of the corresponding nerve and The most important risk factors for PHN are old age
skin dermatome. Pain and unilateral dermatomal rash and severe acute pain. Other predictors of PHN are
are the typical symptoms, often accompanied by severe rash, presence of prodromal pain, viremia at

75
76 Zoster-Associated Pain and Nociceptors

presentation (Scott, F. T. et al., 2003), and more most distressing feature of the pain. Itching and dys-
pronounced immune response (Sato-Takeda, M. esthetic sensations may also be present, being in some
et al., 2004), which reflect more severe acute disease. patients even more annoying than the pain itself
Mechanical allodynia, that is, painful response to (Oaklander, A. L. et al., 2003). Normal sensory func-
normally pain-free mechanical stimulus on the skin, tion is often altered in PHN. Mechanical allodynia is
and pinprick hypoesthesia in acute HZ are associated reported in 6587% of patients with PHN (Watson,
with PHN at 3 months, but due to their insufficient C. P. N. et al., 1988; Nurmikko, T. and Bowsher, D.,
sensitivity and specificity, they cannot be used as 1990; Pappagallo, M. et al., 2000), and some patients
predictors of PHN for an individual patient have cold allodynia (Bowsher, D., 1996). The patients
(Haanpaa, M. et al., 2000). Abnormal findings in quan- may experience paradoxical sensations in the affected
titative sensory testing (QST) are common in HZ: skin area: hot may be felt cold or vice versa, and
warm and cold thresholds are elevated in 20%, heat although light touch may be unpleasant or painful,
pain thresholds are lowered in 10%, and tactile firm pressure may attenuate pain (Nurmikko, T.,
thresholds are elevated in 25% of the patients with 1994). Noordenbos W. (1959) investigated the skin of
HZ. All somatosensory abnormalities tend to normal- the patients with PHN and noted that there were areas
ize with time. The abnormal QST findings are of major sensory loss as well as minimal sensory loss in
associated neither with the severity of acute pain the affected region. Others have suggested that
nor with the development of PHN (Haanpaa, M. extreme allodynia to light touch is often restricted to
et al., 1999b). areas surrounding scarred skin or lying at the border of
HZ has been regarded as a simple model of affected and unaffected dermatomes. Allodynic areas
peripheral unilateral neuritis caused by VZV, but may be often warm termographically, while scarred
evidence from neuropathological reports suggest areas with maximal sensory loss are cool (Rowbotham,
that viral invasion and inflammation may extend to M. C. and Fields, H. L., 1989). When tested with
the central nervous system (CNS) as well (Denny- conventional clinical methods, hypoesthesia to
Brown, D. et al., 1944; Watson, C. P. N. et al., 1991). In touch was present in 90% and hypoesthesia to pin-
a study with 56 immunocompetent HZ patients prick in 92% of the patients in a pain clinic series
without clinical symptoms of CNS infection, cere- (Watson, C. P. N. et al., 1988). In another case series,
brospinal fluid (CSF) samples were obtained from 46 94% of the patients had impaired temperature and/or
patients on days 118 from the eruption of rash, and pinprick sensation and/or allodynia, and about 33%
16 consecutive patients with cranial or cervical HZ had also tactile deficit (Bowsher, D., 1996).
underwent magnetic resonance imaging (MRI) 15 In QST, marked threshold elevations within the
weeks following rash (Haanpaa, M. et al., 1998). In affected dermatome compared with the mirror-
35% of the patients there was evidence of VZV in the image site have been found for touch, pinprick,
CSF either in the form of a positive polymerase chain vibration, warmth, and cold (Bjerring, P. et al., 1990;
reaction (PCR) or anti-VZV-immunoglobulin G Nurmikko, T. and Bowsher, D., 1990; Eide, P. K. et al.,
(IgG). Leucocytosis (range 51440 mL1) was found 1994; Rowbotham, M. C. and Fields, H. L., 1996;
in 46% of the patients. These changes were more Rowbotham, M. C. et al., 1996a; Choi, B. and
common in patients with complications (e.g., periph- Rowbotham, M. C., 1997; Attal N. et al., 1999;
eral motor paresis or ophthalmic complications) but Pappagallo, M. et al., 2000). This indicates damage
they did not predict PHN. Zoster-related MRI to unmyelinated and myelinated sensory fibers. Heat
changes were found in the brainstem in 56% of the pain hypoesthesia is common in patients with PHN,
patients and in the cervical cord in two. Three but a minority of patients has heat pain hyperalgesia
patients had enhancement of the trigeminal nerve (Nurmikko, T. and Bowsher, D., 1990; Rowbotham,
in addition to the brainstem lesions. The presence M. C. and Fields, H. L., 1996; Rowbotham, M. C. et al.,
of brainstem lesions in MRI was associated with the 1996a; Choi, B. and Rowbotham, M. C., 1997). In
development of PHN at 3 months but not with the some patients with PHN, no asymmetry was found
severity of acute pain. in warmth or heat pain thresholds, while the greatest
Those who continue to suffer from prolonged threshold difference observed between the affected
pain after HZ describe three types of pain: a constant and contralateral dermatomes was as high as 12.2  C
deep aching or burning pain, an intermittent pain for warmth and 9.6  C for heat pain (Choi, B. and
with a lancinating quality, and allodynia of the skin. Rowbotham, M. C., 1997). However, noticeable
In an individual patient, any component can be the asymmetry in thresholds between the affected site
 Zoster-Associated Pain and Nociceptors 77

and its mirror-image site was found in different mod- et al., 1991). After HZ, a preponderance of small-dia-
alities from 5% to 13% of HZ patients not meter fibers in the surviving fiber population in
developing PHN (Nurmikko, T. and Bowsher, D., peripheral nerves is suggested (Noordenbos, W.,
1990). Hence, it seems possible to have sensory 1959; Zacks, S. L. et al., 1964; Watson, C. P. N. et al.,
threshold abnormality without PHN, and PHN with- 1991), but the significance of this finding is ambiguous,
out sensory threshold abnormality in the affected as the fibers in question have not been fully identified.
dermatome after HZ. Loss of myelin combined with the preservation of
axons has also been described (Fabian, V. A. et al.,
1997). Loss of axons and myelin in the peripheral
5.2 Neuropathology of Herpes Zoster nerve is similar in cases with and without PHN
and Postherpetic Neuralgia (Zacks, S. L. et al., 1964).
Three studies using skin biopsies report that HZ
The classic report of Head H. and Campbell A. W. lowers the innervation density of skin (Rowbotham,
(1900) is the most comprehensive study of the neu- M. C. et al., 1996a; Oaklander, A. L. et al., 1998;
ropathology of HZ, but it does not report the possible Oaklander, A. L., 2001). These changes are more
presence of PHN. Despite it and later studies, our profound in patients with PHN, possibly because
understanding of the pathophysiology of PHN is far the absence of pain requires the preservation of a
from complete. In acute HZ the dorsal root ganglion minimum density of primary nociceptive neurons
shows hemorrhagic inflammation and subsequent (Oaklander, A. L., 2001). The severity of allodynia
scarring (Head, H. and Campbell, A. W., 1900; is nevertheless associated with well-preserved fiber
Denny-Brown, D. et al., 1944; Reske-Nielsen, E. density (Rowbotham, M. C. et al., 1996a). The results
et al., 1986; Schmidbauer, M. et al., 1992). Typically, of Oaklander A. L. et al. (1998) suggest that unilateral
only one ganglion is severely affected, while mild HZ can cause bilateral segmental damage to primary
patchy inflammatory changes can be found in the sensory neurons. It is of interest that changes in the
neighboring ganglia (Denny-Brown, D. et al., 1944; epidermal innervation in the contralateral unaffected
Schmidbauer, M. et al., 1992). Ganglion cell necrosis skin in patients with PHN correlate with the pre-
is followed by secondary degeneration and fibrosis in sence and severity of pain, but the pathophysiological
the sensory root and the peripheral nerve. Direct explanation for this is unknown.
inflammation can also damage the root and nerve
(Head, H. and Campbell, A. W., 1900; Denny-
Brown, D. et al., 1944; Zacks, S. L. et al., 1964; 5.3 Mechanisms of Acute Zoster
Watson, C. P. N. et al., 1991; Schmidbauer, M. et al., Pain
1992). VZV has been shown in ganglial and satellite
cells; in damaged nerve roots; in the vasa nervorum; Mechanisms of acute zoster pain are not fully estab-
in the distal peripheral nerve endoneurally, perineu- lished. It is obvious that acute zoster pain represents a
rally, and epineurally; and in Schwann cells combination of nociceptive and neuropathic pain.
(Schmidbauer, M. et al., 1992). Inflammation in per- The nociceptive component is caused by inflamma-
ipheral nerves may persist from weeks to months, tion of the skin, which leads to excitation and
which can eventually lead to noticeable fibrosis sensitization of nociceptors. Increased afferent activ-
(Head, H. and Campbell, A. W., 1900; Zacks, S. L. ity transmitted to the spinal cord creates a state of
et al., 1964; Watson, C. P. N. et al., 1991). central hyperexcitability in the dorsal horn neurons,
Only a few studies compare the neuropathological which resolves gradually in concert with the healing
findings of those who recover with the finding of of tissue damage in a majority of patients. The neu-
those who have PHN. In four patients who had HZ ropathic component is caused by direct viral damage
and went on to develop PHN, atrophy of the dorsal and inflammation in the dorsal ganglion, peripheral
horn was found at autopsy whereas those without nerve, and in some cases also in the dorsal root and
PHN did not show similar changes (Watson, C. P. N. spinal cord. According to the study of acute HZ,
et al., 1991). However, one patient without PHN preherpetic pain is not associated with the severity
showed subacute myelopathy. The investigators of rash or QST changes, which suggests that virus-
found no difference in substance P, calcitonin gene- induced irritation in the afferent pathways rather
related peptide, a marker for norepinephrine, or in than dermal inflammation or axonal damage is more
opioid receptor binding studies (Watson, C. P. N. salient in preherpetic pain (Haanpaa, M. et al., 1999a).
78 Zoster-Associated Pain and Nociceptors

Because the abnormal QST findings are not asso- the axon reflex reactions induced by histamine ion-
ciated with the severity of acute zoster pain, tophoresis, Baron R. and Saguer M. (1993) found that
dysfunction of the thin fibers per se seems not to be histamine responses were reduced or abolished
the dominating factor in acute zoster pain. The pre- within allodynic areas, indicating degeneration of
sence of severe acute pain in acute HZ is associated C-fibers. Patients, who had recovered from HZ with-
with the presence of mechanical allodynia but not out PHN, had bilaterally identical histamine
with severe rash, which implies a neuropathic responses, indicating complete recovery of C-fibers.
character of allodynia in acute HZ rather than Additionally, impairment of the C-fiber function cor-
hyperesthesia or hyperalgesia due to cutaneous related positively with the intensity of postherpetic
inflammation (Haanpaa, M. et al., 2000). It is not pain. In contrast, Rowbotham M. C. and Fields H. L.
known to what extent the mechanisms of allodynia (1996) found, using QST measurements, that in the
in acute HZ are the same as the mechanisms of allodynic area of PHN patients, pain severity corre-
allodynia in PHN. The extension of allodynia and lated with the preservation of thermal sensory
somatosensory changes outside the affected derma- function. The same conclusion was drawn from the
tome is not uncommon in acute HZ, which indicates study using skin punch biopsies; the loss of cutaneous
that the CNS is quite frequently involved even in innervation correlated inversely with allodynia, and
mild cases of HZ (Haanpaa, M. et al., 1999a; 1999b; pain intensity was strongly related to the severity of
2000). Although QST changes are common in HZ, allodynia (Rowbotham, M. C. et al., 1996a). In another
they are not associated with the development of study, significant correlation was found between the
PHN. Hence, the prolonged pain in HZ cannot be intensity of ongoing pain and mechanical allodynia in
explained by as a simple consequence of either axo- patients with short-lasting (<1 year) but not in
nal damage (represented as threshold elevation) or
patients with long-lasting (1 year) PHN
sensitization of the nociceptive system (represented
(Pappagallo, M. et al., 2000). According to these
as heat pain hyperalgesia). Transition from acute
results, the underlying pathophysiology may vary
zoster-associated pain to PHN is likely to be a multi-
from patient to patient and with the duration of
factorial process, perhaps involving both peripheral
PHN.
and central changes. The genetic factors regulating
In PHN, three subtypes have been suggested
the inflammatory responses may play an important
(Table 1): the irritable nociceptor subtype, the deaf-
role in this process (Sato-Takeda, M. et al., 2004).
ferented allodynic subtype, and the deafferented
nonallodynic subtype (Fields, H. L. et al., 1998;
5.4 Pathophysiology of Postherpetic Petersen, K. et al., 2000; Rowbotham, M. and
Neuralgia Petersen, K., 2001). In patients with anatomically
intact primary afferent nociceptors, pain and allodynia
Patients with PHN have been studied using quanti- may be due to abnormal hyperactivity of the nocicep-
tative testing of the primary afferent function, skin tors and subsequent central sensitization, respectively
biopsies, and controlled treatment trials. In studies of (irritable nociceptor type). The topical capsaicin
patients with allodynic PHN, the results concerning application test can be used to identify PHN patients
the role of C-nociceptors in PHN are conflicting. in whom the mechanism of pain involves sensitization
Using indirect assessment of C-fibers by quantifying of peripheral afferents (Petersen, K. et al., 2000;

Table 1 Proposed subtypes of PHN

Capsaicin
Subtype Thermal sensory deficit Allodynia Anesthetic infiltration sensitivity

Irritable nociceptors Minimal deficit or hyperalgesic Marked Prolonged relief Severe burning
to heat pain pain
Deafferented nonallodynic Marked to all modalities None No change No sensation
subtype
Deafferented allodynic Marked to all modalities Variable Relief of allodynia short Variably
subtype duration reduced

Fields, H. L., Rowbotham, M., and Baron, R. 1998. Postherpetic neuralgia: irritable nociceptors and deafferentation. Neurobiol. Dis. 5, 209227
and Rowbotham, M. C., Petersen, K. L., and Feilds, H. L. Is postherpetic neuralgia more than one disorder. 1999. IASP New. Let. 37.
 Zoster-Associated Pain and Nociceptors 79

Petersen, K. et al., 2002). In patients with loss of cuta- direct evidence of a major sympathetic contribution
neous C-nociceptor function, evaluated either by to PHN is nonexistent (Wu, C. L. et al., 2000).
using QST of thermal thresholds (Nurmikko, T. and The treatment of HZ aims at minimizing the
Bowsher, D., 1990; Nurmikko, T., 1994; Choi, B. and neural damage with antiviral drugs, maximal pain
Rowbotham, M. C., 1997), C-fiber axon reflex relief with analgesic drugs, and prevention of
(Baron, R. and Saquer, M., 1993), or capsaicin-induced PHN. For those who continue to have prolonged
flare (Morris, G. C. et al., 1995), deafferentation and pain, the current armamentarium of medical
central reorganization may explain the PHN pain treatments, that is, tricyclic antidepressants, gabapen-
(Fields, H. L. et al., 1998). If tactile (A) fibers are tinoids, opioids, and topical lidocaine should be
preserved, allodynia may result from sprouting of the tested (Dubinsky, R. M. et al., 2004). The possibility
spinal terminals of A mechanoreceptors into contact of multiple mechanisms in the same patient may
with the pain transmission neurons. These cases repre- explain why response to any single treatment is so
sent the deafferented allodynic subtype. In patients of often partial and forms a logic base for polytherapy.
deafferented nonallodynic subtype, pain may result The detailed information of the efficacy of the
from the activation of the CNS pain transmission treatments on the various components of PHN is
neurons (Nurmikko, T. and Bowsher, D., 1990; lacking.
Fields, H. L. et al., 1998). In a study of 63 PHN patients,
25% of the cases were of irritable nociceptor type,
50% were of deafferentation with allodynia type, and
References
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(Pappagallo, M. et al., 2000). Attal, N., Brasseur, L., Chauvin, M., and Bouhassira, D. 1999.
Pain relief of peripherally acting treatments, that Effects of a single and repeated applications of a eutectic
is, topical agents, supports the role of peripheral mixture of local anaesthetics (EMLAR) cream on
spontaneous and evoked pain in post-herpetic neuralgia.
nociceptors in PHN. Topical lidocaine, when Pain 81, 203209.
used as a 5% gel under an occlusive dressing Baron, R. and Saguer, M. 1993. Postherpetic neuralgia: are
(Rowbotham, M. C. et al., 1995) or as 5% patch C-nociceptors involved in signalling and maintenance of
tactile allodynia? Brain 116, 14771496.
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Nurmikko, T. 1994. Sensory Dysfunction in Postherpetic Watson, C. P. N., Evans, R., Watt, V. R., and Birkett, N. 1988.
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Disease, Progress in Pain Research and Management, Vol. 3 Watson, C. P. N., Deck, H. J., Morshead, C., Van der Kooy, D., and
(eds. J. Boivie, P. Hansson, and U. Lindblom), pp. 133141. Evans, R. J. 1991. Post-herpetic neuralgia: further post-mortem
IASP Press. studies of cases with and without pain. Pain 44, 105117.
Nurmikko, T. and Bowsher, D. 1990. Somatosensory findings in Watson, C. P., Tyler, K. L., Bickers, D. R., Millikan, L. E.,
postherpetic neuralgia. J. Neurol. Neurosurg. Psychiatry Smith, S., and Coleman, E. 1993. A randomized vehicle-
53, 135141. controlled trial of topical capsaicin in the treatment of
Oaklander, A. L., Romans, K., Horasek, S., Stocks, A., postherpetic neuralgia. Clin. Ther. 15, 510526.
Hauer, P., and Meyer, R. A. 1998. Unilateral postherpetic Wu, C. L., Marsh, A., and Dworkin, R. H. 2000. The role of
neuralgia is associated with bilateral sensory neuron sympathetic blocks in herpes zoster and postherpetic
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 Zoster-Associated Pain and Nociceptors 81

Further Reading Relevant Websites

Helgason, S., Petursson, G., Gudmundsson, S., and http://www.iasp-pain.org International Association for
Sigurdsson, J. A. 2000. Prevalence of postherpetic neuralgia the Study of Pain.
after a first episode of herpes zoster: prospective study with
long term follow up. BMJ 321, 794796. http://www.vzvfoundation.org VZN Research
Foundation.
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 6 Ectopic Generators
M Devor, Hebrew University of Jerusalem, Jerusalem, Israel
2009 Elsevier Inc. All rights reserved.

6.1 Introduction 83
6.2 Nerve Injury and Disease Trigger Ectopia 83
6.2.1 Precipitating Factors 83
6.2.2 Spontaneous Ectopic Discharge 84
6.2.3 Ectopic Mechanosensitivity 84
6.2.4 Abnormal Response to Other Stimuli 85
6.3 Ectopic Firing Contributes to Pain Sensation in Several Ways 85
6.3.1 Ongoing Pain, Tender-Points, and Other Sensitivities 85
6.3.2 Allodynia Pain Evoked by Stimulation of Residual Uninjured Afferents 86
6.4 Cellular Mechanisms of Ectopic Impulse Generation 86
6.4.1 Resonance 86
6.4.2 The Development of Ectopic Pacemaker Capability 87
References 87

Glossary
ectopic generator Pathophysiological site of transduction in the case of afferent endings) and
electrical impulse generation in the peripheral ner- encoding of the generator depolarization into an
vous system (PNS) or central nervous system impulse train.
(CNS). Ectopic excludes locations of impulse gen- microneurographic recording An electrophysio-
eration in normal, healthy tissue. Examples are logical technique in which a fine wire
nerve-end neuromas and dorsal root ganglia microelectrode is inserted transcutaneously into
(DRGs) in the event of nerve injury. the peripheral nerve, permitting recording of
electrogenesis The process of electrical impulse impulse activity in individual nerve fibers. This
generation in nervous system tissue. minimally invasive procedure can be applied to
Electrogenesis usually includes two phases: for- awake humans.
mation of a generator depolarization (sensory

6.1 Introduction 6.2 Nerve Injury and Disease Trigger

Ectopia
Normally, sensation originates in impulse discharge
6.2.1 Precipitating Factors
generated at sensory endings in skin, viscera, and
other peripheral tissues. When there has been injury Ectopic impulse generation develops secondary to
to peripheral nerves, neuropathic pain may arise as a frank injury of the axon or soma of sensory neurons,
result of abnormal impulse discharge generated at or disruption of the myelin sheath that surrounds
ectopic sites. The most common locations of ectopic many axons (dysmyelination and demyelination).
impulse generation are the site of injury and asso- Disruption of the Schwann cell sheath that embraces
ciated dorsal root ganglia (DRGs). This chapter will bundles of nonmyelinated axons (Remak bundles)
touch on the characteristics of ectopic impulse gen- may also be a cause. It is fairly obvious why damage
eration (ectopia), the ways in which ectopia or disease that disrupts the ability of axons to conduct
contributes to neuropathic pain sensation, and the nerve impulses should cause negative sensory
cellular mechanisms that underlie it. abnormalities such as hypoesthesia and numbness. It

83
84 Ectopic Generators

is much less obvious why neural pathology causes vary with the type of injury, source (axon or soma),
positive sensory symptoms and signs such as ongoing and strain and species of animal.
and evoked paresthesias, dysesthesias, and pain. The Firing pattern is usually tonic-rhythmic (i.e., con-
link is the emergence of ectopia. If the axotomy tinuous), bursty, or irregular, with the occasional
happens suddenly, there may be a brief period of neuron showing complex, often cyclic variations in
injury discharge. In general, however, ectopia results firing pattern. The instantaneous firing rate of tonic-
from secondary changes that develop over time rhythmic and bursty activity is typically 1540
(Devor, M., 2006a). The pathophysiological changes impulses per second (ips), with bursts typically occur-
that induce ectopia can be precipitated by a wide ring at intervals of between a few per second and one
variety of events including trauma (frequently iatro- every few seconds (Figure 1). Irregular firing of single
genic), nerve entrapment, infection (bacterial or spikes occurs at the same relatively slow rate as burst-
viral), sterile inflammation, metabolic abnormalities, ing. The reason for these patterns is now known
malnutrition, vascular abnormalities, neurotoxins (Amir, R. et al., 2002b). The basic clock driving ectopia
(including many chemotherapeutic agents), radia- triggers activity at the slow rate. Some cells render
tion, inherited mutations, and autoimmune attack. only singlet spikes. In others, an iterative process based
on postspike depolarizing afterpotentials (DAPs)
yields a short rhythmic burst upon each trigger
6.2.2 Spontaneous Ectopic Discharge event, resulting in bursty discharge. Tonic autorhyth-
A considerable fraction of injured sensory neurons, as micity reflects an unending burst, which can occur in
many as one-third in some experimental preparations, cells that generate DAPs but in which the mechanism
begin to fire spontaneously in the hours and days for ending bursts is relatively ineffective. Interestingly,
following injury. Microneurographic recordings in observations in nerves in which only a part of the
patients with nerve injury have revealed similar results axons have been severed show that spontaneous ecto-
(Nordin, M. et al., 1984; Devor, M., 2006a). The origin pic activity may also develop in neighboring uninjured
of the activity has been identified as the swollen end- axons, particularly in C-fibers (Wu, G. et al., 2001;
bulbs that form in the neuroma just proximal to sites of Shim, B. et al., 2005).
axonal transection (Fried, K. et al., 1991), outgrowing
sprouts, plaques of demyelination, and the cell soma in
6.2.3 Ectopic Mechanosensitivity
the DRG. In the event of sudden traumatic neuropa-
thy activity usually begins earliest in myelinated axons Gentle percussion over sites of nerve injury, areas of
(A-fibers), sometimes as soon as 16 h after axotomy, entrapment or neuromas for example, typically evokes
with a preference for muscle over cutaneous afferents. an intense stabbing or electric shocklike sensation. This
Activity in unmyelinated (C-) fibers tends to appear in is the Tinel sign. Similar sensations can be evoked by
earnest later, after a few weeks. Specific parameters other maneuvers that apply mechanical force to injured

(a)

49 mV

(b)

53 mV

Figure 1 Typical patterns of spike activity generated at ectopic pacemaker sites. (a) Bursty discharge. (b) Irregular singlet
firing. Subthreshold oscillations are visible in the baseline traces. Calibrations: 5 mV/100 ms.
 Ectopic Generators 85

these are response to circulating catecholamines and

R noradrenalin released from nearby (injured) postgan-
glionic sympathetic axons (Devor, M. and Janig, W.,
1981). Ectopic adrenosensitivity of sensory neurons
yields sympatheticsensory coupling, a likely sub-
30
strate of sympathetically maintained chronic pain
Impulses per second

states. Afferent response to local and circulating

20 inflammatory mediators is a second example of ecto-
pic chemosensitivity. Abnormal discharge may also
10 arise from temperature changes, ischemia, hypoxia,
hypoglycemia, and other conditions capable of
0 locally depolarizing afferent neurons at the sites at
10 s which they have developed local ectopic pacemaker
Figure 2 Mechanical afterdischarge. A weak, momentary capability (Devor, M., 2006a). Whereas intact sensory
stimulus to the surface of the DRG triggered a prolonged, endings may be sensitive to these chemical and phy-
self-sustained ectopic discharge burst. sical stimuli, axons in healthy nerve trunks and cell
bodies in sensory ganglia are not. The key change in
nerves or spinal roots such as straight leg lifting in neuropathy is not the presence of excitatory stimuli
sciatica (Lasegues sign), and the signs of Spurling and but rather the ectopic emergence of sensitivity to
Lhermitte. Momentary mechanical stimulation of this them.
sort frequently evokes ectopic discharge that outlasts
the stimulus itself (afterdischarge), with corresponding
aftersensation (Figure 2). This indicates that the under- 6.3 Ectopic Firing Contributes to Pain
lying process is not just activation of sensitized Sensation in Several Ways
nociceptors, or signal amplification, but the triggering 6.3.1 Ongoing Pain, Tender-Points,
of autonomous repetitive firing at ectopic pacemaker and Other Sensitivities
sites. The nature of this local electrical hyperexcitabil-
ity is discussed below. The presence of ongoing ectopic discharge under cir-
In the event of injuries that leave the nerve in- cumstances in which ongoing dysesthesias and pain are
continuity, such as crush or freeze lesions, there is present, and their simultaneous exacerbation by the
usually a second Tinel sign that advances with time, same factors (e.g., mechanical stimulation, sympathetic
marking the farthest position reached by regenerat- efferent activity), constitutes a priori evidence that the
ing sprouts. In some conditions (e.g., diabetic ectopia is the cause of the pain. This conclusion is
neuropathy) paresthesias and pain may occur on further supported by the observation that (diagnostic)
tapping anywhere along the course of a nerve. This nerve blocks using local anesthetics reliably obtund
occurs when there are scattered outgrowing sprouts, pain sensation in cases where the block is placed central
demyelination plaques, or mechanosensitive end- to the site of nerve injury (Devor, M., 2006a). An
bulbs resulting from axonal dying back. Although interesting test case often raised as a counterexample
direct evidence has not yet been sought systemati- is phantom limb pain in amputees. Although this pain is
cally in humans, it follows logically from studies in frequently eliminated (transiently) by blocking stump
animals that pain evoked by deep palpation at tender neuromas, there are reports of phantom limb pain
spots, such as in fibromyalgia, might reflect ectopic persisting despite peripheral nerve block, and this
mechanosensitivity at locations where small nerve observation has formed the basis for the widely held
branches cross through fascial planes, under tendons, belief that phantom limb pain reflects abnormal neural
or are otherwise at risk of being locally pinched. activity originating in the cerebral cortex (Melzack, R.,
1989; Flor, H. et al., 1995). This conclusion is potentially
flawed, however, because it ignores ectopic electrical
impulse generation (electrogenesis) in sensory cell
6.2.4 Abnormal Response to Other
somata in the DRG. Amputation stump revision sur-
Stimuli
gery is a common procedure, and is usually done under
Ectopic pacemaker sites also develop abnormal sen- foramenal or spinal block. Although little published
sitivity to other depolarizing stimuli. Notable among documentation is available (Tessler, M. J. and
86 Ectopic Generators

Kleiman, S. J., 1994), experienced clinicians insist that deep somatic tissues such as muscles and joints.
these blocks reliably eliminate phantom limb pain for Reduced mechanical thresholds, including the
the duration of the pharmacological action of the local recruitment of previously silent C-nociceptors, ren-
anesthetic agent. This would not occur if the pain signal der the sensory endings of these afferents responsive
originated supraspinally. to the (substantial) forces present during weight bear-
ing, for example, or joint rotation (Schmidt, R., 1996).
There is also good evidence for a contribution of
6.3.2 Allodynia Pain Evoked by
central sensitization in deep tissue pain (Banic, B.
Stimulation of Residual Uninjured Afferents
et al., 2004). Relatively little is known about changes
Pain in response to light touch of the skin, tactile in the response of injured afferent neurons, or the
allodynia, is a common symptom in neuropathy. The sensory endings of residual noninjured afferents, in
simplest explanation is reduced response threshold in the presence of neuropathy affecting nerves that
nociceptive C-fiber afferents, the classic excitable supply deep tissues.
nociceptor hypothesis. This explanation is probably
valid for heat allodynia. However, while neuropathy
(and pronociceptive inflammatory mediators) can 6.4 Cellular Mechanisms of Ectopic
significantly reduce mechanical thresholds of sensory Impulse Generation
endings, few fibers that were originally nociceptors
6.4.1 Resonance
come to respond to the very weak tactile stimuli that
are typically sufficient to evoke allodynia (Banik, R. K. The emergence of ectopic hyperexcitability in neuro-
and Brennan, T. J., 2004; Schlegel, T. et al., 2004; pathy is not a simple matter of lowered spike threshold.
Shim, B. et al., 2005). Moreover, the response to Rather, it reflects a more fundamental, qualitative
painful touch in allodynic skin is rapid. There is no change in the injured neurons electrical characteristics.
indication of the long delay between stimulus and Most intact sensory neurons are incapable of generating
response that would be expected from conduction in sustained impulse discharge in midnerve or from
sensitized C-fibers, although sensitized A nocicep- within sensory ganglia, even in the presence of a strong
tors, if they exist, could contribute. sustained depolarizing stimulus. They are designed to
There is considerable evidence that cutaneous fire exclusively in response to stimuli applied to the
tactile allodynia in neuropathy (and inflammation) sensory ending in skin, muscle, etc., where the neuronal
is primarily a sensory response to impulse activity in membrane is specialized for electrogenesis. Alterations
low-threshold mechanosensitive A touch afferents in three cellular processes, gene expression, protein
abnormally amplified in the spinal cord (Campbell, J. trafficking, and channel kinetics (below), actually create
N. et al., 1988; Torebjork, H. et al., 1992). The altered repetitive firing capability de novo at ectopic locations.
signal processing responsible for this A pain is Ectopic repetitive firing capability (pacemaker
believed to result from one or a combination of capability) derives from electrical resonance that
numerous central nervous system (CNS) changes emerges in the cell membrane of the injured afferents,
collectively called central sensitization (Woolf, C. J., and resultant subthreshold oscillations in their nor-
1983; Devor, M., 2006b). Central sensitization, which mally stable resting potential (Amir, R. et al., 1999;
also amplifies ectopic afferent signals (spontaneous Wu, N. et al., 2001). Subthreshold oscillations drive
and evoked), is a dynamic process triggered and sustained ectopic discharge. This takes the form of
maintained by ongoing nociceptive input from irregular singlet spikes in some cells, and as repetitive
the periphery. When the maintaining peripheral bursting in others, depending on the ability of the cell
drive is eliminated, central sensitization fades, along to generate DAPs (Figure 1). Due to subthreshold
with tactile allodynia. In neuropathy, sustained ecto- oscillations, ectopic generator sites become responsive
pic discharge can maintain central sensitization to slow onset and tonic depolarizing stimuli, which
indefinitely, and with it A pain and tactile allodynia previously did not evoke sustained discharge. Some
(Gracely, R. et al., 1992; Koltzenburg, M. et al., 1994). begin to fire spontaneously at rest, in the absence of
This constitutes a second role of ectopia, in addition stimulation. The biophysical processes that lead to
to providing a primary pain signal. membrane resonance, repetitive firing capability,
In contrast to the skin, it is likely that peripherally ectopic discharge, and neuropathic pain are slowly
sensitized nociceptors play an important role in pain coming into focus. Briefly, resonance in neuropathy
evoked by mechanical stimuli in inflamed viscera and appears to reflect reciprocation between inward Na
 Ectopic Generators 87

current carried by fast-activating, inactivating, and

repriming tetrodotoxin (TTX)-sensitive Na chan-
nels, and K current passing through one or more
voltage-insensitive K leak channels, perhaps of the
KCNK K2p family (Abdulla, F. A. and Smith, P. A.,
2002; Amir, R. et al., 2002a; Waxman, S. G., 2002).
Voltage-sensitive K channels tend to hold resonance
in check, and their downregulation after axotomy is
probably an important adjunct to ectopia (Kocsis, J. D.
and Devor, M., 2000). Other channel types/conduc-
tances that might contribute to resonance characteri-
stics of primary sensory neurons include certain Ca2
channels, HCN (the Ih pacemaker channel), KCNQ-
type K channels, the 4 subunit-associated resurgent Figure 3 Immunolabeling shows the accumulation of Na
Na current, and a persistent Na conductance channels at the chronic cut end of hyperexcitable injured
(gNaP), but their role, at this stage, is speculative. axons in an experimental nerve-end neuroma (for details
Many of the first-line analgesic agents used in the see Devor, M. et al., 1989).
treatment of neuropathic pain (McQuay, M. and
Moore, A., 1998), including (certain) anticonvulsants,
2006a). Na channels, for example, accumulate in
tricyclic antidepressants, systemic local anesthetics,
neuroma endbulbs and sprouts, and in areas of
and (certain) antiarrhythmics are thought to act by
demyelination. A third process that contributes to
stabilizing membrane resonance (Catterall, W. A.,
ectopic electrogenesis is altered channel kinetics. It
1987; Deffois, A. et al., 1996).
is not ion channel proteins themselves that determine
cell excitability but the ionic currents they carry.
6.4.2 The Development of Ectopic Increasing mean channel open time, for example,
Pacemaker Capability can have much the same effect on excitability as
increasing the number of channels present.
The cascade of cellular events that lead to pacemaker
Proinflammatory cytokines and other mediators
capability and ectopic neuronal discharge begins
associated with neuropathy can affect channel
with the injured primary sensory neuron. Our cur-
kinetics in this way. For example, cAMP-dependent
rent understanding of the process is as follows.
phosphorylation of Na channel molecules reduces
Axonal transection blocks the normal retrograde
Na current, while dephosphorylation returns it to
flow of neurotrophic signaling molecules (glial
normal (e.g., Li, M. et al., 1992; Gold, M. S. et al., 1996).
cell-derived neurotrophic factor (GDNF), nerve
Because certain hormones, trophic factors, cytokines,
growth factor (NGF), and perhaps others) between
and other inflammatory mediators (notably prosta-
the periphery and the sensory cell body. This triggers
glandins) can activate protein kinases (PKA, PKC),
a change in the quantity of various proteins expressed
they are positioned to affect the basic excitability of
by the cell body and exported by anterograde axo-
afferents, and not just to depolarize and excite sen-
plasmic transport to both peripheral and central axon
sory endings.
endings; some are upregulated and others downregu-
lated (Boucher, T. J. and McMahon, S. B., 2001;
Costigan, M. et al., 2002; Xiao, H. S et al., 2002).
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These changes alter the phenotype of the injured
neuron, increasing its intrinsic resonance and sup-
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porting the emergence of ectopic electrogenesis. currents of rat dorsal root ganglion neurons following
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Amir, R., Liu, C. N., Kocsis, J. D., and Devor, M. 2002a.
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potential oscillations in dorsal root ganglion neurons: role in
erator sites (Figure 3), notably the accumulation of normal electrogenesis and in neuropathic pain. J. Neurosci.
Na channels (Devor, M., et al., 1989; Devor, M., 19, 85898596.
88 Ectopic Generators

Amir, R., Michaelis, M., and Devor, M. 2002b. Burst discharge Injury: Consequences For Chronic Pain. In: Proceedings
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Banic, B., Petersen-Felix, S., Andersen, O. K., Radanov, B. P., pp. 119135. IASP Press.
Villiger, P. M., Arendt-Nielsen, L., and Curatolo, M. 2004. Koltzenburg, M., Torebjork, H., and Wahren, L. 1994.
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Boucher, T. J. and McMahon, S. B. 2001. Neurotrophic factors by cAMP-dependent phosphorylation. Neuron
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Costigan, M., Befort, K., Karchewski, L., Griffin, R. S., with disorders of peripheral nerves, dorsal roots and dorsal
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 7 Sodium Channels
John N Wood, University College London, London, UK
2009 Elsevier Inc. All rights reserved.

7.1 How Do Sodium Channels Work? 89

7.2 Sodium Channels and Pain Pathways 92
7.3 Nav1.3 and Neuropathic Pain 92
7.4 Nav1.7 and Inflammation 93
7.5 Nav1.8 and Nociception 93
7.6 The Diversity of Sodium Channel Subtypes 94
7.6.1 Selective Sodium Channel Blockers as Drugs 95
References 95

7.1 How Do Sodium Channels are resistant to micromolar TTX and are defined as
Work? TTX-resistant (TTXr).
A short sequence found upstream of neuronal
All sodium channels are single proteins made up of sodium channel genes (as well as other neuronal
a-subunits that contain four homologous domains and genes) was identified and named neuron-restricted
form a voltage-gated sodium-selective aqueous pore silencing element (NRSE) or repressor element
(Figure 1). They comprise a family of nine structurally 1(RE-1). Transcription factors that bound to the
related a-subunits (Table 1). They are more than 75% motif were found to act as inhibitors of gene expres-
identical over the amino acid sequences comprising sion in nonneuronal cells. These proteins were
the transmembrane and extracellular domains (Yu, F. named RE-1 silencing transcription factor (REST)
H. and Catterall, W. A., 2003). The a-subunits show or neuron-restrictive silencer factor (NRSF) and
distinct patterns of expression, and are associated with mutational analysis identified a single zinc finger
accessory -subunits that modify channel properties motif in the carboxyl-terminal domain of the factor
and interact with cytoskeletal and extracellular matrix as essential for repressing sodium channel-derived
proteins. The mechanism of channel gating involves reporter genes (Kraner, S. D. et al., 1992). The inhi-
the movement of voltage sensors (shown with positive bitory activity of the complex can be further
charges in Figure 1) in response to changes in mem- modulated by double-stranded RNA molecules that
brane potential. Lysine or arginine residues are found have the same sequence as NRSE/RE-1, and are
at intervals of three amino acids to produce a linear found in developing neuronal precursors. These reg-
array of positive charges in the a-helical S4 transmem- ulatory RNA molecules are able to switch the
brane segments, and the movement of these charged repressor function of the complex to an ac activator
areas of the channel opens the sodium-selective pore, role (Kuwabara, T. et al., 2004). In this way, the
defined by residues within the channel atrium. An assumption of a neuronal phenotype seems to depend
intracellular loop lying between transmembrane in part upon regulatory RNAs driving gene expres-
domains 3 and 4 containing the tripeptide isoleucine sion downstream of NRSE/RE1 motifs. Sodium
phenylalanine methionine (IFM) is subsequently channels are known to be expressed at the very ear-
responsible for channel inactivation on a millisecond liest stages of the appearance of a neuronal
timescale. Sodium channel subtypes can be pharma- phenotype in the mouse. These studies highlight
cologically distinguished by their susceptibility to the significance of sodium channel expression in
block by tetrodotoxin (TTX), a toxin isolated from neuronal function throughout development.
puffer fish, which binds to the ion selectivity pore of The a-subunits of the voltage-gated sodium chan-
some channels (Hille, B., 2002). Most sodium channels nel that form the sodium pore are associated with
are blocked by nanomolar concentrations of TTX and accessory -subunits that modulate channel proper-
are defined as TTX-sensitive (TTXs) whilst others ties and interact with cytoskeletal and extracellular

89
90 Sodium Channels

1/ 3 2/4
-subunit

Figure 1 Structure of sodium channels. Voltage-gated sodium channels comprise a single functional a-subunit, which
is tethered by associated accessory subunits to subcellular locations determined by both extracellular and intracellular
proteins. Phosphorylation sites are shown on intracellular loop 2, and the serine residues in domain 1 that confers
tetrodotoxin sensitivity is highlighted.

Table 1 Names and properties of voltage-gated sodium channel a-subunits

Type Gene symbol Name Tissue TTX-sensitive

NaV1.1 SCN1A Type I CNS/PNS

NaV1.2 SCN2A Type II CNS
NaV1.3 SCN3A Type III CNS
NaV1.4 SCN4A SKM1 Skeletal muscle
NaV1.5 SCN5A SKMII Heart/CNS 
NaV1.6 SCN8A Nach6 CNS/glia
NaV1.7 SCN9A PN1 DRG
NaV1.8 SCN10 SNS DRG 
NaV1.9 SCN11 NAN DRG 

The nomenclature for sodium channel a-subunits, combined with their main site of expression (CNS central nervous system,
PNS peripheral nervous system) and their sensitivity to block by tetrodotoxin (TTX), is shown.

matrix proteins. -subunits are single transmembrane channel activation and inactivation. 3 most closely
domain proteins that contain an extracellular domain related to 1 is the product of a separate gene loca-
that is homologous to the V-set of the immunoglobu- lized to human chromosome 11q23.3. The expression
lin superfamily that includes cell adhesion molecules pattern of 3 differs significantly from 1.
(Isom, L. L., 2002). The biochemical purification of Coexpression of the -subunits with sodium
two proteins, 1 and 2, that associate with a-sub- channel a-subunits in Xenopus oocytes or mammalian
units allowed the properties of these accessory factors cell lines accelerates channel activation, and shifts the
to be investigated in heterologous expression systems. voltage dependence of inactivation to more negative
Molecular cloning has identified a 1-like member of potentials, indicating that -subunits are important
the family named 3 and 2-like subunit named 4, in the assembly, expression, and functional modula-
as well as a splice variant of 1, 1A. tion of the rat brain sodium channel heterotrimeric
1 and 1A are structurally homologous in the complex (Isom, L. L., 2002). However, -subunits do
extracellular immunoglobulin-like loop region. 1A not appear to modulate the biophysical properties of
mRNA is expressed during embryonic development, all sodium channel a-subunits. Nav1.8 a-subunits
and becomes undetectable after birth, concomitant show aberrant properties when expressed in hetero-
with the onset of 1 expression. Immunocyto- logous cells, but coexpression with 1, 1A, or 3
chemical analysis of 1A expression revealed selec- does not recapitulate normal channel behavior. The
tive expression in brain and spinal cord neurons, with existence of factors other than accessory -subunits
high expression in heart and all dorsal root ganglia that can alter inactivation kinetics has also been
neurons. Coexpression of Nav1.2A and 1A-subunits suggested by studies in which sodium channel
results in a 2.5-fold increase in sodium current density a-subunits are expressed in cells lacking endogenous
as a result of an increase in the level of functional sodium channel -subunits. Expression of rat cardiac
sodium channels in expressing cells. 1A-expressing Nav1.5 and rat brain Nav1.2 sodium channels in
cell lines also revealed subtle differences in sodium mammalian cell lines results in sodium channels
 Sodium Channels 91

with rapid activation and inactivation characteristic increase sodium currents by phosphorylation at con-
of native neuronal sodium channel. sensus sites in the cytoplasmic domain III linker.
An additional function of -subunits as cell adhe- Effects on membrane trafficking may also be mediated
sion molecules has been demonstrated by the ability through phosphorylation. In the case of the cardiac
of -subunits to mediate homophilic interactions via channel, activation of PKA causes a slow increase in
their extracellular immunoglobulin-like repeats sodium current in Xenopus oocytes. Chloroquine and
(Isom, L. L., 2002). Interacting -subunits also recruit monensin, both compounds that disrupt plasma mem-
the cytoskeletal protein ankyrin-G to the cell surface brane recycling, reduce sodium currents. Preincubation
via an interaction with their cytoplasmic domains, with these agents also prevented the PKA-mediated
suggesting that these subunits link sodium channel rise in sodium current, indicating that this effect likely
a-subunits indirectly both to the cytoskeleton and to resulted from an increased number of channels in the
the extracellular matrix proteins, such as tenascin-R. plasma membrane.
Tenascin-R is an extracellular matrix molecule that There is strong evidence for specialized roles of
is secreted by oligodendrocytes during myelination the various sodium channel isoforms based on human
and that binds F3-contactin. Transfected cells stably genetic studies as well as the production of knockout
expressing 1- or 2-subunits initially recognize and mice. A perinatal lethal phenotype occurs in the
then are repelled from substrates containing tenas- Nav1.2 sodium channel mouse knockout, which
cin-R. The cysteine-rich amino-terminal domain of shows severe hypoxia as a result of brain stem apop-
tenascin-R appears to be responsible for the repellent tosis. Nav1.8 channel knockouts have selective
effect on -subunit-expressing cells. Application of deficiencies in pain pathways (Akopian, A. N. et al.,
the cysteine-rich N-terminal domain of tenascin-R 1999). Nax, a non-voltage-gated sodium channel
to channels expressed in Xenopus oocytes potentiates homologue has been shown to play an important
expressed sodium currents. The binding of neuronal role in salt homeostasis, whilst loss of Nav1.6 in the
sodium channels to extracellular matrix molecules natural med mutant has been shown to lead to ataxia,
such as tenascin-R may thus play a role both in dystonia, and paralysis. The deletion of accessory
functional regulation and in localizing sodium chan- -subunits, already known to play an important
nels in high density at nodes of Ranvier. role in regulating functional channel expression and
-subunits also interact with the ankyrin-G mem- tethering a-subunits to specific subcellular locations,
brane skeletal protein that is associated with spectrin leads to a complex phenotype resulting in seizures
actin networks and also binds to integral membrane and epileptogenic activity.
proteins including the L1 CAM family of cell adhe- Naturally occurring mutations in both sodium
sion molecules. Ankyrin-G is highly concentrated channel a- and -subunits have been implicated in
along with the L1CAM family members neurofascin various inherited disorders, many of which have been
and NrCAM at nodes of Ranvier and axon initial characterized in muscle sodium channel isoforms.
segments. Voltage-gated sodium channels are thus Brugada syndrome, characterized by ventricular
likely to associate in a complex containing neurofas- fibrillation, heart failure, and sudden death, is asso-
cin/NrCAM and ankyrin-G in myelinated neurons. ciated with mutations in the cardiac sodium channel
As well as the accessory subunits that link sodium Nav1.5. Hyperkalemic periodic paralysis is associated
channels to cytoskeletal and extracellular elements, with mutations in the skeletal muscle channel Nav1.4.
other proteins are associated with sodium channels. However, there is also evidence of a role for neuronal
Voltage-gated sodium channels in brain neurons were sodium channels and -subunits in a variety of
found to associate with receptor protein tyrosine phos- pathologies, including epilepsy, autism, and pain per-
phatase- (RPTP-) (Catterall 200. Both the ception. Generalized epilepsy with febrile seizures
extracellular domain and the intracellular catalytic (GEFS) is associated with mutations in either the
domain of RPTP- interact with sodium channels. 1-subunit (type I) or the a-subunit Nav1.1 (type
Sodium channels were found to be tyrosine phos- II). Some forms of autism associate with mutations
phorylated and dephosphorylation slowed Nav1.1 in SCN1-3A sodium channel genes. The peripheral
inactivation, positively shifting voltage dependence neuron-associated sodium channel Nav1.7 encoded
and increasing whole-cell sodium current. by gene SCN9A has been shown to be the site of
Regulation of sodium channels by protein kinases mutations that lead to the chronic familial pain dis-
(PK) has already been demonstrated in a number of order primary erythermalgia (Yang, Y. et al., 2004).
systems. Protein kinase A (PKA) can decrease or This dominant condition is characterized by burning
92 Sodium Channels

in the extremities, pain evoked by standing and a DRG sensory neurons is shown in Figure 2. The
generalized inflammatory phenotype. Interestingly, persistent current activates at voltages close to the
systemic treatment with the sodium channel blockers resting potential and is likely to play a role in setting
lidocaine or mexiletine leads to dramatic improve- thresholds of activation.
ments in the symptoms associated with this disease. Evidence that altered sodium channel activity in
Neuronal excitability depends to a considerable peripheral neurons is associated with the develop-
extent on sodium channel trafficking, distribution ment of inflammatory and neuropathic pain is strong.
and density, as well as the intrinsic properties of the Altered patterns of sodium channel transcripts as well
channels themselves in terms of thresholds of activa- as changes in posttranslational modifications have
tion and repriming characteristics. The repertoire of been observed. Neuropathic pain that results from
sodium channels expressed by sensory neurons and direct damage to peripheral nerves is the most pro-
the alterations that occur in pain states are now well blematic condition in terms of analgesic therapy. The
documented (Waxman, et al., 2002, Wood, J. N. et al., pain evoked by these conditions seems to result initi-
2004). Unmyelinated C-fiber-associated dorsal root ally from enhanced neuronal excitability that can be
ganglion (DRG) neurons are usually nociceptive blocked by low-dose TTX.
whilst thinly myelinated A-fibers also mediate fast
pain, and a subset of myelinated A-fibers are also
nociceptive. TTXr sodium channels are present on
C-fibers and seem to play a significant role in noci- 7.3 Nav1.3 and Neuropathic Pain
ception (Akopian, A. N. et al., 1999).
Nav1.3 is widely expressed in the adult human cen-
tral nervous system but is normally present at low
7.2 Sodium Channels and Pain levels in adult peripheral nervous system. Axotomy
Pathways
or other forms of nerve damage lead to the reexpres-
sion of Nav1.3 and the associated 3-subunit in
Acute, inflammatory, and neuropathic pain can all be
sensory neurons, but not in primary motor neurons
attenuated or abolished by local treatment with
(Waxman, S. G. et al., 1994). This event can be
sodium channel blockers such as lidocaine. The peri-
reversed in vitro and in vivo by treatment with high
pheral input that drives pain perception thus depends
levels of exogenous glial cell-derived neurotrophic
on the presence of functional voltage-gated sodium
factor (GDNF). Nav1.3 is known to recover (reprime)
channels. Remarkably, two voltage-gated sodium
channel genes (Nav1.8 and Nav1.9) are expressed rapidly from inactivation (Cummins et al., 2001).
selectively in damage-sensing peripheral neurons, Axotomy has been shown to induce the expression of
whilst a third channel (Nav1.7) is found predomi- rapidly repriming TTXs sodium channels in damaged
nantly in sensory and sympathetic neurons. An neurons, and this event can also be reversed by the
embryonic channel (Nav1.3) is also upregulated in combined actions of GDNF and nerve growth factor
damaged peripheral nerves and associated with (NGF). Concomitant with the reversal of Nav1.3
increased electrical excitability in neuropathic pain expression by GDNF, ectopic action potential gen-
states. A combination of antisense and knockout stu- eration is diminished and thermal and mechanical
dies support a specialized role for these sodium pain-related behavior in a rat chronic constriction
channels in pain pathways. injury model is reversed. Moreover, Nav1.3 is upregu-
Small-diameter sensory neurons express a variety lated in dorsal horn neurons following experimental
of sodium channel transcripts as well as a repertoire spinal cord injury and this upregulation is associated
of electrophysiological and pharmacologically dis- with hyperexcitability of these nociceptive neurons
tinct type of sodium currents. Persistent sodium and pain; antisense knockdown of Nav1.3 (and prob-
channels that are resistant to TTX are probably ably other sodium channels) attenuates the dorsal horn
encoded for by Nav1.9 channels, whilst the major neuron hyperexcitability and the pain behaviors in
transient TTXr channel isotype present predomi- spinal cord-injured animals (Hains, B. C. et al., 2003).
nantly in nociceptors is Nav1.8. TTXs currents are It therefore seems likely that Nav1.3 reexpression may
encoded by a number of genes including Nav1.1, play a role in increasing neuronal excitability that
Nav1.7, and Nav1.6. The currentvoltage relationship contributes to neuropathic pain after nerve and spinal
of these different sorts of sodium channels found in cord injury.
 Sodium Channels 93

TTXs and TTXr Na+ currents in small-diameter DRG neurons

(a) (c)

1 nA

2 nA
2 ms

10 ms

(b) 3.5
Current
(nA)

Em (mV)

130 100
1 nA

5 ms

Figure 2 (a) TTX-sensitive (b) -resistant, and (c) persistent currents are shown. Note the different timescales for inactivation
of the currents. The voltage at which the different types of channels activate is shown in panel 4 (red channel (c), blue channel
(a), and black channel (b)). Although those channels that activate at the most negative potentials might seem to render other
channels redundant, the ability to recover from inactivation is also an important determinant of neuronal excitability.

7.4 Nav1.7 and Inflammation pain. Nav1.7 mRNA contributes to an increase in

TTXs sodium current in caraggeenan-inflamed rats,
Nav1.7 encodes a TTXs channel found predomi- further implying a role in altering inflammatory pain
nantly in peripheral sensory and sympathetic thresholds. Recently humans with a loss of functional
neurons. Interestingly, this channel is located at the Nav1.7 have been found to be completely pain free
terminal of sensory neurons, and is regulated in its (Cox, J. J. et al., 2006). Not surprisingly, this has
expression by inflammatory mediators such as NGF, stimulated major interest in developing an Nav1.7-
probably through mechanisms that involve the con- specific channel blocker.
trol of trafficking. The fact that a chronic
inflammatory dominant human disease, primary
erythermalgia, maps to this gene SCN10A suggests 7.5 Nav1.8 and Nociception
that this channel may have an important role in
inflammatory pain (Yang, Y. et al., 2004). Tissue- Nav1.8 is a TTXr sensory neuron-specific channel
specific deletion in mouse nociceptors shows that mainly expressed in nociceptive neurons. This chan-
Nav1.7 plays a major role in inflammatory pain nel contributes much of the sodium current
(Nassar, M. A. et al., 2004). Low-dose lidocaine both underlying the depolarizing phase of the action
inhibits the pain associated with primary erythermal- potential in cells in which it is present and knockout
gia and unmasks the phenotype of the Nav1.8 null, mice lose all the inactivating TTXr sodium channel
further supporting a role for Nav1.7 in inflammatory activity in DRG neurons (Akopian, A. N. et al., 1999).
94 Sodium Channels

Nav1.8 thus underlies the inactivating TTXr sodium defined as nociceptors. However, the phenotype of
currents that have been found to play a critical role in a mouse Nav1.9 knockout is essentially normal.
many aspects of nociceptor function. Functional
expression of TTXr currents encoded by this chan-
nel is regulated by inflammatory mediators, 7.6 The Diversity of Sodium Channel
including NGF. Both antisense and knockout studies Subtypes
support a role for the channel in contributing to
inflammatory pain, as PGE2-induced hyperalgesia is Given the fairly similar properties of voltage-gated
inhibited by antisense oligonucleotides. Antisense sodium channels why are there so many different
studies have also suggested a role for this protein in types of a-subunits present in nociceptive sensory
the development of neuropathic pain (Lai, J. et al., neurons? A number of explanations are possible:
2002). A late-onset deficit in ectopic action propaga- Firstly, the trafficking in sensory neurons of dif-
tion has been described in the Nav1.8-null mutant ferent sodium channels to distinct cellular locations
mouse together with deficits in mechanohyper- (terminals, nodes of Ranvier, etc.) and the regulation
sensitivity. A redistribution of immunoreactive of this process may provide a number of options to
Nav1.8-like material has also been described in ani- control neuronal excitability in different physiologi-
mal models of neuropathic pain, further supporting cal contexts. Thus trafficking of Nav1.8 into the cell
the lobby for Nav1.8 as a key molecule in the patho- membrane through its interaction with p11, which is
genesis of neuropathic pain. However, neuropathic massively upregulated by NGF, could play an
pain behavior at early time points seems to be normal important role in inflammatory pain other channels
in the Nav1.8-null mutant mouse. do not show this behavior. It is important to remem-
Inflammatory mediators are known to alter the ber that there are no external cues such as nodes of
level of TTXr channel expression, and shift their Ranvier to define the topological organization of
sodium channels in C-fibers the relatively even
threshold of activation to more negative potentials
distribution of sodium channels to promote electrical
through the activation of PKA. Prostaglandin E2, ade-
propagation along the length of a very long axon is a
nosine, and serotonin all increased the magnitude of
process that is not understood, and altered channel
Nav1.8-mediated currents, shifted its conductance
density particularly at C-fiber terminals can have
voltage relation in a hyperpolarizing direction, and
important effects on excitability. The two sodium
increased the rates of activation and inactivation of
channels that are uniquely associated with C-fibers
sodium channels in small-diameter sensory neurons
may be involved in these processes.
in culture. These phenomena can be replicated in
Secondly, different structural features of the chan-
vitro with heterologously expressed Nav1.8 channels, nels means that their response to posttranslational
implicating this TTXr channel in setting peripheral modification by enzymes such as PK is quite distinct.
pain thresholds. In support of a role for the Nav1.8 Phosphorylation on intracellular serines in Nav1.8
channel in pain pathways, an Nav1.8-null mutant increases peak current density, whilst in other channels
shows deficits in inflammatory pain processing, in present in DRG neurons, for example Nav1.7, peak
contrast to its responses to neuropathic damage current diminishes. Primary sequence also determines
(Akopian, A. N. et al., 1999). the repriming characteristics of the different channels
This channel also appears to have an important as well as their threshold of activation, which are
role in visceral pain. TTXr sodium currents were crucial determinants of peripheral pain thresholds.
present in every colonic DRG neuron studied Thirdly, the topological relationship between dif-
(Gold, et al., 2002). Modulation of TTXr I(Na) in ferent sodium channels and channels involved in
colonic afferents may be an underlying mechanism sensing noxious stimuli may also play a role in setting
of hyperalgesia and pain associated with inflamma- thresholds of activation and thus pain thresholds. As
tion of the colon. Consistent with this, the Nav1.8- noxious mechanosensation but not heat sensing is
null mutant has deficits in visceral pain and referred deficient in Nav1.8-null mutant mice, we may argue
hyperalgesia. that this channel is closely apposed to potential
Several strands of evidence also support a role for mechanosensors through a sequence-specific interac-
Nav1.9 as an important contributor to the control of tion with a macromolecular complex, and local
excitability in nociceptive afferents. The gene is depolarizations cause this channel to activate first to
expressed predominantly in neurons functionally generate action potentials. Heat sensors may be linked
 Sodium Channels 95

with other sodium channel subtypes. If true, this Gribble, F. M., and Woods, C. G. 2006. An SCN9A
channelopathy causes congenital inability to experience
would suggest that molecules that disrupt specialized pain. Nature 444(7121), 894898.
nociceptive complexes at the terminals of sensory Cummins, T. R., Aglieco, F., Renganathan, M., Herzog, R. I.,
neurons may have a useful analgesic effect. There is Dib-Hajj, S. D., and Waxman, S. G. 2001. Nav1.3 sodium
channels: rapid repriming and slow closed-state inactivation
already evidence that disruption of the cytoskeleton display quantitative differences after expression in a
has antihyperalgesic effects. mammalian cell line and in spinal sensory neurons. J.
Neurosci. 21(16), 59525961.
Gold, M. S., Zhang, L., Wrigley, D. L., and Traub, R. J. 2002.
7.6.1 Selective Sodium Channel Blockers Prostaglandin E(2) modulates TTX-RI(Na) in rat colonic
sensory neurons. J. Neurophysiol. 88(3), 15121522.
as Drugs Hains, B. C., Klein, J. P., Saab, C. Y., Craner, M. J., Black, J. A.,
and Waxman, S. G. 2003. Upregulation of sodium channel
Tetrodotoxin has proved invaluable in discriminating Nav1.3 and functional involvement in neuronal
between two classes of sodium channels. The Nav1.5 hyperexcitability associated with central neuropathic
channel, as well as the peripheral neuron-specific chan- pain after spinal cord injury. J. Neurosci. 23(26), 88818892.
Hille, B. 2002. Ion Channels of Excitable Membranes. Sinauer.
nel Nav1.8, is a subfamily of TTXr sodium channels, Isom, L. L. 2002. Beta subunits: players in neuronal
including the evolutionarily earliest sodium channel hyperexcitability? Novartis Found. Symp. 241, 124138;
Nav1.9. However, little progress has been made in discussion 138143, 226232.
Kraner, S. D., Chong, J. A., Tsay, H. J., and Mandel, G. 1992.
developing specific subtype selective channel blockers. Silencing the type II sodium channel gene: a model for
Conotoxins provide a possible route to this goal. The neural-specific gene regulation. Neuron 9(1), 3744.
venom from marine predatory cone shell snails (Conus) Kuwabara, T., Hsieh, J., Nakashima, K., Taira, K., and
Gage, F. H. 2004. A small modulatory dsRNA specifies the
contains a complex cocktail of neurotoxic disulfide-rich fate of adult neural stem cells. Cell 116(6), 779793.
peptides, termed conotoxins. Conotoxins produce their Lai, J., Lai, J., Gold, M. S., Kim, C. S., Bian, D., Ossipov, M. H.,
toxic effects via the targeted modulation of specific ion Hunter, J. C., and Porreca, F. 2002. Inhibition of
neuropathic pain by decreased expression of the tetrodotoxin-
channels, and there is some evidence that they target resistant sodium channel, Nav1.8. Pain 95(12), 143152.
specific sodium channels. Nassar, M. A., Stirling, L. C., Forlani, G., Baker, M. D.,
Sodium channel subtypes are thus differentially Matthews, E. A., Dickenson, A. H., and Wood, J. N. 2004.
Nociceptor-specific gene deletion reveals a major role for
regulated in different pain states. New subtype selec- Nav1.7 (PN1) in acute and inflammatory pain. Proc. Natl.
tive blockers should therefore prove useful as both Acad. Sci. U. S. A. 101(34), 1270612711.
research tools and potential analgesic drugs. The Ratcliffe, C. F., Qu, Y., McCormick, K. A., Tibbs, V. C., Dixon, J. E.,
Scheuer, T., and Catterall, W. A. 2000. A sodium channel
presence of DRG-specific sodium channel isoforms signaling complex: Modulation by associated receptor protein
presents an exciting opportunity to develop new tyrosine phosphatase beta. Nat. Neurosci. 3(5), 437444.
classes of analgesic drugs that may have quite novel Waxman, S. G., Cummins, T. R., Dib-Hajj, S. D., and Black, J. A.
2000. Voltage-gated sodium channels and the molecular
properties from present-day analgesics with utility in pathogenesis of pain: A review. J. Rehabil. Res. Dev.
some conditions of intractable or chronic pain. 37(5), 517528.
Waxman, S. G., Kocsis, J. D., and Black, J. A. 1994. Type III
sodium channel mRNA is expressed in embryonic but not
adult spinal sensory neurons, and is re-expressed following
Acknowledgments axotomy. J. Neurophysiol. 72(1), 466470.
Wood, J. N., Boorman, J. P., Okuse, K., and Baker, M. D. 2004.
Voltage-gated sodium channels and pain pathways. J.
We thank the Wellcome Trust, the BBSRC, and the Neurobiol. 61(1), 5571.
MRC for support. Yang, Y., Wang, Y., Li, S., Xu, Z., Li, H., Ma, L., Fan, J., Bu, D.,
Liu, B., Fan, Z., Wu, G., Jin, J., Ding, B., Zhu, X., and Shen, Y.
2004. Mutations in SCN9A, encoding a sodium channel
alpha subunit, in patients with primary erythermalgia. J. Med.
References Genet. 41(3), 171174.
Yu, F. H. and Catterall, W. A. 2003. Overview of the voltage-gated
Akopian, A. N., Souslova, V., England, S., Okuse, K., Ogata, N., sodium channel family. Genome Biol. 4(3), 207.
Ure, J., Smith, A., Kerr, B. J., McMahon, S. B., Boyce, S.,
Hill, R., Stanfa, L. C., Dickenson, A. H., and Wood, J. N.
1999. The TTX-R sodium channel SNS has a
specialized function in pain pathways. Nat. Neurosci.
2, 5415489. Further Reading
Cox, J. J., Reimann, F., Nicholas, A. K., Thornton, G.,
Roberts, E., Springell, K., Karbani, G., Jafri, H., Mannan, J., Fauchier, L., Babuty, D., and Cosnay, P. 2000. Epilepsy,
Raashid, Y., Al-Gazali, L., Hamamy, H., Valente, E. M., Brugada syndrome and the risk of sudden unexpected
Gorman, S., Williams, R., McHale, D. P., Wood, J. N., death. J. Neurol. 247(8), 643644.
This page intentionally left blank
 8 Physiology of Nociceptors
M Ringkamp and R A Meyer, Johns Hopkins University, Baltimore, MD, USA
2009 Elsevier Inc. All rights reserved.

8.1 Introduction 97
8.2 Nociceptors in Uninjured Skin 98
8.2.1 C-Fiber Nociceptors 98
8.2.2 A-Fiber Nociceptors 102
8.2.3 Classification of Nociceptors Based on Molecular Markers 103
8.3 Efferent and Trophic Functions of Nociceptors 104
8.4 Acute Pain Sensations and Nociceptor Activity 105
8.4.1 Pain from Heat Stimuli 105
8.4.1.1 Heat pain from glabrous skin is signaled by C-fiber nociceptors 105
8.4.1.2 First pain to heat on hairy skin is signaled by type II A-fiber nociceptors 106
8.4.1.3 Pain to prolonged heat stimuli is signaled by type I A-fiber nociceptors 108
8.4.2 Pain from Controlled Mechanical Stimuli 108
8.4.2.1 Sharp pain is signaled by A-fiber nociceptors 108
8.4.2.2 Pain from tonic pressure is signaled by C-fiber MIAs 108
8.4.3 Cold Pain Sensation 108
8.4.4 Capsaicin-Evoked Pain 108
8.5 Hyperalgesia 109
8.5.1 Primary Hyperalgesia 109
8.5.1.1 Hyperalgesia to heat is mediated by primary afferent sensitization 109
8.5.1.2 Hyperalgesia to mechanical stimuli 109
8.5.1.3 Inflammatory mediators and nociceptor sensitization 111
8.5.2 Secondary Hyperalgesia 111
8.5.2.1 Sensitization of primary afferent nociceptors does not occur 111
5.08.5.2.2 Central mechanisms of secondary hyperalgesia 112
References 113

Glossary
AMH A-fiber mechanoheat-sensitive nociceptor primary hyperalgesia Hyperalgesia that develops
responsive to mechanical and heat stimuli. at the site of injury. Mechanical and thermal hyper-
central sensitization An enhanced neural algesia are present at the zone of primary
response that occurs in the central nervous system hyperalgesia following a cutaneous injury.
(CNS). receptive field Area over which neuron is respon-
CMH C-fiber mechanoheat-sensitive nociceptor sive to mechanical, thermal, chemical, or electrical
responsive to mechanical and heat stimuli. stimulation.
fatigue Decrement in response to repeated secondary hyperalgesia Hyperalgesia that
stimuli. develops in an uninjured tissue surrounding a site of
hyperalgesia Enhanced stimulus-evoked pain. injury. Only mechanical hyperalgesia is present at
MIA Mechanically insensitive afferent. the zone of secondary hyperalgesia.
MSA Mechanically sensitive afferent. sensitization Enhanced neural response.
nociceptor Neuron responsive to noxious or
injurious stimuli.

97
98 Physiology of Nociceptors

8.1 Introduction stimuli of different energy modalities are then applied

to this mechanical receptive field. In early studies, the
A vital function of the sensory nervous system is to response of nociceptors to heat and mechanical stimuli
signal the threat or occurrence of tissue injury. A spe- was tested leading to the nomenclature of C-fiber
cialized class of afferents performs this function. These (CMH) or A-fiber mechanoheat nociceptors (AMH)
afferents respond preferentially to noxious or injurious corresponding to C-fiber or A-fiber nociceptors res-
stimuli and are called nociceptors. Nociceptors are ponsive to heat and mechanical stimuli. Most of these
found in virtually every organ (except the brain), and nociceptors also respond to chemical and/or cold sti-
activity in nociceptive afferents is thought to lead to the muli and therefore are polymodal.
percept of pain. In this chapter, we will focus on noci- Nociceptors that respond to mechanical stimuli are
ceptors innervating skin. Nociceptors from other also called mechanically sensitive nociceptive afferents
somatic tissues have similar properties. (MSAs). Recently, it has become apparent that afferents
The skin contains a number of distinct sensory exist that either are insensitive to mechanical stimuli or
receptors that respond selectively to different types have extremely high mechanical thresholds; these affer-
of innocuous stimuli and encode different aspects of ents are called mechanically insensitive afferents
the sensory experience. For example, several types of (MIAs) or silent nociceptors. These afferents were
low-threshold mechanoreceptors signal different missed in earlier studies of nociceptors that relied on
aspects of touch sensation. Receptors that are exqui- mechanical techniques to search for the receptive field.
sitely sensitive to gentle warming or gentle cooling MIAs have been identified in many different tissues
are responsible for the percepts of warmth and cool- including skin, cornea, knee joint, and viscera. In the
ing, respectively. skin, electrocutaneous stimulation can be used to search
Nociceptors are distinguished from these innocu- for the receptive field of MIAs (see Box 1). As will
ous receptors by their relatively high threshold for become apparent below, cutaneous MIAs differ from
activation. Unlike innocuous receptors, nociceptors MSAs not only in their response to mechanical stimuli
often respond to more than one energy form that is, but also in their response to other stimulus modalities.
they are polymodal. Nociceptors are often subclassi-
fied based on the conduction velocity of their parent
8.2.1 C-Fiber Nociceptors
axon. C-fiber nociceptors are associated with unmye-
linated fibers and have conduction velocities less The response of a typical CMH to a stepped increase
than 2 m s1. A-fiber nociceptors are associated with in temperature in the receptive field is shown in
myelinated fibers and have conduction velocities Figure 1. For stimuli applied to the skin, there are
greater than 2 m s1. five processes that occur to achieve this heat response.
We will first consider the properties of nocicep- First, the heat stimulus needs to be absorbed by the
tors from uninjured skin and how their response is skin. Second, the energy must be transmitted to the
related to sensations of acute pain. We will then receptor terminal that sits below the skin surface.
consider the effects of skin injury on the response of Third, the energy must be converted (or transduced)
nociceptors and how this is related to enhanced pain into a change in membrane potential at the terminal
sensitivity called hyperalgesia. Where possible, elec- of the nociceptor. Molecular mechanisms of heat trans-
trophysiological responses to noxious stimuli will be duction are described in Chapter Molecular Biology of
compared to psychophysical measures of pain. the Nociceptor/Transduction. Fourth, the increase
in membrane potential must be sufficient to generate
an action potential. Finally, the action potential
8.2 Nociceptors in Uninjured Skin must propagate orthodromically to the central
nervous system (CNS) (and to the recording electrode).
In experimental studies of nociceptors, the first task is Thermal modeling studies combined with elec-
to identify where on the skin the afferent is responsive. trophysiological recordings indicate that the heat
This area is called the receptive field. The receptive threshold of CMHs in monkeys depends on the tem-
field is often located based on its response to mechan- perature at the depth of the receptor terminal and not
ical stimuli (e.g., gentle pinching), and then the border on the rate of temperature change (Tillman, D. B.
is mapped with flexible small-diameter cylindrical et al., 1995). Based on this analysis, the majority of
probes that exert a relatively constant force when CMHs have heat thresholds between 39 and 41  C.
they bend (called von Frey probes). Quantitative However, the depth of the heat-responsive terminal
 Physiology of Nociceptors 99

Box 1 Mapping mechanically insensitive afferents

Mechanically insensitive afferents (MIAs) either are completely unresponsive to mechanical stimuli or have such a high threshold
for activation that they cannot be activated by light pinch stimuli applied to the skin. Mechanical stimuli are inadequate to locate
the cutaneous receptive field of these afferents. Furthermore, repetitive noxious mechanical and heat stimuli could sensitize the
nociceptive endings over the course of an electrophysiological experiment. Also, the use of mechanical and thermal stimuli bias
the study toward those afferents that are responsive to these stimuli. Because of these reasons, a different search strategy has
been developed that allows the receptive fields of afferents, including MIAs, to be localized in an unbiased manner. Instead of
using natural stimuli, an electrical stimulus is applied to the skin (see adjacent figure).

Stimulation electrode

eRF Recording electrode

Electrical threshold to initiate action potential

Conduction latency (at threshold)

Latency

Stimulus intensity

A saline-soaked cotton swap is used as a monopolar cathotic search electrode to locate the receptive field. Electrical stimuli are
applied through the cotton swap at the skin over the presumed path of the cutaneous nerve from which the recordings are
obtained. In response to the electrical stimulus, the afferent nerve fiber is excited and an action potential is generated and
conducted to the recording electrode. The latency depends on the conduction velocity of the nerve fiber and the conduction
distance. When the stimulation electrode is moved further distally along the peripheral nerve, the conduction latency (green line) of
the unit increases due to the increase in conduction length. By moving the electrode further distally, the path of the unit can be
traced peripherally toward its receptive field. The receptive field has been located when three phenomena occur. First, the
threshold electrical stimulus intensity (pink line) that is needed to excite the afferent transcutaneously drops to its minimum. This
drop is most likely due to the fact that the afferent nerve fiber ends in peripheral terminals that are located in the superficial layers of
the skin. Therefore, the distance between the stimulation electrode and the nerve endings is minimized. Second, at stimulation
threshold, the conduction latency of the action potential increases to its maximum, since the action potential is generated at the
most distal part of the axon, that is, its peripheral ending. This latency increase is not continuous but a step increase. Third, within
the electrical receptive field (eRF), the latency of the action potential decreases in discrete steps as the stimulus intensity increases.
These sudden latency jumps are thought to be due to a switch of the action potential initiation site, for example, from a distal site of
the axon (like the most distal ending) to a more proximal site (like a branch point) when the stimulus intensity is increased. Skin sites
at which the conduction latency decreases in discrete steps with increasing stimulus intensity belong to the eRF of the unit under
study. The described technique can therefore be used to locate the peripheral receptive field and to delineate its borders.

varies widely (from 20 to 570 mm below the skin The heat response of CMHs increases monotoni-
surface). Since the tissue at the depth of cally with skin temperature above threshold. This
the receptor heats more slowly than the skin surface suprathreshold response varies directly with the
due to thermal inertia, the heat threshold as mea- rate of temperature change at the receptor slow
sured by the surface temperature of the skin can vary temperature changes give lower instantaneous dis-
greatly. charge frequencies than fast temperature changes.
100 Physiology of Nociceptors

1.2

1.0

Normalized heat response

0.8
20 V
1 ms
49 C
0.6

38 C
0.4
Time (s)
Figure 1 Response of a typical C-fiber mechanoheat-
sensitive afferent (CMH) to a heat stimulus. A laser thermal 0.2
stimulator was used to deliver a constant temperature
stimulus (from a 38  C baseline to 49  C for 3 s) to the
receptive field of a CMH on the hairy skin of the hand of an
anesthetized monkey. The large action potential waveform 0.0
corresponds to a CMH that responded with 10 action 1 2 3 4 5
potentials to this stimulus. Inset: High-resolution recording Trial number
of action potential waveforms.
Figure 2 C-fiber mechanoheat-sensitive afferents (CMHs)
exhibit fatigue to repeated heat stimuli. Identical heat stimuli
The heat response of CMHs is strongly influ- were presented every 60 s to the receptive field of CMHs
recorded from the anesthetized monkey. The response
enced by stimulus history. For example, when decreased significantly to the second stimulus and reached a
identical heat stimuli are repeated every 60 s, the plateau after several stimuli. Data were normalized by
response to the second stimulus is about 66% of dividing by the response to the first stimulus. Adapted from
the response to the first (Figure 2). This decrement Peng, Y. B., Ringkamp, M., Meyer, R. A., and Campbell, J.
2003. Fatigue and paradoxical enhancement of heat
in response is called fatigue. The amount of fatigue is
response in C-fiber nociceptors from cross-modal excitation.
inversely related to the interstimulus interval (i.e., J. Neurosci. 23, 47664774, copyright 2003 by the society for
shorter intervals lead to more fatigue) and directly Neuroscience, with permission.
related to the magnitude of the preceding stimulus
(i.e., the larger the preceding stimulus, the greater the
fatigue). As a result of fatigue, the shape of the sti- enhanced, for example, by tissue injury. This sensiti-
mulusresponse function can also vary with stimulus zation may account for aspects of hyperalgesia and is
history. Fatigue can also greatly influence threshold discussed in more detail later in this chapter.
measurements. Complete recovery from fatigue takes CMHs do not respond to mechanical stimulation
more than 10 min. with blunt objects but respond well to application of
A decrease in the heat response can also be pro- sharp objects. Their mean mechanical threshold to
duced by mechanical stimulation of the receptive field von Frey probes is 2.2 bars. The receptive field size is
or electrical stimulation of the parent axon (Peng, Y. B. around 20 mm2 in monkeys but can be much larger in
et al., 2003). Thus, fatigue to one stimulus modality can humans. The receptive field typically consists of
be produced by a stimulus of a different modality. multiple punctate regions of high mechanical sensi-
Interestingly, recovery from cross-modal fatigue is tivity surrounded by areas of lesser sensitivity.
much faster than recovery from fatigue produced by Presumably these hot spots correspond to the loca-
the same stimulus modality. This suggests that cross- tions of the receptor terminals.
modal fatigue does not effect the stimulus transduction For cylindrical probes, the response at a given
apparatus in the same way as unimodal fatigue. force increases as the size of the probe decreases
In contrast to fatigue, which leads to a decreased (Garell, P. C. et al., 1996). This relationship scales
response, the response of nociceptors can also be inversely with a linear dimension of the probe (i.e.,
 Physiology of Nociceptors 101

probe circumference) not probe area. Thus, the ade- exhibit a slowing of conduction with repetitive elec-
quate stimulus for activating nociceptors appears to trical stimulation that is called activity-dependent
be related to force per unit length, and activation of slowing. C-MIAs show significantly more slowing
the receptor terminals appears to occur at the edge of than C-MSAs (Weidner, C. et al., 1999). One possible
the probe where the strain is the greatest. explanation for this finding is that the relative distri-
C-fiber MIAs are by definition insensitive to mecha- bution of ion channels involved with conduction is
nical stimuli but are more responsive to chemical different in these two classes of afferents.
stimuli than CMHs and may be considered as chemo- C-fiber afferents with extremely low thresholds to
sensors. For example, microneurographic recordings in mechanical stimuli have been identified in the hairy
human subjects (see Box 2) have shown that capsaicin skin of rodents, monkeys, and humans (Nordin, M.,
(the active ingredient in hot peppers) evokes a vigorous 1990). These afferents are responsive to light stroking
response in C-fiber MIAs. Histamine also vigorously of the skin but not to heat. They are thought to be
activates a subpopulation of C-MIAs. In contrast, his- involved in the sensation of pleasant touch and there-
tamine and capsaicin evoke weak responses in CMHs. fore may play a role in affiliative behavior.
Interestingly, MIAs can develop a response to mechan- C-fiber afferents with extremely low thresholds to
ical stimuli after skin injury (discussed in more detail heat are prevalent in all species. These afferents do
below). not respond to mechanical stimuli but have ongoing
C-MIAs also differ from C-MSAs with regard to activity at room temperature and respond to gentle
their conductive properties. Most unmyelinated fibers warming of the skin (Darian-Smith, I. et al., 1979).

Box 2 Marking technique

The marking technique is used in recordings that are hampered by a small signal-to-noise ratio that makes the identification of an
action potential based on its waveform impractical. The marking technique is commonly used in human microneurography
experiments, in which the signal-to-noise ratio is usually small. To use the marking technique, the primary afferent is electrically
activated at a constant frequency (e.g., one stimulus every 4 s) through a pair of needle electrodes that are inserted intradermally
close to the receptive field of the unit. In response to the ongoing low-frequency electrical stimulation, an action potential can be
recorded at the recording electrode at a constant, fixed conduction latency (see the following figure).

Electrical stimulation

Cutaneous
stimulation

Time

If the unit is also activated by a cutaneous (mechanical, thermal, and chemical) stimulus (see arrow), the conduction latency of
the following electrically induced action potential will increase. This increase in conduction latency marks the unit as being
activated by the stimulus applied to the receptive field. With ongoing electrical stimulation, the conduction latency slowly recovers
to the original latency. Marking obviously indicates slowing of conduction along the course of the axon and is thought to be due to
the additional action potentials that were initiated by the mechanical (or heat or chemical) stimulus and were conducted along
the axon. Due to these additional action potentials, more sodium channels along the axonal membrane will be in an inactivated
state when the next action potential induced by the ongoing electrical stimulation is generated and conducted. As a consequence
of this inactivation, the conduction velocity for this action potential is decreased, leading to an increase in conduction latency.
102 Physiology of Nociceptors

Table 1 Properties of type I and type II A-fiber nociceptors

Property Type I Type II

Heat threshold (short-duration stimuli) High Low

Heat threshold (long-duration stimuli) Low Low
Response to heat Slow onset, slowly increasing Fast onset, rapidly adapting
Mechanical threshold Low (mostly MSAs) High (mostly MIAs)
Conduction velocity A and A fibers A fibers
Sensitization after heat injury Yes No
Location Hairy and glabrous skin Hairy skin

short-duration heat stimuli after injury. Heat trans-

duction in type I afferents is thought to be subserved
C-fiber by the transient receptor potential channel V2
(TRPV2) ion channel. The mechanical threshold and
receptive field size for type I AMHs are comparable to
those of CMHs. The mean conduction velocity of
Type II A type I fibers is around 25 m s1 with some fibers hav-
ing conduction velocities as high as 55 m s1. Thus,
their conduction velocity overlaps the traditional A
and A myelinated fiber range. Type I AMHs are
found in both hairy and glabrous skin.
Type I A
Type II A-fiber nociceptors are characterized by a
fast response to heat stimuli that quickly adapts
(Figure 3). Their heat thresholds to short-duration
stimuli are much lower than those of type I AMHs.
Heat Most type II fibers have very high mechanical
stimulus thresholds or do not respond to mechanical stimuli
and therefore are MIAs. Their mean conduction
Time velocity (15 m s1) places them within the A myeli-
Figure 3 Response of nociceptors to a long-duration
nated fiber range. In monkeys, they are only found in
intense heat stimulus. The response of C-fiber nociceptors hairy skin.
is quick and slowly adapts. The response of type II A-fibers Similar to CMHs, A-fiber nociceptors do not
is also quick but quickly adapts. The response of type I respond to mechanical stimulation with blunt object
A-fibers is slow and builds during the stimulus. but respond well to sharp objects. The response
to mechanical stimuli varies with position in the
The ongoing activity is suppressed by cooling. These receptive field. When a 100 mm wide blade is
afferents are thought to serve the percept of warmth systematically moved to different positions within
and are often referred to as warm fibers. the receptive field, highly reproducible receptive
field response maps can be obtained that contain
multiple peaks and valleys (Slugg, R. M. et al., 2004).
8.2.2 A-Fiber Nociceptors
As the force increases, the response and width of the
Two distinct types of heat-sensitive A-fiber nocicep- peaks increase, the response in the valleys increases,
tors have been described (Table 1) (Treede, R.-D. and new peaks emerge (Figure 4(a)). Although the
et al., 1998). Type I fibers are responsive to mechanical response increases with force in most positions in
and chemical stimuli but have relatively high heat the receptive field, there are positions where the
thresholds when short-duration stimuli are used. For response reaches a plateau at higher forces
this reason, they have also been called high-threshold (Figure 4(b)). The averaged response across the
mechanoreceptors (HTMs). However, most type I receptive field provides an estimate for the response
A-fibers respond to long-duration heat stimuli in the population of nociceptors that innervates a
(Figure 3), and their thresholds to long-duration heat particular location (Figure 4(c)). The population
stimuli are low. In addition, they develop a response to response function for A- and C-fiber nociceptors
 Physiology of Nociceptors 103

(a) increases monotonically with force. The main differ-

30 ences between A- and C-fiber nociceptors are (1)
Evoked response (AP) 25 A-fibers in general exhibit a larger response than
20 C-fibers, (2) the population response of A-fibers
quadruples with every doubling of force, whereas
15
the response of C-fibers doubles with every doubling
10 of force, and (3) the A-fibers exhibit a better differ-
5 ential response with respect to different probe sizes
and forces (Garell, P. C. et al., 1996; Slugg, R. M. et al.,
0
2000).
0 1 2 3 4
Position (mm) A-fiber afferents with ongoing activity are pre-
sent in monkeys and humans. The neural activity in
(b)
30 these afferents increases with gentle cooling and
decreases with gentle warming (Darian-Smith, I.
Evoked response (AP)

25
et al., 1973). These afferents do not respond to
20
mechanical stimulation with thermally neutral
15 probes. These afferents are thought to subserve the
10 percept of cool.
5
0 8.2.3 Classification of Nociceptors Based
40 80 160 320 640
on Molecular Markers
Force (mN) In addition to a classification that is based on receptive
(c) properties, nociceptive afferents can be categorized
20
using molecular markers that are expressed in their
Evoked response (AP)

15 cell bodies or on their cell membrane. Markers that

have been used for this purpose include neuropep-
10 tides, enzymes, and proteins involved in signal
transduction, receptors for growth factors, and other
5 molecules on the cell surface.
The somata of slowly conducting nociceptive
0 afferents are small. In contrast to large cells that can
40 80 160 320 640 be labeled with RT97, an antibody that is directed
Force (mN) against neurofilament protein (NF200), only some of
Figure 4 Response of a type I A-fiber nociceptor to a
the small cells can be labeled with RT97. Small,
blade stimulus. (a) Maps of response as a function of RT97-positive neurons most likely represent the
position and force. A 100 mm wide blade was somata of A nociceptive fibers. RT97-negative
systematically applied to every position across the cells likely represent unmyelinated neurons.
receptive field. The response shows peaks and valleys Nociceptive afferents are commonly classified as
that grow in height and width as the stimulus strength
increased (forces: 160, 320, and 640 mN). (b) Stimulus
peptidergic or nonpeptidergic (Figure 5). Peptidergic
response function varied with position in the receptive cells express substance P (SP) and/or calcitonin gene-
field. In most positions, the response increased related peptide (CGRP) or somatostatin (SOM). In
monotonically with force, but in one position shown the rats, about 50% of neurons are peptidergic (Lawson,
response reached a plateau at the higher forces. S. N., 1992). Peptidergic cells also express the tyrosine
(Symbols in (a) indicate the positions plotted.) (c) The
average response across the receptive field is plotted.
kinase receptor A (TrkA), the receptor for nerve
This provides an estimate of how the population of growth factor (NGF), suggesting that they are NGF
nociceptors that innervate a single spot on the skin would dependent. In addition, peptidergic cells express the
respond to the stimulus. Adapted from Slugg, R. M., TRPV1 channel that is activated by noxious heat sti-
Campbell, J. N., and Meyer, R. A. 2004. The population muli and protons and also mediates the responsiveness
response of A- and C-fiber nociceptors in monkey ncodes
high-intensity mechanical stimuli. J. Neurosci. 24,
to vanilloids (capsaicin and resiniferatoxin). Centrally,
46494656, copyright 2004 by the Society for peptidergic neurons project to lamina I and the outer
Neuroscience, with permission. lamina II of the dorsal horn.
104 Physiology of Nociceptors

DRG in their peripheral terminations, and in their central,

dorsal horn projections.
IB4, c
While the classification into peptidergic and non-
P2X3 Dorsal horn peptidergic cells is commonly used, it should be
emphasized that there is an overlap of marker expres-
Lamina I
A sion between peptidergic and nonpeptidergic neurons.
Lamina IIo For example, some IB4 neurons express TRPV1or
Lamina IIi CGRP. Furthermore, there are considerable species
differences in the expression and coexpression of these
markers. For example, in mice only a minority of IB4
cells express TRPV1 (Zwick, M. et al., 2002), whereas
Figure 5 Small-diameter dorsal root ganglion neurons
can be divided into two classes based on molecular
in rats the majority of IB4 and IB4 cells express
markers in their cell bodies. The neurons in one class TRPV1 (Michael, G. JC. and Priestley, J. V., 1999).
(shown in pink) are tyrosine kinase receptor A (TrkA), Finally, the expression of these markers is not rigid but
transient receptor potential V1 (TRPV1) channel, calcitonin changes during development and after injury. Thus,
gene-related peptide (CGRP), and substance P (SP)- peripheral inflammation increases the proportion of
positive and project to lamina I and lamina IIo of the dorsal
horn. The neurons in the other class (shown in blue) are IB4,
cells expressing neuropeptides, whereas following
c-ret, and P2X3 positive and project to lamina IIi. nerve injury, the proportion of cells expressing neuro-
peptides decreases.
Nonpeptidergic cells contain fluoride-resistant
acid phospatase (FRAP) and bind the plant lectin
8.3 Efferent and Trophic Functions of
IB4 from Griffonia simplicifolia. About 50% of dorsal Nociceptors
root ganglion cells are IB4 positive (IB4). In contrast
to peptidergic cells, IB4 neurons depend on glial In addition to serving the afferent function of signal-
cell-derived neurotrophic factor (GDNF) and they ing pain sensation, nociceptors also mediate efferent
express the necessary receptors, tyrosine kinase Ret effects such as the flare that surrounds an injury site.
(c-ret), and glial cell-derived neurotrophic factor This flare is thought to be due to an axonreflex
receptors (GDNFR14). IB4 cells express the mechanism, since the flare is absent in denervated
P2X3 receptor, a purinergic ligand-gated ion channel tissue but present when a cutaneous nerve is blocked
that is activated by adenosine triphospate (ATP). at a proximal location. When an action potential is
The central terminals of IB4 neurons terminate in initiated in the cutaneous terminals, it propagates not
the inner sheet of lamina II. Some IB4 neurons also only orthodromically up the axons toward the CNS
express mas-related genes A and D (mrgA and mrgD) but also antidromically into peripheral branches of
or mas-related G protein-coupled receptors A and D the nerve fiber where it causes the release of neuro-
(mrgprA and mrgprD) (Dong, X. et al., 2001). IB4 peptides that are present in the terminals (Figure 6).
and mrgA neurons are mostly P2X 3 and express Since afferent and efferent paths of this reflex arc are
c-ret only at a low level. In contrast, IB4 and mrgD within the same axon, this reflex has been coined an
neurons are also P2X 3 and express high levels of axon reflex. The peripherally released neuropeptides
c-ret (Dong, X. et al., 2001). IB4 fibers innervate include SP, neurokinin A (NKA), CGRP, vasoactive
many tissues of the body, including muscle and visc- intestinal polypeptide (VIP), and SOM.
eral organs. However, IB4/mrgpd fibers are In animals, SP and CGRP are released from
exclusively found in skin, where they terminate in capsaicin-sensitive nerve endings and may induce
the stratum granulosum of the epidermis, whereas vasodilation and plasma extravasation. These neuro-
CGRP-containing nerve fibers end less superficial peptides are also thought to stimulate epidermal cells
in the stratum spinosum (Zylka, M. J. et al., 2005). In (e.g., Langerhans cells and kerotinocytes) and to play
addition, IB4/mrgpd neurons appear to project to a role in immunological processes such as the migra-
a distinct lamina of the dorsal horn, that is, between tion of leukocytes at sites of tissue injury. In human
the outer and the inner sheet of lamina II. Thus skin, the antidromic release of neuropeptides is
peptidergic and nonpeptidergic nociceptive neurons thought to lead to vasodilation, but not plasma extra-
appear to differ in growth factor dependency, in ion vasation (Schmelz, M. and Petersen, L. J., 2001). The
channel expression involved in signal transduction, area of the axon reflexive flare in humans is much
 Physiology of Nociceptors 105

Mast cell
degranulation
Noxious
stimulation Keratinocyte
proliferation 3 Arteriole dilation
2
1 CGRP
SP
Immune cell
stimulation
Postcapillary
venule

Plasma 4
extravasation

Axon
reflex NKA 5

Smooth muscle
contraction
CNS

Figure 6 Nociceptors have an efferent function. Noxious stimulation of the nociceptor terminals leads to action potential
propagation not only to the CNS but also into other peripheral branches of the nociceptor where they cause the release of
neuropeptides such as calcitonin gene-related peptide (CGRP), SP, or NKA. The release of neuropeptides from nociceptors
can lead to (1) proliferation of keratinocytes, (2) stimulation of immune cells or degranulation of mast cells, (3) vasodilation,
(4) plasma extravasation, and (5) smooth muscle contraction. Artwork by Ian Suk, Johns Hopkins University. Reproduced
from Meyer, R. A., Ringkamp, R., Campbell, J. N., and Raja, S. N. 2005. Peripheral Mechanisms of Cutaneous Nociception. In:
Wall and Melzacks Textbook of Pain, 5 edn. (ed. M. K. S. McMahon), pp. 334. Elsevier, with permission from Elsevier.

Table 2 Roles of different afferents in acute pain human subjects are compared to the neuronal response
sensation in nociceptors when both are exposed to identical sti-
muli. A number of different scaling techniques can be
Afferents
Percept responsible
used by humans including a visual analog scale (VAS)
and the magnitude estimation technique (see Box 3)
Heat pain on glabrous skin CMHs
First pain to heat on hairy skin Type II A-fibers
Second pain to heat on hairy skin CMHs
Pain to sustained heat stimulus Type I AMHs 8.4.1 Pain from Heat Stimuli
Pain to sharp mechanical A-fiber nociceptors
8.4.1.1 Heat pain from glabrous skin is
stimulus
Pain to sustained pressure C-MIAs signaled by C-fiber nociceptors
Pain to capsaicin C-MIAs, A-MIAs Near pain threshold, only two classes of fibers
respond to heat stimuli applied to the glabrous skin,
warm fibers, and CMHs. Type II A-fiber nociceptors
larger than the receptive field size of CMHs. C-MIAs are not found on glabrous skin, and type I AMHs
have large receptive field sizes and could therefore have high heat thresholds. Warm fibers respond to
account for the large area of flare. The high threshold gentle warming, but their response reaches a peak at
needed to induce flare by electrocutaneous stimula- temperatures at or below the threshold for pain
tion is also consistent with mediation by MIAs (Figure 7) and then decreases for higher tempera-
(Schmelz, M. et al., 2000). tures. In contrast, the response of CMHs starts at or
below the heat pain threshold, and the response
increases with temperature into the painful range.
8.4 Acute Pain Sensations and The stimulusresponse functions for CMHs and for
Nociceptor Activity human pain ratings match well (Figure 7, Box 3).
There are a number of additional lines of corrobor-
We now address the role of different classes of nocicep- ating evidence that CMHs encode heat pain sensation
tors in acute pain sensation (Table 2). To do this, from glabrous skin: First, the long reaction time in
psychophysical measurements of pain intensity in human subjects to step increases in temperature is
106 Physiology of Nociceptors

Box 3 Psychophysical measurements of pain

A number of different techniques exist to measure the intensity of pain in human subjects. One of the most commonly used
techniques is the visual analog scale (VAS) in which descriptors of pain are placed along the edge of the scale. This technique is
very useful for measuring moderate levels of pain but suffers from ceiling effects for high levels of pain (i.e., once the subjects
rates a given stimulus near the top of the scale, there is little room left for stimuli that may be much more intense).
Another method that is extremely useful for correlations with neural responses is the technique of magnitude estimation. In
this procedure, the subjects are presented a moderately painful stimulus and asked to assign a number (e.g., 100) to their
perceived intensity of that stimulus. Subsequent stimuli are then rated relative to this modulus. For example, if the next stimulus
is twice as intense, the rating would be 200; half as intense, the rating would be 50. This technique does not suffer from ceiling
effects, since the subject can use whatever numbers are needed to indicate the relative intensity of the stimulus. This is
basically a ratio scale, since all ratings are made relative to the modulus. Ideally, the modulus is repeated during the course of
the experiment to refresh the subjects memory.
An example of the use of the magnitude estimation technique to measure the intensity of heat pain in human subjects is
shown in the figure.

2
Normalized response

0
40 42 44 46 48 50
Temperature (C)

In this experiment, the glabrous skin of the hand was exposed to a random presentation of temperatures ranging from 41 to 49  C in
1  C increments. The random sequence was preceded by the presentation of a 45  C stimulus that was used by the subjects to
establish the reference rating (i.e., their modulus). To combine data across subjects who used different numbers for their modulus,
data were normalized by dividing the ratings to a given stimulus by the ratings to the first stimulus. An identical sequence
of temperatures was presented to the receptive field of CMHs recorded from the anesthetized monkey. The data across the
population of CMHs were normalized by dividing the response of the nociceptors to a given temperature by the response to the initial
45  C stimulus. The close correlation between the psychophysical data and the electrophysiological data provides strong evidence
that CMHs encode heat pain from glabrous skin. Figure reproduced from Meyer, R. A. and Campbell, J. N. 1981b. Peripheral neural
coding of pain sensation. Johns Hopkins APL Tech. Dig. 2, 164171, with permission from APLTD.

Human
consistent with peripheral action potential propaga-
pain ratings tion in the slowly conducting unmyelinated fibers.
Second, selective blockade of peripheral conduction
Response

in myelinated fibers does not abolish heat pain sensa-

Warm fibers tion. Third, the fatigue to repeated heat stimuli
C-fibers described above for CMHs is also observed in psycho-
physical studies of pain to repeated heat stimuli.
Fourth, in human microneurography experiments,
30 40 50 intraneural electrical stimulation of presu-mably sin-
Temperature (C) gle CMHs elicits pain sensations.
Figure 7 C-fiber nociceptors signal heat pain sensation from
the glabrous skin. The response of warm fibers reaches a peak 8.4.1.2 First pain to heat on hairy skin is
at relatively low temperatures and decreases as the
signaled by type II A-fiber nociceptors
temperature approaches the painful range. The response of
C-fiber nociceptors starts near the pain threshold and In the hairy skin of the arm, stepped increases in
increases with temperature. The pain ratings of human subjects temperature lead to a double pain sensation; an initial
closely matches the response profile of C-fiber nociceptors. pricking pain is followed after a short lull by a
 Physiology of Nociceptors 107

burning sensation (Figure 8). The reaction time to These findings suggest that the first pain sensation
the pricking pain sensation is too short for conduc- to heat is mediated by myelinated afferents.
tion by slowly conducting unmyelinated fibers in the The type II A-fiber nociceptors are ideally suited
periphery. In addition, first pain sensation is blocked to signal first pain sensation. The quick burst in
during a selective blockade of myelinated fibers. action potential activity at the onset of a heat stimu-
lus seen in type II A-fiber nociceptors is consistent
with the percept of a short-lasting pricking sensation.
(a) (b) The heat threshold of type II A-fiber nociceptors is
also near the heat threshold for first pain sensation.
The lack of type II fibers in glabrous skin is consis-
tent with the absence of first pain to heat on the
glabrous skin (Figure 8).
Figure 9 illustrates the neural responses to
stepped increases in temperature in hairy skin. The
heat stimulus activates a population of CMHs and
Type II A
C-fibers C-fibers type II A-fiber nociceptors with receptive fields that
Pain

Pain

overlap the area of the heat stimulus. At the cuta-

neous terminals, both classes of fibers respond
relatively quickly to the heat stimulus, but the
Time Time
response of the type II A-fibers rapidly adapts. As
Figure 8 Heat pain on hairy and glabrous skin. (a) A heat the neural signal propagates up to the CNS, these
stimulus on hairy skin leads to a dual pain sensation, an initial
pricking pain followed by a burning pain. This is consistent
responses separate in time due to the conduction
with the finding that type II A-fiber nociceptors are found on velocity differences of the myelinated and unmyeli-
hairy skin. (b) A heat stimulus on glabrous skin leads to burning nated fibers. The signal from the type II fibers arrives
pain only. Type II A-fibers are not found on glabrous skin. at the CNS much sooner than the response of the

(e) Pain rating

(d) Neuronal response

in CNS

(c) Neuronal response

in PNS

(b) Nociceptor response

Type ll A at cutaneous terminals
CMH

(a) Heat stimulus

Time
Figure 9 First pain to heat on hairy skin is signaled by A-fiber nociceptors. A stepped heat stimulus on hairy skin (a) leads to
a dual pain sensation, a sharp pain followed after a lull by a burning pain (e). Heat stimuli near pain threshold activate two
classes of nociceptors on hairy skin, type II A-fibers and C-fiber mechanoheat-sensitive afferents (CMHs) (b). The signal that
reaches the central nervous system (CNS) is both delayed and broader than the signal at the receptor terminals. The A-fibers
have a faster conduction velocity than the C-fibers and therefore their signals arrive at the CNS with less delay. The
broadening of the signal is due to the variation in conduction velocity across the population of fibers that respond.
108 Physiology of Nociceptors

CMHs. In addition, the signals from each class of In contrast, C-fiber MIAs develop a response during a
afferents become broader because of the variation in maintained stimulus (Schmidt, R. et al., 2000). In addi-
conduction velocity in each population of fibers that tion, the pain to tonic pressure does not disappear
respond to the stimulus. The double pain response during a selective A-fiber block (Andrew, D. and
reflects the different arrival times at the CNS of Greenspan, J. D., 1999). Thus, it appears that C-fiber
neural signals from these two classes of afferents. MIAs encode the pain to tonic pressure.
This difference in arrival time is more apparent for
stimuli applied to the distal extremities where the
conduction lengths are long. A double pain sensation
is not as apparent at more rostral locations (e.g., the 8.4.3 Cold Pain Sensation
face) where the conduction distances are short. The threshold for cold pain sensation is around 14  C
on hairy skin. This is much farther from resting skin
8.4.1.3 Pain to prolonged heat stimuli is temperature (33  C) than the threshold for heat pain
signaled by type I A-fiber nociceptors (around 45  C). In addition, the slope of the stimulus
A long-duration heat stimulus applied to the glabrous response function for cold pain is shallower than that
skin produces pain that lasts through out the stimu- for heat pain (Morin, C. and Bushnell, M. C., 1998).
lus. However, the response of CMHs adapts to a long Cold pain appears to be signaled, at least in part, from
heat stimulus. In contrast, the response of type I deep receptors since topical anesthetics do not abol-
A-fiber nociceptors builds during the course of a ish it and the latency for the perception of cold pain
prolonged heat stimulus (Figure 3). Therefore, is long.
type I A-fiber nociceptors are thought to account The majority of nociceptors respond to ice
for the pain to sustained high-intensity heat stimuli. applied to the receptive field. A nonselective cation
channel (TRPA1) has been identified that has an
8.4.2 Pain from Controlled Mechanical activation threshold (18  C) near the cold pain
Stimuli threshold and is found in a subset of nociceptors
(Story, G. M. et al., 2003). However, the role of this
8.4.2.1 Sharp pain is signaled by A-fiber channel in signaling the pain to a noxious cold sti-
nociceptors mulus is controversial (Bautista, D. M. et al., 2006;
The force thresholds for sharpness and pain from Kwan, K. Y. et al., 2006). In the presence of an A-fiber
cylindrical objects increase in direct proportion
block, gentle cooling stimuli become painful. It is
with the circumference of the probe (Greenspan, J.
thought that cold-evoked activity in A-delta cold
D. and McGillis, S. L. B., 1991). As noted above, a
fibers normally inhibits pain produced by activation
similar relationship exists between probe circumfer-
of nociceptors.
ence and the response of nociceptors. Sharp pain
sensation is dramatically reduced in the presence of
a selective blockade of A-fiber conduction. In addi-
tion, the reaction time for perception of sharp pain is 8.4.4 Capsaicin-Evoked Pain
short. Thus, A-fiber nociceptors appear to be primar-
ily responsible for encoding the percept of sharp pain. Capsaicin, the active ingredient in hot peppers, pro-
Prolonged topical application of capsaicin leads to a duces a burning percept upon contact with mucous
substantial decrease in heat pain sensitivity but membranes and produces intense pain that lasts for
does not greatly affect mechanical pain sensibility several minutes when injected into the skin. Topical or
(Magerl, W. et al., 2001). This suggests that the A- intradermal capsaicin can also lead to a marked desen-
fibers involved in sharp pain are insensitive to cap- sitization to heat stimuli. The TRPV1 channel
saicin (likely, type I AMHs). expressed by many nociceptors is thought to be
responsible for the pungency of capsaicin. Injection
8.4.2.2 Pain from tonic pressure is of capsaicin into the receptive field of CMHs produces
signaled by C-fiber MIAs a weak response and can lead to a desensitization of
Firm pressure does not hurt if applied for short periods the afferent to heat. Injection of capsaicin into the
of time but becomes painful when sustained for long receptive field of A-fiber and C-fiber MIAs produces
periods of time. The response of mechanically sensitive a long-lasting, strong response, suggesting that these
nociceptors adapts to a constant mechanical stimulus. afferents are responsible for the pain from capsaicin.
 Physiology of Nociceptors 109

8.5 Hyperalgesia heat and mechanical stimuli. Secondary hyperalgesia

occurs in the uninjured tissue surrounding the site of
A tissue injury can lead to an enhancement of pain injury and is characterized by hyperalgesia to
sensibility called hyperalgesia. Hyperalgesia is char- mechanical but not heat stimuli. This dichotomy
acterized by a leftward shift in the stimulusresponse provides evidence that the mechanisms for primary
function relating pain intensity to stimulus intensity and secondary hyperalgesia differ.
(Figure 10(a)). There is a decrease in the threshold
for pain, an increase in the pain to suprathreshold
stimuli, and ongoing pain. The electrophysiological 8.5.1 Primary Hyperalgesia
correlate for hyperalgesia is sensitization of nocicep-
tive neurons (Figure 10(b)), either of peripheral 8.5.1.1 Hyperalgesia to heat is mediated
nociceptors (peripheral sensitization) or of central by primary afferent sensitization
neurons involved in processing nociceptive input Hyperalgesia to heat is prevalent after a cutaneous
(central sensitization) or both. Sensitization is char- injury and after inflammation. Substantial evidence
acterized by a leftward shift in the neural response at indicates that sensitization of primary afferent noci-
different stimulus intensities. There is a decrease in ceptors to heat accounts for this hyperalgesia. For
the threshold for eliciting a response, an increase in example, a burn injury to the glabrous skin of the
the response to suprathreshold stimuli, and sponta- hand leads to marked hyperalgesia to heat and sensi-
neous activity. tization of type I AMHs to heat (Figure 12). Although
Two zones of hyperalgesia are readily identified a burn to the glabrous skin does not lead to sensitiza-
(Figure 11). Primary hyperalgesia occurs at the site of tion of CMHs, a burn injury to the hairy skin does.
injury and is characterized by hyperalgesia to both This suggests that the propensity to sensitize is a
function of skin type. Whether this is due to differ-
(a) Hyperalgesia ences in the milieu or due to differences in the
properties of the neurons is not known.
Hyperalgesia to heat can also develop after inflam-
mation. When a cocktail of inflammatory mediators is
injected into the receptive field of nociceptive affer-
Pain

After ents, marked sensitization to heat stimuli is observed.

Before
8.5.1.2 Hyperalgesia to mechanical
stimuli
Stimulus intensity
Hyperalgesia to mechanical stimuli also occurs after
a cutaneous injury and after inflammation. A number
of possible mechanisms may account for this mechan-
ical hyperalgesia.
(b) Sensitization Nociceptor sensitization. Although a burn injury
leads to sensitization of nociceptors to heat stimuli,
the mechanical threshold of nociceptors is not low-
ered, suggesting that this injury does not produce
sensitization to mechanical stimuli. However, noci-
response
Neuronal

After ceptors are sensitized to mechanical stimuli in

Before
situations of inflammation. For example, injection
into the receptive field of inflammatory substances
leads to a sensitization of MIAs to mechanical stimuli.
Inflammation of the knee joint also leads to a sensi-
Stimulus intensity
tization of MIAs to mechanical stimuli.
Figure 10 Hyperalgesia and sensitization. (a) Spatial summation. When a burn injury is applied at
Hyperalgesia is characterized by a leftward shift in the
stimulus function relating pain intensity to stimulus intensity.
the edge of the receptive field of a nociceptor, the
(b) Sensitization is characterized by a leftward shift in the receptive field expands into the area of injury. As a
stimulus function relating neural response to stimulus result, a mechanical stimulus in the area of injury
intensity. evokes a response in a greater number of nociceptive
110 Physiology of Nociceptors

Heat stimuli Mechanical stimuli

Primary

Pain

Pain
zone
(site of injury)

Stimulus intensity Stimulus intensity

Pain

Pain
Secondary
zone
(uninjured tissue)

Stimulus intensity Stimulus intensity

Figure 11 Primary and secondary hyperalgesia. There are two forms of hyperalgesia. Primary hyperalgesia occurs at
the site of injury and is characterized by hyperalgesia to both mechanical and heat stimuli. Secondary hyperalgesia
occurs in the uninjured tissue surrounding the injury site and is characterized by hyperalgesia to mechanical but not heat
stimuli.

(a) (b)
8
Type I AMHs
3
7 After
2
6 After
Mean normalized response

1
5 Before
0
42 44 46 48
4 Temperature (C)

3
(c)

2 Human judgments
2
CMHs
Before
1 1
Before
After
0 0
42 44 46 48 42 44 46 48
Temperature (C) Temperature (C)

Figure 12 Primary hyperalgesia to heat is signaled by sensitization of primary afferent nociceptors. (a) A burn injury to the
glabrous skin of human subjects leads to a dramatic increase in pain ratings to heat stimuli. (b) The same burn injury leads to a
marked sensitization of type I AMHs in the anesthetized monkey. (c) This burn injury led to a desensitization of CMHs on
glabrous skin. However, burn injuries to CMHs on hairy skin do lead to sensitization. Adapted from Meyer, R. A. and
Campbell, J. N. 1981a. Myelinated nociceptive afferents account for the hyperalgesia that follows a burn to the hand. Science
213, 15271529, with permission from AAAS.
 Physiology of Nociceptors 111

afferents. This will lead to spatial summation in nociceptor terminal sensitivity. Tissue injury can also
the CNS that would be expected to produce more pain. lead to the release or leakage of adenosine and its
Central sensitization. Peripheral sensitization derivatives (e.g., ATP) into the extracellular space and
appears to account, at least in part, for hyperalgesia could therefore activate sensory terminals. Low pH
to mechanical stimuli. In addition, the central sensi- levels found in inflammed tissue activate and modulate
tization phenomena to be described below for nociceptor terminals. During inflammation, cytokines
secondary hyperalgesia may also play a role in pri- (e.g., interleukins and tumor necrosis factor alpha) are
mary hyperalgesia to mechanical stimuli. released by a variety of cells (e.g., macrophages) and are
not only involved in regulating the inflammatory pro-
8.5.1.3 Inflammatory mediators and cess but may also act as algesic substances either by
nociceptor sensitization directly activating nociceptive endings or by increasing
Injury leads to the local release of various endogenous their sensitivity to natural stimuli.
substances that excite or modulate the terminals of As illustrated in Figure 13, these agents can act
primary-afferent nociceptors (Figure 13). For example, directly at specialized receptors on the nociceptor
bradykinin is released on tissue injury (e.g., from the terminal to excite the ending or to alter the sensitivity
plasma) and can excite and sensitize nociceptor term- of the ending. The molecular mechanisms of trans-
inals. Intradermal injection of bradykinin produces pain duction and sensitization are discussed in more
and hyperalgesia. Serotonin, histamine, and prostagla- detail in Chapter Molecular Biology of the
din can also be released from mast cells and alter Nociceptor/Transduction.

Tissue
Platelets damage
Macrophage Immune
CRH cells
lL1

TNF H+
Plasma extravasation lL6 Adenosine
vasodilation LIF
Mast cell
ATP
Glutamate

lL1 NGF ASIC

A2
x3 iGluR Keratinocytes
Bradykinin P2
PAF PGE2 kA mGluR1,5 Endorphins
Tr
Histamine -R
lL1
5HT
2/B1
B
Platelets
EP GlRK
H1 GABAA
Inhibitory
T
5H
PKC SSTR2a
PKA

Ca2+ M2
TTXr
(Nav 1.8/1.9) Gene
H+ TRPV1 regulation
SP

Heat

Figure 13 Inflammatory mediators act on specific receptors located at the nociceptor terminals. Artwork by Ian suk, Johns
Hopkins University. Reproduced from Meyer, R. A., Ringkamp, R., Campbell, J. N., and Raja, S. N. 2005. Peripheral Mechanisms
of Cutaneous Nociception. In: Wall and Melzacks Textbook of Pain, 5 edn. (ed. M. K. S. McMahon), pp. 334. Elsevier.
112 Physiology of Nociceptors

8.5.2 Secondary Hyperalgesia similar to that described above for the axon-reflexive
flare response, the nerve endings could release not only
Secondary hyperalgesia is characterized by hyperal-
vasoactive substances but also substances that produce
gesia to mechanical but not heat stimuli. Two distinct
sensitization of nociceptive terminals. However, a
forms of mechanical hyperalgesia have been reported
number of lines of evidence suggest that spreading
(Figure 14). Punctate hyperalgesia (or static hyperal-
sensitization does not occur. For example, burn or
gesia) is characterized by hyperalgesia to sharp stimuli
that normally cause pain (e.g., stiff von Frey probes mechanical injuries or mustard oil application to one
and pin prick). Stroking hyperalgesia (or dynamic half of the receptive field of nociceptors lead to sensi-
hyperalgesia or allodynia) is characterized by pain to tization of that part of the receptive field but not of the
a light moving touch (e.g., stroking of the skin with a uninjured part of the receptive field. In addition, anti-
soft brush) but not blunt pressure. dromic electrical stimulation of nociceptors, which
should lead to the release of these putative sensitizing
substances from the terminals, does not lead to sensiti-
8.5.2.1 Sensitization of primary afferent
zation. Also, the area of flare is generally smaller than
nociceptors does not occur
the area of hyperalgesia. In addition, flare can be
One possible explanation for secondary hyperalgesia is
induced by substances (e.g., histamine injection) that
that the sensitization process that occurs at the site of
do not lead to the development of hyperalgesia.
injury spreads to adjacent uninjured tissue. In a manner
Finally, flare can spread across the midline of the
body, but hyperalgesia does not.

Injury 8.5.2.2 Central mechanisms of secondary

hyperalgesia
The preponderance of experimental data indicate
that secondary hyperalgesia is the result of plasticity
Flare
in the CNS often called central sensitization
(Figure 15). This central sensitization is initiated by
Stroking hyperalgesia
Ongoing pain Punctate Stroking

Punctate hyperalgesia hyperalgesia hyperalgesia hyperalgesia

(allodynia)

Stroking Punctate Sensitized Sensitized

hyperalgesia hyperalgesia central neuron central neuron
(allodynia) (static
hyperalgesia)
CNS
Adequate Light Punctate PNS
stimulus stroking stimuli
Sensitized Low-threshold
Area Small Large Nociceptor mechanoreceptor
nociceptor

Duration Short Long Skin

Primary zone Secondary zone
Central Yes Yes Figure 15 Secondary hyperalgesia is due to central
sensitization sensitization. An injury or inflammation leads to activation
Primary Low- Nociceptors and sensitization of nociceptors in the zone of primary
afferent threshold hyperalgesia. Activity in the peripheral nociceptors
signal mechano- produces a state of central plasticity in the CNS called
receptors central sensitization (). This sensitization spreads to central
neurons with input from adjacent, uninjured afferents.
Figure 14 Two forms of secondary hyperalgesia to Sensitization to input from nociceptors accounts for
mechanical stimuli. The properties of punctate and stroking punctate hyperalgesia. Sensitization to input from low-
hyperalgesia are different, suggesting that the neural threshold mechanoreceptors accounts for stroking
mechanisms may differ. hyperalgesia (or allodynia).
 Physiology of Nociceptors 113

neural activity in primary-afferent nociceptors and is that selective peripheral blockade of myelinated fibers
already apparent at the level of the dorsal horn. (with conduction maintained in unmyelinated fibers)
Compelling evidence for central sensitization leads to the disappearance of stroking hyperalgesia.
comes from an experiment in which capsaicin was Punctate hyperalgesia appears to be due to central
injected into both the forearms of human volunteers sensitization to input from nociceptors (Figure 15).
(LaMotte, R. H. et al., 1991). In the control arm, the Punctate hyperalgesia, and not stroking hyperalgesia,
capsaicin injection produced intense pain and led to developed when capsaicin was injected into the arm
the development of a flare and a large zone of sec- of a patient with a selective, large-fiber neuropathy,
ondary hyperalgesia. In the test arm, the nerve indicating that small myelinated or unmyelinated
supplying the forearm was anesthetized at a proximal fibers are involved. In addition, enhanced pain to
site prior to the injection. The injection did not hurt woolen fabrics is observed in the zone of secondary
but did lead to the development of an axon-reflexive hyperalgesia. Since the prickliness of woolen fabrics
flare, indicating that the nociceptors had been acti- is encoded by activity in nociceptors, not mechano-
vated and were able to release vasoactive substances receptors, activity in nociceptors likely accounts for
(and also putative sensitizing substances). Three this hyperalgesia. Selective nerve blocks revealed
hours after the capsaicin injection, there was no that punctate hyperalgesia disappeared when A
hyperalgesia on the test arm, but the control arm nerve fibers were blocked, suggesting that A noci-
still had a large zone of secondary hyperalgesia. ceptors are important. The nociceptors appear to be
The characteristics of stroking and punctate capsaicin insensitive since topical application of cap-
hyperalgesia are different, suggesting that the neural saicin produced desensitization to heat but did not
mechanisms are different (Figure 14). For example, eliminate punctate hyperalgesia produced by a sub-
the area of punctate hyperalgesia is typically larger sequent injection of capsaicin adjacent to the treated
than the area of stroking hyperalgesia. In addition, area (Magerl, W. et al., 2001).
the duration of punctate hyperalgesia is longer than
the duration of stroking hyperalgesia. Stroking
hyperalgesia appears to be more dependent on
ongoing input from nociceptors, since cooling or Acknowledgments
anesthetizing the site of injury substantially reduces
the zone of stroking hyperalgesia without greatly This research was supported in part by the National
affecting the zone of punctate hyperalgesia. As Institutes of Health (NS-14447 and NS-026363) and
described below, both forms of secondary hyperalge- by a Stuart S. Janney Fellowship from the Applied
sia are thought to be due to central sensitization Physics Laboratory (RAM).
mechanisms.
Stroking hyperalgesia appears to be due to central
sensitization to input from low-threshold myelinated References
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readily activates low-threshold mechanoreceptors tonic mechanical cutaneous pain: comparison of nociceptor
activity in rat and human psychophysics. J. Neurophysiol.
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fiber population innervating palmar and digital skin of the
adjacent to this area and produced secondary hyper- monkey: responses to cooling pulses. J. Neurophysiol.
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Kenins, P., and Champness, P. 1979. Warm fibers
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mulation of myelinated fibers bypassed the natural to thermal stimuli. J. Neurophysiol. 42, 12971315.
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Anderson, D. J. 2001. A diverse family of GPCRs
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Garell, P. C., McGillis, S. L. B., and Greenspan, J. D. 1996. Schmelz, M., Michael, K., Weidner, C., Schmidt, R.,
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 9 Itch
E Carstens, University of California, Davis, CA, USA
2009 Elsevier Inc. All rights reserved.

9.1 Introduction and Definition of Itch 115

9.2 Is Itch Distinct from Pain? 116
9.3 Pain Inhibition of Itch 117
9.4 Peripheral and Spinal Mechanisms of Itch 117
9.5 Alloknesis (Itchy Skin) and Sensitization 119
9.6 Central Representation of Itch 120
9.7 Animal Models of Itch 120
9.8 Itch Mediators 121
References 123

Glossary
allodynia Pain due to a stimulus that does not nociceptor A receptor preferentially sensitive to
normally provoke pain. a noxious stimulus or to a stimulus, which would
alloknesis Itch elicited by light touching or become noxious if prolonged.
stroking. pain An unpleasant sensory and emotional
hyperalgesia An increased response to a stimulus experience associated with actual or potential
which is normally painful. tissue damage, or described in terms of such
hyperknesis Itch elicited by strong mechanical damage.
stimulation. pruriceptor A sensory receptor preferentially
itch Unpleasant sensation associated with the sensitive to a pruritic stimulus.
desire to scratch.

9.1 Introduction and Definition of such as dry skin and contact or atopic dermatoses
Itch (eczema). Moreover, a variety of systemic disorders
also have itching as a symptom (Krajnik, M. and
Itch is widely considered to be an unpleasant sensa- Zbigniew, Z., 2001), in particular, liver dysfunctions
tion associated with the desire to scratch. The such as cholestasis or biliary cirrhosis (Bergasa, N. V.
definition of itch has recently been refined to include et al., 2000), and renal failure (Murphy, M. and
the following subcategories: pruriceptive (originating Carmichael, A. J., 2000). Neuropathic itch is less
from skin damage), neuropathic (originating from commonly associated with lesions of the spinal cord
nerve damage), neurogenic (originating centrally), (Dey, D. D. et al., 2005) or other central structures.
and psychogenic (Yosipovitch, G. et al., 2003). Itch is While itch associated with, for example, urticaria
frequently associated with common insect bites or (hives) can be relieved by antihistamines acting at
exposure to plants, providing a warning signal to the H1 histamine receptor, itch associated with other
remove the offending stimulus by scratching or skin conditions and systemic diseases is often refrac-
rubbing the affected skin area. Itch can be tory to antihistamines. This suggests that there may
experimentally elicited by cutaneous histamine and be other itch mediators and mechanisms in addition
other chemicals (Schmelz, M. et al., 2003b) and by to histamine, the prototypical itch mediator in nor-
electrical surface stimulation of the skin (Tuckett, R. mal human skin.
P., 1982; Ikoma, A. et al., 2005). However, itch also Itch has historically been considered by some to
commonly occurs under a variety of skin conditions be a low level of pain, and perhaps for this reason

115
116 Itch

the experimental study of itch has until now belied by the following observations. Most impor-
received much less attention compared to pain. tantly, itch can be elicited by focal electrical
Nevertheless, recent studies have provided exciting stimulation of the skin surface (Tuckett, R. P.,
new evidence for the existence of itch-selective 1982; Ikoma, A. et al., 2005) or by intraneural micro-
peripheral receptors and central pathways separate stimulation (Schmidt, R. et al., 1993) and the itch
from those mediating pain (for recent reviews, see becomes more intense but never takes on a painful
Stander, S. et al., 2003; Twycross, R. et al., 2003; quality as the stimulus frequency is increased.
Yosipovitch, G. et al., 2003; 2004; Biro, T. et al., Conversely, pain elicited by intraneural stimulation
2005). This vignette will describe some of the recent never transforms to itch at lower frequencies (Ochoa,
advances in our understanding of itch mechanisms. J. and Torebjork, E., 1989; Handwerker, H. O. et al.,
An overview of the neural basis for itch versus pain 1991). Furthermore, itch and pain rarely coexist and
is provided first, followed by consideration of concurrent noxious stimulation, such as scratching, is
animal itch models and, finally, possible itch medi- well known to inhibit itch. Morphine and other opioids
ators and mechanisms based on human and animal depress pain but frequently elicit or exacerbate itch, as
research. discussed further below. Also, the behavioral manifes-
tation of itch, scratching, is very different from
manifestations of pain that often include withdrawal
9.2 Is Itch Distinct from Pain? of the stimulated limb and/or escape reactions.
Withdrawal from a noxious stimulus serves a protec-
The concept that itch is on a continuum with pain tive function to prevent tissue damage, whereas
prompted the frequency theory (von Frey, M., scratching may represent an attempt to remove a
1922), depicted schematically in Figure 1(a). A low pruritic stimulus that has already impinged upon tis-
firing rate in cutaneous nociceptors connected to a sue. In this regard, humans can localize the site of itch
common central pathway elicits a sensation of itch, elicited by focal histamine application with errors of
while high-frequency firing elicits pain. This idea is less than 10 mm, comparable to localization of punc-
supported by observations that numerous stimuli, tate painful stimuli and only slightly worse than
including histamine (Broadbent, J. L., 1955) elicit localization of tactile stimuli (Koltzenburg, M. et al.,
itch at low intensities and pain at higher intensities, 1993). This would aid the accuracy of scratches direc-
and that both pain and itch are conveyed via the ted to an itchy site. However, two-point discrimination
spinothalamic tract (White, J. C. and Sweet, W. H., for histamine-induced itch is fairly poor, approxi-
1969). The notion that itch and pain are con- mately 1215 cm on forearm skin (Wahlgren, C. F
veyed by a common set of neurons, however, is and Ekblom, A., 1996) consistent with the very large

(a) Skin (b) Skin

Epidermis Pruriceptor Epidermis

H+ H
H+
High freq =
is

pain
is
m

Nociceptor
er

Nociceptor
er
D

Low
freq =
itch
Itch
Pain
Spinal
cord

Figure 1 Theories of itch transmission. (a) Frequency theory. Both pruritic and noxious stimuli excite polymodal
nociceptors in skin, which project to a common population of neurons in the spinal cord. These neurons signal itch at a low
firing rate, and pain at a higher firing rate. H: responsive to histamine. (b) Specificity (or selectivity) theory. Pruritic stimuli
excite a selective population of pruriceptors in superficial epidermis that are histamine-sensitive (H), and that project to a
select population of spinal neurons (blue) in the superficial dorsal horn. Activity in this pathway signals itch. Dermal
nociceptors project to a separate population of pain-signaling spinal neurons (red). Incorporated in this schematic is an
inhibitory connection (black) from pain- to itch-transmitting neurons, reflecting the ability of noxious counterstimuli to
inhibit itch.
 Itch 117

innervation territories of putative pruriceptors -Opioid

(Schmelz, M. et al., 1997) (see below).
For the reasons noted above, it has long been Itch
+
postulated that itch and pain are conveyed via sepa-
rate central pathways, with pain capable of inhibiting -Opioid
itch transmission. This concept is depicted in Pain
Figure 1(b). The separate pathways for itch and Figure 2 Schematic of opioid interactions with itch- and
pain constitute the classic specificity theory. Given pain-signaling spinal neurons. -Opioid agonists (purple)
recent evidence showing a relative, rather than abso- inhibit pain and can induce or exacerbate itch, possibly by
lute, sensitivity of potential itch-signaling fibers to inhibiting interneurons (black) that tonically inhibit itch-
transmitting neurons (blue), i.e., disinhibition. -Opioid
pruritics, the term selectivity is more appropriate. agonists (dark green) also inhibit pain and may excite the
The ability of pain to inhibit itch is incorporated by inhibitory interneuron (black), thus inhibiting itch.
the black inhibitory neuron shown in Figure 1(b),
although interactions between nociceptive and pruri- mediated central mechanisms. One possible scenario
ceptive systems may conceivably occur at supraspinal consistent with this interplay between - and -
as well as spinal levels. opioids is shown in Figure 2.

9.3 Pain Inhibition of Itch 9.4 Peripheral and Spinal

Mechanisms of Itch
Numerous experimental studies have demonstrated
suppression of itch in humans by a variety of counter- Recent studies have provided evidence favoring the
stimuli including cooling, mustard oil, and electrical selectivity theory of itch depicted in Figure 1(b).
cutaneous field stimulation (Bickford, R. G., 1937; Human microneurographic studies have uncovered
Gammon, G. D. and Starr, I., 1941; Murray, F. S. a class of mechanically insensitive cutaneous recep-
and Weaver, M. M., 1975; Frustorfer, H. et al., 1986; tors with slowly conducting unmyelinated afferent
Ward, L. et al., 1996; Nilsson, H. J. et al., 1997). This fibers that respond to cutaneous histamine over a
suppression undoubtedly underlies the antipruritic time course that closely matches that of concomitant
effect of scratching. Opioids acting at the  -opioid itch sensation (Figure 3; Schmelz, M. et al., 1997).
receptor depress pain, yet frequently induce or These potential itch receptors are called pruricep-
exacerbate itch in human patients (Bromage, P., tors. Such histamine-responsive fibers also responded
1981; Bromage, P. R. et al., 1982; Morgan, M., 1987; more weakly to prostaglandin E2, serotonin, and
Ballantyne, J. C. et al., 1988), suggesting that inhibi- acetylcholine, which were capable of inducing weak
tion of tonic nociceptive activity facilitates itch itch when delivered intradermally via microdialysis
transmission. Consistent with this, -opioid antago- (Schmelz, M. et al., 2003b). However, the fibers were
nists reduce histamine-induced itch (Heyer, G. et al., not exclusively pruriceptive, as they also responded
1997a). Intradermal injection of histamine at a site to capsaicin and bradykinin, which induce burning
previously anesthetized by local anesthetic in human pain (Schmelz, M. et al., 2003b), indicating that these
subjects resulted in markedly enhanced itch fibers are itch-selective rather than itch-specific.
(Atanassoff, P. G. et al., 1999). The authors postulated These fibers could also be excited by electrical skin
that histamine excites two types of peripheral recep- stimulation; however, the current intensities were
tor, one conveying itch and the other being much higher compared to electrical currents eliciting
antipruritic (i.e., exciting central itch-inhibitory neu- itch from the wrist (Ikoma, A. et al, 2005) suggesting
rons). Preferential local anesthetic blockade of the that there may be additional pruriceptor subtypes.
antipruritic fibers would unmask pruriceptive input Following the discovery of histamine-selective
leading to increased itch. pruriceptors, a subpopulation of mechanically insensi-
Scratching behavior in mice and monkeys induced tive spinothalamic tract neurons in the cat superficial
by -opioids is reduced by -opioid agonists in mice dorsal horn (lamina I) with temporally similar
(Togashi, Y. et al., 2002; Umeuchi, H. et al., 2003) and responses to histamine was identified (Andrew, D.
monkeys (Ko, M. C. et al., 2003). The agonists are and Craig, A. D., 2001) (Figure 3). Chemical selectivity
also analgesic, suggesting a complex interaction was tested in four cases, with two additionally respond-
between pro- and antipruritic opioid receptor- ing to mustard oil (which elicits burning pain) while the
118 Itch

Primary afferents

Instant frequency (Hz)

10
(human)

0.1

0.01

Histamine iontophoresis (20 mC)

25 Spinal projection neurons
Impulses per 20 s

(cat)
20
15
10
5
0
Itch rating (% VAS)

30 Itch sensation
25 (human)
20
15
10
5
0 2 4 6 8 10 12 14
Time (min)
Figure 3 Time course of histamine-evoked response of an unmyelinated primary afferent fiber (upper panel), and mean responses
of lamina I spinothalamic tract neurons (middle panel) in relation to itch sensation (lower panel). Vertical bar indicates time of delivery
of histamine by iontophoresis to the skin. Adapted from Schmelz, M., Schmidt, R., Bickel, A., Forster, C., Handwerker, H. O., and
Torebjork, H. E. 1997. Specific C-receptors for itch in human skin. J. Neurosci. 17, 80038008 and Andrew, D. and Craig, A. D. 2001.
Spinothalamic lamina I neurons selectively sensitive to histamine: a central neural pathway for itch. Nat. Neurosci. 4, 7276, with
permission from Nature Publishing Group.

other two did not. These latter two units may be dependent expansion of lumbar neuronal receptive
histamine-selective, while the units sensitive to both fields (Jinks, S. L. and Carstens, E., 1998) possibly
histamine and mustard oil resemble lamina I units in reflecting central sensitization of pain rather than
rats that gave prolonged response to serotonin (5-HT; itch, as histamine may be algesic rather than pruritic
also a pruritic) but usually also exhibited phasic in rats (see below; Jinks, S. L. and Carstens, E., 2002).
responses to mustard oil, capsaicin, and other algogens Thus, the bulk of current evidence favors the
(Jinks, S. L. and Carstens, E., 2002). An older study concept of an itch-selective pathway, originating
reported a fraction of wide dynamic range (WDR)- from mechanically insensitive, pruritogen-selective
type neurons to respond to cutaneous application of C-fiber afferents which project to lamina I spinotha-
cowhage (Wei, J. Y. and Tuckett, R. P., 1991). A lamic tract neurons that, in turn, convey itch signals
recent study of identified spinothalamic tract neurons to the lateral thalamus and cortex (Figure 1(b)).
in primate found that about half responded to intra- An alternative view is that spinal WDR neurons
dermal histamine, with all additionally responding to may also contribute to itch, much as they are thought
capsaicin (Simone, D. A. et al., 2004). Moreover, neu- to contribute to signaling pain together with nocicep-
ronal responses to light mechanical stroking were tive-specific neurons. Both pruritic (serotonin,
unchanged after histamine, arguing against a role in histamine) and algesic chemicals (capsaicin, nicotine,
alloknesis for these neurons. The authors conclude formalin) excited neurons in overlapping regions of
that such neurons are more likely to signal pain than the superficial dorsal horn (laminae III) as assessed
itch. Similar findings were reported in the rat for by c-fos (Jinks, S. L. et al., 2002; Nojima, H. et al.,
neurons not identified by ascending projections 2003b). The vast majority of single WDR neurons
(Carstens, E., 1997). Intradermal histamine produced recorded in rat superficial dorsal horn responded to
an N-methyl-D-aspartic acid (NMDA) receptor- both pruritic and algesic chemical stimuli as well as
 Itch 119

noxious heat (Jinks, S. L. and Carstens, E., 2000; 2002).

Importantly, superficial dorsal horn neurons gave antagonists.

Morphine facilitation blocked by -opioid

prolonged responses to serotonin that corresponded

Responses inhibited by -opioid antagonists.
with the time course of serotonin-evoked scratching
behavior in rats (Jinks, S. L. and Carstens, E., 2002) with alloknesis.

Increased mechanical sensitivity consistent

(Figure 4). The neuronal discharge rate during the

response to serotonin was much lower compared to
the phasic responses evoked by capsaicin and other
algesic stimuli. Serotonin elicits dose-related scratch-
9.5 Alloknesis (Itchy Skin) and
ing in rodents that was significantly reduced by opioid
Sensitization
antagonists (Yamaguchi, T. et al., 1999; Nojima, H. and
Intradermal injection of histamine elicits itch sensa-
Carstens, E., 2003a; Nojima, H. et al., 2003b).
tion, a wheal at the injection site, and a surrounding
Furthermore, c-fos expression in the superficial dor-
flare reaction due to histamine-evoked release of
sal horn elicited by intradermal capsaicin, but not
vasodilatory peptides (substance P (SP), calcitonin
serotonin, was attenuated by morphine (Nojima, H.
gene-related peptide). Light stroking in a broad area
et al., 2003b), consistent with serotonin being pruritic
of normal skin surrounding the histamine injection
rather than algesic. If this is the case, then one must
can elicit itch, a phenomenon called itchy skin or
consider the possibility that pruriceptive information
alloknesis (Simone, D. A. et al., 1991). The spread of
is being signaled by superficial WDR neurons, con-
alloknesis was prevented by local anesthesic block
sistent with the frequency theory of itch.
adjacent to the site of histamine injection, indicating
In conclusion, exciting new information strongly
that alloknesis is of neurogenic origin (Simone, D. A.
favors the existence of an itch-selective pathway, but
et al., 1991). Noxious punctuate stimulation in and
it would probably be wise not to completely discount
around the region of alloknesis also elicits itch
the possible added contribution of nonselective neu-
(hyperknesis) (Atanassoff, P. G. et al., 1999).
rons until more complete details about itch signaling
The chronic itch of skin conditions such as atopic
have emerged. Future efforts to characterize itch-
dermatitis suggests the possibility of peripheral or
signaling neurons should test the following proper-
central sensitization along the itch-signaling pathway
ties consistent with itch:
(Ikoma, A. et al., 2004). Moreover, noxious stimuli such
Responses inhibited by scratching and other
antipruritic counterstimuli.
as acetylcholine, acid, heat, and pinprick, which nor-
mally evoke pain, instead elicit itch in atopic
Responses facilitated by morphine (Jinks, S. L.
and Carstens, E., 2000).
dermatitis patients (Vogelsang, M. et al., 1995; Ikoma,
A. et al., 2004). A peripheral sensitization of

(a) (b) 5-HT 1%

5-HT 1% (10 g) (100 g)
30
Scratching bouts per 2 min
Impulses per second

4
20 N = 15

2
10

0 0
0 5 10 15 20 25 30 35 40 0 10 20 30 40
Time (min) Time (min)
Figure 4 Prolonged responses of superficial dorsal horn neurons to 5-HT compared with scratching. (a) Averaged
peristimulustime histogram (PSTH, bin width: 1 s) of responses of superficial dorsal horn neurons to intradermal
microinjection of 5-HT (60 mM; 1 ml) at arrow in pentobarbital-anesthetized rats. Inset shows recordings sites (dots) compiled
on section through L5 dorsal horn. (b) Graph plots mean number of hindlimb scratching bouts (error bars: SEM) directed
toward the site of injection of 5-HT in the nape of the neck in Sprague-Dawley rats. Scratching bouts were counted in 2 min
intervals. Adapted from Jinks, S. L. and Carstens, E. 2002. Prolonged responses of superficial dorsal horn neurons to
intracutaneous serotonin and other irritant chemicals: comparison with scratching behavior. J. Neurophysiol. 87, 12801289.
120 Itch

pruriceptors is suggested by microneurographic application of a noxious cold (5  C) stimulus reduced

recordings from an atopic dermatitis patient, revealing the degree of histamine-evoked cortical activation.
a subset of mechanically insensitive C-fiber afferents Interestingly, coapplication of noxious cold and his-
that exhibited irregular patterns of spontaneous activ- tamine revealed significant activation in midbrain
ity; two of these were activated by pruritic stimulation periaqueductal gray (PAG). This suggests that in
(Schmelz, M. et al., 2003a). Since mechanically insensi- addition to the well-known role of the PAG in pain
tive, histamine-responsive C-fibers normally do not modulation, it may also play a role in modulation of
exhibit any spontaneous activity (Schmelz, M. et al., itch (Carstens, E., 1997; Mochizuki, H. et al., 2003).
1997), their increased firing in atopic dermatitis may These studies indicate that noxious and pruritic sti-
represent sensitization that contributes to spontaneous muli activate overlapping as well as separate cortical
itch. Central sensitization is also likely to contribute to and subcortical regions, lending partial support for
pathological itch (Ikoma, A. et al., 2004). Central sensi- separate central pathways mediating itch versus pain.
tization of pain is mediated partly via excessive
excitation of spinal neurons by nociceptors releasing
glutamate and SP, which act at postsynaptic NMDA 9.7 Animal Models of Itch
and neurokinin-1 receptors to initiate Ca2-dependent
intracellular events resulting in enhanced neuronal Scratching directed toward a site of pruritic stimula-
excitability (Ji, R. R. et al., 2003). Central sensitization tion in rodents is thought to reflect itch sensation
is associated with hyperalgesia (increased pain), allo- (Berendsen, H. H. G. and Broekkamp, C. L. E.,
dynia (pain from a normally nonpainful stimulus), and 1991; Kuraishi, Y. et al., 1995). However, scratching
expansion of mechanosensitive receptive fields. It is is a normal grooming behavior and in some cases may
conceivable that conditions associated with chronic reflect pain rather than itch (DeCastro-Costa, M.
itch may induce central sensitization of itch-signaling et al., 1987). Moreover, hindlimb scratching is elicited
neurons via mechanisms similar to those for central by intrathecal injection of numerous agents including
sensitization of pain (Ikoma, A. et al., 2004). neurokinins, capsaicin, and morphine (Frenk, H. et al.,
1988; Wilcox, G. L., 1988; Ko, M. C. and Naughton,
N. N., 2000), even in spinalized animals (Bossut, D.
9.6 Central Representation of Itch et al., 1988) suggesting direct activation of motor
scratch reflex circuits. Facial scratching is also elicited
Using positron emission tomography (PET), hista- by intracerebroventricular and intramedullary injec-
mine injection in the right upper arm differentially tion of opiates (Konigstein, H., 1948; Thomas, D. A.
activated the contralateral anterior cingulate cortex, et al., 1993; Thomas, D. A. and Hammond, D. L.,
ipsilateral inferior parietal lobule, as well as bilateral 1995; Tohda, C. et al., 1997b; Johnson, M. D. et al.,
supplementary and premotor cortical areas (Hsieh, 1999; Kuraishi, Y. et al., 2000) and other substances
J. C. et al., 1994). Activation of the ipsilateral premotor including adrenocorticotropic hormone (ACTH) and
cortical areas was suggested to represent an urge to many neuropeptides (Share, N. N. and Rackham, A.,
scratch. A more recent PET study using histamine 1981; Van Wimersma Greidanus, T. B., 1986; Van
pin prick on the right arm reported dose-related Wimersma Greidanus, T. B. et al., 1987; Van
activation in premotor and supplementary motor Wimersma Greidanus, T. B. and Maigret, C., 1988;
areas and anterior cingulate gyrus, as well as in con- Van Wimersma Greidanus, T. B. et al., 1988; Van
tralateral primary (but not secondary) somatosensory Wimersma Greidanus, T. B. and Maigret, C., 1991;
and primary motor cortex, prefrontal cortex and Johnson, M. D. et al., 1999).
contralateral insula, and prefrontal and superior Despite the caveats noted above, there is good evi-
temporal cortex (Drzezga, A. et al., 2001). While dence that hindlimb scratching may distinguish
there was some overlap in cortical areas activated between itch and pain. Substances that induce itch in
by itch versus pain, there were notable differences, humans, including SP (Hagermark, O. et al., 1978), the
particularly in the activation of secondary somato- mast cell degranulator compound 48/80 (Fjellner, B.
sensory cortex and thalamus by pain but not itch and Hagermark, O., 1982), serotonin (5-HT;
(Drzezga, A. et al., 2001). A third recent PET study Weisshaar, E. et al., 1997), and platelet activating factor
also reported dose-related activation in many of the (PAF, Fjellner, B. and Hagermark, O., 1985), elicit dose-
same cortical areas by iontophoretically applied his- related scratching in rodents (Berendsen, H. H. G
tamine, but notably also in thalamus. Simultaneous and Broekkamp, C. L. E, 1991; Kuraishi, Y. et al., 1995;
 Itch 121

Woodward, D. F. et al., 1995; Yamaguchi, T. et al., 1999; 2004) have also been developed. One model that
Thomsen J. S. et al., 2001; Jinks, S. L. and Carstens, E., most people can relate to is mosquito-bite-induced
2002). In contrast, capsaicin and formalin, which induce itch. Initial exposure of ICR mice to mosquito bites
burning pain, elicit little or no scratching (Kuraishi, Y. did not result in scratching. However, twice-weekly
et al., 1995; Yamaguchi, T. et al., 1999; Jinks, S. L. exposure of the mice to mosquito bites resulted in a
and Carstens, E., 2002). One notable difference is progressive increase in scratching. Nave mice sensi-
histamine, which is the definitive pruritogen in humans tized by injection of female mosquito salivary gland
(Simone, D. A. et al., 1987; 1991; Hagermark, O., 1995) extract (the presumed antigen) exhibited marked
yet does not elicit scratching in several strains of mice scratching upon the first mosquito bite, indicating
(Kuraishi, Y. et al., 1995; Yamaguchi, T. et al., 1999; that it triggered an acute allergic reaction and asso-
Inagaki, N. et al., 2001) or Sprague-Dawley rats ciated itch (Ohtskua, E. et al., 2001).
(Thomsen, J. S. et al., 2001; Jinks, S. L. and Carstens, E., A potential model of chronic itch is the NC/jic
2002). This might be explained by the much mouse (Tohda, C. et al., 1997a; Yamaguchi, T. et al.,
lower concentration of histamine in cutaneous mast 2001). Within 26 months after birth, a majority of
cells of rodents (Gustafsson, B., 1980; Graziano, F. M., these animals spontaneously develop skin lesions
1988; Purcell, W. M. et al., 1989) in favor of 5-HT (eczema, bleeding, alopecia) associated with exces-
which induces scratching via the 5-HT2 receptor sive scratching that is naloxone-sensitive. These
(Yamaguchi, T. et al., 1999; Nojima, H. and animals did not develop skin lesions, scratching, or
Carstens, E., 2003b). However, other rodent strains increased plasma immonoglobulin G levels when
including the ICR mouse (Maekawa, T. et al., 2000) reared in a specific pathogen-free environment, but
and hairless guinea-pig (Woodward, D. F. et al., 1995) did so within 4 weeks after being transferred to a
exhibit dose-related scratching to histamine, indicat- conventional environment. The NC/jic mouse may
ing the existence of strain differences in the scratch- represent an animal model of itch associated with
inducing potency of itch mediators. chronic dermatitis, affording the possibility to develop
Importantly, opiate antagonists significantly novel antipruritic treatment strategies for clinical cases
attenuated hindlimb scratching elicited by 5-HT of itch that are poorly treated by antihistamines.
(Yamaguchi, T. et al., 1999; Nojima, H. and Carstens,
E., 2003a; Nojima, H. et al., 2003b) or SP (Andoh, T.
et al., 1998). This is consistent with the report that
opiate antagonists reduce histamine-induced itch in 9.8 Itch Mediators
humans (Heyer, G. et al., 1997a) and further supports
scratching as an itch-related response. Histamine is the most important mediator of normal
Additional animal models of acute itch include human itch. It is localized in cutaneous mast cells
hindlimb scratching directed toward the eye in albino along with a variety of other chemicals including
guinea-pigs (Woodward, D. F. et al., 1995). In this serotonin, chymase, and tryptase. Intradermal hista-
model, histamine, 5-HT, PAF, and prostaglandin E2 mine primarily elicits a dose-dependent sensation of
elicited significant, dose-related ocular scratching itch (Melton, F. M. and Shelly, W. P., 1950; Keele,
while even high concentrations of the algesic agents C. A. and Armstrong, D., 1964; Handwerker, H. O.
bradykinin, acetic acid, or saline did not. Another et al. 1987; Simone, D. A. et al. 1987; 1991).
model is biting directed toward the site of 5-HT Microdialysis of histamine in the skin elicited almost
injected into the hind paw of mice (Hagiwara, K. exclusively, itch sensation and strongly excited
et al., 1999). 5-HT (60100 nmol) elicited approxi- mechanically insensitive C-fibers that are considered
mately equal numbers of licks and bites, but only to be pruriceptors as discussed earlier (Schmelz, M.
biting was significantly attenuated by both naloxone et al., 2003b). The itch sensation evoked by histamine
and the 5-HT antagonist, methysergide. In contrast, and other pruritic chemicals (e.g., serotonin, SP, PAF,
formalin elicited hind paw licking but no biting. These vasoactive intestinal polypeptide) is in nearly all cases
results suggest that biting represents a means of nox- prevented by pretreatment with H1 receptor antago-
ious counterstimulation of the paw to relieve itch. nists (Hagermark, O., 1992; Heyer, G. and Magerl, W.,
Animal models of acute allergic (e.g., Inagaki, N. 1995), suggesting a common action via histamine
et al., 2000; Ohtsuka, E. et al., 2001; Nojima, H. and released by mast cell degranulation. However, other
Carstens, E., 2003b) and dry-skin itch (Miyamoto, T. mediators might act independently of histamine
et al., 2002b; Nojima, H. et al., 2003a; Nojima, H. et al., release to induce itch, as discussed further below.
122 Itch

More than 50 years ago, Arthur R. P. and Shelley more frequently pain, and excited most histamine-
W. B. (1955) reported the itch-inducing effects of sensitive C-fiber pruriceptors tested (Schmelz, M.
proteases including mucunain (Shelley W. B. and et al., 2003). Cholinergic agonists elicited dose-related
Arthur, R. P., 1955), a peptidase extracted from spi- scratching in mice that was reduced by opiate and
cules of the velvet bean plant, cowhage, which elicit muscarinic M3 receptor antagonists; nicotine did not
itch upon contact with skin. The serine proteases elicit scratching (Miyamoto,T. et al., 2002).
trypsin and chymase and the cystein protease papain Intradermal injection of the neuropeptide SP eli-
induce itch that may be independent of histamine cits itch (Hagermark, O. et al., 1978; Thomsen, D. A.
(Rajka, G., 1969; Hagermark, O. et al., 1972). Mast cell et al., 2002) via mast cell degranulation (Hagermark,
chymase elicited scratching in mice, and a chymase O. et al., 1978). However, a recent study reported no
inhibitor reduced allergic scratching (Imada, T. et al., itch or pain sensations with up to 105 M SP deliv-
2002). An exciting new development is the discovery ered by intradermal microdialysis, and concluded
of the proteinase-activated receptor (PAR). One sub- that physiological levels of SP are insufficient to
type, PAR-2, is targeted by trypsin and mast cell induce mast cell degranulation (Weidner, C. et al.,
tryptase, is expressed in sensory neurons and other 2000). In rodents, SP elicited dose-dependent
tissues (Steinhoff, M. and Roosterman, D., 2005) and scratching that was reduced by opiate and neuroki-
may be involved in itch (Steinhoff, M. et al., 2003). In nin-1 antagonists, and was at least partly independent
particular, PAR-2 expression is increased fourfold in of mast cell degranulation (Andoh, T. et al., 1998).
patients with atopic dermatitis, and these patients A variety of other mediators may also have some
experience itch upon intralesional application of role in itch. PAF elicits histamine-dependent itch in
PAR-2 agonists that is histamine-independent humans (Fjellner, B. and Hagermark, O., 1985), and
(Steinhoff, M. et al., 2003). Thus, PAR-2 represents scratching behavior in guinea-pigs, which, however,
a novel target for antipruritic drugs, particularly for was reduced by PAF antagonists but not by histamine
conditions like atopic dermatitis in which the itch is H1 antagonists (Woodward, D. F. et al., 1995).
refractory to antihistamines. Leukotriene B4 (LTB4) elicited scratching in mice
Serotonin (5-HT) delivered to the skin induces weak that was attenuated by an LTB4 antagonist (Andoh,
itch in humans (Hagermark, O., 1992; Weisshaar, E. et al., T. and Kuraishi, Y., 1998). LTB4 also plays a role in
1997) and may be involved in the pruritus of polycythe- scratching elicited by intradermal injection of SP
mia vera (Fjellner, B. and Hagermark, O., 1979; (Andoh, T. et al., 2001) and nociceptin (Andoh, T.
Fitzsimons, E. J. et al., 1981) and cholestasis (Schworer, et al., 2004). Endothelin delivered by intradermal
H. et al., 1995; Weisshaar, E. et al., 1999; Jones, E. A. and microdialysis or injection induced short-lasting burn-
Bergasa, N. V., 2000). Intradermal microdialysis of 5- ing itch accompanied by release of histamine and
HT elicited weak-to-moderate itch in some subjects nitric oxide (Wenzel, R. R et al., 1998; Katugampola,
and pain in others, but excited mechanically insensitive, R. et al., 2000). Interleukin-2 (IL-2), but not tumor
histamine-sensitive C-fiber pruriceptors (Schmelz, M. necrosis factor-alpha, given intradermally induced
et al., 2000b; 2003a) in support of a role as itch mediator. weak itch (Wahlgren, C. F. et al., 1995; Darsow, U.
As noted earlier, 5-HT potently induces scratching in et al., 1997) and there is indirect evidence that IL-4
rodents via the 5-HT2 receptor (Yamaguchi, T. et al., and IL-6 may also have some role in pathological itch
1999; Nojima, H. and Carstens, E., 2003b), whereas the (Nordlind, K. et al., 1996; Chan, L. S. et al., 2001).
5-HT3 receptor may be involved under certain chronic The preceding list is by no means comprehensive,
itch conditions in humans (Schworer, H. et al., 1995; but certainly we have now at least scratched the
Weisshaar, E. et al., 1999). surface in terms of identifying peripheral mediators,
Intradermal injection (Hagermark, O. and receptors, and neural pathways involved in signaling
Strandberg, K., 1977) or microdialysis (Neisius, U. the enigmatic sensory quality of itch.
et al., 2002; Schmelz, M. et al., 2003) of prostaglandin
E2 also elicited weak itch in some subjects and
strongly excited histamine-sensitive C-fiber pruri- Acknowledgments
ceptors (Schmelz, M. et al., 2003).
Acetylcholine normally elicits pain, but instead The authors work has been supported by research
induced itch in patients with atopic dermatitis grants from the NIH (#DE13685) and the State
(Heyer, G. et al., 1997b). Intradermal microdialysis of California Tobacco-Related Disease Research
of acetylcholine induced itch in some subjects but Program (#11RT-0053).
 Itch 123

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 10 Thermal Sensation (Cold and Heat) through
Thermosensitive TRP Channel Activation
Makoto Tominaga, National Institutes of Natural Sciences, Okazaki, Japan
2009 Elsevier Inc. All rights reserved.

10.1 Introduction 127

10.2 Heat Receptors 127
10.2.1 TRPV1 (VR1) 127
10.2.1.1 Cloning of TRPV1 and its characterization 127
10.2.1.2 Reduction of temperature threshold for TRPV1 activation: A mechanism for
inflammatory pain 128
10.2.2 TRPV2 (VRL-1) 129
10.2.3 TRPV3 and TRPV4 129
10.3 Cold Receptors 129
10.3.1 TRPM8 129
10.3.2 TRPA1 130
10.4 Thermonociception by Non-TRP Proteins 130
10.5 Conclusion 130
References 131

10.1 Introduction to be activated by cold stimuli (Jordt, S. E. et al., 2003;

Patapoutian, A. et al., 2003). Between the two channels,
We feel a wide range of temperatures spanning from TRPA1 is perhaps more likely to be involved in
cold to heat. Within this range, temperatures over nociception because the temperature threshold for its
about 43  C and below about 15  C evoke not only a activation is about 17  C, close to the reported thresh-
thermal sensation, but also a feeling of pain. The old for cold nociceptors.
neurons that allow us to sense temperatures are
located in the dorsal root ganglia (DRG) and within
cranial nerve ganglia such as trigeminal ganglion 10.2 Heat Receptors
(TG). It has been hypothesized that cutaneous noci-
ceptor endings detect temperature and other physical 10.2.1 TRPV1 (VR1)
stimuli by means of ion channels responsive to these 10.2.1.1 Cloning of TRPV1 and its
stimuli. Insight into the molecular nature of the tem- characterization
perature-activated channels came with the cloning of One characteristic shared by many nociceptive neu-
the capsaicin receptor, TRPV1 (Caterina, M. J. et al., rons is sensitivity to capsaicin, the main pungent
1997). TRPV1 is a member of the TRP (transient ingredient of hot chili peppers (Szallasi, A. and
receptor potential) superfamily of ion channels, Blumberg, P. M., 1999). A capsaicin receptor was
whose prototypical member, TRP, was found to be isolated in 1997 and designated vanilloid receptor
deficient in a Drosophila mutant exhibiting abnormal subtype 1 (VR1) (Caterina, M. J. et al., 1997), and
responsiveness to continuous light (Montell, C., 2005). now it is called TRPV1 in TRPV subfamily of a
Three other TRPV channels, TRPV2, TRPV3, and large TRP ion channel superfamily (Figure 1).
TRPV4, have been cloned and characterized as heat Patch-clamp recordings (Caterina, M. J. et al., 1997;
thermosensors (Jordt, S. E. et al., 2003; Patapoutian, A. Tominaga, M. et al., 1998) revealed that TRPV1 is a
et al., 2003). The threshold temperatures for activation nonselective cation channel with high Ca2 perme-
of these channels range from relatively warm (TRPV3 ability and that TRPV1 can be activated not only by
and TRPV4) to extremely hot (TRPV2). In contrast to capsaicin but also by protons and heat over 43  C, all
the four thermosensitive TRPV channels, two other of which are known to cause pain in vivo. Single
TRP channels, TRPM8 and TRPA1, have been found channel openings of TRPV1 by any of the three

127
128 Thermal Sensation (Cold and Heat) through Thermosensitive TRP Channel Activation

(a)
TRPA1 TRPV3 and TRPV4 TRPV2

TRPM8 TRPV1
Cold Heat
5 10 15 20 25 30 35 40 45 50 C
Pain Pain

Temperature
(b) threshold for
Receptor Expression Other effective stimuli
activation
Capsaicin/ proton
Sensory neurons
TRPV1 >43 C lipids
brain/ skin?
2-Aminoethoxydiphenyl borate (2APB)
Sensory neurons/ brain/
Mechanical stimulus
TRPV2 >52 C spinal cord/ lung/
liver/ spleen/ colon 2-Aminoethoxydiphenyl borate (2APB)
Sensory neurons/ 2-Aminoethoxydiphenyl borate (2APB)
TRPV3 >3239 C brain/ spinal cord/ camphor
skin/ stomach/ colon
Sensory neurons/ Hypotonic stimulus/ 4PDD
TRPV4 >2735 C hypothalamus/ skin/ Mechanical stimulus
kidney/ lung/ inner ear Epoxyeicosatrienoic acids
TRPM8 <2528 C Sensory neurons Menthol/ icilin
Allyl isothiocyanate
TRPA1 <17 C Sensory neurons 9-tetrahydrocannabinol
cinnamaldehyde
mechanical stimulus

Figure 1 (a) Temperatures causing pain and activating six thermosensitive transient receptor potential (TRP) channels.
Dotted lines indicate that threshold temperatures for activation of TRPV1 and TRPM8 are not fixed but changeable in the
presence of other stimuli (see text). (b) Comparison of the mammalian six thermosensitive TRP channels.

stimuli in a membrane-delimited fashion indicate of the knockout mouse thermosensory phenotype was a
that the three stimuli gate TRPV1 directly and that virtual absence of thermal hypersensitivity in the setting
TRPV1 is, itself, a heat sensor. These stimuli are of inflammation. These findings indicate that TRPV1 is
likely to work in concert to regulate the activity essential for selective modalities of pain sensation and
of TRPV1 in vivo, especially under pathological for tissue injury-induced thermal hyperalgesia.
conditions where tissue acidosis and elevated tem-
perature may come into play. TRPV1 transcript and 10.2.1.2 Reduction of temperature
protein were found to be most highly expressed in threshold for TRPV1 activation: A
sensory neurons (Caterina, M. J. et al., 1997; mechanism for inflammatory pain
Tominaga, M. et al., 1998). Moreover, in situ hybridi- Inflammatory pain is characterized by hypersensi-
zation and immunostaining revealed that, within tivity both at the site of damage and in the adjacent
DRG and TG, TRPV1 expression predominated in tissue, and one underlying mechanism is the modula-
small-diameter cell bodies, most of which give rise to tion (sensitization) of ion channels such as TRPV1.
unmyelinated C fibers. Sensitization is triggered by extracellular inflamma-
To determine whether TRPV1 really contributes to tory mediators that are released in vivo from
the detection of these noxious stimuli in vivo, mice surrounding damaged or inflamed tissues and from
lacking this protein were generated and analyzed for nociceptive neurons themselves (Julius, D. and
nociceptive function. Sensory neurons from mice lack- Basbaum, A. I., 2001). Tissue acidification is also
ing TRPV1 were deficient in their responses to each of induced in pathological conditions such as ischemia
the reported noxious stimuli capsaicin, proton, and or inflammation, and such acidification exacerbates
heat (Caterina, M. J. et al., 2000). Consistent with this or causes pain. Among the inflammatory mediators,
observation, behavioral responses to capsaicin were extracellular ATP, bradykinin, prostaglandins (PGE2
absent, and responses to acute thermal stimuli were and PGI2), or tryptase or trypsin have been reported
diminished in these mice. The most prominent feature to potentiate TRPV1 responses through their
 Thermal Sensation (Cold and Heat) through Thermosensitive TRP Channel Activation 129

Gq-coupled P2Y2, B2, EP1, IP, or PAR2 (proteinase preferentially expressed in A fibers. TRPV2 tran-
activated receptor 2) receptors, respectively, script and protein were found not only in sensory
mainly in a protein kinase C (PKC)-dependent man- neurons but also in motoneurons and in many non-
ner in both a heterologous expression system and neuronal tissues that are unlikely to be exposed to
native DRG neurons (Tominaga, M. and Caterina, temperatures above 50  C (Caterina, M. J. et al.,
M. J., 2004; Moriyama, T. et al., 2005). In addition to 1999). These results indicate that TRPV2 undoubt-
potentiating capsaicin- or proton-evoked currents, in edly contributes to numerous functions in addition to
the presence of the inflammatory mediators, the nociceptive processing.
threshold temperature for heat activation of TRPV1
was reduced to as low as 30  C, such that normally
nonpainful thermal stimuli (i.e., normal body tem- 10.2.3 TRPV3 and TRPV4
perature) are capable of activating TRPV1. Under TRPV3 and TRPV4 have been found to be activated
these circumstances, the inflammatory mediators by warm temperatures, 3438  C for TRPV3 and
seem to act as direct activators of TRPV1. This 2735  C for TRPV4, in heterologous expression
represents a novel mechanism through which the systems and mouse keratinocytes, and to be
large amounts of the mediators released from differ- expressed in multiple tissues, including, among
ent cells in inflammation might trigger a sensation of others, sensory and hypothalamic neurons and kera-
pain. Interactions between the Gq-coupled receptors tinocytes ( Jordt, S. E. et al., 2003; Patapoutian, A. et al.,
and TRPV1 in relation to inflammatory pain have 2003; Tominaga, M. and Caterina, M. J., 2004)
been proven at behavioral level using the Gq-coupled (Figure 1). Among the tissues, both TRPV3 and
receptor- or TRPV1-deficient mice (Tominaga, M. TRPV4 were most predominantly expressed in the
and Caterina, M. J., 2004; Moriyama, T. et al., 2005). A skin keratinocytes and have been reported to be
PKA-dependent and PIP2-mediated pathways also involved in thermosenation by keratinocytes, based
seem to be involved in TRPV1 sensitization on the studies using wildtype and TRPV3- or
(Tominaga, M. and Caterina, M. J., 2004). TRPV4-deficient keratinocytes (Chung, M. K. et al.,
Tissue acidification is induced in pathological 2004; Lee, H. et al., 2005; Moqrich, A. et al., 2005).
conditions such as ischemia or inflammation, and Thermosensing ability of TRPV3 or TRPV4 were
such acidification exacerbates or causes pain. In addi- proven at animal level because both TRPV3- and
tion to the direct activation of TRPV1, acidification TRPV4-deficient mice exhibited some different
also shifts temperatureresponse curve of TRPV1 to thermal preference from that in wildtype mice.
the left so that the channel can be activated at lower TRPV4 has also been shown to be involved in
temperatures (lower than body temperature) and mechanical stimulus- and hypotonicity-induced
responses to heat are bigger at a given suprathreshold nociception in rodents at baseline or following
temperature (Tominaga, M. et al., 1998). This phe- hypersensitivity induced by prostaglandin injection
nomenon might also contribute to inflammatory pain. or taxol hypersensitivity. Furthermore, TRPV4-
deficient mice reportedly exhibit deficits in inflam-
10.2.2 TRPV2 (VRL-1) mation-induced thermal hyperalgesia (Todaka, H.
et al., 2004). TRPV3 has been reported to function
A protein, which is 49% identical to TRPV1, was as a camphor receptor (Moqrich, A. et al., 2005).
isolated and designated VRL-1 and later renamed
TRPV2 (Caterina, M. J. et al., 1999; Jordt, S. E. et al.,
2003; Patapoutian, A. et al., 2003; Tominaga, M. and
Caterina, M. J., 2004) (Figure 1). TRPV2 can be 10.3 Cold Receptors
activated by high temperatures with a threshold of
10.3.1 TRPM8
52  C. TRPV2 currents showed similar properties
to those of TRPV1 such as high Ca2 permeability. The cooling sensation of menthol, a chemical agent
Intense TRPV2 immunoreactivity was observed found in mint, is well established, and both cooling
in medium- to large-diameter cells in rat DRG neu- and menthol have been suggested to be transduced
rons (Caterina, M. J. et al., 1999). Together with the through a nonselective cation channel in DRG neu-
fact that many of the TRPV2 immunoreactive cells rons (Figure 1). Two groups independently cloned
in rat DRG costained with the marker for myelinated and characterized a cold receptor, TRPM8, which
neurons, it was concluded that TRPV2 is can be also activated by menthol (McKemy, D. D.
130 Thermal Sensation (Cold and Heat) through Thermosensitive TRP Channel Activation

et al., 2002; Peier, A. M. et al., 2002). In heterologous et al., 2003). Furthermore, TRPA1 is frequently coex-
expression systems, TRPM8 could be activated by pressed with TRPV1, raising the possibility that
menthol or by cooling, with an activation tempera- TRPA1 and TRPV1 mediate the function of a class
ture of 2528  C. TRPM8 could alternatively be of polymodal nociceptors. Such coexpression might
activated by other cooling compounds, such as also explain the paradoxical hot sensation experi-
menthone, eucalyptol, and icilin. There also appears enced when one is exposed to a very cold stimulus.
to be interaction between effective stimuli for
TRPM8, in that subthreshold concentrations of
menthol increased the temperature threshold for 10.4 Thermonociception by Non-TRP
TRPM8 activation from 25  C to 30  C. This is remi- Proteins
niscent of TRPV1, whose activation temperature is
reduced under mildly acidic conditions that do not The discovery of thermosensitive TRP channels
open TRPV1 alone (Tominaga, M. et al., 1998). indicates that thermosensation is mediated in part
Whole-cell recording in HEK293 cells expressing by a common molecular mechanism involving ion
TRPM8 revealed that TRPM8 is a nonselective channels such as TRP channels as primary transdu-
cation channel with relatively high Ca2 cers of thermal stimuli. However, it is still possible
permeability. that other proteins are involved in thermosensation
TRPM8 is expressed in a subset of DRG and TG and thermonociception. Indeed, TREK-1, a member
neurons that can be classified as small-diameter C of a family of mammalian two-pore domain K chan-
fiber-containing neurons (McKemy, D. D. et al., 2002; nels, is expressed in mouse DRG neurons and
Peier, A. M. et al., 2002). Interestingly, however, activated by heat. Because opening of TREK-1
TRPM8 is not coexpressed with TRPV1. causes hyperpolarization of the membrane potential
by efflux of K ions, TREK-1 might be involved in
pain relief through the reduction of firing upon nox-
10.3.2 TRPA1
ious heat stimulus. Alternatively, inhibition of
TRPA1 was reported as a distantly related TRP chan- TREK-1 by cold stimuli might lead to nociception
nel which is activated by cold with a lower activation through depolarization. Further studies will be
threshold as compared with TRPM8 (Story, G. M. necessary to determine whether TREK-1 is involved
et al., 2003) (Figure 1). In heterologous expression in thermonociception. In addition to K channels,
systems, TRPA1 was activated by cold stimuli with other membrane proteins such as Na/K ATPase,
an activation temperature of about 17  C, which is members of degenerin/epithelial sodium channels
close to the reported noxious cold threshold. This (DEG/ENaC), and P2X3 receptors have been
finding led to the suggestion that TRPA1 is involved reported to be thermosensative. However, the invol-
in cold nociception. Whole-cell recording in Chinese vement of those proteins in thermonociception
hamster ovary (CHO) cells expressing TRPA1 remains to be established.
revealed cationic permeability with similar prefer-
ences for monovalent and divalent cations. Whether
TRPA1 is gated directly by cold remains to be eluci- 10.5 Conclusion
dated. A study from another group failed to reproduce
cold responsiveness in TRPA1 (Jordt, S. E. et al., 2004). Significant advances in thermonociception research
The reason for this apparent discrepancy is unclear. have been made in the last several years with the
However, both groups have demonstrated that cloning and characterization of thermosensitive TRP
TRPA1 can be activated by pungent isothiocyanate channels. It is not known how multiple thermosensi-
compounds such as those found in wasabi, horseradish, tive TRP channels are used within the same or
cinammon, and mustard oil (Tominaga, M. and different sensory neurons to provide us with the
Caterina, M. J., 2004). Thus, several of the thermo- ability to evaluate temperature precisely over a
sensitive TRP channels likely to be involved in broad range. Central integration of the thermal infor-
nociception can be activated by stimuli other than mation acquired by peripheral nerve endings is
temperature. almost certainly a critical component of this process.
Unlike TRPM8, TRPA1 is specifically expressed Furthermore, regulatory mechanisms similar to those
in a subset of sensory neurons that express the noci- emerging for TRPV1 are likely to exist for most of
ceptive markers CGRP and substance P (Story, G. M. the thermosensitive TRP channels. Therefore, an
 Thermal Sensation (Cold and Heat) through Thermosensitive TRP Channel Activation 131

understanding of the molecules surrounding thermo- McKemy, D. D., Neuhausser, W. M., and Julius, D. 2002.
Identification of a cold receptor reveals a general role for TRP
sensitive TRP channels from the periphery to the channels in thermosensation. Nature 416, 52.
central nervous system will be indispensable to a Montell, C. 2005. The TRP superfamily of cation channels. Sci.
complete understanding of thermonociception. STKE 2005, re3.
Moqrich, A., Hwang, S. W., Earley, T. J., Petrus, M. J.,
Murray, A. N., Spencer, K. S., Andahazy, M., Story, G. M.,
and Patapoutian, A. 2005. Impaired thermosensation in mice
lacking TRPV3, a heat and camphor sensor in the skin.
References Science 307, 1468.
Moriyama, T., Higashi, T., Togashi, K., Iida, T., Segi, E.,
Caterina, M. J., Leffler, A., Malmberg, A. B., Martin, W. J., Sugimoto, Y., Tominaga, T., Narumiya, S., and Tominaga, M.
Trafton, J., Petersen-Zeitz, K. R., Koltzenburg, M., 2005. Sensitization of TRPV1 by EP1 and IP reveals
Basbaum, A. I., and Julius, D. 2000. Impaired nociception peripheral nociceptive mechanism of prostaglandins. Mol.
and pain sensation in mice lacking the capsaicin receptor. Pain 1, 3.
Science 288, 306. Patapoutian, A., Peier, A. M., Story, G. M., and Viswanath, V.
Caterina, M. J., Rosen, T. A., Tominaga, M., Brake, A. J., and 2003. ThermoTRP channels and beyond: mechanisms of
Julius, D. 1999. A capsaicin-receptor homologue with a high temperature sensation. Nat. Rev. Neurosci. 4, 529.
threshold for noxious heat. Nature 398, 436. Peier, A. M., Moqrich, A., Hergarden, A. C., Reeve, A. J.,
Caterina, M. J., Schumacher, M. A., Tominaga, M., Rosen, T. A., Andersson, D. A., Story, G. M., Earley, T. J., Dragoni, I.,
Levine, J. D., and Julius, D. 1997. The capsaicin receptor: a McIntyre, P., Bevan, S., and Patapoutian, A. 2002. A TRP
heat-activated ion channel in the pain pathway. Nature channel that senses cold stimuli and menthol. Cell 108, 705.
389, 816. Story, G. M., Peier, A. M., Reeve, A. J., Eid, S. R.,
Chung, M. K., Lee, H., Mizuno, A., Suzuki, M., and Mosbacher, J., Hricik, T. R., Earley, T. J., Hergarden, A. C.,
Caterina, M. J. 2004. TRPV3 and TRPV4 mediate warmth- Andersson, D. A., Hwang, S. W., McIntyre, P., Jegla, T.,
evoked currents in primary mouse keratinocytes. J. Biol. Bevan, S., and Patapoutian, A. 2003. ANKTM1, a TRP-like
Chem. 279, 21569. channel expressed in nociceptive neurons, is activated by
Jordt, S. E., Bautista, D. M., Chuang, H. H., McKemy, D. D., cold temperatures. Cell 112, 819.
Zygmunt, P. M., Hogestatt, E. D., Meng, I. D., and Julius, D. Szallasi, A. and Blumberg, P. M. 1999. Vanilloid (Capsaicin)
2004. Mustard oils and cannabinoids excite sensory nerve receptors and mechanisms. Pharmacol. Rev. 51, 159.
fibres through the TRP channel ANKTM1. Nature 427, 260. Todaka, H., Taniguchi, J., Satoh, J., Mizuno, A., and Suzuki, M.
Jordt, S. E., McKemy, D. D., and Julius, D. 2003. Lessons from 2004. Warm temperature-sensitive transient receptor
peppers and peppermint: the molecular logic of potential vanilloid 4 (TRPV4) plays an essential role in thermal
thermosensation. Curr. Opin. Neurobiol. 13, 487. hyperalgesia. J. Biol. Chem. 279, 35133.
Julius, D. and Basbaum, A. I. 2001. Molecular mechanisms of Tominaga, M. and Caterina, M. J. 2004. Thermosensation and
nociception. Nature 413, 203. pain. J. Neurobiol. 61, 3.
Lee, H., Iida, T., Mizuno, A., Suzuki, M., and Caterina, M. J. Tominaga, M., Caterina, M. J., Malmberg, A. B., Rosen, T. A.,
2005. Altered thermal selection behavior in mice lacking Gilbert, H., Skinner, K., Raumann, B. E., Basbaum, A. I., and
transient receptor potential vanilloid 4. J. Neurosci. Julius, D. 1998. The cloned capsaicin receptor integrates
25, 1304. multiple pain-producing stimuli. Neuron 21, 531.
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 11 The Development of Nociceptive Systems
G J Hathway and M F Fitzgerald, University College London, London, UK
2009 Elsevier Inc. All rights reserved.

11.1 The Development of Nociceptors and Their Peripheral and Spinal Connections 133
11.2 The Development of Spinal Nociceptive Circuits 135
11.3 The Development of Ascending Pain Pathways 136
11.4 Pain and the Developing Cortex 136
11.5 Pain and the Developing Limbic System 137
11.6 The Development of Descending Pain Pathways 137
11.7 Development of Neurotransmitter/Receptor Function in Descending Pathways 138
11.8 The Development of Persistent Pain 140
11.9 Concluding Remarks 140
References 141

Glossary
A fibers Large-diameter myelinated primary afferent receptive field The area on the body surface that
sensory fibers with large cell bodies in the dorsal root when stimulated evokes action potentials in a given
ganglion. The largest diameter A fibers are mainly neuron.
low-threshold mechanoceptors and the smaller A periaqueductal gray An area of the brainstem that
fibers are both mechanoreceptors and nociceptors. surrounds the aqueduct connecting the third and
C fibers Small-diameter unmyelinated primary fourth ventricles. This area projects to the medul-
afferent sensory fibers with small cell bodies in the lary raphe region, which in turn sends projections
dorsal root ganglion. The majority are nociceptors. down the dorsolateral funiculus of the spinal cord to
They divide into a neuropeptide containing, Trk- the dorsal horn. This pathway is known to strongly
receptor expressing group and a lectin IB4 binding modulate spinal pain processing.
group although the functional implications of this receptor subunits Ion channels are generally
are still unclear. made up of several glycoprotein subunits around a
embryonic (E) age These are dated from the time central pore. These subunits can confer special
of fertilization. Rat gestation is 21.5 days, mouse a characteristics upon a channel, such as increased
little shorter. Rats are born relatively early in terms calcium permeability or longer opening times. The
of CNS development and the early postnatal period subunits of many channels change with develop-
is often paralleled with the final gestation of devel- ment, thereby altering the channel properties.
opment in man.

11.1 The Development of The two main subtypes of primary sensory neurons, A
Nociceptors and Their Peripheral and cells and C cells (future nociceptors) are specified at an
Spinal Connections early stage of development under the control of spe-
cific transcription factors. In the rat, lumbar DRG cells
Pain is usually, although not always, triggered by tissue are born in two waves from embryonic day (E)12E15
damage and consequently a discussion of developing (gestation 21.5 days); C cells are born in the second
nociceptive pathways begins with the maturation of wave, later than the larger-diameter A cells. There is
primary sensory neurons in the dorsal root ganglia evidence also that the future peptidergic nociceptors
(DRG) and their peripheral and central connections. are born before the nonpeptidergic group (Hall, A. K.

133
134 The Development of Nociceptive Systems

et al., 1997; Jackman, A. and Fitzgerald, M., 2000). The growth of C-fiber terminals into the spinal dorsal
Young C cells immediately send out centrally and horn occurs just before birth in the rat and from the
peripherally directed processes, again lagging some outset, these fibers terminate in a somatotopically
days behind A cells. However, from the time that precise manner in laminae III of the dorsal horn.
they innervate peripheral skin, nociceptors are capable Neuropeptide containing nocieptive terminals in
of transmitting information about noxious cutaneous laminae I and II can be detected prenatally, steadily
stimulation (Box 1). increasing in density over the first 10 postnatal days,
The study of sensory neurogenesis, axon out- while IB4ve terminals appear a few days later, sug-
growth, and target finding is a fast-moving and gesting that these fibers may form central connections
exciting area of developmental biology. Not only are later than peptidergic fibers and indeed be involved in
there a number of key regulatory genes controlling a separate central pain pathway (Braz, J. M. et al., 2005).
these events but the many aspects of nociceptor devel- The immature C-fiber terminals are functional at
opment are dependent upon access to neurotrophic birth but neurotransmitter release is of low frequency
factors (Fitzgerald, M., 2005). A key neurotrophic fac- and the synapses are too immature to evoke substan-
tor affecting nociceptor development is nerve growth tial spike activity in vivo until the second postnatal
factor, NGF, which not only determines nociceptor week. This in contrast to the A fiber which, when
fate but is also critically required for nociceptor sur- stimulated, can evoke robust action potentials in
vival (Davies, A. M., 2000; Fitzgerald, M., 2005) dorsal horn cells from before birth (Fitzgerald, M.
during embryonic life and continues to influence and Jennings, E., 1999; Baccei, M. L. et al., 2003). As in
nociceptor development postnatally by the adult, immature C fibers terminate exclusively in
1. NGF regulates axon outgrowth, independently of laminae I and II. However, in the first postnatal
cell survival (Markus, A. et al., 2002). weeks, they share this termination area with large-
2. NGF determines nociceptor physiological prop- diameter A afferents whose terminals extend dor-
erties. NGF signaling in the early postnatal period sally into laminae II and I and form transient synaptic
is required for C nociceptors to respond to nox- connections. This is followed by a gradual withdra-
ious heat and for the normal development of wal from the superficial laminae over the first 3
myelinated high-threshold mechanoreceptors postnatal weeks (Fitzgerald, M. et al., 1994). The
(A fibers) (Ritter, A. M. et al., 1991; Lewin, G. R. postnatal withdrawal of A fibers from the superficial
and Mendell, L. M., 1994). dorsal horn is an activity-dependent process, which
3. NGF influences nociceptor expression of the that can be prevented by intrathecal application of
neuropeptides substance P (SP) and calcitonin- the NMDA receptor antagonist MK801 (Beggs, S.
gene-related peptide (CGRP) (Davies, A. M., et al., 2002). The functional implications of the pre-
2000; Hall, A. K. et al., 2001). sence of A fibers in newborn lamina II are not known
4. NGF regulates skin innervation density. Excess but they may contribute to the larger receptive fields
epidermal NGF leads to hyperinnervation and increased excitability to tactile stimulation
(Davis, B. M. et al., 1997). described below.

Box 1 Postnatal development of C nociceptor properties

Although all of the broad functional classes of primary afferents are present in the rat hindlimb at birth, the maturation
process varies according to the individual stimulus modality (Koltzenburg, M., 1999). From birth, polymodal nociceptors
respond to noxious chemical, mechanical, and thermal stimuli with thresholds and firing frequencies that are characteristic
of mature C fibers (Fitzgerald, M., 1987; Keller, A. F. et al., 2004). This is consistent with early expression of TRPV1,
critical for the detection of painful thermal and chemical stimuli, and the ATP-gated P2X3 receptor (Guo, A. et al., 2001)
that plays an important role in thermal and mechanical hyperalgesia following inflammation or nerve injury and the
tetrodotoxin-resistant sodium channel Nav1.8, necessary for the normal detection of noxious mechanical and thermal
stimuli and the sensitization of nociceptive sensory neurons (Benn, S. C. et al., 2001).
Nociceptive cells require activation of trkA receptors by nerve growth factor (NGF) for survival during the embryonic period,
but during the postnatal period, a subset of these neurons downregulate trkA and become dependent on glial-cell-line-derived
neurotrophic factor (GDNF) via the expression of the GDNF receptor Ret (Bennett, D. L. et al., 1996; Molliver, D. C. et al., 1997).
Thus the clear distinction between (1) the trkAve, neuropeptide containing C fibers and (2) the IB4ve, nonpeptidergic
nociceptors appears postnatally, although the functional implications of this in terms of nociception is not clear.
 The Development of Nociceptive Systems 135

11.2 The Development of Spinal horn neurons, fewer of which are nociceptive in the
Nociceptive Circuits newborn cord, especially of the wide dynamic range
(WDR) variety. Receptive field areas of dorsal horn
At birth, strong spinal nociceptive reflexes can be cells, mapped by natural mechanical stimulation of
evoked by mechanical stimulation, which are exagger- skin, are large at birth and decrease in size with age.
ated compared to the adult for the first 23 postnatal Pinching or brushing the receptive field of many
weeks (Ekholm, J., 1967; Fitzgerald, M. et al., 1988) with neonatal dorsal horn cells results in long-lasting
lower thresholds and larger, more synchronized and after-discharges (3090 s) which on days 03 may be
prolonged muscle contractions. A prick on the hind more pronounced than the initial evoked response but
foot in the newborn rat can evoke limb withdrawal, which decrease in duration and amplitude of these
wriggling, rolling, and simultaneous responses from all responses with age (Fitzgerald, M. and Jennings, E.,
four limbs. Receptive fields of hindlimb flexor muscles 1999; Torsney, C. and Fitzgerald, M., 2002). This
are large and disorganized such that the limb with- predominantly A-fiber-induced spike activity appear
drawal can be evoked that is not always appropriate to to have a critical role in the formation of nociceptive
the stimulus (Holmberg, H. and Schouenborg, J., 1996). circuits. Newborn rat pups have a high error rate in
As the pup matures, the response decreases in magni- the direction of a tail flick on noxious stimulation than
tude and becomes restricted to an isolated leg or foot adults, which gradually improves over 3 weeks
movement. Thresholds for withdrawal from heat sti- (Waldenstrom, A. et al., 2003). The maturation is
muli are also lower in younger animals. In contrast with unaffected by daily noxious stimulation but blocking
mechanical reflexes, but consistent with the slow synap- low-intensity tactile inputs from the tail with local
tic maturation of C-fiber-evoked responses, cutaneous anesthetic, during the critical 10-day period, prevents
application of the selective C-fiber irritant, mustard oil, this postnatal tuning. Low-intensity tactile input
evokes little or no spinal reflex response until postnatal from, for example, spontaneous twitching during
day 10 (Baccei, M. L. and Fitzgerald, M., 2005). sleep could play a role in shaping nociceptive circuits
The excitability of mechanical neonatal cutaneous in early life (Petersson, P. et al., 2003).
reflexes is attributable to underlying changes in dorsal Postnatal maturation and tuning of nociceptive
horn sensory processing and is likely to be due to circuits critically depends upon the postnatal
absence of fine-tuning of sensory and nociceptive development of excitatory and inhibitory neuro-
neuronal circuitry in the newborn. The earlier transmitter/receptor function in the neonatal dorsal
maturation and widespread presence of functional horn. The developmental pharmacology of spinal
A-fiber terminals in both the substantia gelatinosa pain pathways has been an area of intense recent
and deeper laminae in the first weeks of life appear interest (Pattinson, D. and Fitzgerald, M., 2004;
to adequately compensate for the weak C-fiber input Fitzgerald, M., 2005) and some important features
and dominate the physiological responses of dorsal are shown in Box 2. The properties of immature

Box 2 The development of excitatory and inhibitory transmission in developing spinal pain circuits
Glutamate AMPA and NMDA receptors are highly expressed in the newborn dorsal horn and their expression decreases
and becomes spatially organized with age. This is accompanied by changes in subunit composition leading to greater
AMPA-dependent Ca2 influx, altered receptor kinetics, greater NMDA receptor affinity, and increased Mg2 sensitivity in
neonatal spinal neurons (Watanabe, M. et al., 1994; Jakowec, M. W. et al., 1995; Albuquerque, C. et al., 1999; Bardoni, R.,
2001; Green, G. M. and Gibb, A. J., 2001; Stegenga, S. L. and Kalb, R. G., 2001). Presynaptic Ca2-permeable KA receptors
found on IB4ve C cells switch to a Ca2-impermeable form early in the first postnatal week (Lee, C. J. et al., 2001).
Despite early expression of GABA and glycine and their receptors, newborn lamina II neurons exhibit only GABA mIPSCs
(Baccei, M. L. and Fitzgerald, M., 2004) probably due to low levels of gephyrin, the postsynaptic scaffolding protein (Bremner
et al., 2005). There is also a fourfold acceleration in the decay rate of GABA mIPSCs in laminae III neurons between P8 and P23
triggered by local neurosteroid release (Keller, A. F. et al., 2004). GABAAR activation in the developing dorsal horn can be
excitatory rather than inhibitory, due to the high intracellular Cl concentration in immature neurons producing a positive ECl.
GABAAR activation depolarizes some lamina II cells but the response becomes exclusively hyperpolarizing by P6P7 (Baccei, M.
L. and Fitzgerald, M., 2004). This is probably due to the onset of expression of the KCl cotransporter KCC2 which decreases
[Cl]i over the postnatal period. However, ECl is always more negative than action potential threshold at P0P1 suggesting that
GABA will still inhibit the firing of newborn superficial dorsal horn neurons (albeit less effectively than in the adult).
Both presynaptic and postsynaptic GABABRs are functional from birth in the neonatal dorsal horn, but seem to be regulated
differently over the early postnatal period (Baccei, M. L. and Fitzgerald, M., 2004).
136 The Development of Nociceptive Systems

synaptic transmission tends to enhance calcium influx somatosensory (whisker) pathways in rodents and
into developing dorsal horn neurons compared to their relevance to nociceptive processing can only
adult neurons, which may be important for a variety be inferred. Recently, however, a study by Man S.
of developmental processes including synapse forma- H. et al. (2005) has shown the presence at birth of the
tion and remodeling. NK1ve lamina 1 ascending projection from the DH
to the parabrachial area of the brainstem using retro-
grade tracing and c-fos immunostaining. The
11.3 The Development of Ascending outgrowth of axons from the thalamus to the cortex
Pain Pathways begins at E13E14 in rodents just before the arrival of
the first cells migrating into the preplate zone. The
Following integration in the dorsal horn, sensory and migration of cortical neurons from the proliferative
nociceptive information is conducted to supraspinal neuroepithelium lining the telencephalic ventricle to
centers via ascending tracts to targets in the brainstem, the cortical plate is highly complex and tightly regu-
midbrain, hypothalamus, thalamus, and amygdala. A lated (Meyer, G., 2001). Thalamic axons and preplate
key ascending pathway involved in pain and hyperal- fibers meet in the lateral part of the internal capsule
gesia arises from lamina I of the dorsal horn of the at E15 and grow in association with early corticofugal
spinal cord, particularly neurokinin 1 (NK1) expressing axons, right up to the cortical subplate. The topogra-
neurons receiving A- and C-fiber nociceptive input, phy of thalamic axons is maintained throughout the
which project to the parabrachial area of the brainstem. pathway and they reach the cortex by associating
These neurons then project to the amygdala (affective with the projections of a number of preexisting
components of pain) and the hypothalamus (autonomic cells, including the preplate scaffold. These cells
functions such as changes in heart rate and blood invade the cortical plate 2448 h later and form func-
pressure). Outputs from these structures are passed tional synaptic connections in the cortical plate
via the periaqueductal gray (PAG), which in turn around E18E19. At birth, thalamic axons have
sends descending projections back to the dorsal horn extended into cortical layers IV, V, and VI in a
in a spinobulbar spinal loop. Descending projections to topographically precise manner and exhibit a periph-
the dorsal horn arise from the brainstem nuclei, nucleus ery-related pattern in layer IV (Schlaggar, B. L. and
raphe magnus (NRM), raphe dorsalis, reticularis, para- OLeary, D. D., 1994; Molnar, Z. et al., 2003). The
gigantocellularis (NRPG), as well as locus coeruleus, spatial distribution of thalamic synapses onto cortical
PAG, and mesencephalic reticular formation (Todd, A. neurons (predominantly found at dendritic spines) is
J. et al., 2000; Mantyh, P. W. and Hunt, S. P., 2004; established at least as early as P11 in the somatosen-
Suzuki, R. et al., 2004; Ding, Y. Q. et al., 1995; Marshall, sory cortex of the mouse (Lev, D. L. et al., 2002).
G. E. et al., 1996).
Little is known about the development of ascend-
ing spinothalamic and spinoparabrachial tracts. In the
rat, neurogenesis of dorsal horn projection neurons is 11.4 Pain and the Developing Cortex
complete by E16 having followed a systematic pat-
tern of migration and settlement in the dorsal horn The development of specific cortical areas is depen-
related to their axon trajectories (Nissen, U. V. et al., dent upon both extrinsic influences from
2005). The projecting axons grow up the spinal cord thalamocortical input and intrinsic genetic controls
prenatally (Bice, T. N. and Beal, J. A., 1997); spi- such as the transcription factors Emx2 and Pax6
nothalamic tract fibers reach the thalamus just (OLeary, D. D. and Nakagawa, Y., 2002). This pro-
before birth at E19. In the rat, afferents appear to cess extends over several months in man compared to
reach the thalamus by E19 (Higashi, S. et al., 2002) the mouse cortex that requires a week (Angevine, J. B.
while in the sheep, projection fibers from dorsal horn and Sidman, R. L., 1961). In man, the generation and
cells are in the thalamus approximately two-thirds migration of cortical neurons begins in the fifth gesta-
through gestation (Rees, S. et al., 1994; Ding, Y. Q. tional week (ORahilly, R. and Muller, F., 1994) and
et al., 1995). The thalamocortical axon (TCA) projec- continues until week 22 (Sidman, R. L. and Rakic, P.,
tion originates in dorsal thalamus, conveys sensory 1973). Following the arrival of neurons at their cor-
input to the neocortex, and has a critical role in rect destination, they begin to differentiate and to
cortical development. Information about the devel- extend dendritic fields. It is in the last 2 months of
opment of these pathways is entirely from visual and gestation that the characteristic folding of the cortex
 The Development of Nociceptive Systems 137

into sulci and gyri takes place (Fees-Higgins, A. and Gallagher, M. and Schoenbaum, G., 1999; Maren,
Larroche, J. C., 1987). S., 1999; Ledoux, J. E., 2000b; Cardinal, R. N. et al.,
One area of interest is the anterior cingulate cor- 2002; Davidson, R. J., 2002; Zald, D. H., 2003). In the
tex (ACC) that appears to play an important role in rat, the amygdala is a complex of 13 nuclei and
the affective and cognitive aspects of pain, but little is cortical regions and their subdivisions. There is
known of the development of this region. Ultrasonic some debate as to whether the amygdala should be
scans taken from preterm neonates have confirmed considered a single entity or as a collection of func-
that at week 28 the ACC is fully developed tionally diverse nuclei that share some anatomical
(Huang, C. C., 1991). and physiological similarities (Aggleton, J. P. and
Subplate neurons (SPn) play an important role in Saunders, R. C., 2000). There are both direct and
the formation of thalamocortical connections during indirect nociceptive inputs to the laterocapsular
early development and show glutamatergic and part of the central nucleus of the amygdala (the
GABAergic spontaneous synaptic activity. Whole- nociceptive amygdala) and the whole central area
cell recordings from SPn in somatosensory corti- supplies the major output forming widespread con-
cal slices of postnatal day 03 rats, showed that nections with forebrain areas, the hypothalamus and
SPn receive distinct functional synaptic inputs the brainstem (Pitkanen, A. et al., 1998; Davis, M.,
arising from the thalamus, cortical plate, and other 1998b; Aggleton, J. P. and Saunders, R. C., 2000;
SPn mediated by AMPA and NMDA receptors Ledoux, J. E., 2000a; Bourgeais, S. et al., 2001;
(Hanganu, I. L. et al., 2002; Luhmann, H. J. et al., Neugebauer, V. and Li, W., 2002; 2003; Li, W. and
2003). GABA activity is depolarizing at this age (see Neugebauer, V., 2004).
Box 2). Electrophysiological analysis of cortical cells Neurons in the amygdala of the rhesus monkey are
at P7 shows them to be organized in columns as in the generated during a 3-week period very early
adult but to have larger receptive fields, similar to in gestation (E30E50 of a 165-day gestation;
that discussed above for the spinal cord (Armstrong- Kordower, J. H. et al., 1992). This neurogenesis appears
James, M. A., 1970; Armstrong-James, M., 1975). to occur simultaneously within all the amygdalal
Human somatosensory potentials evoked by electri- nuclei. This pattern has also been reported in humans
cal stimulation are observed at 29 weeks (Klimach, (Humphrey, T., 1968; Nikolic, I. and Kostovic, I.,
V. J. and Cooke, R. W., 1988) while preliminary 1986). In the rodent, LOM-homeodomain genes have
studies using infrared spectroscopy during routine been shown to play a role in the segregation of
clinical heel lances suggest that a cortical blood embryonic amygdalal cells into appropriate groupings
flow response is present as early as 25 weeks (Slater, and this family of genes plays a fundamental role in
R. et al., 2006). the development of every single amygdalal nucleus
While axonal growth and early topographic (Remedios, R. et al., 2004). The distribution of opiate
arrangement of thalamocortical fiber pathways do receptors in the amygdala of a 1-week-old monkey
not rely on activity-dependent mechanisms requiring appears the same as in the adult indicating early
evoked neurotransmitter release (Molnar, Z. et al., neurochemical maturation (Bachevalier, J. et al.,
2002), the formation of cortical circuits is highly 1986). Widespread connections to and from the mon-
dependent upon normal sensory experience. The key amygdala at 1 week of age appear to be pruned
postnatal plasticity of cortical circuits has been stu- over the postnatal period (Webster, M. J. et al., 1991).
died in great detail for the visual (Yao, H. and Dan, The same connections are likely to occur over the first
Y., 2005) and somatosensory whisker barrel cortex 2 postnatal weeks in rats.
(Fox, K., 2002), but little is known about the effect
of nociceptive activity upon cortical development
(Anand, K. J., 2000). 11.6 The Development of
Descending Pain Pathways

11.5 Pain and the Developing Limbic Brainstem descending pathways form a major
System mechanism in the control of pain transmission
(Pitkanen, A. et al., 1998; Ren, K. and Dubner, R.,
The amygdala is a part of the limbic system and plays 2002; Gebhart, G. F., 2004) and there is more
a key role in the emotional evaluation of sensory known about the development of these descending
stimuli (Davis, M., 1998a; Aggleton, J. P., 1999; controls compared to the ascending systems.
138 The Development of Nociceptive Systems

Brainstem nuclei differentiate between E11 and E16 of the spinal cord at various postnatal stages suggests
in the rat and all other major brain systems are that descending axon collaterals innervate the inter-
morphologically identifiable by E18 (Altman, J. and mediate and central gray from birth but that they are
Bayer, S. A., 1984; Momose-Sato, Y. et al., 2001). In not found in the dorsal horn until P15 (Gilbert, M.
the rat embryo, the parabrachial area can be visua- and Stelzner, D. J., 1979).
lized histochemically from at least E17 (Cholley, B. Descending inhibitory pathways traveling from
et al., 1989) (Figure 1 ). the brainstem via the dorsolateral funiculus of the
As with ascending tracts, axons from brainstem spinal cord to the dorsal horn grow down the spinal
nuclei appear to grow down the spinal cord well cord early in fetal life, but they do not extend collat-
before birth (Cabana, T. and Martin, G. F., 1984). eral branches into the dorsal horn for some time.
Injections of horseradish peroxidase into lumbosacral Stimulation of the dorsolateral funiculi (DLF) fails
spinal cord of the neonatal rat label brainstem nuclei to produce descending inhibition of C-fiber-evoked
with similar density to that seen in the adult (Leong, activity in neonatal rats before P9, but high-intensity
S. K., 1983; Fitzgerald, M. et al., 1987) but it is unclear stimulation produced descending inhibition at P18.
at what stage these axons begin producing collaterals By P22P24 more typical adult responses were
that innervate the dorsal horn and make synapses observed (Fitzgerald, M. and Koltzenburg, M.,
with their target cells. Mapping degenerating axons 1986). Similarly, the inhibition of C and A activity
and synaptic endings following thoracic hemisections in the dorsal horn by PAG stimulation via the DLF
that is clearly observed in adults (Basbaum, A. I. and
Fields, H. L., 1978) cannot be produced in rat pups
cc Cortical before P21 (van Praag, H. and Frenk, H., 1991).
?
Hip projections
Amygdala Po and connections Diffuse noxious inhibitory control (DNIC), mea-
connections
Ic
VPM/VPL
beginning sured as a reduction in noxious-evoked dorsal horn
unknown VMH c-fos expression in the presence of an inflammatory
Ce

Spinothalamic stimulus elsewhere on the body, also develops

connections between P12 and P21 (Boucher, T. et al., 1998).
in place The delayed postnatal onset of functional descend-
Spinopara- ing inhibition, despite the presence of DLF terminals
brachial
connections PB LC
in the dorsal horn may be due to immaturity of neu-
bc
in place rotransmitter/receptor interactions (see below) or
delayed maturation of critical interneurons. It has
C fibers been suggested that the maturation of descending
Brainstem-
descending in place; inhibition is dependent upon afferent C-fiber activity,
connections synaptic because rats treated with capsaicin at birth have
in place; V maturation
A5
RVM
underway
reduced inhibitory controls as adults (Cervero, F.
synaptic Py and Plenderleith, M. B., 1985). The lack of descending
maturation
beginning inhibition in the neonatal dorsal horn means that there
is no endogenous analgesic system to dampen noxious
A fibers inputs as they enter the CNS and their effects may
in place; therefore be more profound than in the adult. It also
pruning
underway explains why stimulus-produced analgesia from the
PAG is not effective until P21 in rats (van Praag, H.
Figure 1 Diagram summarizing the state of and Frenk, H., 1991).
development of ascending and descending pathways
involved in the transmission of pain in the newborn rat. It
is important to remember that changes in the anatomy of
the pathways depicted here are also accompanied by 11.7 Development of
changes in the physiology and neurochemistry of these Neurotransmitter/Receptor Function
pathways also. Additionally, the maturation of in Descending Pathways
human nociceptive pathways occurs over different
and as yet unknown time courses from those discussed
here. Adapted from Hunt, S. and Mantyh, P. 2001. The It is widely accepted that 5-HT is the most relevant
molecular dynamics of pain control. Nat. Rev. Neurosci. neurotransmitter with respect to the descending
2, 8391. control of spinal cord excitability. Descending
 The Development of Nociceptive Systems 139

serotonergic pathways predominantly mediate des- postnatally. As with 5-HT, the development of the
cending inhibitory control over spinal cord noradrenergic system in the ventral horn precedes
excitability, but 5-HT is involved in both antinoci- that in the dorsal horn. Expression of the 2A receptor
ceptive and pronociceptive effects depending upon mRNA is observed in high levels in the dorsal
the subtype of receptor that it is acting upon (Millan, horn from E19 and is downregulated postnatally
M. J., 2002; Gebhart, G. F., 2004). An excitatory, (Winzer-Serhan, U. H. et al., 1997), which has led to
pronociceptive role has been attributed to the the proposal that noradrenaline has a trophic role in
5-HT3 receptor (Suzuki, R. et al., 2002). In the the developing cord (Huang, Y. et al., 2002). Recent
adult, descending inhibition and excitation are con- functional studies have shown that the local
veyed rostrocaudally via different spinal pathways, spinal cord application dexmedetomidine, the potent
the DLF and ventral/ventrolateral funiculi (VLF), 2A agonist, has antinociceptive and antihyperalgesic
respectively (Millan, M. J., 2002), and since both of actions from at least P3 (Walker, S. 2005).
these tracts appear in the embryonic period, Dopamine is also released by descending tracts; the
both types of control may also be possible in the main source is from the A11 (posterior) region of the
newborn. Neurons that produce 5-HT (serotonin) hypothalamus (Commissiong, J. W. et al., 1978;
are amongst the earliest to be detected in the devel- Commissiong, J. W. and Neff, N. H., 1979; Swanson,
oping CNS (Rubenstein, J. L., 1998) with the L. W. and Kuypers, H. G., 1980), which provide
mesencephalon containing detectable levels by E12 extensive networks of dopamine-containing fibers in
(E12; Aitken, A. R. and Tork, I., 1988). 5-HT contain- the dorsal horn and lamina X of the spinal cord
ing descending neurons is present in the rat between (Skagerberg, G. et al., 1982; 1988). In the rat, dopamine
E15 and E19 (Lidov, H. G. and Molliver, M. E., 1982; containing neurons appear in the mesencephalon at
Lakke, E. A., 1997), those arising from the raphe nuclei E12E13 (Olson, L. and Seiger, A., 1972) and levels of
are present in the cervical cord by E14 and in the dopmaine appear to increase throughout development
lumbar cord by E16E17. 5-HT fibers were not found with particularly high levels being found in the soli-
to innervate the dorsal horn until E19 (Rajaofetra, N. tary tract in the brainstem (Moore, R. Y. and Bloom, F.
et al., 1989). Examination of the pre- and postnatal E., 1978).
development of the descending serotonergic pathways Numerous other nonmonoaminergic transmitters
to the spinal cord show that the maturation of 5-HT are found in the terminals of descending pathways in
terminals occurs in a ventral to dorsal direction such the spinal cord including histamine, vasopressin, and
that innervation of the lumbar dorsal horn does not oxytocin. Other transmitters released by descending
occur until P14 and the adult pattern of high-density projections are also present in intrinsic DH neurons.
terminals in lamina II at P21 (Bregman, B. S., 1987). Of these, acetylcholine is the most prominent, having
Direct application of 5-HT to the neonatal spinal cord both a pro- and antinociceptive effect depending on
in vitro is effective in very young superficial dorsal the subtype of the receptor activated. Other neuro-
horn cells, suggesting that at least some 5-HT recep- transmitters in the category are GABA and glycine.
tors are functional from birth (Li, P. and Zhuo, M., Cholinergic neurons in the brainstem generally
1998). develop later than catecholaminergic systems with
Although 5-HT is the most important neurotrans- levels of cholinergic markers such as choline acetyl-
mitter, other monoaminergic and peptidergic transferase being only a few percent of the levels seen
compounds must also be considered with regard to in the adult rat at birth but start to increase rapidly
the descending control of spinal cord excitability. after P10 (Diebler, M. F. et al., 1979).
The major noradrenergic input to the dorsal horn is A major peptidergic descending input from the
from the locus coeruleus (LC) and adjacent noradre- brainstem to the dorsal horn of the spinal cord is
nergic nuclei in the brainstem. Activation of this via enkephalin and other opioid peptides (pre-
pathway leads to the release of noradrenaline that proenkephalin, pre-prodynorphin, pre-proendomor-
acts postsynaptically on 2-adrenergic receptors to phin, pre-pro-opiomelanocortin (POMC), and
produce a decrease in excitability and a concomitant orphaninFQ). Opioids play an important role in
analgesia (Sagen, J. and Proudfit, H. K., 1984; Jones, pain transmission both by initiating the descending
B. E. 1991; Jasmin, L. et al., 2002). Noradrenaline con- controls from the brainstem and by directly reducing
taining terminals appear earlier in the dorsal horn than the evoked activity of C and A fibers in the dorsal
serotonin (Loizou, L. A., 1972; Commissiong, J. W., horn via presynaptic opiate receptors. There is
1983) and levels are generally mature by 2 weeks considerable postnatal development of this receptor
140 The Development of Nociceptive Systems

expression, with neonatal rats not only having a 2001). This is interesting in view of the substantial cell
higher density but also a less selective distribution death that occurs in the DRG of axons damaged in
of -opioid receptors on DRG cells (Nandi, R. et al., early life compared to adults (Yip, H. K. et al., 1984;
2004). Morphine has a markedly higher analgesic Benn, S. C. et al., 2002) followed by transganglionic
efficacy in young rats in mechanical but not thermal degeneration and subsequent withdrawal of central
noxious tests but it is not known whether this results terminals (Bondok, A. A. and Sansone, F. M., 1984;
from central or peripheral regulation of opioid path- Aldskogius, H. and Risling, M., 1989), sprouting of
ways (Nandi, R. and Fitzgerald, M., 2005). adjacent, intact axon collaterals into the denervated
region (Fitzgerald, M., 1985; Fitzgerald, M. et al.,
1990; Shortland, P. and Fitzgerald, M., 1994) and dis-
ruption of the somatotopic organization of central
11.8 The Development of Persistent terminals within the dorsal horn and cortex (Kaas, J.
Pain H. et al., 1983). Clearly these changes do not trigger
pain. One possibility is that the central microglial
While it is essential to understand the underlying and immune responses that follow nerve injury in the
development of nociception, tissue injury, inflamma- adult and may contribute to neuropathic pain
tion, and nerve damage can lead to more persistent (Marchand, F. et al., 2005) are not activated in the
pain that lasts for days, weeks, or months. Such pains immature spinal cord.
arise from central sensitization and changes in gene There is also evidence that injury at a critical period
expression as triggered by intense and sensitized of development may lead to long-term alterations in
peripheral nociceptor inputs (Woolf, C. J. and sensory processing resulting in hypo- or hyperalgesia
Salter, M. W., 2000). (Lidow, M. S., 2002). The mechanisms underlying this
Cutaneous mechanical thresholds fall in neonatal rat are poorly understood. It is possible that early tissue
pups following local carrageenan injection (Marsh, D. damage, which is known to release excess neurotro-
et al., 1999) and mustard oil application (Jiang, M. C. phins into the wounded area (Constantinou, J. et al.,
and Gebhart, G. F., 1998) but the effect is less at P3 than 1994) not only sensitizes peripheral nociceptors, but
at P21. This may partly reflect peripheral nociceptor also permanently influences nociceptor development
terminal immaturity. C fibers are capable of peripheral as described above. Alternatively central changes in
sensitization from before birth (Koltzenburg M., and activity-dependent connectivity are produced because
Lewin, G. R., 1997) but peripheral C-fiber terminals do of the increased barrage of C-fiber nociceptor input at a
not release sufficient SP to produce neurogenic extra- time when synapses are still forming (Fitzgerald, M.
vasation until P10 (Fitzgerald, M., and Gibson, S., and Walker, S. M., 2003). Central changes in transmit-
1984). Electrophysiological studies in vivo show that terreceptor expression may also occur. Early
sensitization of dorsal horn cells can be observed inflammatory pain leads to an increase in expression
25 h after carageenan inflammation of the hindpaw in the PAG of serotonin receptors (5HT1A, 5HT1D,
at all ages although the exact pattern of effects is 5HT2A, 5HT2C, 5HT4), NR2D and NR3A subunits
age dependent (Fitzgerald, M. and Jennings, E., 1999; of the NMDA receptor, GABAA2 and GABAAy sub-
Torsney, C. and Fitzgerald, M., 2003). In slice prepara- units, the NK1, and the CCK-2 receptor. As well
tions, NMDA-dependent C-fiber-evoked depolariza- as being hypoalgesic, these animals exhibited decreased
tion of spinal cord cells and windup of cells to repeated anxiety and greater stress-coping ability; this was
C-fiber stimulation has been demonstrated in the indexed by better performance on the elevated
young (814 days) spinal cord (Sivilotti, L. G, et al., plus maze and decreased basal and stress-induced neu-
1993). Recently, the role of descending brainstem roendocrine markers of anxiety and stress (Anseloni, V.
excitatory pathways in maintaining persistent pain C. et al., 2005). For further review of this area, see
have been emphasized (Gebhart, G. F., 2004), but the Baccei M. L. and Fitzgerald M. (2005).
developmental regulation of this system has not been
investigated.
In adults, partial nerve injury can trigger neuro- 11.9 Concluding Remarks
pathic pain, charcterized by a lowered mechanical
sensitivity or allodynia. In postnatal animal models of In this chapter we have attempted to briefly review
neuropathic pain, this allodynia is not observed until what is currently known about the development and
the animals are over 3 weeks old (Howard, R. F. et al., maturation of pain pathways. While our knowledge of
 The Development of Nociceptive Systems 141

Box 3 Development of pain pathways in humans

Because human infants cannot report pain, we have to use indirect physiological and behavioral methods to assess its
existence and severity and there is considerable debate on how best to measure pain in infants (Stevens, B. J. and Franck, L.
S., 2001).
From 23 weeks of gestational age, human fetuses mount a hormonal response when needles are inserted into the
innervated intrahepatic vein (Giannakoulopoulos, X. et al., 1994). Noxious activation of nerve endings stimulates central
pathways (most likely through the fetal hypothalamicpituitaryadrenal axis), producing a biochemical stress response,
although the link between perceived pain and the hormonal stress response is unpredictable, even in adults. Hormonal
responses are also used to measure pain in the youngest preterm infants (2426 weeks), along with many other physiological
measures such as heart rate variability, crying and palmar sweating and motor activity (Stevens, B. J. and Franck, L. S., 2001),
and characteristic facial expressions (Craig, K. D. et al., 2002). Many of these measures can be applied to pain related to a
particular noxious event, such as a needle prick, but are harder to use for persistent pain. Studies using cutaneous reflexes
evoked by stimulation in and around an area of injury, show that infants display cutaneous sensitization or hyperalgesia lasting
days and weeks (Andrews, K. and Fitzgerald, M., 2002; Andrews, K. et al., 2002), consistent with laboratory studies. Nothing is
known of the maturation of descending brainstem controls in human infants.
True pain experience requires functional maturation of higher brain centers and recent studies using infrared spectroscopy
during routine clinical heel lances suggest that a cortical blood flow response is present as early as 25 weeks (Slater, R. et al.,
2006).

the development of spinal nociceptive pathways has Aitken, A. R. and Tork, I. 1988. Early development of serotonin-
containing neurons and pathways as seen in wholemount
increased considerably in recent years (Fitzgerald, M., preparations of the fetal rat brain. J. Comp. Neurol.
2005), the understanding of the structural and func- 274, 3247.
tional maturation of supraspinal centers involved in Albuquerque, C., Lee, C. J., Jackson, A. C., and
MacDermott, A. B. 1999. Subpopulations of GABAergic and
the affective and nociceptive components of pain is non-GABAergic rat dorsal horn neurons express Ca2-
severely lacking. With rapid improvements in medical permeable AMPA receptors. Eur. J. Neurosci.
knowledge and technologies, it is now possible to treat 11, 27582766.
Aldskogius, H. and Risling, M. 1989. Number of dorsal root
children born at increasing premature stages of devel- ganglion neurons and axons in cats of different ages. Exp.
opment. This requires long periods of hospitalization Neurol. 106, 7073.
and large numbers of often invasive procedures that in Altman, J. and Bayer, S. A. 1984. The development of the rat
spinal cord. Adv. Anat. Embryol. Cell Biol. 85, 1164.
an adult or older child would be considered painful Anand, K. J. 2000. Pain, plasticity, and premature birth: a
(Box 3), leading to the administration of analgesic and prescription for permanent suffering? Nat. Med. 6, 971973.
sedative compounds. We know that neonates respond Andrews, K. and Fitzgerald, M. 2002. Wound sensitivity as a
measure of analgesic effects following surgery in human
to pain but the pathways differ in important ways from neonates and infants. Pain 99, 185195.
those in adults. Until we understand more of the Andrews, K. A., Desai, D., Dhillon, H. K., Wilcox, D. T., and
functional development and maturation of the central Fitzgerald, M. 2002. Abdominal sensitivity in the first year of
life: comparison of infants with and without prenatally
neuronal elements involved in the experience of pain, diagnosed unilateral hydronephrosis. Pain 100, 3546.
the best course of analgesic action in infancy remains Angevine, J. B. and Sidman, R. L. 1961. Autoradiogrpahic study
unclear. of cell migration suring histogenesis of cerebral cortex in the
mouse. Nature 192, 766768.
Anseloni, V. C., He, F., Novikova, S. I., Turnbach, R. M.,
Lidow, I. A., Ennis, M., and Lidow, M. S. 2005. Alterations in
stress-associated behaviors and neurochemical markers in
Acknowledgments adult rats after neonatal short-lasting local inflammatory
insult. Neuroscience 131, 635645.
Armstrong-James, M. A. 1970. Spontaneous and evoked single
The authors would like to thank The Wellcome
unit activity in 7-day rat cerebral cortex. J. Physiol.
Trust, The Wellcome London Pain Consortium, 208, 10P11P.
and Professor S. P. Hunt (UCL). Armstrong-James, M. 1975. The functional status and columnar
organization of single cells responding to cutaneous
stimulation in neonatal rat somatosensory cortex S1.
J. Physiol. 246, 501538.
Baccei, M. L. and Fitzgerald, M. 2004. Development of
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 12 Appropriate/Inappropriate Developed Pain Paths
J Schouenborg, Lund University, Lund, Sweden
2009 Elsevier Inc. All rights reserved.

12.1 General Background 147

12.2 Modular Organization of Sensorimotor Circuits in the Spinal Cord 147
12.3 Action-Based Sensory Encoding 149
12.4 Functional Adaptation of Sensorimotor Circuits During Development 149
12.5 Concluding Remarks 152
References 152

12.1 General Background (nociception, pressure, temperature, joint angles,

muscle force, and length) arises from a complex
The topographical organization of the somatosensory body constitution. Understanding how sensory infor-
input to the spinal cord has traditionally been mation is processed by spinal sensorimotor circuits
regarded as a somatotopically organized map of the and how the adult organization emerges during
body (Molander, C. and Grant, G., 1985; Mirnics, K. development is therefore a major task. This review
and Koerber, H. R., 1995; Silos-Santiago, I. et al., 1995; will deal with the current literature on the modular
Wilson, P. and Kitchener, P. D., 1996; Brown, P. B. organization of the spinal cord and the role of soma-
et al., 1997; 2004; 2005), with different body parts tosensory imprinting in establishing an action-based
represented in different central nervous system sensory encoding in these modules.
(CNS) areas and the size of the representation
being a function of receptor density and, to some
extent, practise (see Blake, D. T., et al., 2002). Most 12.2 Modular Organization of
studies on the body representation have been focused Sensorimotor Circuits in the Spinal
on the sensory discriminative aspects (Brown, P. B. Cord
et al., 2004; 2005). However, a major function of the
spinal cord is to use the sensory information to con- During the last 10 years, the concept of a modular
trol skeletal muscles. It has therefore been suggested organization of the spinal cord has grown stronger.
that the somatosensory input may be primarily orga- The idea of a modular organization of motor circuits
nized in a motor frame of reference (Levinsson, A. in CNS is not new and, for the spinal cord, was
et al., 2002). Such a view might explain some of the proposed already in 1981 by Grillner, S. for locomo-
oddities with the known topography. For example, in tor circuits (unit bursters causing rhythmic activity
the dorsal horn, there is an extensive overlap of around a joint) and later for circuits controlling
afferent terminals from different body parts, and the scratch reflexes in the turtle (Mortin, L. I. et al.,
sequence of the representation from distal to prox- 1985; Stein, P. S. et al., 1998; Berkowitz, A., 2001).
imal body parts is neither continuous nor sequential However, although a modular organization is con-
(Levinsson, A. et al., 2002). Moreover, the phenom- ceivable, it is not yet clear exactly what constitutes a
enon of somatotopically inappropriate terminations, module in these rhythm-generating systems, the
that is, primary afferents terminating outside the extent to which the different modules overlap in
main projection area, might be explained if different space, and how sensory information is related to the
motor systems requiring sensory information from function of these modules (Tresch, M. C. et al., 2002).
same body region have different locations in the A modular type of reflex organization in the mam-
spinal cord. malian spinal cord was first demonstrated for the
To be useful in motor control, somatosensory nociceptive withdrawal reflex (NWR) system in the
information must be encoded with respect to body rat (Schouenborg, J. and Kalliomaki, J., 1990;
anatomy and movement patterns produced by the Schouenborg, J. et al., 1992), but subsequently also
sensorimotor circuits. This is a computationally in cats (Levinsson, A. et al., 1999), mice (Thelin,
demanding task since the multisensory information J. M. and Schouenborg, J., 2003), and humans

147
148 Appropriate/Inappropriate Developed Pain Paths

(Sonnenborg, F. A. et al., 2000). Here, the word mod- mediolateral sequence reminiscent of the corre-
ular is used synonymous to the term functional unit sponding topographical organization of the
of a system, that is, not alluding to the existence of motoneuron columns in the ventral horn. Hence,
different motor systems, such as stepping, standing, the reflex encoders appear to be located in discrete
scratching, or withdrawal reflex systems. For the pools that have a musculotopic organization.
NWR system, each excitatory module preferentially A corresponding set of inhibitory reflex modules
acts on a single muscle and performs a detailed sen- also exists. In this case, the receptive fields correspond
sorimotor transformation resulting in a graded to the graded movement of the skin area toward
withdrawal of the limb (or part of the limb) from its external stimulation (i.e., increase in load) on contrac-
receptive field. For each excitatory NWR module, tion of the muscle in the module (Weng, H.-R. and
the input strength has a characteristic pattern on the Schouenborg, J., 1996). As a result of this organization,
skin that mimics the pattern of withdrawal efficacy in the excitatory and inhibitory modules are engaged to a
a standing position when the principal output muscle degree that is proportional to their respective with-
of the module contracts (Figure 1; Schouenborg, J. drawal/loading efficacy on skin stimulation.
and Weng, H.-R., 1994). In a sense, the pattern of The same principles of sensorimotor transforma-
withdrawal (or unloading) efficacy is imprinted on tion as found for rat withdrawal reflexes also apply to
the receptive field of the module. Neurones that the cat, mice, and humans (Levinsson, A. et al., 1999 in
encode sensory input in this motor frame of refer- cats; Sonnenborg, F. A. et al., 2000; Thelin, J. M. and
ence, termed reflex encoders (Schouenborg, J. et al., Schouenborg, J., 2003). Interestingly, muscles with
1995), are mainly located in deep dorsal horn laminae different function in two different species exhibit
IVVI. These neurones receive and weight the mul- corresponding differences in the receptive fields of
tisensory input from both tactile A and nociceptive the withdrawal reflexes. Thus, withdrawal reflexes
A and C afferent fibers in proportion to the with- elicited in dorsiflexors are much weaker in the cat
drawal/unloading action of the modules in a standing than in the rat and vice versa for reflexes to plantar
position (Figure 2). It is worth noting that, although flexors. Cats stand on their digits, whereas rats stand
the nociceptive input is stronger than that from tac- on the whole plantar surface. In the standing position,
tile input, the strength of the input mediated by the dorsiflexors of cat digits are much less effective in
different types of afferent fibers is weighted in the withdrawing the plantar skin than the corresponding
same way. In fact, a large proportion of the wide muscles in rats and vice versa for plantar flexors.
dynamic range neurones, that is, neurones receiving These findings are consistent with the view that
a convergent input from tactile and nociceptive these reflexes consist of adaptive modules (see
receptors, often referred to as wide dynamic range Section 12.4).
neurons, in the deep layers of the fifth lumbar seg-
ment, appear to be of the reflex encoder type. Within
the L45 segments, reflex encoders for the interossei,
N
flexor digitorum longus, gastrocnemius, peronei, and
extensor digitorum longus muscles are located in a N Re M

N
Gastrocnemius Peroneus longus Tibialis anterior
Withdrawal
action

Receptive Figure 2 Schematic circuitry organization of a withdrawal

field reflex module. Nociceptive afferents from the receptive field
(right) synapse with nociceptive neurones (N) in the
Figure 1 Action-based sensory encoding. Typical superficial laminae of the dorsal horn. Reflex encoder
distributions of sensitivity and corresponding withdrawal neurones (Re) receive a weighted nociceptive input from
action for three muscles are shown. Low, medium, and high superficially located neurones. Re project either directly or
dot density indicates areas of the skin from which the evoked indirectly to motoneurones essentially controlling the action
responses were 030%, 3070%, and 70100% of maximal of a single muscle. Withdrawal efficacy of the module is
response, respectively. Adapted from Schouenborg, J. and shown to the right. Sensitivity in strength of primary afferent
Weng, H.-R. 1994. Sensorimotor transformation in a spinal input and sensitivity in receptive fields is coded by color
motor system. Exp. Brain. Res. 100, 170174. intensity. Tactile input is omitted for clarity.
 Appropriate/Inappropriate Developed Pain Paths 149

Based on microstimulation in the dorsal horn of fibers encode the sensory input in the same way as
the spinal cord, a somewhat different modular orga- individual spinal modules, that is, as a function of the
nization of sensorimotor circuits acting on synergistic unloading pattern caused by single muscle
muscle groups was later proposed in frogs and rats contraction.
(Tresch, M. C. and Bizzi, E., 1999; Bizzi, E. et al., 2000,
Saltiel, P. et al., 2001). According to these authors,
electrical and glutamatergic stimulation of the deep 12.4 Functional Adaptation of
dorsal horn often results in a movement toward an Sensorimotor Circuits During
equilibrium point, independent of the starting posi- Development
tion of the limb. Here, the output of a module would
include activity in two or more different muscles. Given that the adult sensorimotor transformations
Whether these findings reflect the existence of a performed by the spinal withdrawal reflex circuits
fundamentally different type of reflex system than reflect precisely weighted connections in modules,
that of the withdrawal reflex system or the outcome how can this weighting be achieved during develop-
of a combined stimulation of many withdrawal reflex ment? While the gross topographical organization of
modules is not clear at present. In a recent study, interneurons of the spinal cord is likely to be guided
Avella and colleagues suggested, by analyzing electro- by gradients of trophic substances during the devel-
myography (EMG) in frog hindlimbs during kicking, opment (Albright, T. D. et al., 2000, Chen, A. I. et al.,
that a combination of a limited number of synergistic 2006; Chen, C. L. et al., 2006; Kramer, I. et al., 2006), it
units was used for this behaviour (Avella, A. et al., is difficult to see how such mechanisms could encode
2003). the detailed strength of every connection in the net-
works. Recent studies on the NWR system provide
some clues to this problem. The sensorimotor trans-
12.3 Action-Based Sensory formations performed by its modules are functionally
Encoding adapted during the first postnatal weeks in the rat
(Holmberg, H. and Schouenborg, J., 1996a). During
By mapping tactile input to the dorsal horn, long- this time, the strength of the erroneous (or inap-
itudinal cigar-shaped zones that weight the propriate) connections becomes weaker, whereas
monosynaptic cutaneous input as a function of the the strength of the adequate connections becomes
unloading pattern caused by individual modules proportional to the unloading effect on the skin of
were found (Figure 3; Levinsson, A. et al., 2002). muscle contraction. Moreover, during this time,
This was a surprising finding in that it indicates NWR can adapt to both altered innervation of the
that sensory input to the spinal cord gets organized skin caused by nerve sections (Holmberg, H. and
and weighted in a motor frame of reference already at Schouenborg, J., 1996b) and to altered movement
the level of the first-order synapses and that there patterns caused by tendon transfer in the neonatal
may, consequently, not be a true map of the body in rat (Holmberg, H. et al., 1997). Blocking the sensory
the spinal cord as previously thought (Molander, C. input during the time of functional adaptation
and Grant, G., 1985; Mirnics, K. and Koerber, H. R., abolishes the learning (Waldenstrom, A. et al., 2003),
1995; Silos-Santiago, I. et al., 1995; Brown, P. B. et al., indicating a role for sensory input. Notably, a selec-
1997). Given that a main function of the spinal cord is tive block of the nociceptive input by EMLA salve
to translate sensory information into movement cor- did not have an effect as compared with vehicle
rections, an action-based sensory encoding do, treatment. The functional adaptation is not blocked
however, appear to be an economical and fast way unless tactile input is also blocked, indicating a key
of processing the sensory information. role for tactile input in tuning nociceptive connec-
There are many reasons to think that action-based tions (Figure 4(a)). This conclusion is strongly
sensory encoding is fundamental also for higher- supported by the finding that tactile feedback ensu-
order motor systems. For example, the C1, C3, and ing on spontaneous motility in spinal sensorimotor
X zones in the anterior cerebellar lobe is divided into circuits alters the nociceptive connection strengths in
microzones, where each microzone is defined by its NWR modules during postnatal development
climbing fibre input from a specific receptive field on (Figures 4(b)4(c)); Petersson, P. et al., 2003). Thus,
the skin (Garwicz, M., 2002; Garwicz, M. et al., 2002; the pattern of tactile inputs arriving in conjunction
Apps, R. and Garwicz, M., 2005). These climbing with the spontaneous movements has an instructive
150 Appropriate/Inappropriate Developed Pain Paths

I
II
III
IV

VI

VII X

IX VIII

Figure 3 Schematic figure of proposed modular organization of the spinal cord. Columns of the dorsal horn receive a
cutaneous input that has a specific weight distribution. This weight distribution is the same as that of nociceptive input to
reflex encoders (interneurones that can encode the withdrawal reflex strength of individual modules) in deep dorsal horn. The
reflex encoders are assumed to project to single muscles and weight the input according to the withdrawal efficacy of the
output muscle. Sensitivity in strength of primary afferent input and sensitivity in receptive fields is coded by color intensity.
Upper recordings, receptive fields of monosynaptic tactile field potentials; middle recordings, receptive fields of reflex
encoders, bottom recordings, receptive fields of single muscles. Schematic indication of Rexeds laminae to the left.

role in the functional adaptation of the reflex mod- solves the puzzle of how a pain-related system can be
ules. Uncorrelated input (given at random time learned during postnatal development, despite the rare
points) does not cause a learning effect. Since this occurrence of noxious events. Notably, this novel form
learning process results in an imprint of the with- of unsupervised learning occurs during active sleep,
drawal efficacy on the reflex modules, it was termed characterized by atonia in the musculature. This state
somatosensory imprinting (Holmberg, H. et al., 1997; may be particularly advantageous for learning since the
Petersson, P. et al., 2003). sensory feedback on muscle contraction stands out
The finding that the strength of the nociceptive from a more or less silent background.
connections to the NWR network can be functionally A hitherto neglected problem is how the myriad of
adapted by tactile input discloses a hitherto unknown different types of somatosensory primary afferents,
form of cross-modality learning in the spinal cord that conveying information from different sensory
 Appropriate/Inappropriate Developed Pain Paths 151

N
(a) (b) (c) **
T
Skin pressure 80
Twitch
Skin pressure
T

Error rate (%)

75
N
Aberrant Normal
4 3
W W 70
RE
65
1 2
60
M Before After Before After Before After
Aberrant Normal Uncorrelated

Figure 4 Somatosensory imprinting. (a) Proposed learning cycle underlying somatosensory imprinting. A learning cycle
consists of the following chain of events. (1) Spontaneous bursts in reflex encoders (REs). (2) Motoneuron (M) activation leading
to a muscle twitch that causes skin movement. (3) Increased or decreased skin pressure resulting in altered sensory input to pre-
reflex encoder interneurons. Black lines (T) represent afferents from skin areas from where an increase and decrease,
respectively, in low-threshold mechanoreceptor input would occur. Red lines (N) symbolize nociceptive afferents. (4) The
strengths of erroneous connections (receiving increased tactile input) between pre-RE interneurons and REs are weakened and
that of appropriate ones (receiving reduced tactile input) is strengthened (Dw change in connection strength) (modified from
Waldenstrom, A. et al., 2003). (b) Behavioural experiments. Schematic training set-up depicting aberrant and normal training
protocol. Distal tail of a sleeping rat pup is shown. Air puffs directed to the skin surface moving toward or away from the air
stream is termed aberrant or normal conditioning, respectively. One session of conditioning stimulation lasted for 2 h. Arrow
indicates direction of spontaneous tail flick (modified from Petersson, P. et al., 2003). (c) The nociceptive withdrawal reflex
(NWR) error rate (%) before and after aberrant or normal tactile stimulation (mean SE). The NWR error rate was significantly
increased (KruskalWallis and Dunnetts test P < 0.01) in rats given aberrant tactile feedback for 2 h (47 training sessions).
Normal air-puff stimulation (53 training sessions) had no effect. Control experiments, using random air-puff stimulation (23
training sessions) or no stimulation (23 training sessions) did not affect NWR adaptation (Wilcoxon signed rank test P 0.65 and
P 0.37, respectively), ruling out direct effects of the set-up, test stimulations, or of uncorrelated stimulation. (Adapted from
Petersson, P., Waldenstrom, A., Fahreaus, C., and Schouenborg, J. 2003. Spontaneous muscle twitches during sleep guide
spinal self-organization. Nature 424(6244), 7275.

modalities and locations, gets organized in the dorsal Importantly, these findings suggest that the adult
horn so that spinal multisensory modules receive topo- action-based sensory encoding emerges by sensorimo-
graphically ordered information. Considering the tor circuits probing the patterns of tactile input that
apparent lack of somatotopic organization in the dorsal ensue on movements caused by the effector muscles.
root ganglia and that different nerve fibre types enter Spontaneous movements are a ubiquitous phe-
the spinal cord by different routes and at different nomenon during embryonic development in all
times during development, this is not a trivial problem. vertebrates and mammals (De Vries, J. I. et al., 1982;
Indeed, recent data indicate that the adult body repre- Blumberg, M. S. and Lucas, D. E., 2000). However,
sentation in the dorsal horn arises from an initially while a role in maturation of motoneurones, moto-
floating and plastic organization with many inap- neuronal axonal path finding, and contacts with
propriate connections through profound activity- skeletal muscles appears likely (Hanson, M. G. and
dependent rewiring, involving both sprouting and Landmesser, L. T., 2004; 2006), their role in the CNS
elimination of afferent connections. In this process, and for sensorimotor learning in particular has, how-
tactile input seems to guide the alignment of tactile ever, not been known. The activity appears to be
and nociceptive termination since disturbed tactile caused by spontaneous endogenous activity in neu-
input patterns results in disturbed termination patterns ronal circuits in the spinal cord and brain stem
of both tactile and nociceptive afferent fibers (Granmo, (Blumberg, M. S. and Lucas, D. E., 2000). Although
G. et al., 2002; Petersson, P. et al., 2004; Granmo, G. et al., present classifications tend to lump the spontaneous
2005; 2007). The activity-dependent changes in the motility broadly into a few categories, detailed stu-
organization of the primary afferent termination in dies in humans distinguished 16 different types (De
the spinal cord including changes in laminar termina- Vries, J. I. et al., 1982). The prevalence and
tion patterns of tactile afferents are paralleled in time complexity of these movements lead us to suggest
by reflex sensorimotor transformations, suggesting that that all major spinal motor systems contribute to
the emerging action-based body representation is at the spontaneous movements during development.
least partly imprinted on the first-order synapses. Furthermore, since somatosensory imprinting is
152 Appropriate/Inappropriate Developed Pain Paths

highly effective, it may well be that all major groups functional assembly of the dorsal horn. The elimina-
of spinal motor systems learn relevant aspects of the tion of inappropriate connections and strengthening
body anatomy and biomechanics during develop- of appropriate connections that normally occurs dur-
ment by probing the sensory feedback after ing development is likely to be a reflection of such a
spontaneous endogenous activation. It has also been learning-dependent process.
suggested that the sensory input arising after sponta- Besides providing insights into how the wiring of
neous movements guide the tuning of the the spinal cord is accomplished, the reviewed find-
somatotopic organization of the somatosensory cor- ings may well have clinical relevance. While the
tex during development (Khazipov, R. et al., 2004). It repair of the spinal cord after injury has so far been
may thus be proposed that sensory input arising as a relatively unsuccessful, some advances indicate that
consequence of spontaneous movements during regeneration across spinal lesions is possible and that
development plays a key role also in the organization some functional recovery may occur after such
of higher-order sensorimotor systems. regeneration (Li, Y. et al., 1997; Zgraggen, W. J.
As yet, little is known about the molecular et al., 1998; Levinsson, A. et al., 2000). A major pro-
mechanisms underlying somatosensory imprinting. blem, once the obstacle of limited regeneration has
While calcium transients (Perrier, J. F. and been overcome, will be to eliminate the erroneous
Hounsgaard, J., 2000; Russier, M. et al., 2003) and/or connections arising from regeneration (Bareyre, F.
spontaneous fluctuations of glutamate release M. et al., 2004). Such connections may be devastating
(ODonovan, M. J. et al., 1998; ODonovan, M. J., for the patient. For example, patients with partially
1999) have been suggested to play a role for the transected spinal cords often report chronic pain that
initiation of spontaneous movements, N-methyl-D- is very difficult to alleviate (Sjolund, B. H., 2002). If
aspartate (NMDA) receptors may also be involved adaptive mechanisms, such as somatosensory
in the learning mechanisms since the rewiring of the imprinting, which decrease the gain in erroneous
tactile afferent fiber termination in the dorsal horn connections and increase the gain in adequate con-
during development is blocked by topically applied nections can be utilized, true functional recovery
MK-801 (Mendelson, B., 1994; Beggs, S. et al., 2002; may be attainable also from chaotically regenerated
Granmo, G. et al., 2002; 2005). Moreover, mice with a connections. A combination therapy based on train-
disturbed long-term potentiation (LTP) in the hip- ing using feedback stimulation to tune the
pocampus exhibit disturbed receptive fields of NWR, regenerated connections and pharmacological treat-
suggesting that LTP-like mechanisms in the spinal ment that disinhibits regeneration might thus prove
cord may be involved in somatosensory imprinting to be extremely useful.
(Thelin, J. M., 2005). To further unravel the mechan-
isms that underlie the somatosensory imprinting is
clearly an important task in future. Acknowledgments

This work was supported by Swedish MRC project

12.5 Concluding Remarks no 01013 and a Linne grant, Knut and Alice
Wallenbergs Foundation, Kocks Foundation.
The embryonic development of the spinal cord is
characterized by a ventral to dorsal temporal
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 13 Pain Control: A Child-Centered Approach
Patricia A McGrath, The University of Toronto, Toronto, ON, Canada
2009 Elsevier Inc. All rights reserved.

13.1 Introduction 155

13.2 Modifying a Childs Pain 155
13.3 Guidelines for Treating Acute Pain 157
13.4 Guidelines for Treating Chronic Pain 161
References 163

Glossary
behavioral factors These factors refer to the spe- emotional factors These factors include parents
cific behaviors of children, parents, and staff during and childrens feelings about the pain itself or
pain episodes and also to parents and childrens responsible health condition and its adverse impact
broader actions in response to a chronic pain on the family, as well as any associated emotional
problem. disorders (such as anxiety or depression).
cognitive factors These factors include parents situational factors These factors represent a
and childrens understanding of the pain problem, unique interaction between the child and the con-
knowledge of effective therapies, and expectations text in which the pain is experienced; they can vary
for continuing pain or pain relief. dynamically, depending on the specific circum-
stances and setting.

13.1 Introduction with pain, identifying contributing psychological and

contextual factors, and then targeting interventions
During the last two decades, unprecedented scientific accordingly. This chapter describes such a treatment
and clinical attention has focused on the unique pain approach, based on the unique factors that modify a
problems of infants, children, and adolescents childs pain.
(Schechter, N. L. et al., 2003; McGrath, P. A., 2005;
Schechter, N. L., 2007). Our knowledge of how chil-
dren perceive pain and how we can alleviate their 13.2 Modifying a Childs Pain
suffering has dramatically improved. Like adults,
childrens pain is modified both by ascending systems Certain modifying factors are relevant for all
activated by sensory stimuli and by descending pain- childrens pain experiences, irrespective of etiology.
suppressing systems activated by psychological fac- The model shown in Figure 1 lists the key factors
tors. However, childrens pain seems more plastic or that can increase pain, intensify distress, and exacer-
modifiable in comparison to adults, so that environ- bate pain-related disability. A childs characteristics,
mental and psychological factors may exert a more such as cognitive level, sex, gender, temperament,
powerful influence on childrens pain perceptions previous pain experience, family, and cultural
(McGrath, P. A. and Dade, L. A., 2004). background (listed in the shaded box), generally
Plasticity, from both a biological and a psycholo- shape how children interpret and cope with pain
gical perspective, is an essential concept in treating a (Blount, R. L. et al., 1991; Schechter, N. L. et al.,
childs pain. Our treatment approach has shifted from 1991; Bennett-Branson, S. and Craig, K., 1993;
an almost exclusive disease-centered focus detecting Schanberg, L. E. et al., 1998; Chambers, C. T., 2003,
and treating the putative source of tissue damage to a Uman, L. S. and Chambers, C. T., 2007). However,
broader child-centered perspective, assessing the child other factors (listed in the open boxes) represent a

155
156 Pain Control: A Child-Centered Approach

Cognitive Behavioral Emotional

Lack of information about cause and Overt distress responses Anticipatory anxiety
prognosis
Inconsistent use of drug and nondrug Heightened distress
Lack of knowledge about practical therapy
drug and nondrug therapy Fear re. undiagnosed condition and
Child or family responses that prolong continuing pain
Unrealistic or negative expectations pain and disability
about treatment efficacy Situation-specific stress (school,
Failure to fully resolve stressful sports, social)
Inaccurate identification of pain situations
triggers Frustration re. disruption to
Withdrawal from activities (school, activities
Failure to recognize or know how to sports, social)
resolve stress Underlying anxiety or depression

Tissue damage Child

or Age Pain
stressful situation Gender
Cognitive level
Previous pain experience
Family learning
Culture

Figure 1 Biopsychosocial model for pain.

unique interaction between the child and the context Behavioral factors refer to the specific behaviors of
in which the pain is experienced. These situational children, parents, and staff during pain episodes and
factors can vary dynamically, depending on the spe- also to parents and childrens broader actions in
cific circumstances and setting. For example, a child response to a chronic pain problem. Emotional factors
receiving treatment for cancer may have repeated include parents and childrens feelings about the pain
injections, central venous port access, or lumbar itself or responsible health condition and its adverse
punctures, any of which can cause pain. Yet, even impact on the family, as well as any associated emo-
when tissue damage from these procedures is consis- tional disorders (such as anxiety or depression).
tent, the situational factors present for a particular Parents understanding of the cause of pain, possible
treatment are unique for each child. treatments, and long-term prognosis guides their beha-
The beliefs, behaviors, and emotional states of the viors toward children and shapes childrens emotional
children, parents, and clinicians all play important roles responses to the pain problem.
in modifying childrens pain. Certain factors can inten- Situational factors may affect children even more
sify pain and distress, whereas others can eventually than adults. Adults typically have experienced a wide
trigger pain episodes, prolong pain-related disability, variety of pains differing in etiology, intensity, and
or maintain the cycle of repeated pain episodes for quality; their diverse experiences create a broad base
children with recurrent pain syndromes (McGrath, P. of knowledge, realistic expectations, and versatile cop-
A. and Hillier, L. M., 2003). While they may be unable ing behaviors. In contrast, children, especially very
to change the more stable child characteristics, health young children, have considerably limited pain experi-
care providers can dramatically improve childrens ence. They rely mainly on information conveyed
pain by modifying situational factors. Cognitive factors within the immediate situation to interpret pain and
include parents and childrens understanding of the to respond appropriately. Thus, children are particu-
pain problem, knowledge of effective therapies, and larly sensitive to environmental cues and to the
expectations for continuing pain or pain relief. behaviors of the adults who are present. A childs pain
 Pain Control: A Child-Centered Approach 157

is not predetermined by the extent of injury, disease et al., 2003; Brown, S. C. 2006.) Premature and term
progression, or invasiveness of a medical procedure. newborns show reduced clearance of most opioids
When health care providers ignore the powerful (Wong, C. M. et al., 2003). The differences in pharma-
mediating impact of situational factors, they can easily, cokinetics and pharmacodynamics among neonates,
albeit inadvertently, increase a childs distress and pain. preterm infants, and full-term infants warrant special
Generally, accurate information, realistic expectations, dosing considerations for infants and close monitoring
choices, control, and the use of some independent pain- when they receive opioids.
reducing strategies will result in less pain. Recommended starting doses for analgesic medi-
cations for children are listed in Tables 1 and 2.
Starting doses for adjuvant analgesic medications to
13.3 Guidelines for Treating Acute control pain, drug-related side effects, and other
Pain symptoms are listed in Table 3.
Analgesics should be complemented by a practical
Although distraction and parental support can mini- cognitive-behavioral approach to ensure optimal pain
mize the pain caused by minor bumps and scrapes of relief, especially when treating acute pain from inva-
childhood, pharmacological therapy (analgesics, sive medical procedures. First, health care providers
adjunct analgesics, anesthetics, anxiolytics, and seda- should appreciate that children learn about pain
tives, as reviewed in other chapters) is the foundation because of their own injury-related acute pains.
for controlling childrens acute pain (Finley, G. A. and They perceive acute pains more as the occasionally
McGrath, P. J., 2001; Dalens, B., 2003; Krane, E. J. et al., inevitable result of daily activities than as something
2003; Maunuksela, E.-L. and Olkkola, K. T., 2003; to fear. They quickly learn that pain is caused by
Yaster, M., 2003; Wilder, R. T., 2003; Brown, S. C. injury, often easily visible, is relatively brief, and is
et al., 2005; Bean-Lijewski, J. D., 2007; Schneeweiss, S., relieved by many drug and nondrug interventions.
2007). Children require appropriate analgesics, i.e., Thus, they usually have accurate developmentally
drugs selected according to type and severity of appropriate information about why something
pain, doses determined by a childs weight based on hurts and positive expectations for pain relief. In con-
the drugs duration of action to provide consistent trast, when children experience pain during invasive
pain relief and prevent breakthrough pain throughout medical and dental procedures or after surgery, they
the recovery period, and doses administered at regular often feel that they have no control, they may not
dosing intervals. (For dosing guidelines, please see know what to expect, and they may not know any
Finley, G. A. and McGrath, P. J., 2001; Krane, E. J. simple pain control methods that they can use in these

Table 1 Nonopioid drugs to control pain in children

Drug Dosage Comments

1
Acetaminophen 1015 mg kg PO, Lacks gastrointestinal and hematological side effects; lacks anti-inflammatory
every 46 h effects (may mask infection-associated fever)
Dose limit of 65 mg kg 1 day1 or 4 g day1, whichever is less
Ibuprofen 510 mg kg1 PO, Anti-inflammatory activity
every 68 h Use with caution in patients with hepatic or renal impairment, compromised
cardiac function or hypertension (may cause fluid retention, edema), history of
GI bleeding or ulcers, may inhibit platelet aggregation
Dose limit of 40 mg kg 1 day1; maximum dose of 2400 mg day1
Naproxen 1020 mg kg1 day1 Anti-inflammatory activity. Use with caution and monitor closely in patients with
PO, divided impaired renal function. Avoid in patients with severe renal impairment
every 12 h Dose limit of 1 g day1
Diclofenac 1 mg kg1 PO, Anti-inflammatory activity. Similar GI, renal, and hepatic precautions as noted
every 812 h above for ibuprofen and naproxen
Dose limit of 50 mg dose1

Note: Increasing the dose of nonopioids beyond the recommended therapeutic level produces a ceiling effect, in that there is no additional
analgesia but there are major increases in toxicity and side effects.
PO, by mouth; GI, gastrointestinal.
Brown, S. C. and McGrath, P. A. In Press. Pain Control in Children. In: Oxford Textbook of Palliative Medicine 4/e (eds. G. Hanks, N.
Cherny, N. Christakis, M. Fallon, S. Kaasa, R. Portenoy). Oxford University Press.
158 Pain Control: A Child-Centered Approach

Table 2 Opioid analgesics usual starting doses for children

Equianalgesic IV:PO Starting dose PO/ Duration

Drug dose (parenteral) Starting Dose IV ratio transdermal of action

Morphine 10 mg Bolus dose 0.05 1:3 0.150.3 mg kg1 dose1 34 h

0.1 mg kg1 every every 4 h
24 h
Continuous
infusion 0.01
0.04 mg kg1 h1
Hydromorphone 1.5 mg 0.0150.02 mg kg1 1:5 0.06 mg kg1 every 34 h 24 h
every 4 h
Codeine 120 mg Not recommended 1.0 mg kg1 every 4 h (dose 34 h
limit 1.5 mg kg1dose1)
Oxycodone 510 mg Not recommended 0.10.2 mg kg1 every 34 h 34 h
Meperidinea 75 mg 0.51.0 mg kg1 every 1:4 1.02.0 mg kg1 every 34 h 13 h
34 h (dose limit 150 mg)
Fentanylb 100 mg 12 mg kg1h1 as 25 mg patch 72 h
continuous infusion (patch)
a
Avoid use in renal impairment. Metabolite may cause seizures.
b
Potentially highly toxic. Not for use in acute pain control.
Principles of opioid administration:
1. If inadequate pain relief and no toxicity at peak onset of opioid action, increase dose in 50% increments.
2. Avoid IM administration.
3. Whenever using continuous infusion, plan for hourly rescue doses with short onset opioids if needed. Rescue dose is usually 50200%
of continuous hourly dose. If greater than 6 rescues are necessary in 24-h period, increase daily infusion total by the total amount of rescues
for previous 24 h/24. An alternative is to increase infusion by 50%.
4. To change opioids because of incomplete cross-tolerance: if changing between opioids with short duration of action, start new opioid at
50% of equianalgesic dose. Titrate to effect.
5. To taper opioids anyone on opioids over 1 week must be tapered to avoid withdrawal: taper by 50% for 2 days, and then decrease by
25% every 2 days. When dose is equianalgesic to an oral morphine dose of 0.6 mg kg 1 day1, it may be stopped. Some patients on
opioids for prolonged periods may require much slower weaning.
PO, by mouth; IV, intravenous.
Brown, S. C. and McGrath, P. A. In Press. Pain Control in Children. In: Oxford Textbook of Palliative Medicine 4/e (eds. G. Hanks,
N. Cherny, N. Christakis, M. Fallon, S. Kaasa, and R. Portenoy). Oxford University Press.
NOTE: Doses are for opioid-naive patients. For infants under 6 months, start at one-quarter to one-third the suggested dose and titrate to
effect.

new situations. In fact, health care providers may relevance of the situation (i.e., injury, disease, treat-
instruct children how to cope (i.e., behave) during a ment) and consequent pain. The manner in which
painful procedure in a manner that makes it easier for health care providers introduce themselves, the
the person administering the procedure, but does not words they choose to explain what will happen, and
necessarily make it better for the child. whether they confidently show children what they
Health care providers often focus on explaining to could do to lessen the pain profoundly impact what
parents and inadvertently neglect the child. Time children will experience.
constraints necessitate juggling who has priority These situational factors can intensify childrens
and, understandably, concerned parents are a prior- pain and distress, even when children receive anxio-
ity. Yet, children should remain the number 1 lytics and analgesics. As an example, when children
priority because it is their pain that is directly mod- with cancer at our center first started receiving topi-
ified. Every word and action conveys powerful cal anesthetic creams before intravenous insertions
information to children. As an example, a young and portacather injections, considerably more
boy became very distressed when he learned that a children were referred to our pain clinic. We learned
local anesthetic should freeze his arm so he wouldnt that busy clinical staff no longer focused on an indi-
feel anything. Finally, he explained that he was afraid vidual child in the manner that they had previously
his arm would become like a Popsicle. Health care encouraging the child to understand from their
providers have tremendous power in influencing specific perspective (i.e., age, gender, culture) and
what children know, how they can respond, and the use their preferred close your pain gate methods.
Table 3 Adjuvant analgesics doses for children

Drug category Drug, dosage Indications Comments

Antidepressants Amitriptyline, Neuropathic pain Usually improved sleep and pain

0.20.5 mg kg1 PO. (i.e., vincristine-induced, relief within 35 days
radiation plexopathy,
tumor invasion, CRPS-1)
Titrate upward by 0.25 mg kg1 Insomnia Anticholinergic side effects are
every 23 days dose-limiting. Use with caution for
children with increased risk for
Maintenance: cardiac dysfunction
0.23.0 mg kg1
Alternatives: nortriptyline,
doxepin, imipramine
Anticonvulsants Gabapentin, Neuropathic pain, Side effects: gastrointestinal upset,
5 mg kg1day1 PO. especially shooting, ataxia, dizziness, disorientation,
stabbing pain. somnolence
Titrate upward over Monitor for hematological, hepatic,
37 days. Maintenance: and allergic reactions with
up to 1550 mg kg1day1 carbamazepine
PO divided TID.
Carbamazepine, initial dosing:
10 mg kg1day1 PO divided OD
or BID. Maintenance: up to
2030 mg kg1day1
PO divided every 8 h.
Increase dose gradually
over 24 weeks.
Alternatives: phenytoin,
clonazepam

(Continued )
Table 3 (Continued)

Drug category Drug, dosage Indications Comments

Sedatives, Diazepam, Acute anxiety, muscle Sedative effect may limit opioid use.
hypnotics, 0.0250.2 mg kg1 spasm Other side effects include
anxiolytics PO every 6 h. depression and dependence with
prolonged use
Lorazepam, Premedication for
0.05 mg kg1dose1 SL painful procedures
Midazolam, 0.5 mg kg1dose1
PO administered 1530 min
prior to procedure;
0.05 mg kg1dose1 IV for
sedation
Antihistamines Hydroxyzine, 0.5 mg kg1 Opioid-induced pruritus, anxiety, nausea Sedative side effects may be helpful
PO every 6 h.
Diphenhydramine, 0.51.0 mg kg1
PO/IV every 6 h
Psychostimulants Dextroamphetamine, methylphenidate, Opioid-induced somnolence Side effects include agitation, sleep disturbance,
0.10.2 mg kg1 BID. and anorexia.
Escalate to 0.30.5 mg kg1 as needed Potentiation of opioid analgesia Administer second dose in afternoon to avoid
sleep disturbances

Corticosteroids Prednisone, prednisolone, and dexamethasone Headache from increased intracranial pressure, Side effects include edema, dyspeptic symptoms,
dosage depends on clinical situation (i.e. spinal or nerve compression; widespread and occasional gastrointestinal bleeding
dexamethasone initial dosing: 0.2 mg kg1 IV. metastases
Dose limit 10 mg. Subsequent dose 0.3 mg
kg1day1 IV divided every 6 h)

CRPS-1, complex regional pain syndrome, type 1; PO, by mouth; IV, intravenous; SL, sublingual.
Brown, S. C. and McGrath, P. A. In Press. Pain Control in Children. In: Oxford Textbook of Palliative Medicine 4/e (eds. G. Hanks, N. Cherny, N. Christakis, M. Fallon, S. Kaasa, and R. Portenoy).
Oxford University Press.
 Pain Control: A Child-Centered Approach 161

Instead, they applied the magic cream and sent procedure is inevitable and a cycle of heightened
children to a play area until it was time to conduct anxiety, increased pain, and emotional distress
the invasive procedure. Despite the proven anes- ensues.
thetic properties of the cream, many children Many cognitive-behavioral programs have been
reported increased pain and exhibited increased dis- designed to complement drug therapies for children
tress in comparison to their pretopical procedures with cancer (Hilgard, J. R. and LeBaron, S., 1984;
(McGrath, P. A., 1990). LeBaron, S. and Zeltzer, L. K., 1996; Olness, K. and
Although psychological therapies are reviewed in Kohen, D. P., 1996; Liossi, C., 2002; Collins, J. and
other chapters, even the simple provision of informa- Weisman, S. J., 2003; Kuttner, L. and Solomon, R.,
tion about what is happening, why the procedure is 2003). These child-centered programs usually com-
conducted in a certain manner, what equipment will bine basic education with psychological therapies
be used, who will be present, the nature of the childs (i.e., attention and distraction, hypnosis or hypnotic-
sensations during different phases of the procedure, like suggestions for analgesia, desensitization proce-
and some straightforward instructions about how a dures, behavioral management, relaxation exercises,
child might lessen pain will dramatically reduce the and biofeedback) to minimize childrens anxiety and
pain caused by any invasive procedure to minimize pain. The specific program components vary for dif-
anxiety and pain. As listed in Table 4, health care ferent children because therapies are selected to
providers should explain what will happen and target the unique child, family, and situational factors
emphasize what sensations a child may feel, e.g., that were identified from the childs pain assessment
warmth, coolness, pressure, rather than the frighten- as the factors that intensified childrens pain, anxiety,
ing label of pain. They should teach children a few and distress. Generally, as children receive develop-
basic attention and distraction methods to reduce mentally appropriate accurate information, gain
pain and guide families to recognize the particular realistic expectations, have more choices and control,
and use independent pain control strategies, they
circumstances that exacerbate pain and distress.
have less pain and distress.
Children who receive multiple invasive proce-
dures over a prolonged period are at particular risk
for developing anxiety and fear. Inaccurate under-
standing of the disease or injury, parents anxiety 13.4 Guidelines for Treating Chronic
about the underlying condition and painful treat- Pain
ments, inconsistency in treatment administration,
lack of control and coping strategies, and fears Effective treatment of chronic pain usually requires
about the effects of the disease or side effects of an integrated treatment approach comprising drug
treatment can intensify a childs treatment-related therapy, physical therapy, and psychological coun-
pain. Regrettably, some inconsistencies in how pro- seling, rather than a single therapy. Chronic pain
cedures are conducted are common in clinical generally has multiple sources affecting both periph-
practice. The more procedures a child has, the eral and central nervous systems, with both
more likely that they will experience a difficult pro- nociceptive and neuropathic components. Certain
cedure more pain, more nausea, etc. A child then cognitive, behavioral, and emotional factors may
becomes fearful and expects that another difficult intensify pain, increase distress, and exacerbate a

Table 4 Preparing children for invasive procedures

Remember that anxiety and distress can increase pain.

Explain what will happen clearly, in simple developmentally appropriate language.
Emphasize the qualities of the sensations they may experience such as cold, sharp, tingling, and pressure so that children
focus on what they are feeling, rather than only on the hurting aspect.
If the procedure will hurt, describe what children might feel using more familiar examples of pains that they have experienced
during play and sports.
Use versatile pain control methods that involve children such as attention and distraction, deep breathing, counting
exercises, or guided imagery.
Give children as many choices and as much control as possible such as choosing which arm for injections, whether to watch
or look away, and which pain control method to use.
162 Pain Control: A Child-Centered Approach

childs disability. The pain adversely affects all pain varies widely among different children or in the
aspects of childrens lives. Parents are distressed by same child at different times. To relieve ongoing
the pain itself, its implications for their childrens pain, opioid doses should be increased steadily until
future, its life-threatening potential (if any), and the comfort is achieved, unless the child experiences
prospect of continuing pain and progressive disabil- unacceptable side effects such as somnolence or
ity. Parents and adolescents tend to emphasize and respiratory depression. Adjuvant analgesics (espe-
fear the future consequences of childrens physical cially gabapentin and tricyclic antidepressants) have
condition, whereas young children are more preoc- become the main drugs for treating neuropathic pain
cupied by the immediate consequences and (Berde, C. B. et al., 2003).
disruption to their daily activities. The dynamics Some children, irrespective of what may have
within the family (both for siblings and for extended caused their pain initially, develop significant func-
family members) inevitably change as chronic pain tional disability that far exceeds their reported pain
prevents children from pursing their normal level. These children do not attend school, partici-
activities and as family schedules adjust accordingly pate in sports, or socialize with peers. Families are
to the health care needs of the child with pain (for frustrated because they may have tried to follow
review, please see McGrath, P. J. and Finley, G. A., treatment recommendations and reintegrate children
1999; Berde, C. B. et al., 2003; McGrath, P. A. and back into their normal activities. Parents may believe
Dade, L. A., 2004). that health care providers do not believe the child
In order to treat chronic pain effectively, health and do not understand the severity of the pain pro-
care providers should carefully determine the physical blem. Families believe that children will return to
etiology (if possible) and also critically appraise the normal (even when children have been disabled for
unique situational factors that may influence pain. years) as soon as health care providers cure the pain
Interdisciplinary teams, comprising physicians, nurses, problem. A futile cycle ensues, with health care pro-
psychologists, psychiatrists, and physical therapists, viders attempting to restore normal functioning and
are better equipped to conduct a more comprehensive families refusing to cooperate until the pain is
biopsychosocial assessment than a single pain specia- relieved. Children with severe disability constitute a
list. After consultation among members, the team unique clinical challenge. Gradually, I have learned
should provide a diagnosis that includes both the to identify the problem to families as twofold a
etiology and the relevant factors influencing childrens chronic pain problem and a disability problem.
pain. The biopsychosocial model shown in Figure 1 Usually disability is the more serious problem.
provides a concrete framework for explaining these Thus, treatment should immediately focus on the
factors and subsequently presenting the rationale for both aspects we must deal with the pain, but, con-
an integrated treatment approach comprising pharma- currently, we must deal with the increasing disability.
cology, physiotherapy, and psychology. The specific Almost all children who have a major disability pro-
drugs and therapies within this child-centered blem will require additional psychological treatment
approach derive from the assessment results and are and assistance to reenter school and resume their
selected to target the responsible central and periph- previous activities.
eral mechanisms and to mitigate the pain-exacerbating For any child with chronic pain, health care pro-
impact of situational factors. vides should recognize the common situational
NSAIDs and opioids in effective doses and regular factors that influence parents responses to childrens
dosing schedules can relieve pain with a primary continuing pain complaints, especially when pain is
nociceptive etiology. Drugs should be selected not caused by active disease or injury. Parents usually
according to their analgesic potency based on the try to understand their childs pain from an acute
childs pain level acetaminophen to control mild disease perspective, where pain is due to a single
pain, codeine to control moderate pain, and mor- cause and can be relieved by a single treatment.
phine for strong pain (World Health Organization, They do not understand that chronic pain, unlike
1998). Even when children require opioid analgesics, most pains that they have already experienced, may
they should continue to receive acetaminophen (and have several interrelated causes. Thus, parents
NSAIDs, if appropriate) as supplemental analgesics. request further medical investigations as they search
Health care providers should monitor a childs pain for the clear-cut physical etiology and concentrate on
regularly and adjust analgesic doses as necessary to finding one treatment that will immediately stop the
control the pain. The effective opioid dose to relieve pain. Parents may reject potentially effective
 Pain Control: A Child-Centered Approach 163

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New York. Springer-Verlag. and R. Portenoy). Oxford University Press.
Uman, L. S. and Chambers, C. T. 2007. Impact of Familial Schanberg, L. E., Gil, K. M., Anthony, K. K., Yow, E., and
Factors on Childrens Chronic Pain. In: Encyclopedic Rochon, J. 2005. Pain, stiffness, and fatigue in juvenile
Reference of Pain (eds. R. F. Schmidt and W. D. Willis), polyarticular arthritis: contemporaneous stressful events and
pp. 961964. Springer-Verlag. mood as predictors. Arthritis Rheum. 52(4), 11961204.
 14 Assaying Pain-Related Genes: Preclinical and
Clinical Correlates
V E Scott, R Davis-Taber, and P Honore, Global Pharmaceutical Research and Development, Abbott
Park, IL, USA
2009 Elsevier Inc. All rights reserved.

14.1 Introduction 165

14.2 Microarray Analysis of Whole Dorsal Root Ganglion from the Spinal Nerve
Ligation Model 165
14.3 Microarray Data Analysis and Follow-up Studies 167
14.4 Microarray Analysis of Subpopulations of Neurons Isolated from Dorsal Root Ganglion 167
14.5 Evaluation of Additional Neuropathic Pain Models by Microarray 169
14.6 The Strengths and Limitations of Microarray Analysis for Pain Research 170
14.7 The Genetics of Pain 171
References 172

14.1 Introduction studies. Microarray studies are clearly just the start-
ing point. This chapter will be divided into sections
Microarray gene chip technology is a powerful tool to detailing the studies that have been reported evalu-
evaluate changes in multiple transcript levels simulta- ating gene expression changes from neuropathic pain
neously in different tissue preparations. In the pain models using microarray gene chip analysis, includ-
field, efforts have focused on characterizing gene tran- ing discussion in each section on study design,
script changes that occur in chronic pain states such as sample selection, isolation of RNA, choice of micro-
neuropathic pain (Wang, H. et al., 2002; Xiao, H. S. et al., array gene chip platforms, and data analysis. The
2002; Davis-Taber, R. et al., 2003; Thimmapaya, R. follow-up to microarray studies outlining the expec-
et al., 2003; Valder, C. R. et al., 2003; Davis-Taber, R. tations and limitations of these studies will also be
et al., 2004) and this will be the main focus of this described. Lastly, the use of microarray gene chips
chapter. Neuropathic pain is initiated or caused by a for pinpointing genetic variation in disease states that
primary lesion or dysfunction of the nervous system. will direct suitable therapeutics based upon genetic
As this technology emerged, it was anticipated that it predispositions for specific patient populations will
could be used to determine the key molecular players be discussed.
in both the development and maintenance of neuro-
pathic pain, which would enhance our understanding
of this disease and provide insights into the identifica- 14.2 Microarray Analysis of Whole
tion of potential therapeutic targets for the treatment of Dorsal Root Ganglion from the Spinal
pain. Prior to microarray approaches, studies were Nerve Ligation Model
limited to evaluation of a set of specific preselected
genes/proteins in samples from animal pain models or Several different studies have evaluated the expression
human tissues using techniques such as quantitative changes that occur in the rat whole dorsal root gang-
reverse transcriptase polymerase chain reaction (RT- lion (DRG; Wang, H. et al., 2002; Thimmapaya, R.
PCR), Western blot analysis, and immunocytochemis- et al., 2003; Valder, C. R. et al., 2003) and dorsal horn
try. Dependent upon the gene chips used, this analysis of the spinal cord (Sun, H. et al., 2002; Wang, H. et al.,
permits evaluation of both known and unknown gene 2002) following spinal nerve ligation (SNL) using
transcripts (expressed sequence tags (ESTs)) using microarray analysis. The overall goal of each of these
relatively small amounts of RNA. In each of the studies studies was to obtain a better understanding of the
highlighted in this chapter a wealth of transcriptional molecular pathophysiology of neuropathic pain and
information was obtained. However, determination of through this analysis identify potential new targets for
which gene product would offer the best therapeutic the treatment of this chronic pain state. In some stu-
potential will require extensive additional follow-up dies following tight ligation of the lumbar 5 (L5) and

165
166 Assaying Pain-Related Genes: Preclinical and Clinical Correlates

lumbar 6 (L6) DRG, the development of allodynia was both strains, that is, Nav1.8 and protein kinase C
shown to develop 3 days following surgery and epsilon highlighting the limitations of this study.
remained established for weeks ipsilateral to the insult A critical consideration when designing these
(Wang, H. et al., 2002; Valder, C. R. et al., 2003). In experiments is to ensure that sufficient samples are
these studies each animal was assessed for the devel- collected to reflect the true changes that arise due to
opment of tactile allodynia by behavioral testing prior neuropathic pain and animal variation accurately. To
to being included/excluded from the study. This was this end assessment of DRG from L5/L6 region was
performed using von Frey filaments and to be performed using at least ten animals that were pooled
included in the study animals needed to have a paw prior to isolation of RNA for each of these studies. The
withdrawal threshold of less than 3 g in the neuro- comparison was made between the injured L5/L6 and
pathic rats and greater than 10 g in the control rats. either the contralateral L5/L6 (Wang, H. et al., 2002) or
While some studies focused on evaluating the changes sham L5/L6 (Thimmapaya, R. et al., 2003). In one of
that occur both during the development and in fully the spinal cord studies nine animals were divided into
established neuropathic pain (Wang, H. et al., 2002), three groups and in each group the dorsal and ventral
others assessed only established pain (Sun, H. et al., spinal cords were pooled separately to form each pair
2002; Thimmapaya, R. et al., 2003; Valder, C. R. et al., of samples (three pairs). The comparison that was
2003; Davis-Taber et al., 2003). Several experimental made in this tissue was between the dorsal and ventral
design differences exist in each of these reports. In spinal cord of neuropathic rats (Wang, H. et al., 2002).
some studies since the contralateral paw did not dis- In the other study, the global gene expression in the
play significant allodynia similarly to the sham ipsilateral lumbar spinal cord was compared with the
animals, the L4/L5/L6 (Wang, H. et al., 2002) or L5/ contralateral lumbar spinal cord from the neuropathic
L6 (Valder, C. R. et al., 2003) DRG contralateral to the rats and to the sham control (Sun, H. et al., 2002).
injury were selected for comparison with the equiva- Typically protocols for isolating total RNA have
lent injured DRGs from the same animals. In a parallel been comparable throughout these studies using the
study to avoid any potential issues that could arise TRizol protocol (Life Technologies/Invitrogen;
during surgery to alter gene expression contralateral Carlsbad, CA, USA) followed by RNeasy (Qiagen;
to the injury and not revealed by allodynia assessment, Valencia, CA, USA) RNA clean-up step. The RNA
the L5/L6 ipsilateral to the surgery were compared quality was assessed either by denatured gel electro-
with the L5/L6 from sham-operated animals phoresis or capillary electrophoresis on the Agilent
(Thimmapaya, R. et al., 2003). 2100 Bioanalyzer (Applied Biosystems; Foster City,
Another interesting comparison examined the CA, USA) to ensure that the 28S:18S ribosomal RNA
expression differences between the injured L5/L6 (rRNA) ratio was greater than 1.0 for each sample,
DRG from Harlan and Holtzman Sprague-Dawley indicating that the sample was pure.
rats (Valder, C. R. et al., 2003). The Holtzman rats The microarray gene chip platform that was
were selected for this analysis as they do not develop selected for each of these studies was the Affymetrix
the tactile allodynia observed in Harlan rats follow- rat RG-U34a GeneChip. On these microarray chips,
ing SNL (Luo, Z. D. et al., 1999). The goal of this each gene transcript is represented by probe sets con-
study was to compare the expression profiles taining 16 oligonucleotides, eight of which are matched
between these two strains and then extend these and eight mismatched to the sequence of the transcript
analyses to assess changes that were regulated follow- of interest. The oligonucleotides are 20 base pairs long,
ing injury in one or both strains. This study did with a 39 bias and may be to the coding and noncoding
identify differentially expressed genes in the two regions of the gene. Hybridization probes from each of
strains, providing new information on neuroplasticity these samples were generated and hybridized to the
in DRG of these rats in response to nerve injury chips as recommended by Affymetrix. The microarray
(Valder, C. R. et al., 2003) in addition to those asso- data was analyzed using the SAFER algorithm
ciated with neuropathic pain. However, cautious (Holder, D. et al., 2002), Microarray Suite Expression
analysis of these data is important since similar reg- Algorithm (Affymetrix; Santa Clara, CA, USA) or
ulation of gene changes in both strains is not an Rosetta Resolver (Rosetta Biosoftware; Seattle, WA,
eliminating factor in defining whether the altered USA). Several microarray analysis software packages
gene transcript is related to neuropathic pain devel- are available. Some utilize only a statistical algorithm
opment. Several transcripts that are known to be (Affymetrix analysis software) while other analysis
centrally involved in pain processing did change in packages incorporate error models for normalization
 Assaying Pain-Related Genes: Preclinical and Clinical Correlates 167

of intensity between arrays and adjustments for probe- synaptobrevin) were downregulated, whereas genes
specific biases (SAFER and Resolver). Following a involved in the immune response (e.g., MHCII) and
paired t-test and analysis of variance (ANOVA) analy- cell regeneration/proliferation (e.g., Reg-2 and
sis to determine a ratio for the expression changes GADD45) were upregulated. A similar pattern of
between both samples a P value is generated and changes has been observed in the different neuro-
represents the probability that an observed gene pathic pain studies reported (Wang, H. et al., 2002;
expression change was due to a measurement error. Xiao, H. S. et al., 2002; Valder, C. R. et al., 2003).
To be considered significant, the change in gene
expression had to occur in each microarray experiment
and had to exhibit a value of P  0.05 within each 14.4 Microarray Analysis of
experiment. Another consideration that has been Subpopulations of Neurons Isolated
shown in each of these studies is how much does the from Dorsal Root Ganglion
transcript need to change to be considered significant
enough to play a role in pain? In all the studies dis- All the studies described above examine the gene
cussed in this chapter a cut-off of twofold was selected. changes that occur in the entire DRG, which is
This arbitrary cut-off was chosen since a large number composed of heterogenous subpopulations of cells
of significant changes were observed but it is known (neurons and glial cells). Examining specific subpo-
that the expression of P2X3, a validated pain target, pulations of neurons would provide more precise
only changed by 1.8-fold and would be excluded from information on the transcript expression changes.
this analysis. Therefore it is important not to exclude To accomplish this, LCM was used. This is a novel
all small changes due to magnitude but if their changes technology that permits the isolation and capture
are significant with P  0.05, further analysis is of specific cells from a tissue such as DRG in a
warranted. native state. Determining the gene expression profile
of different cell types has been hampered until
recently by the inability to recover enough high-
14.3 Microarray Data Analysis and quality RNA from these cells for use in microarray
Follow-up Studies experiments. Application of a new technique, anti-
sense RNA (aRNA) amplification, has increased the
Following microarray analysis, the changes that were yield of RNA to accommodate the requirements
observed must be confirmed using either quantitative necessary for complementary DNA (cDNA) spotted
RT-PCR (Q-RTPCR), in situ hybridization, or, if and high-density oligonucleotide microarrays. DRG
reagents are available, immunocytochemistry. The are composed of multiple neuronal and glial cell
latter two are preferable as the microarray studies types and in particular, small-diameter neurons med-
described so far have all examined heterogeneous iate nociceptive responses whereas large neurons
populations of cells (neurons and glial cells) and facilitate mechanosensation. Two studies have pro-
evaluation of specific tissue sections would provide filed the differential gene expression of these
a more precise profile of where the expression nociceptive and mechanosensitive naive neurons by
changes have occurred. Following confirmation of a microarray (Luo, L. et al., 1999; Davis-Taber, R. et al.,
subset of transcripts identified by microarray, data 2003). DRGs were flash frozen in cryomolds, sec-
analysis has focused on classifying the gene tran- tioned to 8 mm thickness and visualized with a
scripts into groups based upon the function of the Nissl (cresyl violet) stain to identify the neurons.
gene product if known. An example of such a classi- Naive small (<25 mm and an identifiable nucleus)
fication comes from the studies examining the and large (>40 mm) diameter neurons were captured
changes in the L5/L6 DRG following SNL com- (PixCell II LCM system; Arcturus, Mountain View,
pared to sham L5/L6 DRG whereby gene products CA, USA) and total RNA extracted from cellular
were grouped into cellular homeostasis and remodel- material adherent to the thermoplastic film on the
ing, ion channels and receptors, enzymes, immune LCM caps. Multiple RNA isolation and amplifica-
response, neuronal markers, synaptic plasticity, tran- tion reagents and kits are commercially available
scription factors, and a miscellaneous category from a variety of vendors (e.g., Arcturus, Mountain
(Thimmapaya, R. et al., 2003). Following this classi- View, CA, USA; Stratagene, San Diego, CA, USA;
fication, it became evident that several examples of Epicentre Technologies, Madison, WI, USA). aRNA
genes involved in synaptic plasticity (e.g., Narp and was amplified two to three times prior to
168 Assaying Pain-Related Genes: Preclinical and Clinical Correlates

hybridization onto a custom cDNA spotted array gene expression suggests that there was minimal
or an Affymetrix high-density oligonucleotide glial contamination in the capturing process.
GeneChip array (Luo, L. et al., 1999; Davis-Taber, R. To understand the gene expression changes that
et al., 2003). occur in neuropathic pain states further, neuronal
There are several aspects of experimental design subpopulations were isolated from DRG following
that should be considered prior to initiation of an SNL or sham surgery and profiled by microarray.
LCM microarray study. First, a sufficient number of Similar to the naive DRG studies, small (1000
cells should be captured. For each of the LCM stu- cells) and large (1000 cells) neurons were captured
dies discussed, approximately 1000 small and 1000 from SNL and sham L5/L6 ipsilateral DRGs (4000
large DRG neurons were isolated for microarray total) by LCM and evaluated by Affymetrix rat RG-
profiling. An alternative to capturing a greater U34a GeneChip arrays (Davis-Taber, R. et al.,
amount of cellular material is increasing the number 2004). Microarray analysis showed altered expression
of rounds of RNA amplification. One trade-off to patterns between SNL and sham animals especially in
higher yields is a shortening of the RNA transcripts large DRG neurons where the majority of gene
with each successive round of amplification. One other changes occurred. Although some genes showed a
consideration is the 39 bias of the aRNA transcripts common expression signature across both small and
generated. This is important when determining the large neurons, a number of genes were dysregulated
microarray platform to use. Many high-density in only one neuronal subtype. A comparison of the
oligonucleotide arrays already incorporate a 39 bias whole DRG SNL microarray data (Thimmapaya, R.
in their probe design (e.g., Affymetrix) but this issue et al., 2003) with the SNL small and large DRG
may be more significant when using custom spotted neuron study revealed only a small number of genes
arrays. that were commonly altered in all three samples. Also,
The initial studies profiling naive small and large only a small percentage of gene expression changes
DRG neurons observed a differential gene expres- observed in the small (35%) or large (22%) neurons
sion pattern for these two neuronal subpopulations. were dysregulated in the whole DRG microarray
In the first study, naive cervical DRG aRNA from study. This suggests that within the individual neu-
adult rats were hybridized to a custom cDNA spotted ronal subtypes, the distinct expression patterns that
microarray (Luo, L. et al., 1999). The microarray chip were observed may have been masked or minimized
had 477 cDNAs that were obtained from two differ- in the whole DRG microarray study. Microarray
ential display experiments: (1) identified genes analysis of the small and large DRG neuronal sub-
uniquely expressed in DRG compared to brain, populations isolated from the injured DRG following
liver, and kidney; and (2) genes that were differen- SNL has provided a deeper understanding of the
tially expressed in L5/L6 between ipsilateral (SNL) differential gene expression associated with neuro-
and contralateral (control). Approximately 40 genes pathic pain (Davis-Taber, R. et al., 2004).
were found to be enriched in either neuronal sub- A potential limitation of the LCM microarray stu-
population with a greater majority detected in the dies is the ability to collect a pure subpopulation of
small DRG neurons. The second study collected neurons. The time constraints of standard immunohis-
similar sized neuronal cells from rat lumbar L5/L6 tochemical staining procedures would allow tissue
DRG and hybridized the aRNA to the Affymetrix sections to rehydrate which could hinder precise
Neurobiology GeneChip (Davis-Taber, R. et al., capture of individual cells so a modified cresyl violet
2003). In the small and large DRG neurons, 100 staining protocol developed by Arcturus Engineering,
genes were detected and classified into functional Mountain View, CA, USA, was used to visualize both
groups (channels and receptors, enzymes, cell signal- small and large diameter DRG neurons. Without a
ing, and cytoskeletal). In the categories of cell way to distinguish the different small DRG neuron
signaling and channels and receptors, there was an subtypes, a mixed population of peptidergic and non-
equal number of genes differentially enriched in each peptidergic neurons were captured and pooled
neuronal subpopulation, however, large DRG neu- together. TRPV1 expression is an example of this
rons predominantly expressed cytoskeletal proteins limitation whereby it was shown to be downregulated
and enzymes. Examining the results reported in both in the L5/L6 small DRG neurons in the LCM SNL
experiments has shown that similar expression pat- study but was not detected in the naive, L5/L6 small
terns and fold enrichment were observed for both DRG neurons. Localization studies have shown that
neuronal subpopulations. A lack of glial-specific TRPV1 is found on peptidergic small-diameter
 Assaying Pain-Related Genes: Preclinical and Clinical Correlates 169

neurons and the receptors inconsistent detection may axotomy (Xiao, H. S. et al., 2002). The L4/L5 DRG
result from the level of sensitivity of RNA amplifica- were then probed against this chip at various time-
tion and microarray techniques from a mixed points to assess the changes in expression that occur
population of small DRG neurons. However, evalua- both during the development and following estab-
tion of the expression levels of RNA from lished pain. Some genes changed expression
subpopulations of neurons has provided a depth of immediately by day 2 (neuropeptide Y, heat shock
information on transcripts that was previously not protein 27, LIM domain protein CLP36) while others
possible when whole DRG samples were assessed. showed a more significant change at 14 days (neuro-
nal nitric oxide synthase (nNOS), calcitonin gene-
related peptide (CGRP)) following axotomy. The
14.5 Evaluation of Additional changes were confirmed for a subset of genes by
Neuropathic Pain Models by in situ hybridization. Interestingly many of the
Microarray genes that changed expression were the same as
those observed in the SNL model (Table 1) indicat-
In addition to studies on the expression changes in ing that these neuropathic pain models share some
DRG from SNL model of neuropathic pain, micro- underlying molecular components. A second study
array analysis has also been performed on DRG used the Affymetrix rat RG-U34a GeneChip gene
following peripheral axotomy. In one study a custom chip and also examined the L4/L5 DRG following
microarray chip was generated that contained genes sciatic nerve axotomy over a time course of 114
that were overexpressed in DRG 14 days following days postinjury (Costigan, M. et al., 2002). This

Table 1 Representative gene expression changes identified by microarray analysis from rat L5/L6 DRG following
peripheral nerve injury

SNL (L5/L6)
Accession (Thimmapaya, Published changes:
number Description R. et al., 2003) preclinical neuropathic models

X59864 ASM15 4.84 " Valder C. R. et al. (2003)

AI009268 Ca2/calmodulin-dependent protein 2.29 " Xiao H. S. et al. (2002)
kinase 2 delta
X81193 Cysteine-rich protein 3 (muscle LIM 42.57 " Wang H. et al. (2002); " Xiao H. S. et al.
protein) (2002); " Valder C. R. et al. (2003)
L21192 GAP43 2.76 " Wang H. et al. (2002); " Xiao H. S. et al.
(2002); " Valder C. R. et al. (2003)
AF028784 Glial fibrillary acidic protein (GFAP) 5.54 " Valder C. R. et al. (2003)
M98049 Pancreatitis-associated protein 20.36 " Wang H. et al. (2002); " Valder C. R.
(Reg-2) et al. (2003)
AA799645 Phospholemman chloride channel 2.45 " Costigan M. et al. (2002)
X80290 Pituitary adenylate cyclase activating 3.04 " Costigan M. et al. (2002); " Wang H.
peptide (PACAP) et al. (2002); " Xiao H. S. et al. (2002)
M18331 Protein kinase C epsilon 5.55 " Xiao H. S. et al. (2002); # Valder C. R.
et al. (2003)
U48246 Protein kinase C-binding protein NELL1 12.15 # Wang H. et al. (2002); " Xiao H. S. et al.
(2002); # Valder C. R. et al. (2003)
AB003991 SNAP-25A 2.50 # Costigan M. et al. (2002); # Wang H.
et al. (2002); # Xiao H. S. et al. (2002);
# Valder C. R. et al. (2003)
AF059030 Sodium voltage-gated channel, Nav1.9 23.85 # Wang H. et al. (2002); # Valder C. R.
et al. (2003)
X52772 Synaptotagmin I 3.03 $ Xiao H. S. et al. (2002)
X59737 Ubiquitous mitochondrial creatine kinase 6.33 # Wang H. et al. (2002)

Genes that show enhanced or diminished expression from injured dorsal root ganglia (DRG) versus sham-operated DRG with a twofold
change and P  0.05. The fold increases and decreases in gene expression observed with DRG from spinal nerve ligated (SNL) rats are
given, together with published microarray analysis of gene changes from preclinical neuropathic pain models (Xiao, H. S. et al., 2002,
axotomy; Wang, H. et al., 2002, SNL; Costigan, M. et al., 2002, axotomy; Valder, C. R. et al., 2003, SNL). Genes were upregulated ("),
downregulated (#), or unchanged ($).
170 Assaying Pain-Related Genes: Preclinical and Clinical Correlates

analysis compared naive DRG to the injured DRG assays are not suitable for detection of post-transla-
and focused on the evaluation of data following this tional modifications of functional proteins including
comparison. The data presented supported the use of a phosphorylation/dephosphorylation of ion channels,
P < 0.05 significance threshold for detecting regulated membrane proteins, and transcription factors.
genes. When a cut-off of a twofold change in expres- Functional modifications of pain-relevant proteins
sion was applied to select gene transcripts to assess, that do not alter their transcriptional levels include
this led to an estimated error rate of 16%. Combining activation of N-methyl-D-aspartate receptors by
a more than 1.5-fold expression change and a P < 0.05 injury-induced protein kinase C-mediated phos-
reduced the estimated error to 5%. Using the latter phorylation (Chen, L. and Huang, L. Y., 1992) and
analysis, 240 genes changed expression significantly the downstream regulatory element antagonistic
and many had not been previously described as chan- modulator (DREAM)-mediated derepression of pro-
ging following axotomy. Changes in expression of a dynorphin expression that is regulated by elevated
subset of the genes were confirmed by in situ and intracellular calcium and protein kinase A activation
Northern blots analysis. This study also identified (Carrion, A. M. et al., 1998). DREAM-deficient mice
increases in gene expression of genes mostly involved have been shown to have elevated spinal cord pro-
in immune responses and inflammation and decreases dynorphin levels and reduced inflammatory pain
in those involved in neurotransmission similar to that behaviors (Cheng, H. Y. et al., 2002).
reported for the SNL model of neuropathic pain. Additionally, translation of the data arising from
Taken together the microarray studies identified these studies to identify potential novel therapeutic
genes that are dysregulated in neuropathic pain states targets is also a challenge. The approach undertaken
from different pain models providing an opportunity in the studies highlighted in this chapter, has been
to unveil potential neuropathic molecular signatures to examine the literature for any data that would
that are common between these models. indicate that the protein encoded by a dysregulated
transcript plays a role in pain. A complicating
factor studying nociception and associated gene reg-
14.6 The Strengths and Limitations ulation in pain states is that unlike metabolic diseases
of Microarray Analysis for Pain where pathways are well understood, little is
Research known about the specific pathways underlying pain.
Application of programs that assess gene ontology
Our knowledge of the complexity of neuropathic (i.e., GenMAPP, GoSurfer, GOTM, WebGestalt,
pain has heightened following the use of microarray GoMiner, OntoExpress, and ErmineJ) and known
gene chip technology. This approach provided pathways (i.e., Pathway Assist) can provide some
opportunities to examine multiple gene transcripts insights into nociceptive signaling.
simultaneously in pain states. Interestingly in these As mentioned earlier, microarray studies have gen-
different studies similar gene changes have been erated a wealth of information but to understand the
observed in the DRG from diverse peripheral nerve role of each gene product in pain will require exten-
injury models. A subset of these changes are given in sive follow-up studies. Another approach that could be
Table 1, which shows that in most cases the gene taken would be to evaluate the effects of administering
products were dysregulated in a similar direction. target specific agonists or antagonists in preclinical
However, in a couple of examples the transcripts pain models to determine if these proteins offer poten-
were altered differently between the different models tial opportunities for novel therapeutic intervention if
(Nell1; protein kinase C epsilon). The studies that such agents already exist. Gene silencing using anti-
have been performed have focused on gene changes sense or small interfering RNA (siRNA) approaches
that occur in the spinal cord and DRG but it is also would also be other ways to establish if a protein that
known that areas of the brain are important for pain was dysregulated in a pain state would be a useful pain
processing. Changes in gene expression can also target (Hemmings-Mieszczak, M. et al., 2003; Tan, P.
occur in brain areas such as the rostral ventral H. et al., 2005). Pain microarray studies as predicted
medulla and periaqueductal gray, which participate have deepened our understanding of key players in
in ascending and descending nociceptive signaling neuropathic pain. However, deciphering the precise
(Miki, K. et al., 2002) in chronic pain states. role of specific gene products in pain and identifica-
In addition to the strengths of microarray analysis tion of novel therapeutic targets awaits further
detailed above there are also limitations. These investigation and is eagerly anticipated.
 Assaying Pain-Related Genes: Preclinical and Clinical Correlates 171

14.7 The Genetics of Pain as low-pain sensitivity, average-pain sensitivity, and

high-pain sensitivity were identified, that encompass
The pain microarray gene chip analysis detailed ear- 96% of the examined genotypes and lead to altered
lier in this chapter provides a snapshot of the changes enzyme activity. The more active the COMT
in gene expression that occur in chronic pain states enzyme inversely correlated with pain susceptibility
with the ultimate goal of understanding pain mechan- and the lower risk of developing TMD (Diatchenko, L.
isms and identifying potential novel therapeutic et al., 2005).
targets for pain treatment. Additional efforts to iden- Another interesting aspect of human genetics
tify human genes associated with pain have examined examined the sex differences in pain and analgesic
congenital insensitivity to pain (CIP) that has been sensitivity has been an area of research focus
linked to the gene encoding serine palmitoyl-transfer- and increasing amounts of evidence suggest that sex
ase (Nagasako, E. M. et al., 2003). A subtype of this differences may reflect the activation of gender-
disorder CIP IV has been linked to single changes in specific neural mechanisms. Since clinically -opioid
nucleotide sequence (single nucleotide polymorph- receptor agonists show more efficacy in women than
isms (SNPs)) of the TrkA receptor rendering the men, a recent report investigated the possible sex
protein nonfunctional (Indo, Y. et al., 1996). While specificity of the genetic mediation of -opioid
nerve growth factor binding to the TrkA receptor is analgesia in mice using quantitative trait locus
known to contribute to the survival and regulation of (QTL) mapping (Mogil, J. S. et al., 2003). QTL map-
small diameter neurons, loss-of-function phenotypes ping is a technique in which the co-inheritance of
observed in these individuals with SNPs are similar to continuous traits and polymorphic DNA markers
(e.g., microsatellites or SNPs) can be used to broadly
the knockout animals. Another example is the gene
identify the chromosomal location of genes respon-
responsible for familial hemiplegic migraine that has
sible for trait variability (Lander, E. S. and Schork, N.
been linked to human chromosome 19p13 and more
J., 1994). Three convergent lines of evidence point to
recently identified as the gene encoding a calcium
the mouse melanocortin-1 (MCR1) gene as the
channel -subunit (CACNA1A; Joutel, A. et al.,
female-specific QTL. Furthermore analysis of
1993). Specific patients suffering from migraines have
women with red hair and fair skin containing two
been shown to have missense mutations in this gene
variant MCR1 alleles displayed significantly greater
that lead to gain-of-function phenotypes. This calcium
analgesia from the -opioid pentazocine than all
channel has been shown to be associated with seroto- other groups (Mogil, J. S. et al., 2003).
nin release, magnesium levels, and the phenomenon of With the completion of the human genome pro-
cortical spreading depression, each of which contri- ject and ongoing efforts to sequence the genomes of
bute to the pathophysiology of migraine (Ophoff, R. A. a variety of other species, a plethora of sequence
et al., 1996). information is now available. New microarray appli-
Differences in human pain perception have also cations and chip platforms are being developed
been associated to a common SNP (V158M) in the to pinpoint genetic variation in disease states,
gene of the enzyme catechol-o-methyltransferase isolate biomarkers that may indicate a predisposition
(COMT). Individuals homozygous for the Met158 for a given disease and identify treatment strategies
allele of COMT showed diminished regional and/or drug candidates that would have maximal
-opioid responses to pain compared with heterozy- benefit for a patient population with a specific
gotes, which was opposite to the effects observed with genetic make-up.
the Val158 homozygote (Zubieta, J. K. et al., 2003). The first of these new array platforms are com-
Individuals with the more active form of the enzyme parative genomic hybridization (CGH) arrays. These
(Val158) are less susceptible to pain than people with microarrays are designed across large areas of
Met158 at the same position. COMT activity is also sequence (i.e., a single chromosome) and even an
associated with a pathological pain condition known entire genome to provide broad coverage for geno-
as temporomandibular joint disorder (TMD) that typing. CGH arrays have been generated from
impacts 515% of the adult population. The relation- bacterial artificial chromosome (BAC) clones con-
ship between COMT polymorphism, pain sensitivity, taining large genomic inserts and require a
and risk of TMD development has recently been reference genome for comparison to the sample gen-
reported (Diatchenko, L. et al., 2005). Three genetic ome (Herr, A. et al., 2005). These arrays have been
variants (haplotypes) of the COMT gene, designated used to determine differences in copy number
172 Assaying Pain-Related Genes: Preclinical and Clinical Correlates

of particular genes and identify chromosomal imbal- without the proper analysis tools. With the imple-
ances found in cancer, genetic disorders, and deletion mentation of these new array platforms, software has
mutants with unique phenotypes used for scientific also been designed to aid in genotyping and linkage
research (Hughes, T. R. et al., 2000; Bruder, C. E. G. mapping studies. In the future, microarray may dee-
et al., 2001; Herr, A. et al., 2005). In contrast, to CGH pen our understanding of the genomic landscape and
arrays, resequencing and genome tiling arrays are perhaps shed light on the origins of disease states.
designed to smaller, specific regions of a chromosome This genetic-driven trend could lead to individua-
or genomic sequence and have been used to detect lized medical diagnoses and therapeutic treatment
unknown mutations or to narrow down the area strategies.
where a mutation may occur (Chee, M. et al., 1996;
Galvin, P. et al., 2004; Lin, S. M. et al., 2006).
Resequencing arrays have probes designed to a par-
ticular sequence which when combined together will References
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section of sequence with each probe representing Segraves, R., Hamilton, G., Zhang, X. X., Evans, D. G.,
Wallace, A. J., Baser, M. E., Zucman-Rossi, J.,
one of the four nucleotides at a single nucleotide
Hergersberg, M., Boltshauser, E., Papi, L., Rouleau, G. A.,
position. The probes are designed in sets of four. At Poptodorov, G., Jordanova, A., Rask-Andersen, H.,
a certain nucleotide position in the middle of the Kluwe, L., Mautner, V., Sainio, M., Hung, G., Mathiesen, T.,
Moller, C., Pulst, S. M., Harder, H., Heiberg, A., Honda, M.,
probe, each of the four probes will represent a differ- Niimura, M., Sahlen, S., Blennow, E., Albertson, D. G.,
ent nucleotide (A, C, G, or T). The next four Pinkel, D., and Dumanski, J. P. 2001. High resolution
overlapping probes will have a mismatch (A, C, G, deletion analysis of constitutional DNA from
neurofibromatosis type 2 (NF2) patients using microarray-
or T) at the next nucleotide position in succession
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from the mismatch in the previous probe set. The Carrion, A. M., Mellstrom, B., and Naranjo, J. R. 1998. Protein
overall result from both these array platforms is a lack kinase A-dependent derepression of the human
prodynorphin gene via differential binding to an intragenic
of hybridization and a subsequent decrease in signal
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intensity for the probes surrounding a mutation or Chee, M., Yang, R., Hubbell, E., Berno, A., Huang, X. C.,
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Fodor, S. P. A. 1996. Accessing genetic information with
Linkage analysis studies using restriction-frag- high-density DNA arrays. Science 274, 610614.
ment length polymorphisms (RFLP) and Chen, L. and Huang, L. Y. 1992. Protein kinase C reduces Mg2
polymorphic microsatellites have been useful in block of NMDA-receptor channels as a mechanism of
modulation. Nature 356, 521523.
understanding mendelian-linked diseases. However,
Cheng, H. Y., Pitcher, G. M., Laviolette, S. R., Whishaw, I. Q.,
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more complex disorders (i.e., schizophrenia, manic Crackower, M., Goncalves, J., Sarosi, I., Woodgett, J. R.,
Oliveira-dos-Santos, A. J., Ikura, M., van der Kooy, D.,
depression, and multiple sclerosis) that may have Salter, M. W., and Penninger, J. M. 2002. DREAM is a critical
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inheritance (Risch, N. J., 2000). More recently, Costigan, M., Befort, K., Karchewski, L., Griffin, R. S.,
DUrso, D., Allchorne, A., Sitarski, J., Mannion, J. W.,
SNPs, single nucleotide changes in DNA sequence, Pratt, R. E., and Woolf, C. J. 2002. Replicate high-density rat
have been identified and catalogued in a number of genome oligonucleotide microarrays reveal hundreds of
genomic databases. Several SNP microarray chips regulated genes in the dorsal root ganglion after peripheral
nerve injury. BMC Neurosci. 3, 16.
have been developed (e.g., Affymetrix GeneChip
Davis-Taber, R., Choi, W., Kroeger, P., Seiler, F., Gagne, G.,
Human Mapping 10K array) and used in concert Kage, K., Faltynek, C., Gopalakrishnan, M., and Scott, V.
with high-density microsatellite linkage sets to iden- 2003. Differential gene expression in small and large
diameter DRG neurons isolated by laser capture
tify susceptibility or candidate genes that may be microdissection. Am. Pain Soc. 2003; poster .
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(Riley, J. H. et al., 2000; Risch, N. J., 2000; Roses, A. D., Gagne, G., Kage, K., Kroeger, K., Honore, P., Fagerland, J.,
Faltynek, C., Gopalakrishnan, M., Surowy, C., and
2000; Sawcer, S. J. et al., 2004; Schaid, D. J. et al., 2004). Scott, V. E. 2004. Transcriptional profiling in small and large
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 15 Evolutionary Aspects of Pain
E T Walters, University of Texas at Houston, Medical School, Houston, TX, USA
2009 Elsevier Inc. All rights reserved.

15.1 Introduction 176

15.2 Evolution of Nociceptive Mechanisms 176
15.2.1 Nociceptors 176
15.2.1.1 Identification of nociceptors in diverse phyla 176
15.2.1.2 Conserved mechanisms of nociceptive transduction 177
15.2.1.3 Nociceptor-specific gene expression 178
15.2.2 Nociceptive Networks and Antinociceptive Modulation 178
15.2.2.1 Common nociceptive functions mediated by diverse neural circuits 178
15.2.2.2 Widespread occurrence of antinociceptive mechanisms 179
15.2.2.3 Recent evolution of opiate systems 179
15.2.3 Nociceptive Plasticity and Sensitization 179
15.2.3.1 N-Methyl-D-asparate-receptor-dependent long-term synaptic potentiation 180
15.2.3.2 Activation of the transcription factor, Ca2 and cAMP Response Element-Binding
Protein 180
15.2.3.3 Local protein synthesis and primitive memory mechanisms 180
15.3 Evolution of Emotional Components of Pain 181
15.3.1 Emotional Awareness is not Required for Nociceptive Responses 181
15.3.2 Even Snails Display Complex Motivational Effects after Noxious Stimulation 181
15.3.3 Cognitive and Emotional Responses to Noxious Stimulation Differ across Species 181
15.4 Conclusions 182
References 182

Glossary
CREB Ca2 and cAMP response element-binding NMDA-receptor-dependent LTP N-Methyl-D
protein, a transcription factor that binds to specific asparate-receptor-dependent long-term synaptic
enhancer elements in DNA and enhances the potentiation, a form of use-dependent
expression of corresponding genes when certain synaptic plasticity induced by Ca2 influx through
Ca2- or cAMP-activated protein kinases, such as the class of glutamate receptors that is
CaMK or PKA, phosphorylate the CREB protein. selectively activated (pharmacologically) by the
degenerin/epithelial Na (DEG/ENaC) drug, NMDA.
channels A family of ion channels that conduct RFamide peptides A family of peptide neuro-
Na ions and can be constitutively active or gated transmitters sharing the same C terminal RFamide
by mechanical distention or protons. Channels that (arginine-phenylalanine-NH2) sequence.
are gated by protons are called acid-sensing ion transient receptor potential (TRP) A family of ion
channels (ASICs). channels, many of which conduct cations nonse-
MAPK (ERK) Mitogen-activated protein kinase, a lectively. Different channels in this family are
family of protein kinases that includes the extra- opened by various inputs, including heat, cooling,
cellular signal-regulated kinase (ERK). ERK has protons, and intracellular signals.
many functions related to neural plasticity, both in TTX-resistant, voltage-gated Na
the cytoplasm and nucleus. channels Members of the voltage-gated Na
MRG Mas-related genes, a family of genes channel family that are relatively insensitive to
encoding G-protein-coupled receptors, some of blockade by the specific antagonist, tetrodotoxin
which are expressed selectively in mammalian (TTX). These channels (NaV1.8 and NaV1.9) are
nociceptors. Also called sensory neuron-specific expressed selectively in nociceptive dorsal root
G-protein-coupled receptors (SNSRs). ganglion neurons.

175
176 Evolutionary Aspects of Pain

15.1 Introduction (2) the conservation of homologous mechanisms des-

cending from common ancestors. Figure 1 shows a
Pain, according to a widely accepted definition simplified view of evolutionary relationships among
(Merskey, H. and Bogduk, N., 1994), is an unpleasant selected species mentioned in this essay that have
sensory and emotional experience associated with been used to study nociceptive biology. Those that
actual or potential tissue damage. This definition is have had their genome completely sequenced are
intended to capture essential features of human indicated by asterisks (human, mouse, fruit fly, and
experiences linked to tissue damage and makes no roundworm). The current explosion of molecular
claims about the evolutionary roots or possible bio- genetic information from a growing number of
logical functions of pain. Nonetheless, it raises two species promises to illuminate evolutionary relation-
distinct sets of evolutionary questions. First, how did ships among nociceptive mechanisms with a level of
sensory responses to actual or potential tissue detail and precision that could only be dreamed of a
damage evolve? What biological adaptations do few years ago.
these responses represent, and to what extent are
the underlying mechanisms conserved across the 15.2.1 Nociceptors
animal kingdom? Second, how did the unpleasant
emotional components of pain evolve? This essay Nociceptors are defined as sensory neurons that are
considers comparative functional and molecular activated preferentially by tissue-damaging stimuli,
findings from diverse species in the animal kingdom either immediately or when a noxious stimulus is
that begin to address these questions. Because this sufficiently prolonged. Because of the medical,
task requires reference to far more reports than can social, and economic importance of pain, most
be cited in a brief account, citations are made pri- studies of nociceptors have been in selected mam-
marily to review articles or to recent reports that malian species rather than animals more distantly
provide additional references to this diverse litera- related to humans. Thus, the comparative physiolo-
ture. Although no recent, comprehensive review of gical and molecular information needed to discern
nociceptive biology in invertebrates exists, highly evolutionary patterns in nociceptive function is still
comprehensive reviews of nociceptive mechanisms quite limited. However, the availability of extensive
in mammals can be found in other chapters in this genomic and proteomic data from a growing
volume, and in a recent book (Willis, W. D. and number of species should soon shed light on the
Coggeshall, R. E., 2004). evolution of specific molecular components of
nociceptive systems.

15.2.1.1 Identification of nociceptors

15.2 Evolution of Nociceptive in diverse phyla
Mechanisms Physiological identification and characterization of
nociceptors is challenging because the somata are
Because an animals reproductive success is clearly usually dispersed among those of many other types
enhanced if it can detect and avoid pervasive threats of neurons, and the fine peripheral and central fibers
to its structural integrity that otherwise would impair are essentially inaccessible for direct intracellular
or end its ability to reproduce, selection pressures for examination in living preparations. Nevertheless,
mechanisms that recognize noxious stimuli and trig- nociceptors have now been demonstrated convin-
ger adaptive physiological and behavioral responses cingly in representatives of all the major animal
must be very strong (Walters, E. T., 1994). Indeed, phyla (Figure 1), including nematode worms
animals in several phyla have now been shown to (Tobin, D. M. and Bargmann, C. I., 2004), leeches
possess nociceptors connected to neural networks (Nicholls, J. G. and Baylor, D. A., 1968), insects
controlling defensive responses, and these varied (Tracey, W. D. Jr., et al., 2003), mollusks (Illich, P. A.
nociceptive systems share numerous features includ- and Walters, E. T., 1997), and various chordates,
ing distinctive forms of plasticity. Common including fish (Sneddon, L. U. et al., 2003), amphibians
properties in diverse species are of interest for evolu- (Lynn, B. and OShea, N. R., 1998), reptiles (Liang, Y.
tionary questions because they point to (1) common F. et al., 1995), birds (Holloway, J. A. et al., 1980), and
selection pressures that have shaped similar but inde- mammals (Light, A. R., 1992). Additional species,
pendently derived (analogous) adaptations; and/or including snails (Malyshev, A. Y. and Balaban, P. M.,
 Evolutionary Aspects of Pain 177

Homo sapiens *
Phylogenetic relationships Mammalia
55 Ma
of a few species used
Mus musculus *
to investigate mechanisms
Amniota (mouse)
of nociceptive plasticity 300 Ma
Gallus domesticus
Tetrapoda 240 Ma (chicken)
Trimeresurus flavoviridis
320 Ma (Habu pit viper)
Gnathostomata Rana pipiens
400 Ma (leopard frog)
Chordata
Deuterostomia Oncorhynchus mykiss
560 Ma
520 Ma (rainbow trout)
Petromyzon marinus
600 Ma (sea lamprey)
Arthropoda
Drosophila melanogaster *
Bilateria
Ecdysozoa (fruit fly)
Nematoda
Caenorhabditis elegans *
(soil roundworm)
600 Ma
Animalia 700 Ma Annelida
Hirudo medicinalis
Lophotrochozoa (medicinal leech)
Molluska Aplysia californica
(sea hare)
Cnidaria
Scyphozoa Aglantha digitale
(jellyfish)

Figure 1 Greatly simplified phylogeny of a few selected species (right) that have been used to investigate nociceptive
biology. Numbers indicate approximate time of divergence (millions of years ago, Ma) of the indicated groups (Dawkins, R.,
2004). Groupings reflect recent molecular phylogenetics rather than traditional views based on morphological evidence. The
traditional phyla indicated on this chart are chordata, arthropoda, nematoda, annelida, molluska, and cnidaria. Because of the
paucity of fossil evidence prior to the Cambrian period (more than 540 Ma) and uncertainties in molecular clocks over longer
periods, the older times of divergence on the chart are much rougher estimates than the more recent ones. Asterisks indicate
species for which complete genome sequence information is currently available.

2002) and lampreys (Christenson, J. et al., 1988) have depolarizing current entry through channels in the
mechanosensory neurons that are likely to be noci- degenerin/epithelial Na channel (DEG/ENaC)
ceptive, despite their relatively low mechanical family. Several of these channels are required for acti-
thresholds, because the tissue they innervate is deli- vation of low-threshold mechanosensory neurons in the
cate enough to be damaged by modest pressures (see nematode, C. elegans (Tobin, D. M. and Bargmann, C. I.,
Illich, P. A. and Walters, E. T., 1997). 2004), while in mice some channel subunits within this
same family are acid sensitive (DRASIC/ASIC3) and
15.2.1.2 Conserved mechanisms of contribute to the responses of some nociceptive
nociceptive transduction neurons both to acid and to noxious pinch
Tissue damage can be produced by various mechanical (Price, M. P. et al., 2001). Another family of channels,
forces and chemical agents as well as by excessive the transient receptor potential (TRP) superfamily, has
temperature changes, so one would expect that demonstrated its importance for nociceptive transduc-
nociceptors utilize diverse stimulus transduction tion in both vertebrates and invertebrates. TRPV
mechanisms. Combined molecular and physiological channels in C. elegans are required for nociceptive,
analyses of nociceptive transduction have revealed osmosensory, mechanosensory, and some chemosensory
important mechanisms in a few species. Many forms responses (Tobin, D. M. and Bargmann, C. I., 2004). In
of mechanotransduction, perhaps including the trans- mammals, TRPV1 channels in sensory neurons are
duction of noxious forces in some cells, depend upon activated by heat, acid, capsaicin (the pungent vanilloid
178 Evolutionary Aspects of Pain

ingredient of chili peppers), and other chemicals species) with mice MRG genes. Little is known
(Krause, J. E. et al., 2005). Mice lacking TRPV1 channels about the functions of these receptors, although the
show impaired sensory and behavioral responses to heat, diversity of ligands implicated for them include a
vanilloids, and inflammation (Caterina, M. J. et al., 2000). variety of RFamide peptides thought to be antinocicep-
TRP channels also transduce noxious stimuli in the fly, tive (Dong, X. et al., 2001 and see below). The recency
Drosophila; mutations of the painless gene (which and rapidity of change in the MRG family suggests
encodes a TRPA family member) nearly eliminate that strong selection pressures may be acting on
nociceptive behavioral responses of larvae to noxious antinociceptive modulation of nociceptors in verte-
heat or mechanical stimuli (Tracey, W. D., Jr., et al., brates, including primates (Choi, S. S. and Lahn, B. T.,
2003). TRPA family channels are found in vertebrate 2003; Zylka, M. J. et al., 2003; Zhang, L. et al., 2005).
nociceptors as well, where they are activated by noxious Further investigation of the distribution across
cold, various pungent compounds, and bradykinin diverse taxa of genes found to be preferentially
(Bandell, M. et al., 2004). These recent results show expressed in mammalian nociceptors should provide
that nociceptive transduction in at least three different interesting insights into the evolution of nociceptive
phyla makes use of shared families of ion channels, systems.
notably TRPV and TRPA channels, and perhaps
DEG/ENaC channels. However, these channel families
15.2.2 Nociceptive Networks and
are not specific to nociception; each family also has
Antinociceptive Modulation
members that are involved in innocuous mechanosensa-
tion (including hearing in Drosophila) and various forms The capability for rapid detection of noxious stimuli
of chemosensation (Tobin, D. M. and Bargmann, C. I., could only have been selected during evolution if
2004). Thus, nociception has exploited a number of systems to translate this information into adaptive
ancient and versatile ion channel families that have behavior had also been selected. Almost every animal
also been used during evolution for additional forms of that has been examined, from jellyfish to humans,
sensory transduction. displays withdrawal and/or escape responses to nox-
ious stimulation (Walters, E. T., 1994), although
15.2.1.3 Nociceptor-specific gene these responses are inhibited under some circum-
expression stances (see further ahead).
Although evolution is opportunistic and tends to find
multiple uses for individual gene products, some 15.2.2.1 Common nociceptive functions
remain functionally specialized. Several genes have mediated by diverse neural circuits
been found to be expressed relatively selectively in Nociceptive responses are mediated by neural cir-
mammalian nociceptors, suggesting specialized func- cuits that range enormously in complexity across
tions related to nociceptive processing. These include different animals. In cnidarians (hydra, jellyfish, sea
the TTX-resistant, voltage-gated Na channels, anemones), some simple withdrawal reflexes may be
NaV1.8 and NaV1.9, which show alterations after mediated in part by just a two-cell pathway involving
nerve injury that are likely to contribute to neuropathic a multifunction sensory-motor neuron synaptically
pain (Lai, J. et al., 2004). They also include a family of connected to epithelial-muscular cells. However,
receptors called Mas-related genes (MRG) (Dong, X. even in cnidarians, which lack a brain or central
et al., 2001) or sensory neuron-specific G-protein- ganglia, relatively complex withdrawal and escape
coupled receptors (SNSR) (Lembo, P. M. et al., 2002). responses can be mediated by a neural net containing
These sensory neuron receptors have displayed sensory, motor, and interneurons (e.g., Mackie, G. O.,
exceptionally rapid recent evolution in some mammals 2004). At the other end of the behavioral spectrum,
(Choi, S. S. and Lahn, B. T., 2003), and might have nociceptive circuits in mammals involve not only
first appeared in the chordate lineage. Rapid changes diverse types of sensory, motor, and interneurons at
within the MRG family are illustrated dramatically the spinal level, but vast numbers of neurons in
by the variation in nociceptor-specific MRG genes higher brain structures (e.g., Craig, A. D., 2003).
within a single mammalian order; mice have 22 Different populations of neurons process nociceptive
MRGA and 14 MRGC genes, whereas gerbils, and sensory information, generate defensive motor pat-
rats only have one of each (Zylka, M. J. et al., 2003). terns, and modulate nociceptive responses depending
Only six of the 13 MRG genes in humans are clearly upon the state of the organism and its previous
orthologous (displaying sequence homology across experience. These basic nociceptive/defensive tasks
 Evolutionary Aspects of Pain 179

are also mediated by functionally similar, but much and immune systems of diverse species, including
simpler, neural circuits in several invertebrate spe- invertebrates and even unicellular organisms
cies that have been investigated, including C. elegans (Stefano, G. B. et al., 2003). However, genes within
(Wittenburg, N. and Baumeister, R., 1999), the med- the opioid/orphanin family have not been detected
icinal leech (Shaw, B. K. and Kristan, W. B., Jr. , 1995), in the genomes of C. elegans or Drosophila (nor have
and several gastropod mollusks Aplysia (Cleary, L. J. genes within the corresponding melanocortin recep-
et al., 1995), Tritonia (Getting, P. A., 1977), and Helix tor family), which supports the hypothesis that this
(Balaban, P. M., 2002). Although the organization of gene family first appeared in a deuterostome ancestor
these circuits differs greatly among the mammalian around the time that the chordate lineage diverged
and invertebrate species examined, common (Dores, R. M. et al., 2002) (Figure 1). The only inver-
mechanisms of nociceptive plasticity appear to be tebrate opioids that have been sequenced are proteins
shared by these diverse animals (see further ahead). related to pro-opiomelanocortin (POMC), which have
been found in immunocytes in a leech (Salzet, M.
15.2.2.2 Widespread occurrence of et al., 1997) and in the mussel, Mytilus edulis (Stefano,
antinociceptive mechanisms G. B. et al., 1999). Paradoxical features of these POMC
One of the most important functions of nociceptive sequences led to the suggestion that the vertebrate-like
circuits is to modulate nociceptive responses, so that POMC in these invertebrates may have come somehow
the threshold, selection, and intensity of defensive by cross-phylum gene transfer (Dores, R. M. et al.,
responses are well matched to the nature and context 2002), and these unusual features reinforce indications
of a noxious threat. The form of modulation that has that antinociceptive opioids may be a chordate innova-
received the most experimental attention in nocicep- tion. In contrast, another neuropeptide family having
tive systems is antinociception, which produces antinociceptive functions is widespread in the animal
analgesia or hypoalgesia in humans. Many authors kingdom: the RFamide family which is expressed in
have argued that it is adaptive for an animal to inhibit many phyla, including cnidarians, nematodes, arthro-
responses, including pain and recuperative responses pods, mollusks, and chordates (Li, C. et al., 1999;
evoked by injury, during active defense in life-threa- Nichols, R., 2003). In mammals some RFamides acti-
tening encounters with predators or aggressive vate MRG receptors (Dong, X. et al., 2001) (and see
conspecifics (see Walters, E. T., 1994). Indeed, most above), while in Aplysia FMRFamide can produce both
animals examined thus far, including various inverte- short-term and long-term inhibition of synaptic trans-
brates, transiently suppress appetitive, recuperative, mission from nociceptive sensory neurons (Montarolo,
and other nondefensive behaviors that might com- P. G. et al., 1988). However, RFamides also have pro-
pete with urgent fight-or-flight responses evoked nociceptive actions (Yudin, Y. K. et al., 2004) as well as
immediately by noxious stimuli. In mammals many actions unrelated to nociceptive modulation.
(Light, A. R., 1992) and at least one invertebrate,
Aplysia (Cleary, L. J. et al., 1995), inhibitory effects of
15.2.3 Nociceptive Plasticity and
noxious stimulation are exerted at multiple stages
Sensitization
within sensory-motor pathways. Strong pre- and/or
postsynaptic inhibition during noxious stimulation As just discussed, many animals show a transient
occurs at the first synapse in these pathways (from reduction in responsiveness to somatic stimuli during
somatic primary afferent neurons) in various species, active defensive behavior. However, if an animal is
including mammals (Light, A. R., 1992) and Aplysia injured and escapes, it often displays enhanced
(Mackey, S. L. et al., 1987). responsiveness to somatosensory stimuli while it
recuperates. This nociceptive sensitization (and
15.2.2.3 Recent evolution of opiate hyperalgesia in some species) is targeted to the
systems injured region and can be very long lasting. It facil-
There has been much interest in the phylogenetic itates defensive responses, helping to protect an
distribution of antinociceptive neuropeptides, injured animal during a period when it may be wea-
and especially the opioid family because of its kened and emitting wound-related signals that can
well-known analgesic functions in humans. attract the attention of predators, parasites, and
Pharmacological, immunocytochemical, and bio- aggressive competitors (Walters, E. T., 1994). The
chemical studies have suggested that opiate-like persistence of nociceptive sensitization during recup-
substances and opiate receptors occur in nervous eration periods that can last weeks or longer raises
180 Evolutionary Aspects of Pain

the possibility that nociceptive sensitization may (Silva, A. J. et al., 1998; Kandel, E. R., 2001). Later,
utilize mechanisms that are shared with other long- hyperalgesia was found to be accompanied by CREB
lasting neural alterations, such as learning and phosphorylation in the rat spinal cord (Ji, R. R. and
memory. Demonstrations that the same plasticity Rupp, F., 1997), and interference with CREB binding
mechanisms contribute both to nociceptive sensitiza- to DNA was shown to attenuate sensitized behavioral
tion and to learning and memory, and evidence responses in a rat neuropathic pain model (Ma, W.
that these mechanisms have common functions et al., 2003). Interestingly, the sensory neurons
in very distantly related animal species, indicate from Aplysia originally used to demonstrate possible
that evolutionary links between injury and memory roles of CREB in long-term memory formation
may be quite primitive (Walters, E. T., 1994; (Kandel, E. R., 2001) are nociceptive and exhibit
Weragoda, R. M. et al., 2004). Of several apparent long-term hyperexcitability following noxious cuta-
commonalities between injury-induced and learn- neous stimulation, and this hyperexcitability is
ing-related mechanisms in diverse phyla, three will blocked by injection of CRE oligonucleotides to pre-
be considered here. vent CREB binding (Lewin, M. R. and Walters, E. T.,
1999). Cellular signals involved in activating
15.2.3.1 N-Methyl- D -asparate- CREB are also common to both memory and noci-
receptor-dependent long-term synaptic ceptive sensitization/hyperalgesia in various phyla.
potentiation Particular attention has been paid to the roles of PKA
As in mammals ( Ji, R. R. et al., 2003), somatosensory and MAPK (ERK) in CREB-related plasticity in
afferents in several intensively studied invertebrate mammalian hippocampus and forebrain (Waltereit, R.
species appear to release glutamate, which acts upon and Weller, M., 2003), mammalian spinal cord
both NMDA and non-NMDA postsynaptic receptors. (Kawasaki, Y. et al., 2004), and Aplysia sensory neurons
These invertebrates include C. elegans (Brockie, P. J. (Kandel, E. R., 2001).
et al., 2001), Aplysia (Dale, N. and Kandel, E. R., 1993),
Helix (another snail) (Bravarenko, N. I. et al., 2003), and 15.2.3.3 Local protein synthesis and
the leech (Burrell, B. D. and Sahley, C. L., 2004). In primitive memory mechanisms
mammals (Ji, R. R. et al., 2003), Aplysia (Lin, X. Y. The observations mentioned above suggest that
and Glanzman, D. L., 1994), Helix (Nikitin, V. P. evolution made use of common basic mechanisms,
et al., 2002), and the leech (Burrell, B. D. and including NMDA-receptor-dependent LTP and
Sahley, C. L., 2004), intense activation of a sensory CREB-dependent transcriptional regulation, to
pathway (which under normal conditions would occur induce long-lasting neuronal alterations underlying
during noxious stimulation of the periphery) strongly quite different functions: nociceptive sensitization
depolarizes second-order neurons and activates and learning. Because strong selection pressures
NMDA receptors, leading to LTP in the nociceptive would have favored adaptive responses to peripheral
pathway, which contributes to nociceptive sensitiza- injury at the earliest stages of animal evolution (prior
tion. As in mammals (Martin, S. J. et al., 2000), in to the evolution of brains complex enough for asso-
Aplysia (Murphy, G. G. and Glanzman, D. L., 1997), ciative learning), it has been suggested that adaptive
Helix (Nikitin, V. P. et al., 2002), and the leech responses to peripheral injury, including nociceptive
(Burrell, B. D. and Sahley, C. L., 2004), NMDA- sensitization, emerged first as an adaptation to tissue
receptor-dependent LTP has been implicated as a damage and later evolved into mechanisms of learn-
mechanism of learning and memory. ing and memory (Walters, E. T., 1994). In ancient,
soft-bodied animals that were ancestral to most
15.2.3.2 Activation of the transcription contemporary animals (Figure 1), some of these
factor, Ca 2 and cAMP Response Element- mechanisms may have functioned initially at sites of
Binding Protein peripheral injury in axons and terminals. A vestige of
A second commonality is a role for a transcription early memory-like mechanisms linked to peripheral
factor, the Ca2 and cAMP response element-bind- injury might be represented today by localized, long-
ing protein (CREB), in inducing long-term neuronal term hyperexcitability of peripheral axon segments
alterations that require changes in gene transcription of nociceptive sensory neurons found in Aplysia
and protein synthesis. A key role for this transcription (Weragoda, R. M. et al., 2004). This sensitizing axonal
factor was first implicated in long-term memory alteration is induced by nerve crush or local depolar-
formation in Aplysia, Drosophila, mice, and rats ization and, like the late phase of LTP in mammalian
 Evolutionary Aspects of Pain 181

hippocampus (Tang, S. J. et al., 2002), it requires local, responses, and these too can be observed throughout
rapamycin-sensitive protein synthesis (Weragoda, R. M. the animal kingdom. Pain motivates humans and other
et al., 2004). It has not yet been determined whether mammals to avoid contexts or actions associated with
this apparently primitive mechanism contributes to the onset of the pain, and similar motivational effects
neuropathic pain in mammals, but evidence for intra- of noxious stimulation (often electric shock that acti-
axonal protein synthesis following peripheral nerve vates nociceptors) have been used experimentally to
injury has been found in axons of mammalian DRG produce aversive learning in a wide variety of inverte-
neurons (e.g. Zheng, J. Q. et al., 2001). brates, including flies, worms, and snails (reviewed by
Krasne, F. B. and Glanzman, D. L., 1995; Waddell, S.
and Quinn, W. G., 2001; Rankin, C. H., 2004). Some of
15.3 Evolution of Emotional these motivational effects appear remarkably similar to
Components of Pain those produced by noxious stimuli in mammals. For
example, pairing an odor with strong cutaneous shock
Perhaps the most commonly asked question related causes the previously neutral odor to evoke a condi-
to the evolution of pain is one that has yet to be tioned fear-like state in Aplysia, so that the odor comes
answered by scientists: Which animals feel pain? to evoke a freezing response, inhibit appetitive
This question is critical for animal welfare issues, responses, and facilitate defensive responses to tactile
and is also of considerable philosophical interest stimuli (Walters, E. T. et al., 1981).
(Allen, C., 2004). Many philosophers of science
have argued that a human observer cannot know for
15.3.3 Cognitive and Emotional
sure whether another being or species has subjective,
Responses to Noxious Stimulation Differ
conscious experiences such as pain. The existence of
across Species
emotional pain in other species may defy proof, but
evolutionary and neurobiological considerations Although relatively simple invertebrates show func-
indicate strongly that these components of pain did tional similarities to vertebrates (sometimes with
not emerge de novo in humans, while also suggesting conserved molecular mechanisms) in detecting
that the capacity to experience subjective pain is not noxious stimuli, activating defensive behavior, and
a universal animal trait. motivating learned and unlearned responses to a
noxious event, vast differences in the size and
structure of their brains indicate that major differ-
15.3.1 Emotional Awareness is not
ences exist between most invertebrates and many
Required for Nociceptive Responses
vertebrates in the cognitive and possible emotional
As discussed above, most animals have nociceptive processing of information about tissue damage. For
systems that detect noxious stimuli and activate example, in Aplysia, the body surface is represented
defensive responses. Nociceptive reflexes and even by a somatotopic map formed by nociceptor somata
nociceptive plasticity, however, can almost certainly in central ganglia (Walters, E. T. et al., 2004), and
operate without conscious, emotional experience peripheral injury causes nociceptor soma hyperex-
because these responses are expressed not only in citability selectively in nociceptors representing the
the simplest animals but also in greatly simplified injured region. These and other sensory changes
physiological preparations, such as a spinalized rat are sufficient to alter reflexive responses to stimuli
or an Aplysia ganglion connected to part of the ani- received by the injured region, but they do not cause
mals tail. Moreover, in human patients nociceptive these normally silent sensory neurons to become
reflexes can be expressed without conscious aware- spontaneously active. If one assumes that ongoing
ness below the level of a spinal transection. pain requires ongoing neural activity somewhere in
a nervous system, then an absence of ongoing activ-
ity may be taken as evidence that tissue damage
15.3.2 Even Snails Display Complex
fails to produce long-lasting, pain-like states. In
Motivational Effects after Noxious
relatively simple invertebrates, one can systemati-
Stimulation
cally sample the activity of individual neurons and
In addition to the abilities to detect noxious stimuli use optical imaging methods to sample the activity
and activate defensive responses, adaptations to actual of populations of neurons to test this possibility
or incipient tissue damage include motivational following strong noxious stimulation. Although
182 Evolutionary Aspects of Pain

many more neurons need to be sampled before firm 15.4 Conclusions

conclusions can be made, preliminary observations
in Aplysia suggest that a persistent (lasting days or Nociceptive mechanisms that permit an animal to
longer) increase in ongoing neural activity within detect, respond to, and remember peripheral injury
the CNS does not accompany long-term, nocicep- are found throughout the animal kingdom. Some of
tive sensitization of defensive reflexes. Instead, these are likely to represent convergent (analogous)
sensitization lasting days or weeks appears to be evolutionary responses to similar injury-related
maintained largely by synaptic facilitation and selection pressures in diverse organisms, which may
hyperexcitability of sensory neurons, which remain depend upon different molecular substrates in differ-
silent in the absence of peripheral stimulation. ent animal groups. For example, while most animals
An apparent lack of ongoing activity in the Aplysia show transient, antinociceptive responses to noxious
CNS would contrast with increases in neural activ- stimulation, the use of opiates and melanocortin
ity that probably occur in some regions of the receptors for antinociception may largely be
human brain during persistent pain states (Casey, restricted to vertebrates. On the other hand, emer-
K. L., 2000). ging molecular genetic information indicates that
Investigating the evolution of emotional aspects other nociceptive mechanisms have been highly
of pain is an enormous challenge because the biolo- conserved from mechanisms that must have already
gical functions of conscious pain (as opposed to been present around 600 million years ago in worm-
unconscious nociceptive responses) are not obvious like animals that were the last common ancestors of
and the neural substrates are very difficult to define. modern roundworms, flies, leeches, snails, mice, and
Nevertheless, there are many intriguing observa- humans (Figure 1). Conserved (homologous)
tions in these areas. For example, much research nociceptive mechanisms that have been revealed
has been stimulated by the suggestion that conscious thus far are related to nociceptive transduction (e.g.,
awareness of an animals own bodily states might TRP channels), plasticity of nociceptor synapses
help it predict the behavioral states (read the mind) (e.g., NMDA-receptor-dependent LTP), and regula-
tion of gene transcription in nociceptive pathways
of other animals (Keenan, J. et al., 2003). Conscious
(e.g., CREB activation). Because of dramatic
awareness of bodily states, including the emotional
increases in molecular genetic information from all
awareness of pain, involves dynamic neural repre-
the major phyla and the coupling of this information
sentations of the body (e.g. Craig, A. D., 2003). Even
with continuing physiological studies of various spe-
the most complex brains of invertebrates, such as the
cies, it seems likely that knowledge gained from
octopus or honeybee, as well as the brains of non-
investigating highly conserved mechanisms in dispa-
mammalian vertebrates, have much smaller
rate creatures will provide insights that might help
numbers of neurons and less obviously organized
control intransigent pain states in humans.
brain structures than found in behaviorally sophis- Comparative studies may also shed some light on
ticated mammals such as primates and cetaceans, the selection pressures that shaped the evolution of
and thus presumably less capacity for such emotional aspects of pain, even though the subjective
representations. Indeed, extremely complex neu- experience of pain may not be amenable to direct
roanatomical substrates support the multiple experimental investigation in other species.
representations and meta-representations of the
body that are associated with emotional awareness
and suffering in humans, which has suggested to
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 16 Redheads and Pain
J S Mogil, McGill University, Montreal, QC, Canada
2009 Elsevier Inc. All rights reserved.

16.1 Finding Genes Associated with Phenotypic Variability 185

16.2 MC1R and the Melanocortin-1 Receptor 185
16.3 MC1R and Analgesia 186
16.4 MC1R and Anesthesia 188
16.5 MC1R and Pain 188
16.6 Future Directions 190
References 190

Glossary
allele One possible form (or variant) of a phenotype An observable or measurable biologi-
gene, defined by its particular nucleotide cal trait.
sequence. pleiotropy The involvement of a single gene in
genotype The inherited alleles (one from each more than one phenotype
parent) at a genetic locus (e.g., a gene). polymorphism see Allele.

16.1 Finding Genes Associated with (transgenic knockout) mouse, and in others simply
Phenotypic Variability by the known involvement of the protein in pain
processing.
With recent developments in classical and molecu- There are a finite number of human genes, current
lar genetic techniques, identifying genes, and the estimates are less than 30 000, but a virtually unlim-
DNA sequence variants in or near them, responsi- ited number of biological phenotypes. It is thus
ble for disease susceptibility and normal variation necessarily true that every gene will eventually be
in biological phenotypes is becoming increasingly shown to be genetically associated with multiple (and
tractable. Two main methodologies are employed possibly unrelated) traits. This principle is known as
to this end: genetic linkage mapping and genetic pleiotropy. Currently, one of the best-known exam-
association studies (see Lander, E. S. and Schork, ples of a gene with pleiotropic effects is MC1R, the
N. J., 1994). These approaches have begun to be gene coding for the melanocortin-1 receptor
applied to the study of pain (see Mogil, J. S., 2004). (MC1R).
For example, variable sensitivity of humans to
experimental and clinical pain has been provision-
ally associated with the genes COMT (catechol-O- 16.2 MC1R and the Melanocortin-1
methyltransferase) (Zubieta, J. K. et al., 2003; Receptor
Diatchenko, L. et al., 2005), OPRD (-opioid recep-
tor; Kim, H. et al., 2004), TRPV1 (transient receptor The melanocortin receptor family consists of five
potential vanilloid type 1; Kim, H. et al., 2004), subtypes, MC1RMC5R. These receptors bind the
OPRM (-opioid receptor gene) (Klepstad, P. melanocortin peptides, including -melanocyte-
et al., 2004; Fillingim, R. B. et al., 2005), IL6 (inter- stimulating hormone (-MSH), -MSH, -MSH,
leukin-6; Noponen-Hietala, N. et al., 2005), IL1 and adrenocorticotrophic hormone (ACTH), all
(interleukin-1; Solovieva, S. et al., 2004), and others. cleaved from the proopiomelanocortin (POMC)
These association studies were inspired in some gene along with -endorphin (see Tatro, J. B., 1996;
cases by a mouse linkage mapping finding, in Wikberg, J. E. S., 1999; Abdel-Malek, Z. A., 2001;
others by the phenotype of a null mutant Starowicz, K. and Przewlocka, B., 2003). The

185
186 Redheads and Pain

receptors are transcribed from five separate genes, lentigines (i.e., freckles; Bastiaens, M. et al., 2001a),
and each receptor has differential tissue distributions reduced tanning ability (Healy, E. et al., 2000), and
and affinities for the melanocortins. All melanocortin skin cancer (malignant melanoma, squamous cell, and
receptor subtypes couple positively to adenylate basal cell carcinoma) susceptibility even in indivi-
cyclase when activated (Chhajlani, V. and Wikberg, duals without red hair (Palmer, J. S. et al., 2000;
J. E. S., 1992). The MC1R, previously known as the Bastiaens, M. et al., 2001b). It should be noted that
-MSH receptor, actually has equal affinity for MC1R genetics is rather complex, with a large num-
ACTH, and is found primarily in melanocytes of ber of known rare (<1% allele frequency) mutations
the skin. It is also been demonstrated in a variety of and at least three common variants (Val60Leu,
glial cells, the testes, placenta, pituitary (see Tatro, J. Val92Met, and Arg163Gln) that are only partially
B., 1996; Wikberg, J. E. S., 1999; Abdel-Malek, Z. A., penetrant or completely silent, producing no impair-
2001; Starowicz, K. and Przewlocka, B., 2003), and in ment of MC1R functioning (see Schaffer, J. V. and
neurons of the midbrain periaqueductal gray (Xia, Y. Bolognia, J. L., 2001).
et al., 1995). The role of -MSH acting on melano-
cyte MC1Rs to regulate pigmentation of skin
and hair by stimulation of eumelanin synthesis has 16.3 MC1R and Analgesia
been understood in great detail for some time (see
Eberle, A. N., 1988). Other -MSH effects are also Using genetic linkage (formally, quantitative trait
well known, and have been attributed to actions locus or QTL) mapping, a gene affecting the magni-
at the MC1R. For example, -MSH has potent tude of swim stress-induced analgesia (SIA) in female
anti-inflammatory and antipyretic activity (see but not male mice was localized to distal mouse chro-
Tatro, J. B., 1996), and may play a role in nerve mosome 8 (Mogil, J. S. et al., 1997). This SIA was
regeneration (see Starowicz, K. and Przewlocka, B., previously demonstrated to be sex-specific in its neu-
2003) and female sexual behavior (Pfaus, J. G. et al., rochemical mediation (e.g., Mogil, J. S. et al., 1993;
2004). Injection of -MSH (or ACTH) into the brain Kavaliers, M. and Galea, L. A. M., 1995). Specifically,
has been reported to affect nociceptive sensitivity pharmacological blockade of the N-methyl-D-aspar-
(see Starowicz, K. and Przewlocka, B., 2003), usually tate (NMDA) excitatory amino acid receptor
in a manner opposed to the actions of endogenous abolishes SIA in male rodents, but does not do so (at
opioids, although the literature is contradictory. It is any dose or in any estrous phase) in females. Kavaliers
possible that some or all of these effects are mediated at M. and Choleris E. (1997) found that analgesia from a
spinal MC4Rs, since the MC4R-selective antagonist, -opioid-selective agonist, NPC12626, was similarly
SHU9119, reduces neuropathic hypersensitivity states sexually dimorphic in its neurochemical mediation,
when injected into the spinal cord (Vrinten, D. H. being blocked by NMDA antagonists only in males.
et al., 2001). The possible sex-specificity of -opioid analgesia was
Given the known role of MC1Rs in pigmentation then a hot topic, as data from Gear R. W. et al. (1996)
and the highly polymorphic nature of MC1R, the suggested that -opioid-acting drugs may only pro-
hypothesis that the MC1R gene might be associated duce efficacious analgesia, in the third molar
with variability in human hair color and skin tone extraction model, in women. Mogil J. S. et al. (2003)
was palpable. Indeed, Valverde P. et al. (1995) demon- hypothesized that if SIA and -opioid analgesia were
strated that the inheritance of loss-of-function MC1R both sexually dimorphic, and a female-specific linkage
alleles was genetically associated with red hair and on chromosome 8 had been already demonstrated for
fair skin in humans. Subsequent work has determined SIA, one would likely be seen for -opioid analgesia as
that approximately 6085% of natural redheads are well. This was quickly confirmed using the -opioid-
so because of their inheritance of variant alleles of selective compound, U50,488 (Mogil, J. S. et al., 2003),
MC1R, especially those producing the very common but the confidence interval containing the gene(s)
Arg151Cys, Arg160Trp, and Asp294His amino acid responsible for the linkage was quite large. Of the
substitutions (see Schaffer, J. V. and Bolognia, J. L., 200 murine genes in this region, however, one
2001). Red hair is much more likely if multiple alleles emerged as a high-probability candidate: Mc1r.
are inherited, and the number of variant alleles is Evidence for its candidacy included the possible role
important in determining the precise shade of red. of -MSH and/or ACTH as antiopioid peptides (see
In addition, it has been subsequently demonstrated above) and the demonstration of neuronal MC1Rs
that MC1R is the major gene responsible for solar in the periaqueductal gray (Xia, Y. et al., 1995),
 Redheads and Pain 187

a supraspinal region of well-known importance in pain specific QTL for U50,488 analgesia. First, null mutant
modulation (see Basbaum, A. I. and Fields, H. L., recessive yellow (C57BL/6-Mc1r e/e, or e/e) mice,
1984). It should also be noted that dynorphin can which arose spontaneously rather than being engi-
bind with reasonable affinity in in vitro assays to neered using transgenic technology (see Cone, R. D.
MC1R (Quillan, J. M. and Sadee, W., 1997). et al., 1996), were tested for U50,488 analgesia along
Using a three-pronged strategy, Mogil J. S. and with their C57BL/6 wild-type controls. We found that
colleagues have provided considerable supporting evi- female but not male e/e mutants displayed signifi-
dence that the mouse Mc1r gene is indeed the female- cantly greater U50,488 analgesia (see Figure 1(a)),

(a) Male Female

80 80
B6 B6 *
e /e e /e
60 60
% Analgesia

% Analgesia
40 40
*

20 20

0 0
10 30 50 70 10 30 50 70
U50,488 dose (mg kg1) U50,488 dose (mg kg1)

(b) (c)
40 120
*
* 100
Total rating change

30
% Analgesia

80

20 60

40
10
20

0 0
Male Female Male Female
U50 + Saline 0 or 1 variant
2+ variants
U50 + Antagonist
Figure 1 Female-specific effect of MC1R functional status on -opioid analgesia in mice and humans. (a) Response on the
49  C tail-withdrawal test to systemic injection of various doses (1070 mg kg1, subcutaneous (s.c.)) of the selective -opioid
agonist, U50,488, in male and female spontaneous mutant recessive yellow (e/e) mice and their wild-type counterparts,
C57BL/6 (B6). The e/e mutants of both sexes have nonfunctional MC1Rs due to a frameshift mutation at position 183.
Symbols represent mean standard error of the mean (SEM) percent of total analgesia (based on area under the timeeffect
curve) over a 60 min testing period postdrug administration. (b) Female-specific potentiation of U50,488 (U50) analgesia
(70 mg kg1, s.c.), on the 49  C tail-withdrawal test, by injection of a peptidergic MC1R antagonist, Ac-Nle-Asp-Trp-DPhe-
Nle-Trp-Lys-NH2 (20 mg/mouse in 2.5 ml), into the lateral ventricles of outbred mice. Bars represent mean SEM percent of
total analgesia (based on area under the timeeffect curve) over a 60-min testing period postdrug administration. (c) Female-
specific influence of MC1R genotype on pentazocine (0.5 mg kg1, intravenous) analgesia in humans. Subjects of both sexes
were studied in two experimental sessions as described in detail in Mogil J. S. et al. (2003). Bars represent mean SEM
difference scores for summed pain intensity ratings (0100; 5 ) during the ischemic pain procedure, computed by
subtracting postdrug from predrug ratings, in subjects with consensus sequence or one MC1R variant (0 or 1 variant) and
subjects with two or more MC1R variants (2 variants). The 2 variant subjects possessed nonfunctional MC1Rs, and were
unanimously redheaded. P < 0.05 compared to other condition within-sex. Adapted from Mogil, J. S. et al. 2003. The
melanocortin-1 receptor gene mediates female-specific mechanisms of analgesia in mice and humans. Proc. Natl. Acad. Sci.
U. S. A. 100, 48674872.
188 Redheads and Pain

and perhaps more importantly, that this analgesia was Section 16.5 below), we found that e/e mice were
sensitive to NMDA blockade in the female mutants, significantly more sensitive to analgesia from both
suggesting that they had switched over to the malelike morphine and its bioactive metabolite, morphine-
system (Mogil, J. S. et al., 2003). This switching has 6-glucuronide (Mogil, J. S. et al., 2005a). Curiously,
been observed before and since with other hormonal the effect of MC1R functional status on -opioid
disruptions of the female system (Mogil, J. S. et al., analgesia did not appear to be sex-specific, with
1993; Kavaliers, M. and Galea, L. A. M., 1995; both male and female mutants showing increased
Sternberg, W. F. et al., 2004). Second, injection of a sensitivity (see Figure 2(b)). Again, data collected in
selective peptidergic MC1R antagonist, Ac-Nle-Asp- humans were entirely analogous, with redheads of
Trp-DPhe-Nle-Trp-Lys-NH2, into the cerebral ven- both sexes showing robustly higher morphine-6-
tricles of (outbred) male and female mice produced a glucuronide inhibition of noxious electrical stimuli
potentiation of U50,488 analgesia only in females (see (Mogil, J. S. et al., 2005a; see Figure 2(d)).
Figure 1(b)), and furthermore rendered that analgesia
newly sensitive to inhibition by an NMDA antagonist
(Mogil, J. S. et al., 2003). Finally, Austin and Mogil have 16.4 MC1R and Anesthesia
demonstrated sex and strain differences in periaque-
ductal gray expression of the Mc1r gene using It has long been clinical lore among anesthesiologists
quantitative polymerase chain reaction (J. S. Austin that redheads are harder to anesthetize with inhalant
and J. S. Mogil, unpublished data) in directions entirely anesthetics than others, but these anecdotal impres-
consistent with the hypothesis that lower MC1R func- sions were never tested experimentally until recently.
tionality is associated with higher -opioid analgesia. Liem E. B. et al. (2004) recruited 20 young adult
These data, supportive of the sex-specific invol- women (10 with red hair, 10 with dark hair), and
vement of Mc1r in -opioid analgesia in the mouse, studied the minimum alveolar concentration of des-
inspired a small association study in humans flurane required to prevent movement in response to
designed to test the same hypothesis in our species. intradermal electrical stimulation of the anterior
College-age subjects were tested for thermal and thigh. They found that red-haired women required
ischemic pain before and after administration of pen- almost 20% more anesthetic to abolish responding.
tazocine (Talwin; 0.5 mg kg1) or saline, and then This finding regarding anesthetic sensitivity cannot
genotyped at the human MC1R gene by direct easily be reconciled with the aforementioned findings
sequencing of exons containing common variants. regarding analgesia. If the greater response of MC1R
We found that pentazocine analgesia was modest in mutants (i.e., redheads) to - and -opioid analgesics
all male subjects, and also in women with functional is reflective of higher endogenous activity of a toni-
MC1Rs (i.e., those with the consensus DNA sequence cally active analgesic system, one might have
or only one variant). Women with nonfunctional expected red-haired women to require less anesthetic,
MC1Rs (i.e., those with two or more variants; all not more. Unfortunately, only women were tested in
phenotypic redheads) displayed robust pentazocine the study by Liem E. B. et al. (2004), so sex differences
analgesia (Mogil, J. S. et al., 2003; see Figure 1(c)). could not be evaluated. In apparent support of this
These data represent the first time a genetic find- finding, this same group tested e/e mutants for sensi-
ing in the mouse has ever been directly translated to tivity to four inhaled anesthetic compounds
humans within a single published study. More impor- (isoflurane, sevoflurane, desflurane, and halothane),
tantly, though, the data are the first to suggest that, and found that although mutant mice were not sig-
like the mouse, humans possess qualitatively (i.e., nificantly less sensitive to any one anesthetic
neurochemically) distinct mechanisms of pain mod- compared to controls, when data from all anesthetics
ulation, since the functional status of the MC1R were combined the genotypic difference achieved
affected analgesic output in one sex but not the statistical significance (P 0.023; Xing, Y. et al., 2004).
other. A follow-up study was performed, in order to
determine whether Mc1r and MC1R play a role in
-opioid analgesia in mice and humans, respectively. 16.5 MC1R and Pain
-Opioid analgesia is of much greater clinical rele-
vance than -opioid analgesia due to the high - One limitation of the study by Liem E. B. et al. (2004)
selectivity of most clinically used opiate drugs. In is that no attempt was made to study nociceptive
addition to differences in pain sensitivity per se (see sensitivity of the subjects in the absence of anesthetic,
 Redheads and Pain 189

so it was impossible to tell whether the differences These data are in contrast with an earlier study,
observed involved differential sensitivity among gen- showing no MC1R genotypic differences in thermal
otypes to the anesthetic, or differential sensitivity to pain sensitivity (Mogil, J. S. et al., 2003), although that
the noxious stimulus itself. It has recently been study was probably too lightly powered to assess the
demonstrated in mice that the latter can easily be issue. However, the data of Liem E. B. et al. (2005) are
confused for the former (Mogil, J. S. et al., 2005b). In a also entirely contradictory with a larger follow-up
recently published follow-up study by their group study (Mogil, J. S. et al., 2005a), which observed in e/e
(Liem, E. B. et al., 2005), women (30 with red hair, 30 mutant mice of both sexes (across a broad range of
with dark hair) were tested for sensory thresholds, noxious stimulus modalities) and humans of both
pain thresholds, and pain tolerance to electrical and sexes a decreased sensitivity to noxious electrical
thermal stimulation. No differences were seen in stimulation (see Figures 2(a) and 2(b)). There are a
sensitivity to electrical stimulation, but heat and number of possible reasons for the discrepancy.
cold pain perception was higher in redheads. Methodological differences can be found between
Redheads were also found to be resistant to subcuta- the three experiments, of course. It is difficult to
neous (but not liposomal) lidocaine anesthesia blind experimenters to hair color, and it is possible
against electrical stimulation (Liem, E. B. et al., 2005). that this lack of blinding was a greater issue in one

(a) (b) Male Female

10 B6 100 B6 100 B6
Withdrawal latency (s)

* *
8 e /e e /e e /e *
75 75

% Analgesia
% Analgesia

*
6 *
50 50
4 *

25 25
2

0 0 0
Male Female 5 10 15 5 10 15
M6G dose (mg kg1) M6G dose (mg kg1)

(c) (d)
* *
25 0 or 1 variant 1.75 0 or 1 variant
Pain tolerance (mA)

2+ variants 1.50 2+ variants

20
1.25
AUEC (mA)

15 1.00
10 0.75
0.50
5
0.25
0 0.00
Male Female Male Female
Figure 2 Effect of MC1R functional status on pain sensitivity and -opioid analgesia in mice and humans of both sexes. (a)
Sensitivity of male and female spontaneous mutant recessive yellow (e/e) mice and their wildtype counterparts, C57BL/6 (B6),
on the thermal paw-withdrawal test. Bars represent mean standard error of the mean (SEM) latency (in s) to withdraw from a
radiant heat stimulus positioned under the ventral hind paws (right and left hind paw data combined). (b) Response of B6 and
e/e mutants on the 49  C tail-withdrawal test to systemic injection of various doses (515 mg kg1, subcutaneous) of the
selective -opioid agonist (and morphine metabolite), morphine-6-glucuronide (M6G). Symbols represent mean SEM
percent of total analgesia (based on area under the timeeffect curve) over a 300-min testing period postdrug administration.
(c) Baseline tolerance to electrical current (10 Hz; 0.1 ms; increased by 0.5-mA steps) applied to the skin in men and women
with functional (0 or 1 variant) or nonfunctional (2 variants) MC1Rs. Bars represent mean SEM tolerated stimulus intensity.
(d) Influence of MC1R genotype on M6G analgesia (0.3 mg kg1, intravenous) in men and women. Subjects were studied as
described in detail in Mogil J. S. et al. (2005a). Bars represent mean SEM analgesia as expressed as area under the time
effect curve (AUEC relative to baseline current shown in (c)). P < 0.05 compared to other genotype. Adapted from Mogil, J. S.
et al. 2005a. Melanocortin-1 receptor gene variants affect pain and -opioid analgesia in mice and humans. J. Med. Genet.
42, 583587.
190 Redheads and Pain

study than another. Liem E. B. and colleagues studied Bastiaens, M. T., ter Huurne, J. A. C., Kielich, C., Gruis, N. A.,
Westendorp, R. G. J., Vermeer, B. J., and Bavinck, J. N. B.
women only, whereas Mogil J. S. and colleagues 2001b. Melanocortin-1 receptor gene variants determine the
studied men and women. Most importantly, perhaps, risk of nonmelanoma skin cancer independently of fair skin
Liem E. B. and colleagues compared phenotypes and red hair. Am. J. Hum. Genet. 68, 884894.
Belfer, I., Wu, T., Kingman, A., Krishnaraju, R. K., Goldman, D.,
(women with dark or red hair) whereas Mogil J. S. and Max, M. B. 2004. Candidate gene studies of human pain
and colleagues compared MC1R genotypes (subjects mechanisms: a method for optimizing choice of
with functional or nonfunctional MC1Rs). It is not polymorphisms and sample size. Anesthesiology
100, 15621572.
the case that all redheads are so because of inheri- Chhajlani, V. and Wikberg, J. E. S. 1992. Molecular cloning and
tance of MC1R variants (see Section 16.2 above); that expression of the human melanocyte stimulating hormone
needs to be determined empirically by sequencing. receptor. FEBS Lett. 309, 417420.
Cone, R. D., Lu, D., Koppula, S., Vage, D. I., Klungland, H.,
Thus it is possible that some subjects in Liem E. B. Boston, B., Chen, W., Orth, D. N., Pouton, C., and
et al. (2005) were misclassified in their inferred Kesterson, R. A. 1996. The melanocortin receptors:
genotype. agonists, antagonists, and the hormonal control of
pigmentation. Rec. Prog. Horm. Res. 51, 287317.
Finally, a human genetic association study has Diatchenko, L., Slade, G. D., Nackley, A. G., Bhalang, K.,
revealed a possible association with MC1R variants Sigurdsson, A., Belfer, I., Goldman, D., Xu, K.,
and vulvar vestibulitis, with sufferers 3.5-fold more Shabalina, S. A., Shagin, D., Max, M. B., Makarov, S. S., and
Maixner, W. 2005. Genetic basis for individual variations in
likely to carry loss-of-function alleles at the MC1R pain perception and the development of a chronic pain
(Foster, D. C. et al., 2004). It is unclear whether this condition. Hum. Mol. Genet. 14, 135143.
risk is related to altered pain processing or, as sug- Eberle, A. N. 1988. The Melanotropins. Chemistry, Physiology
and Mechanisms of Action. S. Karger.
gested by the authors, to the known anti- Fillingim, R. B., Kaplan, L., Staud, R., Ness, T. J., Glover, T. L.,
inflammatory role of MC1Rs. Campbell, C. M., Mogil, J. S., and Wallace, M. R. 2005. The
A118G single nucleotide polymorphism of the mu-opioid
receptor gene (OPRM1) is associated with pressure pain
sensitivity in humans. J. Pain 6, 159167.
Foster, D. C., Sazenski, T. M., and Stodgell, C. J. 2004. Impact
16.6 Future Directions of genetic variation in interleukin-1 receptor antagonist and
melanocortin-1 receptor genes on vulvar vestibulitis
It is obvious that future work is needed to resolve the syndrome. J. Reprod. Med. 49, 503509.
Gear, R. W., Miaskowski, C., Gordon, N. C., Paul, S. M.,
true directions of MC1R genotypic effect on pain and Heller, P. H., and Levine, J. D. 1996. Kappa-opioids produce
pain inhibition phenotypes, and to clarify which phe- significantly greater analgesia in women than in men. Nature
notypes are sex-specific and which are not. Med. 2, 12481250.
Healy, E., Flanagan, N., Ray, A., Todd, C., Jackson, I. J.,
Nonetheless, the fact that a gene like MC1R would Matthews, J. N. S., Birch-Machin, M. A., and Rees, J. L.
be involved in pain, analgesia, or anesthesia should be 2000. Melanocortin-1-receptor gene and sun sensitivity in
considered quite a surprise. It does not appear on any individuals without red hair. Lancet 355, 10721073.
Kavaliers, M. and Choleris, E. 1997. Sex differences in N-
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Klepstad, P., Rakvag, T. T., Kaasa, S., Holthe, M., Dale, O.,
Borchgrevink, P. C., Baar, C., Vikan, T., Krokan, H. E., and
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-opioid receptor gene may increase morphine
Abdel-Malek, Z. A. 2001. Melanocortin receptors: their requirements in patients with pain caused by malignant
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Annu. Rev. Neurosci. 7, 309338. Increased sensitivity to thermal pain and reduced
Bastiaens, M., ter Huurne, J., Gruis, N., Bergman, W., subcutaneous lidocaine efficacy in redheads.
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The melanocortin-1-receptor gene is the major freckle gene. Liem, E. B., Lin, C. M., Suleman, M. I., Doufas, A. G.,
Hum. Mol. Genet. 10, 17011708. Gregg, R. G., Veauthier, J. M., Loyd, G., and Sessler, D. I.
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2004. Anesthetic requirement is increased in redheads. Quillan, J. M. and Sadee, W. 1997. Dynorphin peptides:
Anesthesiology 101, 279283. antagonists of melanocortin receptors. Pharm. Res.
Mogil, J. S. (ed.) 2004. The Genetics of Pain, p. 349. IASP 14, 713719.
Press. Schaffer, J. V. and Bolognia, J. L. 2001. The melanocortin-1
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Mitchell, S. R., Kest, B., and Belknap, J. K. 1997. 137, 14771485.
Identification of a sex-specific quantitative trait locus Solovieva, S., Leino-Arjas, P., Saarela, J., Luoma, K.,
mediating nonopioid stress-induced analgesia in female Raininko, R., and Riihimaki, H. 2004. Possible association of
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 17 Autonomic Nervous System and Pain
Wilfrid Janig, Physiologisches Institut, Christian-Albrechts-Universitat zu Kiel, Germany
2009 Elsevier Inc. All rights reserved.

17.1 Introduction 194

17.2 Neurobiology of the Autonomic Nervous System and Pain 196
17.2.1 Reactions of the Sympathetic Nervous System in Pain 196
17.2.2 Visceral Afferents, Autonomic Nervous System, and Pain 198
17.2.3 Hyperalgesia and Sympathetically Mediated Changes in Referred Zones During
Visceral Pain 199
17.3 Role of the Sympathetic Nervous System in the Generation of Pain 201
17.3.1 Clinical Background: Sympathetically Maintained Pain 201
17.3.2 Sympathetically Maintained Pain Following Nerve Lesion Simulated in Behavioral
Animal Models 202
17.3.3 Direct Involvement of the Sympathetic Nervous System in the Generation of Pain
Following Nerve Trauma 205
17.3.3.1 Coupling between lesioned postganglionic and afferent nerve terminals 205
17.3.3.2 Coupling between unlesioned postganglionic and afferent nerve terminals following
partial nerve lesion 206
17.3.3.3 Coupling in the dorsal root ganglion and collateral sprouting following peripheral nerve
lesion 206
17.3.3.4 Adrenoceptors involved in chemical sympathetic afferent coupling following nerve
lesion 208
17.3.3.5 Synopsis 208
17.3.4 Indirect Involvement of the Sympathetic Nervous System in the Generation of Pain 209
17.3.4.1 Changes of neurovascular transmission and development of hyperreactivity of blood
vessels 209
17.3.4.2 Sensitization of nociceptors mediated by sympathetic terminals independent of
excitation and release of noradrenaline 211
17.3.4.3 Sensitization of nociceptors initiated by nerve growth factor or cytokines possibly
mediated by sympathetic terminals 212
17.3.4.4 Sympatho-adrenal system and nociceptor sensitization 213
17.4 Sympathetic Nervous System and Central Integrative Mechanisms in the Control of
Hyperalgesia and Inflammation 216
17.4.1 The Complex Regional Pain Syndrome Type I as Model 216
17.4.2 Sympathetic Nervous System and Acute Experimental Inflammation 218
17.4.3 Sympathetic Nervous System and Immune System 221
References 221

Glossary
allodynia, hyperalgesia Hyperalgesia denotes or cold receptors (cold allodynia). The mechanism of
increased pain generated by a stimulus that is nor- allodynia is central (central sensitization generated
mally painful and excites nociceptors. It has a by persistent excitation of nociceptors). Secondary
peripheral (sensitization of nociceptors) and/or a allodynia is pain elicited by stimulation of low-
central component (sensitization of central neurons, threshold mechanoreceptors in an area of skin that
e.g., in the dorsal horn of the spinal cord). Allodynia surrounds a territory with sensitized nociceptors
is pain generated by stimuli that activate low- (e.g., generated by inflammation) (Merskey, H. and
threshold mechanoreceptors (mechanical allodynia) Bogduk, H., 1994; Meyer, R. A. et al., 2005).

193
194 Autonomic Nervous System and Pain

causalgia See Chapter Complex Regional Pain or deep body structures where the pain is referred
Syndromes. to is innervated by nerves of the same spinal seg-
complex regional pain syndrome See Chapter ments as the affected deep body structures
Neuropathic Pain: Clinical. (Giamberardino, M. A., 1999; Janig, W., 1993;
enteric nervous system The enteric nervous sys- Janig, W. and Habler, H. J., 2002).
tem (Janig, W., 2006). sympathetic block, sympathectomy See
homeostasis The maintenance of physiological Chapter Sprouting in Dorsal Root Ganglia.
parameters such as concentration of ions, blood sympathetically maintained pain (SMP) See
glucose, arterial blood gases, and body core tem- Chapter Complex Regional Pain Syndromes.
perature in a narrow range and around sympatho-adrenal system The sympatho-
predetermined set-points is called homeostasis. adrenal system is the adrenal medulla and its
Homeostatic regulation involves autonomic sys- innervation by sympathetic preganglionic neurons.
tems, endocrine systems, and the respiratory The adrenal medulla consists of cells that release
system (Janig, W., 2006). either adrenaline or noradrenaline upon impulses in
neuropathic pain See Chapters Neuropathic the preganglionic neurons. The sympatho-adrenal
Pain: Basic Mechanisms (Animal) and Complex system is functionally distinct from the various
Regional Pain Syndromes. types of sympatho-neural systems (Janig, W.,
referred pain Pain from a particular visceral organ 2006).
or a particular deep somatic structure is perceived viscero-somatic convergent neurons (dorsal
to arise from the body surface (skin), from deep horn of the spinal cord) See Chapter Spinal Cord
somatic structures, or from other viscera. The area Mechanisms of Hyperalgesia and Allodynia.

17.1 Introduction aspects of pain, hyperalgesia, and inflammation. This

is discussed elsewhere ( Janig, W., 2005; Janig, W. and
Research on the relationship between the autonomic Levine, J. D., 2006) and summarized in Janig W. (2007).
nervous system and pain is a controversial field with The topic Autonomic Nervous System and Pain cannot
many interesting facets that are important from the be reduced to peripheral local processes in which
biological, pathobiological, and clinical point of view. peripheral nociceptive afferent fibers (sensitization of
(I prefer to use the terms biology and pathobiology, nociceptors and generation of ectopic impulses after
instead of physiology and pathophysiology, because nerve lesion), sympathetic fibers, and nonneural cells
they include physiological, morphological, biochem- (e.g., cells of the immune system and the vasculature)
ical, and molecular aspects.) I will discuss this field in are involved. The question to be asked is: How are the
a broad context and focus mainly on the sympathetic cellular and subcellular processes that lead to ongoing
nervous system. I will argue that the peripheral sympa- pain, allodynia, and hyperalgesia in neuropathic pain
thetic noradrenergic neuron may have, in addition to and inflammatory pain orchestrated by the brain in the
its conventional function to transmit signals generated continuous protection of the body against agents from
in the brain to peripheral target cells (e.g., smooth outside as well as from within the body? The brain is an
muscle cells, secretory epithelia, heart cells, and neu- important player to organize and control these protec-
rons of the enteric nervous system), quite different tive body reactions via neuroendocrine (hypothalamo-
functions that are directly or indirectly related to pro- pituitary-adrenocortical and sympatho-adrenal) and
tection of body tissues and pain. These functions have autonomic (preferentially the sympathetic) systems.
not been studied as extensively as those related to the Feedback to the brain occurs via various signaling
regulation of autonomic target cells (Janig, W., 2006). molecules (e.g., interleukines), hard-wired nociceptive
The parasympathetic nervous system is not involved in afferents, and other afferent neurons (Figure 1).
the generation of pain, yet it may also be important in Table 1 lists different functions of the sympathetic
protection of body tissues (e.g., the gastrointestinal tract nervous system related to pain and protection of
(GIT)). Vagal afferents are involved in integrative body tissues in biological and pathobiological
 Autonomic Nervous System and Pain 195

feelings of emotions may require explicitly the afferent

Cortex
limbic system
feedback from body responses generated by the efferent
neural autonomic and somatomotor signals (Damasio,
A., 2003). These central representations act back on the
peripheral tissues, the immune and nociceptive primary
afferent neurons, via the endocrine and autonomic ner-
Hypothalamus
vous systems (Figure 1). The central circuits are also
the origin of illness responses that include aversive
feelings, pain, and hyperalgesia. This central integration
Brain stem is related or identical to the integrative processes, invol-
ving neuroendocrine, immune, and neural systems,
NTS
Cytokines HPA occurring during environmental challenges such as
other system Vagal viral and bacterial infection (Arkins, S. et al., 2001;
afferents
Miller, A. H. et al., 2001). Thus, the autonomic nervous
Small system is one component in the generation of pain and
SA system
intestine protective body reactions that are organized and regu-
Inflammation
SN system lated by the brain and disease occurs when this neural
Nociceptor
sensitization regulation fails.
Ectopic neural
impulse activity I will distinguish three aspects of the topic
Nociceptive
Spinal cord Sympathetic nervous system and pain (Table 1):
afferent system
Neural Control Components 1. Reactions of the sympathetic nervous system dur-
Endocrine Telencephalon
Spinal cord, brainstem,
ing pain. This includes the autonomic integrated
HPA Hypothalamic hypothalamus and adaptive reactions during pain, reflexes to
pituitaryadrenal Neuroendocrine systems
SA Sympathoadrenal (hypothal., SA system) noxious stimulation, and reactions mediated by
SN Sympathoneural
Vagal afferents sympathetic systems in referred zones.
Figure 1 Spinal cord, brainstem, and hypothalamus 2. Coupling (cross-talk) from sympathetic (noradre-
contain neuronal circuits (pink shaded area) that modulate nergic) neurons to afferent neurons in the
nociceptor sensitivity, inflammation, and ectopic activity in generation of pain. Four broad categories of cou-
lesioned afferent neurons in the periphery of the body via the pling will be discussed: Direct coupling related to
sympatho-adrenal (SA) system, the sympatho-neural
excitation of the sympathetic postganglionic neu-
system, and the hypothalamicpituitaryadrenal (HPA)
system. Feedback information from the peripheral rons and release of noradrenaline; indirect
inflammatory process occurs via nociceptive primary coupling related to influence of inflammatory pro-
afferent neurons and cytokines. Abdominal vagal afferents cesses by the sympathetic nervous system;
signal events from the inner defense line of the body (GALT, coupling independent of excitation and noradre-
gut-associated lymphoid tissue) to the lower brainstem
naline release by sympathetic neurons. A fourth
(NTS, nucleus tractus solitarii). The telencephalon controls
inflammation and sensitivity of nociceptors via the circuits in category is the effect of adrenaline released by the
the neuraxis (see shaded double arrows). adrenal medulla on the sensitivity of nociceptors.
3. Sympathetic nervous system and central mechan-
isms. Based on observations made on patients and
conditions. It shows that peripheral as well as central animal experimentation, it is likely that activity in
mechanisms have to be considered and that the topic sympathetic noradrenergic neurons or in the sym-
includes not only pain and hyperalgesia but also neural patho-adrenal system, both generated in the brain,
and neuroendocrine regulation of inflammation and contributes directly or indirectly to pain. This
central aspect includes the various forms of sym-
the immune system.
pathetic afferent coupling discussed before.
The integration of the control of protective neural,
endocrine, and immune mechanisms occurs in the brain
(brainstem, hypothalamus, limbic system, and neocor- The role of the sympathetic nervous system and
tex). Perception of sensations, feeling of emotions, its transmitters in chronic inflammation (such as
autonomic, endocrine, and somatomotor responses are rheumatoid diseases) or in systemic chronic pain
coordinated and are, therefore, parallel readouts of the diseases (e.g., fibromyalgia) and its interaction with
central representations, although the evolvement of the peptidergic afferent innervation as well as the
196 Autonomic Nervous System and Pain

Table 1 Sympathetic nervous system and pain

Reactions of the sympathetic nervous system in paina

Protective spinal reflexes
Fight, flight, and quiescence organized at the level of the periaqueductal gray
Hyperalgesia and sympathetically mediated changes in referred zones during visceral pain
Role of the sympathetic nervous system in the generation of painb
Sympatheticafferent coupling in the periphery
Coupling after nerve lesion (noradrenaline, -adrenoceptors)
Coupling via the micromilieu of the nociceptor and the vascular bed
Sensitization of nociceptors mediated by sympathetic terminals independent of excitation and release of noradrenaline
Sensitization of nociceptors initiated by cytokines or nerve growth factor and mediated by sympathetic terminals
Sympatho-adrenal system and nociceptor sensitization
Sympathetic nervous system and central mechanismsc
Control of inflammation and hyperalgesia by sympathetic and neuroendocrine mechanisms
Complex regional pain syndrome and sympathetic nervous system
Immune system and sympathetic nervous system
Rheumatic diseases and sympathetic nervous system
Systemic chronic pain (e.g., fibromyalgia) and sympathetic nervous system
a
Bandler R. and Shipley M. T. (1994); Bandler R. et al. (2000a; 2000b); Janig W. (1993; 2006).
b
Janig W. and Baron R. (2001); Janig W. (2005); Janig W. and Habler H. J. (2000b); Janig W. and Koltzenburg M. (1991); Janig W. and
Levine J. D. (2005).
c
Janig W. and Baron R. (2002; 2003; 2004); Janig W. and Habler H. J. (2000a); Janig W. et al. (2000); Janig W. and Levine J. D. (2005);
Straub R. H. and Harle P. (2005); Straub R. H. et al. (2005); Vierck C. J. Jr. (2006).

hypothalamo-pituitary-adrenal (HPA) system will in particular of the sympathetic nervous system, in the
not be discussed. We are at the beginning of the periphery and in the brain, mostly receives insufficient
experimental investigation of these control mechan- attention in studies of the functioning of this system
isms (Straub, R. H. and Harle, P., 2005; Straub, R. H. under pathobiological conditions. I emphasize three
et al., 2005; Vierck, C. J. Jr., 2006). points (Janig, W. and McLachlan, E. M., 2002; Janig,
W., 2006): First, the sympathetic nervous system
is functionally as well differentiated as the parasym-
pathetic nervous system. It does not react as a unitary
17.2 Neurobiology of the Autonomic
system under normal physiological conditions.
Nervous System and Pain
Second, the terms sympathetic and parasympathetic
are defined anatomically and not functionally.
The peripheral autonomic nervous system is by defi-
Thus, to use these terms functionally is misleading.
nition an efferent system consisting of many
Third, the long-held view that sympathetic and para-
functionally distinct pathways that transmit the
sympathetic nervous system function universally
impulse activity from the spinal cord or brainstem to
antagonistically is also misleading. Both systems com-
the effector cells. These pathways are separate from
plement each other in their functions.
each other and functionally distinct with respect to the
effector cells, as is the impulse transmission in the
autonomic ganglia and at the neuroeffector junctions
17.2.1 Reactions of the Sympathetic
to the effector cells. Each peripheral sympathetic or
Nervous System in Pain
parasympathetic pathway is connected to distinct neu-
ronal networks in the spinal cord, brainstem, and Any acute, but possibly also chronic tissue-damaging
hypothalamus that generate the typical discharge pat- stimulus affects the sympathetic nervous system.
terns in the autonomic neurons and are responsible for Neurons of sympathetic systems exhibit generalized
the precise regulation of the target organs (cardiovas- and specific reactions to these stimuli. The general-
cular regulation, thermoregulation, regulation of ized reactions probably occur only in certain types of
pelvic organs, regulation of GIT, and so forth). The sympathetic system (e.g., muscle vasoconstrictor,
autonomic regulations are represented in these central visceral vasoconstrictor, sudomotor neurons, or sym-
networks. The anatomical, histochemical, and func- pathetic cardiomotor neurons) but are weak or absent
tional organization of the autonomic nervous system, in other systems (e.g., sympathetic systems to pelvic
 Autonomic Nervous System and Pain 197

organs). They are organized in spinal cord, brainstem and chronic pain occurring particularly in the deep
(medulla oblongata and mesencephalon), and and visceral body domains. It is represented in the
hypothalamus and can be conceptualized as compo- ventrolateral periaqueductal gray, activated from the
nent parts of the different patterns of defense deep (somatic and visceral) body domains, and con-
behavior, such as confrontational defense, flight, or sists of hyporeactivity, hypotension, bradycardia, and
quiescence (Figure 2). Confrontational defense and an endogenous opioid analgesia. These stereotyped
flight are typical of an active defense strategy when preprogrammed elementary behaviors and their
animals encounter threatening stimuli that are poten- association with the endogenous control of analgesia
tially injurious for the body, the former potentially enable the organism to cope with dangerous situa-
leading to fight and the latter to forward avoidance. tions that are always accompanied by pain or
Both patterns are represented in the lateral and dor- impending pain. The dorsolateral, lateral, and ven-
solateral periaqueductal gray of the mesencephalon, trolateral columns of the periaqueductal gray have
activated from the body surface or cortex, and asso- distinct reciprocal connections with the autonomic
ciated with endogenous nonopioid analgesia, centers in the lower brainstem and hypothalamus
hypertension, and tachycardia. Quiescence is similar that differentially regulate the activity in neurons of
to the natural reactions of mammals to serious injury the autonomic pathways. They are under differential

Active coping strategies

evoked from the lPAG and the dlPAG
Confrontational Defense/Threat
Hypertension and tachycardia
Extracranial vasodilation
Hindlimb and renal vasoconstriction
Nonopioid-mediated analgesia

dm Flight
dl Hypertension and tachycardia
Hindlimb vasodilation
dm Extracranial and renal vasoconstriction
dl Nonopioid-mediated analgesia
l dm
dl

l
dm
vl dl
Passive coping strategies
l
evoked from the vlPAG

vl
Quiescence/Hyporeactivity
Hypotension
Bradycardia
Opioid-mediated analgesia

Figure 2 Representation of defense behaviors in the dorsolateral, lateral, and ventrolateral periaqueductal gray. Schematic
illustration of the dorsolateral (dl), lateral (l), and ventrolateral (vl) columns within the rostral, intermediate, and caudal
periaqueductal gray (PAG). The dorsomedial neural PAG columns are indicated too. Stimulation of neuron populations in the
dlPAG, lPAG, and vlPAG by microinjections of the excitatory amino acid glutamate that excites only cell bodies of neurons but
not axons evokes distinct defense behaviors: Confrontational defense is elicited from the rostral portion of the dlPAG and
lPAG; flight is elicited from the caudal part of the dlPAG and lPAG; quiescence (cessation of spontaneous motor activity) is
elicited from the vlPAG in the caudal portion of the PAG. These defense behaviors include typical autonomic cardiovascular
reactions (changes of blood pressure, heart rate, and blood flows) and sensory changes (nonopioid- or opioid-mediated
analgesia). The representations of confrontational defense and flight are the basis for active coping strategies produced by
the cortex. The representation of quiescence is the basis for passive coping strategies produced by the cortex. Modified from
Bandler R. and Shipley M. T. (1994) and Bandler R. et al. (2000b).
198 Autonomic Nervous System and Pain

control of the medial and orbital prefrontal cortex milieu and the regulation of the organs to the beha-
and other telencephalic centers (Bandler, R. and vior of the organism. They are anatomically and
Shipley, M. T., 1994; Bandler, R. et al., 2000a; 2000b; functionally closely associated with the autonomic
Keay, K. A. and Bandler, R., 2004). nervous system. However, they should not be labeled
There also exist more localized reactions of the sympathetic, parasympathetic, or autonomic afferent
sympathetic nervous system to noxious stimuli that neurons. These functional labels are misleading since
are organized within the spinal cord and trigeminal they imply that the visceral afferent neurons have
nucleus and in the periphery. These localized reac- functions that uniquely pertain to the sympathetic or
tions are reflected in various somato-sympathetic and parasympathetic autonomic nervous system, the
viscero-sympathetic reflexes, the afferent neurons of exception being that afferent neurons of the enteric
these reflexes are nociceptive (skin, deep somatic nervous system are per definition enteric afferent
tissues, and viscera) and the efferent neurons sympa- neurons. No functional, morphological, histochem-
thetic innervating blood vessels (in skin, deep ical, or other criteria do exist to associate any type of
somatic tissues, and viscera), sweat glands, erector visceral afferent neuron with only one autonomic
pili muscles, enteric nervous system, or other effec- system ( Janig, W., 2006).
tors in the viscera. Both afferent neurons and efferent Usually, vagal afferent neurons are described and
pathways are synaptically connected by various characterized by adequate physiological stimuli
groups of interneurons, establishing circuits that exciting them and by the main reflexes associated
probably are the basis of the changes occurring in with the cardiovascular system, respiratory system,
referred zones during chronic stimulation of deep GIT, or other regulatory systems (Undem, B. J. and
nociceptive primary afferents (see below). For exam- Weinreich, D., 2005). However, several observations
ple, cutaneous vasoconstrictor neurons exhibit show that groups of vagal afferents innervating visc-
distinct inhibitory reflexes to noxious stimuli of the eral organs in the thoracic and abdominal cavity are
territories innervated by these neurons. involved in the control of nociception and pain, in
The hypothalamo-mesencephalic and the spinal neuroendocrine control of nociceptors, in the control
level of integration are presumably protective under of inflammation, and in the control of general pro-
normal biological conditions and are associated with tective body reactions including illness responses.
the activation of the hypothalamo-pituitary adreno- Subgroups of vagal afferent neurons may signal
cortical system ( Janig, W., 2006). events from visceral organs, in particular those
occurring at the internal defense line of the body in
the GIT represented in the liver and in the small
17.2.2 Visceral Afferents, Autonomic
intestine (which contains the largest immune system
Nervous System, and Pain
of the body, the gut associated lymphoid tissue
Visceral organs in the thoracic, abdominal, and pelvic (GALT)), to the central nervous system that triggers
cavities are innervated by two sets of extrinsic protective body reactions not only in the viscera but
primary afferent neurons: spinal visceral afferent also in the superficial and deep somatic body tissues
neurons and vagal visceral afferent neurons (the (see Chapters Visceral Pain; Vagal Afferent Neurons
latter also including afferent neurons from arterial and Pain; Janig, W., 2005). These vagal afferent neu-
baro- and chemoreceptors that project through the rons may be activated or sensitized by cellular
glossopharyngeal nerve). Spinal visceral afferent processes of these inner body defense lines and
neurons have their cell bodies in the dorsal root their signaling molecules. Stimulation of vagal affer-
ganglia (DRG) and vagal afferent neurons in the ents may generate sickness behavior that is
inferior (nodose) ganglion of the vagus nerve (a few characterized by several protective illness responses
in the superior (jugular) ganglion of the vagus nerve (immobility, decreased social interaction, decrease in
and some arterial baro- and chemoreceptor afferents food intake and of digestion, formation of taste aver-
in the petrosal ganglion). Visceral afferent neurons sion against novel food, loss of weight, fever, increase
are involved in specific organ regulations, multiple of sleep, change in endocrine functions (e.g., of the
organ reflexes, neuroendocrine regulations, and visc- activation of the hypothalamicpituitary system),
eral sensations (including visceral pain), shaping and malaise; Watkins, L. R. et al., 1995; Maier, S. F. and
eliciting emotional feelings and other functions. Watkins, L. R., 1998; Watkins, L. R. and Maier, S. F.,
They monitor the inner state of the body and serve 1999; 2000; Goehler, L. E. et al., 2000), activates the
to maintain homeostasis and to adapt the internal inhibitory control of nociceptive impulse transmission
 Autonomic Nervous System and Pain 199

in the spinal and trigeminal dorsal horn (Foreman, R. Little attention has been given to the generation of
D., 1989; Randich, A. and Gebhart, G. F., 1992; Janig, autonomic changes (reflected in changes of blood
W., 2005), inhibits nociceptiveneuroendocrine flow, sweating, and piloerection) and of trophic
reflex circuits that control inflammation (via the changes in the skin and its appendages, subcutis,
sympatho-adrenal system and the hypothalamic joint capsules, and fascia of the referred hyperalgesic
pituitary system (Green, P. G. et al., 1995; 1997; (Heads) zones during visceral diseases (Janig, W. and
Janig, W. et al., 2000; Miao, F. J.-P. et al., 2000; Morrison, S. F., 1986). These changes are very vari-
2001)), and inhibits the central circuits that control able between individuals but reported to be relatively
via the adrenal medulla and release of adrenaline the stable for each individual. The clinical diagnosis of a
sensitivity of nociceptors (Khasar, S. G. et al., 1998a; visceral disease (e.g., in the GIT, of the evacuative
1998b; Janig, W. et al., 2000; Khasar, S. G. et al., 2003; organs, or of the heart) may be predicted from these
Janig, W. et al., 2006; see below). For these hypothetic changes with a probability of about 70% (Beal, M. C.,
functions of vagal afferents involving neuroendocrine 1983; Cox, J. M. et al., 1983; Beal, M. C., 1985;
and autonomic systems, in the protection of the body, Nicholas, A. S. et al., 1985).
an umbrella concept in the frame of body protection, It is important to note that the trophic changes
pain, and inflammation has to be formulated. This occur in regions that are outside of the site of injury
has to include the central circuits (see Janig, W., 2005; or trauma, that the first sign of the trophic changes is
Janig, W. and Levine, J. D., 2006). probably the edema, and that the severe signs of these
changes develop relatively slowly and late. These
changes are believed to be generated by the segmen-
17.2.3 Hyperalgesia and Sympathetically
tal sympathetic outflow to the visceral reference
Mediated Changes in Referred Zones During
zones at the trunk. However, the trophic changes in
Visceral Pain
the referred zones may also or additionally be related
Chronic stimulation of spinal afferents from visceral to changes in the retrograde axonal transport in
organs may elicit pain, hyperalgesia, and allodynia in afferent neurons that innervate the referred zones
the referred zones of the body surface and of the deep (see discussion in Janig, W., 1993). We need more
somatic domain (skin, subcutaneous tissue, muscula- reliable and quantitative data about the changes
ture, tendons, fascia, and bones). Hyperalgesia and occurring in the referred somatic zones in patients
allodynia that are elicited in the superficial and deep with various visceral diseases to develop testable
somatic domains by visceral diseases are equivalent hypotheses.
to secondary hyperalgesia and allodynia in the skin The mechanisms explaining referred visceral pain
caused by ongoing activity in polymodal C-nocicep- and the changes observed in the referred zones
tors (Cervero, F. and Janig, W., 1992; Vecchiet, L. that probably are associated with the sympathetic
et al., 1993). The degree and spatial spread of referred neurons are barely understood. Classical central the-
pain, hyperalgesia, and allodynia depend on the ories trying to explain referred visceral pain are
intensity and duration of the visceral stimulation the convergence-projection theory of Ruch T. C.
and the type of visceral organ. They are abolished (1965) and the convergence-facilitation theory of
after blockade of the impulse traffic in spinal visceral MacKenzie, J. (1920). (The convergence-projection
afferents to the spinal cord. They may be generated theory of Ruch T. C. (1965) is based on the concept
by sensitization of neurons in the spinal cord that are that spinal neurons in the dorsal horn projecting to
involved in the generation of sensations (e.g., spino- the thalamus obtain convergent synaptic input from
thalamic tract neurons), in the descending control of primary afferent neurons innervating viscera or
spinal circuits (e.g., spinal neurons projecting to the somatic body tissues. The telencephalon localizes
brainstem or interneurons mediating signals of the the pain generated by activity in visceral afferents
endogenous control system), and in the regulation of erroneously into somatic tissues of the corresponding
sympathetic target organs and skeletal musculature dermatomes, myotomes, and sklerotomes. The con-
constituting spinal and supraspinal viscero-sympathetic vergence-facilitation theory of MacKenzie J. (1920)
and viscero-somatic reflex pathways (Figure 3). The proposes that afferent inputs from the viscera to the
most likely candidate of these neurons is the broad class spinal cord are not directly related to pain sensation
of viscero-somatic convergent neurons in the spinal but that they produce a generalized increase of excit-
cord (Cervero, F. and Janig, W., 1992; Cervero, F., ability in the area of the spinal cord, which is
1995; Bielefeldt, T. K. and Gebhart, G. F., 2006). related to the respective visceral organ generating
200 Autonomic Nervous System and Pain

Observed phenomena:

BS, thal 1) Referred pain,

Viscus hyperalgesia, allodynia
2) Autonomic changes:
blood vessels, sweat glands
DH
3) Muscle tension
4) Trophic changes

Possible mechanisms:

1) Sensitization of
spinal neurons
2) Changes of balance
Blood vessels between spinal circuits
Sweat glands and descending
Other control systems
3) Viscerosympathetic
reflexes
4) Viscerosomatic
reflexes
5) Changes in regulation of
peripheral microcirculation
6) Changes of retrograde
axonal transport in
afferent neurons

Figure 3 Observed phenomena generated in referred zones during visceral trauma (e.g., inflammation) and the putative
mechanisms underlying these changes. Sympathetic pathways (indicated in red) mediate changes of blood flow, sweating, or
piloerection and possibly changes of tissue structure (edema, trophic changes). The mechanisms underlying the latter are not
understood. Continuous arrows, excitation; dashed arrows, retrograde axonal transport. BS, brainstem; DH, dorsal horn; thal,
thalamus. Modified from Janig W. (1993).

an irritable focus. This was to promote activity in activity in visceral afferents (Herrero, J. F. et al.,
neurons projecting to the thalamus and obtaining 2000; Janig, W. and Habler, H. J., 2002).
convergent synaptic input from somatic tissue (lead- This invites speculation that not only spinal
ing to referred pain and referred hyperalgesia), in mechanisms but also, or in particular, supraspinal
motoneurons (leading to muscle spasms), and in sym- mechanisms are responsible to explain the phenom-
pathetic neurons (leading to autonomic changes enology of referred pain. Supraspinal mechanisms
in the referred zones) (Ness, T. J., 1995).) However, controlling synaptic impulse transmission in the dor-
both theories explain the mechanisms underlying sal horn are inhibitory or excitatory (Urban, M. O.
referred pain rather incompletely and the mechanisms and Gebhart, G. F., 1999; Porreca, F. et al., 2002).
of the other changes in the referred zones in which These mechanisms are represented in the periaque-
the sympathetic nervous system is involved not at all ductal gray, the caudal raphe nuclei, the
(see Ness, T. J. and Gebhart, G. F., 1990; Janig, W. ventromedial medulla, and the dorsolateral pontine
and Koltzenburg, M., 1993; Vecchiet, L. et al., 1993; tegmentum (Fields, H. L. et al., 2006). They do con-
Ness, T. J., 1995; Giamberardino, M. A., 1999, for trol not only nociceptive impulse transmission in the
discussion). It is suggested that persistent sensitiza- dorsal horn but also various autonomic regulations
tion and activation of visceral afferent neurons (e.g., and possibly neuroendocrine regulations (Mason, P.,
during chronic inflammation) sensitize the second- 2001) and are in turn under the control of the cere-
order viscero-somatic convergent neurons in the dor- bral hemispheres. Thus, the reference of visceral
sal horn of the spinal cord that can be synaptically affections (e.g., chronic inflammation in the viscera)
activated from viscera as well as from somatic tissues into somatic zones may be dependent on these endo-
(skin and deep somatic tissues). However, it has not genous protective control systems. These referred
been convincingly demonstrated that these viscero- phenomena (that include sensory, autonomic, and
somatic convergent neurons can be sensitized by motor changes) may therefore be the biological
 Autonomic Nervous System and Pain 201

expression of protective mechanisms operating in 2. Intradermal injection of noradrenaline in physio-

spinal cord and brainstem in the somatic body logically relevant doses in patients with SMP
domains. evokes greater pain in the affected limb regions
than in the contralateral unaffected limb or in
control subjects. Most patients whose pain
increased after injection of noradrenaline in the
17.3 Role of the Sympathetic affected extremity report a decrease in pain fol-
Nervous System in the Generation of lowing systemic injection of the -adrenoceptor
Pain blocker phentolamine (Ali, Z. et al., 2000; see also
Arner, S., 1991).
17.3.1 Clinical Background:
3. In CRPS type I patients with SMP, pain is
Sympathetically Maintained Pain
relieved after blockade of sympathetic activity to
Pain being dependent on activity in sympathetic neu- the affected extremity by a local anesthetic
rons is called sympathetically maintained pain (SMP). applied to the appropriate sympathetic paraver-
SMP is a symptom and includes generically sponta- tebral ganglia. This pain relief lasts significantly
neous pain and pain evoked by mechanical or thermal longer than the short-lasting pain-relieving effect
stimuli. It may be present in the complex regional pain of saline injected close to the same paravertebral
syndrome (CRPS) type I and type II and in other pain ganglia (control injection) in the same group of
syndromes. The idea about the involvement of the CRPS patients showing that the real pain-reliev-
(efferent) sympathetic nervous system in the genera- ing effect generated by blockade of sympathetic
tion of pain is based on various clinical observations activity can be discriminated from the placebo
that have been documented in the literature since tens effect and the duration of pain relief outlasts the
of years (Bonica, J. J., 1990). Representative for these duration of conduction block generated by the
multiple observations on patients with pain in which local anesthetic by an order of magnitude (Price,
the sympathetic nervous system is causally involved D. D. et al., 1998; see Figure 15).
are experimental investigation on patients with CRPS 4. Pain (spontaneous, mechanical allodynia) in the
who had SMP (14) and other less well-controlled hand of chronic CRPS I patients with SMP, which
observations on patients with pain. is clamped to a temperature of about 36  C, is
enhanced when activity in cutaneous vasoconstric-
1. Spontaneous pain, mechanical allodynia, and cold tor neurons is increased by central cooling
allodynia in the upper extremity of patients with (Figure 5), yet not in chronic CRPS I patients with-
CRPS II, which are alleviated by stellate ganglion out SMP (Baron, R. et al., 2002). This experiment
block, can be rekindled, under the condition of demonstrates that an increase of activity in sympa-
sympathetic block, by injection of noradrenaline thetic neurons by a physiological intervention
into the skin area that was painful before sympa- (central cooling) can increase pain, possibly inde-
thetic block (Figure 4; Torebjork, H. E. et al., pendent of a vascular component. An important side
1995). By the same token, it has been shown in effect of this experimental investigation is that the
patients with causalgia, who had been successfully peripheral mechanisms underlying SMP in CRPS I
treated either by sympathectomy or by transcuta- patients occur mainly in deep somatic tissues.
neous nerve stimulation, that ionophoretic 5. Injection of adrenaline around a stump neuroma
application of noradrenaline to the previously that has developed long after limb amputation
causalgic skin area rekindles spontaneous pain may be intensely painful (Chabal, C. et al., 1992).
and mechanical allodynia (Wallin, G. et al., 1976). 6. In patients with CRPS II, but not in those with
Rekindling of spontaneous and evoked pain or hyperhidrosis, intraoperative electrical stimula-
enhancement of both by noradrenaline injected tion of the sympathetic chain may increase
into the skin area, which was painful before sym- spontaneous pain (Walker, A. E. and Nulsen, F.,
pathetic block or before successful treatment, took 1948; White, J. C. and Sweet, W. H., 1969).
10 min or longer to develop (Figure 4b). This long 7. In patients with SMP phentolamine (-adreno-
latency indicates that noradrenaline does not act ceptor blocker) given intravenously may relieve
possibly directly on the afferent fiber terminals, pain but not propranolol (-adrenoceptor blocker)
but by some unknown mechanism via changes in (Arner, S., 1991; Raja, S. N. et al., 1991; Dellemijn,
the micromilieu of the nociceptors. P. L. et al., 1994).
202 Autonomic Nervous System and Pain

controls 5 g noradrenaline
(a) (b)
50 10 20 40 min
Area of

temperature (C)
Skin surface
mechanical
allodynia

Thermal
0 stimulation

1000

amplitude (m)
Injection of Vibration

Indentation
noradrenaline 100

Area of
vasoconstriction
(pallor)
0

Pain threshold Pain threshold Warm/cold

uninjured side injured side difference limen
Figure 4 Mechanical and cold allodynia in CRPS type II patients with sympathetically maintained pain (SPM) elicited by
intradermal injection of noradrenaline. Noradrenaline was injected intradermally at the site of mechanical and cold allodynia
when the patients were temporarily relieved from their pain by a stellate ganglion block with lidocaine (1%) or bupivacaine
(0.25%). (a) Area of pallor and mechanical allodynia 20 min after intradermal injection of 2 mg noradrenaline. (b) Sequential
quantitative sensory testing of patients before and after intradermal injection of 5 mg noradrenaline during a time without
blockade of the stellate ganglion. This patient had mechanical allodynia (indentation amplitude of a mechanical vibration
stimulus to evoke pain about 90 mm, lower panel) and cold hyperalgesia/allodynia (cold pain threshold about 15  C, upper
panel). The mechanical allodynia (further decrease of indentation amplitude to elicit pain) and cold allodynia (further decrease
of cold pain threshold) progressively worsened starting approximately 20 min after intradermal injection of 5 mg noradrenaline.
Modified from Torebjork H. E. et al. (1995) with permission.

8. In some patients with postherpetic neuralgia, The interpretation of these data is that the nocicep-
spontaneous pain and mechanical allodynia tors are excited and possibly sensitized by
are enhanced after intracutaneous injection of noradrenaline released by the sympathetic fibers.
adrenaline or the 1-adrenoceptor agonist pheny- Either the nociceptors have expressed adrenoceptors
lephrine (Choi, B. and Robotham, M. C., 1997). or the excitatory effect is generated indirectly, e.g., via
changes in blood flow and subsequent ischemia or by
I recommend to read Mitchell S. W. (1872),
other components modulated by sympathetic fibers.
Livingston W. K. (1943/1976; here in particular
The ways of sympathetic afferent coupling to be dis-
chapters 5, 6, 14, and 15), and Bonica J. J. (1953; cussed are illustrated in Figure 6. Sympathetically
Chapter 28). These texts are not only of historical maintained activity in nociceptive neurons may gen-
interest (here the literature before 1953 is discussed) erate and maintain a state of central sensitization/
but also the reader will find (1) a wealth of carefully hyperexcitability, leading to spontaneous pain and sec-
reported clinical observations and (2) ideas devel- ondary evoked pain (mechanical and possibly cold
oped on the basis of these clinical observations that allodynia). This is schematically illustrated in Figure 7.
are still quite modern, showing that the progress in
this field is not as impressive as it appears to be.
These key experiments conducted on human patients
with SMP demonstrate that (1) activation of sympa- 17.3.2 Sympathetically Maintained Pain
thetic postganglionic neurons can produce pain, (2) Following Nerve Lesion Simulated in
blockade of sympathetic activity can relieve the pain, Behavioral Animal Models
(3) noradrenaline injected intracutaneously rekindles In animals (rats), signs of pain are measured by quan-
the pain, and (4) -adrenoceptor blockers or guanethi- tifiable behavioral components, e.g., paw-withdrawal
dine (which depletes noradrenaline from its stores) can latency to thermal noxious stimulation (heat and
relieve the pain ( Janig, W. and Baron, R., 2001; 2002; cold), frequency and threshold of paw withdrawal to
Janig, W., 2002; Janig, W. and Baron, R., 2003). mechanical stimulation, degree of self-mutilation
 Autonomic Nervous System and Pain 203

(a) SYMPATHETIC AFFERENT

(a)

Affected limb
3
300
Skin blood flow (PU)

mediated by
Activity- 2 adrenoceptors
dependent blood vessels
200
1

100
(b)
Unaffected limb
0 Nociceptive
Brady ?
kinin B2 PGE2 terminal
(b) Affected limb
Skin temperature (C)

34

29 (c)
Nerve Nociceptive
growth ?
Inflammatory terminal
24 Unaffected limb factor trkA
mediator(s)
0 40 80 0 40 80

Duration of cooling (%) Duration of warming (%)

(d) Adrenal
medulla
Sympathetic Nociceptive
High sympathetic activity Low sympathetic activity Adrenaline terminal
(c) Preganglionic
neurons

Figure 6 Ways hypothesized to couple sympathetic and

Dynamic primary afferent neurons following peripheral nerve lesion
mechanical (a) or during inflammation (bd). (a) These types of coupling
hyperalgesia depend on the activity in the sympathetic neurons and on
the expression of functional adrenoceptors by the afferent
neurons or mediation indirectly via the blood vessels (blood
flow). It can occur in the periphery (1), in the dorsal root
ganglion (3), or possibly also in the lesioned nerve (2). (b)
Figure 5 Experimental changes of activity in cutaneous The inflammatory mediator bradykinin acts at B2 receptors
sympathetic vasoconstrictor neurons by physiological in the membrane of the sympathetic varicosities or in cells
thermoregulatory reflex stimuli in 13 complex regional pain upstream of these varicosities, inducing release of
syndrome (CRPS) patients. With the help of a thermal suit, prostaglandin E2 (PGE2) and sensitization of nociceptors.
whole-body cooling and warming was performed to alter This way of coupling is probably not dependent on activity
sympathetic skin nerve activity. The subjects were lying in a suit in the sympathetic neurons. (c) Nerve growth factor
supplied by tubes, in which running water at 12  C and 50  C, released during an experimental inflammation reacts with
respectively (inflow temperature), was used to cool or warm the the high-affinity receptor trkA, in the membrane of the
whole body. By these means, sympathetic activity can be sympathetic varicosities, inducing release of an
switched on and off. (a) High sympathetic vasoconstrictor inflammatory mediator or inflammatory mediators and
activity during cooling induces considerable drop in skin blood sensitization of nociceptors. This effect is probably not
flow on the affected and unaffected extremity (laser Doppler dependent on activity in the sympathetic neurons. (d)
flowmetry). Measurements were taken at 5-min intervals Activation of the adrenal medulla by sympathetic
(mean SD). (b) On the unaffected side, a secondary decrease preganglionic neurons leads to release of adrenaline that
of skin temperature was documented. On the affected side, the generates sensitization of nociceptors. The ? in (b) and (c)
forearm temperature was clamped at 35  C by a feedback- indicates that PGE2 or other inflammatory mediators may
controlled heat lamp to exclude temperature effects on the be released by cells other than the sympathetic
sensory receptor level. Measurements were taken at 5-min varicosities. Modified from Janig W. and Habler H. J.
intervals (mean SD). (c) Effect of cutaneous sympathetic (2000b).
vasoconstrictor activity on dynamic mechanical hyperalgesia in
one CRPS patient with sympathetically maintained pain (SMP).
Activation of sympathetic neurons (during cooling) leads to an
increase of the area of dynamic mechanical hyperalgesia.
Reprinted with permission from Baron R. et al. (2002).
204 Autonomic Nervous System and Pain

Central sensitization
Central hyperexcitability

PNS C CNS
Inflammation: Amplification of
sensitization A, A physiological
impulse transmission
STIMULI PAIN
C
Nerve lesion: Changes in
ectopic impulses A , A central
representations

Sympathetic nervous system

Neuroendocrine systems
Figure 7 Concept of generation of peripheral and central hyperexcitability during inflammatory pain and neuropathic pain.
The upper interrupted arrow indicates that the central changes are generated (and possibly maintained) (a) by persistent
activation of nociceptors with C-fibers (e.g., during chronic inflammation) called here central sensitization or (b) after trauma
with nerve lesion by ectopic activity and other changes in lesioned afferent neurons called here central hyperexcitability. The
lower interrupted arrow indicates the efferent feedback via the sympathetic nervous system and neuroendocrine systems
(e.g., the sympatho-adrenal system). Primary afferent nociceptive neurons (in particular those with C-fibers) are sensitized
during inflammation. After nerve lesion, all lesioned primary afferent neurons (unmyelinated as well as myelinated ones)
undergo biochemical, physiological, and morphological changes that become irreversible with time. These peripheral
changes entail changes of the central representation (of the somato-sensory system) that become irreversible if no
regeneration of primary afferent neurons to their target tissue occurs. The central changes, induced by persistent activity in
afferent nociceptive neurons during inflammation or induced after nerve lesion, are also reflected in the efferent feedback
systems that may establish positive feedback loops to the primary afferent neurons.

(autotomy), or spontaneous lifting and licking of paw. (e.g., phentolamine (1,2), prazosin (1), or yohimbin
These reactions are interpreted as equivalent to (2)) or of adrenoceptor agonists (e.g., phenylephrine
mechanical or thermal hyperalgesia or allodynia or (1) and clonidine (2)), and intraperitoneal applica-
spontaneous pain in humans. Behavioral models allow tion of guanethidine (that is taken up by the
to test whether the sympathetic nervous system noradrenergic terminals and depletes noradrenaline).
contributes to these hyperalgesic/allodynic behaviors Overall, the results obtained on these rat beha-
or to spontaneous pain behavior. Animals are given vioral models of nerve injury are somewhat
clinically relevant nerve lesions, and behavioral disappointing since they did not lead to straightfor-
experiments are performed before and after interven- ward answers. Thus, the controversial results on
tions aimed at the sympathetic supply of the affected these models provide conflicting evidence on the
extremity. The strength of these models includes role of the sympathetic nervous system in generat-
the range of possible experimental manipulations ing pain behavior, indicating that behavioral animal
performed at the sympathetic nervous system models must be designed with utmost care. Small
and the use of different strains of animals or trans- changes in the experimental procedures may create
genic animals. Quantitative data obtained in these major behavioral changes. Behavioral testing should
behavioral experiments lead to the formulation of be performed in a blinded fashion if possible, and
hypotheses that can be tested in reduced experiments the models must be designed in close association
in vivo and in vitro using various methodical with the clinical situation to be certain to test what
approaches. is intended to be tested. The results of systemic
Several rat models of neuropathic pain involving pharmacological interventions (e.g., application of
controlled nerve lesions have been used to determine adrenoceptor agonists or antagonists, guanethidine,
the contribution of the sympathetic nervous system to or chemical sympathectomy with 5-hydroxy-dopa-
neuropathic pain. The results obtained in these mine) do not necessarily allow functional
experiments are summarized in Table 2. The inter- interpretations, partly because of the widespread
ventions include surgical or chemical sympathectomy, systemic effects of these agents. Finally, the con-
systemic or local application of adrenoceptor blockers troversial results obtained from the animal behavior
 Autonomic Nervous System and Pain 205

Table 2 Nerve lesion animal (rat) models to study sympathetically maintained pain (SMP)

Behavioral Interventions at the

Model Nerve lesion measurements sympathetic supply Results

1. Autotomy modela Ligation and Autotomy of Intraperitoneal Autotomy #

transection of sciatic affected paw guanethidine
and saphenous nerve (self-mutilation)
Subcutaneous Autonomy "
noradrenaline
2. Partial lesion of Ligation of 3350% of Heat HA/ Intraperitoneal Heat HA #, Mechanical
the sciatic nerveb sciatic nerve mechanical HA guanethidine, HA , Heat HA ",
Noradrenaline local Mechanical HA "
3. Chronic Loose ligation of sciatic Heat HA/ Surgical Heat HA #, Mechanical
constriction nerve mechanical HA sympathectomy, HA (#),
injury of the intraperitoneal
sciatic nervec guanethidine
4. Spinal nerve Ligation and Mechanical HA/ Surgical Mechanical HA # or f
ligation modeld transection of spinal allodynia heat sympathectomy heat HA #
nerve L5, L5/L6 HA
5. Cryolysis of the Cryolysis of sciatic Mechanical Surgical Mechanical allodynia
sciatic nervee nerve allodynia sympathectomy
a
Wall P. D. et al. (1979a; 1979b); Coderre T. J. et al. (1984; 1986); Coderre T. J. and Melzack R. (1986).
b
Seltzer Z. et al. (1990); Shir Y. and Seltzer Z. (1991); Tracey D. J. et al. (1995).
c
Bennett G. J. and Xie Y. K. (1988); Neil A. et al. (1991); Desmeules J. A. et al. (1995); Kim K. J. et al. (1997).
d
Kim S. H. and Chung J. M. (1991); Kim S. H. et al. (1993); Kim K. J. et al. (1997); Kinnman E. and Levine J. D. (1995); Ringkamp M. et al.
(1999).
e
Willenbring S. et al. (1995).
f
The negative effects of surgical sympathectomy reported by Ringkamp M. et al. (1999: Pain 79, 142) were obtained independently by two
different groups.
HA, hyperalgesia; #, decrease; ", increase; , no effect.

models may argue that the models do not reflect deep somatic tissues in mammals. The effects that
SMP as defined clinically. have been measured under experimental conditions
This controversial situation does not mean that on sensory receptors with myelinated or unmyeli-
behavioral animal models are useless and cannot nated axons were weak and can in part be explained
simulate the clinical situation. On the contrary, we by changes of the effector organs (erector pili muscles
are dependent on the design of these animal models if and blood vessels) induced by the activation of sym-
we want to unravel the mechanisms behind the dif- pathetic neurons. These rather negative results do
ferent types of pathological pain in which the not rule out that noradrenaline or colocalized sub-
sympathetic nervous system is involved. However, stances released by the postganglionic terminals have
the same model cannot be expected to represent secondary long-term effects on the excitability of
SMP in different groups of patients. Furthermore, sensory receptors although we have no experimental
the animal models so far tested are related to nerve evidence for this ( Janig, W. and Koltzenburg, M.,
lesions. Rat models simulating SMP in patients with 1991).
pain states developing after trauma without nerve
lesion (Table 1) practically do not exist (e.g., for
patients with CRPS I). 17.3.3.1 Coupling between lesioned
postganglionic and afferent nerve
terminals
Coupling may occur between sympathetic fibers and
17.3.3 Direct Involvement of the
afferent terminals in a neuroma, following nerve sec-
Sympathetic Nervous System in the
tion or ligation. Some myelinated as well as
Generation of Pain Following Nerve Trauma
unmyelinated nerve fibers in the neuroma can be
Under physiological conditions, there exists almost excited following electrical stimulation of the sym-
no acute influence of sympathetic postganglionic pathetic supply or by noradrenaline or adrenaline
neurons on sensory neurons projecting to skin or injected systemically. The coupling has been
206 Autonomic Nervous System and Pain

observed in young neuromas but less so in old ones, afferent axons that project through the lesion site
weeks and months after nerve lesion. Furthermore, and nonlesioned postganglionic axons. It is suggested
the frequency of electrical stimulation applied to the that the unlesioned unmyelinated afferents develop
sympathetic trunk in order to excite the lesioned hyper-reactivity to catecholamines following degen-
afferent nerve fibers was high, generating activity in eration of the sympathetic postganglionic axons.
the sympathetic neurons that does not occurs under Expression of adrenoceptors in afferent fibers may
physiological conditions ( Janig, W., 2006). This cor- be triggered by collateral sprouting of both afferent
responds to the clinical experience showing that fibers and postganglionic fibers in the target tissue
neuroma pain is usually not dependent on sympa- (Sato, J. and Perl, E. R., 1991). However, sympathetic
thetic activity. It also corresponds to histological afferent coupling leading to activation or sensitiza-
observations showing that catecholamine-containing tion of nociceptors requires sympathetic stimulation
axon profiles are rare within the neuroma and for at high nonphysiological frequencies.
several centimeters proximal to it but may be present
in the scar tissue around the neuroma), many weeks 17.3.3.3 Coupling in the dorsal root
after cutting and ligating the nerve (McLachlan, E. ganglion and collateral sprouting following
M., unpublished data). Thus, this coupling is peripheral nerve lesion
chemical and occurs via noradrenaline acting on Chemical sympathetic afferent coupling following
-adrenoceptors, although other mediator substances peripheral nerve lesion (e.g., a spinal nerve, the scia-
may also be involved (Janig, W., 1990). Ephaptic tic nerve, or another hindlimb nerve) may occur
coupling between sympathetic fibers and afferent proximally to the injury site in the nerve or in the
fibers has so far not been observed in a neuroma DRG (Figure 6a3). Sympathetic postganglionic fibers
(Devor, M. and Janig, W., 1981; Blumberg, H. and reach the spinal nerves via gray rami. Most of these
Janig, W., 1982; 1984; Janig, W. and Koltzenburg, M., fibers project distally to peripheral target cells, and
1991). others project proximally (e.g., to the DRG) and are
The situation is different when afferent and sym- normally found along blood vessels. This situation
pathetic fibers are allowed to regenerate to the target changes after an experimental nerve lesion (e.g.,
tissue. This has been shown experimentally for the transection and ligation of the sciatic nerve in rats).
chronic situation more than a year after cross-union Now many perivascular catecholamine-containing
of nerves (e.g., adaptation of the proximal stump of axons of the unlesioned proximally projecting neu-
the sural or superficial peroneal nerve to the distal rons start to sprout in the DRGs that contain somata
stump of the tibial nerve, in the cat) and after rein- with lesioned axons. The extent of this collateral
nervation of appropriate and inappropriate target sprouting increases with time after nerve lesion.
tissues. Now unmyelinated afferent fibers may be Some somata are partially or almost completely sur-
vigorously excited by electrical low-frequency sti- rounded by varicose catecholaminergic terminals
mulation of the sympathetic supply (Habler, H. J. several weeks after nerve lesion; the frequency of
et al., 1987). Also this excitation is adrenoceptor- these catecholamine-fluorescent structures increases
mediated (Figure 8). It can be mimicked by adrena- for more than 70 days after nerve lesion. The nora-
line or noradrenaline and blocked by -adrenoceptor drenergic axons sprout preferentially to axotomized
blockers (e.g., phentolamine). large-diameter afferent cell bodies (McLachlan, E.
M. et al., 1993; Chapter Sprouting in Dorsal Root
17.3.3.2 Coupling between unlesioned Ganglia).
postganglionic and afferent nerve Neurophysiological experiments on rats with
terminals following partial nerve lesion nerve lesion (ligation and transection of a spinal
Intact C-fiber polymodal nociceptors in the skin may nerve, the sciatic nerve, or other hindlimb nerves)
develop sensitivity to catecholamines following par- show that afferent neurons projecting in the lesioned
tial nerve injury. Sympathetic nerve stimulation and nerve can be excited or depressed in their activity
noradrenaline may excite the polymodal nociceptors affected via the DRG by electrical stimulation of
or sensitize them for heat stimuli. The activation and sympathetic neurons and by catecholamines: (1) In
sensitization is already seen 410 days after partial the first 2 to 3 weeks after nerve lesion, most A-fiber
nerve lesion and is maintained for at least 150 days. neurons with spontaneous activity and only very few
This sympathetic afferent coupling apparently silent A-fiber neurons are excited. At later time per-
involves the nonlesioned polymodal nociceptive iods, when the catecholaminergic sprouting in the
 Autonomic Nervous System and Pain 207

(a) (c) 1 Hz
pre post lesion

2 Hz

stim LST rec afferent stim neuroma

3 Hz
(b)
stim neuroma
4 Hz

* 5 Hz
100 V LST 100 V
5 ms 5s

(d) mm Hg (e)
imp
160 400

140 200

imp
50
5

0 0
1 min 0 5 Hz 10
5 g adrenaline i.v. 0.2 g angiotensin i.v.
Figure 8 Sympathetic afferent coupling after nerve lesion with subsequent fiber regeneration. Excitation of unmyelinated
afferent fibers by electrical stimulation of sympathetic fibers following nerve injury. Unmyelinated primary afferents were
recorded in cats 1120 months following a nerve lesion. The central cut stump of a cutaneous nerve innervating hairy skin
(sural or superficial peroneal nerve) had been imperfectly adapted to the distal stump of a transected mixed nerve (tibial
nerve). This preparation was designed to mimic the consequences of lesion of a mixed nerve. A neuroma-in-continuity at the
lesion site was present and cutaneous nerve fibers had regenerated into skin and deep somatic tissue supplied by the mixed
nerve. (a) Experimental setup. post, postganglionic; pre, preganglionic; LST, lumbar sympathetic trunk. (b) The afferent fibers
were identified as unmyelinated by electrical stimulation of the neuroma with single impulses. The signal indicated by the dot
was recorded from the same afferent fiber as in (c) and (d); the afferent fiber conducted at 1.3 m s1. (c) Record from a single
unmyelinated afferent unit. Supramaximal stimulation of the LST with trains of 30 pulses at 15 Hz (trains and stimulation
artifacts indicated by bars). Note that the afferent unit had some low rate of ongoing activity (impulses before the trains at 1
and 4 Hz) and a second fiber was activated at 5 Hz stimulation (marked by an asterisk). (d) Adrenaline (5 mg injected i.v.)
activated the fiber. Angiotensin (0.2 mg injected i.v.) generated a large increase of blood pressure (MAP, mean arterial blood
pressure) but did not activate the afferent fiber. (e) Stimulus response curves for the single unit (&) and four filaments
containing two to three (l, N /H ) or more than five (&) afferent units. Ordinate scale is the total number of impulses exceeding
ongoing activity in response to variable stimulation frequency of the LST. Modified from Habler H. J. et al. (1987).

DRG is more prominent, most spontaneously active fibers (Michaelis, M. et al., 2000). (4) Activation of
A-fiber neurons are inhibited during stimulation of the afferent neurons, via the DRG, requires high-
the sympathetic supply but rarely excited. (2) Only frequency stimulation of the sympathetic neurons at
very few afferent neurons with unmyelinated axons 5 to 20 Hz. Sympathetic pre- and postganglionic
respond to electrical stimulation of the sympathetic neurons rarely discharge at these frequencies under
trunk; most of them are inhibited in their activity physiological conditions. (5) Mechanical allodynic
(McLachlan, E. M. et al., 1993; Devor, M. et al., 1994; and hyperalgesic behavior shown by rats with L5
Michaelis, M. et al., 1996). (3) The coupling from spinal nerve injury (transection and ligation) is not
sympathetic postganglionic fibers to the afferent cell dependent on the innervation of the DRG by sympa-
bodies in the DRG probably occurs only to sponta- thetic neurons (Ringkamp, M. et al., 1999; see point 4
neously active muscle afferent neurons with A- in Table 2). The rate of ectopic activity originating in
208 Autonomic Nervous System and Pain

the DRG, 15 to 45 days after spinal nerve injury, is useful in the design of more specific treatment mod-
not maintained by activity in sympathetic neurons alities for neuropathic pain conditions involving
(Liu, X. et al., 2000). (6) Afferent neurons in the DRG sympathetic efferent activity (Janig, W. et al., 1996;
may be activated indirectly by ischemia generated by Janig, W. and Habler, H. J., 2000b).
decrease of blood flow during vasoconstriction in the
DRG following electrical stimulation of the sympa- 17.3.3.5 Synopsis
thetic trunk and not directly via adrenoceptors Peripheral trauma with nerve injury may lead to
expressed in the DRG cells (Habler, H.-J. et al., sensitization and activation of nociceptive and acti-
2000). (7) Axotomized medium- to large-diameter vation of other primary afferent neurons. These
DRG neurons express 2A-adrenoceptors after scia- processes may depend on and are maintained by the
tic nerve injury (Birder, L. A. and Perl, E. R., 1999). sympathetic nervous system. Several ways of cou-
However, these upregulated adreceptors disappear pling between sympathetic postganglionic neurons
after several weeks as perineural catecholaminergic and primary afferent neurons are possible and have
sprouting develops and only small depolarizations been worked out experimentally on animal models
can be generated by high concentrations of noradre- with controlled nerve lesions, showing that there may
naline in axotomized DRG somata (Lopez de develop intimate relationships between sympathetic
Armentia, M. et al., 2003). and afferent neurons under pathophysiological con-
The mechanism leading to the collateral sprout- ditions (Figure 9):
ing of postganglionic noradrenergic fibers in the
1. The sympathetic postganglionic neuron may
DRG is possibly related to neurotrophic signals and
develop chemically mediated cross talk to primary
their receptors generated by afferent cells with
afferent neurons. Whether this occurs probably
lesioned axons in the DRG and the surrounding
depends on the time after nerve lesion as well as
satellite glia cells (see Chapter Sprouting in Dorsal
on the type of nerve lesion (partial or complete).
Root Ganglia). Whether these aberrant pathological
2. Following peripheral nerve lesion, remote collat-
connections can account for spontaneous pain and
eral sprouting of unlesioned postganglionic fibers
allodynia in some patients after peripheral nerve
occurs in the DRG, preferentially toward the
lesions is doubtful and awaits further investigations
large-diameter sensory cells. Collateral sprouting
( Janig, W. et al., 1996).
of unlesioned postganglionic fibers may also occur
in the peripheral target tissue (in particular after
17.3.3.4 Adrenoceptors involved in
partial nerve lesion).
chemical sympathetic afferent coupling
3. The transmission from sympathetic neurons to
following nerve lesion
afferent neurons is mediated by noradrenaline,
Excitation or depression of lesioned primary afferent
but additional mediator substances cannot be
neurons (in the DRG, in their lesioned terminals in
excluded.
the neuroma or in their regenerating sprouts), or of
4. Primary afferent neurons express functional adre-
unlesioned collaterally sprouting primary afferents
noceptors. The type of adrenoceptor involved is
generated by activation of the sympathetic innerva-
preferentially 2 in animal models.
tion, is mimicked by systemic injection of
5. The signals that initiate the sprouting and the
noradrenaline or adrenaline and prevented by block-
functional expression of adrenoceptors in the
ade of -adrenoceptors (e.g., phentolamine). Thus,
afferent neurons may be related to neurotrophic
both excitation and depression are suggested to be
substances that derive from the Schwann cells or
mediated by -adrenoceptors. The cellular mechan-
other cells. Their upregulation is related to the
isms underlying the increased sensitivity are
plastic changes of the primary afferent and sym-
unknown. Novel expression or upregulation of adre-
pathetic postganglionic neurons and possibly to
noceptors occurs; alternatively, normally present
decrease of the density of the noradrenergic inner-
adrenoreceptors that are not functional become
vation and/or to activity in the afferent and
uncovered and effective during the response to
sympathetic neurons (see Chapter Sprouting in
damage. The subtype of -adrenoreceptor being
Dorsal Root Ganglia).
involved in the sympatheticafferent coupling in
the different rat models is predominantly 2 (Chen, Y. These plastic changes of primary afferent and sym-
et al., 1996). Knowledge about the subtypes of adre- pathetic postganglionic neurons following peripheral
noceptor following nerve trauma may turn out to be trauma with nerve lesion may explain some of the
 Autonomic Nervous System and Pain 209

Production/uncovering
LESION
of -adrenoceptors
dependent on:
Biochemical signals,
Activity?

NAd

NAd
Postganglionic

Preganglionic
Collateral sprouting
dependent on:
Biochemical signals,
Activity?
Figure 9 Relation between afferent and sympathetic neurons following peripheral nerve lesion. Collateral sprouting of
unlesioned sympathetic neurons in the dorsal root ganglion (DRG) and in the peripheral target tissue. Upregulation or
uncovering of functional adrenoceptors () by afferent neurons after nerve lesion. It is unclear in which way these processes
are related to the biochemical signals (e.g., neurotrophins) synthesized by neurons, Schwann cells, and other cells in the DRG
and the expression of their receptors. In which way are these processes dependent on activity in the afferent neurons, on the
presence/absence of postganglionic noradrenergic neurons, or on the activity in the postganglionic neurons? NAd,
noradrenaline. Modified from Janig W. et al. (1996).

clinical sensory phenomena in patients with SMP, neuropeptides from the receptive terminals (e.g., sub-
e.g., in CRPS type II. However, they do not explain stance P (SP) and calcitonin-gene-related peptide
(1) the slow development of spontaneous pain and (CGRP)). Both afferent-induced changes are called
allodynia after noradrenaline is injected into the neurogenic inflammation (McDonald, D. M., 1997;
skin that was painful before sympathetic block or Holzer, P., 1998). Thus, there are possibilities for
before sympathectomy (Torebjork, H. E. et al., 1995) indirect coupling between sympathetic and afferent
(Figure 4); (2) the long-lasting pain relief in patients nerve terminals (Figure 10): (1) Vascular perfusion of
with SMP after sympathetic blocks Figure 15 and (3) the micromilieu surrounding the nociceptors after
the mechanism of SMP in patients without nerve nerve trauma may change as consequences of dener-
lesion, e.g., in patients with CRPS type I. vation and reinnervation by postganglionic
vasoconstrictor neurons and afferent nociceptive
neurons and hyper-reactivity of blood vessels to
17.3.4 Indirect Involvement of the nerve impulses and circulating catecholamines may
Sympathetic Nervous System in the develop (Jobling, P. et al., 1992; Koltzenburg, M. et al.,
Generation of Pain 1995). (2) Nociceptors may be sensitized by inflam-
matory mediators such as prostaglandins and
Nociceptive afferents are embedded in a complex
interleukins, the release of which from nonneuronal
micromilieu (Figure 10). The state of this micromi-
cells of the micromilieu may be mediated by nora-
lieu surrounding the receptive terminals depends on
drenergic nerve terminals.
mediator substances that are released during inflam-
matory processes following trauma from non-neural
cells such as mast cells, polymorphonuclear leuco-
17.3.4.1 Changes of neurovascular
cytes, macrophages, fibroblasts, endothelial cells, or
transmission and development
other cells. The microcirculation is under neural
of hyperreactivity of blood vessels
control of sympathetic vasoconstrictor neurons.
Neural control of blood vessels can change dramati-
Moreover, activation of subgroups of nociceptive
cally after trauma with nerve lesion, but possibly also
primary afferents causes not only orthodromic
after trauma without (Koltzenburg, M. et al., 1995):
impulse traffic but also arteriolar (precapillary) vaso-
dilation and (in most tissues, but not in human skin) 1. Cutaneous blood vessels that are reinnervated
venular plasma extravasation by release of after a nerve lesion exhibit stronger than normal
210 Autonomic Nervous System and Pain

(a)
Sympathetic
1 fibers MP 5

Varocosities

5
A? Capillaries MP
Receptive
Afferent 2 endings
fibers 4
C

3
Venules
Arterioles 4
MC

(b)

SP
Afferent
2
5

1
NAd
MP
TNF- Sympathetic
IL-1

NAd
3 SP

NK receptors

BV NAd Adrenoceptors
Figure 10 (a) The micromilieu of nociceptors. The microenvironment of primary afferents is thought to affect the properties
of the receptive endings of myelinated (A) and unmyelinated (C) afferent fibers. This has been particularly documented for
inflammatory processes, but one may speculate that pathological changes in the direct surroundings of primary afferents may
contribute to other pain states as well. The vascular bed consists of arterioles (directly innervated by sympathetic and afferent
(A, C) fibers), capillaries (not innervated and not influenced by nerve fibers), and venules (not directly innervated but
influenced by nerve fibers). The micromilieu depends on several interacting components: Neural activity in postganglionic
noradrenergic fibers (1) supplying blood vessels (3, BV) causes release of noradrenaline (NAd) and possibly other substances
causing vasoconstriction. Excitation of primary afferents (A- and C-fibers) (2) causes vasodilation in precapillary arterioles
and plasma extravasation in postcapillary venules (C-fibers only) by the release of SP and other vasoactive compounds (e.g.,
calcitonin gene-related peptide, CGRP). Some of these effects may be mediated by nonneuronal cells such as mast cells
(MC, 4) and macrophages (MP, 5). Other factors that affect the control of the microcirculation are the myogenic properties of
arterioles (3) and more global environmental influences such as a change of the temperature and the metabolic state of the
tissue (modified from Janig, W. and Koltzenburg, M., 1991). (b) Hypothetical relation between sympathetic noradrenergic
nerve fibers (1), peptidergic afferent nerve fibers (2), blood vessels (3, BV), and macrophages (4, MP). The activated and
sensitized afferent nerve fibers activate macrophages (via SP release). The immune cells start to release cytokines, such as
tumor necrosis factor  (TNF-) and interleukin 1 (IL-1), which further activate afferent fibers by enhancing sodium influx into
the cells. SP (and CGRP) released from the afferent nerve fibers reacts with neurokinin 1 (NK1) receptors (CGRP receptors) in
the blood vessels (arteriolar vasodilation, venular plasma extravasation; neurogenic inflammation). The sympathetic nerve
fibers interact with this system potentially on three levels: (1) via adrenoceptors (mainly ) on the blood vessels
(vasoconstriction), (2) via adrenoceptors (mainly ) on macrophages (further release of cytokines), and (3) via adrenoceptors
(mainly ) on afferents (further sensitization of these fibers). Modified from Janig W. and Baron R. (2003).
 Autonomic Nervous System and Pain 211

vasoconstriction to impulses in sympathetic Sham sympathectomy

neurons. 120
Sympathectomized
2. The sympathetically reinnervated cutaneous Decentralization
blood vessels show stronger than normal vasocon-
strictions to systemic catecholamines and appear

Paw-withdrawal threshold (g)

100
to be hyper-reactive.
3. Blood vessels exhibit changes in vasodilation to * *
antidromic activation of reinnervated peptidergic
unmyelinated afferents. 80 Decentralization
The reinnervated blood vessels may therefore be
Surgical
under stronger than normal vasoconstrictor influence sympathectomy

that can no longer be counteracted by an afferent- Skin

mediated vasodilation (see Habler, H. J., et al., 1997). 60

Nociceptive
In vitro investigation of the rat tail artery has shown afferents

that the functional recovery of neurovascular trans-

Baseline
mission is permanently disturbed after a nerve lesion threshold
40
( Jobling, P. et al., 1992).
The altered neural and nonneural control of blood 1 0 1 2 3
vessels, following trauma with nerve lesion, can con- Log (bradykinin) ng
tribute to the abnormal regulation of blood flow Figure 11 Mechanical hyperalgesic behavior and
through skin and deep somatic tissues and possibly sympathetic innervation. Cutaneous nociceptors on the
to the trophic changes (including the edema) that are dorsum of the paw are stimulated by a linearly increasing
seen in patients with CRPS. They may furthermore mechanical force. Paw-withdrawal threshold (PWT) is
defined as the mean ( SEM) minimum force (in grams,
be a permissive factor in the generation of afferent ordinate scale) at which the rat withdraws its paw. Abscissa
nociceptive impulse activity and therefore in the scale: log dose of bradykinin (BK, in nanograms) injected in
sensitization of nociceptors and in the generation of a volume of 2.5 ml of saline into the dermis of the skin.
pain. Decrease of PWT to mechanical stimulation induced by
Finally, it is commonly believed that skin tem- intradermal injection of BK in sham-sympathectomized
(control) rats (l, 6 hindpaws), in surgically
perature changes observed in painful disorders sympathectomized rats (lumbar paravertebral ganglia L2
following trauma with or without nerve lesion (e.g., L4 removed 8 days before; &, 12 hindpaws), and in rats with
in CRPS I or II) reflect changes of activity in cuta- decentralized lumbar sympathetic chains (white rami with
neous vasoconstrictor neurons, in the sense that cold preganglionic axons to paravertebral ganglia L2 and L3 and
skin is associated with a high level and warm skin chain rostral to ganglion L2 transected 8 days before; ^, 10
paws). Abscissa scale: log dose of BK (in nanograms).
with a low level of activity in these neurons. This is Sham sympathectomy and sympathetic decentralization
probably a misconception, and there is no proof for were both significantly different from sympathectomy
this relation between skin temperature and activity in groups at doses of BK of 102 and 103 ng (, P < 0.01).
sympathetic neurons under these pathophysiological Modified from Khasar S. G. et al. (1998a).
conditions.

single injection of bradykinin, this decrease lasts for

17.3.4.2 Sensitization of nociceptors more than 1 h for mechanical stimulation. This type
mediated by sympathetic terminals of mechanical hyperalgesic behavior is mediated by
independent of excitation and release of the B2 bradykinin-receptor and is not present when
noradrenaline bradykinin is injected subcutaneously (Khasar, S. G.
Withdrawal threshold to stimulation of the rat hind- et al., 1993; 1995; Janig, W. and Habler, H. J., 2000b).
paw with a linearly increasing mechanical stimulus In normal rat, bradykinin-induced hyperalgesic
applied to the dorsum of the paw decreases dose- behavior is blocked by the cyclooxygenase inhibitor
dependently after intradermal injection of the indomethacin and therefore mediated by a prosta-
inflammatory mediator bradykinin (an octapeptide glandin (probably PGE2) that sensitizes nociceptors
cleaved from plasma 2-globulins, by kallikreins cir- for mechanical stimulation. However, in vagoto-
culating in the plasma) (Figure 11). Following a mized rats in which the bradykinin-induced
212 Autonomic Nervous System and Pain

mechanical hyperalgesia is significantly enhanced experiments in the cat, not yet in humans for cuta-
(by activation of the adrenal medulla, see below and neous pain (Manning, D. C. et al., 1991); (4) in skin
Figure 13a), indomethacin has almost no effect on there is a spatial dissociation between afferent noci-
bradykinin-induced hyperalgesia. This failure is not ceptive fibers and sympathetic fibers, many of the
related to a switch from B2- to B1-receptor subtype former ending in the epidermis and subepidermal
because the selective B1-receptor agonist des-Arg9- plexus, whereas most of the latter terminate in dee-
bradykinin fails to produce hyperalgesia in vagoto- per dermal layer, raising the question as to the nature
mized rats (Khasar, S. G. et al., 1998a). The same of the interaction between both groups of fibers.
applies to rats that were sound-stressed for 4 days These points argue that the dependence of brady-
(Khasar, S. G. et al., 2005). Thus, bradykinin-induced kinin-induced cutaneous mechanical behavior on
hyperalgesia may not be mediated by prostanoids in the sympathetic innervation has to be reinvestigated
vagotomized or sound-stressed rats. Furthermore, using a rigorous experimental approach. Neurophy-
convincing sensitization to mechanical stimulation siological experiments are required to demonstrate
by bradykinin has only been demonstrated for afferent that sensitization of nociceptors by bradykinin is
fibers innervating the cat knee joint (Neugebauer, V. dependent on the presence of sympathetic terminals.
et al., 1989) or the cat skeletal muscle (Mense, S. and
Meyer, H., 1988). Manning D. C. et al. (1991) demon- 17.3.4.3 Sensitization of nociceptors
strated in humans that intradermal injection of initiated by nerve growth factor or
bradykinin (100 ng to 10 mg) generates hyperalgesia cytokines possibly mediated by
to heat stimulation but not to mechanical stimulation. sympathetic terminals
The decrease in paw-withdrawal threshold to 17.3.4.3.(i) Nerve growth factor Systemic injec-
mechanical stimulation generated by intradermal tion of nerve growth factor (NGF) is followed by a
injection of bradykinin is significantly reduced after transient thermal and mechanical hyperalgesia in rats
surgical sympathectomy. Decentralization of the (Lewin, G. R. et al., 1993; 1994) and humans (Petty, B.
lumbar paravertebral sympathetic ganglia (denervat- G. et al., 1994). During experimental inflammation
ing the postganglionic neurons by cutting the (evoked by Freunds adjuvant in the rat hindpaw),
preganglionic sympathetic axons) does not abolish NGF increases in the inflamed tissue paralleled by
the bradykinin-induced mechanical hyperalgesic the development of thermal and mechanical hyper-
behavior (Figure 11). This indicates that the sensitiz- algesia. Both are prevented by anti-NGF antibodies
ing effect of bradykinin is not dependent on activity in (Lewin, G. R. et al., 1994). The mechanisms respon-
the sympathetic neurons innervating skin and there- sible are sensitization of nociceptors via high-affinity
fore not on release of noradrenaline (Khasar, S. G. NGF-receptors (trkA receptors) and an increased
et al., 1998a) and that bradykinin stimulates the release synthesis of CGRP and SP in the afferent cell bodies
of prostaglandin from the sympathetic terminals or induced by NGF taken up by the afferent terminals
from other cells in association with the sympathetic and transported to the cell bodies. The NGF-
terminals in the skin (Figure 6b). induced sensitization of nociceptors or part of it
Mechanical hyperalgesic behavior generated by seems to be mediated indirectly by the sympathetic
intracutaneous injection of bradykinin and its depen- postganglionic terminals. In rats, heat and mechanical
dence on the sympathetic innervation of the skin is hyperalgesic behavior generated by local injection of
an interesting phenomenon. However, that mechan- NGF into the skin is prevented or significantly
ical hyperalgesia is due to prostanoids sensitizing reduced after chemical or surgical sympathectomy
nociceptors since indomethacin prevents it appears (Woolf, C. J. et al., 1996). These experiments suggest
to be too simple. The reasons are (1) the finding that that the NGF released during inflammation by
sympathetic fibers mediate this hyperalgesia inde- inflammatory cells acts on the sympathetic terminals
pendent of neural activity and release of via high-affinity trkA receptors, inducing the release
noradrenaline; (2) indomethacin does not block this of inflammatory mediators (the source of which is
behavior under certain conditions (e.g., when the unknown) and subsequently sensitization of nocicep-
adrenal medullae are activated after vagotomy (see tors for mechanical and heat stimuli (Figure 6c;
below) or after sound stress); (3) sensitization of McMahon, S. B., 1996; Woolf, C. J., 1996; Janig, W.
nociceptive afferents innervating deep somatic tis- and Habler, H. J., 2000b). It is unclear whether this
sues by bradykinin to mechanical stimulation sensitization of nociceptors mediated by terminal
has been demonstrated in neurophysiological sympathetic nerve fibers (1) is dependent on activity
 Autonomic Nervous System and Pain 213

in the sympathetic neurons and release of noradrena- Stimulus

line and on adrenoceptors expressed in the (Pathogenic challenge)
nociceptive afferent neurons or (2) is independent
Resident cells
of neural activity and release of noradrenaline.
Bradykinin
17.3.4.3.(ii) Proinflammatory cytokines Based on other
behavioral experiments conducted on rats (studying
*TNF-#
mechanical and heat hyperalgesia), it has been shown
that tissue injury, injection of the bacterial cell wall *IL-I# *IL-8
endotoxin lipopolysaccharide, or injection of carra- IL-6
geenan (a plant polysaccharide) stimulates tissue
inflammation and leads to sensitization of nocicep- *COX-2# Symp. terminal
tors. Systematic pharmacological interventions using Eicosanoids
blockers or inhibitors of the various mediators
demonstrate that the proinflammatory cytokines, Nociceptive terminal
tumor necrosis factor  (TNF-), interleukin-1
(IL-1), IL-6, and IL-8 may be involved in this process
of nociceptor sensitization and therefore in the gen- Cytokines and inflammatory
eration of hyperalgesia (Woolf, C. J. et al., 1996; 1997; hyperalgesia
Poole, S. et al., 1999). Pathogenic stimuli lead to (Sensitization of nociceptors)
activation of resident cells and release of Figure 12 Hypothetical role of cytokines in sensitization of
the inflammatory mediator bradykinin and other nociceptors during inflammation and the underlying putative
mediators. The inflammatory mediators and the peripheral mechanisms leading to hyperalgesia. Pathogenic
stimuli activate resident cells and lead to release of
pathogenic stimuli themselves activate macrophages, inflammatory mediators (such as bradykinin).
monocytes, and other immune-related cells that Proinflammatory cytokines are synthesized and released by
release TNF-alpha. Nociceptors are believed to be macrophages and other immune or immune-related cells.
sensitized by two possible pathways (Figure 12): (1) Nociceptors are postulated to be sensitized by two pathways
TNF- induces production of IL-6 and IL-1 by involving the cytokines: (1) Tumor necrosis factor  (TNF-)
induces synthesis and release of interleukin 1 (IL-1) and IL-6
immune cells, whereby IL-6 enhances the production which, in turn, induce the release of eicosanoids
of IL-1. These interleukins stimulate cyclooxygen- (prostaglandin E2 and I2 (PGE2, PGI2)) by activating
ase 2 (COX 2) and the production of prostaglandins cyclooxygenase-2 (COX-2). (2) TNF- induces synthesis and
(PGE2, PGI2) that in turn react with the nociceptive release of IL-8. IL-8 activates sympathetic terminals that
terminal via E-type prostaglandin receptors. (2) sensitize nociceptors via 2-adrenoceptors. Glucocorticoids
inhibit the synthesis of the cytokines and the activation of
TNF- induces the release of IL-8 from endothelial COX-2 (indicated by asterisks). Anti-inflammatory cytokines
cells and macrophages. IL-8 reacts with the sympa- (such as IL-4 and IL-10) that are also synthesized and
thetic terminals that are supposed to mediate released by immune cells inhibit the synthesis and release of
sensitization of nociceptive afferent terminals by proinflammatory cytokines (indicated by #). This scheme is
release of noradrenaline to act via 2-adrenoceptors. fully dependent on behavioral experiments and
pharmacological interventions. The different steps will need
These two peripheral pathways by which nociceptive to be verified experimentally using neurophysiological
afferents are hypothesized to be sensitized involving experiments. Modified from Poole S. et al. (1999).
cytokines are under inhibitory control of circulating
glucocorticoids (indicated by asterisks in Figure 12)
and of other, anti-inflammatory, interleukins (e.g., fibers on the afferent nerve fibers are lacking. These
IL-4 and IL-10 indicated by # in Figure 12). experiments have to be done.
It is important to emphasize that the mechanisms of
sensitization of nociceptive afferents, involving NGF,
proinflammatory cytokines, and noradrenergic sym- 17.3.4.4 Sympatho-adrenal system and
pathetic postganglionic fibers, have been deduced on nociceptor sensitization
the basis of behavioral and pharmacological experi- Activity of the sympatho-adrenal system (adrenal
ments. Proof of such interaction by directly assessing medulla) is regulated by central circuits that are
the activity of nociceptors with electrophysiological different from those regulating other sympathetic
techniques and of the effect of noradrenergic nerve systems (Morrison, S. F. and Cao, W. H., 2000;
214 Autonomic Nervous System and Pain

Morrison, S. F., 2001; Janig, W., 2006). Activation mechanical stimulation of the skin, compare closed
of this system (e.g., by release of the central sympa- circles with open triangles in Figure 13a). Both develop
thetic circuits, connected to preganglionic neurons slowly over 7 to 14 days during continuous activation
that innervate the adrenal medulla, from vagal inhi- of the adrenal medullae (e.g., induced by subdiaphrag-
bition by subdiaphragmatic vagotomy (Figure 13c; matic vagotomy) and are maintained over 5 weeks
Khasar, S. G. et al., 1998a; 1998b), but not of the tested (open triangles in Figure 13b). In rats with
sympatho-neural system, generates mechanical hyper- denervated adrenal medullae (adrenal nerves with
algesia and enhances bradykinin-induced mechanical preganglionic axons innervating the cells of the adrenal
hyperalgesia (decrease of paw-withdrawal threshold to medulla sectioned (Figure 13c)), subdiaphragmatic

(a) AM denervation (c) NTS

AM denerv. + vagot.
Sham vagotomy
120 Vagotomy
Vagot. + AM denerv.

Vagus nerve
Paw-withdrawal threshold (g)

100

Diaphragm
Baseline
threshold
80 Subdiaphragm.
Denervation
vagotomy

Small
Preggl. intestine
60 Adrenal
medulla

Skin
40

1 0 1 2 3 Nociceptive
log (BK) ng afferents

(b) 120
Paw-withdrawal threshold (g) 1ng BK

* + +
100

+#
+#
80

* # #
60

Vagotomy Denervation of the AM

40

0 7 14 21 28 35
days
 Autonomic Nervous System and Pain 215

vagotomy has only a small effect (compare open given intracutaneously or systemically. Adrenaline
squares with closed squares in Figure 13a). This probably does not act directly on the cutaneous
decrease of paw-withdrawal threshold can be explained nociceptors but on cells in the microenvironment
by removal of inhibition of nociceptive impulse trans- of the nociceptors inducing slow changes, which
mission in the spinal dorsal horn (Figure 13c; see result in nociceptor sensitization. Candidate cells
Randich, A. and Gebhart, G. F., 1992; see Chapter may be mast cells, macrophages, or keratinocytes,
Descending Control Mechanisms). Denervation of which then release substances that generate sensiti-
the adrenal medullae, 14 days after vagotomy when zation (Khasar, S. G. et al., 1998b; Janig, W. et al.,
the decrease of paw-withdrawal threshold is maxi- 2000; Janig, W. and Habler, H. J., 2000b). The
mal, is followed by a slow increase of paw- change of sensitivity of a population of nociceptors
withdrawal threshold taking more than 3 weeks generated by adrenaline released by the adrenal
(compare closed triangles with open triangles in medullae that is regulated by the brain would be a
Figure 13b). Thus, the decrease of paw-withdrawal novel mechanism of sensitization. This novel
threshold following vagotomy is reversed after mechanism would be different from mechanisms
denervation of the adrenal medullae. Application of that lead to activation and/or sensitization of noci-
adrenaline through a minipump for 3 to 14 days ceptors by sympathetic afferent coupling as
slowly enhances the development of mechanical discussed above (Figure 6d).
hyperalgesia induced by bradykinin injected intra- This peripheral mechanism of pain and hyperal-
dermally. Furthermore, continuous application of a gesia involving adrenaline released by the sympatho-
2-adrenoceptor blocker over 714 days by way of a adrenal system and acting on nociceptors may oper-
minipump prevents the enhancement of this hyper- ate in ill-defined pain syndromes such as irritable
algesic behavior generated by activation of the bowel syndrome, functional dyspepsia, fibromyalgia,
adrenal medullae (Khasar, S. G. et al., 2003). and chronic fatigue syndrome (Wolfe, F. et al.,
The results of this experiment are interpreted in 1990; Mayer, E. A. et al., 1995; Clauw, D. J. and
the following way: Adrenaline released by persistent Chrousos, G. P., 1997). The conclusions about
activation of the adrenal medullae sensitizes cuta- long-term sensitization of nociceptors for mechanical
neous nociceptors for mechanical stimuli. This stimulation by adrenaline fully rest on behavioral
sensitization of nociceptors and its reversal are experiments. Neurophysiological experiments in
slow and take days to develop. The slow time course vivo on primary afferent nociceptive neurons are
implies that the nociceptor sensitization cannot be required to test directly whether all or only a sub-
acutely blocked by an adrenoceptor antagonist population of nociceptors are sensitized.

Figure 13 Long-term enhancement of bradykinin (BK)-induced mechanical hyperalgesia after vagotomy and its
disappearance after denervation of the adrenal medullae (AM). (a) Decrease of paw withdrawal threshold (PWT) to mechanical
stimulation of the dorsum of the rat hindpaw induced by BK injected intradermally (BK-induced behavioral mechanical
hyperalgesia), in sham-vagotomized rats (l, n 18), in vagotomized rats (, n 16; 7 days after subdiaphragmatic vagotomy), in
rats with denervated adrenal medullae (&, n 6), and in vagotomized rats with denervated AM (&, n 6). For details see legend
of Figure 11. (b) Total change of PWT (baseline plus BK-induced) in response to intradermal injection of 1 ng BK in rats before and
7 to 35 days after vagotomy (, n 6), before and 7 to 35 days after sham vagotomy (l, n 8) and in rats whose AM are
denervated 14 days after vagotomy and measurements taken up to 35 days after initial surgery (N , rats tested after vagotomy and
denervation of the AM (n 6); H , rats only tested after additional denervation of the AM (n 4)). Note that 1ng BK injected
intradermally does not significantly change the PWT in rats with intact vagus nerves (see (a)). P < 0.01, N / versus l on day 7;

P > 0.05 N /H versus l on days 28 and 35; #P < 0.01, N /H versus  on days 28 and 35. (c) Schematic diagram showing the
proposed neural circuits in spinal cord and brainstem which modulate nociceptor sensitivity via the sympatho-adrenal system.
Sensitivity of cutaneous nociceptors for mechanical stimulation is modulated by adrenaline released by the AM. Activation of the
AM increases the sensitivity of the nociceptors. Activity in preganglionic neurons innervating the AM depends on activity in vagal
afferents from the small intestine that has an inhibitory influence on the central pathways to these preganglionic (preggl) neurons.
Thus, interruption of the vagal afferents leads to activation of the AM. It is hypothesized that these neuronal (reflex) circuits in the
brainstem are under the control of upper brainstem, hypothalamus, and telencephalon. Dotted thin lines: Axons of sympathetic
premotor neurons in the brainstem that project through the dorsolateral funiculi of the spinal cord to the preganglionic neurons of
the AM.
, excitation; , inhibition. Modified from Khasar S. G. et al. (1998b).
216 Autonomic Nervous System and Pain

17.4 Sympathetic Nervous System On the basis of clinical observations and research
and Central Integrative Mechanisms in on human patients and animals, the hypothesis has
the Control of Hyperalgesia and been put forward that CRPS (in particular type I) is a
Inflammation systemic disease involving the central and peripheral
nervous system (Figure 14). Various traumas can
The brain is suggested to modulate inflammation via trigger variable combinations of clinical phenomena
the sympatho-neural and/or the sympatho-adrenal in which the somatosensory system, the sympathetic
system although the mechanisms underlying these nervous system, the somatomotor system, and per-
influences are little understood. Any influence of ipheral (vascular and inflammatory) systems are
this type will affect indirectly the sensitivity of noci- involved (Harden, R. N. et al., 2001; Janig, W. and
ceptors. Here I will briefly discuss the hypothetical Baron, R., 2002; 2003). The central changes are
role of both sympathetic systems: (1) in CRPS that is reflected in changes of somatic sensations (increase
characterized by a peripheral inflammatory process of detection thresholds for mechanical, cold, warm,
(in addition to changes in the central nervous system) and heat stimuli), of motor performances, and of
suggested to be dependent on the sympathetic ner- neural regulation of sympathetic effector systems
vous system; (2) in experimental inflammation (vasculature, sweat glands, inflammatory cells, etc.).
(bradykinin-induced synovial plasma extravasation); Thus, it is hypothesized that the central representa-
and (3) in the potential modulation of the immune tions of the sensory, motor, and sympathetic systems
tissue. are changed. The peripheral changes consist of
inflammation involving blood vessels, inflammatory
cells, peptidergic afferent nerve fibers, and sympa-
thetic afferent coupling (see above and Figures 6
17.4.1 The Complex Regional Pain and 10). The peripheral changes cannot be seen
Syndrome Type I as Model
independently of the central ones; both interact
CRPS are painful disorders that may develop as a with each other via afferent and efferent signals.
consequence of trauma typically affecting the limbs. Furthermore, the mechanisms that underlie CRPS
Clinically, they are characterized by pain (sponta- cannot be reduced to one system or to one mechan-
neous, hyperalgesia, and allodynia), active and ism only (e.g., to sympathetic afferent coupling, to an
passive movement disorders including an increased adrenoceptor disease, to a peripheral inflammatory
physiological tremor, abnormal regulation of blood disease, and to a psychogenic disease) (Harden, R. N.
flow and sweating, edema of skin and subcutaneous et al., 2001; Janig, W. and Baron, R., 2002; 2003).
tissues, and trophic changes of skin, appendages of The centrally generated activity in sympathetic
skin, and subcutaneous tissues (Stanton-Hicks, M. neurons may be involved in various ways in the
et al., 1995; Janig, W. and Stanton-Hicks, M., 1996; pathogenesis of CRPS type I, the main argument
Harden, R. N. et al., 2001; Janig, W. and Baron, R., being that the peripheral changes are reversed or
2002; 2003). CRPS type I (previously called reflex aggravated after intervention at the peripheral sym-
sympathetic dystrophy) usually develops after minor pathetic nervous system (sympathetic blocks,
trauma with a small or no obvious nerve lesion at an stimulation of the sympathetic innervation;
extremity (e.g., bone fracture, sprains, bruises or skin Figure 14). Three points that are relevant in the
lesions, and surgeries) and rarely also after remote present context of sympathetic nervous system,
trauma in the visceral domain or after a lesion of the pain, and body protection need to be emphasized
central nervous system (e.g., stroke). An important for CRPS type I with SMP:
feature of CRPS I is that the severity and combina-
tion of clinical symptoms are disproportionate to the
In a subgroup of CRPS I patients, pain or a
component of pain is obviously dependent on activ-
severity and type of trauma with a tendency to spread ity in sympathetic neurons and related to activation
in the affected distal limb. The symptoms are not or sensitization of nociceptors by noradrenaline
confined to the innervation zone of an individual released by the sympathetic fibers (SMP). The noci-
nerve. CRPS type II (previously called causalgia) ceptors have expressed adrenoceptors and/or the
develops after trauma with a mostly large nerve excitatory effect is generated indirectly, e.g., by way
lesion (Stanton-Hicks, M. et al., 1995; Janig, W. and of changes in blood flow in deep somatic tissues of an
Stanton-Hicks, M., 1996; Harden, R. N. et al., 2001). extremity (Baron, R. et al., 2002). Sympathetically
 Autonomic Nervous System and Pain 217

PAIN SOMATOSENSATION
Spontaneous Mechanical perception
Hyperalgesia Warm perception
Allodynia Cold perception

Dynamic changes
in Forebrain
central representations
of
Somatosensory,
Somatomotor and
Sympathetic systems

Brainstem

Initiation
maintenance Dynamic
changes of Spinal cord
spinal circuits

Afferent

Sympathetic

Somatomotor

PERIPHERAL TISSUES: SYMPATHETIC NS: MOTOR SYSTEM:

Swelling and edema Regulation of blood Motor force
Inflammation flow and sweating Tremor
Trophic changes Sympathetic-afferent Dystonia
coupling

Figure 14 Schematic diagram summarizing the sensory, autonomic, and somatomotor changes in complex regional pain
syndrome I (CRPS I) patients. The figure symbolizes the CNS (forebrain, brainstem, and spinal cord). Changes occur in the
central representations of the somatosensory, the motor, and the sympathetic nervous system (which include the spinal
circuits) and are reflected in the changes of the sensory painful and nonpainful perceptions, of cutaneous blood flow and
sweating, and of motor performances. They are triggered and possibly maintained by the nociceptive afferent input from the
somatic and visceral body domains. It is unclear whether these central changes are reversible in chronic CRPS I patients.
These central changes affect the endogenous control system of nociceptive impulse transmission possibly too. Coupling
between the sympathetic neurons and the afferent neurons in the periphery (see red arrow) is one component of the pain in
CRPS I patients with sympathetically maintained pain (SMP). However, it seems to be unimportant in CRPS I patients without
SMP. Modified from Janig W. and Baron R. (2002; 2003).

maintained activity in nociceptive neurons may gen- inflammatory cells (macrophages and mast cells) by
erate a state of central sensitization/hyperexcitability way of noradrenaline and adrenoceptors on the
that is responsible for spontaneous pain and second- inflammatory cells (Figure 10) (Janig, W. and
ary evoked pain (mechanical and cold allodynia; see Baron, R., 2002; 2003).
above and Figures 5 and 7). Trophic changes in skin, appendages of skin,
Swelling (edema) and inflammation may be
generated by sympathetic and peptidergic afferent
and subcutaneous tissues (including joints and so
forth) are believed to be dependent, at least in part,
fibers interacting with each other at the arteriolar on the sympathetic innervation. The nature of this
site (influencing blood flow) and venular site neural influence on the tissue structure is unknown
(influencing plasma extravasation) of the vascular (Janig, W. and Stanton-Hicks, M., 1996; Harden, R.
bed. Sympathetic fibers are suggested to influence N. et al., 2001; Janig, W. and Baron, R., 2003).
218 Autonomic Nervous System and Pain

These three groups of observation made on patients (a) Saline

with CRPS I (who have no nerve lesion and therefore

Intensity of pain sensation (VAS)

6
by definition not neuropathic pain) argue that the
change of activity in sympathetic neurons entails
peripheral changes which in turn may result in sec- 4
50%
ondary activation and/or sensitization of primary
afferent nociceptive neurons. The underlying
2
mechanism(s) of this activation/sensitization of noci-
ceptive afferent neurons may involve short- and
long-term processes as discussed above 0
As already mentioned above, an important obser- 1 0 2 4 6
vation, in CRPS I patients with SMP (but possibly Days
also in CRPS II patients with SMP), is that pain relief (b) Lidocaine/bupivacaine
following conduction block of sympathetic neurons

Intensity of pain sensation (VAS)

by a local anesthetic applied to the sympathetic chain 6
mostly outlasts the conduction block by at least an
order of magnitude (days) (Figure 15; Price, D. D. 4 50%
et al., 1998). The long-lasting pain-relieving effect of
sympathetic blocks suggests that activity in sympa-
thetic neurons, which is of central origin, maintains a 2
positive feedback circuit via the primary afferent
neurons that are probably nociceptive in function.
0
This positive feedback circuit maintains a central
1 0 2 4 6
state of hyperexcitability (e.g., of neurons in the
Days
spinal dorsal horn; Figure 7), via excitation of afferent
neurons started by an intense noxious event. The Figure 15 Effect of sympathetic blocks with a local
anesthetic (lidocaine/bupivacaine) or of injection of saline
persistent afferent activity needed to maintain such close to the corresponding paravertebral sympathetic
a state of central hyperexcitability is switched off ganglia on pain in seven patients with CRPS I. Double-blind
during temporary block of conduction in the sympa- crossover study. Effect on pain following both interventions
thetic chain lasting only a few hours and cannot be at the sympathetic supply was measured in the same group
immediately switched on again when the conduction of CRPS I patients. Pain was systematically measured
repeatedly using the visual analogue scale (VAS) on the day
block wears off and the activity in the sympathetic of the injection and on 7 days after the injection. Both
postganglionic neurons (and therefore probably also interventions produced pain relief (see 50% value of pain
the sympathetically induced activity in afferent neu- relief). However, the mean relief of pain to injection of the
rons) returns. It is hypothesized that the afferent local anesthetic lasted for 6 days and was significantly
activity has to act over a long time period to initiate longer than the mean pain relief following local injection of
saline that lasted for 6 h (placebo block). The initial maximal
and maintain the central state of hyperexcitability peaks of analgesia were statistically not different.
(via the positive feedback). The mechanism under- Means SEM. Modified from Price D. D. et al. (1998).
lying this important and interesting phenomenon
needs to be studied experimentally in patients with
SMP as well as in animal models that have to be et al., 2006; Meyer, R. A. et al., 2005). Inflammation is
designed. potentially controlled by the brain via the HPA
system, the sympatho-neural system, and the
sympatho-adrenal system (Figure 16c), resulting in
17.4.2 Sympathetic Nervous System an enhancement or inhibition of the inflammatory
and Acute Experimental Inflammation (protective) responses depending on the behavioral
Inflammation of body tissues is accompanied by sen- state of the organism. Any change in neural and neu-
sitization of nociceptors leading to ongoing pain and roendocrine control of inflammation should indirectly
hyperalgesia (Figure 7). The sensitization of nocicep- affect the sensitization of nociceptors innervating the
tors is generated by various signaling molecules inflamed tissue and therefore ongoing pain and
released by the inflammatory cells including cells of hyperalgesia. Although we know relatively little
the immune system (Figures 10 and 12; McMahon, S. B. about the role of both sympathetic systems in the
 Autonomic Nervous System and Pain 219

(a) BK alone (c) Anterior pituitary Hypothalamus

0.16 BK with stim C-fibers gland
BK with stim C-fibers and adrenalectomy
Absorbance at 620 nm

Brain stem

0.08

HPA (d) Inflow

system 30 gauge needle

Electrical stimulation
BK Outflow
Adrenal cortex 25 gauge needle
0
0 20 40 60 80 100 120 Adrenal medulla Evans blue dye
concentration
Time (min) Preggl measured by
absorbance at 620 nm

(b) BK alone
BK with capsaicin Spinal cord
BK with capsaicin and adrenal denervation
3 g 10 g 30 g Sympa-
% max absorbance in control

thetic
outflow
100

3 2 1 4
Nociceptive
50 afferent
neuron
BK intra-articular infusion
Inflammation
0
0 20 40 60 80 100
Time (min)

Figure 16 Experimental inflammation is controlled by the hypothalamo-pituitary system and the sympatho-adrenal system.
Bradykinin (BK)-induced plasma extravasation (PE) was determined in the knee joint of anesthetized rats (d). Skin overlying
the knee was excised to expose the joint capsule, and rats were then given an intravenous injection of Evans blue dye
(50 mg kg 1). The 30-gauge inflow and 25-gauge outflow needles were then inserted into the knee joint cavity for the inflow of
perfusion fluid (0.9% saline at 250 ml min1). Perfusion fluid was collected over 5-min periods for up to 120 min and dye
concentration determined spectrophotometrically at 620 nm. The absorbance at this wavelength is linearly related to the dye
concentration and therefore to the degree of PE of the synovia (see ordinate scales in (a) and (b)). Following collection of the
first three samples to establish baseline PE levels when the joint cavity is perfused with saline, BK (160 nM; and other
substances) is (are) added to the perfusing fluid. This produces a rapid and sustained increase in the magnitude of PE (l in (a)
(n 8) and (b) (n 16)). (a) Noxious transcutaneous electrical stimulation (25 mA, 3 Hz, 0.25 ms pulses) exciting C-fibers
applied to the contralateral hindpaw after sample 8 produces a rapid decrease in the magnitude of PE (N , n 7). This inhibition
produced by noxious electrical stimulation is prevented in rats in which the adrenal gland had been removed 1 week prior to
the perfusion study (H , n 7). (b) Stimulation of cutaneous nociceptors by capsaicin injected into the palmar skin of the rat
forepaw (3, 10, 30 mg in 2.5 ml) decreased the magnitude of PE (N , n 8). This inhibition produced by capsaicin was prevented
in rats with denervated adrenal medullae (section of the adrenal nerves; H , n 8). The rats were vagotomized. (c) Schematic
diagram showing the neural and neuroendocrine components being involved in the control of BK-induced synovial PE: (1)
Terminals of sympathetic postganglionic neurons mediate 6070% of BK-induced synovial PE and the inhibitory effect of
corticosterone on BK-induced synovial PE. (2, 3) Control of BK-induced PE by the HPA system and the sympatho-adrenal
system. (4) Change of sensitivity of nociceptive afferents during inflammation as well as modulation of experimental
inflammation by peptides released these afferents. The central nociceptiveneuroendocrine reflex circuits are not shown (see
Green, P. G. et al., 1995; Miao, F. J.-P. et al., 2000; 2001). Mean SEM in (a) and (b). (a) after Green P. G. et al. (1995); (b) after
Miao F. J.-P. et al. (2000); (d) after Janig W. et al. (2006).

control of inflammation (see Straub, R. H. et al., 2005; Continuous perfusion of bradykinin through the
Straub, R. H. and Harle, P., 2005), I will summarize rat knee joint (Figure 16d) produces a large, sustained
the role of both systems in the control of acute experi- increase in plasma extravasation with a decay of
mental inflammation generated in the rat knee joint about 10% over 12 h (Figure 16a and b, filled cir-
perfused with the inflammatory mediator bradykinin cles). This bradykinin-induced plasma extravasation
(bradykinin-induced plasma extravasation). response is largely dependent on the postganglionic
220 Autonomic Nervous System and Pain

sympathetic neuron terminal: surgical sympathect- Capsaicin injected into the skin or into the peritoneal
omy reduces the magnitude of response by 60 cavity excites only C-fibers (and a few A-fibers; but
70%, but acute sympathetic decentralization (section not A-fibers) asynchronously. This activation is
of the preganglionic axons) has no significant effect. similar to physiological activation, e.g., by heat.
Similarly, acute interruption of the lumbar sympa- These modes of afferent stimulation can be used to
thetic chains during ongoing bradykinin-induced differentially activate both neuroendocrine systems
plasma extravasation does not reduce this extravasa- experimentally whatever the central underlying
tion. Finally, the sodium channel blocker tetrodotoxin mechanisms are for this differential activation.) The
coperfused with bradykinin into the knee joint cavity nociceptiveneuroendocrine reflex circuits in spinal
does not reduce the plasma extravasation (Miao, F. J.-P. cord and brainstem are enhanced after subdiaphrag-
et al., 1996a; 1996b). Thus, bradykinin-induced synovial matic vagotomy; thus, they are under inhibitory
plasma extravasation is dependent on the presence of control of central circuits activated by abdominal
sympathetic terminals in the synovia, but not on activ- vagal afferents. Both the central nociceptiveneu-
ity in these terminals and not on release of roendocrine reflex circuits and their inhibitory
noradrenaline (function 1 in Figure 16c). This depen- control exerted by activity in abdominal vagal affer-
dence on the sympathetic terminals is particularly large ents are described elsewhere (Miao, F. J.-P. et al.,
at bradykinin concentrations that have been measured 1997a; 1997b; 2000; 2001; 2003).
in inflamed tissues (108 to 107 M; Hargreaves, K. M. Transcutaneous electrical stimulation of C-fibers
et al., 1993; Swift, J. Q. et al., 1993). This is a novel inhibits bradykinin-induced plasma extravasation
peripheral function of postganglionic sympathetic (normal triangles in Figure 16a). This inhibition is
fibers, at least in the context of the model of synovial mediated by the HPA system since it can be prevented
inflammation, produced by the potent endogenous by removal of the adrenal glands (inverted triangles in
inflammatory mediator bradykinin. This function Figure 16a), by hypophysectomy, by blockade of the
occurs at the postcapillary venules by release of a corticosterone metabolism, and by blockade of corti-
chemical substance (possibly prostaglandin E2 and/or costerone receptors. It can be mimicked by
related substances), which is independent of the elec- intravenous infusion of corticosterone (Green, P. G.
trical activity in the sympathetic neurons. Whether this et al., 1997). The inhibition of bradykinin-induced
substance is released by the sympathetic terminals or plasma extravasation generated by activation of the
by other cells in association with these terminals is HPA system or by intravenous infusion of corticoster-
unknown. Future investigations will have to show one is dependent on the presence of sympathetic
whether this unusual function of sympathetic postgan- postganglionic nerve fibers in the synovia: The brady-
glionic terminals also applies to other inflammatory kinin-induced plasma extravasation remaining after
models. sympathectomy (removal of paravertebral ganglia)
Bradykinin-induced synovial plasma extravasa- cannot any longer be inhibited by activation of the
tion is under control of the brain mediated by the hypothalamicpituitary system or by corticosterone
sympatho-adrenal system and the HPA system (func- injected intravenously. This shows that the sympa-
tions 2 and 3 in Figure 16c). Both the neuroendocrine thetic fibers are in a strategic, yet unexpected,
control systems can be reflex activated by noxious position in the control of synovial plasma extravasa-
stimuli. The HPA system is most efficiently activated tion by the brain (Green, P. G. et al., 1997).
by continuous transcutaneous electrical stimulation Reflex inhibition of bradykinin-induced plasma
of afferent C-fibers and the sympatho-adrenal system extravasation generated by stimulation of nociceptors
during stimulation of nociceptors by capsaicin (e.g., by capsaicin (normal triangles in Figure 16b) is pre-
injected into the plantar or palmar skin or into the vented by denervation of the adrenal medullae
peritoneal cavity). (inverted triangles in Figure 16b). Thus, it is mediated
The differential activation of both neuroendo- by adrenaline released by the adrenal medullae.
crine systems, the hypothalamicpituitary system The mechanisms by which corticosterone released
and the sympatho-adrenal system, generated by the by the adrenal cortex and adrenaline released by the
modes of afferent stimulation is interesting but puz- adrenal medulla depress bradykinin-induced synovial
zling. During transcutaneous electrical stimulation, plasma extravasation and the role of the sympathetic
C-fibers are stimulated continuously and synchro- nerve fibers innervating the synovia in this depression
nously at 14 Hz together with A-fibers. This are unknown. Corticosterone seems to act via the pep-
activation of C-fibers is certainly unphysiological. tide annexin possibly released by inflammatory cells in
 Autonomic Nervous System and Pain 221

the synovia (Green, P. G. et al., 1998). Here it is impor- Supraspinal

tant to emphasize that the control of experimental pathways
synovial inflammation by both neuroendocrine systems Primary
afferent
affects indirectly the sensitivity of nociceptors inner- inputs
vating the knee joint and, therefore, also pain and
hyperalgesia and that this change of sensitivity is there- Preganglionic
fore under the control of the brain. neurons

17.4.3 Sympathetic Nervous System

and Immune System
The hypothalamus can influence the immune system by
way of the sympathetic nervous system and can there-
fore control protective mechanisms of the body at the
cellular level (Besedovsky, H. O. and del Rey, A., 1995;
Hori, T. et al., 1995; Madden, K. S. and Felten, D. L.,
1995; Madden, K. S. et al., 1995). The parameters of
the immune tissues potentially controlled are prolif-
eration, trafficking and circulation of lymphocytes,
Immune cell
functional activity of lymphoid cells (e.g., activity of target Visceral target
natural killer cells) and cytokine production, hema-
topoesis of bone marrow, mucosal immunity, Vascular target
thymocyte development, etc. (for details of potential
Figure 17 A separate sympathetic pathway to the
mechanisms, see Elenkov, I. J. et al., 2000). The immune tissue: a hypothesis. Demonstrated are three
mechanisms of this influence remain largely unsolved functional types of sympathetic pathway, one each to the
(Ader, A. and Cohen, N., 1993; Saphier, D., 1993; vasculature (e.g., to blood vessels in skeletal muscle or skin
Besedovsky, H. O. and del Rey, A., 1995). It is or viscera), to the viscera (e.g., to the gastrointestinal tract
unknown (1) whether there exists a functionally dis- being involved in the regulation of motility or secretion), or to
the immune tissue. This neural link to the immune tissue
tinct sympathetic system from the brain to the may indirectly lead to sensitization of nociceptors and pain
immune system or (2) whether the modulation of (modified from Janig, W., 2006).
the immune system is a general function of the sym-
pathetic nervous system. The more likely hypothesis
favored by me is that the immune system is modulated Acknowledgment
by the brain by way of a functionally and anatomically
distinct sympathetic pathway (Figure 17; for discus- Supported by the Deutsche Forschungsgemeinschaft
sion, see Janig, W. and Habler, H. J., 2000a; Janig, W., and the German Ministry of Research and Education
2006). The reflex discharge characteristics of the (German Research Network on Neuropathic Pain).
neurons of this hypothetical sympathetic pathway
innervating the immune tissue and the neural circuits
in spinal cord, brainstem, and hypothalamus being References
involved in regulation of the activity of this pathway
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Biophysics (eds. T. C. Ruch and H. D. Patton), pp. 345363. activation: the role of pro-inflammatory cytokines in
Saunders. inflammation, illness responses and pathological pain states.
Saphier, D. 1993. Psychoimmunology: The Missing Link. Pain 63, 289302.
In: Hormonally Induced Changes in Mind and Brain White, J. C. and Sweet, W. H. 1969. Pain and the Neurosurgeon:
(ed. J. Schulkin), pp. 191224. Academic Press. A Forty Year Experience. Springfield.
Sato, J. and Perl, E. R. 1991. Adrenergic excitation of cutaneous Willenbring, S., Beauprie, I. G., and DeLeo, J. A. 1995. Sciatic
pain receptors induced by peripheral nerve injury. Science cryoneurolysis in rats: a model of sympathetically
251, 16081610. independent pain. Part 1: Effects of sympathectomy. Anesth.
Seltzer, Z., Dubner, R., and Shir, Y. 1990. A novel behavioral Analg. 81, 544548.
model of neuropathic pain disorders produced in rats by Wolfe, F., Smythe, H. A., Yunus, M. B., Bennett, R. M.,
partial sciatic nerve injury. Pain 43, 205218. Bombardier, C., Goldenberg, D. L., et al. 1990. The American
Shir, Y. and Seltzer, Z. 1991. Effects of sympathectomy in a College of Rheumatology (1990) Criteria for the Classification
model of causalgiform pain produced by partial sciatic nerve of Fibromyalgia. Report of the Multicenter Criteria
injury in rats. Pain 45, 309320. Committee. Arthritis Rheum. 33, 160172.
Stanton-Hicks, M., Janig, W., Hassenbusch, S., Haddox, J. D., Woolf, C. J. 1996. Phenotypic modification of primary sensory
Boas, R., and Wilson, P. 1995. Reflex sympathetic dystrophy, neurons: the role of nerve growth factor in the production
changing concepts and taxonomy. Pain 63, 127133. of persistent pain. Phil. Trans. R. Soc. Lond. B
Straub, R. H. and Harle, P. 2005. Sympathetic 351, 441448.
neurotransmitters in joint inflammation. Rheum. Dis. Clin. Woolf, C. J., Allchorne, A., Safieh-Garabedian, B., and Poole, S.
North Am. 31, 4359. 1997. Cytokines, nerve growth factor and inflammatory
Straub, R. H., Baerwald, C. G., Wahle, M., and Janig, W. 2005. hyperalgesia: the contribution of tumour necrosis factor
Autonomic dysfunction in rheumatic diseases. Rheum. Dis. alpha. Br. J. Pharmacol. 121, 417424.
Clin. North Am. 31, 6175. Woolf, C. J., Ma, Q.-P., Allchorne, A., and Poole, S. 1996.
Swift, J. Q., Garry, M. G., Roszkowski, M. T., and Peripheral cell types contributing to the hyperalgesic action
Hargreaves, K. M. 1993. Effect of flurbiprofen on tissue levels of nerve growth factor in inflammation. J. Neurosci.
on immunoreactive bradykinin and acute postoperative pain. 16, 27162723.
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Koltzenburg, M. 1995. Noradrenaline-evoked pain in
neuralgia. Pain 63, 1120.
Tracey, D. J., Cunningham, J. E., and Romm, M. A. 1995. Further Reading
Peripheral hyperalgesia in experimental neuropathy:
mediation by alpha 2-adrenoreceptors on post-ganglionic Miao, F. J.-P., Green, P. G., and Levine, J. D. 2004.
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Undem, B. J. and Weinreich, D. (eds.). 2005. Advance in Vagal modulation of inflammation in the rat. J. Physiol.
Afferent Neurobiology. CRC Press. 554, 227235.
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 18 Sympathetic Blocks for Pain
A Sharma, Columbia University, New York, NY, USA
J N Campbell and S N Raja, Johns Hopkins University, Baltimore, MD, USA
2009 Elsevier Inc. All rights reserved.

18.1 Introduction 227

18.1.1 Sympathetically Maintained Pain 228
18.2 Techniques of Sympathetic Blockade 229
18.3 Sympathetic Block for Diagnosis 229
18.3.1 Local Anesthetic Sympathetic Blocks 230
18.3.2 Intravenous Regional Sympathetic Blockade 231
18.3.3 Systemic Alpha-Adrenergic Blockade 231
18.4 Sympathetic Block for Therapy of Chronic Pain States 232
18.4.1 Local Anesthetic Sympathetic Blocks and Intravenous Regional
Sympathetic Blockade 232
18.4.2 Spinal Cord Stimulation 233
18.4.3 Chemical Neurolysis and Radiofrequency Denervation Techniques 233
18.4.4 Surgical Sympathectomy 233
18.5 Conclusions 234
References 234

18.1 Introduction allodynia (enhanced pain to natural stimuli such as a

pin prick or light stroking of the skin, respectively),
Traditionally, the sympathetic nervous system (SNS) autonomic features, and motor dysfunction. Some
is considered as an efferent system that controls peri- patients in addition have a motor impairment not
pheral blood flow, sweating, and piloerection. This readily explained by a lesion in the peripheral nervous
autonomic part of the nervous system, however, may system. Edema at the time of presentation or at an
play a pivotal role in certain pain conditions. Over the earlier time point is a common feature. CRPS may be
last decade, much knowledge has been gained on the present in cases with (CRPS type II) or without a
mechanisms of interaction between the sensory affer- nerve lesion (CRPS type I). After the initial descrip-
ent and sympathetic efferent systems. Pain dependent tion of this disease by Weir Mitchell in 1864 (who
on the discharge of sympathetic nerves is referred to as called it causalgia) and decades later by Paul Sudeck
sympathetically maintained pain (SMP). Insights into (Sudecks syndrome), Leriche advocated its treatment
the pathophysiology of SMP help its treatment. The by sympathetic interruption in 1916. Leriche (vicious
concept of SMP stemmed from clinical observations cycle hypothesis, 1939) and Mandl (1947) were pio-
that certain patients with persistent pain after trau- neers in proposing the connection between pain and
matic nerve lesions received dramatic pain relief from SNS. The concept that the SNS was involved in the
blockade of sympathetic ganglion with local anesthetic mechanisms of pain in CRPS was further propagated
agents or perivascular sympathectomy. by Evans, who coined the term reflex sympathetic
A pain syndrome often associated with SMP is now dystrophy. In view of the explosion of research regard-
termed complex regional pain syndrome (CRPS). ing mechanisms of pain in RSD and lack of uniform
Only subsets of patients with CRPS have SMP. diagnostic criteria, major taxonomical changes were
Formerly termed as reflex sympathetic dystrophy proposed by the International Association for the
(RSD), CRPS is an incompletely understood pain Study of Pain (IASP) Consensus Team, and the term
disorder that usually results from a traumatic injury complex regional pain syndrome was introduced
and clinically presents as nondermatomal distribution (Stanton-Hicks, M. et al., 1995). According to the
of pain and sensory abnormalities in an extremity. IASP classification, two distinct subtypes are now
Symptoms and signs include hyperalgesia and identified. Patients in whom the above-described

227
228 Sympathetic Blocks for Pain

conglomeration of symptoms and signs develop after 18.1.1 Sympathetically Maintained Pain
injury to a major peripheral nerve are considered to
The clinical picture of CRPS with vascular (vasodi-
have CRPS type II (previously known as causalgia).
lation or vasoconstriction, skin temperature
The remaining patients in whom injuries to a limb or
asymmetries, or skin color changes) and sudomotor
lesions in remote body areas precede the onset of
abnormalities (swelling, hyper-, or hypohidrosis)
symptoms without any identifiable nerve damage are
perhaps promoted the initial idea of SNS dysfunction
classified as CRPS type I (previously termed reflex
as the primary etiology of the disease. Several lines of
sympathetic dystrophy) (Merskey, H. and Bogduk, N.,
evidence suggest a dysregulation of autonomic func-
1995). Since the initial proposal of diagnostic criteria
tion in the initial phase of the disease. Furthermore,
for CRPS, further modifications have been made
(Harden, R. N. and Bruehl, S. P., 2006). Presently, enhancement of spontaneous pain and hyperalgesia
the clinical signs and symptoms of CRPS are grouped after physiological activation of SNS (Baron, R. et al.,
into the following categories: 2002; Drummond, P. D. and Finch, P. M., 2004) and
resolution of pain, physiological tremor, and swelling
1. Positive sensory abnormalities (spontaneous pain, in some patients after sympathetic blocks strengthen
mechanical, somatic or deep somatic hyperalgesia). this view. The detection of autoantibodies against the
2. Vascular abnormalities (vasodilation or vasocon- autonomic nervous system in certain CRPS patients
striction, skin temperature asymmetries, or skin provides further evidence to this concept.
color changes). The relief of pain by blockade of the SNS initially
3. Sudomotor abnormalities (swelling, hyper-, or invited speculation of an increased SNS discharge as
hyperhidrosis). a potential mechanism for SMP. Several observations
4. Motor changes (weakness, tremor, dystonia, or were against this hypothesis. First, patients with
coordination deficits) and trophic changes (nail SMP, particularly in the early period of the disease,
or hair changes, skin atrophy, joint stiffness, or may present with warm rather than cold skin. Under
soft-tissue changes). conditions of increased sympathetic discharge, vaso-
The criteria are being further validated as no gold constriction should lead to a cold skin. Also others
standard diagnostic tool has been established for noted that catecholamine levels in the venous return
CRPS (Baron, R. and Janig, W., 2004; Harden, R. N. of the affected extremity were below those in normal
and Bruehl, S. P., 2006). The 1993 IASP consensus subjects or their unaffected side.
team also acknowledged the term sympathetically In 1948, Walker et al. made an important clinical
maintained pain (SMP) and defined it as Pain that observation that provided convincing evidence for
is maintained by sympathetic efferent innervation or SMP. At that time, sympathectomy was performed
by circulating catecholamines. A role for circulating to treat limb ischemia and also to relieve pain in
catecholamines has not been clearly established patients with causalgia. Walker and colleagues per-
though. SMP is traditionally diagnosed by relief formed a preganglionic sympathectomy and placed
of allodynia, hyperalgesia, ongoing pain, and a selec- electrodes on the ganglion. Postoperatively, the gang-
tive blockade of sympathetic function. Notably, lion was stimulated. Since the ganglion was
patients with CRPS, as previously indicated, may dissociated from the spinal cord, electrical stimulation
have both SMP and sympathetically independent would only induce an efferent discharge in the peri-
pain (SIP) in varying proportions (Stanton-Hicks, pheral sympathetic fibers. Stimulation induced pain
M. et al., 1995). in patients with causalgia, but not in patients with
Even after decades of animal and human research, limb ischemia. This indicated that SMP was due to
the pathophysiology of CRPS remains uncertain. efferent function of the SNS. In a further experiment
Though it is common to refer to CRPS as a neuro- performed decades later, norepinephrine was injected
pathic pain disorder, it is not clear that pathology of in patients with SMP. In these patients, the sympa-
the nervous system is at the root of this disorder as thetic ganglia were blocked with local anesthetic. If
patients with or without nervous system lesions may SNS dysfunction does exist, does it involve under- or
manifest with symptoms of CRPS ( Janig, W. and overactivity? Earlier description of the disease pro-
Baron, R., 2002; 2003; 2004). Moreover, it is clear moted the concept of distinct phases with an initial
that CRPS is a diagnosis of exclusion, as other diseases phase involving warm, erythematous skin (stage I or
(e.g., nerve entrapment and diabetic neuropathy) may acute stage) presumably due to SNS underactivity, an
present with similar signs and symptoms. intermittent phase of both warm and cold sensation
 Sympathetic Blocks for Pain 229

(stage II or dystrophic stage) followed by chronic cold 18.2 Techniques of Sympathetic

sensation (stage III or atrophic stage) due to SNS Blockade
hyperactivity. Although the existence of distinct
phases of CRPS is debated at present (Bruehl, S. The techniques to achieve sympathetic blockade
et al., 2002), many studies have shown that vasomotor include the following:
tone is reduced in very early stages of CRPS
1. Local anesthetic regional block (LASB) of sympa-
(Kurvers, H. A. et al., 1995; Wasner, G. et al., 2001),
thetic ganglion or trunk, for example, stellate
but not in later stages. The finding of reduced plasma
ganglion block or lumbar sympathetic block.
norepinephrine and neuropeptide Y concentration on
2. Neuraxial techniques, for example, epidural or
the affected extremity in early phase of the disease
spinal block.
confirms this proposition (Raja, S. N. et al., 1995). It
3. Intravenous regional sympathetic blockade
seems that the apparent increase in vasomotor tone in
(IRSB) with phentolamine, guanethidine, brety-
later stages of CRPS (as evident by vasoconstriction
lium, clonidine, or lidocaine using tourniquet
or cold skin) is due to increased sensitivity (hyper- or
(Bier block technique).
supersensitivity) of skin microvessels to catechola-
4. Intravenous infusion of systemic alpha-adrenergic
mines (Kurvers, H. A. et al., 1995) or secondary
antagonists.
changes in neurovascular transmission while actual
5. Radiofrequency denervation of sympathetic gang-
sympathetic tone is still depressed. Possible mechan-
lion or trunk.
isms for this supersensitivity might be decreased
6. Surgical sympathectomy.
neuronal uptake of norepinephrine in the sympathetic
neuroeffector junction (Raja, S. N. et al., 1995) or While LASB, IRSB with sympatholytic or local
increased number of peripheral -receptors (Davis, anesthetic medications, neuraxial techniques, and intra-
K. D. et al., 1991; Ali, Z. et al., 2000). Studies in venous infusion of adrenergic antagonist can be used
experimental animals report an increased expression for both diagnostic and therapeutic benefits, neurolytic
of -adrenoceptors in the neuromas that develop procedures (e.g., 5 and 6) are reserved for treatment of
after nerve damage. Norepinephrine causes increased selected patients who show consistent but short-lasting
excitation of the neuroma afferents. Peripheral nerve response to initial diagnostic modalities.
injury may also result in sprouting of adrenergic fibers
into the dorsal root ganglion and forms contact with
sensory neurons. Thus, sympathetic afferent interac-
tions might exist in the vicinity of the sensory 18.3 Sympathetic Block for
receptors in partially denervated peripheral tissues, Diagnosis
the peripheral nerve at the site of injury, and the
dorsal root ganglion where cell bodies of injured The effectiveness of any diagnostic test or procedure
sensory nerves are located. is measured by its sensitivity and specificity. A good
Although SNS dysfunction is a fascinating con- diagnostic test, when performed appropriately,
cept, it does not explain all features of CRPS. should give consistent results in a given patient and
Inflammatory and central origin of pain in CRPS should help guide therapy or predict outcome. Before
patients has been suggested, and significant animal any meaningful conclusions can be drawn from a
and human research support their contributions. The sympathetic block by any technique, optimum inhi-
readers are advised to review the articles by Janig W. bition of sympathetic outflow to the involved
and Baron R. (2003; 2004) for further understanding extremity needs to be achieved. This requires knowl-
in this matter. Readers are also encouraged to read edge of the anatomical distribution of sympathetic
the conflicting views of Ochoa J. and Verdugo R. J. innervation and the common pitfalls associated with
(2001) on the existence of SMP as well as the value of the conduct and interpretation of these procedures.
diagnostic phentolamine test or sympathetic blocks. For instance, in performing stellate ganglion block to
The authors have negated the existence of SMP achieve an upper extremity sympathetic blockade, it
based on their own studies (Verdugo, R. J. and is imperative to block contributions from upper thor-
Ochoa, J. L., 1994), which in our opinion, had meth- acic ganglions that are anatomically separate from
odological deficiencies. A detailed discussion of these the stellate ganglion. This could be achieved by
mechanisms and opposing views is beyond the scope modifications in the previous technique. Similarly,
of this chapter. injection of local anesthetic solution into nearby
230 Sympathetic Blocks for Pain

vascular structures could cause failure to achieve 18.3.1 Local Anesthetic Sympathetic
sympatholysis. This can be avoided with the use of Blocks
intravenous contrast agent and live fluoroscopy to
LASBs are technically challenging procedures that
rule out intravascular placement of needle tip.
require training. They have the advantage of selec-
Following sympathetic blockade, determination of tively blocking stellate or lumbar sympathetic ganglion
adequacy of sympatholysis is imperative. Monitoring and/or chain with a small dose of local anesthetic
skin temperature change is commonly used in clinical solution without concomitant blockade of sensory or
practice. Adequate sympathetic blockade is inferred by motor fibers (Table 1). Using fluoroscopic or CT gui-
approximation of core body temperature and skin tem- dance, the needle tip is positioned in close proximity to
perature of the ipsilateral limb. Accepting a rise in skin the sympathetic ganglia and a local anesthetic solution
temperature by an arbitrary number in comparison is injected. Although numerous complications of LASB
with the contralateral limb leads to higher false-positive have been reported in the literature, these techniques
blocks (Schurmann, M. et al., 2001), probably due to are safe in experienced hands. Efficacy of the block is
bilateral increase in blood flow following sympathetic assessed as previously described by monitoring objec-
block techniques. Many authors recommend further tive evidence of sympathetic blockade, for example,
assessment of sympathetic function by sweat test cutaneous temperature monitoring. The patient is
(Stevens, R. A. et al., 1998) or skin conductance response. asked to report the intensity of pain using a numeric

Table 1 Comparison of diagnostic modalities for sympathetically maintained pain

Technique Advantages Disadvantages

Interventional regional 1. Well-localized sympathetic ganglion or 1. Requires technical expertise and has
sympathetic block with chain can be anesthetized (blocked). higher cost
local anesthetic agents Objective signs of sympathectomy are 2. Requires fluoroscopy or CT guidance
(LASBs) seen with small doses of LA 3. Complication from contrast or
2. Systemic side effects from intravenous medication allergy
medications are avoided
4. Complication from technical reasons;
e.g., infection, bleeding, and nerve
damage
5. Poorly tolerated in pediatric population
and patients with needle phobia
6. Sedatives used during the
interventional technique might hinder
with postprocedure pain responses
7. False negative
 LA fails to anesthetize the
sympathetic ganglion adequately
 Intravascular injection
 Central sympatheticnociceptive
interactions may be missed
8. False positive
 LA anesthetizing somatic afferent
fibers in the vicinity and causing pain
relief from concurrent somatic
blockade
 Regional ischemic pain will improve
with sympathetic block
 Placebo effect

Intravenous regional 1. Easy to perform 1. Side effects of medications, e.g.,

sympathetic blockade 2. Better patient acceptance phentolamine can cause positive
(IRSB) 3. Low cost chrontropic and inotrophic effects
4. Intravenous medications like 2. Achieving complete sympathectomy at
phentolamine can cross bloodbrain clinically tolerable doses might be
barrier and act on spinal cord -2 difficult
adrenergic receptors, which are involved 3. Placebo effect cannot be ruled out
in pain modulation
 Sympathetic Blocks for Pain 231

pain score before and after sympathetic blockade. The 18.3.3 Systemic Alpha-Adrenergic
response is usually considered to be positive when a Blockade
50% or greater improvement in pain scores is reported
Intravenous alpha-adrenergic blockade with phento-
in the light of adequate sympathectomy. LASB is tra-
lamine (Phentolamine block, PhB) has been proposed
ditionally considered as the gold standard against
as a safe and effective method of diagnosing SMP
which other techniques have been tested in past
(Raja, S. N. et al., 1991; Dellemijn, P. L. et al., 1994).
(Dellemijn, P. L. et al., 1994). Beyond CRPS, LASBs
This technique is different from IRSB as no tourni-
are also used to block efferent sympathetic fibers at
quet is used to isolate the affected limb. In the initial
thoracic sympathetic trunk, celiac plexus, superior
study by Raja S. N. et al. (1991), 20 patients were
hypogastric plexus, and ganglion impar to confirm
randomly assigned to 2535 mg of intravenous phen-
improvement in pain prior to more invasive therapeu-
tic blocks. These invasive procedures are used in the tolamine (being administered in a double-blinded
treatment of cancer pain, which may be somatic or fashion) or LASB with 0.25% bupivacaine, and simi-
visceral, and neuropathic or mixed in nature. A posi- lar pain relief patterns were observed. Although
tive response to a temporary diagnostic local anesthetic 57  C temperature difference was seen following
block is a useful predictor of the success of subsequent LASB, the changes in cutaneous temperature after
neurodestructive procedures. Differential spinal or PhB were variable (1.8 0.7  C). Similar results were
epidural blocks have also been described to distinguish obtained by Dellemijn P. L. et al. (1994) who con-
somatic or central mechanisms from sympathetic etiol- cluded that phentolamine infusion is less sensitive
ogies. However, it is difficult to achieve selective but more specific test of SMP than LASB. Both
sympathetic blockade with these techniques. groups (Raja, S. N. et al., 1991; Dellemijn, P. L. et al.,
1994) mentioned that the mechanism of pain relief
during sympathetic blockade was independent of
cutaneous vasodilatation or skin temperature
18.3.2 Intravenous Regional Sympathetic changes. In an attempt to find optimum test dose,
Blockade Raja S. N. et al. (1996) then compared the effect of
Intravenous regional anesthesia, a technique two different doses of phentolamine (0.5 and 1 mg
described by August Bier in 1908, involves injection kg1) on skin blood flow (using a laser Doppler
of a local anesthetic solution in a limb, isolated by blood flow monitor) and sympathetically mediated
tourniquet. Based on this concept, Hannington-Kiff vasoconstrictor response (a reflex decrease in periph-
proposed IRSB using guanethidine. Initially, gua- eral blood flow in response to deep inhalation) (Raja,
nethidine (with or without a local anesthetic agent) S. N. et al., 1996). They concluded that the higher
was used to perform this block and later other med- dose of phentolamine (1 mg kg1 over 10 min)
ications were tried. These include sympatholytics resulted in more complete adrenoceptor blockade
like reserpine, bretylium, or clonidine; local anes- and recommended this higher dose with cutaneous
thetics like prilocaine or lidocaine; and others like temperature monitoring to evaluate the sympa-
kitanserin (5-HT2 antagonist). Although IRSBs have thetically mediated component in neuropathic pain
been used therapeutically in SMP, their use in diag- states.
nostic blocks has been limited. The ischemic There is a paucity of research in recognition of
tourniquet used during the procedure can itself placebo responsiveness to sympathetic blocks.
relieve hyperalgesia in neuropathic pain patient Randomization of patients for interventional techni-
(Campbell, J. N. et al., 1988) and cause false-positive ques not only is difficult but also has ethical
results. Certain patients tolerate tourniquet poorly. implications. Clinical observation frequently shows
With IRSBs, guanethidine is often coinjected with prolonged benefit in some patients beyond the dura-
local anesthetics since the release of norepinephrine tion of action of local anesthetic agents. Although
by guanethidine is usually associated with the reversal of central sensitization in the spinal cord
exacerbation of pain. As a result, diagnosis is often and reduced coupling of sympathetic and sensory
confounded by the effects of local anesthetics. neurons have been put forward as a possible explana-
Sudden release of tourniquet might release high tion, other possibilities need further consideration. In
doses of medications in systemic circulation and order to reduce placebo responsiveness, patients
cause significant side effects. This technique is, should have different diagnostic procedures like
hence, not useful as a diagnostic tool. LASB and PhB on separate days. These two
232 Sympathetic Blocks for Pain

procedures provide complementary information and peer review journals from 1916 through 1999. Only
both may be needed to confirm the diagnosis of SMP 29 studies met the inclusion criteria of a sample size
(Dellemijn, P. L. et al., 1994). of at least 10 patients undergoing LASB with con-
vincible evidence of CRPS. Interestingly, of these 29
studies, only 10 studies evaluated the technical suc-
18.4 Sympathetic Block for Therapy cess of block and only two of those assessed pain and
of Chronic Pain States outcome. Based on the data from 14 studies (454
patients) that quantified the magnitude of patients
In the past, once diagnosis of SMP was established, responses and also reported the number of patients
patients were often offered a series of regional or with different degrees of responses, authors found
intravenous blocks in attempt to reduce pain and that 29% patients undergoing LASB obtain full pain
vasomotor symptoms. At present, they are considered relief (>75% improvement) while 41% obtain partial
as an element of multidisciplinary treatment plans. relief (2575% improvement). Authors concluded
Patients who get significant benefit from initial that less than one-third positive response (full pain
blocks are often subjected to more aggressive inter- relief) is consistent with placebo effect and that the
ventional therapies. These treatment modalities are efficacy of sympathetic blocks for treatment of CRPS
presumed to have enduring sympathectomy or mod- is inconclusive. Many of these 14 studies were pub-
ulation of SNS. They include spinal cord (dorsal lished prior to 1960 and only two actually identified
column) stimulation, chemical neurolytic proce- technical success of block and pain scores.
dures, radiofrequency denervation of sympathetic Long-term usefulness of IRSB is even less certain.
trunk, or even surgical sympathectomy. Jadad A. R. et al. (1995) attempted a small RCT (nine
patients) of the effects of repeated intravenous gua-
nethidine blocks on pain intensity and relief, adverse
18.4.1 Local Anesthetic Sympathetic
effects, mood, duration of analgesia, and global
Blocks and Intravenous Regional
scores. The trial was stopped prematurely because
Sympathetic Blockade
of the severity of the adverse effects (hypotension).
LASBs provide short-term benefits in relieving No significant difference was found between gua-
spontaneous and evoked pain and reducing vasomo- nethidine and placebo on any of the outcome
tor symptoms. Their long-term efficacy is unproven. measures. The authors also presented a systematic
In contrast, IRSB does not require any special review of seven studies on the effects of IRSB in
expertise, is easier to perform and, comparatively, CRPS. Two of these studies, one using ketanserin
has better patient acceptance (Table 1). These and one bretylium, with 17 patients in total, showed
procedures have the risk of tourniquet-related pain some advantage over control but few other RCTs
(if used) and significant side effects from the medica- showed lack of efficacy of guanethidine. Authors
tions. Both techniques can get false-positive results concluded that the use of guanethidine in IRSBs for
from placebo effect. patients with CRPS was not supported by any avail-
Randomized controlled trials (RCTs) are consid- able literature. A recent study by Livingstone et al.
ered as gold standard to evaluate the efficacy of any again showed no significant analgesic advantage of
therapeutic modality, but not many of these studies guanethidine over a normal saline placebo block in
have been conducted to assess the usefulness of the treatment of early CRPS type I.
LASB in SMP. Most randomized trials have been Difficulty in conducting RCTs for LASBs or
done in CRPS patients (includes both SMP and SIP IRSBs for treatment of CRPS or SMP is explicable.
patients) and not specifically in SMP states. SIP Randomization to placebo is almost impossible.
patients are not expected to get any long-term ben- Patients or physicians cannot be blinded as outcomes
efits from sympathetic blocks and their inclusion in of adequate sympathectomy are obvious. Also, ethical
the study biases the results. Cepeda M. S. et al. (2002) issues might arise in performing technically challen-
attempted a systematic review of RCTs of the effect ging procedures like stellate ganglion block with
of LASB in CRPS (not SMP) patients but were saline. But randomization can be done in a different
unable to pool data due to significant differences in fashion. The new concept of expertise-based RCTs
their design. The authors then reviewed nonrando- might be useful in this condition when patient out-
mized controlled studies, case series, and RCTs with comes can be compared between physicians who
acceptable designs, published in English-language perform repeat LASBs or IVRBs to those who solely
 Sympathetic Blocks for Pain 233

impose conservative management. But again, such syndrome, wound infection, and spinal cord injury.
studies should compare patients with documented There is a theoretical advantage of lesser incidence of
diagnosis of SMP and not CRPS. Based on available postsympathetic neuralgia and more precise sym-
data in literature, efficacy and duration of LASB is patholysis with radiofrequency technique in
variable and, indeed, unpredictable. Short duration of comparison with chemical neurolysis. Outcome data
pain relief and reduction in vasomotor symptoms is for these techniques vary. While Wilkinson H. A.
often seen in clinical practice and should be utilized (1996) reported almost 90% partial or complete evi-
to improve mobility, range of motion, and motor dence of sympathectomy after 2 years, most authors
strength by physiotherapy. Repeated blocks are ben- report sustained pain relief in less than two-thirds of
eficial in selected patients with willingness to actively patients at 2 years, and about one-third at 5 years. In a
participate in physiotherapy and are showing signs of systematic review of the effects of percutaneous neu-
continuing improvement. rodestructive procedures for neuropathic pain, Mailis
A. and Furlan A. (2003) concluded that the practice of
surgical and chemical sympathectomy is based on
18.4.2 Spinal Cord Stimulation
poor quality evidence, uncontrolled studies, and per-
Spinal cord stimulation (SCS) has recently gained sonal experience. Therefore, more clinical trials of
acceptance for a wide variety of neuropathic pain syn- sympathectomy are required to establish the overall
dromes. A recent RCT (Kemler, M. A. et al., 2004) and effectiveness and potential risks of this procedure.
meta-analysis of literature (Grabow, T. S. et al., 2003) Because of the limited long-term outcomes, consid-
showed that SCS results in a long-term pain reduction eration should be given to neuromodulatory methods
and health-related quality-of-life improvement, but no for SMP involving extremities. For other SMP syn-
clinically important improvement of functional status, dromes, like visceral neuropathic pain in pancreatic
in chronic RSD. The technique is more effective and cancer, chemical neurolyis continues to be the pro-
less expensive when compared with the standard treat- cedure of choice.
ment protocol for chronic CRPS (Mekhail, N. A. et al.,
2004). The mechanism of action by which the device
18.4.4 Surgical Sympathectomy
provides pain relief is unclear. In general, patients with
good response to sympathetic blocks are likely to have Surgical sympathectomy has been tried in patients
long-term pain relief (Hord, E. D. et al., 2003) with SCS, who achieve good pain relief with a series of sympa-
although it does not affect skin microcirculation. The thetic block. Better outcomes are predicted when
readers are encouraged to read a recent review on this sympathectomy is performed early in the disease.
subject for a detailed discussion (Meyerson, B. and Concerns about recurrence or development of new
Linderoth, B., 2001). CRPS or disabling compensatory sweating syndrome
have been raised after surgical sympathectomy, but
current data suggests low incidence (7%) of these
18.4.3 Chemical Neurolysis and
complications (Bandyk, D. F. et al., 2002). Other dis-
Radiofrequency Denervation Techniques
turbing complication includes transient (<3 months)
Patients who show good response to initial sympa- postprocedural sympathalgia (in one-third of the
thetic blocks are often subjected to radiofrequency patients following cervicodorsal sympathectomy and
denervation or chemical neurolytic destruction of 20% of the patients after lumbar sympathectomy). At
sympathetic innervation. The technique of radiofre- 1 year postprocedure, one-quarter of the patients are
quency denervation has gone through various expected to continue experience significant improve-
modifications by Noe et al., Rocco, and Sluitjer. At ment (pain severity score <3) and an additional 50%
present, destruction of T2 and T3 ganglions is of the patients would have continued but reduced pain
recommended for upper extremity involvement severity and an increase in daily/work activities
while guidelines for lower extremity vary. Most (Bandyk, D. F. et al., 2002). Many surgeons prefer
authors suggest destruction of at least the first three video-assisted minimally invasive techniques instead
ganglions (L1, L2, and L3), whereas others believe of open sympathectomy to reduce other complication
that the L4 and even L5 must be included for rates and promote early postoperative recovery.
good long-term results. Complications of neurodes- With the increasing popularity of radiofrequency pro-
tructive procedures include postsympathectomy cedure, rates of surgical sympathectomy have
sympathalgia, compensatory hyperhidrosis, Horners decreased.
234 Sympathetic Blocks for Pain

18.5 Conclusions Grabow, T. S., Tella, P. K., and Raja, S. N. 2003. Spinal cord
stimulation for complex regional pain syndrome: an
evidence-based medicine review of the literature. Clin. J.
SNS plays an intriguing role in maintaining pain in Pain 19(6), 371383.
certain conditions. Complex regional pain syndromes Harden, R. N. and Bruehl, S. 2006. Diagnosis of complex
regional pain syndrome: signs, symptons, and new
are prime example of such states. Selective sympa- empirically derived diagnostic criteria. Clin. J. Pain
thetic blocks are commonly used to diagnose a subset 22(5), 415419.
of patients with a predominant sympathetically Hord, E. D., Cohen, S. P., Cosgrone, G. R., Ahmed, S. U.,
Vallejo, R., Chang, Y., and Stojanovic, M. P. 2003. The
maintained pain state. In order to avoid false-positive predictive value of sympathetic block for the success of
diagnosis, at least two different diagnostic tests spinal cord stimulation. Neurosurgery 53(3), 626632.
should be implicated on separate days to confirm Jadad, A. R., Carroll, D., Glynn, C. J., and McQuay, H. J. 1995.
Intravenous regional sympathetic blockade for pain relief in
the diagnosis. Treatment options that have shown reflex sympathetic dystrophy: a systematic review and a
some promising results include local anesthetic sym- randomized, double-blind crossover study. J. Pain Symp.
pathetic blocks, SCS, radiofrequency techniques, and Manage. 10(1), 1320.
Janig, W. and Baron, R. 2002. Complex regional pain syndrome
surgical sympathectomy in early stages. Visceral pain is a disease of the central nervous system. Clin. Auton. Res.
of pancreatic origin responds well to chemical neu- 12(3), 150164.
rolytic blocks. More randomized trials are warranted Janig, W. and Baron, R. 2003. Complex regional pain syndrome:
mystery explained? Lancet Neurol. 2(11), 687697.
to prove long-term efficacy of neuromodulation as Janig, W. and Baron, R. 2004. Experimental approach to CRPS.
well as neurodestructive techniques. Pain Mar. 108(12), 37.
Kemler, M. A., De Vet, H. C., Barendse, G. A., Van Den
Wildenberg, F. A., and Van Kleef, M. 2004. The effect of
spinal cord stimulation in patients with chronic reflex
sympathetic dystrophy: two years follow-up of the
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Kurvers, H. A., Jacobs, M. J., Beuk, R. J., Van den
Ali, Z., Raja, S. N., Wesselmann, U., Fuchs, P. N., Meyer, R. A., Wildenberg, F. A., Kitslaar, P. J., Slaaf, D. W., and
and Campbell, J. N. 2000. Intradermal injection of Reneman, R. S. 1995. Reflex sympathetic dystrophy: evolution
norepinephrine evokes pain in patients with sympathetically of microcirculatory disturbances in time. Pain 60(3), 333340.
maintained pain. Pain 88(2), 161168. Mailis, A. and Furlan, A. 2003. Sympathectomy for neuropathic
Bandyk, D. F., Johnson, B. L., Kirkpatrick, A. F., pain. Cochrane Database Syst. Rev. (2), CD002918.
Novotney, M. L., and Back, M. R. 2002. Surgical Mekhail, N. A., Aeschbach, A., and Stanton-Hicks, M. 2004.
sympathectomy for reflex sympathetic dystrophy Cost benefit analysis of neurostimulation for chronic pain.
syndromes. J. Vasc. Surg. 35(2), 269277. Clin. J. Pain 20(6), 462468.
Baron, R. and Janig, W. 2004. Complex regional pain Merskey, H. and Bogduk, N. 1995. Classification of Chronic
syndromes how do we escape the diagnostic trap? Lancet Pain: Descriptions of Chronic Pain Syndromes and Definition
364(9447), 17391741. of Terms. IASP Press.
Baron, R., Schattschneider, J., Binder, A., Siebrecht, D., and Meyerson, B. and Linderoth, B. 2001. Spinal Cord Stimulation.
Wasner, G. 2002. Relation between sympathetic In: Bonicas Management of Pain, 3rd edn.
vasoconstrictor activity and pain and hyperalgesia in (eds. J. D. Loeser, S. D. Butler, C. R. Chapman, et al.),
complex regional pain syndromes: a case-control study. pp. 18571876. Lippincott Williams and Wilkins.
Lancet 359(9318), 16551660. Ochoa, J. and Verdugo, R. J. 2001. Mechanisms of neuropathic
Bruehl, S., Harden, R. N., Galer, B. S., Saltz, S., Backonja, M., pain: nerve, brain, and psyche: perhaps the dorsal horn but
and Stanton-Hicks, M. 2002. Complex regional pain not the sympathetic system. Clin. Auton. Res.
syndrome: are there distinct subtypes and sequential stages 11(6), 335339.
of the syndrome? Pain 95(12), 119124. Raja, S. N., Choi, Y., Asano, Y., Holmes, C., and Goldstein, D. S.
Campbell, J. N., Raja, S. N., Meyer, R. A., and Mackinnon, S. E. 1995. Arteriovenous differences in plasma concentrations of
1988. Myelinated afferents signal the hyperalgesia catechols in rats with neuropathic pain. Anesthesiology
associated with nerve injury. Pain 32(1), 8994. 83(5), 10001008.
Cepeda, M. S., Lau, J., and Carr, D. B. 2002. Defining the Raja, S. N., Treede, R. D., Davis, K. D., and Campbell, J. N.
therapeutic role of local anesthetic sympathetic blockade in 1991. Systemic alpha-adrenergic blockade with
complex regional pain syndrome: a narrative and systematic phentolamine: a diagnostic test for sympathetically
review. Clin. J. Pain 18(4), 216233. maintained pain. Anesthesiology 74(4), 691698.
Davis, K. D., Treede, R. D., Raja, S. N., Meyer, R. A., and Raja, S. N., Turnquist, J. L., mEleka, S., and Campbell, J. N.
Campbell, J. N. 1991. Topical application of clonidine 1996. Monitoring adequacy of alpha-adrenoceptor blockade
relieves hyperalgesia in patients with sympathetically following systemic phentolamine administration. Pain
maintained pain. Pain 47(3), 309317. 64(1), 197204.
Dellemijn, P. L., Fields, H. L., Allen, R. R., McKay, W. R., and Schurmann, M., Gradl, G., Wizgal, I., Tutic, M., Moser, C.,
Rowbotham, M. C. 1994. The interpretation of pain relief and Azad, S., and Beyer, A. 2001. Clinical and physiologic
sensory changes following sympathetic blockade. Brain evaluation of stellate ganglion blockade for complex regional
117(Pt 6), 14751487. pain syndrome type I. Clin. J. Pain 17(1), 94100.
Drummond, P. D. and Finch, P. M. 2004. Persistence of pain Stanton-Hicks, M., Janig, W., Hassenbusch, S., Haddox, J. D.,
induced by startle and forehead cooling after sympathetic Boas, R., and Wilson, P. 1995. Reflex sympathetic
blockade in patients with complex regional pain syndrome. dystrophy: changing concepts and taxonomy. Pain
J. Neurol. Neurosurg. Psychiatry 75(1), 98102. 63(1), 127133.
 Sympathetic Blocks for Pain 235

Stevens, R. A., Stotz, A., Kao, T. C., Powar, M., Burgen, S., and dystrophy (CRPS I): mechanisms and diagnostic value.
Kleinman, B. 1998. The relative increase in skin temperature Brain. 124(Pt 3), 587599.
after stellate ganglion block is predictive of a complete Wilkinson, H. A. 1996. Percutaneous radiofrequency upper
sympathectomy of the hand. Reg. Anesth. Pain Med. thoracic sympathectomy. Neurosurgery 38(4), 715725.
23(3), 266270.
Verdugo, R. J and Ochoa, J. L. 1994. Sympathetically
maintained pain. I. Phentolamine block questions the
concept. Neurology 44(6), 10031010.
Verdugo, R. J., Campero, M., and Ochoa, J. L. 1994. Further Reading
Phentolamine sympathetic block in painful
polyneuropathies. II. Further questioning of the concept of Janig, W., Levine, J. D., and Michaelis, M. 1996. Interactions of
sympathetically maintained pain. Neurology sympathetic and primary afferent neurons following nerve
44(6), 10101014. injury and tissue trauma. Prog. Brain Res. 113, 161184.
Wasner, G., Schattschneider, J., Heckmann, K., Maier, C., and Nathan, P. W. 1983. Pain and the sympathetic system. J. Auton.
Baron, R. 2001. Vascular abnormalities in reflex sympathetic Nerv. Syst. 7(34), 363370.
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 19 Sprouting in Dorsal Root Ganglia
E M McLachlan, Prince of Wales Medical Research Institute, Randwick, NSW, Australia
2009 Elsevier Inc. All rights reserved.

19.1 The Discovery 238

19.2 Sprouting of Sympathetic Axons within Dorsal Root Ganglia 239
19.3 Sprouting of Primary Afferent Neurons within Dorsal Root Ganglia 239
19.4 Formation of Perineuronal Rings or Baskets 239
19.5 Underlying Cellular Mechanisms 240
19.6 Other Functional Considerations 241
19.7 Other Forms of Sympathetic Plasticity after Nerve Injury 241
References 242

Glossary
A nociceptors They have often been ignored but p75 It is the low-affinity nerve growth factor (NGF)
there is accumulating evidence that a subpopula- receptor recognized by NGF, brain-derived neuro-
tion of large-diameter fast-conducting (A) sensory trophic factor (BDNF), neurotrophin-3 (NT-3), and
neurons are nociceptive (Djouhri, L. and Lawson, S. neurotrophin-4/5 (NT-4/5). p75 interacts with each
N., 2004). neurotrophin but with slightly different binding prop-
calcitonin gene-related peptide (CGRP) It is a erties. It has striking structural homology with a family
neuropeptide present in the majority of peptidergic of receptors, including tumor necrosis factor (TNF)
primary afferent nociceptor neurons. CGRP is also receptors, CD40, and Fas, consistent with it having
expressed in some medium to large diameter many functions.
afferent neurons and appears de novo in others. peptidergic sensory neurons These are small-
including gracile-projecting mechanosensitive diameter dorsal root ganglion neurons that contain
neurons, after nerve injuries (Ma, W. et al., 1999). a known neuropeptide (such as substance P, cal-
galanin It is a neuropeptide present in a subpo- citonin gene-related peptide, galanin,
pulation of CGRP-containing primary afferent somatostatin, and vasoactive intestinal polypep-
nociceptor neurons. It is upregulated both in these tide) and are usually nociceptors (Lawson, S. N.,
neurons and in sympathetic postganglionic neu- 2002).
rons after a peripheral nerve injury. perineuronal rings or baskets These are
neuropeptide Y (NPY) It is a neuropeptide that is arrangements of varicose nerve terminals formed
present in many sympathetic postganglionic vaso- by the endings of sympathetic neurons and pepti-
constrictor neurons in most species. It is dergic and nonpeptidergic sensory neurons within
upregulated in gracile-projecting low-threshold dorsal root ganglia that project into a lesioned
mechanosensitive primary afferent neurons after nerve trunk.
peripheral nerve lesions. retrograde reaction Refers to the response in the
nonpeptidergic sensory neurons These are soma of a neuron when its axon is severed.
small-diameter dorsal root ganglion neurons that Following axotomy, either a positive signal of injury
lack any known neuropeptide but express GFR1- or the arrest of the normal retrograde traffic from
2 and c-RET, components of the receptor complex the axon terminals reaches the soma within a few
for glial-derived neurotrophic factor, and bind iso- hours via the retrograde axoplasmic transport
lectin B4 (derived from Griffonia simplicifolia) mechanism. The neuron soma swells and the
(Bennett, D. L. et al., 1998), neither of which are nucleus may assume an eccentric position. These
features of peptidergic sensory neurons. structural changes are referred to chromatolysis as
Nonpeptidergic sensory neurons are usually noci- the chromophilic Nissl bodies are disrupted.
ceptive (Lawson, S. N., 2002). Protein synthesis is modified so that synthesis of
neurotransmitter substances is reduced and other

237
238 Sprouting in Dorsal Root Ganglia

activities such as regeneration are supported. The is upregulated after nerve injury prior to the cells
satellite glia around the soma also respond and proliferating around damaged neurons.
begin to release cytokines and chemokines. They substance P (SP) It is a neuropeptide present in a
proliferate and start to synthesize neurotrophins. subpopulation of peptidergic primary afferent noci-
satellite glia These are the support cells that sur- ceptor neurons. It is upregulated in gracile-projecting
round the cell bodies of primary afferent neurons low-threshold mechanosensitive primary afferent
within dorsal root ganglia. They normally express neurons after peripheral nerve lesions.
low levels of glial fibrillary acidic protein (GFAP) which

19.1 The Discovery Following early studies triggered by military inju-

ries, an explosion of research in the last 25 years has
Clinicians observe that, in a subgroup of patients with utilized mainly rodents with injuries to one sciatic
chronic neuropathic pain after peripheral nerve nerve that lead to diverse alterations in the damaged
injury of varying degrees of severity, sympathetic neurons, their supporting glia, the tissues they pre-
activity exacerbates or initiates pain, usually viously supplied, and the related vasculature. In
described as burning or stabbing (Bonica, J. J., 1990). addition inflammatory cells invade, not only the
The pain can be referred to superficial and/or deep lesion site, but also retrogradely along the nerve
sites and is often associated with allodynia and trunk and within L4/5 dorsal root ganglia (DRGs),
hyperalgesia. It has been postulated that, after the L3/4 sympathetic paravertebral ganglia, and the ven-
injury, nociceptors are activated by norepinephrine tral horn of L4/5 spinal cord (Aldskogius, H. and
released by sympathetic activity, for example, during Kozlova, E. N., 1998; Hu, P. and McLachlan, E. M.,
emotional stress or exposure to low environmental 2002; 2004). These changes persist for many months
temperatures. This idea was based on evidence that unless the damaged axons can regenerate to their
pain was relieved by blockade of (1) the activity of target tissues.
sympathetic postganglionic neurons by injecting Of relevance to injury-induced spontaneous pain
local anesthetics into paravertebral ganglia supplying is the generation of spontaneous activity in 2030%
the affected limb (Bonica, J. J., 1990), (2) norepi- of primary afferents within the damaged nerve. This
nephrine release by injecting guanethidine into the activity is thought to contribute to central sensitiza-
limb (Wahren, L. K. et al., 1991), or (3) the effects of tion in the dorsal horn leading to abnormal discharge
norepinephrine by injecting -adrenoceptor antago- of nociceptive pathways (Mannion, R. J. and Woolf,
nists such phentolamine intravenously (Raja, S. N. C. J., 2000). The ectopic activity arises at two sites: in
et al., 1991). Sympathetically maintained pain usually unmyelinated (C) neurons, it is initiated at the site of
develops days to weeks after the injury. the injury (Michaelis, M. et al., 1995); in myelinated
These observations are mysterious because sym- (A) neurons, it arises from oscillations in the mem-
pathetic nerve terminals are not co-located with brane potential of the somata in the DRG (Kajander,
nociceptor neurons or their terminals. In the skin, K. C. et al., 1992; Liu, C. N. et al., 2000). When the
sympathetic axons terminate on arteriolar vessels DRG and its connections are intact in vivo, activity in
deep within the dermis whereas nociceptor terminals A axons arises in muscle but not cutaneous afferents
lie below and within the epidermis. Sensory and (Michaelis, M. et al., 2000).
sympathetic terminals do not usually co-locate in The ectopic discharge in some DRG neurons can
deeper tissues, other than rare axons containing sub- be generated or modulated by sympathetic activity.
stance P (SP) and calcitonin gene-related peptide Wilfrid Janig recorded from single myelinated afferent
(CGRP) that run among the sympathetic perivascu- axons projecting down a ligated sciatic nerve
lar terminals on blood vessels (Holzer, P., 1992). (McLachlan, E. M. et al., 1993). Stimulation of the
There is no evidence for -adrenoceptors on noci- sympathetic supply at high frequency (50 Hz for 10 s)
ceptor terminals in normal skin and subcutaneous excited a subgroup of afferents. Intravenous application
introduction of norepinephrine produces blanching of phentolamine completely inhibited the response,
but no pain. confirming that the sympathetic-sensory link was
 Sprouting in Dorsal Root Ganglia 239

located proximally. Histochemical demonstration of sprouts appear sooner and are more prolific when the
norepinephrine revealed perivascular terminals out- sciatic nerve is only partially transected (Shir, Y. and
side the ganglion sprouting through the DRG Seltzer, Z., 1991) or subjected to chronic constriction
(McLachlan, E. M. et al., 1993). After a few weeks, (CCI) (Bennett, G. J. and Xie, Y. K., 1988) rather than
some sprouting axons had formed interwoven baskets transection. These injuries, like spinal nerve transec-
or rings of varicose terminals around a few large-dia- tion, leave intact axons lying in regions of Wallerian
meter neurons. These novel structures appeared to be degeneration and the ensuing inflammatory reaction
the anatomical basis for a functional link between damages more axons. Such injuries produce mechanical
sympathetic and sensory systems after injury. allodynia and thermal hyperalgesia within a few days.

19.2 Sprouting of Sympathetic Axons 19.3 Sprouting of Primary Afferent

within Dorsal Root Ganglia Neurons within Dorsal Root Ganglia

Sprouting of norepinephrine-containing terminals One possible trigger for sprouting of perivascular

into DRGs after injury was soon confirmed in many terminals is an increased local concentration of neu-
laboratories (e.g., Chung, K. et al., 1996; Kim, H. J. rotrophins, such as nerve growth factor (NGF)
et al., 1996; Ramer, M. S. and Bisby, M. A., 1997), (Herzberg, U. et al., 1997; see below). Consistent
mainly using immunohistochemistry to demonstrate with this, sensory neurons containing SP and
tyrosine hydroxylase (TH), the rate-limiting enzyme CGRP which bear the appropriate receptors (trkA)
for the synthesis of norepinephrine. The time course sprout within the lesioned DRG, forming similar
of development of sprouts varies with the site and numbers of perineuronal rings (McLachlan, E. M.
nature of the lesion (Ramer, M. S. et al., 1999). In rats, and Hu, P., 1998). Galanin appears in some sprouts
the sprouts develop over a few weeks after sciatic of both sympathetic and peptidergic origin (Hu, P.
transection and form increasing numbers of perineur- and McLachlan, E. M., 2001).
onal rings after 4 weeks. The number of rings There is one report (Li, L. and Zhou, X. F., 2001)
reaches a maximum (2.5% of neurons or 7.5% that the nonpeptidergic primary afferent axons that
of A neurons) between 8 and 12 weeks and then bind Griffonia simplicifolia I isolectin B4 (IB4) also
gradually declines over 1 year or more (Hu, P. and sprout within L5 DRG after spinal nerve transection.
McLachlan, E. M., 2001). While sympathetic sprouts Increasingly from 5 weeks after the lesion, IB4
appear in normal aged animals (Ramer, M. S. and axons were found to encircle mainly large-diameter
Bisby, M. A., 1998b), this cannot account for the neuron somata where they were intermingled with
raised proportion of rings on the side ipsilateral to proliferated satellite cells that also bound IB4. In
the lesion. Some perineuronal rings may persist inde- some cases, TH axons were present around the
finitely after injury, probably accounting for their same neurons. What is amazing is that tenfold more
presence in humans as described by Ramon y Cajal neurons received IB4 perineuronal rings than
(Garcia-Poblete, E. et al., 2003). The extent of sprout- TH, CGRP, or galanin rings at the same post-
ing is generally greater if the lesion is closer to the operative time (McLachlan, E. M. and Hu, P., 1998;
DRG (Kim, H. J. et al., 1996; Ramer, M. S. and Bisby, Hu, P. and McLachlan, E. M., 2001). However, only a
M. A., 1998a) and if more axons are cut (Kim, H. J. few of the IB4 axons also stained for protein gene
et al., 2001), although the relationships are not linear product (PGP) 9.5, a pan-neuronal marker (ubiquitin
and may depend on the proportions of cutaneous and C-terminal hydroxylase).
muscle axons involved (Hu, P. and McLachlan, E. M.,
2003). Sprouting of sympathetic terminals into DRGs
after sciatic transection does not occur in guinea-pigs 19.4 Formation of Perineuronal Rings
that lack sympathetic terminals on nearby vessels or Baskets
(Hu, P. and McLachlan, E. M., 2000).
Sympathetic sprouts are already quite extensive Perineuronal rings form more often at either pole
after 7 days if the spinal nerve is transected (Chung, of the DRG where large-diameter neurons are
K. et al., 1996; Ramer, M. S. et al., 1998). Cutting L5 clustered (McLachlan, E. M. et al., 1993). The
spinal nerve leads to hyperalgesia within a few days in neurons that receive them can be axotomized or
the territory of the intact root (L4). Sympathetic intact (Jones, M. G. et al., 1999; Hu, P. and
240 Sprouting in Dorsal Root Ganglia

(a) (b) rats with nerve transection reduced the amount of

sympathetic sprouting and the number of TH rings
(Zhang, J. M. et al., 2004), as did its application locally
at the time of the lesion. These data suggest that
activity derived from the neuroma is important for
initiating the sprouting response.
Sympathetic axons sprout from two sources: (1)
from intact perivascular terminals, which occur after
a distant lesion, and are dependent on NGF and (2) by
retrograde growth along the spinal nerve, which
occurs when anterograde growth is obstructed at the
lesion, and is NGF-independent. The former predo-
minates after a distant injury, and the latter after spinal
nerve ligation (Ramer, M. S. and Bisby, M. A., 1999).
Figure 1 Perineuronal rings in rat L5 dorsal root ganglia 10 However, a common response to axotomy is the
weeks after sciatic nerve transection and ligation. Two pairs sprouting of collaterals from a transected axon within
of micrographs (a) and (b) show immunofluorescence for a short distance from the soma (Kelly, M. E. M. et al.,
tyrosine hydroxylase (TH, green) and synaptophysin (Syn, 1989) and this may occur after lesions to either sym-
red). (a) TH terminals that lack Syn (thin arrows) wrap
around a neuron soma adjacent to another (A) that bears a
pathetic or sensory axons.
dense basket of Syn terminals. (b) A neuron receives a Upregulation of neurotrophin messenger ribonu-
TH/Syn perineuronal ring (thick arrows). TH axons in the cleic acid (mRNA) by satellite glia leads to their local
small nerve bundle to the left in (b) do not express Syn but a production within the DRG, stimulating growth of
sprouting axon in the main axon bundle (arrowhead) is Syn. axons bearing appropriate receptors. This has been
shown for NGF and neurotrophin-3 (NT-3) which
McLachlan, E. M., 2003). The varicose terminals are synthesized by the proliferated satellite glia that
intertwine around the somata but normally remain surround large-diameter neurons. Treatment with
within the layers of proliferated satellite glia, very antibodies to NGF or NT-3 reduces the length of
rarely forming contact with neuron somata (Chung, TH sprouts after a spinal nerve lesion (Ramer, M.
K. et al., 1997; Shinder, V. et al., 1999). S. and Bisby, M. A., 1999; Zhou, X. F. et al., 1999) as
Sometimes isolated neurons are targeted, or more both trkA and trkC are present on sympathetic and
than one type of axon can encircle the same neuron sensory axons. Brain-derived neurotrophic factor
(McLachlan, E. M. and Hu, P., 1998) or, in other (BDNF) is upregulated in lesioned DRG neurons
cases, adjacent neurons receive a ring from one or and antibodies to BDNF also inhibit sympathetic
other axon type (Figure 1). Synaptophysin, a non- sprouting (Deng, Y. S. et al., 2000). These approaches
specific marker of sprouting axons, appears in rings at do not seem to have been applied to examine sprout-
a similar density to those with TH, CGRP, or galanin ing of sensory neurons. Glial cell-derived
(Hu, P. and McLachlan, E. M., 2003). This may result neurotrophic factor (GDNF) mRNA is present in
because synaptophysin is only partly co-located with satellite cells in DRGs and is upregulated after injury
these substances or because of nonpeptidergic axon (Hammarberg, H. et al., 1996), providing a basis for
sprouts. As synaptophysin is also a synaptic vesicle the sprouting of IB4 axons (Li, L. and Zhou, X. F.,
protein, its presence in some but not all varicosities 2001). Intrathecal GDNF does not elicit sympathetic
within rings supports them being release sites. sprouts (Jones, M. G. et al., 1999) but leukemia inhi-
Unfortunately, it is likely to be impossible to measure bitory factor does (Thompson, S. W. and Majithia, A.
release directly from these terminals. A., 1998), perhaps by an effect that modifies NGF
production.
The formation of perineuronal rings has been stu-
19.5 Underlying Cellular died in trigeminal ganglia of transgenic mice. When
Mechanisms these animals overexpress NGF (Davis, B. M. et al.,
1994), TH terminals preferentially target trkA
Systemic lidocaine is successful in treating some CGRP sensory neurons that contain NGF (Walsh,
forms of neuropathic pain (Wallace, M. S. et al., G. S. and Kawaja, M. D., 1998; Walsh, G. S. et al.,
2000). Delivery of lidocaine by osmotic pump to 1999). The data indicate that CGRP axons do not
 Sprouting in Dorsal Root Ganglia 241

sprout in these animals. If NGF or trkA is deleted, the mechanoreceptors. The absence of sympathetic
TH axons do not enter the ganglion. If p75 is rings around peptidergic neurons (Ramer, M. S. and
deleted but NGF is overexpressed, abundant TH Bisby, M. A., 1998c) excludes the gracile-projecting A
sprouts wander through the DRG without associating neurons which express SP, CGRP, NPY, and galanin
with specific neuron somata (Walsh, G. S. et al., 1999). several weeks after a peripheral injury (Zhang, X.
These elegant experiments do not help to explain et al., 1993; Noguchi, K. et al., 1995; Ma, W. and
sprouting or the formation of rings after nerve injury Bisby, M. A., 1997; Ma, W. et al., 1999). Varicose
in normal adult animals. As mentioned above, NGF peptidergic rings do not form around peptidergic
is synthesized by satellite glia that express p75 neurons (McLachlan, E. M. and Hu, P., 1998).
whereas trkA is downregulated on neuron somata However, 20% of A neurons in rat L5 DRG are
(Verge, V. M. et al., 1989). Further, no sprouts appear A-nociceptors (Djouhri, L. and Lawson, S. N., 2004)
in the rat trigeminal ganglion after transection of the that is, 6% of all neurons, and at least some have large
infraorbital or inferior alveolar nerve quite close to diameters. Allowing that TH terminals make up
the ganglion (Bongenhielm, U. et al., 1999) or chronic only 50% of rings, rings may occur on 6% of
constriction injury (CCI) of the mental nerve (Grelik, neurons, making it feasible that A-nociceptor neu-
C. et al., 2005). This seems surprising as sympathetic rons are the targets. While trkA is currently the only
axons join the nerve immediately distal to the tri- specific marker of these A-neurons (Fang, X. et al.,
geminal ganglion as for the DRG. Perhaps the 2005), this is downregulated after axotomy (Verge, V.
absence of large-diameter proprioceptive neurons M. et al., 1989; Li, L. et al., 2000) so that it may not be
(located in the mesencephalic nucleus V) prevents possible to test this idea.
the changes that trigger sympathetic sprouting in Despite evidence that -adrenoceptors are
lesioned DRGs. Whether peptidergic neurons in tri- expressed de novo by a small proportion of largely
geminal ganglia sprout after nerve lesions does not medium-diameter neurons in lesioned DRGs
seem to have been examined. (Birder, L. A. and Perl, E. R., 1999), there is limited
Nevertheless it seems likely that the expression of evidence that norepinephrine can excite lesioned A
p75 on proliferated glia in rat DRGs is important for neurons. Although norepinephrine can depolarize a
ring formation. In guinea-pigs, in which sympathetic small proportion of both control and damaged neu-
sprouting does not occur (see above), peptidergic rons in intact DRGs in vitro (Jones, M. G. et al., 1999),
axons sprout into the DRG but do not form perineur- this does not seem to be -adrenoceptor mediated
onal rings after sciatic transection. This might be (Lopez de Armentia, M. et al., 2003; cf. Xing, J. L. et al.,
explained by both the absence of glial reaction to 2003). For neuropeptides, neuron somata with mye-
axotomy (i.e., no upregulation of glial fibrillary acidic linated axons lack CGRP binding sites (Segond von
protein) and the failure of glia to express p75 in this Banchet, G. et al., 2002), although they can develop
species (Hu, P. and McLachlan, E. M., 2000). sensitivity to galanin and SP after axotomy (Xu, Z. Q.
What initiates neurotrophin production? It might et al., 1997; Abdulla, F. A. et al., 2001). This has not
follow the retrograde reaction in satellite glia and the been demonstrated in situ.
subsequent invasion of the DRG by macrophages and
lymphocytes (Hu, P. and McLachlan, E. M., 2002).
Following sciatic transection, macrophage and T-cell
density peaks after 1 week and remains raised for 19.7 Other Forms of Sympathetic
several months. Lymphocytes and macrophages Plasticity after Nerve Injury
secrete neurotrophins and proinflammatory cyto-
kines. Neurotrophins trigger sprouting and cytokines Is there another explanation for the enhanced
sensitize DRG neurons. Further, T cells contribute to responses to sympathetic activation after injury?
injury-induced pain (Moalem, G. et al., 2004). The normal consequence of sympathetic activity is
vasoconstriction which is abolished by phentolamine.
Stimulation paradigms that excite DRG neurons
19.6 Other Functional (>10 Hz) elicit marked vasoconstriction in most
Considerations vascular beds. Stimulation of the lumbar sympathetic
chain augmented discharge in a proportion of
The somata around which perineuronal rings myelinated dorsal root axons from spinal nerve-
form have been thought to be low-threshold injured rats only when vascular resistance in the
242 Sprouting in Dorsal Root Ganglia

DRG was near maximal (Habler, H. J. et al., 2000). sprouting fails to occur in the trigeminal ganglion after
peripheral nerve injury in the rat. Pain 82, 283288.
Discharge was also increased by vasoconstrictor Bonica, J. J. 1990. The Management of Pain. Lea and Febiger.
agents and blockade of nitric oxide synthase. These Chung, K., Lee, B. H., Yoon, Y. W., and Chung, J. M. 1996.
data imply that enhanced neurovascular responses Sympathetic sprouting in the dorsal root ganglia of the
injured peripheral nerve in a rat neuropathic pain model. J.
after a lesion jeopardize perfusion of the DRG, Comp. Neurol. 376, 241252.
and raise the possibility that ischemia may be Chung, K., Yoon, Y. W., and Chung, J. M. 1997. Sprouting
responsible for increasing excitability of sensory sympathetic fibers form synaptic varicosities in the dorsal
root ganglion of the rat with neuropathic injury. Brain Res.
neurons. 751, 275280.
Finally, the phenomenon of sympathetic sprout- Davis, B. M., Albers, K. M., Seroogy, K. B., and Katz, D. M.
ing in lesioned DRGs has had attention because of its 1994. Overexpression of nerve growth factor in transgenic
mice induces novel sympathetic projections to primary
possible relation to sympathetically maintained neu- sensory neurons. J. Comp. Neurol. 349, 464474.
ropathic pain. It is, however, important to note the Deng, Y. S., Zhong, J. H., and Zhou, X. F. 2000. Effects of
failure of sympathectomy to modify ectopic activity endogenous neurotrophins on sympathetic sprouting in the
dorsal root ganglia and allodynia following spinal nerve
in anesthetized rats (Liu, X. G. et al., 2000) or injury. Exp. Neurol. 164, 344350.
mechanical allodynia (Ringkamp, M. et al., 1999), Djouhri, L. and Lawson, S. N. 2004. A-fiber nociceptive
and the lack of relation between the extent of sympa- primary afferent neurons: a review of incidence and
properties in relation to other afferent A-fiber neurons in
thetic sprouting and pain behavior (Kim, H. J. et al., mammals. Brain Res. Rev. 46, 131145.
1999). Thus, although sympathetic perineuronal Fang, X., Djouhri, L., McMullan, S., Berry, C., Okuse, K.,
rings have been postulated to underlie ectopic activ- Waxman, S. G., and Lawson, S. N. 2005. trkA is expressed in
nociceptive neurons and influences electrophysiological
ity, evidence for this in experimental animals is not properties via Nav1.8 expression in rapidly conducting
convincing. Nevertheless the experiments on sprout- nociceptors. J. Neurosci. 25, 48684878.
ing have provided insights into plasticity in the Garcia-Poblete, E., Fernandez-Garcia, H., Moro-Rodriguez, E.,
Catala-Rodriguez, M., Rico-Morales, M. L., Garcia-Gomez-
damaged nervous system. de-las-Heras, S., and Palomar-Gallego, M. A. 2003.
Sympathetic sprouting in dorsal root ganglia (DRG): a recent
histological finding? Histol. Histopathol. 18, 575586.
Grelik, C., Bennett, G. J., and Ribeiro-da-Silva, A. 2005.
Acknowledgments Autonomic fibre sprouting and changes in nociceptive
sensory innervation in the rat lower lip skin following chronic
constriction injury. Eur. J. Neurosci. 21, 24752487.
Work in the authors laboratory was supported by the Habler, H. J., Eschenfelder, S., Liu, X. G., and Janig, W. 2000.
National Health & Medical Research Council and Sympathetic-sensory coupling after L5 spinal nerve lesion in
the rat and its relation to changes in dorsal root ganglion
the Motor Accidents Authority of New South Wales. blood flow. Pain 87, 335345.
Hammarberg, H., Piehl, F., Cullheim, S., Fjell, J., Hokfelt, T., and
Fried, K. 1996. GDNF mRNA in Schwann cells and DRG satellite
cells after chronic sciatic nerve injury. Neuroreport 7, 857860.
Herzberg, U., Eliav, E., Dorsey, J. M., Gracely, R. H., and
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 20 Vagal Afferent Neurons and Pain
W Janig, Christian-Albrechts-Universitat zu Kiel, Kiel, Germany
2009 Elsevier Inc. All rights reserved.

20.1 Introduction 246

20.2 Vagal Afferents, Pain, and Nociception 246
20.2.1 Thoracic Visceral Organs 246
20.2.2 Pelvic and Abdominal Organs 247
20.3 Abdominal Vagal Afferents, Protection of the Body, and Illness Responses 247
References 249

Glossary
allodynia, hyperalgesia See Chapter Autonomic with small-diameter myelinated (A delta) fibers)
Nervous System and Pain. generate upon excitation precapillary (arteriolar)
angina (pectoris) A paroxysmal thoracic pain, vasodilation (by release of calcitonin gene-related
with a feeling of suffocation and impending death, peptide (CGRP) supported by release of substance
due, most often but not always, to anoxia of the P and neurokinin A) and postcapillary (venular)
myocardium. plasma extravasation (by release of substance P
GALT (gut-associated lymphoid tisue) Immune and neurokinin A). Originally venular plasma extra-
system of the gastrointestinal tract, in particular in vasation was called neurogenic inflammation, but
the small intestine. It includes the immune cells of both precapillary vasodilation and postganglionic
Peyers patches, M-cells of the epithelial lining plasma extravasation are collectively now called
modified entestinal epithelial cells (enterocytes), neurogenic inflammation.
lyphocytes in the almina propria, macrophages, referred pain See Chapter Autonomic Nervous
and mast cells. Its function is to defend the body System and Pain.
against invading antigens from food, bacteria, vagal afferents Afferents projecting through the
parasites, and toxins. vagal nerves. Most of their cell bodies are located in
neurogenic inflammation Peptidergic primary the inferior vagal (nodose) ganglion and a few in the
afferent neurons with unmyelinated fibers (some superior vagal (jugular) ganglion.

1. Vagal afferents innervating the heart may be interleukin-1 (IL-1) or tumor necrosis factor
involved in pain during angina referred to the  (TNF-)) into the peritoneal cavity. One com-
upper cervical dermatomes and myotomes. ponent of the illness responses is hyperalgesic
2. Vagal afferents innervating the stomach and being behavior.
excited by acid are involved in nociceptive pro- 5. Activity in abdominal vagal afferents innervating
tective reactions, but probably not in conscious the small intestine and probably sensing toxic and
perception of pain. other events in the gastrointestinal tract (GIT)
3. Transmission of nociceptive impulses in the spinal may be involved in reflex modulation of inflam-
or trigeminal dorsal horn is under inhibitory con- mation and sensitivity of nociceptors in somatic
trol generated by activity in cardiopulmonary tissues via the sympatho-adrenal system (releasing
and/or abdominal vagal afferents. adrenaline) and possibly the hypothalamo-pitui-
4. Vagal afferents innervating the liver (thus passing taryadrenal system.
through the hepatic branch of the abdominal
The response properties of vagal afferents to physio-
vagus nerves) are involved in illness responses
logical stimuli being involved in the reflex
generated by lipopolysaccharides (LPS) (bacterial
modulation of nociception, pain, hyperalgesia,
antigens) or proinflammatory cytokines (e.g.,
inflammation, and illness responses are unknown. It

245
246 Vagal Afferent Neurons and Pain

is likely that not one functionally specific type of esophagus and trachea as well as from the bronchi
vagal afferent neuron is involved but several different may be generated by stimulation of spinal visceral
types. afferent neurons yet not of vagal afferents. However,
this situation is unclear and needs further
experimentation.
20.1 Introduction Cardiac pain (e.g., during ischemic heart disease)
is considered to be mediated by activation of spinal
The vagus nerves contain 8085% visceral afferent visceral afferents having their cell bodies in the dorsal
fibers, most of them innervating the GIT, and 15 root ganglia C8T9 (mainly T26). However,
20% preganglionic parasympathetic fibers, at least in attempts to relieve cardiac angina pain by surgical
rats (Precht, J. C. and Powley, T. L., 1990; Berthoud, interventions (cervico-thoracic sympathectomy,
H. R. et al., 1991). These afferent neurons are dorsal rhizotomy, injection of alcohol into the
involved not only in organ regulation by the auto- sympathetic chain) consistently showed that only
nomic nervous system (Janig, W., 2006) but also in 5060% of patients report complete relief from
the control of protective body reactions that include angina while the remaining patients report either
nociception and pain. These functions are inferred partial relief or no relief at all. Some failures to relief
from various types of experimental investigations pain may have been attributed to incomplete spinal
and from some clinical observations. It is unclear denervation of the heart; but vagal afferents innervat-
whether functionally specific groups of vagal affer- ing particularly the inferiorposterior part of the
ents are involved in protective body reactions or heart may mediate cardiac pain too (see Meller,
whether vagal afferents being involved in organ reg- S. T. and Gebhart, G. F., 1992).
ulation are also responsible for protective body Electrical stimulation of cervical vagal afferent
reactions. In this chapter I will summarize: fibers in rats modulates the nociceptive tail reflex as
well as the responses of dorsal horn neurons in the
1. data related to the role of vagal afferents in gen- lumbar spinal cord. Low-intensity electrical vagal sti-
erating pain and in modulating central processing mulation, probably exciting mainly myelinated
of nociceptive impulse activity and afferents, facilitates both whereas high-intensity stimu-
2. data about the role of vagal afferents in protective lation, probably exciting mainly C-fibers, attenuates
reactions of the body including inflammation both (Randich, A. and Gebhart, G. F., 1992; Ren, K.
and hyperalgesia (Janig, W., 2005; Janig, W. and et al., 1993). Nociceptive reflexes in neurons of the
Levine, J. D., 2006). trigeminal nucleus caudalis or oralis of rats were
mainly suppressed during electrical stimulation of cer-
vical vagal afferents (Bossut, D. F. and Maixner, W.,
20.2 Vagal Afferents, Pain, and 1996; Takeda, M. et al., 1998). Neurophysiological
Nociception investigations in monkeys and rats show that some
spino-thalamic tract (STT) neurons in the superficial
20.2.1 Thoracic Visceral Organs
dorsal horn and deeper laminae of the cervical seg-
Pain elicited from the proximal esophagus and prox- ments C1C2 (C3) can be synaptically activated by
imal trachea is probably mediated by vagal afferents electrical stimulation of cardiopulmonary spinal and
innervating the mucosa of these organs. These affer- vagal afferents or by injection of algogenic chemicals
ents are peptidergic (i.e., contain calcitonin gene in the pericardial sac via both afferent pathways. The
related peptide (CGRP) and/or substance P); their activation of the STT neurons by vagal afferents is
cell bodies are probably located in the jugular gang- relayed through the nucleus tractus solitarii. These
lion (superior vagal ganglion; see Berthoud, H. R. and STT neurons are also synaptically activated by
Neuhuber, W. L., 2000) cells in the nodose ganglion mechanical stimulation of the somatic receptive fields
(inferior vagal ganglion) are almost exclusively pep- in the corresponding segments from the head, jaw,
tide-negative, whereas neurons in the jugular neck, and shoulder (dermatomes, myotomes); this is
ganglion may contain peptides). Activation of these fully in line with clinical observations showing that
afferents generates neurogenic inflammation in the cardiac pain may be referred to neck, shoulder, and
mucosa (venular plasma extravasation and vasodila- jaw (Lindgren, I. and Olivecrona, H., 1947; White, J. C.
tion (McDonald, D. M. et al., 1988; McDonald, D. M., and Bland, E. F., 1948; Meller, S. T. and Gebhart, G. F.,
1990)). Pain elicited from the more distal sections of 1992). The dorsal horn of the same high cervical spinal
 Vagal Afferent Neurons and Pain 247

segments contains neurons with similar convergent meal. But acid reflux or an obstructed or distended
synaptic inputs from vagal, spinal visceral, and somatic viscus usually does not generate discomfort and pain
afferents that project to more caudal spinal thoracic, (Juler, G. L. and Eltorai, I. M., 1985; Strauther, G. R.
lumbar, and sacral segments. These dorsal horn neu- et al., 1999). An acute perforation of a duodenal ulcer
rons are involved in the inhibitory control of may be accompanied by violent pain in the right or left
nociceptive impulse transmission (see Foreman, R. D., shoulder (page 284 in Guttmann, L., 1976), indicating
1999; Chandler, M. J. et al., 2002; see also Section 20.3, that vagal afferents may be involved.
point 2). It is generally accepted that stimulation of vagal
Experimental studies on animals showing that afferents innervating the stomach elicits emesis, bloat-
nociceptive behavior and inhibition of nociceptive ing, and nausea, all three being protective reactions.
impulse transmission in the spinal and trigeminal dor- Experiments on rats show that influx of acid into and
sal horn is inhibited by electrical stimulation of the other chemical insults of the gastroduodenal mucosa
cervical vagus nerve have led to the idea that contin- lead to a host of locally and centrally organized pro-
uous electrical stimulation of vagal afferents could be tective reactions that are mediated by spinal visceral
used to treat chronic pain syndromes in patients. and vagal afferent neurons. Activation of vagal affer-
Electrical stimulation of the vagus nerve via a pair of ents by these chemical stimuli excites neurons in the
electrodes implanted at the left cervical vagus nerve nucleus tractus solitarii, area postrema, lateral para-
has been used recently in patients with some success to brachial nucleus, subceruleus nucleus, thalamic and
treat drug-resistant refractory epilepsy. The mechan- hypothalamic paraventricular nuclei, supraoptic
isms underlying this antiepileptic effect are unknown nucleus, and central amygdala, but not in the insular
(Schachter, S. C., 2005). A few experimental studies cortex (the major central representation area of the
have been conducted on these patients to test whether stomach) as shown by the expression of the marker
experimental pain can be attenuated by vagal nerve protein Fos (Michl, T. et al., 2001). Furthermore,
stimulation. Furthermore, in patients with chronic neurons in the spinal dorsal horn do not seem to be
headache (migraine, tension-type headache, cluster activated. Thus, vagal afferents innervating the gas-
headache) electrodes have been implanted at the left troduodenal mucosa (Holzer, P. and Maggi, C. A.,
cervical vagus nerve. In these experiments, in pilot 1998; Holzer, P.; 2002; 2003; 2006) (1) seem to be
studies, as well as in some case reports it has been involved in chemonociception, local protective reac-
shown that electrical vagal nerve stimulation may tions, and generation of autonomic, endocrine, and
reduce experimental pain or relieve chronic headache behavioral protective reactions but (2) not in pain
(for literature and discussion see Multon, S. and perception, yet in the emotional aspect of pain and
Schoenen, J., 2005). therefore indirectly in upper abdominal hyperalgesia.
These fascinating ideas, formulated by Holzer,
need verification by further experimentation.
20.2.2 Pelvic and Abdominal Organs
For example, it is unclear in which way (1) activity
Visceral pain elicited from pelvic and most abdominal in spinal visceral afferents and vagal afferents
organs is triggered by stimulation of spinal visceral is centrally integrated in vivo so as to elicit
afferent neurons and not by stimulation of vagal affer- the protective reflexes, protective behavior, and
ent neurons (Cervero, F., 1994; see Chapter Spinal pain sensations including visceral hyperalgesia and
Cord Mechanisms of Hyperalgesia and Allodynia). (2) activity in vagal afferents is responsible for
However, the situation is not entirely clear for the the emotional aspects of visceral pain, but spinal
gastroduodenal section of the GIT. Whether patients visceral afferents for the conscious perception of
with complete interruption of spinal ascending visceral pain.
impulse transmission at the thoracic level T1 or at a
more rostral segmental level can experience pain from
the gastroduodenal section (e.g., during gastritis, a 20.3 Abdominal Vagal Afferents,
peptic ulcer, or distension of the stomach) has never Protection of the Body, and Illness
been systematically investigated. Patients with com- Responses
plete lesion of the cervical spinal cord experience
abdominal hunger, dread, and nausea (Crawford, J. P. The small intestine and liver are very vulnerable
and Frankel, H. L., 1971). Furthermore, these patients portals of entries into the body. Both have potent
may experience vague sensations of fullness after a hot local defense systems and serve as internal defense
248 Vagal Afferent Neurons and Pain

lines of the body. The small intestine contains a deleterious for the GIT and for the body (Walls,
powerful immune system (the gut associated lym- E. K. et al., 1995a; 1995b). Additionally, some vagal
phoid tissue (GALT) (Shanahan, F., 1994)) that is afferents that innervate the small intestine and the
innervated by vagal afferents projecting through the liver respond to cytokines (e.g., IL-1); these
celiac branches of the abdominal vagus nerve. afferents may encode events that are related to
Specific modulation of activity in these afferents in the immune system of the GIT and liver
conjunction with the reaction of the GALT may act (Niijima, A., 1996).
as an early warning system to the rest of the body by 2. Electrical stimulation of abdominal vagal afferents
transmitting important information to the brain about exerts inhibition or facilitation of central nocicep-
toxic events and agents in the small intestine that are tive impulse transmission in the spinal dorsal horn
dangerous for the organism (Berthoud, H. R. and and depresses nociceptive behavior depending on
Neuhuber, W. L., 2000). Thus, particularly vagal as to whether unmyelinated or myelinated vagal
afferents in the celiac branches of the abdominal afferents are excited (Randich, A. and Gebhart, G.
vagus nerves that innervate the small intestine (in F., 1992). Electrical stimulation of cervical vagal
addition to the distal duodenum and the proximal afferents in monkeys suppresses transmission of
colon) and vagal afferents in the hepatic branch that impulse activity in STT neurons with nociceptive
innervate the liver (in addition to the proximal duo- function at all levels of the spinal cord. Electrical
denum, pancreas, and pylorus) may be important for stimulation of subdiaphragmatic vagal afferents
protective functions of the GIT and the body: has no effect on STT neurons in this species,
arguing that particularly cardiopulmonary vagal
1. Vagal afferents in the celiac branches of the afferents are involved in this inhibitory control.
abdominal vagus nerve monitor chemical and The central pathways mediating the inhibitory
mechanical events that occur in the intestine effect are neurons in the subceruleusparabrachial
under physiological and pathophysiological complex (noradrenergic) and neurons in the
conditions (for review see Grundy, D. and nucleus raphe magnus of the rostroventromedial
Scratcherd, T., 1989), that is, are related to medulla (serotonergic) that project to the spinal
meals, ingestion of toxic substances, inflammation, cord (Foreman, R. D., 1989). The central pathways
obstruction. These afferents respond to distension mediating the facilitatory effect are mediated by
or contraction of the small intestine or to intra- suprapontine pathways (Gebhart, G. F. and
luminal chemical stimulation (e.g., maltose, Randich, A., 1992; Randich, A. and Gebhart, G.
glucose, protein products of long-chain lipids, or F., 1992). The functional types of vagal afferents
intraluminal osmotic stimuli). The responses to being involved in inhibitory and facilitatory con-
chemical stimuli are mediated by enterochromaf- trol of nociceptive impulse transmission are
fin cells releasing 5-HT and by 5-HT3 receptors unknown.
on the terminals of the vagal afferents (Zhu, J. X. 3. Activity in vagal afferents innervating the small
et al., 2001) or enteroendocrine cells releasing intestine (i.e., projecting through the celiac
cholecystokinin (CCK) and the CCKA receptor branches of the abdominal vagus nerves) is impor-
on the terminals of the vagal afferent neurons tant in reflex modulation of inflammatory processes
(Richards, W. et al., 1996; Lal, S. et al., 2001). (e.g., in the knee joint) and mechanical hyperalgesic
Vagal afferent neurons may be associated with behavior (by sensitization of nociceptors) in remote
the GALT and may be excited by inflammatory body tissues involving the sympatho-adrenal sys-
processes and toxic processes, also probably tem and possibly the hypothalamo-pituitary
mediated by enterochromaffin cells releasing ser- adrenal system (Green, P. G. et al., 1995; 1997;
otonin (5-HT), enteroendocrine cells releasing Miao, F. J.-P. et al., 1997a;1997b; Khasar, S. G.
CCK, or mast cells releasing histamine and other et al., 1998a; 1998b; Janig, W. et al., 2000;
compounds (Kirkup, A. J. et al., 2001; Kreis, M. E. Miao, F. J. -P. et al., 2000, Miao; F. J. -P.et al., 2001;
et al., 2002). The functional specificity of these Khasar, S. G. et al., 2003; Miao, F. J. -P. et al., 2003a;
afferents with respect to the different types of 2003b; see Section 5.17.3.4.4 and 5.17.4 in Chapter
intraluminal stimuli is unknown. The vagal affer- Autonomic Nervous System and Pain.
ents are important for preabsorptive detection of 4. Intraperitoneal injection of illness-inducing
energy-yielding molecules and probably for other agents, such as the bacterial cell wall endotoxin
properties of nutrient solutions that are toxic and LPS in rats, produces behavioral hyperalgesia.
 Vagal Afferent Neurons and Pain 249

This is mediated by activity in subdiaphragmatic immune system (proinflammatory cytokines

vagal afferents, specifically afferents running IL-1, TNF-, IL-6) trigger via different centers
in the hepatic branch. It is suggested that in brain stem and hypothalamus illness responses,
LPS activates hepatic macrophages (Kupffer one component being pain and hyperalgesia.
cells), dendritic cells, and leukocytes that release The underlying mechanisms of these illness
IL-1 and TNF-. This in turn activates vagal responses have been discussed (Maier, S. F.
afferents from the liver. IL-1 and TNF- and Watkins, L. R., 1998; Watkins, L. R. and
injected intraperitoneally themselves generate Maier, S. F., 1999, Goehler, L. E. et al., 2000;
behavioral hyperalgesia that is abolished by Watkins, L. R. and Maier, S. F., 2000). The phy-
vagotomy (Watkins, L. R. et al., 1994a; 1994b; siology of the vagal afferents involved in the
1995a; 1995b; 1995c; 1995d). The proinflamma- communication between the immune system,
tory cytokines either activate the vagal afferents which operates as a diffuse sensory system to
directly or bind to glomus cells in the abdominal detect chemical constituents associated with dan-
paraganglia that are innervated by vagal afferents; gerous microorganisms and their toxins, and the
the activated vagal afferents signal the peripheral brain has to be worked out.
events to the brain, leading to hyperalgesia (as
shown by an enhanced thermal nociceptive tail-
flick reflex). Acknowledgments
5. Pain behavior mediated by vagal afferents, acti-
vated by intraperitoneal injection of LPS or of Supported by the Deutsche Forschungsgemeinschaft
proinflammatory cytokines, is part of a general and the German Ministry of Research and Education
sickness behavior characterized by various protec- (German Research Network on Neuropathic Pain).
tive illness responses (e.g., immobility, decreased
social interaction, decrease in food intake, forma-
tion of taste aversion to novel foods, decrease of
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 21 Sex, Gender, and Pain
R B Fillingim, University of Florida College of Dentistry, Community Dentistry and Behavioral Science
Gainesville, FL, USA
2009 Elsevier Inc. All rights reserved.

21.1 Sex Differences in Pain Responses 253

21.2 Sex Differences in Responses to Pain Treatment 253
21.3 Mechanisms Underlying Sex Differences in Pain 254
21.4 Conclusions and Future Directions 255
References 255

Glossary
biopsychosocial model A model of pain that posits social institutions on the basis of that individuals
that the experience of and response to pain results gender presentation.
from complex, dynamic interactions among biologi- sex The classification of living things, generally as
cal, psychological, and sociocultural variables. male or female, according to their reproductive
gender A persons self-representation as male or organs and functions assigned by the chromoso-
female, or how that person is responded to by mal complement.

21.1 Sex Differences in Pain (Boccardi, L. and Verde, M., 2003). In addition,
Responses women display greater perceptual responses to labora-
tory-induced pain relative to men, although the
Considerable evidence suggests that females and magnitude of these differences varies across studies,
males differ in their experience of both clinical and perhaps due to methodological differences (e.g., pain
experimentally induced pain (for reviews, see assay, stimulus characteristics, sample selection;
Fillingim, R. B. and Maixner, W., 1995; Unruh, A. M., Fillingim, R. B. and Maixner, W., 1995; Riley, J. L.
1996; Berkley, K. J., 1997; LeResche, L., 1999; et al., 1998). Interestingly, consistent with the human
Fillingim, R. B., 2003). For example, several chronic literature, greater behavioral responses to noxious
pain disorders are more prevalent among women, and stimuli have been reported among female compared
in population surveys women report more pain than to male rodents (e.g., Barrett, A. C. et al., 2002b; 2003;
men (Fillingim, R. B. and Maixner, W., 1995; Unruh, A. Terner, J. M. et al., 2003; also for reviews, see Bodnar, R.
M., 1996; Berkley, K. J., 1997). Moreover, some evi- J. et al., 1988; Berkley, K. J., 1997), with some exceptions
dence indicates that clinical pain may be more severe (Mogil, J. S. et al., 1993; Kayser, V. et al., 1996). In
for women than men. For example, women have general, these findings suggest that females exhibit
reported greater postsurgical pain than men in some greater perceptual responses to pain than males; how-
(Savedra, M. C. et al., 1993; Averbuch, M. and Katzper, ever, the mechanisms underlying these sex differences
M., 2000; Coulthard, P. et al., 2000; Taenzer, A. H. et al., have not been determined.
2000; Morin, C. et al., 2000; Averbuch, M. and
Katzper, M., 2001; Kalkman, C. J. et al., 2003) but not
all (e.g., Gordon, N. C. et al., 1995; Gear, R. W. et al., 21.2 Sex Differences in Responses to
1996a; Chia, Y. Y. et al., 2002) studies. Women have Pain Treatment
also reported higher pain ratings in acute cancer-
related pain (Cepeda, M. S. et al., 2003), procedural In addition to basal pain sensitivity, sex-related influ-
pain such as colonoscopy (Froehlich, F. et al., 1997), ences on responses to pharmacologic and non-
and conditions presented in emergency rooms pharmacologic pain treatments have received

253
254 Sex, Gender, and Pain

increasing attention in recent years. The rodent litera- sex differences in pain responses could refer to the
ture fairly consistently demonstrates greater analgesic same fundamental processes described at different
responses to opioid agonists among males versus female levels of analysis.
animals (Bodnar, R. J. et al., 1988; Kepler, K. L. et al., Several biological variables have been implicated
1989; 1991; Kiefel, J. M. and Bodnar. R. J., 1992; Islam, in sex differences in both clinical and experimental
A. K. et al., 1993; Cicero, T. J. et al., 1996a; 1997; Kest, B. pain responses. Abundant preclinical and clinical evi-
et al., 2000; Craft, R. M., 2003a; 2003b). However, the dence suggests that gonadal hormones are important,
human literature regarding sex differences in analgesic because hormonal influences on basal pain sensitivity
responses has shown mixed results. Some clinical stu- as well as responses to analgesic compounds have been
dies demonstrate greater analgesic responses among well documented (Riley, J. L. III. et al., 1999; Fillingim,
women, others show no sex differences, and others R. B. and Ness, T. J., 2000; Bodnar. R. J. et al., 2002).
report greater analgesia among men (Fillingim, R. B. Also, endogenous pain inhibitory systems may func-
and Gear, R. W., 2004). Among the most consistent tion differently in females and males, including the
findings is a series of studies indicating more robust endogenous opioid system. Zubieta and colleagues
analgesic responses -opioid-acting drugs after oral have reported sex differences in both basal and pain-
surgery (Gear, R. W. et al., 1996a; 1996b; 1999; 2003). evoked -opioid receptor binding in certain brain
Laboratory studies in humans have shown mixed regions (Zubieta, J. K. et al., 1999; 2002). Genetic
results, as well (Sarton, E. et al., 2000; Fillingim, R. B. factors may also contribute to sex differences in pain.
et al., 2004; Romberg, R. et al., 2004; Olofsen, E. et al., In rodents, sex differences in nociceptive and antino-
2005; Fillingim, R. B. et al., 2005c). Sex differences in ciceptive responses are often strain-dependent (Mogil,
response to nonpharmacologic interventions for pain J. S. et al., 2000; Barrett, A. C. et al., 2002a; Terner, J. M.
has received less empirical attention, though a few et al., 2003), and pain-related quantitative trait loci
laboratory (Keogh, E. et al., 2000; Sternberg, W. F. (QTLs) are often sex-specific (Mogil, J. S. et al., 1997;
et al., 2001) and clinical (Hansen, F. R. et al., 1993; Mogil, J. S., 2000). Recent findings provide corrobor-
Krogstad, B. S. et al., 1996; Jensen, I. B. et al., 2001) ating results in humans (Kim, H. et al., 2004). Also,
studies suggest that sex differences in treatment Mogil J. S. et al. (2003) demonstrated in both mice and
responses may emerge. It is possible that the consistent humans that the melanocortin-1-receptor gene
sex differences in analgesic responses are specific to the (MC1R) was associated with analgesic responses to -
oral surgery pain model, perhaps due to its inflamma- opioids among females but not males. Also, the A118G
tory component or based on interactions with other single nucleotide polymorphism of the -opioid
medications administered perioperatively. The clinical receptor gene (OPRM1) was significantly associated
relevance of these results beyond this acute pain model with pressure pain thresholds, but this association
remains unknown, and this is an area ripe for future was only significant among males (Fillingim, R. B.
investigation. et al., 2005b). Thus, several biological factors may
contribute to sex differences in pain.
Numerous psychosocial variables also contribute
21.3 Mechanisms Underlying Sex to sex differences in pain responses. Anxiety and
Differences in Pain other negative emotions have been more strongly
related with both clinical and experimental pain
It is important to recognize that sex differences in responses among men than women (Fillingim, R. B.
pain are inevitably determined by interactions et al., 1996; Edwards, R. R. et al., 2000; Riley, J. L., III
among multiple biopsychosocial processes. While et al., 2001; Jones, A. and Zachariae, R., 2002;
these mechanisms are typically described as either Robinson, M. E. et al., 2005) Moreover, positive affect
psychosocial or biological, this dualistic conceptuali- predicted lower experimental pain sensitivity and
zation is artificial and potentially counterproductive. fewer side effects in responses to pentazocine (a
For instance, sex differences in expression of pain are mixed action opioid) among men, but not women
often attributed to the effects of stereotypic sex roles, (Fillingim, R. B. et al., 2005a). Additional psychosocial
typically considered a psychosocial issue. However, factors that may contribute to sex differences in pain
neurobiological and hormonal correlates of mascu- include cognitive and behavioral pain coping strate-
line versus feminine sex roles may well contribute to gies (Affleck, G. et al., 1999; Unruh, A. M. et al., 1999;
differences in nociceptive processing. Thus, various Keefe, F. J. et al., 2000; Mercado, A. C. et al., 2000;
psychosocial and biological mechanisms underlying Osman, A. et al., 2000), gender roles (Otto, M. W. and
 Sex, Gender, and Pain 255

Dougher, M. J., 1985; Myers, C. D. et al., 2001; Barrett, A., Smith, E., and Picker, M. 2002b. Sex-related
differences in mechanical nociception and antinociception
Robinson, M. E. et al., 2003), and family history of produced by mu- and kappa-opioid receptor agonists in
pain (Edwards, P. W. et al., 1985; Fillingim, R. B. et al., rats. Eur. J. Pharmacol. 452, 163.
2000). Barrett, A. C., Smith, E. S., and Picker, M. J. 2003. Capsaicin-
induced hyperalgesia and -opioid-induced
antihyperalgesia in male and female Fischer 344 rats.
J. Pharmacol. Exp. Ther. 307, 237245.
Berkley, K. J. 1997. Sex Differences in Pain. Behav. Brain Sci.
21.4 Conclusions and Future 20, 371380.
Directions Boccardi, L. and Verde, M. 2003. Gender differences in the
clinical presentation to the emergency department for chest
The evidence reviewed indicates consistent sex dif- pain. Ital. Heart J. 4, 371373.
Bodnar, R. J., Commons, K., and Pfaff, D. W. 2002. Central
ferences in pain-related responses, to which multiple Neural States Relating Sex and Pain. Johns Hopkins
biopsychosocial factors contribute. The literature University Press.
Bodnar, R. J., Romero, M. T., and Kramer, E. 1988. Organismic
focuses more heavily on quantitative sex differences
variables and pain inhibition: roles of gender and aging. Brain
(i.e., does the magnitude of pain or analgesia differ in Res. Bull. 21, 947953.
women vs. men?); however, of equal if not greater Cepeda, M. S., Africano, J. M., Polo, R., Alcala, R., and
importance are qualitative sex differences (i.e., are Carr, D. B. 2003. Agreement between percentage pain
reductions calculated from numeric rating scores of pain
the mediators of pain and analgesia different in intensity and those reported by patients with acute or cancer
women vs. men?). Some examples are provided pain. Pain 106, 439442.
above, including findings demonstrating sex-specific Chia, Y. Y., Chow, L. H., Hung, C. C., Liu, K., Ger, L. P., and
Wang, P. N. 2002. Gender and pain upon movement are
genetic and psychosocial associations with pain. The associated with the requirements for postoperative patient-
clearest challenge for the future is to determine the controlled iv analgesia: a prospective survey of 2,298
Chinese patients. Can. J. Anaesth. 49, 249255.
practical significance of these sex differences and to
Cicero, T. J., Nock, B., and Meyer, E. R. 1996. Gender-related
integrate this information into clinical practice. It differences in the antinociceptive properties of morphine.
seems plausible that expanding research into qualita- J. Pharmacol. Exp. Ther. 279, 767773.
tive sex differences may be more informative in this Cicero, T. J., Nock, B., and Meyer, E. R. 1997. Sex-related
differences in morphines antinociceptive activity:
regard, because such differences imply that some relationship to serum and brain morphine concentrations.
components of the pain processing system may be J. Pharmacol. Exp. Ther. 282, 939944.
fundamentally different in women versus men, which Coulthard, P., Pleuvry, B. J., Dobson, M., and Price, M. 2000.
Behavioural measurement of postoperative pain after oral
calls for sex-specific treatment approaches. It is surgery. Br. J. Oral Maxillofac. Surg. 38, 127131.
hoped that continued research into sex, gender and Craft, R. M. 2003a. Sex differences in drug- and non-drug-
induced analgesia. Life Sci. 72, 26752688.
pain will ultimately enhance pain management for
Craft, R. M. 2003b. Sex differences in opioid analgesia: from
both women and men. mouse to man. Clin. J. Pain 19, 175186.
Edwards, R. R., Augustson, E., and Fillingim, R. B. 2000. Sex-
specific effects of pain-related anxiety on adjustment to
chronic pain. Clin. J. Pain 16, 4653.
Acknowledgment Edwards, P. W., Zeichner, A., Kuczmierczyk, A. R., and
Boczkowski, J. 1985. Familial pain models: the relationship
between family history of pain and current pain experience.
This work was supported by NIH grant NS41670. Pain 21, 379384.
Fillingim, R. B. 2003. Sex-related influences on pain: a review of
mechanisms and clinical implications. Rehabil. Psychol.
48, 165174.
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 22 Neurotrophins and Pain
Lorne M Mendell, State University of New York, Stony Brook, NY, USA
2009 Elsevier Inc. All rights reserved.

22.1 The Neurotrophin Family and Its Receptors 260

22.2 Neurotrophins and Development of Nociceptors 260
22.3 Nerve Growth Factor Affects Electrophysiological Properties of Nociceptors that
Express TrkA 262
22.4 Exogenous Nerve Growth Factor Elicits Hyperalgesia 262
22.4.1 Chronic Changes 262
22.4.2 Acute Changes 262
22.4.3 Mechanisms of Nerve Growth Factor Action in the Skin 263
22.5 Acute Sensitization by Nerve Growth Factor of the Response of Dissociated
Nociceptors 264
22.6 TrkA Signaling Pathways Responsible for Acute Effects of Nerve Growth Factor 265
22.7 p75 Receptor Influence on Sensory Neuron Function 266
22.8 Effects of Nerve Growth Factor on Expression of Channels and Receptors in
Nociceptors 267
22.8.1 TRPV1 Receptors 267
22.8.2 P2X3 Receptors 268
22.8.3 ASIC Channels 268
22.8.4 BK Receptors 268
22.8.5 Na Channels 268
22.8.6 PGE2 Receptors 269
22.9 Neurotrophins and Sensitization of Tissues Other than Skin 269
22.9.1 Visceral Structures 269
22.9.2 Muscle Afferents 269
22.9.3 Joint Receptors 269
22.10 Nerve Growth Factor, Enhances Peptide Expression in the Dorsal Root Ganglion,
and Central Sensitization 270
22.11 Role of Brain-Derived Neurotrophic Factor, Neurotrophin-4, and Neurotrophin-3 in
Peripheral Sensitization 270
22.12 Neurotrophins and Neuropathic Pain 271
22.13 Neurotrophins and the Clinic 272
22.14 Discussion, Conclusions, and Open Questions 272
References 273

Glossary
immunoadhesin A chimeric molecule consisting of skin nerve preparation In vitro preparation in
a IgG backbone (Fc region) linked to the binding site which the skin is removed with its nerve supply
of a receptor that binds to a molecule to be seques- (e.g., ankle skin and saphenous nerve) and placed
tered, e.g., trkA-IgG binds nerve growth factor (NGF). in a chamber. The nerve is led into a second
keratinocyte Immune-competent cell in the chamber separated from the skin chamber so that
epidermis. the skin can be maintained in oxygenated Ringer,
mast cell Immune-competent cell that stores whereas the nerve can be placed in oil for extra-
inflammatory mediators. cellular recording.
neutrophil A type of white blood cell that phago- tachyphylaxis A process whereby the response to
cytoses microorganisms (i.e., due to infection) and an agent applied repeatedly becomes smaller and
breaks them down. smaller.

259
260 Neurotrophins and Pain

22.1 The Neurotrophin Family and Its 22.2 Neurotrophins and

Receptors Development of Nociceptors

Neurotrophin molecules are 13.5-kDa proteins that The early history of the discovery of NGF has been
come in four major forms that show a substantial over- extensively reviewed (Cowan, W. M., 2001). It began
lap in their primary structure. These are known as with the observation that extracts from mouse salivary
nerve growth factor (NGF), brain-derived neuro- glands induce proliferation of processes of explanted
trophic factor (BDNF), neurotrophin-3 (NT-3), and dorsal root ganglion (DRG) and sympathetic gang-
neurotrophin-4 (also known neurotrophin-5 or neuro- lion cells. Later, biochemical studies revealed that the
trophin-4/5, NT-4/5). The molecular biology active factor is a peptide that became known as nerve
underlying the synthesis of these molecules has been growth factor. NGF was subsequently identified as
reviewed elsewhere (Teng, K. K. and Hempstead, B. L., an important factor assuring survival of developing
2004). Neurotrophins bind to two classes of receptors: sensory neurons since its absence induced by treating
the tropomyosin-related kinase (trk) receptors, trkA, the animal with an NGF antibody resulted in the
trkB, and trkC, as well as the p75 receptor that is a survival of fewer cells in the DRG as well as in
member of the tumor necrosis factor (TNF) super- sympathetic ganglia. This led to the neurotrophic
family. Although these two receptor types are hypothesis (Davies, A. M., 1996), which states that a
often considered to function together as a complex soluble trophic factor is present in limiting amounts
(Chao, M. V. and Hempstead, B. L., 1995; He, X. L. in the target tissue of a population of neurons, which
and Garcia, K. C., 2004), there is also evidence that require it in order to survive. Nerve terminals that
they can function independently (see Section 22.7). are successful in internalizing this factor and trans-
The trk receptors are known as the high-affinity porting it to the cell body for its metabolic use
neurotrophin receptors and selectively bind the differ- survive; the unsuccessful ones die. More detailed
ent neurotrophin molecules. NGF binds selectively to studies of the DRG revealed that selective neutrali-
trkA, BDNF and NT-4/5 to trkB, and NT-3 to trkC zation of NGF via its antibody results in depletion of
(Huang, E. J. and Reichardt, L. F., 2001). Some cross- a limited population of DRG cells, specifically those
binding with lower affinity also occurs (Barbacid, M., that express peptides such as substance P and calci-
1994). The p75 receptor is generally known as the tonin gene-related peptide (CGRP) and project into
low-affinity receptor, although it actually binds neu- superficial laminae of the spinal cord. i.e., those that
rotrophins with high affinity but with rapid kinetics. are identified with nociception (reviewed in
It binds all known neurotrophins with similar affinity. Mendell, L. M., 1995). Animals treated in this way
Thus, the selective expression of trk receptors in exhibit a higher threshold for painful stimuli.
different cell types (MCMahon, S. B. et al., 1994) is Genetically modified animals have proven very useful
a major factor underlying the selective effects of in demonstrating these relationships with those under-
neurotrophins in the nervous system. or overexpressing NGF in the skin being hypo- or
Ligands binding to the extracellular component of hyperalgesic, respectively (Davis, B. M. et al., 1993),
trk receptors induce autophosphorylation of the trk and those underexpressing the NGF receptor trkA
molecule, which leads to downstream effects via multi- being hypoalgesic (Smeyne, R. J. et al., 1994).
ple signaling pathways (Kaplan, D. R. and Miller, F. D., In all these manipulations, NGF was considered
2000). For example, phospholipase C (PLC) is phos- to affect the number of nociceptors, permitting more
phorylated subsequent to its recruitment to the cell or fewer to survive. However, it has become apparent
membrane as a consequence of phosphorylation of that NGF influences the properties of nociceptors in
tyrosine 785 on trkA (Obermeier, A. et al., 1993). This the immediate postnatal period, beyond the time it
is very likely an important mechanism in mediating can affect DRG cell number. Ritter A. M. et al. (1991;
neurotrophin-induced effects (see below). However, see also Lewin, G. R. et al., 1992a) demonstrated that a
other intracellular signaling pathways are also activated class of nociceptors known as A high-threshold
by neurotrophin binding to trk receptors including the mechanoreceptors (HTMRs) is severely depleted in
RasRafMekErk pathway and the PI-3K pathway rats treated postnatally with an antibody to NGF
(Kaplan, D. R. and Cooper, E., 2001). Some of these (anti-NGF). The proportion of A low-threshold
have also been implicated in mediating the functional mechanoreceptors known as down hair receptors
effects of neurotrophins (see Section 22.6). (D hairs) is elevated by a similar proportion.
 Neurotrophins and Pain 261

Significantly, the proportion of a third population of 1997). Cells undergoing this conversion do not express
afferent fibers also conducting in the A range and p75 even in neonates, i.e., they are p75/trkA as
innervating subcutaneous tissues is unchanged. Thus, distinct from cells that are p75/trkA (Figure 1).
HTMRs are not selectively depleted since if this Receptors for GDNF consist of an intracellular sig-
occurs, the proportion of all other fiber types would naling receptor domain RET, which is a tyrosine
be expected to increase. Furthermore, when cell kinase receptor. There are four different ligand-
counts were undertaken in the DRG, no change in binding domains, GFR1GFR4, that bind different
cell number was noted. Consistent with these find- members of the GDNF family. GDNF itself binds
ings, the magnitude of the vasodilator response to GFR1. Other agonists are neurturin (NTN)
elicited by antidromic activation of cutaneous nerves binding to GFR2, artemin binding to GFR3
is reduced in proportion to the depletion of A and persepherin that binds to GFR4 (Saarma, M.
HTMRs (Lewin, G. R. et al., 1992b).
These findings suggest that the postnatal devel-
opment of nociceptors depends on the availability of
NGF and that in its absence sensory neurons nor-
mally destined to be HTMRs become D hairs. This
has been referred to as a phenotypic shift (Lewin, G.
R. et al., 1992a). Cell number in the DRG declines if
the antibody is delivered between P0 and P2,
whereas the phenotypic shift from HTMR to D
hair occurs after anti-NGF treatment between P4
and P11. Attention was drawn to evidence that sen-
sory axons normally first innervate the epidermis,
with many terminals later withdrawing to the dermal
layer as hair follicles and other structures develop
(Reynolds, M. L. et al., 1991). NGF is expressed by a
limited population of cells in the epidermis, for
example in basal keratinocytes (Di Marco, E. et al.,
1991), and it was hypothesized that the availability of
NGF there would cause trkA-expressing neurons to
differentiate into nociceptors with epidermal term-
inals. In the absence of NGF due to anti-NGF
treatment, these terminals would recede into the der-
mal layers of the skin, as occurred with most non-
trkA-expressing neurons (reviewed in Lewin, G. R.
and Mendell L. M., 1993; Mendell, L. M. et al., 1999).
Exogenous NGF delivered postnatally does not
elevate the relative proportion of HTMRs (Lewin,
G. R. et al., 1993). However, enhancing NGF levels in
the skin beginning during development (E11) in
transgenic mice via the keratin promoter increases
the number of sensory neurons well above normal
levels (Albers, K. M. et al., 1994). Electrophysiological
studies have confirmed that nociceptors are selec-
Figure 1 Dorsal root ganglion (DRG) tissue section from
tively rescued by the availability of the additional 10-day-old rat immunostained for trkA (red), TRPV1 (green),
NGF (Stucky, C. L. et al., 1999). and p75 (blue). Most small cells are at least weakly positive
The DRG cell population undergoes substantial for trkA, TRPV1, and p75 (small down and horizontal
changes in the expression of trkA postnatally. A sub- arrows). In one case (large up arrow), the cell was trkA/
TRPV1/p75; this cell was likely to lose its trkA expression
stantial number lose trkA expression over the first 3
and gain receptors for GDNF. The cell denoted by an
postnatal weeks, and there is a parallel increase in the arrowhead is negative for all three markers used. Scale bar
number of cells exhibiting receptors for glial-derived (lower left) is 20 mm. Data from Petruska J. C. and Mendell
neurotrophic factor (GDNF) (Molliver, D. C. et al., L. M. (unpublished).
262 Neurotrophins and Pain

and Sariola, H., 1999). In rats, the population Individual A HTMRs in rats treated systemically
expressing cRet also express the lectin IB4 (in adults over the first 2 postnatal weeks with NGF exhibit a
trkA cells are IB4). In mice, this separation is not significantly diminished mechanical threshold (i.e.,
as distinct. In rats, IB4/cret/trkA cells have are more sensitive) up to 9 weeks of age after which
been shown to project more deeply into lamina II it returns to control values (Lewin, G. R. et al., 1993).
(lamina IIi) and are considered to be sensitive to Neonatal treatment with anti-NGF elevates mechan-
nonnoxious stimulation. These neurons, unlike ical threshold (Lewin, G. R. et al., 1992). Treatment of
trkA/IB4 neurons, do not contain CGRP. older rats with NGF has no measurable effect on
GDNF has actions on GFR/Ret-expressing noci- mechanical threshold. A similar study on C-fibers
ceptors that only partially overlap those of NGF (Lewin, G. R. and Mendell, L. M., 1994) has revealed
on trkA-expressing nociceptors (see Section 22.8). that treatment with NGF increases the fraction sen-
sitive to noxious heat, whereas anti-NGF reduces it.
Unlike the changes in mechanical sensitivity in A-
fibers, these changes are apparently permanent. The
22.3 Nerve Growth Factor Affects
mechanical sensitivity of these C-fiber units is
Electrophysiological Properties of
unchanged, although a population of mechanically
Nociceptors that Express TrkA
sensitive units with an unusually low threshold is
observed in anti-NGF-treated preparations. Redu-
The finding that treatment with anti-NGF affects
cing NGF levels in the skin in adult rats using the
properties of nociceptors suggested that trkA is
immunoadhesin trkA-IgG 1012 days before record-
expressed selectively in nociceptors. This has been
ing (Bennett, D. L. et al., 1998a) also causes a
confirmed in a number of electrophysiological stu-
substantial decline in the response of individual
dies (Ritter, A. M. and Mendell, L. M., 1992; Djouhri,
units to noxious heat with no change in the response
L. et al., 2001) where application of NGF or anti-
to mechanical stimulation. The number of units sen-
NGF was found to affect properties of the action
sitive to bradykinin also decreases after trkA-IgG
potential in nociceptors only, regardless of axon size
treatment, suggesting an important contribution of
(A, A, or C). Interestingly, both these studies found
NGF to bradykinin sensitivity of nociceptors (see
effects on action potential fall time although the
Section 22.8.4).
effects differed, probably because of the very differ-
ent conditions of the experiments. Both studies also
revealed that NGF does not affect the duration of 22.4.2 Acute Changes
after hyperpolarization. The selective expression of
Examination of the acute effects of a single adminis-
trkA in electrophysiologically characterized nocicep-
tration of NGF (i.p.) in adult rats revealed that
tors has recently been confirmed directly by Fang X.
thermal hyperalgesia becomes significant virtually
et al. (2005).
immediately, unlike mechanical hyperalgesia that
develops gradually and reaches significance only
after a few hours. Hyperalgesia elicited by a single
22.4 Exogenous Nerve Growth Factor i.p. injection of NGF lasts several days. Localized
Elicits Hyperalgesia small injections of NGF into the skin elicit thermal
and mechanical hyperalgesia of much shorter dura-
22.4.1 Chronic Changes
tion (Woolf, C. J. et al., 1994; Shu, X. et al., 1999).
Neonatal rats are more sensitive to being handled These local injections indicate the likelihood that
after an initial injection of NGF (Lewin, G. R. et al., NGF can act directly on cells located in the skin.
1993). It was surmised that changes had occurred in NGF itself does not evoke any behavioral response; it
the nociceptive system. Behavioral studies in merely sensitizes the response to subsequent noci-
5-week-old rats treated with NGF every other day ceptive stimuli.
from birth revealed both mechanical and thermal Although administration of NGF to the skin eli-
hyperalgesia (Lewin, G. R. et al., 1993). Additional cits hyperalgesia, it does not cause inflammation. On
studies with serial observations on rats treated either the other hand, treatments or injuries that elicit
as neonates (02 weeks old) or as juveniles (25 inflammation, such as complete Freunds adjuvant
weeks old) demonstrated that the sensitizing effect (CFA), enhance NGF levels in the skin (Weskamp,
of NGF lasts several weeks after the end of treatment. G. and Otten, U., 1987; Aloe, L. et al., 1992).
 Neurotrophins and Pain 263

Inflammatory cytokines, specifically TNF and IL- tested the response to noxious heat and mechanical
1, known to be expressed during inflammation, stimulation under control conditions and 10 min later
enhance NGF levels when delivered separately into with NGF applied to the corium surface of the unit
the skin (Woolf, C. J. et al., 1997). These studies have or under control conditions (no NGF). They
suggested a temporal sequence: TNF ! IL- observed a significant decline in threshold to noxious
1 ! NGF (Figure 2). Cells responsible for producing heat, but no acute change in threshold to mechanical
NGF include keratinocytes (Di Marco, E. et al., 1991) stimulation. The sensitivity to NGF was abolished if
and mast cells (Leon, A. et al., 1994). mast cells were degranulated over a period of several
The requirement for NGF as an essential inter- days prior to treatment (see Section 22.4.3).
mediate in causing inflammatory hyperalgesia has
been demonstrated in experiments where its levels
22.4.3 Mechanisms of Nerve Growth
are experimentally diminished using an antibody to
Factor Action in the Skin
NGF (Lewin, G. R. et al., 1994; Woolf, C. J. et al.,
1994) or an immunoadhesin such as trkA-IgG A number of pharmacological approaches have been
(McMahon, S. B. et al., 1995; Koltzenburg, M. et al., undertaken to explore the identity of cells and
1999), both of which deplete endogenous NGF. mechanisms underlying sensitization of nociceptive
These agents prevent the development of hyperalge- responses by NGF. A contribution by the sympa-
sia to mechanical or thermal stimuli in response to thetic nervous system to the sensitizing effect of
inflammatory agents such as CFA. In the case of NGF has been identified since guanethidine treat-
thermal hyperalgesia, these agents can also substan- ment substantially reduces NGFs acute hyperalgesic
tially reduce ongoing inflammatory pain within an effect (Andreev, N. Yu. et al., 1995) by delaying its
hour. However, mechanical hyperalgesia is reduced onset (Woolf, C. J. et al., 1996).
only after 2448 h (Woolf, C. J. et al., 1994; 1996). Mast cells also participate in NGF-induced sensi-
This difference between thermal and mechanical tization, particularly of noxious thermal stimuli. Mast
hyperalgesia is consistent with the conclusion, based cells express trkA receptors (Horigome, K. et al.,
on the latency of onset after exposure to NGF, that 1993), and NGF causes them to degranulate their
thermal hyperalgesia is due to a peripheral action, contents consisting of serotonin, histamine, and
whereas the mechanical hyperalgesia is indirect, NGF itself (reviewed in Mendell, L. M. et al., 1999).
probably involving transcriptional changes at the Degranulation of mast cells in advance of NGF
level of the DRG (but see Section 22.7). administration delays the development of thermal
Rueff A. and Mendell L. M. (1996) recorded the hyperalgesia for about 3 h but does not prevent the
activity of single nociceptor axons in an excised skin later phase of thermal hyperalgesia, nor does it abol-
nerve preparation with the corium side up where ish mechanical hyperalgesia (Lewin, G. R. et al., 1994;
NGF could be applied directly to the electrophysio- see also Woolf, C. J. et al., 1996). Behavioral experi-
logically identified receptive field of the fiber. They ments using pharmacological antagonists indicate that
serotonin is an important contributor to mast cell-
mediated sensitization initiated by NGF (Lewin, G.
Injury R. et al., 1994). Under some special conditions, NGF
can elicit sensitization in the absence of mast cell
TNF
Neutrophil
activity (Amann, R. et al., 1996a), suggesting the exis-
IL-1
accumulation tence of other pathways (Figure 2), or perhaps that
Leukotriene B4
8(R),15(S)- NGF acts directly on trkA receptors expressed by
diHETE Nociceptor other cells, for example, the nociceptors themselves
NGF terminal
(Averill, S. et al., 1995; Galoyan, S. M. et al., 2003; see
MC NGF
Section 22.5).
Another action of NGF is enhancement of the
Figure 2 Major mechanisms underlying acute action of level of leukotrienes, specifically leukotriene B4
nerve growth factor (NGF) on sensory neurons. Exogenous (LTB4), via upregulation of the enzyme 5-lipoxygen-
NGF sensitizes sensory neurons via neutrophils, mast cells
ase (Amann, R. et al., 1996b; Bennett, G. et al., 1998b).
(MCs), and perhaps directly. Other possible mechanisms
are mentioned in the text. NGF is released endogenously Leukotrienes are known to sensitize nociceptors
after injury via the action of cytokines TNF and IL-1 (Martin, H. A. et al., 1988). This occurs as a result of
(rectangle). neutrophil accumulation and their subsequent
264 Neurotrophins and Pain

release of 8(R), 15(S)-diHETE. If neutrophils are 22.5 Acute Sensitization by Nerve

depleted pharmacologically, NGF-induced hyperal- Growth Factor of the Response of
gesia is abolished (Bennett, G. et al., 1998b). Dissociated Nociceptors
Keratinocytes may also play an important role in
NGF-induced hyperalgesia. These cells are known to Shu X. Q. and Mendell L. M. (1999a; 1999b) have
express trkA receptors and to contain NGF (Terenghi, investigated the effect of NGF on the response of
G. et al., 1997) and thus may act in a manner similar to small (<30 mm) dissociated DRG cells to capsaicin,
that described for mast cells (see above). They are also known to elicit the sensation of noxious heat
known to release ATP (Koizumi, S. et al., 2004) that (Lamotte, R. H. et al., 1992) via the VR1 (now referred
activates nociceptor terminals (Cook, S. P. and to as TRPV1) receptor (Caterina, M. J. et al., 1997).
McCleskey, E. W., 2002). Enhancement of NGF NGF sensitizes the response to capsaicin (Shu, X. and
expression in these cells in genetically modified mice Mendell, L. M., 1999b), measured as an increased
enhances nociceptive behavior (Davis, B. M. et al., 1993). inward current in the presence of NGF, in a dose-
The multiplicity of cell types that express trkA dependent manner over the range of 2100 ng ml1.
receptors and the number of substances capable of Its effect is blocked by the protein kinase inhibitor
sensitizing nociceptors released by these cells in K252a known to antagonize the response of trkA,
response to NGF indicate the complexity of periph- although not in a highly selective manner (Knusel,
eral sensitization by neurotrophins. They also B. and Hefti, F., 1992). Not all cells tested are sensi-
indicate the difficulty of using behavioral methods tized by NGF, indicating that not all these small cells
and preparations such as the skin nerve preparation, express the trkA receptor, as predicted from immu-
where all these cells remain in close contiguity, to nohistochemical studies (Averill, S. et al., 1995;
arrive at cellular mechanisms of sensitization. Thus, Bennett, D. L. et al., 1996a; Figures 1 and 3).
many studies have made use of dissociated DRG cells The duration of NGF action is difficult to deter-
to examine the effects of NGF directly on trkA- mine because the test input, capsaicin, is associated
expressing neurons. Such studies have the obvious with long-lasting tachyphylaxis. To circumvent this
advantage of providing a relatively unobstructed problem, Shu X. and Mendell L. M. (2001) have
view of cellular mechanisms but at the expense of examined the duration of NGF action under condi-
the intercellular interactions that take place in vivo. tions of reduced Ca2 that eliminates tachyphylaxis

Time (s) Time (s)

0 20 40 60 0 20 40 60 0 20 40 60 0 20 40 60
0 0
Temp. (C) response (pA)

100 200
200 400
300 600
50 5 min 50 5 min
45 45
40 40
35 35

Figure 3 The response of two dissociated dorsal root ganglion (DRG) cells to administration of nerve growth factor (NGF)
and demonstration that sensitization of the response requires expression of trkA. Cell on left was tested with two heat pulses
(bottom traces). The second response 5 min later exhibited tachyphylaxis despite the presence of NGF in the bathing solution
for 5 min between heat pulses. The cell (diameter 26 mm; thick arrow) was recovered and immunostained for TRPV1 and trkA.
It was classified as TRPV1/trkA. Another cell in the same field denoted by white arrowhead is trkA/TRPV1 and a third cell
denoted by white-bordered arrowhead is classified trkA/ TRPV1. Cell on right showed sensitization of the response to heat
after 5 min exposure to NGF. The cell (diameter 25 mm; arrow) was recovered, immunostained, and classified trkA/TRPV1.
Cell denoted by arrowhead was trkA/TRPV1. Adapted from Galoyan et al. (2003) with permission.
 Neurotrophins and Pain 265

(Koplas, P. A. et al., 1997). They found that a 10-min correspondence between the expression of trkA and
exposure to NGF sensitizes a neuron to repeated the ability of NGF to sensitize the TRPV1 response,
puffs of capsaicin for as long as 70 min, suggesting i.e., trkA is expressed in cells that fail to be sensitized
that the effect of NGF involves more than a direct by NGF. However, cells in neonates failing to be
action on the conductance of a membrane channel or sensitized by NGF can be sensitized by bradykinin,
the sensitivity of a receptor (see Section 22.6). i.e., the failure for NGF to sensitize the cells is not a
Galoyan S. M. et al. (2003) have carried out similar generalized failure for sensitization to occur. It is not
experiments in small dissociated DRG cells from presently known what factor develops during the
adult rats using noxious heat instead of capsaicin as neonatal period that permits NGF to sensitize the
the test stimulus (Figure 3). One advantage of the heat-evoked response.
noxious heat stimulus over capsaicin is the ease with NGF-induced sensitization develops gradually
which threshold can be established. These cells from P4 to P10 (Zhu, W. et al., 2004). This is the
respond an inward current whose threshold is 43 time at which trkA-expressing cells lose their require-
44  C which is characteristic of the TRPV1 receptor ment for NGF in order to survive (Lewin, G. R.
expressed in hetrologous cells (caterina, M. J., et al., et al., 1992). It has been hypothesized that the
1997). NGF-induced sensitization of the response to inability for NGF to sensitize cells in neonates may
noxious heat has properties that are similar to sensi- be a protective mechanism to prevent undue noci-
tization of the response to capsaicin. NGF does not ceptive input due to sensitization at a time that NGF
alter temperature threshold; its major effect is to is required to assure survival of nociceptors (Zhu, W.
enhance the magnitude of the heat evoked current. et al., 2004).
Recent studies suggest that the acute increase of GDNF, like NGF, acutely elevates cytosolic
TRPV1 current elicited by exposure to NGF is due Ca2 in DRG cells (Lamb, K. and Bielefeldt, K.,
to increased trafficking and subsequent insertion of 2003). Recent experiments suggest that exposure of
the additional TRPV1 receptors into the plasma dissociated sensory neurons to GDNF, neurturin or
membrane rather than an increase in channel unitary artermin for several minutes rapidly enhances the
conductance (Zhang, X., et al., 2005; Stein, A. T., et al., sensitivity of TRPV1 receptors to capsaicin in iso-
2006). These cells respond with a threshold of about lated DRG cells (Malin, S. A., et al., 2006). Artemin
4344  C, which is characteristic of the TRPV1 mRNA, but not neurturin or GDNF, is upregulated
receptor (Caterina, M. J. et al., 1997). NGF-induced in the skin in response to inflammatory conditions; its
sensitization of the response to noxious heat has expression increases faster, and to a greater extent,
properties that are similar to sensitization of the than that of NGF. Administration of these agents
response to capsaicin. One of the advantages of nox- elicits thermal hyperalgesia lasting several hours
ious heat over capsaicin as the stimulus is the ability which is substantially prolonged if NGF is co-
to establish a threshold value. This has permitted the injected. These findings broaden our understanding
conclusion that NGF does not alter temperature of the role of trophic factors in sensitization of
threshold; its major effect is to enhance the magni- nociceptors.
tude of the heat-evoked current.
As observed with capsaicin, NGF does not sensi-
tize the heat-evoked response in some neurons.
Immunohistochemical characterization of the 22.6 TrkA Signaling Pathways
recorded cells has revealed that the NGF-induced Responsible for Acute Effects of
sensitization of the heat-evoked inward current is Nerve Growth Factor
closely associated with the expression of trkA
(Figure 3). This is not unexpected since studies in Signaling initiated by trk receptor activation is
heterologous cells have demonstrated that the ability diverse, and for a general treatment of this subject
for NGF to sensitize the TRPV1 response requires from a more molecular perspective, the reader is
that the cell express trkA (Chuang, H. H. et al., 2001). referred to a recent review (e.g., Huang, E. J. and
Zhu W. et al. (2004) have recently demonstrated Reichardt, L. F., 2001; Nicol, G. D. and Vasko, M. R.,
that the capsaicin- or noxious heat-induced TRPV1 2007). Many studies carried out on NGF-induced
response in DRG cells isolated from neonatal rats signaling have been directed toward determining
cannot be sensitized by NGF unlike the response in the signaling pathways involved in cell survival and
cells from adults. Thus, in neonates, there is no close axon outgrowth. Here, we provide a more focused
266 Neurotrophins and Pain

treatment related to NGFs role as a sensitizing agent current. These studies point out the complexity
in sensory neurons. of studies on signaling pathways, specifically the
Chuang H. H. et al. (2001) reported that phospha- need to define the conditions under which they are
tidyl-4.5-inositol biphosphate (PtdIns(4,5)P2  PIP2) investigated. It may be necessary to carry out such
exerts a tonic inhibitory effect on the inward current studies in vivo where conditions are more tightly
produced by TRPV1 via its interaction with the C- regulated in order to understand which of the many
terminal domain of TRPV1. They suggested that possible pathways are operating under physiological
NGF sensitizes TRPV1 by activating PLC which conditions.
inhibits PIP2, thereby disinhibiting TRPV1, a
mechanism corroborated by the demonstration that
the PLC inhibitor U-73122 antagonizes the sensitiz- 22.7 p75 Receptor Influence on
ing effect of NGF in adult rat DRG neurons Sensory Neuron Function
(Galoyan, S. M., et al., 2003). This mechanism has
not been confirmed in subsequent studies (see The p75 receptor is expressed in all cells that also
below). Furthermore, recent findings suggest that express trkA and trkB receptors, but in only about
PIP2 facilitates rather than inhibits TRPV1 receptor half the cells expressing trkC. Cells not expressing trk
activity (Stein, A. T. et al., 2006). A related finding do not express p75 (Wright, D. E. and Snider, W. D.,
indicates that PIP2 is required for recovery from 1995). p75 binds all neurotrophins (see reviews by
desensitization (Lin, B., et al., 2005) known to occur Patapoutian, A. and Reichardt, L. F., 2001; Huang, E. J.
after TRPV1 receptor activation. and Reichardt, L. F., 2001). The functional role of
Chuang H. H. and coworkers pointed out that the trk and p75 in mediating the effects of neurotrophins
activation of PLC would also be expected to enhance has received greatest attention in developmental
levels of protein kinase C (PKC) via diacylglycerol studies as determinants of neuronal survival and apop-
(DAG). However, they found that inhibitors of PKC tosis (Hempstead, B. L. et al., 1991; Miller, F. D. and
fail to abolish NGF-induced sensitization of TRPV1. Kaplan, D. R., 2001). For example, p75 acts as a med-
This has been confirmed by Shu X. and Mendell L. M. iator of sympathetic neuron apoptosis, whereas trkA
(2001) in adult rat DRG cells using a different pair of mediates cell survival (Majdan, M. et al., 2001).
PKC blockers. Interestingly, other investigators Relatively little attention was paid to the p75
working in dissociated DRG neurons have obtained receptor initially in evaluating the physiological
very different results on the signaling pathways effects of NGF because of evidence that sensitization
involved in NGF-induced sensitization. Bonnington to noxious heat by NGF is normal in p75 knockouts
J. K. and McNaughton P. A. (2003), working in neo- (Bergmann, I. et al., 1998, but see von Schack, D. et al.,
natal mouse DRG cells, used a different inhibitor of 2001, for evidence that this knockout is not com-
PLC, neomycin, and found no inhibition of NGF- plete). However, the finding that activation of p75
induced sensitization of the capsaicin response and enhances ceramide via activation of the sphingomye-
also that the most robust inhibition of NGF-induced lin pathway (Dobrowsky, R. T. et al., 1994) prompted
potentiation occurred by blocking the activity of PI- a search for physiological effects elicited by
3K and PKC, the latter in contrast to the earlier NGF binding to the p75 receptor (reviewed by
findings (see above). Zhuang Z.-Y. et al. (2004), work- Nicol, G. D. and Vasko, M. R., 2007). Zhang Y. H.
ing in adult rat (see also Zhu, W. and Oxford, G. S., et al. (2002) reported that NGF-induced activation of
2007) have confirmed the role for PI-3K in NGF- the ceramide pathway via the p75 receptor enhances
induced sensitization, agreeing with Bonnington J. K. membrane excitability (membrane gain-number of
and McNaughton P. A., but also reported that the APs per nanoampere of current applied across the
MEK inhibitor PD98059 blocked the effect of NGF cell membrane) in capsaicin-sensitive DRG neurons
unlike the findings of both Bonnington J. K. and (presumptive nociceptors) by enhancing a TTX-
McNaughton P. A. (2003) and Shu X. and Mendell insensitive Na conductance and decreasing the
L. M. (2001). magnitude of the K-rectifier. These experiments
It is important to note that there are several dif- were carried out first by short circuiting p75 by
ferences among these experiments including the use administering ceramide itself, and second by demon-
of rats and mice of different strains, cell culture con- strating that the effects of NGF could be blocked by
ditions, protocols (e.g., number of capsaicin pulses), administering glutathione, an inhibitor of neutral
different assays such as Ca2 imaging, and membrane sphingomyelinase, the enzyme responsible for
 Neurotrophins and Pain 267

producing ceramide (Liu, B. and Hannun, Y. A., 22.8 Effects of Nerve Growth Factor
1997). A p75 blocking antibody also prevents cera- on Expression of Channels
mide facilitation of membrane gain (Zhang, Y. H. and and Receptors in Nociceptors
Nicol, G. D., 2004). Thus, NGF might sensitize the
response of the cell by enhancing its depolarization The acute changes in sensory neuron function con-
(trkA) and independently by increasing the number sidered heretofore are very rapid and hence
of action potentials produced per unit of depolariza- considered to be posttranscriptional changes.
tion (p75) (Mendell, L. M., 2002). In this context, it However, there is now substantial evidence that
is interesting that one of the major deficits in p75- NGF can also regulate the expression of numerous
knockout mice is a substantial reduction of dis- receptors and channels via transcriptional mechan-
charge frequency of certain mechanoreceptors isms and in this way exert much more profound and
(Stucky, C. L. and Koltzenburg, M., 1997). long-lasting effects on the properties of nociceptors
Cellular mechanisms for interaction between p75 (Figure 4). Furthermore, in the case of several of the
and trkA have also been described. Working in channels (TRPV1, BK, P2X3, and TTX-resistant
human neuroblastoma cells, Plo I. et al. (2004) Na channels), there is evidence for differential reg-
reported that activation of trkA reduces sphingo- ulation by NGF and GDNF.
myelinase levels and thus reduces ceramide
production (i.e., reduced p75 signaling) via PKC.
Similarly, Bilderback T. R. et al. (2001) demonstrated 22.8.1 TRPV1 Receptors
in PC12 cells that PI-3K, activated by NGF via Chronic exposure of DRG cells in culture to NGF
trkA, acutely inhibits acidic sphingomyelinase acti- increases their response to capsaicin (Bevan, S. and
vated by NGF via the p75 receptor. Since PI-3K Winter, J., 1995; Ganju, P. et al., 1998); this is attributed
under certain conditions has also been shown to play to an increase in expression of TRPV1 associated with
a role in NGF-induced activation of TRPV1 via enhanced levels of TRPV1 mRNA (Winston, J. et al.,
trkA (see Section 22.6), NGF appears to have the 2001). In vivo studies have documented that peripheral
potential to affect cell discharge in a complex man- administration of NGF enhances expression of
ner involving interaction of the signaling pathways TRPV1 via activation of p38 MAP kinase in trkA
activated by p75 and trkA. Also in PC12 cells, sensory neurons in the DRG and in sciatic nerve and
MacPhee I. and Barker P. A. (1997; 1999) demon- especially in the skin by selective centrifugal transport
strated a reciprocal action of NGF, i.e., brief (Ji, R. R. et al., 2002). A recent report (Amaya, F. et al.,
activation of p75 acutely reduces trkA activity via a 2004) has shown that the elevated expression of
ceramide-dependent mechanism, whereas extended TRPV1 begins 1 day after CFA administration into
p75 activation facilitates trkA. Another recent report the paw and lasts for up to 1 week, returning to normal
(He, X. L. and Garcia, K. C., 2004) suggests that at 2 weeks. Both NGF and GDNF levels increase in
NGF can bind to p75 in a 2:1 molecular ratio and DRG neurons after CFA treatment, with NGF
that this complex might bind to trkA. Such a scheme
would predict cooperativity between p75 and trkA Sensitivity to noxious Sensitivity to
at the level of the cell surface in addition to intra- heat and lowered pH lowered pH
TRPV1 ASIC3
cellular signaling. Furthermore, there is now
evidence that p75 can modulate the action of the Activation of
BDNF
trk receptors by increasing trk receptor autopho- cells in lamina II
sphorylation (Berg, M. M. et al., 1991; Verdi, J. M. NGF DRG neuron

et al., 1994), by increasing the concentration of the BK1, BK2

Sensitivity
to bradykinin
neurotrophin in the vicinity of its trk receptor
(Barker, P. A. and Shooter, E. M., 1994), and/or by P2X3 SNS/PN3
increasing the selectivity of the trk receptor for its Sensitivity Membrane
to ATP excitability
ligand (Rodriguez-Tebar, A. et al., 1992; Benedetti,
M. et al., 1993). Thus, even though there is evidence Figure 4 The variety of possible long-term actions of
nerve growth factor (NGF) on expression of various
for independent effects of neurotrophins via trkA channels, receptors, and neurotrophins in nociceptors. In
and p75, the possibility that these pathways could each case, the major functional effect is listed. Further
interact in nociceptive neurons must be considered. details in text.
268 Neurotrophins and Pain

exhibiting a relatively rapid rise and fall (peak at 1 22.8.4 BK Receptors

day) and GDNF reaching its peak level at 1 week and
Another receptor upregulated by NGF is a member
falling to control levels by the second week.
of the bradykinin (BK) receptor family. Nociceptors
Selectively interfering with NGF and GDNF upre-
express the B2 form of the BK receptor constitu-
gulation using antibodies has clarified that NGF and
tively; the B1 form is expressed only after
GDNF elevate TRPV1 expression in different popu-
inflammation (Dray, A. and Perkins, M., 1993).
lations of nociceptive sensory neurons, trkA and
McNaughton P. A. and coworkers have reported
IB4 neurons, respectively. Furthermore, restricting
that freshly dissociated mouse DRG cells undergo
elevation of TRPV1 expression to either population of
upregulation of B2 mRNA (confirmed at the protein
DRG cells reduces the thermal nociception elicited
level immunocytochemically) when exposed to NGF
during CFA-induced inflammation.
for 3 days; GDNF is much less effective (Lee, Y. J.
et al., 2002). These studies are consistent with pre-
22.8.2 P2X3 Receptors vious observations that cultured cells are more
responsive to bradykinin when cultured in the pre-
NGF also enhances expression of the P2X3 receptor
sence of NGF (Petersen, M. et al., 1998). The identity
(Bradbury, E. J. et al., 1998; Ramer, M. S. et al., 2001)
of the receptor responsible for this action of NGF
considered important in pain produced by ATP
(trkA or p75) is controversial (see discussion in Lee,
(Cook, S. P. and McCleskey, E. W., 2002). One
Y. J. et al., 2002). Interestingly, expression of the B1
week after intrathecal administration, GDNF ele-
receptor can be enhanced in cultured DRG neurons in
vates P2X3 expression in spinal terminals of Ret/
the presence of GDNF but not NGF (Vellani, V. et al.,
IB4 neurons in which it is normally expressed.
2004). However, in vivo the effects of NGF may be
NGF has a more profound effect since it causes
much more complex as suggested by Rueff A. et al.
novel expression of P2X3 in neurons terminating in
(1996) who hypothesized that NGF might indirectly
more superficial laminae of the spinal cord where
cause upregulation of B1 receptors on DRG neurons
P2X3 is normally not expressed. Although these lat-
via release of cytokines such as TNF and IL-1
ter changes are relatively small, it was concluded that
from inflammatory cells.
they might be a significant component in NGF-
mediated inflammatory pain.
22.8.5 Na Channels
22.8.3 ASIC Channels
Waxman, S. G., Kocsis, J. D., and coworkers have
ASIC channels are Na channels gated by H. The shown that NGF can affect Na and K channel
ASIC3 isoform is expressed in DRG cells identified expression of nociceptive afferent fibers (Fjell, J.
as nociceptors by coexpression of markers character- et al., 1999; Everill, B. and Kocsis, J. D., 2000). Fjell J.
istic of such neurons. Along with TRPV1 receptors, and coworkers noted changes in the properties of
ASIC3 receptors are considered to be important small-diameter DRG cells after peripheral axotomy
mediators of the pain associated with tissue acidosis that could be reversed by administration of NGF.
(Voilley, N. et al., 2001). Mamet J. et al. (2003) have Specifically, they observed that axotomy diminishes
shown in culture that NGF enhances the expression the TTX-resistant Na current in these neurons and
of the ASIC3 isoform in DRG cells. They describe a that provision of NGF partially reverses this decline
physiological control mechanism, suggesting that low due to an effect on SNS/PN3 TTX-resistant Na
levels of NGF regulate ASIC3 expression via trkA channel (Nav 1.8). NGF is unable to reverse the
and the PLC/PKC signaling pathway. During axotomy-induced attenuation of activity in NaN,
inflammation, NGF levels rise and the increased another TTX-resistant Na channel (Nav 1.9).
activation of p75 removes inhibition of the down- However, GDNF is able to reverse the decline in
stream effects of JNK and p38 that are also both NaN and SNS/PN3. Interestingly, GDNF but
activated by NGF via trkA (Ji, R. R. et al., 2002). not NGF enhances TTX-resistant currents in small
Since these downstream effects include expression DRG neurons studied in culture, suggesting that the
of ASIC3, this mechanism can act as a switch ability to restore function after axotomy does not
whereby the enhanced level of NGF switches on predict effects under all conditions. However, given
production of ASIC3 by removing tonic inhibition the likely importance of TTX-resistant Na chan-
of JNK and p38. nels in the function of nociceptors (Ritter, A. M. and
 Neurotrophins and Pain 269

Mendell, L. M., 1992; Fang, X. et al., 2005), the finding 22.9.2 Muscle Afferents
that neurotrophins can influence their function
Recent studies have indicated that neurotrophins can
makes this system a potentially important target in
influence the discharge of small-diameter muscle
achieving pain control.
afferents. Hoheisel U. et al. (2005) evaluated the effect
of intramuscular NGF on the discharge of small-
diameter (Group IV) muscle afferents and found
22.8.6 PGE2 Receptors that only units with high mechanical threshold are
facilitated. They reported that threshold is
The multiplicity of channels affected by NGF sug-
unchanged, but discharge frequency in response to
gests that this growth factor acts as a master switch in
adequate stimulation increases virtually immediately
regulating the sensitivity of sensory neurons to noci- after NGF (0.8 mM) administration. Interestingly,
ceptive stimuli. However, its control is not universal they also observed that administration of BDNF
since NGF does not control expression of the PGE2 (1 mM) causes a decline in discharge frequency, but
receptor (Southall, M. D. and Vasko, M. R., 2000) that only several minutes after administration.
mediates the hyperalgesia caused by prostaglandins. The sensitization of high-threshold muscle affer-
ents is consistent with the sensitization observed in
response to intramuscular administration of NGF to
the masseter muscle in humans (Svensson, P. et al.,
22.9 Neurotrophins and Sensitization 2003). Changes were observed 1 and 7 days after
of Tissues Other than Skin administration but not after 1 h and were described
as consisting of both mechanical allodynia and hyper-
The skin has attracted most of the attention in studies algesia. Under pathological conditions, it seems likely
of the peripheral effects of neurotrophins. This is in that the cytokine TNF is upregulated in muscle and
keeping with the focus on this organ in investigations that this leads to production of NGF (Schafers, M.
of somatosensory mechanisms. However, there is et al., 2003).
considerable evidence that NGF contributes to the
sensitization of other tissues, specifically viscera,
muscle, and joints, all of which are sites where 22.9.3 Joint Receptors
inflammatory pain can occur.
Synovial fluid from arthritic joints in humans
expresses elevated levels of NGF (Falcini, F. et al.,
1996; Halliday, D. A. et al., 1998), which has been
22.9.1 Visceral Structures attributed to elevated levels of cytokines such as
Several investigators have observed enhanced levels TNF and IL1- (Manni, L. et al., 2003), much as
of NGF in the bladder in models of interstitial cysti- has been reported in skin (see Section 22.4.3). There
tis, a normally painful condition (Oddiah, D. et al., have been no reports of enhanced sensitivity of joint
1998; Toma, H. et al., 2000; Bielefeldt, K. et al., 2003). afferents to mechanical stimulation in response to
In human studies, the level of NGF determined in NGF although sensitization is known to occur in
models of experimental arthritis (Schaible, H. G.
bladder biopsies was found to be higher in indivi-
and Schmidt, R. F., 1985). Joints treated with CFA
duals whose condition is painful (Lowe, E. M. et al.,
have a more extensive innervation (Shinoda, M. et al.,
1997). Using viral transfer methods, Lamb K. et al.
2003), possibly as a result of the enhanced levels of
(2004) demonstrated that enhancing NGF levels in
NGF known to promote terminal axonal sprouting in
rat bladder leads to overactivity measured cystome-
the skin (Diamond, J. et al., 1992). An enhancement in
trographically. If NGF is neutralized using
peripheral axon terminals of trkA nociceptive affer-
immunological methods, the bladder overactivity
ents might be expected to increase sensitivity to
produced by external irritants is reduced (Lamb, K.
peripheral stimuli as a result of increased numbers of
et al., 2003). NGF has also been demonstrated to be receptor molecules. Recently, Shelton D. L. et al. (2005)
upregulated in other visceral organs such as the have demonstrated that administration of a monoclonal
bowel in association with inflammatory conditions antibody against NGF (mAb 911, Rinat Neuroscience)
(di Mola, F. F. et al., 2000) or in an animal model of can eliminate the pain associated with an experimental
pancreatitis (Toma, H. et al., 2000). autoimmune arthritic condition in rats.
270 Neurotrophins and Pain

22.10 Nerve Growth Factor, different neurotrophins act to promote peripheral

Enhances Peptide Expression in the and central sensitization (but see Section 22.11, indicat-
Dorsal Root Ganglion, and Central ing that BDNF and NT-4 can also elicit peripheral
Sensitization sensitization).
Direct physiological evidence for the action of
The ability of NGF to elicit rapid sensitization via BDNF in the superficial dorsal horn has come from
posttranscriptional changes and to increase the experiments in spinal slices where it has been reported
expression of receptors and channels involved in that BDNF facilitates the synaptic response of cells in
setting the sensitivity and activity of nociceptors is lamina II to electrical stimulation of C-fibers both
indicative of its ability to enhance peripheral sensiti- in neonates (Garraway, S. M. et al., 2003) and in
zation. However, NGF also elevates expression of adults (Garraway, S. M. et al., 2005). This facilitation
peptides in the DRG ganglion and these are believed of AMPA/kainate responses (measured in voltage
to enhance the central effects of glutamate released by clamp) is virtually immediate after superfusion of
nociceptors, i.e., to contribute to central sensitization BDNF (100 ng ml1) and lasts for at least a few
(McMahon, S. B. et al., 1993). Lindsay R. M. and hours after washout of the BDNF. The effect of
Harmar A. J. (1989) were among the first to demon- BDNF requires participation of NMDA receptors
strate that DRG cells maintained in culture exhibit since it is blocked by APV. Additional pharmacologi-
enhanced levels of substance P and CGRP after cal experiments have demonstrated a role for PKC in
exposure to NGF in the medium. These peptides this process and also that administration of BDNF
are considered to be important mediators in central results in phosphorylation of the NMDA receptor in
sensitization because of the very long-lasting depo- neurons in the superficial dorsal horn (Slack, S. E. et al.,
larization that they produce in postsynaptic cells, 2004). It is important to note that BDNF itself does not
leading to activation of the NMDA receptor elicit any detectable effect on lamina II neurons; it
only acts to sensitize the response to stimulation of
(McMahon, S. B. et al., 1993). Both NGF and
small-diameter sensory afferents. The neurotrophin
GDNF enhance capsaicin-stimulated release of
effect is specific; superfusion of NT-3 does not sensi-
CGRP from cultured trigeminal ganglion neurons;
tize this response (Garraway, S. M. et al., 2003).
however, the effect of GDNF is due entirely to the
The requirement for functional NMDA receptors
greater survival of these neurons in its presence (i.e.,
in order for BDNF to be effective in sensitizing the
no change in response of individual neurons to
response of lamina II neurons to C-fiber input is
capsaicin), whereas the effect of NGF is attributed
consistent with the behavioral finding that pretreat-
in part to increased CGRP release per neuron, i.e.,
ing rats for several days with systemic MK-801, an
NGF sensitizes the response to capsaicin (Price, T. J. antagonist of NMDA receptors, selectively blocks the
et al., 2005). late phase of thermal and mechanical hyperalgesia
The possibility for direct participation of neurotro- elicited by NGF (Lewin, G. R. et al., 1994). This late
phins in central sensitization was initially advanced by phase was attributed to central sensitization, whereas
the finding in vivo that BDNF is also upregulated in the the acute phase, not affected by MK-801 treatment,
cell body of trkA receptor-expressing DRG cells several was identified with peripheral sensitization. Thus,
hours to days after exposure to NGF (Apfel, S. C. et al., NGF exerts its effects by peripheral and central
1996; Michael, G. J. et al., 1997). This suggested that sensitization which together have been advanced as
BDNF would be released into the spinal cord, a predic- mechanisms to explain different components of
tion that has been borne out experimentally (Lever, I. J. hyperalgesia (McMahon, S. B. et al., 1993).
et al., 2001). The high-affinity receptor for BDNF, trkB,
is found on neurons throughout the superficial dorsal
horn (Bradbury, E. J. et al., 1998; Garraway, S. M. et al., 22.11 Role of Brain-Derived
2003). A physiological role for BDNF has been estab- Neurotrophic Factor, Neurotrophin-4,
lished in models of inflammatory pain by demonstrating and Neurotrophin-3 in Peripheral
that sequestration of spinal BDNF using intrathecally Sensitization
delivered trkB-IgG diminishes the response to noxious
heat (Kerr, B. J. et al., 1999; Mannion, R. J. et al., 1999; There is increasing evidence that acute sensitization
Pezet, S. et al., 2002). In these experiments, it was also of nociceptors can be affected by neurotrophins other
demonstrated that NGF has no central action, i.e., than NGF. Specifically, BDNF and NT-4 sensitize
 Neurotrophins and Pain 271

nociceptive function acutely at the behavioral level associated with nerve manipulations that are
(Shu, X. et al., 1999), at the level of nociceptor dis- accepted as models for neuropathic pain. Thus,
charge measured in isolated skin nerve preparations Zhou X. F. et al. (2000) have demonstrated that anti-
(Rueff, A. and Mendell, L. M., 1996; Shu, X. et al., bodies to NGF, BDNF, and NT-3 delivered to the
1999), and at the level of capsaicin-activated TRPV1 DRG can reduce the pain associated with spinal
currents measured in dissociated cells (Shu, X. and nerve axotomy. Interestingly, antibodies to NGF eli-
Mendell, L. M., 1999b). The proportion of small cells cit their effect in the early stages after injury, whereas
(d  30 mm) sensitized by NT-4 is similar to that BDNF and NT-3 antibodies exert their effects only
sensitized by NGF. Therefore, it seems unlikely later. In addition, Zhou X. F. et al. demonstrated that
that trkA expression is required for cells to be sensi- BDNF and NGF, but not NT-3, delivered to the
tized by NT-4 since as a population, relatively few DRG in intact preparations elicit mechanical allody-
(10%) small trkA sensory neurons also express trkB nia. It is worth contrasting these findings with those
(Karchewski, L. A. et al., 1999). Thus, activation of of Kerr B. J. et al. (1999; see above) who showed that
trkB alone can sensitize the response to capsaicin. NGF delivered intrathecally elicits no nociceptive
It is not known where trkB ligands would come behavior. These differences presumably reflect the
from naturally in the periphery since there is no differences in the expression of trkA that is known to
evidence at present that BDNF or NT-4 is upregu- be present in the DRG, but not in the spinal cord.
lated in the skin immediately after inflammation. Theodosiou M. et al. (1999) carried out similar
However, BDNF is upregulated in nerves under- experiments with neurotrophin antibodies in rats
going Wallerian degeneration, opening the subject to chronic partial axotomy of the sciatic
possibility that trkB ligands sensitize neurons after nerve. They observed both mechanical and thermal
damage to neighboring nerve trunks. Another possi- hyperalgesia in these rats that could be relieved by
bility is that BDNF, which is known to be released antibodies to NGF or BDNF but not NT-3.
from the central terminals of nociceptors, would also Fukuoka T. et al. (2001) have studied uninjured
be released from their peripheral terminals in the neurons in the L4 DRG in rats subjected to L5 spinal
skin and that this would be the normal source of nerve ligation (Chung model). They observed an
trkB ligands. Such a line of reasoning suggests that increase in BDNF mRNA levels in L4 associated
BDNF might act peripherally to reinforce the central with increased levels of expression as well as recruit-
sensitization it also elicits. ment of additional neurons into the population of
NT-3 does not elicit any acute sensitization of the DRG neurons expressing BDNF mRNA. They also
response to noxious heat measured either electro- found that the enhanced sensitivity to noxious heat
physiologically or behaviorally (Shu, X. et al., 1999; associated with this injury could be diminished tem-
Shu, X. and Mendell, L. M., 1999b) despite evidence porarily by intrathecal application of an antibody to
that it can bind to trkA (Lindsay, R. M., 1996). BDNF. Significantly, NGF appears to play a role in
However, 7-day exposure of trkA/trkCsensory this enhancement of BDNF levels since NGF is
neurons to NT-3 results in a substantial reduction increased in the DRG whose spinal nerve is ligated,
in trkA receptor expression (Gratto, K. A. and Verge, and an antibody to NGF delivered to the injured
V. M., 2003). Accompanying this loss of trkA is a DRG can prevent the upregulation of BDNF in the
decline in substance P levels. These studies suggest neighboring ganglion as well as the associated ther-
that NT-3 might have therapeutic value in diminish- mal hyperalgesia.
ing painful states initiated by the activation of trkA. A recent study (Wilson-Gerwing, T. D. et al.,
2005) indicates that NT-3 can prevent the long-last-
ing increased expression of TRPV1 observed in the
22.12 Neurotrophins and chronic constriction injury (CCI) model of neuro-
Neuropathic Pain pathic pain. They further demonstrated that NT-3
reduces expression of p38 MAPK shown to be a
Most considerations of neurotrophins and pain have primary factor in the delayed elevation of TRPV1
centered on their role in inflammatory pain since expression in the skin (Ji, R. R. et al., 2002). Treatment
they are naturally upregulated in the both periphery with NT-3-reduced thermal hyperalgesia, but not
and DRG and play a prominent role in peripheral the mechanical hypersensitivity associated with CCI.
and central sensitization. However, there is also evi- A different role for neurotrophins in neuropathic
dence that neurotrophins may contribute to the pain pain was reported by Bennett D. L. et al. (1996b) who
272 Neurotrophins and Pain

demonstrated that sprouting of peripherally axoto- localized injections of NGF (13 mg per patient) in
mized A-fibers into the superficial laminae of the the arm resulted in localized allodynia and thermal
spinal cord, considered to be a possible mechanism hyperalgesia lasting several weeks (Dyck, P. J. et al.,
for neuropathic pain after such damage (Woolf, C. J. 1997). These experiments in humans with verbal
et al., 1995), is reduced by application of NGF to the reports provide significant confirmation of the results
damaged nerve. They argued that this was not a from animal experiments. They also suggest that the
direct effect of NGF on the sprouting of A-fibers use of NGF to increase cell survival or for other
since it would imply that sprouting is paradoxically purposes will require measures to overcome the
reduced by administration of NGF. Also, large non- nociceptive side effects.
nociceptive A-fibers do not express trkA, and so it is As discussed above, peripheral NGF levels are
not clear how NGF would affect their sprouting. upregulated as a consequence of inflammation result-
They proposed that NGF elicits sprouting of axoto- ing in hyperalgesia. Furthermore, the hyperalgesia
mized C-fibers that would reverse the A-fiber associated with inflammation can be largely elimi-
sprouting caused initially by the peripheral axotomy. nated by reducing NGF levels using either a
The pain associated with peripheral nerve damage sequestering antibody or a trkA immunoadhesin.
has been shown to also have a microglial component. This has prompted the development of NGF anti-
ATP released from damaged nociceptive afferents bodies for eventual use in humans. Recently, Sevcik
activates microglial P2  4 receptors; the activated M. A. et al. (2005) reported that an antibody mAb 911
microglia release BDNF which inhibits the KCC2 (Rinat Neuroscience Corporation) is effective in
chloride transporter in lamin I cells (Coull, J. A., et al., attenuating the pain in a mouse model of bone cancer
2005). This shifts the chloride equilibrium potential whose sarcoma tumor cells are a source of high levels
in the depolarizing direction which changes the sign of NGF. As shown above (Section 22.9), bladder pain
of inhibitory input toward excitation. This increases associated with models of cystitis is also associated
activity in nociceptive pathways contributing to pain. with elevated expression of NGF, as is arthritic pain.
Thus, microglial BDNF as well as neuronal (DRG) Thus, we can expect that clinical trials using will be
BDNF can contribute to central sensitization. undertaken as soon as antibodies have been rendered
Interaction between these two sources of BDNF safe for human use.
remains to be discovered. Several clinical conditions reported to be asso-
Despite the lack of consistency, these experiments ciated with alterations in NGF or trkA expression
indicate several potential roles for neurotrophins in have been reviewed by Anand P. (2004). The trkA
neuropathic pain. These are based on their ability to receptor is dysfunctional in individuals with conge-
directly sensitize neuronal responses as well as their nital insensitivity to noxious stimulation, probably as
ability to elicit structural changes in the sensory a result of a mutation. Leprosy, a condition associated
nervous system. with hypoalgesia, is accompanied by reduced expres-
sion of NGF. Similar findings have been made in
diabetic neuropathy. Conversely, the pain associated
22.13 Neurotrophins and the Clinic with injury to nerves has been correlated with
enhanced expression of NGF in the associated
The very strong sensitizing effect of neurotrophins nerve trunks. Together, these findings suggest that
on nociceptive pathways both peripherally and in the manipulation of neurotrophin levels may turn out to
spinal cord has given them a very high profile in pain be a useful therapeutic approach to various condi-
research. One of the earliest indications of a clinical tions associated with disorders of nociception.
effect emerged from Phase I clinical trials to deter-
mine the safety of NGF delivered systemically
(Petty, B. G. et al., 1994). NGF was administered 22.14 Discussion, Conclusions, and
subcutaneously or intramuscularly in human volun- Open Questions
teers in amounts ranging from 0.03 to 1.0 mg kg1,
dosages that are much lower than typically used in The past 15 years have seen an important expansion
rats (1 mg g1). These subjects experienced myalgia in our understanding of the role of neurotrophins as it
in truncal and bulbar muscles as well as hyperalgesia has become generally accepted that they function
at the site of injection. The latter could persist for up throughout the life of the animal, not only during
to 7 weeks. Similar human trials using single development. With regard to their effect on
 Neurotrophins and Pain 273

nociception, it is clear that the survival of nociceptors Finally, the fact that NGF has so many different
is dependent on the integrity of the NGF/trkA sys- possible actions on nociceptive function in terms of
tem during the perinatal period. Beyond this period, the variety of cells that express trkA (Section 22.4.3)
the influence of NGF on nociceptors is very wide as well as the multiple effects that NGF can elicit in
ranging (Petruska, J. C. and Mendell, L. M., 2004). individual nociceptors (Sections 22.8 and 22.10) raises
NGF sensitizes nociceptors both acutely via post- important questions about how all these mechanisms
transcriptional mechanisms (Figure 2) and interact to cause sensitization. If the scheme outlined in
chronically by enhancing the expression of numerous Figure 2 is a complete representation of the peripheral
channels associated with nociceptors (Figure 4). mechanism of NGF action, one would expect that
These effects are exerted on nociceptors innervating elimination of mast cells would reduce but not abolish
many different tissues, including skin, muscle, joints, NGF-induced sensitization. However, acute sensitiza-
and viscera although at this time it is not yet tion is abolished (Lewin, G. R. et al., 1994), suggesting
known whether these neurons are regulated identi- that NGF activation of the neutrophil pathway
cally by NGF. (Bennett, G. et al., 1998b) or direct activation of
The discovery of the capsaicin receptor VR1/ nociceptor terminals is not sufficient to mediate the
TRPV1 (Caterina, M. J. et al., 1997), the revelation acute phase of NGF-induced hyperalgesia. Similarly,
that this receptor mediated the response to noxious inhibiting neutrophil accumulation completely elimi-
heat (Tominaga, M. et al., 1998), and the finding that nates the hyperalgesic effect of NGF (Bennett, G. et al.,
NGF could sensitize the response to capsaicin (Shu, X. 1998b), indicating that the mast cell or direct effects are
and Mendell, L. M., 1999b) or to noxious heat not sufficient either. Together, these findings suggest
(Galoyan, S. M. et al., 2003) seemed to be entirely an interaction between these different mechanisms,
consistent with the behavioral evidence that NGF and it will be important to dissect these interactions
sensitizes the response to noxious heat. However, in order to more fully understand how NGF (and
several experiments have raised difficulties with this neurotrophins more generally) acts as a sensitizing
simple picture. A substantial fraction of cell mem- agent for nociceptors. In view of increasing interest in
brane patches from small-diameter sensory neurons manipulating NGF levels in a clinical setting, it will be
were found to be sensitive to capsaicin (indicating necessary to understand its physiology at a more inte-
TRPV1 expression) or noxious heat, but not both grative level.
(Nagy, I. and Rang, H. P., 1999), suggesting that the
receptor for capsaicin and for noxious heat are not
the same. Furthermore, although two studies in dis- Acknowledgment
sociated cells from TRPV1/ mice revealed the
expected deficit in noxious heat sensitivity I thank Dr. Jeff Petruska for useful comments on an
(Caterina, M. J. et al., 2000; Davis, J. B. et al., 2000), early draft of the manuscript and for help with fig-
the same reports demonstrated that less reduced pre- ures. The authors research was supported by NIH
parations (skin nerve or in vivo preparations) (NS39420; NS16996) and the Christopher Reeve
displayed considerable residual sensitivity to noxious Paralysis Foundation Consortium on Spinal Injury.
heat. This raises questions concerning our model of
how NGF acts to sensitize the response to noxious
heat specifically whether other noxious heat recep-
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 23 Morphological and Neurochemical Organization
of the Spinal Dorsal Horn
A Ribeiro-da-Silva, McGill University, Montreal, QC, Canada
Y De Koninck, Centre de recherche Universite Laval Robert-Giffard, Quebec, QC, Canada
2009 Elsevier Inc. All rights reserved.

23.1 Introduction 280

23.2 Overall Organization of Dorsal Horn: Rexed Lamination 280
23.3 Primary Afferent Fibers 283
23.3.1 Types of Afferents, Neurochemistry, and Termination in Spinal Cord 283
23.3.2 Synaptic Arrangements of Primary Afferents 284
23.3.3 Receptors on Primary Sensory Fibers 286
23.3.4 Expression of Voltage-Gated Sodium Channels by Sensory Fibers 288
23.4 Dorsal Horn Neurons 288
23.4.1 Lamina I Neurons 289
23.4.1.1 Morphological classification of lamina I neurons 289
23.4.1.2 Correspondence with function 289
23.4.1.3 Expression of NK-1r in subpopulations of lamina I neurons 291
23.4.2 Lamina II Neurons 292
23.4.3 Laminae IIIVI 293
23.4.4 Neurochemistry of Dorsal Horn Neurons 294
23.4.4.1 Neurokinins 294
23.4.4.2 Neurokinin receptors 295
23.4.4.3 CGRP and CGRP receptors 295
23.4.4.4 Somatostatin 295
23.4.4.5 Opioid peptides 295
23.4.4.6 Opioid receptors 296
23.4.4.7 Glutamate 296
23.4.4.8 Glutamate receptors 297
23.4.4.9 Inhibitory amino acids 297
23.4.4.10 GABA and glycine receptors 298
23.4.4.11 Other classical transmitters and other neuropeptides 298
23.5 Identified Neuronal Circuits 299
23.6 Conclusion and Future Directions 301
References 302

Glossary
bouton Dilated portion of an axon possessing translucent and gelatinous appearance when
synaptic vesicles and establishing synapses examined in fresh tissue.
with adjacent dendrites or other neuronal compo- superficial dorsal horn Region of the dorsal
nents. Synonyms axonal varicosity or axon horn corresponding to laminae I and II of
terminal. Rexed.
presynaptic dendrite Dendrite possessing synaptic glomerulus (of the spinal
synaptic vesicles, and which is presynaptic to other cord) Complex synaptic arrangement in which a
neuronal processes, mainly dendrites. In the dorsal central axonal bouton, of primary sensory afferent
horn, presynaptic dendrites occur in islet cells. origin, establishes synaptic contacts with several
substantia gelatinosa (of Rolando) Synonym of processes from dorsal horn neurons.
lamina II of Rexed. So named because of the

279
280 Morphological and Neurochemical Organization of the Spinal Dorsal Horn

23.1 Introduction the 1970s to establish criteria to identify the laminae

at the ultrastructural level (for reviews see Light, A.
The superficial laminae of the dorsal horn of the R., 1992; Ribeiro-da-Silva, A., 2004).
spinal cord, particularly the marginal layer (or lamina Despite some recent criticism (Woodbury, C. J.
I of Rexed) and the substantia gelatinosa (or lamina II et al., 2000), the Rexed lamination remains the refer-
of Rexed), represent the area of the central nervous ence. Unfortunately, it is not always followed
system (CNS) where the first modulation of pain- accurately, and many published micrographs show
related information occurs. Although some progress laminar limits that were based more on speculation
has been made in recent years concerning our knowl- rather than on objective evaluation using the proper
edge of the anatomical and neurochemical approaches. This is true particularly for ultrastruc-
characteristics of the relevant cells and systems, our tural studies and also for those using radioactive
understanding of this area is far from complete. In ligand binding. This is unfortunate because, as we
particular, despite significant recent progress, our will discuss below, it is easy to set up the limits of the
understanding of the synaptic circuitry and how neu- laminae with acceptable accuracy (see Figures 1 and
rotransmitters/neuromodulators in this region 2 and below).
interact with their receptors is still incomplete, and Besides neuroglia and vessels, the spinal dorsal
even less is known about the changes that occur in horn contains several elements of neuronal origin:
acute and chronic pain (for recent review see Todd, (a) the final arborization and termination of primary
A. J. and Ribeiro-da-Silva, A., 2005). This chapter afferent fibers; (b) local circuit neurons, some excita-
presents an overview of what is known concerning tory and others inhibitory; (c) projection neurons to
the morphology and neurochemistry of the spinal the brain; (d) propriospinal neurons, which intercon-
dorsal horn, attempting to extract an emerging inte- nect different levels of the spinal cord; and (e)
grated view of the morphological and neurochemical descending axons from several supraspinal sources.
organization in this area of the CNS from the avail- Lamina I of the dorsal horn is also known as the
able literature. marginal layer and lamina II as the substantia gelati-
Because most of the work on the structure of the nosa (of Rolando). These two laminae together
spinal cord has been carried out on the rat, we have represent what is frequently called the superficial
based our description on this species, but when pos- dorsal horn and are of particular importance for the
sible have made comparison to other species. We spinal processing of pain-related information and its
have focused on the superficial dorsal horn because forwarding to higher levels. This is true in particular
of its relevance in the transmission and modulation of for lamina I and the outer 2/3 of lamina II, as the
pain-related information. inner 1/3 is mostly nonnociceptive. In contrast, lami-
nae IIIIV represent what is called the nucleus
proprius, which was thought to represent a nonnoci-
ceptive region of the spinal cord. While this view is
23.2 Overall Organization of Dorsal mostly valid, this region does contain neurons that
Horn: Rexed Lamination are known to respond to noxious stimuli and project
to the brain, which were initially thought to occur
The first subdivision of the dorsal horn into horizon- only in lamina V (De Koninck, Y. et al., 1992; Ma, W.
tal laminae, based on the morphological properties of et al., 1996).
the cells in a Nissl-type staining, was performed in Figure 1 shows the laminar organization of the rat
the cat dorsal horn (Rexed, B., 1952; 1954). It was spinal cord at the C4 level, as detected using a Nissl
later verified that this lamination could be adapted method on thick sections, and the correlation with
with minor modifications to almost any mammalian laminae detected on semithin plastic sections.
species. In particular, it has been adapted to Although the criteria are quite different, there is a
the monkey (Ralston, H. J., III, 1979), human very good match between the laminae when identi-
(Harmann, P. A. et al., 1988), rat (Steiner, T. J. and fied by the Nissl approach and the semithin sections
Turner, L. M., 1972; Molander, C. et al., 1984; 1989), approach. When using a Nissl staining, lamina I can
and mouse (Ma, W. Y. et al., 1995), besides nonmam- be recognized by a rather low cell density
malian species such as the pigeon (Leonard, R. B. and (Figure 1(a) and (1b)), with an occasional large neu-
Cohen, D. H., 1975). Through a correlation of light ron among mostly small cells. Lamina II is a layer
and electron microscopy, it has been possible since with a higher density of cells than lamina I; these cells
 Morphological and Neurochemical Organization of the Spinal Dorsal Horn 281

(a) (b)

20 m DC

DC
I

IIo
I
IIo
IIi
IIi
III
IV

V
VI
III

40 m IV

(c)
DC

I
IIo

IIi

III

IV
40 m

Figure 1 Rexeds laminae at the C4 level of the adult rat spinal cord. (a) and (b) represent micrographs from frozen cross
sections, 50 mm in thickness, stained with toluidine blue as described by Rexed B. (1952). In (a), note the overall laminar
organization of the dorsal horn. In (b), note that lamina I (I) has a low density of cells, whereas outer lamina II (LIIo) stands out
because of the clustering of small neurons, which differentiate it from the inner part of lamina II (IIi). Both LIIo and inner lamina
II (LIIi) have numerous small neurons, of rather uniform size. The presence of some slightly larger neurons separates lamina III
from LIIi, whereas lamina IV can be easily distinguished from lamina III by the lower cell density and the presence of some
large neurons. (c) represents a micrograph from an Epon-embedded semithin (2 mm thick) transverse section of the rat dorsal
horn at the C4 level, stained with toluidine blue and Azur II, to illustrate that Rexeds laminae can be identified in these
sections, which are used as reference for ultrastructural observations; notably, lamina IIi can be clearly delineated from lamina
III by the absence of myelin (see text for details). DC, dorsal columns.

are rather small in size and occur in a considerably scheme as it should and some authors simply con-
higher density in outer lamina II (LIIo) than in inner sider LIIo as the dorsal half of lamina II. Although in
lamina II (LIIi). It should be noted that in many rat we have used lamina IIA to refer to LIIo and
articles the subdivision of lamina II into outer and lamina IIB to refer to LIIi (see Ribeiro-da-Silva, A.,
inner parts does not follow a cytoarchitectonic 2004, for rationale), in the current chapter we will use
282 Morphological and Neurochemical Organization of the Spinal Dorsal Horn

the nomenclature followed in most of the literature. can be distinguished from LIIo by the presence of a
The border between LIIi and lamina III is difficult to higher density of small myelinated fibers, whereas
distinguish in Nissl-type preparations, as the main LIIi is virtually devoid of myelin (Figure 1(c)). In
features are similar except for the presence of some contrast, lamina III is easily distinguished from IIi by
larger cells in lamina III. Lamina IV, in contrast, is the appearance of numerous thinly myelinated fibers
easy to distinguish from lamina III because of the (Figure 1(c)).
lower cell density and the presence of some rather At mid-lumbar levels, which represent the region
large neurons (Figure 1(b)). innervating the hind limbs and which has been stu-
The superficial laminae can also be recognized in died extensively, the thickness of the superficial
Epon-embedded 12-mm-thick semithin sections laminae is greater in the medial 2/3 than in the
stained with toluidine blue, as this approach has lateral third. Overall the thickness of LI and LIIo is
been validated by integrated light and electron twice the value of mid-cervical levels. Figure 2 illus-
microscopic studies (Ribeiro-da-Silva, A. and trates the laminar subdivisions at the level of the L5
Coimbra, A., 1982). In these preparations, lamina I segment of the young rat spinal cord.

UV Nissl

White matter
LI
LIIo

LIIi

LIII

Postnatal day 20

IB4 CGRP

40 m

Figure 2 Rexeds laminae at the L5 level of the spinal cord of a young rat (20-day old). Micrographs represent confocal
images that result from the fusion of several optical sections obtained using a multitrack approach on a 50-mm-thick
transverse section. Section was immunostained for calcitonin gene-related peptide (CGRP) and incubated for IB4 binding
and a Nissl-type labeling using an ultraviolet fluorescent marker. The Nissl-type labeling was used to identify the Rexeds
laminae and to create an overlay, which was applied to the other images, and is shown isolated in the upper left picture. This
figure clearly illustrates that CGRP immunostaining occurs in lamina I (LI), outer lamina II (LIIo), and part of inner lamina II (LIIi),
whereas IB4 labeling is most intense in the dorsal part of LIIi. Because the thickness of each lamina is rather constant at the
same level of the spinal cord, overlays created like this are useful to identify the laminae even in material not stained with a
Nissl method.
 Morphological and Neurochemical Organization of the Spinal Dorsal Horn 283

23.3 Primary Afferent Fibers Merighi, A. et al., 1991). In the spinal cord, sensory
fibers that express SP and CGRP terminate mostly in
23.3.1 Types of Afferents,
laminae I, outer II, and lamina V; those that colocalize
Neurochemistry, and Termination in Spinal
CGRP and SOM terminate in laminae I and II, and
Cord
those that contain FRAP, bind the lectin GSA-IB4 or
The great majority of afferents that transmit pain- express the purinergic P2X3 receptor, terminate
related information (nociception) are of small dia- mostly in the middle third of lamina II (see Figure 2;
meter and have unmyelinated (C) or thinly for reviews see Bradbury, E. J. et al., 1998; Ribeiro-da-
myelinated (A) axons. Most such small-diameter Silva, A., 2004). Not all the nociceptive afferents that
fibers convey pain-related information and, therefore, express SP are C fibers; some are thinly myelinated
are often named nociceptors. These small-diameter (A) afferents. There is a significant population of
fibers represent the central processes of pseudounipo- nociceptive fibers that are A and were shown to
lar neurons of small or medium size located in the represent high-threshold mechanoreceptors (HTM)
dorsal root ganglia (DRG) and terminate in the super- after being characterized electrophysiologically and
ficial laminae of the dorsal horn of the spinal cord filled intracellularly with horseradish peroxidase in
(for reviews see Grant, G. and Robertson, B., cat and monkey; they were shown to terminate in
2004; Ribeiro-da-Silva, A., 2004). The smaller- laminae I and V (Light, A. R. and Perl, E. R., 1979).
diameter (unmyelinated) nociceptive afferents can be The morphological and neurochemical identification
divided into two major subpopulations, the of these A HTM fibers is difficult because they do
peptidergic and the nonpeptidergic. The peptidergic not contain peptides, do not bind IB4, and do not
expresses substance P (SP) and calcitonin gene-related express P2X3 receptors. However, there is evidence
peptide (CGRP), and the nonpeptidergic possesses from an in situ hybridization study that the 5-HT3
fluoride-resistant acid phosphatase (FRAP) activity, may be highly expressed by A HTM fibers (Zeitz, K.
P. et al., 2002). Although the prevailing view is that A
binds the lectin GSA-IB4, and expresses purinergic
fibers are nonnociceptive, it has been suggested that
P2X3 receptors (Hunt, S. P. and Rossi, J., 1985;
some of the myelinated nociceptors conduct in the A
Alvarez, F. J. and Fyffe, R. E., 2000; Hunt, S. P. and
range. In rat, these A nociceptors might represent up
Mantyh, P. W., 2001). These two populations differ in
to 20% of all A somatic afferents according to some
neurotrophic support in the adult. In fact, during
reports (for review see Djouhri, L and Lawson, S. N.,
development, both populations require nerve growth
2004).
factor (NGF) for survival, but shortly after birth only
Although a small number of C fibers are nonno-
the peptidergic continues to respond to NGF, whereas
ciceptive and convey either innocuous mechanical or
the nonpeptidergic population starts to respond thermal information, most nonnociceptive mechan-
instead to glial cell line-derived neurotrophic factor osensitive afferents are either thinly myelinated (A)
(GDNF) (Bennett, D. L. H. et al., 1998). Accordingly, or thick afferents (A) (Alvarez, F. J. and Fyffe, R. E.,
the peptidergic population expresses the NGF high- 2000). Most innocuous A afferents innervate down
affinity receptor, trkA, whereas the nonpeptidergic hairs (D hair afferents) and terminate in the deeper
expresses GDNF receptors. It has also been shown part of lamina II and in lamina III, a distribution that
that the latter population expresses the purinergic contrasts with the preferential termination of noci-
receptor P2X3 (Bradbury, E. J. et al., 1998; Snider,W. ceptive A afferents in laminae I and V (Light, A. R.
D. and McMahon, S. B., 1998). Although the distinc- and Perl, E. R., 1979). Like the nociceptive fibers,
tion between two populations of primary sensory the nonnociceptive afferents are glutamatergic
fibers, peptidergic and nonpeptidergic, seems attrac- (Battaglia, G. and Rustioni, A., 1988; De Biasi, S.
tive, it is not fully accurate as a small proportion of and Rustioni, A., 1988; Merighi, A. et al., 1991).
peptidergic sensory fibers (those that colocalize Curiously, the T-type calcium channel Cav3.2
CGRP and somatostatin (SOM)) do not respond to seems to be expressed exclusively by D hair afferents,
NGF in the adult and bind the lectin GSA-IB4 as detected by in situ hybridization at the level of the
(Alvarez, F. J. and Fyffe, R. E., 2000; Priestley, J. V. DRG (Shin, J. B. et al., 2003).
et al., 2002). It should be noted that, in all of the above It should be noted that other neuropeptides have
putative nociceptive fibers, the classical synaptic been localized in the fibers that contain SP and
transmitter is very likely glutamate (Battaglia, G. and CGRP: neurokinin A (Dalsgaard, C. J. et al., 1985),
Rustioni, A., 1988; De Biasi, S. and Rustioni, A., 1988; galanin (Ju, G. et al., 1987; Zhang, X. et al., 1993), and
284 Morphological and Neurochemical Organization of the Spinal Dorsal Horn

Table 1 Overview of the classical transmitter and peptidergic innervation of the spinal dorsal horn of the rat, as described
in text

Origin of neurotransmitter/neuropeptide

Local circuit Neurons projecting to

>Primary afferents neurons brain Descending fibers

Glu SP CGRP End-2

a
Glu SP ENK Glu SP ENK 5-HT SP TRH
(Galb) (CCKc) (VIPc)
Glu SOM CGRP Glu NKB Glu dynorphins (?) 5-HT ENK
Glu dynorphin B (at sacral levels) ? Glu SOM Glu Gal CCK GABA
Glu ? Endomorphin-1 ? GABA Gly
Glu (NPYc) GABA ENK Noradrenaline
GABA Ach
GABA NPY
Glu Gal CCK (?)
Glu dynorphins (?)
Glu neurotensin
a
All neurons that express SP colocalize NKA in rat. Neurochemical in parenthesis are not expressed or expressed in a reduced number of
afferents in the absence of lesion.
b
Galanin is strongly upregulated after nerve lesion.
c
These peptides are expressed only in a significant number of primary afferents following peripheral nerve lesion.
This table does not intend to be a complete list. It is restricted to the peptides mentioned in the text.
Abbreviations: Ach, acetylcholine; CCK, cholecystokinin; CGRP, calcitonin gene-related peptide; End-2, endomorphin 2; ENK,
enkephalin; Gal, galanin; Glu, glutamate; NKA, neurokinin A; NKB, neurokinin B; NPY, neuropeptide Y; SP, substance P; SOM,
somatostatin; TRH, thyrotropin-releasing hormone; VIP, vasocative intestinal peptide.

the opioid peptide endomorphin-2 (Martin-Schild, S. dorsal horn neurons. They also constitute important
et al., 1997; Martin-Schild, S. et al., 1998). Dynorphin modulatory devices as the primary afferent terminals
B has been detected in some primary sensory fibers are often postsynaptic to other neuronal profiles.
from visceral afferents from pelvic viscera at sacral Since the central terminals of glomeruli are all of
levels in the cat (Basbaum, A. I. et al., 1986). Some primary afferent origin, as has been shown by their
neuropeptides are not expressed in any significant complete disappearance following multiple dorsal
amount in primary sensory fibers in nave rats but rhizotomies (Coimbra, A. et al., 1984), they provide a
are upregulated following peripheral axotomy: very useful means of identifying these afferents with
vasoactive intestinal peptide (VIP) and cholecystoki- electron microscopy, without the need for transported
nin (CCK) appear on small, peptidergic afferents, or immunocytochemical markers. The detailed fea-
whereas neuropeptide Y (NPY) appears in large-dia- tures of synaptic glomeruli in the rat have been
meter afferents (for review see Hokfelt, T. et al., reviewed in a recent publication (Ribeiro-da-Silva,
1994). Table 1 gives an overview of classical trans- A., 2004). To be classified as a synaptic glomerulus,
mitters and peptides in the dorsal horn, including a synaptic arrangement must meet simultaneously all
those in the terminals of primary sensory fibers. of following criteria when viewed in a single ultrathin
section: (a) it should possess a central terminal (C)
with round synaptic vesicles; (b) this C terminal has to
23.3.2 Synaptic Arrangements of Primary be surrounded by at least four neuronal profiles,
Afferents representing either dendritic profiles or axonal bou-
In the dorsal horn, primary afferent boutons establish ton profiles; (c) there have to be at least two visible
mostly simple axodendritic, and to a lesser extent, synaptic contacts between the central bouton profiles
axosomatic synapses. However, a significant propor- and the peripheral profiles. Two main types of synap-
tion of primary afferent endings participate in tic glomeruli have been identified in rat by Ribeiro-
complex arrangements named synaptic glomeruli, da-Silva A. and Coimbra A. (1982), and are named
where they form the central terminal (Coimbra, A. type I and type II. Glomeruli of type I have a central
et al., 1974; Ribeiro-da-Silva, A. and Coimbra, A., bouton of scalloped contour, with an electrondense
1982). Glomeruli represent multiplier systems, i.e., matrix, with rather densely packed synaptic vesicles
devices that transmit sensory information to several and a low density of mitochondria, whereas type II
 Morphological and Neurochemical Organization of the Spinal Dorsal Horn 285

glomeruli are larger and possess a central terminal,

which has a rounder contour, a lighter cytoplasmatic
matrix, synaptic vesicles, which are more uniform in
D G
size and less densely packed, and a higher density of
mitochondria (Ribeiro-da-Silva, A. and Coimbra, A.,
1982). This classification is applicable to the mouse D
V1
and, with some modifications, to the cat and the
monkey (unpublished observations). A comparable
classification was established for the monkey dorsal V2
horn (Knyihar-Csillik, E. et al., 1982b), in which three
V2
glomerular types were described, based on the prop-
erties of their central terminals. Of the three types C
described in the monkey by Knyihar-Csillik E. et al.
(1982b), the dense sinusoid axon (DSA) type corre-
sponds to the type I of Ribeiro-da-Silva A. and D
D
Coimbra A. (1982) and the regular synaptic vesicles
(RSV) type corresponds to type II. Although the large G
dense-core vesicle (LDCV) type of Knyihar-Csillik E.
et al. (1982b) has apparently no equivalent in the V1
classification of Ribeiro-da-Silva A. and Coimbra A.
(1982), this type has been considered in rat as a sub- Excitatory synapse
type of type I (subtype Ib), as in this species most of Inhibitory synapse
their central terminals share the dark axoplasm, the
sinuous contour, and small size with type I central Figure 3 Diagram of a synaptic glomerulus of type Ia, with
representation of the complex synaptic arrangements of the
boutons, although they display numerous dense-core triadic type. The drawing was inspired by an electron
vesicles (Ribeiro-da-Silva, A. et al., 1989; Ribeiro-da- micrograph of a type Ia glomerulus and shows the
Silva, A., 2004). However, the incorporation of these morphological features and the synaptic circuits involving
glomeruli with dense-core vesicles in type I is not the central bouton, of primary sensory origin, and the
legitimate in cat and monkey, as in these species they peripheral profiles. C, Central bouton; D, regular dendrite;
V1, presynaptic dendrite; V2, peripheral axonal bouton; G,
have larger central boutons, with more mitochondria glial profile.
and a lighter matrix than in rat. Therefore, for the
sake of simplicity, we propose that in cat and pri-
Silva, A., 1995). Concerning nonnociceptive boutons,
mates, glomeruli can be classified into type I, type II,
many terminate as central boutons of type II synaptic
and peptide-type (because all the central boutons of
glomeruli (e.g., the D-hair afferents), but many are
this later type express neuropeptides see below). In
nonglomerular, including the ones corresponding to
rat both type Ia and type II glomeruli have complex
the termination of G-hair afferents, based on the data
synaptic arrangements in which the central bouton is
obtained in the cat (for review see Maxwell, D. J. and
postsynaptic to presynaptic dendrites and axonal bou-
Rethelyi, M., 1987). Figure 4 shows a diagrammatic
tons and presynaptic to both normal dendrites and
representation of the different types and subtypes of
presynaptic dendrites (see Figure 3 for details). One
main feature of peptide-type synaptic glomeruli (type synaptic glomeruli in rat. There are considerable spe-
Ib in rat) is that their central terminal is seldom post- cies differences in the type of synaptic arrangements of
synaptic to presynaptic dendrites and peripheral axons. primary sensory fibers. For instance, lamina I and LIIo
Of the nociceptive afferents, the nonpeptidergic, are virtually devoid of synaptic glomeruli in rat
IB4-binding afferents terminate mostly as central bou- (Ribeiro-da-Silva, A. and Coimbra, A., 1982), but not
tons of synaptic glomeruli of type I (or more precisely in cat or monkey; in this latter species, there are, in
in rat of subtype Ia), whereas the peptidergic endings laminae I and IIo, numerous type II glomeruli corre-
are glomerular in only a small proportion, particularly sponding to the termination of the HTM myelinated
in rat (Ribeiro-da-Silva, A. et al., 1989); when they are nociceptors (Rethelyi, M. et al., 1982), which are non-
glomerular, the central boutons (subtype Ib) contain glomerular in rat, as well as a considerable number of
dense-core vesicles and peptide immunoreactivity peptide-type glomeruli (Knyihar-Csillik, E. et al.,
(CGRP, together with either SP or SOM; Ribeiro-da- 1982b), which are sparse in rat.
286 Morphological and Neurochemical Organization of the Spinal Dorsal Horn

Type Ia Type Ib Type IIa Type IIb

IIo ++ +

IIidorsal ++++++ ++ +

IIiventral +++ +++++ ++

III ++++ ++++

Afferent Nonpeptidergic Peptidergic A fibers A fibers (?)

innervation C fibers (CGRP + SP D-Hair (?)
(IB4 and P2X3) CGRP + SOM)
Figure 4 Types of synaptic glomeruli and their laminar incidence in laminae IIII of the rat dorsal horn. The drawings on top
were inspired by actual electron micrographs of representative samples of each glomerular type. Note that in rat, lamina I (I) is
virtually devoid of glomeruli and that outer lamina II (LIIo) has only a low number of them. This is a considerable difference
when comparing to the cat and primate, which have glomeruli in significant numbers in lamina I and outer II. In contrast, the
other laminae have abundant synaptic glomeruli. Most type Ia glomeruli are condensed in a rather narrow band (20 mm thick
at cervical levels), which represents the dorsal part of inner lamina II (IIidorsal). Most of the inner lamina II (IIiventral) has a
predominance of type IIa glomeruli, whereas lamina III is normally devoid of type I glomeruli and has similar numbers of both
subtypes of type II glomeruli. The data concerning laminar incidence of glomeruli are semiquantitative and are based on the
data collected from other publications (Ribeiro-da-Silva, A. and Coimbra, A., 1982; Ribeiro-da-Silva, A. et al., 1989). The
information on the types of afferent fibers, which terminate as central boutons of each type of glomerulus, is either based on
evidence obtained in rat (for type I glomeruli) or extrapolated from the data obtained in cat and primate. See text for details on
glomerular classification.

It is important to stress that of the populations of et al., 1982a). Similarly, in rat the HTM myelinated
nociceptive primary sensory fibers, the peptidergic is nociceptors are not glomerular and not postsynaptic
not postsynaptic to GABAergic neurons in the spe- to other structures. The above indicates different
cies studied (Alvarez, F. J. et al., 1993; Ribeiro-da- presynaptic control of the different type of nocicep-
Silva, A., 2004). In contrast, in rat, the IB4-binding tive inputs, as well as likely species differences in
population (which terminates as central boutons of such presynaptic control.
glomeruli of type Ia) is postsynaptic to GABAergic
neurons (Todd, A. J. and Lochhead, V., 1990;
Ribeiro-da-Silva, A., 2004). In cat and primates, the
23.3.3 Receptors on Primary Sensory
HTM myelinated nociceptors that terminate in
Fibers
synaptic glomeruli are also postsynaptic to dorsal
horn neurons at synapses, which have the morpholo- Receptors expressed on primary afferent terminals in
gical properties of inhibitory junctions or in which the spinal dorsal horn include neurotransmitter/
the profiles have been shown to express GABA modulator receptors and neurotrophic factor recep-
(Rethelyi, M. et al., 1982; Alvarez, F. J. et al., 1992). tors. It is important to point out that most of the past
There seem to be species differences though, as a studies of the neurotransmitter/modulator receptors
primate study has shown that the terminals that have on primary afferents have been performed in the
the morphology of the IB4 nonpeptidergic popula- dorsal root ganglia. Information on localization of
tion of the rat did not show them to be involved in receptors on the terminals of primary sensory fibers
axo- or dendroaxonic synapses (Knyihar-Csillik, E. in the spinal cord is recent and still rather limited as it
 Morphological and Neurochemical Organization of the Spinal Dorsal Horn 287

required the development of specific antibodies in situ hybridization studies, which have shown that
against the neurotransmitter/modulator receptors. mRNA for each of the main opioid receptors (, ,
Concerning glutamate receptors, -amino-3- and ) occurs in DRG neurons (Minami, M. et al.,
hydroxy-5-methylisoxazole-4-propionic acid (AMPA) 1995).
receptors have a widespread distribution in DRG neu- There is both immunocytochemical and in situ
rons (Tachibana, M. et al., 1994; Lu, C. R. et al., 2002). It hybridization evidence that the SP receptor, the neu-
has been shown at the light and ultrastructural levels rokinin 1 receptor (NK-1r), occurs in DRG cells and
that N-methyl-D-aspartic acid (NMDA) receptors on sensory fibers in the periphery (Ruocco, I. et al.,
occur in sensory fibers in the dorsal horn (Liu, H. 1997; Li, H. S. and Zhao, Z. Q., 1998). Surprisingly,
et al., 1994b). A presynaptic localization of sensory fibers the NK-1r cannot be detected by immunocytochem-
in the dorsal horn has also been shown for kainate istry on terminals of primary sensory fibers in the
(Hwang, S. J. et al., 2001) and metabotropic glutamate spinal dorsal horn, an observation that suggests that
receptors (Ohishi, H. et al., 1995). It is important to note the NK-1r synthesized in DRG neurons is targeted
that while ionotropic glutamate receptors are typically preferentially toward the periphery. In contrast, the
viewed as excitatory, their activation on primary affer- NK-1r occurs in well-defined populations of dorsal
ent terminals leads to inhibition of transmitter release, horn neurons, as described below.
likely via primary afferent terminal depolarization The capsaicin receptor, TRPV1, previously
(PAD) akin to the action of presynaptic GABAA recep- named VR1 (Caterina, M. J., 2003), occurs in small
tors (Kerchner, G. A. et al., 2001; Lee, C. J. et al., 2002; diameter primary sensory afferents. Unfortunately,
Bardoni, R. et al., 2004). This contrasts with the apparent there is conflicting evidence in the literature regard-
excitatory role that glutamate receptors appear to play ing which afferent populations express the receptor.
at the peripheral endings of primary afferents (Cairns, The in situ hybridization data of Michael G. J. and
B. E. et al., 1998; Svensson, P. et al., 2003). To further Priestley J. V. (1999) concluded that it is expressed by
complicate the matter, PAD appears to have an oppo- the majority of small- and medium-sized dorsal root
site effect on spontaneous transmitter release ganglion neurons, which label with the lectin IB4 or
(facilitation) versus evoked release (inhibition) (for are CGRP-immunoreactive (IR). In contrast, using
review see Engelman, H. S. and MacDermott, A. B., immunocytochemistry Guo A. et al. (1999) concluded
2004). Subunits of the GABAA receptor have also been that TRPV1 occurred mainly in the nonpeptidergic,
identified by in situ hybridization in DRG neurons IB4-binding afferents, and to a minor extent in the
(Persohn, E. et al., 1991), consistent with the functional peptidergic as well. An important observation is that
evidence of a role for these receptors in presynaptic there was a phenotypic switch in the postnatal period
inhibition of primary afferents (for review see leading to the progressive expression of TRPV1 in
Rudomin, P. and Schmidt R. F., 1999). Intriguingly, DRG neurons that bind the lectin IB4, whereas the
however, to date it has not yet been possible to immu- population that colocalized peptides did not change
nocytochemically label GABAA receptor subunits at (Guo, A. et al., 2001). Furthermore, a very significant
axoaxonic synapses in the dorsal horn, although they species difference does occur because in mouse
have been labeled in dorsal horn neurons (Alvarez, F. J. TRPV1 is not expressed by the IB4-binding popula-
et al., 1996; Todd, A. J. et al., 1996). There also appears to tion of afferents but rather by the peptidergic
be a difference in the strength of GABAA receptor- population (Zwick, M. et al., 2002), suggesting that
mediated responses between subtypes of small-dia- the phenotypic switch, which occurs postnatally in
meter afferents (Labrakakis, C. et al., 2003). Subunits of the rat, does not take place in the mouse. This is a
the metabotropic GABAB receptor have been identified particularly important point as it suggests that the
in DRG neurons and, in the dorsal horn, in dendrites results obtained with TRPV1 knockout mice are not
and central boutons of glomeruli (Poorkhalkali, N. et al., transposable to the rat.
2000; Ribeiro-da-Silva and Shigemoto, unpublished The purinergic receptor P2X3 has been detected
observations). in the nonpeptidergic population of nociceptive
Concerning opioid receptors, it is well established afferents (Bradbury, E. J. et al., 1998) and, in agree-
that they occur in primary sensory fibers. The evi- ment with this, in the central boutons of glomeruli of
dence originates from studies using radioactive type I (Llewellyn-Smith, I. J. and Burnstock, G.,
ligand binding, which show that both - and -opioid 1998). Because of the variability with the conditions
receptors in the dorsal horn are reduced following of incubation in a number of afferents that are labeled
dorsal rhizotomy (Fields, H. L. et al., 1980), and from by IB4 binding (see Alvarez, F. J. and Fyffe, R. E.,
288 Morphological and Neurochemical Organization of the Spinal Dorsal Horn

2000, for a discussion), P2X3 immunostaining may the DRG (Djouhri, L. et al., 2003) and by immunocy-
represent a better way of identifying the nonpepti- tochemistry, which shows that Nav1.9 channel
dergic subpopulation of C fibers. immunoreactivity is restricted to laminae III whereas
Concerning cholinergic receptors on primary sen- some Nav1.8 signal was detected in the deeper laminae
sory fibers, both nicotinic and muscarinic receptors as well (Amaya, F. et al., 2000). Regarding small-dia-
have been identified (Flores, C. M. et al., 1996; meter afferents that express these channels, both
Haberberger, R. et al., 1999). Concerning adrenergic channels seem to be expressed equally by peptidergic
receptors, both 1 and 2 receptors have also been and nonpeptidergic (IB4-positive) afferents, although
localized in the DRG neurons (Kinnman, E. et al., such expression was present only in about 50% of
1997; Birder, L. A. and Perl, E. R., 1999; Xie, J. et al., trkA-positive and 50% IB4-positive neurons (Amaya,
2001), and there is also some evidence that they occur F. et al., 2000). However, a recent study in the mouse
in nociceptive primary afferents in the spinal cord indicated that, in this species, the Nav1.8 channel is
(Stone, L. S. et al., 1998). Because -adrenergic recep- expressed by 80% of the IB4-binding population of
tors in the DRG are upregulated after peripheral sensory fibers and only by 20% of the CGRP-IR
nerve lesion, these receptors may be implicated in (Braz, J. M. et al., 2005), which would suggest that
the sensitization of primary afferents in sympatheti- there are significant species differences in the relative
cally maintained pain. expression of these channels. Of the TTX-sensitive
There is abundant literature on the expression of channels, the Nav1.7 channel is also expressed selec-
trophic factor receptors by primary sensory fibers tively in nociceptive, small-diameter afferents,
(for review see Priestley, J. V. et al., 2002). All three although it also occurs in sympathetic neurons, and
high-affinity neurotrophin receptors, trkA, trkB. and seems to play a major role in acute and inflammatory
trkC, have been detected in DRG neurons. Of these, pain (Nassar, M. A. et al., 2004). Interestingly, muta-
trkB and trkC occur in large-diameter neurons, cor- tions that cause loss of function of Nav1.7 were
responding to low-threshold mechanoreceptors, recently found in three consanguineous families from
whereas trkA is found in nociceptive neurons that northern Pakistan characterized by complete inability
express the neuropeptides CGRP and SP. The low- to sense pain (Cox, J. J. et al., 2006).
affinity neurotrophin receptor, p75, has been identi-
fied in the populations of DRG neurons that express
one of the high-affinity neurotrophin receptors. The 23.4 Dorsal Horn Neurons
nonpeptidergic, IB4-binding population, including
its subpopulation that colocalizes CGRP and SOM, The great majority of neurons in each lamina of the
expresses in the adult components of the GDNF dorsal horn are local circuit neurons (interneurons),
receptor family, but not in neurotrophin receptors with axons that remain in the spinal cord. It should be
(Alvarez, F. J. and Fyffe, R. E., 2000). noted that laminae I and V also possess predomi-
nantly interneurons, despite the fact that they are
normally regarded as the origin of major ascending
23.3.4 Expression of Voltage-Gated
nociceptive pathways to the brain. These interneur-
Sodium Channels by Sensory Fibers
ons participate in synaptic circuits, which
In recent years, there has been a developing interest unfortunately are still very incompletely known. To
on voltage-gated sodium channels expressed by pri- complicate matters, there is evidence of considerable
mary sensory fibers, as changes in some have been interspecies differences. These differences affect the
linked to pain states (for review see Wood, J. N. et al., distribution and synaptic connections of primary sen-
2004). Channels with  subunits coded by the Nav1.8 sory fibers as well as the neurotransmitter/
and Nav1.9 genes are sensory neuron-specific and neuropeptide colocalization and the ascending path-
commonly referred to as tetrodotoxin (TTX)-resistant ways. It certainly does not help that most
because higher doses of TTX are required to block immunocytochemical studies on physiologically
them (Amaya, F. et al., 2000). Although the two chan- characterized, intracellularly filled neurons were
nels are mostly colocalized in the same DRG neurons performed in the cat, whereas most of those based
in the rat (Amaya, F. et al., 2000), the Nav1.8 channel on a combination of tract tracing and immunocyto-
also occurs in some larger-diameter afferents, as was chemistry are from the rat and the monkey, and
demonstrated in a study in which the physiological studies on transgenic animals were performed in the
properties of the neurons expressing it was studied in mouse.
 Morphological and Neurochemical Organization of the Spinal Dorsal Horn 289

The primary sensory fibers terminate in the dorsal 23.4.1 Lamina I Neurons
horn in contact with both interneurons and projec-
23.4.1.1 Morphological classification
tion neurons. Unfortunately, our knowledge
of lamina I neurons
concerning the details of the termination of specific
Four morphological types of neurons have been
fiber populations on identified dorsal horn neurons is
described in the rat by Lima D. and Coimbra A.
extremely limited, because a systematic study com-
(1983; 1986): fusiform, flattened, multipolar, and pyr-
bining intracellular recordings and labeling of
amidal (Figure 5). Most of these neurons have their
sensory fibers with labeling of dorsal horn neurons
perikarya and dendritic arborization extending pre-
has never been done for the nociceptive afferents.
dominantly in the rostrocaudal axis, but with some
This lack of information from intracellular techni-
processes extending along the mediolateral axis. For
ques is not a major issue for the SP afferents that
most cells, the dendrites remain within the limits of
innervate neurons that express the NK-1r because
lamina I. However, some specific cell subtypes have
the available antibodies label the NK-1r-IR dorsal
been described with dendrites extending into lami-
horn neurons extremely well, in a Golgi-like manner,
nae II and III (Figure 6); these represent a small
which allows their morphological characterization.
proportion of the total number of cells that may
Because of this, we have rather detailed information
represent local circuit neurons (Yu, X. H. et al.,
on NK-1r-positive neurons innervated directly by
2005), although the possibility remains that they pro-
SP, including their supraspinal projection sites (for
ject to another supraspinal site different from the
recent review see Todd, A. J. and Ribeiro-da-Silva,
thalamus and the parabrachial nucleus (Lima, D.
A., 2005). Unfortunately, the same is not true for the
and Coimbra, A., 1990; Almarestani, L. et al., 2007).
termination of the nonpeptidergic population of
Antidromic recording and retrograde labeling with
nociceptive fibers in the dorsal horn, despite the
horseradish peroxidase or cholera toxin subunit b
fact that their endings can be identified directly on
have demonstrated that lamina I neurons project to
electron micrographs (EM) without the need of any
several supraspinal areas including the thalamus, the
label, because of their unique morphology as type I
periaqueductal gray, and the parabrachial nucleus
glomeruli central boutons (Ribeiro-da-Silva, A.,
(for reviews see Lima, D. and Coimbra, A., 1988;
2004). In fact, it is still unknown if the central boutons
Spike, R. C. et al., 2003; Yu, X. H. et al., 2005). A
of type I glomeruli synapse directly on the projection
comparable classification of lamina I projection neu-
neurons or if there is an interposed interneuron.
rons has been carried out in the cat (Zhang, E. T. et al.,
Recent evidence demonstrates a pathway for non-
1996) and the monkey (Zhang, E. T. and Craig, A. D.,
peptidergic afferents connecting to lamina I neurons
1997; Yu, X. H. et al., 1999) based on the analysis of
via lamina IIi interneurons (Lu, Y. and Perl, E. R.,
horizontal sections (Figure 5). In this latter classifica-
2005) (given that the great majority of the C fibers
tion, the multipolar and flattened types were merged
that innervate lamina IIi are from the nonpeptidergic
into one category (multipolar neurons). Because of
subpopulation; see also below). A challenging alter-
the correlation with classifications done in other spe-
native has been proposed recently in a study using a
cies and correlation with function (see below), we
transgenic mouse endogenously expressing a tracer
have found it more appropriate to classify lamina I
transported transsynaptically in the nonpeptidergic
neurons in rat into the general types of fusiform,
afferents that express the NaV1.8 channel. This study
multipolar, and pyramidal (Yu, X. H. et al., 1999;
has shown that these afferents are connected to
2005; Almarestani, L. et al., 2007) (Figure 6). While
lamina II interneurons that link primarily to lamina
to a large extent these cell types can be recognized in
V projection neurons that, in turn, target fourth-
horizontal sections (Figure 5), complete classification
order neurons in the amygdala, hypothalamus, bed
of these cells require a 3D reconstruction of their
nucleus of the stria terminalis, and globus pallidus
dendritic tree (Figures 5 and 6) (Yu, X. H. et al.,
(Braz, J. M. et al., 2005). The Nav1.8 subclass of non-
1999; 2005).
peptidergic, IB4-binding afferents demonstrated in
mice would thus likely be distinct from the putative
subclass that would connect to lamina I via lamina IIi 23.4.1.2 Correspondence with function
based on the studies by Lu Y. and Perl E. R. (2003; Recent evidence suggests a correspondence between
2005) in the rat. It remains to be determined whether the morphological characteristics of lamina I neurons
this is a species difference and how these pathways and their functional properties (Han, Z. S. et al., 1998).
transpose to other species. Three main functional types of lamina I neurons
290 Morphological and Neurochemical Organization of the Spinal Dorsal Horn

(a)

Fusiform
Pyramidal
R

a
20 m
a

Multipolar
Flattened

a
20 m

20 m

a
20 m
a

(b) (c)

Fusiform Multipolar

Parasagittal

Pyramidal

Horizontal

Unidentified
Transverse

Figure 5 Representative drawings of lamina I neurons in the rat and the monkey spinal dorsal horn. The drawings in (a) are
based on camera lucida reconstructions from Golgi-impregnated cells viewed in the horizontal plane (modified from Lima, D.
and Coimbra, A., 1986). (b) The basic scheme was extended to higher species, such as the monkey, with minor nomenclature
adjustments (modified from Zhang, E. T. and Craig, A. D., 1997; see text). (c) Three-dimensional reconstructions from serial
confocal images of retrogradely labeled lamina I spinothalamic neurons in rats (from Yu, X. H. et al., 2005). Note that lamina I
projection neurons in rat (Yu, X. H. et al., 2005), cat (Zhang, E. T. et al., 1996; Han, Z. S. et al., 1998), and monkey (Zhang, E. T.
and Craig, A. D., 1997) do not appear to have ventrally directed dendrites (see also text) and are thus properly identified only in
the horizontal plane as shown in (c) (modified from Yu, X. H. et al., 2005).

have been identified on the basis of their response to fusiform cells were all NS, pyramidal cells were all
cutaneous sensory inputs: nociceptive-specific (NS) COLD, and multipolar cells were divided between
neurons, responsive only to noxious heat and pinch; HPC and NS (Han, Z. S. et al., 1998). These results
polymodal nociceptive neurons, responsive to nox- are also consistent with previous reports that many
ious heat and pinch as well as to innocuous and pyramidal and multipolar cells possess large axons in
noxious cold (HPC); and innocuous thermoreceptive the cat (Gobel, S., 1978a) and the rat (Lima, D. and
neurons responsive only to innocuous cooling Coimbra, A., 1986). In contrast, fusiform cells are
(COLD) (Craig, A. D. and Kniffki, A. I., 1985; Craig, characterized by thin, likely unmyelinated axons
A. D. and Bushnell, M. C., 1994; Dostrovsky, J. O. and (Lima, D. and Coimbra, A., 1986), consistent with
Craig, A. D., 1996; Han, Z. S. et al., 1998). A corre- the observation that HPC and COLD cells have
spondence between these functional classes of lamina faster antidromic conduction velocities (Craig, A. D.
I and the morphological types (fusiform, pyramidal, and Kniffki, K. D., 1985) than NS neurons (Craig, A.
and multipolar) has been found in the cat whereby D. and Kniffki, K. D., 1985). Recent evidence using
 Morphological and Neurochemical Organization of the Spinal Dorsal Horn 291

Fusiform

Pyramidal

Multipolar

Horizontal

IIo

Parasagittal IIi

IIo

Transverse IIi

Figure 6 Morphological classification of the neuronal populations in lamina I. The classification is mainly based on cell
body shape and primary dendrite orientation. Adaptation of the original classification by Lima D. and Coimbra A. (1986) to
take into account recent findings on projection neurons (see Figure 5 and text) and interneurons (see text). Most cells types
(especially the projection neurons) are best identified in the horizontal plane. Sagittal and transverse views may be useful to
test for ventrally directed dendrites.

whole cell recording in slices indicates the presence innocuous thermal stimuli was of particular interest
of distinct classes of lamina I neurons on the basis of because of the evidence that nonnociceptive neurons
their intrinsic firing properties (Prescott, S. A. and De fail to respond to SP (Henry, J. L., 1976) and do not
Koninck, Y., 2002; Ruscheweyh, R. et al., 2004). In receive significant amount of SP input (De Koninck,
addition, a correspondence between intrinsic electro- Y. et al., 1992; Ma, W. et al., 1996). We therefore
physiological properties and morphology in lamina I decided to test whether pyramidal neurons express
has been shown in rat (Prescott, S. A. and De NK-1r (Yu, X. H. et al., 1999). The results indicated
Koninck, Y., 2002) whereby tonic cells were typically that, while normally one-third of lamina I neurons
fusiform, phasic cells were pyramidal, and delayed are pyramidal in the cervical and lumbar enlarge-
onset and single spike cells were multipolar. In the ments of the spinal cord (Zhang, E. T. et al., 1996;
latter study, the multipolar cell subtype probably Zhang, E. T. and Craig, A. D., 1997; Yu, X. H. et al.,
correspond to the small multipolar subtype with ven- 1999), only 5% of the NK-1r-expressing neurons in
trally directed dendrites and therefore are likely not these areas were pyramidal, most being either fusi-
projection neurons. form or multipolar. This type of differential
distribution gives further support to the concept
23.4.1.3 Expression of NK-1r in that there is a correlation between the functional
subpopulations of lamina I neurons properties of neurons and their morphological fea-
The finding that pyramidal neurons corresponded to tures. In a recent study (Yu, X. H. et al., 2005), we
a class of lamina I neurons that only respond to obtained data in the rat consistent with that in the
292 Morphological and Neurochemical Organization of the Spinal Dorsal Horn

monkey whereby retrogradely labeled lamina I spi- Stalked cells

nothalamic tract (STT) cells were evenly distributed
among the three main cell types (Zhang, E. T. and
Craig, A. D., 1997; Yu, X. H. et al., 1999), and NK-1r
immunoreactivity was also preferentially expressed
in fusiform and multipolar lamina I STT neurons.
A high proportion (6080%), but not all, of lamina Large islet cells
I projection neurons are IR for NK-1r (Marshall, G.
E. et al., 1996; Yu, X. H. et al., 1999; 2005). In contrast,
however, only a subset of NK-1r-IR neurons are
projection neurons (49% after the injection of tracer
in the thalamus; Yu, X. H. et al., 2005). A large pro-
portion of lamina I neurons project to the
parabrachial nucleus. Importantly, although it had
been reported that very few lamina I neurons in rat
project to the thalamus at lumbar levels (Spike, R. C.
et al., 2003), we found that a significant number, about
9%, of lamina I neurons project to the thalamus (Yu,
X. H. et al., 2005), a figure comparable to that found in Small islet cells
primate by us (Yu, X. H. et al., 1999) and others
(Andrew, D. et al., 2003). Our recent data on spino-
parabrachial neurons also indicate that most
pyramidal cells in this subpopulation do not express
NK-1r (Almarestani, L. et al., 2007).

23.4.2 Lamina II Neurons

Figure 7 Representative drawings of lamina II neurons in
Lamina II neurons have been classified originally by the rat spinal dorsal horn. Three main cell types are
Ramon y Cajal (1909) into two main types, the cen- represented in these camera lucida drawings of Golgi-
tral cell, which occurs throughout the lamina, and the impregnated cells: the stalked, small islet, and large islet
limiting cell, which is restricted to a region close to cells (see text for further details). Modified from Todd A. J.
and Spike R. C. (1993).
the laminae III border. These same morphological
types were identified in the cat by Gobel S. (1975;
1978b), and were renamed islet cells and stalked cells, cells described in other species (Todd, A. J. and
respectively. A Golgi method study by Todd A. J. and Lewis, S. G., 1986).
Lewis S. G. (1986) identified cells with the character- Recently, Grudt T. J. and Perl E. R. (2002) per-
istics of stalked and islet cells, as described by Gobel, formed a detailed morphological analysis of the
in lamina II of the rat (Figure 7). Concerning inci- different cells types in lamina II following intracel-
dence of these cells, the stalked cells were shown to lular labeling in spinal cord slices of the hamster.
represent about half of the stained cells in LIIo; in They identified five morphological types of lamina
contrast, islet cells were found throughout lamina II II neurons: islet, central, mediallateral, radial, and
and corresponded to about one-third of the entire vertical (Figure 8). Most of lamina II neurons had
stained neuronal population (Todd, A. J. and Lewis, dendritic arbors that were primarily orientated in the
S. G., 1986). It should be noted that these authors rostrocaudal dimension and relatively flattened in
reported other cell types in lamina II; in fact, about the mediolateral and dorsoventral directions (consis-
half of the cells in the ventralmost part of lamina II tent with Scheibel, M. E. and Scheibel, A. B., 1968).
did not fit in either the stalked or islet cell categories, The islet cells of Grudt T. J. and Perl E. R. (2002)
although they could be subdivided into groups based appear to correspond well with the islet cells of
on their dendritic arborization. The axons of these Gobel S. (1975; 1978b) and the large islet cells of
cells either crossed to lamina III or remained within Todd A. J. and Lewis S. G. (1986) (Figures 7 and 8);
the limits of lamina II. Some of these neurons may the radial cells resembled the star-shape neurons of
correspond to the stellate and laminae IIIII border Bicknell H. R. and Beal J. A. (1984); or the stellate
 Morphological and Neurochemical Organization of the Spinal Dorsal Horn 293

C fiber
A fiber
To brain C1 fiber
C2 fiber

Vertical cell Radial cell

IIo
(stalked)

IIi Central cell Islet cell

(small islet) (large islet)
Excitatory
Inhibitory
Figure 8 Diagrammatic representation of morphological types of neuron in lamina II and their primary afferent input and
interconnections. A correspondence between the classification by Todd A. J. and Spike R. C. (1993) (words in parenthesis)
and that of Grudt T. J. and Perl E. R. (2002) and Lu Y. and Perl E. R. (2003; 2005) is attempted as described in the text. Scheme
for interconnection is based on Lu Y. and Perl E. R. (2003; 2005) (see text for a complete description). Note that distinct
classes of C fibers appear to contact the two classes of islet cells based on the conduction velocity data. Note also the feed-
forward inhibition between the islet cell and the central cell, which is unidirectional (Lu, Y. and Perl, E. R., 2003).

neurons described by Schoenen J. (1982). The central Gobel, S. et al., 1980). These findings concur with
cells were similar to islet cells, but with a much previous studies in the cat by Light A. R. et al.
smaller dendritic expansion in the rostrocaudal (1979). It interesting to note that in the cat, according
direction, thus appearing to correspond to Ramon y to Light A. R. and collaborators (1979), the type of
Cajals central cells and Todds small islet cells response elicited from lamina II cells, either nocicep-
(Figures 7 and 8). The mediallateral cells were the tive or nonnociceptive, had little correlation with
sole type with dendrites extending substantially in their morphology but was related to where the den-
the mediolateral axis. The vertical cells appear to dritic tree arborized. This observation was
correspond to the limitroph neurons of Ramon y interpreted as a consequence of the type of primary
Cajal and stalked cells of Gobel. sensory fibers, which contacted the dendrites of the
Using paired recordings in slices and stimulation cells, as the afferents that terminated in deep lamina
of primary afferents in rat spinal cord slices, Lu Y. II did not seem to be nociceptive in the cat (Light, A.
and Perl E. R. (2003; 2005) were able to describe a R. and Perl, E. R., 1979).
specific connection path between subclasses of
C-fiber afferents and the network of large islet cells,
23.4.3 Laminae IIIVI
small islet cells (central), and stalked cells (vertical),
as summarized in Figure 8. Notably, their results Laminae IIIVI possess both interneurons and pro-
show that distinct subclasses of C fibers connect to jection neurons and have a rather heterogeneous
different laminae II neurons subtypes and that population of cells with dendrites that cut across
stalked cells are indeed feed-forward excitatory these laminae (Brown, A. G., 1982). Cells of some of
cells to lamina I neurons, confirming what had been the major ascending pathways (spinothalamic, spino-
postulated in previous reports (Bennett, G. J. et al., cervical, and dorsal column postsynaptic pathway)
1980; Gobel, S. et al., 1980). have cell bodies located in these laminae (Brown, A.
Concerning islet cells, it has been found that their G., 1982). Lamina III can be considered as a conti-
electrophysiological properties differed depending nuation of the ventralmost part of lamina II, with
on their localization in lamina II, as those situated some small neurons with characteristics similar to
in deep lamina II were nonnociceptive, while those in those of that part of lamina II, as described in one
LIIo were nociceptive (Bennett, G. J. et al., 1980; study (Wall, P. D. et al., 1979). In cat, cells in lamina
294 Morphological and Neurochemical Organization of the Spinal Dorsal Horn

III have been described as nonnociceptive and het- 23.4.4.1 Neurokinins

erogeneous (Maxwell, D. J. et al., 1983) based on All three main mammalian neurokinins (SP, neuro-
intracellular injections of physiologically character- kinin A, and neurokinin B (NKB)) occur in dorsal
ized neurons. However, this concept that all lamina horn neurons. SP-containing cell bodies have been
III cells are nonnociceptive is not correct based on detected in laminae I and II, both with immunocy-
the detection of wide dynamic range cells in this tochemistry (Ljungdahl, A. et al., 1978; Ribeiro-da-
laminae IIIIV of the cat, which possess dorsally Silva, A. et al., 1991) and in situ hybridization
oriented dendrites branching in the superficial lami- (Warden, M. K. and Young, W. S., 1988). It is inter-
nae (De Koninck, Y. et al., 1992; Ribeiro-da-Silva, A. esting to note that most, if not all, SP-IR cell bodies
et al., 1992; Ma, W. et al., 1996; 1997), as well as on the in the rat dorsal horn colocalize enkephalin (ENK)
detection of neurons in the same laminae of the rat, immunoreactivity (Ribeiro-da-Silva, A. et al., 1991),
which express the NK-1r and possess dorsally and they seem to be glutamatergic as well because
oriented dendrites that branch into laminae I and II they express the vesicular glutamate transporter 2
(Liu, H. et al., 1994a; Brown, J. L. et al., 1995; (VGLUT2) (Todd, A. J. et al., 2003). It has been
Littlewood, N. K. et al., 1995; Naim, M. et al., 1997). speculated in the past that some of these neurons
colocalizing SP and enkephalin immunoreactivities
represent stalked cells, based on their laminar locali-
zation in LIIo, absence of presynaptic dendrites, and
23.4.4 Neurochemistry of Dorsal Horn synapses on lamina I neurons (Ribeiro-da-Silva, A.
Neurons et al., 1991). However, neurons colocalizing SP and
enkephalin likely belong to other cell types as well
In spite of a large number of studies that identified
and some of these project to higher levels. In parti-
neurochemicals in dorsal horn neurons, the available
cular, SP-IR lamina I and lamina V cells have
information is less than satisfactory because, with a
been shown to project to the thalamus in rat and
few exceptions, the information on the morphologi-
cat (Battaglia, G. and Rustioni, A., 1992), and it is
cal properties of the neurons is either incomplete or
possible that they also project to other targets such as
simply lacking. This is a consequence of the fact that,
the midbrain parabrachial nucleus. Based on the
in most cases, little more is known than the size and
finding that spinal cord SP-IR neurons normally
the localization of the neurochemically characterized
colocalize enkephalin and glutamate, we can specu-
cell bodies. There are some exceptions, however. late that such projection neurons colocalize all three
The most striking exception is likely the excellent neurochemicals.
characterization of the population that expresses the Virtually all SP-containing neurons in the rat
NK-1r because a Golgi-like staining of the cells is express neurokinin A (NKA) as well (Carter, M. S.
obtained. With most neurochemicals, either the and Krause, J. E., 1990), which means that their dis-
staining is inadequate to allow a proper characteriza- tribution is essentially the same. This applies to the
tion of the cell (e.g., cell body faintly labeled) or the primary sensory fibers that also express SP, which
number of immunolabeled cells is so high that it is represent the main source of SP in the dorsal horn
impossible to characterize individual cells. In the case (see Section Types of Afferents, Neurochemistry,
of in situ hybridization studies, only cell bodies are and Termination in Spinal Cord). Another source of
labeled, and therefore the information obtained with SP and NKA in the dorsal horn are axons originating
the use of this approach is significant but limited. The from cell bodies located in the brain stem (Hokfelt, T.
only approaches that can deliver an integrative view et al., 1978; Gilbert, R. F. et al., 1982); although most
of morphological and neurochemical properties of such fibers terminate in the ventral horn, some ter-
the cells are those that involve the combination of minate in the dorsal horn.
intracellular filling of neurons with immunocyto- NKB derives from a different precursor, and in
chemistry, but they have been sparsely used to contrast to SP and NKA, does not occur in primary
date. Therefore, in the text below little correlation sensory neurons (Ogawa, T. et al., 1985). A light and
to cell populations described above will be made electron microscopic immunocytochemical study
because such data are simply lacking. Table 1 gives using a marker of NKB neurons revealed that the
an overview of classical transmitter/neuropeptide signal occurs in axon terminals in laminae III, with a
colocalizations in the dorsal horn, as described in peak in lamina IIi, and in cell bodies and dendrites
the text. mostly in lamina IIi (McLeod, A. L. et al., 2000).
 Morphological and Neurochemical Organization of the Spinal Dorsal Horn 295

Lamina III showed much less immunolabeling. Signal were sparse in lamina II (Yashpal, K. et al., 1992).
for NKB was detected in dendrites of type I glomer- Curiously, following peripheral nerve lesion, CGRP
uli, suggesting a participation in the modulation of receptors were upregulated in lamina II, indicating
nociception (McLeod, A. L. et al., 2000). Neurons that lamina II cells have the capacity to synthesize
expressing NKB seem to be excitatory and glutama- the receptor (Kar, S. et al., 1994).
tergic (Polgar, E. et al., 2006).
23.4.4.4 Somatostatin
23.4.4.2 Neurokinin receptors The neuropeptide SOM occurs both in primary sen-
Originally, NK-1r immunoreactivity was described sory fibers (see above) and in dorsal horn neurons
in neurons with cell bodies residing in lamina I and in (Hokfelt, T. et al., 1976; Alvarez, F. J. and Priestley, J.
laminae IIIIV, but not in lamina II (Liu, H. et al., V., 1990; Ribeiro-da-Silva, A. and Cuello, A. C.,
1994a; Nakaya, Y. et al., 1994). More recent studies, 1990b). Cells bodies from SOM-IR neurons occur
however, have identified NK-1r immunoreactivity in mainly in lamina II (Alvarez, F. J. and Priestley, J.
some neurons in LIIo (McLeod, A.L. et al., 1998; V., 1990; Ribeiro-da-Silva, A. and Cuello, A. C.,
Ribeiro-da-Silva, A. et al., 2000). Most of the NK- 1990b) and seem to be glutamatergic as they also
1r-IR neurons project to higher levels, having targets express VGLUT2 (Todd, A. J. et al., 2003). The
such as the thalamus (Marshall, G. E. et al., 1996), the receptors for SOM are G protein-coupled and form
parabrachial nucleus (Ding, Y.-Q. et al., 1995; Todd, a family of five receptors (sst1 to sst5) (Dournaud, P.
A. J. et al., 2000), the lateral reticular nucleus, the et al., 2000). The distribution of these receptor sub-
dorsal part of caudal medulla, and, to a minor extent, types has been studied with immunocytochemistry,
the periaqueductal gray (Todd, A. J. et al., 2000). and they have been found in cell bodies and processes
The receptor for neurokinin A (the neurokinin-2 in the superficial dorsal horn (Schulz, S. et al., 1998;
receptor) hardly occurs in the CNS (for review see Von Banchet, G. S. et al., 1999).
Ribeiro-da-Silva, A. et al., 2000). Immunoreactivity
for the neurokinin-2 receptor in the superficial spinal 23.4.4.5 Opioid peptides
dorsal horn was rather weak and restricted to a nar- 23.4.4.5.(i) Enkephalins Although there are two
row band in the lateral part of lamina I, and seemed main enkephalins, met- and leu-enkephalin, no dif-
to be localized in glial cells (Zerari, F. et al., 1998). ferences in distribution have been detected with
This is an unexpected finding that indicates that antibodies generated against each of the two, and as
either neurokinin A acts through another receptor they share a common precursor, in situ hybridization
in the CNS or that it acts mainly in the periphery. studies do not reveal differences in distribution
There is evidence that NKA coreleased with SP from either. Therefore, as in most studies, we refer to
primary sensory fibers acts through the NK-1r and enkephalin immunoreactivity, not discriminating
may be an important component of signaling through whether studies were done with antibodies raised
peptidergic afferent fibers (Trafton, J. A. et al., 2001). against leu- or met-enkephalin. Enkephalin immu-
The preferential receptor for NKB, the neuroki- noreactivity is intense in axons and cell processes in
nin-3 receptor, was identified in cell bodies located in laminae I and II (Hokfelt, T. et al., 1977; Hunt, S. P.
lamina I and, mostly, in lamina II (Ding, Y. Q. et al., et al., 1981) and has also been detected in neuronal
1996; Zerari, F. et al., 1997; Mileusnic, D. et al., 1999; cell bodies in laminae IIII (Del Fiacco, M. and
Ding, Y. Q. et al., 2002). Interestingly, the cell popula- Cuello, A. C., 1980; Hunt, S. P. et al., 1981; Miller
tions that express the neurokinin-3 receptor and Seybold, 1989). In situ hybridization studies have
coexpress either the -opioid receptor or GABA not detected any significant evidence for the pre-
and nitric oxide synthase (NOS) (Ding, Y. Q. et al., sence of the enkephalin precursor signal in DRG
2002). neurons, beyond a few isolated cells (Ruda, M. A.
et al., 1989), and sensory deafferentation does not
23.4.4.3 CGRP and CGRP receptors result in any decrease of enkephalin levels in the
Immunoreactivity for CGRP in the spinal dorsal dorsal horn (Ruda, M. A. et al., 1986). Therefore,
horn is restricted to primary sensory fibers in rodents, almost all the enkephalin in the dorsal horn seems
and likely in others species as well. Concerning to be from local circuit neurons, with a minor com-
CGRP receptors, their distribution has been studied ponent colocalized in axons from serotonergic
with ligand binding and have been shown to occur in neurons of the raphe nuclei that project to the spinal
high density in lamina I and in deeper laminae but cord (Hokfelt, T. et al., 1979); most such axons,
296 Morphological and Neurochemical Organization of the Spinal Dorsal Horn

however, terminate in the ventral horn (Tashiro, T. cells (Cruz, L. and Basbaum, A. I., 1985; Cho, H. J.
et al., 1988; Menetrey, D. and Basbaum, A. I., 1987). A and Basbaum, A. I., 1989). The dynorphins are not
small percentage of enkephalinergic neurons project restricted to local circuits neurons, as some of the
to the brain (Standaert, D. G. et al., 1986). Concerning neurons that express them project to the parabrachial
enkephalin in neurons with cell bodies located in the nucleus (Standaert, D. G. et al., 1986).
dorsal horn, it occurs in two different populations of
neurons: (a) cells that are SP-IR (Tashiro, T. et al., 23.4.4.5.(iii) Endormorphins Of the two endo-
1987; Senba, E. et al., 1988; Ribeiro-da-Silva, A. et al., morphins, endomorphin-2 is restricted to primary
1991) and that likely are glutamatergic (Todd, A. J. sensory fibers, where it is colocalized with SP
et al., 2003); and (b) cells that are GABAergic (Todd, (Martin-Schild, S. et al., 1998; Sanderson Nydahl K.,
A. J. et al., 1992b). An interesting conclusion is that as et al., 2004). In contrast, endomorphin-1 is intrinsic to
all SP-IR neurons in the dorsal horn seem to possess the CNS and occurs in fibers in laminae I and II
enkephalin immunoreactivity, the colocalization of (Martin-Schild, S. et al., 1999).
both peptides can be used as a marker to identify
SP-IR fibers that are intrinsic to the dorsal horn 23.4.4.6 Opioid receptors
(Ribeiro-da-Silva, A., 2004). The ultrastructural stu- Besides occurring on primary sensory fibers (see
dies of SP and enkephalin double labeling have above), opioid receptors are located on dorsal horn
confirmed the colocalization of the two peptides in neurons as well. In situ hybridization cytochemistry
both rat and cat and provided evidence that enke- has confirmed the occurrence of opioid receptors in
phalin-IR boutons or presynaptic dendrites are never dorsal horn neurons (Minami, M. and Satoh, M.,
presynaptic to SP-IR boutons (Ribeiro-da-Silva, A. 1995). The -opioid receptor occurs both on cell
et al., 1991; Ma, W. et al., 1997). In the cat, enkephalin- bodies and dendrites of dorsal horn neurons and in
IR boutons have been shown to contact spinothala- axon terminals (Cheng, P. Y. et al., 1995; 1997); some
mic neurons (Ruda, M. A. et al., 1984) and neurons of of the axon terminals expressing the receptor colo-
the dorsal column postsynaptic pathway (Nishikawa, calize enkephalin (Cheng, P. Y. et al., 1995). In
N. et al., 1983). Our own data on physiologically contrast, the receptor has been localized mostly in
characterized, intracellularly filled dorsal horn neu- dendrites and cell bodies (Arvidsson, U. et al., 1995).
rons, some of which likely projected to the brain, has Immunoreactivity for the  receptor occurs mostly in
shown that these cells had appositions from boutons lamina II (Honda, C. N. and Arvidsson, U., 1995;
colocalizing SP and enkephalin immunoreactivities Kemp, T. et al., 1996), axon terminals, dendritic pro-
and from boutons IR for only one of the peptides, files, and cell bodies (Cheng, P. Y. et al., 1996; 1997).
which likely represented the population colocalizing Most of the cell bodies displaying -opioid receptor
enkephalin and GABA and the population colocaliz- immunoreactivity are located in lamina II and do not
ing SP and CGRP (primary sensory in origin) (Ma, colocalize GABA or glycine immunoreactivities,
W. et al., 1997). which suggests that the neurons that express the 
receptor may be mostly excitatory interneurons
23.4.4.5.(ii) Dynorphins Most of the dynorphin in (Kemp, T. et al., 1996).
the dorsal horn is from spinal cord neurons, with a
minor component at sacral levels from visceral affer- 23.4.4.7 Glutamate
ents (see above). The distribution of the dynorphins It has been known for a long time that a high percen-
in the dorsal horn has been studied both by immu- tage of neurons in the dorsal horn are glutamatergic.
nocytochemistry with antibodies raised against However, the evidence was mostly indirect, as glu-
dynorphin A and dynorphin B as well as by in situ tamate immunostaining is not a reliable marker of
hybridization using probes for the dynorphin precur- glutamatergic cells bodies (Ottersen, O. P. and
sor. The available data indicate that the dynorphins Storm-Mathisen, J., 1984). Currently, the most
occur in axons and cells bodies in laminae I, IIo, and reliable way of identifying glutamatergic dorsal
V, in both rat and cat (Cruz, L. and Basbaum, A. I., horn neurons is the immunocytochemical detection
1985; Miller, K. E. and Seybold, V. S., 1987; Leah, J. of one of the vesicular glutamate transporters.
et al., 1988; Ruda, M. A. et al., 1988; Cho, H. J and However, because the immunoreactivity is located
Basbaum, A. I., 1989). These neurons have been mostly in axonal terminals and is low in dendrites
reported as forming an heterogeneous population, and cell bodies, we have to rely on colocalization
although some neurons in LIIo resembled stalked studies to identify the glutamatergic neurons.
 Morphological and Neurochemical Organization of the Spinal Dorsal Horn 297

Colocalization studies have revealed that most non- 46% of the overall neuronal population (Todd, A. J.
primary sensory axonal varicosities in the dorsal horn and McKenzie, J., 1989; Todd, A. J. and Sullivan, A.
that expressed SP, SOM, or neurotensin were IR for C., 1990). Through a combination of GABA immu-
the vesicular glutamate transporter (VGLUT2), nostaining and the Golgi method, some of the
strongly indicating that these cell populations are GABAergic neurons of lamina II were identified as
glutamatergic (Todd, A. J. et al., 2003). It is also islet cells, but never as stalked cells (Todd, A. J. and
assumed that most of the projection neurons, includ- McKenzie, J., 1989). This observation is consistent
ing the cells of lamina I, are glutamatergic, although with the observation of islet cells with presynaptic
direct immunocytochemical evidence is lacking. dendrites (Gobel, S. et al., 1980; Spike, R. C. and
Systematic studies combining in situ hybridization Todd, A. J., 1992), which indicate an inhibitory func-
for vesicular glutamate transporters are required to tion. Results from Lu Y. and Perl E. R. (2003; 2005)
confirm this point. suggest that only the large islet cells of Todd A. J.
(1986) (Figures 7 and 8) are GABAergic while small
23.4.4.8 Glutamate receptors islet cells (central cell) are glutamatergic. Large islet
Besides occurring in primary sensory fibers (see cells appear to act as feed-forward inhibitory neurons
section Receptors on Primary Sensory Fibers to small islet cells (Lu, Y. and Perl, E. R., 2003)
above), glutamate receptors have also been identified (Figure 8).
in dorsal horn neurons. Concerning the ionotropic In the spinal cord, GABA participates in both
glutamate receptors, their distribution has been stu- presynaptic inhibition on terminals of primary affer-
died with both in situ hybridization and ents and postsynaptic inhibition of dorsal horn
immunocytochemistry. The predominant subunits neurons. Morphological support for the involvement
of AMPA receptors in the dorsal horn are GluR1 of GABAergic interneurons in presynaptic inhibition
and GluR2 (Furuyama, T. et al., 1993; Jakowec, M. was obtained by demonstrating that these are presy-
W. et al., 1995a; 1995b), and the labeling of many cell naptic to primary sensory fibers; this evidence was
bodies has been detected by immunocytochemistry. obtained by means of a combination of GAD immu-
An interesting observation is that the GluR1 subunit nocytochemistry with nerve lesions in the rat
is particularly associated with GABAergic neurons, (Barber, R. P. et al., 1978) or with the intracellular
while the GluR2 is mainly found in cells that lack filling of identified afferent fibers in the cat
GABA (Spike, R. C. et al., 1998). Concerning NMDA (Maxwell, D. J. and Noble, R., 1987). The identifica-
receptors, the NR1 subunit is highly expressed in tion of both presynaptic dendrites and peripheral
dorsal horn neurons (Tolle, T. R. et al., 1993; axons in glomeruli presynaptic to their central bou-
Watanabe, M. et al., 1994) and may be expressed by ton, of known primary sensory origin, provided
all cells. Less is known about the expression of the further morphological support to the participation
NR2 subunit of the NMDA receptor and of kainate of GABAergic dorsal horn neurons in the presynaptic
receptors (Furuyama, T. et al., 1993; Tolle, T. R. et al., inhibition of primary sensory fibers (Todd, A. J.,
1993; Watanabe, M. et al., 1994). Metabotropic gluta- 1996; Ribeiro-da-Silva, A., 2004). In these presynap-
mate receptors are also expressed by dorsal horn tic structures, GABA is colocalized with other
neurons and of these the mGluR5 is the most pre- neurochemicals, such as enkephalin, acetylcholine
valent (Berthele, A. et al., 1999; Jia, H. et al., 1999). (Ribeiro-da-Silva, A., 2004), glycine (Todd, A. J.,
1991; 1996), NPY, or galanin (Polgar, E. et al.,
23.4.4.9 Inhibitory amino acids 1999b). A minor component of the GABAergic term-
23.4.4.9.(i) GABA The distribution of GABAergic inals in the dorsal horn originates from the brain stem
neurons in the dorsal horn was originally studied by (Antal, M. et al., 1996), and a recent in vivo electro-
autoradiography using 3H-GABA high-affinity physiological study provides evidence for descending
uptake and by immunocytochemistry using anti-glu- monosynaptic GABAergic and/or glycinergic inputs
tamic acid decarboxylase (GAD) antibodies (Ribeiro- from the rostral ventromedial medulla to the sub-
da-Silva, A. and Coimbra, A., 1980; Hunt, S. P. et al., stantia gelatinosa (Kato, G. et al., 2006).
1981; Barber, R. P. et al., 1982). Later studies used
immunoreactivity for GABA itself (Todd, A. J. and 23.4.4.9.(ii) Glycine Glycinergic neurons are
McKenzie, J., 1989). A study in rat showed that found in most laminae of the dorsal horn; in laminae
GABAergic neurons were evenly distributed in lami- IIII, their proportion in the overall is considerably
nae IIII, where they represented 2433% or 28 higher in lamina III (30%) than in lamina I (9%) or in
298 Morphological and Neurochemical Organization of the Spinal Dorsal Horn

lamina II (14%) (Todd, A. J. and Sullivan, A. C., Mitchell, K. et al., 1993). One of the glycine receptor
1990). Virtually all glycinergic neurons in laminae subunits (GlyR 3) has been found to be specifically
IIII are also GABAergic (Todd, A. J. and Sullivan, A. expressed in the superficial laminae of the spinal
C., 1990; Todd, A. J., 1991); however, only about half dorsal horn, particularly in lamina II (Harvey, R. J.
of the GABAergic cells colocalize glycine immunor- et al., 2004), although the most abundant subunit in
eactivity (Todd, A. J. and Sullivan, A. C., 1990; Todd, the adult spinal cord is the 1 (Becker, C. M. et al.,
A. J., 1991). Terminals colocalizing GABA and gly- 1988). One interesting feature relative to the distri-
cine immunoreactivities are presynaptic mostly to bution of GABA and glycine receptors in the spinal
dendrites and cell bodies, except in LIIi and deeper cord is that many synapses colocalize GABA and
dorsal horn where they are also presynaptic to pri- glycine, and the specificity of the mixed GABA/
mary sensory fibers in type II but not in type I glycine synapses seems to be determined by the
glomeruli (Todd, A. J., 1996). There is indirect evi- expression, properties, and the subsynaptic localiza-
dence that glycine is likely colocalized with GABA in tion of the GABAA, GABAB, and glycine receptors in
some descending fibers from the brain stem (Kato, G. the target cells (Chery, N. and De Koninck, Y., 1999;
et al., 2006). 2000) and their changes during development (Keller,
A. F. et al., 2001).
23.4.4.10 GABA and glycine receptors It should be kept in mind that the great majority of
GABAA receptors seem to be ubiquitous in dorsal GABA- and glycine-containing terminals are presy-
horn neurons, where their subunit precursors have naptic to spinal neurons, not to primary afferent
been identified by in situ hybridization (Persohn, E. terminals (Todd, A. J. and Spike, R. C., 1993), and
et al., 1991) and their proteins by immunocytochem- GABAergic and glycinergic interneurons control an
istry (Alvarez, F. J. et al., 1996; Bohlhalter, S. et al., important network of polysynaptic connections
1996). The combination of 3, 3, and 2 subunits between primary afferents and dorsal horn neurons.
appears to occur in cells throughout the dorsal horn, For example, blockade of GABAergic and glyciner-
while 1 subunits are expressed by neurons in deeper gic inhibition unmasks low-threshold input to dorsal
laminae (Bohlhalter, S. et al., 1996). An electron horn neurons that do not receive monosynaptic input
microscopic study using an antibody-recognizing from low-threshold afferents (Baba, H. et al., 2003;
subunits 23 of the GABAA receptor revealed Torsney, C. and MacDermott, A. B., 2006). Impaired
predominantly dendrite and cell body staining, with inhibition is thus a likely substrate for cross talk
no direct evidence of labeling of sensory fibers between sensory modalities and appear to underlie
(Alvarez, F. J. et al., 1996). allodynia characteristic of neuropathic pain syn-
A study of the distribution of immunoreactivity dromes (Moore, K. A. et al., 2002; Coull, J. A. et al.,
for the two subunits of the metabotropic GABAB 2003; Coull, J. A. et al., 2005).
receptor, GABABR1 and GABABR2, revealed that
they occurred in highest concentrations in laminae I 23.4.4.11 Other classical transmitters and
and II, where they were detected both in cell bodies other neuropeptides
and in the neuropile (Margeta-Mitrovic, M. et al., One of the better-defined transmitter-specific sys-
1999). The localization of the cell bodies that express tems in the dorsal horn is the cholinergic one. In
the subunits of the GABAB receptor was confirmed fact, neuronal perikarya IR for choline acetyltrans-
by in situ hybridization (Towers, S. et al., 2000). ferase (ChAT) have been identified in the spinal
Interestingly, in the dorsal horn, glycine receptors dorsal horn (Kimura, H. et al., 1981; Barber, R. P.
seem restricted to intrinsic neurons. Glycine recep- et al., 1984; Todd, A. J., 1991). These neurons are
tors are ionotropic receptors made up of a not numerous, occur mainly in laminae IIIIV, and
combination of  and  subunits. The initial studies are presynaptic to primary sensory fibers in synaptic
were performed with antibodies against gephyrin (a glomeruli and to neurons (Ribeiro-da-Silva, A. and
glycine- and GABA receptor-associated protein). Cuello, A. C., 1990a). Interestingly, as mentioned
These studies were performed at the time when it above, most, if not all, of these cholinergic cell bodies
was thought that this protein was associated only and boutons colocalize GABA.
with the glycine receptor. Surprisingly, these studies Regarding other transmitters, spinal serotonin ori-
were rather accurate, as in the spinal cord, gephyrin- ginates from neurons with cell bodies in the brain
IR sites match the distribution of glycine receptors stem (for review see Ruda, M. A. et al., 1986).
more closely than that of GABAA receptors (see e.g. Interestingly, in the cat, these serotonin-IR profiles
 Morphological and Neurochemical Organization of the Spinal Dorsal Horn 299

have been shown to synapse on projection neurons synapses or close to the site of release, diffusion of
(Ruda, M. A., 1986). These fibers colocalizes other neurochemicals over longer distances cannot be
transmitters such as enkephalin (Hokfelt, T. et al., excluded, particularly in the case of neuropeptides.
1979) (see above) or SP and thyrotropin-releasing Nevertheless, it should be kept in mind that the
hormone (TRH) ( Johansson, O. et al., 1981). neuropile represents a significant diffusion barrier,
Another transmitter-specific system originating and it is unlikely that even peptides diffuse large
from the brain stem is the noradrenergic one. distances when released at physiological concentra-
Although their light microscopic distribution and tions. Therefore, a detailed description of the
their origin in the brain stem are well known chemical neuroanatomy of the dorsal horn circuits
(Westlund, K. N. et al., 1983; Fritschy, J. M. and is important to understand sensory processing.
Grzanna, R., 1990), there is still not much informa- As most synapses in the dorsal horn are axoden-
tion about these noradrenergic fibers except that they dritic, it is not surprising that the majority of
are presynaptic to spinal dorsal horn neurons interneurons appear to act on their targets through
(Hagihira, S. et al., 1990; Doyle, C. A. and Maxwell, a postsynaptic mechanism; however, some cells
D. J., 1991). Interestingly, noradrenergic analgesia clearly participate in presynaptic interactions.
appears to involve presynaptic inhibition of gluta- GABAergic axons, which are thought to originate
mate release from 2c-adrenergic terminals of local from local interneurons, establish axoaxonic synapses
excitatory interneurons (Olave, M. J. and Maxwell, on the central boutons of synaptic glomeruli, except
D. J., 2002; 2003a) contacting NK-1r-IR projection for those in peptidergic (type Ib) glomeruli that are
neurons (Olave, M. J. and Maxwell, D. J., 2003b). seldom postsynaptic to other profiles. There is a
Neurotensin immunoreactivity occurs in neurons in fundamental difference between the types of
laminae I and II (Hunt, S. P. et al., 1981; Seybold, V. S. GABAergic neurons that innervate type Ia and type
and Elde, R. P., 1982), which do not colocalize GABA II glomeruli, because boutons containing GABA and
(Todd, A. J. et al., 1992a) but do express VGLUT2, glycine are presynaptic to the central boutons of type
and are likely glutamatergic (Todd, A. J. et al., 2003). II, but not to type Ia glomeruli (Todd, A. J., 1996). In
Furthermore, most of the neurotensin-IR neurons in contrast, neurons colocalizing GABA and enkephalin
LIIi express protein kinase C gamma (Polgar, E. et al., immunoreactivities seem to innervate central axons
1999a). of type I and II glomeruli equally (through both
axoaxonic and dendroaxonic synapses) (Ribeiro-da-
Silva, A., 2004). The population of neurons with
23.5 Identified Neuronal Circuits GABA and ChAT is relatively small (Todd, A. J.,
1991); however, their axons contribute a significant
While our knowledge of synaptic circuits in the contingent of axoaxonic synapses in type II, and to a
dorsal horn remains limited, some patterns are emer- lesser extent in type Ia, glomeruli (Ribeiro-da-Silva, A.
ging based on recent findings. Figure 9 attempts to and Cuello, A. C., 1990a; Ribeiro-da-Silva, A., 2004).
summarize some observations described above. It The neurons with GABA and acetylcholine
should be noted here that there are significant inter- (revealed by ChAT immunoreactivity) have cell
species differences in the neurochemical and bodies in laminae III and IV and dendrites that
anatomical organization of the region. Thus some of extend superficially, where they receive synaptic
the proposed circuits may not hold across all species. input from primary afferents in both type Ia and
One should stress here that, while we propose type II glomeruli. Cells that colocalize GABA and
well-defined circuits, not all chemical communica- NPY appear to selectively target NK-1r-IR laminae
tion involves hard-wired synapses (Zoli, M. et al., III/IV projection neurons, with which they make
1999). Nonsynaptic or paracrine transmission is numerous axodendritic and axosomatic synapses
thought to occur particularly with peptides and (Polgar, E. et al., 1999b).
monoamines; however, it probably occurs for As discussed above, most (if not all) of the
amino acid transmitters also acting both on their SP-containing neurons in the superficial laminae
ionotropic and metabotropic receptors (Rossi, D. are also enkephalin-IR (Ribeiro-da-Silva, A. et al.,
and Hamann, M., 1998; Chery, N. and De 1991). They are located in laminae I and II and
Koninck, Y., 1999; 2000). Whereas under physiolo- receive synaptic input from nonpeptidergic C fibers
gical conditions, most interneuronal communication in type Ia glomeruli and from SP-containing afferents
in the dorsal horn probably takes place either at outside glomeruli. Interestingly, based on evidence
300 Morphological and Neurochemical Organization of the Spinal Dorsal Horn

Cell bodies Boutons

Central gic
Dendrites glomerular d er
pti gic
boutons pe er
C d
+ pti
A gic
Boutons or presynaptic dendrites
- pe er
n on pti
d
C pe
n- +A
no ir
C ha
D-
To brain A

NK-1r
I +Glu SP+ENK
+Glu

IIo

CIb Glu
CIa
GABA+ENK
CIa
or GABA+Gly
IIi GABA
+Gly
Glu GABA
+ENK
CII
GABA
+NPY
CII
GABA
III-V
+ChAT
NK-1r
+Glu
To brain
Glu Glu

Figure 9 Summary diagram of neuronal circuits in the dorsal horn. Different synaptic arrangements involve the different
classes of primary afferents endings in the dorsal horn (glomerular and nonglomerular). In particular, the glomerular structures
are exquisite sites of presynaptic modulation of afferent inputs. Yet, it should be kept in mind that, while this diagram
highlights the different presynaptic arrangements in which the different populations of GABAergic interneurons are involved,
most GABAergic and glycinergic synapses remain axodendritic and axosomatic in the dorsal horn. While physiological
studies indicate that A high-threshold mechanoreceptors (HTM) provide input to laminae I and V, further information on the
synaptic arrangements they are involved in is unavailable due to the lack of specific markers for these types of afferents.
Peptidergic A and C nociceptors are known to contact spinal projection neurons that express the NK-1 receptor both in
laminae I and V. The substance P (SP)enkephalin (ENK)Glu interneuron positioned in lamina IIo represents a prototype
stalked cell, which, for the sake of simplicity, accounts for both the circuitry illustrated in Figure 8 based on the physiological
data (see Lu, Y. and Perl, E. R., 2003; 2005) and that based on what is known of the peptidergic circuitry (Ribeiro-da-Silva, A.
et al., 1991). It remains to be confirmed, however, whether a single type of stalked cell, expressing SP, ENK, and glutamate,
constitutes a common pathway to lamina I projection neurons or whether parallel circuits are involved. Two nonpeptidergic
afferent pathways are illustrated to account for the possibility that parallel circuits with no cross talk at the level of the dorsal
horn may connect to ascending pathways (see text for further details; Braz, J. M. et al., 2005).

from studies in the cat (Ma, W. et al., 1997), these parallel pathway via laminae IIV (Braz, J. M. et al.,
neurons seem to preferentially innervate nociceptive 2005) (see above) that exclude NK-1r-expressing
neurons in lamina I and in the deep dorsal horn, cells, suggest that distinct subclasses of nonpeptider-
which probably express the NK-1r (Figure 9). gic afferents connect to laminae I and V ascending
Finally, these findings, together with those of Lu pathways with virtually no cross talk at the level of
Y. and Perl E. R. (2003; 2005) and the evidence of a the dorsal horn (Figure 9).
 Morphological and Neurochemical Organization of the Spinal Dorsal Horn 301

23.6 Conclusion and Future 1992; Ribeiro-da-Silva, A. et al., 1992; Ma, W. et al.,
Directions 1996; 1997; Lawson, S. N. et al., 1997; 2002).
An important challenge for the future will also be
An important conclusion of this review is that, to better unravel the functional significance of the
despite all the progress, we are still unable to answer multiple transmitter signals originating from the
a few very basic questions regarding the synaptic same cells and the same synaptic terminals. For
organization of the dorsal horn. This is in part a example, the functional meaning of the corelease of
consequence of the heterogeneity of neuronal popu- GABA and glycine (Chery, N. and De Koninck, Y.,
lations in each layer and therefore the difficulty to 1999) two very similar neurotransmitters from the
properly identify subtypes of neurons in functional same terminals remains elusive in a large part.
studies (in contrast with highly organized structures Several synapses also appear to use opposing trans-
like the cerebellum, hippocampus, and even mitters, such as SP and enkephalin (Ribeiro-da-Silva,
neocortex). A. et al., 1991) or GABA and ATP (Jo, Y. H. and
For example, while a significant amount of data on Schlichter, R., 1999); whether these involve local
the organization of inhibitory circuits is available feedback systems, differential release conditions, or
from purely morphological and immunocytochem- nonlinear modes of action (Kupfermann, I., 1991;
ical studies, little is known at the functional level, in Nusbaum, M. P. et al., 2001) remains to be deter-
part because of the difficulty to identify GABAergic mined. Several of these questions continue to be
and glycinergic interneurons for targeted electrophy- elusive simply because the modes of action of several
siological recording. This is likely to change, thanks neuropeptides remain incompletely understood.
to the growing availability of transgenic mice that Target-selective expression of receptors may deter-
express enhanced green fluorescent protein (GFP) in mine the functional meaning of the mixed transmitter
specific subpopulations of inhibitory interneurons signals released from the same neuron. For example,
when GABA and glycine are colocalized in axoaxo-
(e.g., under the control of the promoter for glutamic
nic contacts with primary afferent terminals, only
acid decarboxylase or the glycine transporter 2)
GABAA and GABAB are expressed on those term-
(Hantman, A. W. et al., 2004; Heinke, B. et al., 2004;
inals; the glycinergic signal must only be meaningful
Dougherty, K. J. et al., 2005; Zeilhofer, H. U. et al.,
at other synapses from the same presynaptic neuron
2005). The same is true for subpopulations of inter-
or acting on receptors located on dendrites neighbor-
neurons that are associated with other specific
ing the release site (Rossi, D. and Hamann, M., 1998).
markers (e.g., neuropeptides and calbindin).
In the same line, fine subcellular segregation of
Emerging classification schemes, based on combi-
receptors in the target cells may have very significant
nations of morphological features, transmitter and
impact on how mixed transmitter signals are decoded
receptor phenotypes, and electrophysiological prop-
by the postsynaptic cell (see e.g., Chery, N. and De
erties (Thomson, A. M. et al., 1989; Yoshimura, M. Koninck, Y., 1999; 2000; Keller, A. F. et al., 2001).
and Jessell, T. M. 1989; Lopez-Garcia, J. A. and King, While important information has arisen from the
A. E., 1994; Han, Z. S. et al., 1998; Jo, Y. H. et al., 1998; use of markers of specific primary afferent subclasses,
Yu, X. H. et al., 1999; Grudt, T. J. and Perl, E. R., 2002; it is necessary to develop more refined classification
Prescott, S. A. and De Koninck, Y., 2002; schemes. For example, both on anatomical and phy-
Ruscheweyh, R. and Sandkuhler, J., 2002; siological grounds, the current classification schemes
Ruscheweyh, R. et al., 2004; Hantman, A. W. and of peptidergic and nonpeptidergic C-fiber afferents
Perl, E. R., 2005; Yu, X. H. et al., 2005), will also be (e.g., binding IB4 or expressing CGRP) remains too
critical to better understand the organization of neu- general to account for the existing diversity of sub-
ronal circuits in the dorsal horn. Indeed, studies classes. The search for newer cell surface markers or
combining electrophysiological recording and intra- the development of transgenic models expressing
cellular labeling with immunocytochemistry remain GFP in specific subpopulations of cells will be
critical to obtain a finer identification of neuronal important to unravel the synaptic circuitry and func-
subpopulations and their function as well as to obtain tional role of each subclass of afferents and their
a more complete picture of the afferent and efferent spinal dorsal horn targets.
connections of transmitter-specific interneurons; An important issue to keep in mind that may be an
however, such approaches are technically demanding important limitation in our current understanding is
and remain insufficiently used (De Koninck, Y. et al., the occurrence of interspecies differences. For
302 Morphological and Neurochemical Organization of the Spinal Dorsal Horn

example, many of the original studies on neurons processing as well as of the neural basis of altered
characterized physiologically and labeled intracellu- nociception in chronic pain conditions.
larly with injections of markers have been performed
in the cat, whereas tract-tracing and immunocyto-
chemical studies have generally been carried out in
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 24 Spinal Cord Physiology of Nociception
A R Light, University of Utah, Salt Lake City, UT, USA
S Lee, Korea Institute of Science and Technology, Seoul, Korea
2009 Elsevier Inc. All rights reserved.

24.1 Organization of the Spinal Cord 311

24.2 Ventral Horn: Motor 311
24.3 Dorsal Horn: Sensory 311
24.4 Laminar Organization 311
24.5 Laminae I and II (the Substantia Gelatinosa) 312
24.6 The Adequate Stimulus 312
24.7 Physiology of Neurons in the Dorsal Horn by Laminae 313
24.8 Lamina I 313
24.8.1 Modality of Input to Lamina I 313
24.8.2 Receptive Field Characteristics 314
24.9 Projections and Function of Lamina I Neurons 314
24.10 Lamina II 315
24.11 Outer Lamina II 315
24.11.1 Modality of Input to Outer Lamina II 315
24.11.2 Receptive Field Characteristics 316
24.12 Projections and Function of Outer Lamina II Neurons 316
24.12.1 Inner Lamina II (IIi) 316
24.12.1.1 Projections and function of inner lamina II neurons 320
24.12.2 Lamina III 320
24.12.3 Lamina IV 320
24.12.4 Lamina V 320
24.13 The Ventral Horn 321
24.14 Somatotopic Organization 321
24.15 Physiological Properties of Spinal Neurons 321
24.16 Intrinsic Properties of Spinal Dorsal Horn Neurons 322
24.16.1 Burst Firing Neurons 323
24.16.2 Delayed-Onset Firing Neurons 325
24.16.3 Gap Firing Neurons 325
24.16.4 Phasic Firing Neurons 325
24.16.5 Single-Spike Firing Neurons 325
24.17 Relationship between Intrinsic Firing Patterns and Physiological Types of Dorsal
Horn Neurons 325
24.18 Summary 326
References 327

24.1 Organization of the Spinal structure. The neurons of the spinal cord are clustered
Cord together in a butterflylike arrangement, with the each
of the two wings containing neurons representing or
The spinal cord consists of a wrapping of axons inside controlling muscles and skin in the right or left side of
of which are neurons comprising the first relay for the body, respectively. The more posterior (or dorsal)
sensory information and motoneurons with axons portion of each wing is specialized for sensory function
that connect directly with skeletal muscles. In addition and is called the dorsal horn, while the more anterior
to sensory and motor integration, considerable sympa- (or ventral) portion is specialized for motor function
thetic nervous system integration occurs in this and is called the ventral horn (see Figures 1 and 2).

311
312 Spinal Cord Physiology of Nociception

(Mauderli, A. P. et al., 2000). This chapter will not

consider the ventral horn in detail. The axons sur-
rounding the ventral gray matter are of interest in
the physiology of pain systems, as the axons of many
nociceptive (and non-nociceptive) neurons in the
dorsal horn travel in the white matter of the ventral
horn. Some ascend all the way to the thalamus, or
even hypothalamus.

Figure 1 Cut-away view of lumbar spinal cord. From

Light, A.R. 1992. Pain and Headache (Vol. 12): the Initial 24.3 Dorsal Horn: Sensory
Processing of Pain and its Descending Control: Spinal and
Trigeminal Systems. Karger, with permission.
The dorsal horn consists of the terminations of pri-
mary afferent neurons whose cell bodies lie in the
dorsal root ganglia, and many interneurons with ter-
Posterior (Dorsal)
minations within the spinal cord. The dorsal horn
also contains many transmission neurons with axons
that leave the spinal cord and terminate in the brain-
stem (medulla and pons), thalamus, and
l
hypothalamus.
ll outer
The dorsal horn also receives many inputs from
lll ll inner
Dorsal higher brain centers that can modify, or in some
lV horn cases, completely inhibit transmission from primary
V
Vl afferent neurons to other brain centers, including
the ventral horn.
Ventral
horn

24.4 Laminar Organization

Many scientists have described the organization

Anterior (Ventral) of the dorsal horn as laminar. Rexed B. (1952) divided
the dorsal horn into six layers or laminae defined by
Figure 2 Laminae of the spinal cord dorsal horn.
the sizes and shapes of the cells within them (see
Figure 2). Other defining characteristics of these
laminae are the orientation of axons and dendrites
24.2 Ventral Horn: Motor of the neurons within them, and the amount of
myelin that coats the axons in each layer (Clarke,
The ventral horn contains all of the motoneurons J. L., 1859). The most dorsal, or superficial laminae
whose axons leave the ventral horn in the ventral (laminae I and II) as well as one of the deepest layers
roots and directly innervate skeletal muscles of the (lamina V) are most often associated with nociception
body. However, most of the neurons in the ventral (Light, A. R., 1992). While these laminae were
horn have axons that remain in the spinal cord originally defined using anatomical criteria they
and are, therefore, interneurons and are involved prove to also segregate some of the physiology of
in motor integration. For the purposes of this chap- sensory systems of the spinal cord.
ter, the ventral horn is most important for the
integration of reflexes. The most important of
which are nociceptive reflexes. The most notable 24.5 Laminae I and II
of the nociceptive reflexes is the flexion reflex (the Substantia Gelatinosa)
which is often used as a surrogate for a response to
pain, although the flexion reflex can often be regu- Because of its peculiar appearance in fresh tissue, the
lated in different ways than the perception of pain gelatinous translucent region that caps the dorsal
 Spinal Cord Physiology of Nociception 313

horn has been called the substantia gelatinosa. This 24.7 Physiology of Neurons in
region was defined as lamina II by Rexed B. (1952). the Dorsal Horn by Laminae
Capping the substantia gelatinosa is the even more
superficial lamina I, also called the substantia spon- Being largely sensory in nature, the types of periph-
giosa or marginal zone (Waldeyer, W., 1889; Ramon eral stimuli that activate the intrinsic neurons of the
Y Cajal, S., 1909). These regions have been associated dorsal horn are strongly related to the physiological
with nociception since at least the time of Ranson types of primary afferents that synapse on the spinal
(Ranson, S. W., 1913; 1914; 1915; Ranson, S. W. and neurons. A discussion of the properties of nociceptive
von Hess, C. L., 1915; Ranson, S. W. and Billingsley, primary afferents and their responses to peripheral
P. R., 1916; Ranson, S. W. and Davenport, H. K., stimuli of various types is found in chapter 4.
1931). Ranson made the association between the Integration of these primary afferent inputs occurs
translucent appearance of these regions and the lack to various degrees in the spinal neurons. Some neu-
of myelinated axon terminations in these regions and rons retain the modality selectivity of the primary
the likely termination of unmyelinated axons, which afferents, while others integrate the input of more
even then he associated with pain. than one type of primary afferent.
Fifty years later, lamina I and II were definitively
shown to contain neurons that specifically responded
to noxious (noxious meaning tissue damaging) sti-
muli and others responding to innocuous thermal 24.8 Lamina I
stimuli (Christensen, B. N. and Perl, E. R., 1969; 24.8.1 Modality of Input to Lamina I
1970; Mosso, J. A. and Kruger, L., 1973). Some of
these neurons were shown to project through the Most investigators agree that a substantial portion of
anterolateral tract to the thalamus (Kumazawa, T. lamina I (see Figure 2 for location in spinal cord)
et al., 1971; 1975; Craig, A. D. and Kniffki, K. D., neurons respond specifically or preferentially to var-
ious forms of noxious stimuli (Light, A. R., 1992). In
1985). However, these investigators also showed that
studies using cats, Light A. R. found that 63% of
laminae I and II integrated more than purely noci-
neurons identified as lamina I neurons by intracellu-
ceptive inputs with innocuous thermal neurons
lar labeling were nociceptive while 12% responded
found in lamina I and innocuous mechanical
to innocuous cooling (4%) or warming (cooling and
neurons found in lamina II.
warming neurons are referred to collectively as ther-
moreceptive neurons); 14% responded to innocuous
mechanical stimuli, and 5% responded to both
innocuous mechanical stimuli and noxious stimuli.
24.6 The Adequate Stimulus Of the neurons with nociceptive inputs, 28%
responded only to noxious mechanical stimuli while
Physiological descriptions of nociceptive neurons 35% responded to noxious mechanical stimuli, and
in the spinal cord dorsal horn are often based on also to noxious heat and chemical stimuli. Only some
the adequate stimulus. We define this as the best of these neurons were tested with noxious cold.
(most effective) natural (most likely to occur in an Figure 3 shows some characteristic responses of
environment not manipulated by humans) stimulus lamina I cells.
that excites (evokes action potentials) in a neuron. Andrew D. and Craig A. D. (2002) determined that
For example, while punctate pressure that does not the types of inputs observed for lamina I neurons
cause immediate harm to the skin may occasionally projecting to the thalamus were similar to those
evoke an action potential in a spinal neuron, if that described for unidentified lamina I neurons. Some dif-
neuron fires a train of action potentials at 100 Hz to ferences for identified spinothalamic neurons were a
punctuate pressure that damages the skin, then the disproportionate percentage of neurons with thermal
adequate stimulus is presumed to be noxious inputs in the spinothalamic population. Of 125 lamina I
mechanical stimulation. If this same neuron responds spinothalamic neurons, 61% were nociceptive and
to electrical shock, its adequate stimulus is not pre- 39% were thermoreceptive (most responded to cool-
sumed to be electrical stimulation. Electrical shock is ing, 35%, with the rest responding to innocuous
unlikely to occur in a world without human warming, 4%). Andrew D. and Craig A. D. also found
intervention. that the nociceptive neurons were classifiable into
314 Spinal Cord Physiology of Nociception

(a) nociceptive neurons (see Figure 4). The data presented

in these publications clearly indicate that lamina I
neurons are capable of reliably detecting and transmit-
ting precise quantitative information about noxious
Brush Needle Forceps
pressure to higher centers (Craig, A. D. and Andrew,
(b)
D., 2002). These authors showed the quantitative
responses of lamina I neurons to activation by noxious
heat. The responses correlate very well with human
pain ratings of the same stimuli (see Figure 5).
In addition to nociceptive and thermal inputs,
Warming Noxious heat lamina I neurons also encode stimuli interpreted as
(c)
itch and metabolic signals produced by contracting
muscles.
Craigs laboratory reported recording neurons in
lamina I that specifically respond to stimuli provok-
ing itch (Andrew, D. and Craig, A. D., 2001; see
32 C
Figure 6), and others that responded only to actively
7 C contracting muscles (Wilson, L. B. et al., 2002; see
(d) Figure 7). Some of the neurons responding only to
actively contracting muscles may represent metabor-
eceptors that encode the metabolites produced by
actively contracting muscles, and may be the periph-
Slow brush Slow brush Slow brush
eral receptors that are capable of encoding the signals
Figure 3 Responses of lamina I neurons (a) Neuron
interpreted as muscle fatigue, and/or muscle pain.
activated only by noxious mechanical stimulation of skin. (b) These neurons may also be the sensory arm of sym-
Neuron activated by both noxious mechanical stimuli (not pathetic reflexes known to be activated by actively
shown) and noxious heating of the skin. (c) Neuron activated contracting muscles (Rotto, D. M. and Kaufman,
only by innocuous cooling of the skin. (d) Neurons activated M. P., 1988).
only by innocuous mechanical stimulation of the skin.
This neuron responded only to brushing at less than 10
While the detailed neuronal properties presented
cm s1. Scale bar = 10 s for (a), (b), and (d); 20 s for (c). From here were obtained from recordings in cats, similar
Light, A. R. 1992. Pain and Headache (Vol. 12): the Initial percentages of these various types of neurons in
Processing of Pain and its Descending Control: Spinal and lamina I have been found in monkeys (Kumazawa,
Trigeminal Systems. Karger. Reprinted with permission T. et al., 1975; Kumazawa, T. and Perl, E. R., 1978;
from Light, A. R., Trevino, D. L., and Perl, E. R. 1979.
Morphological features of functionally defined neurons in
Light, A. R. et al., 1979; Price, D. D. et al., 1979; Chung,
the marginal zone and substantia gelatinosa of the spinal J. M. et al., 1986; Craig, A. D., 2004) and rats (Giesler,
dorsal horn. J. Comp. Neurol. 186, 151171. Copyright G. J., Jr. et al., 1976; Menetrey, D. et al., 1977; Light,
1979, Wiley-Liss of John Wiley & Sons, Inc. A. R. and Willcockson, H. H., 1999) although some of
those referenced here report many more neurons
responding to both noxious and non-noxious stimuli
somewhat different groups than those summarized by in the wide-dynamic-range (WDR) fashion. Very
Light A. R. (1992). Of the 61% nociceptive neurons, a recently, similar types of neurons have also been
minority (43%) were described as nociceptive specific. reported in lamina I of the mouse, in an ex vivo
Of the nociceptive-specific neurons, most (80%) were preparation in mouse (Koerber, H. R., personal com-
responsive to both noxious mechanical and noxious munica- tion, 2007).
heat stimuli, with only 20% responsive to only
mechanical stimuli. The majority (57%) of the noci-
24.8.2 Receptive Field Characteristics
ceptive neurons was classified as responding to heat,
pressure, and cold (HPC). These neurons responded The receptive fields of lamina I neurons range from
not only to noxious mechanical pressure, but also quite small (1 mm2) to quite large (1000 mm2)
responded to innocuous cooling temperatures. Craig depending on the body part encoded. The smallest
(Andrew, D. and Craig, A. D., 2002) has also determined receptive fields are found on the digits, while the
the quantitative response properties of lamina I largest are found on the back and hip. The receptive
 Spinal Cord Physiology of Nociception 315

NS neurones HPC neurones

25 25
0.1 mm2

Mean firing rate (imp s1)

20 20 0.5 mm2
5 mm2
15 15

10 10

5 5

0 0

0 20 40 60 80 100 120 0 20 40 60 80 100 120

Stimulus intensity (g) Stimulus intensity (g)
10 g 60 g
25 25 20 g 90 g
Mean firing rate (imp s1)

30 g 120 g
20 20 45 g

15 15

10 10

5 5

0 0

5 0.5 0.1 5 0.5 0.1

Probe size (mm2) Probe size (mm2)
Figure 4 Quantitative responses of lamina I neurons Mean population stimulusresponse curves to graded mechanical
stimulation (10120 g) for NS (n 20) and HPC (n 19) neurones (top row). The same data replotted with respect to toprobe
size are shown in the bottom row. NS, nociceptive-specific neurons; HPC, heat, pressure, and cold. From Andrew, D. and
Craig, A. D. 2002. Quantitative responses of spinothalamic lamina I neurones to graded mechanical stimulation in the cat. J.
Physiol. 545, 913931, reprinted with permission.

field sizes and pattern indicates considerable conver- 24.9 Projections and Function
gence from primary afferent neurons. Thus, the of Lamina I Neurons
smallest excitatory receptive fields for lamina I neu-
rons can approximate the size of C-primary afferent While many of the neurons in lamina I most likely
polymodal nociceptors (<1 mm). The largest excita- function as integrative interneurons within lamina I,
tory receptive fields, however, are many times the a large proportion of these neurons project to other
size of the largest primary afferent receptive field. spinal cord and brain regions.
Lamina I neurons with A-fiber nociceptive input do Many of the neurons in lamina I have been shown
not have the spotlike receptive fields of the primary to project to higher brain centers including the tha-
afferents, but have a more uniform receptive field, lamus, the parabrachial region, the medullary
indicating convergence of several A-nociceptors onto reticular formation, and even the hypothalamus. A
the lamina I neurons. much larger proportion of lamina I cells have been
Excitatory receptive fields often have an inhibi- shown to project to higher brain centers than neurons
tory surround region, stimulation of which with located in laminae II-IV. Craig A. D. (1996; 2003) has
noxious stimuli causes inhibitory post synaptic also found lamina I projection neurons responding
potentials (IPSPs)in the recorded neuron and can specifically to itch inputs, and others responding
inhibit activation induced by stimulation of the specifically to ergoreception (inputs from actively
more central excitatory receptive field (see Figure 8 contracting muscles; see above). He has attempted
for explanation of how these receptive fields are to generalize these inputs as homeostasis inputs, that
formed). is, inputs necessary to maintain body integrity.
316 Spinal Cord Physiology of Nociception

90
3 s ISI, 53 24.11 Outer Lamina II
Second pain
80
70 First pain
24.11.1 Modality of Input to Outer
Lamina II
Verbal rating

60
50
40
The outer region contains mostly nociceptive neu-
30 rons while the inner region contains mostly
20 innocuous mechanically responsive neurons (see
10 Figure 9 for recordings of common responses). This
0
0 5 10 15 20 relationship has been shown in cats, monkeys, and rats
Stimulus number (Kumazawa, T. and Perl, E. R., 1978; Bennett, G. J.
2 s ISI, 58 C
et al., 1979; Light, A. R. et al., 1979; Bennett, G. J.
14 et al., 1980; Light, A. R. et al., 1981; Light, A. R. and
12 HPC Kavookjian, A. M., 1984; Steedman, W. M. et al.,
10 NS
Frequency

1985; Light, A. R. and Willcockson, H. H., 1999).

8
6
In a summary of data from experiments in the
4 Light and Perl laboratory (Light, A. R., 1992), 70%
2 of the neurons identified as having cell bodies in
0 lamina IIo were nociceptive. Forty-eight per cent
0 2 4 6 8 10 12 14 16 responded only to mechanical nociceptive inputs,
Stimulus number while 32% responded to at least noxious heat and
Figure 5 Comparison of the mean response curves of noxious mechanical inputs, and many that were
heat, pressure, and cold (HPC) and nociceptive-specific tested, also responded to noxious chemicals, making
(NS) lamina I spinothalamic tract cells (bottom) to the them polymodal nociceptive inputs. Of the remain-
maximal repeated brief contact heat stimulus trial with the der, 20% responded to both innocuous and noxious
mean report of human subjects to the maximal trial in the
study of Vierck et al. (1997) (top). In the top panel, 53
mechanical inputs and 5% each were thermorecep-
DEG = 53 C; ISI = Interstimulus interval. From Craig, A. D. tive (most to cooling) and 5% responded only to
and Andrew, D. 2002. Responses of spinothalamic lamina I innocuous mechanoreceptive stimuli.
neurons to repeated brief contact heat stimulation in the cat. The neurons in the outer region of lamina II have
J. Neurophysiol. 87, 19021914, reprinted with permission. small receptive fields, and neurons can receive only
noxious mechanical inputs, only noxious heat or cold
inputs, and mixtures of mechanical and heat, and
Thus, one of the major functions of lamina I is to noxious chemical inputs. Summarizing data from several
convey information about nociceptive inputs, itch, publications (Light, A. R. et al., 1979; Light, A. R.
and thermal inputs, to higher brain centers . Lamina and Kavookjian, A. M., 1984; Light, A. R. and
I also integrates and relays inputs necessary for sym- Kavookjian, A. M., 1988; Light, A. R. and
pathetic functions. Craig has grouped these functions Willcockson, H. H., 1999), some 70% of lamina
as homeostatic and suggests the view that lamina I is IIo neurons are nociceptive, 5% are thermorecep-
critical for relatively objective information about tive, 5% are responsive to innocuous mechanical
body integrity necessary to maintain homeostasis, stimuli, and 20% respond to both innocuous
and thus, life (Craig, A. D., 1996; 2003). mechanical stimuli and nociceptive stimuli.
Similar to lamina I neurons, some of the nocicep-
tive neurons respond selectively to noxious
mechanical inputs, with perhaps more of these
24.10 Lamina II than other types. Other nociceptive neurons
respond to noxious heat as well as to noxious
Almost 30 years after the first definitive recordings mechanical stimuli.
were made from the substantia gelatinosa, the func-
tion of this region is still not understood. Recordings
combined with intracellular labeling suggested that
24.11.2 Receptive Field Characteristics
the anatomically observable division of lamina II into
an inner and outer region had an apparent functional The receptive fields of lamina IIo neurons tend to
correlation. be smaller than those of the overlying lamina I
 Spinal Cord Physiology of Nociception 317

(a)

10 Hz

Histamine + 1 mA DC 30 s
3 min
(b)
30
impulses per 20 s
Mean number of

20

10 Histamine

0
0 5 10 15 20 25
Time (min)

(c)
30
impulses per 20 s
Mean number of

20

10
Vehicle

0
0 5 10 15 20 25
Time (min)
Figure 6 Histamine- and vehicle-evoked responses from histamine-sensitive lamina I spinothalamic tract neurons. (a) The
response of a single neuron to histamine. Top histogram, binned firing rate of the neuron (1-s bins). The middle trace shows
the analog record of neuronal activity; the thickening of the baseline in this record during the iontophoresis (indicated by the
lower trace) is due to the current-evoked activation of several neighboring neurons. (b) Mean response of all 10 histamine-
sensitive lamina I spinothalamic tract neurons to histamine application. (c) Mean responses of the same 10 neurons to vehicle
application. Error bars, 1 standard deviation; bin size, 20 s; DC, direct current. With permission from Andrew, D. and Craig,
A. D. 2001. Spinothalamic lamina I neurons selectively sensitive to histamine: a central neural pathway for itch. Nat. Neurosci.
4, 7277.

neurons (Price, D. D. et al., 1979). This may be due also have fewer complex receptive fields, some-
to the less extensive mediolateral spread of den- times lacking an inhibitory surround region.
drites of lamina II neurons. Lamina II neurons may These two observations, along with some
318 Spinal Cord Physiology of Nociception

(a) 1.3
kg
0

5 Hz

110
mm Hg
68

10 s
(b) 1.4
kg

0.3

5 Hz

10 s
(c) 2.7
kg
0.5

5 Hz

10 s
Figure 7 Examples of the variety of responses of lamina I neurons that were excited by muscle contraction. The traces show
from the top downward: the tension developed in the triceps surae muscle, the discharge of the single neuron histogrammed in 1 s
bins and either mean blood pressure (a) or the neural recording ((b) and (c)). (a) Response from a heat, pressure, and cold (HPC)
lamina I neuron with unidentified projections that was excited by contraction that paralleled the tension developed in the muscle
and the centrally evoked cardiovascular reflex. (b) Response of another HPC lamina I neuron with unidentified projections that was
only excited after the contraction. (c) Response of a spinobulbar lamina I neuron (central conduction velocity 2.2 m s1) that was
excited during and after the contraction; this neuron had no identifiable cutaneous receptive field but responded to stretching the
triceps muscle and also to rhythmic contractions at 5 Hz. From Wilson, L. B., Andrew, D., and Craig, A. D. 2002. Activation of
spinobulbar lamina I neurons by static muscle contraction. J. Neurophysiol. 87, 16411645, reprinted with permission.
 Spinal Cord Physiology of Nociception 319

(a) 25 mV
(a) Slow brush
0.5 s
(b)

(b) Pinch

(c) Cooling 25 mV
1s
(c)

Figure 9 Low-gain, direct-current patch-clamp

recordings in vivo from the most common types of
responsive neurons encountered in the substantia
gelatinosa of rats. Examples of slow brush (a), pinch (b), and
innocuous cooling (c) (ethyl chloride spray applied to brush
and brushed on skin) responsive neurons. From Light, A. R.
and Willcockson, H. H. 1999. Spinal laminae III neurons in
(d) rat recorded in vivo in whole cell, tight seal configuration:
properties and opioid responses. J. Neurophysiol. 82,
33163326, reprinted with permission.

Figure 8 Diagram of mechanisms of integration of

nociceptive inputs in lamina I neurons. (a) Indicates the
receptive field of a single primary afferent nociceptor with an A
anatomical findings suggest that at least some
axon. Each spot represents a point at which noxious lamina II neurons project inputs to overlying
mechanical stimulation evokes a discharge. (b) Left panel lamina I neurons.
indicates the receptive fields of three mechanical nociceptors
with overlapping receptive fields. The three primary afferents
terminate on the same lamina I neuron in the middle panel. The
right panel indicates the resulting receptive field. The receptive 24.12 Projections and Function of
field is somewhat larger, and more responsive points are found Outer Lamina II Neurons
within the responsive area. Thus, unlike primary afferent
receptors, spinal nociceptive neurons have nearly continuous In addition to projections to lamina I, lamina IIo
receptive fields with less unresponsive regions between
neurons have been shown to project to deeper
responsive spots. (c) Left panel indicates the receptive fields of
three nonoverlapping primary afferent mechanical laminae, including laminae III, IV, and V (Light, A.
nociceptors. The middle panel indicates that the lamina I R. and Kavookjian, A. M., 1988; Eckert, W. A., III,
neuron receives excitatory input from the axons whose et al., 2003). Substantial intralaminar projects have
receptive field is indicated with open circles. The other two also been noted for many years (Szentagothai, J.,
axons excite inhibitory neurons (this is a simplification), and in
1964; Matsushita, M., 1969; Rethelyi, M. and
all cases, both excitatory and inhibitory neurons are excited by
all primary afferent neurons). The result is that the receptive Szentagothai, J., 1969; Rethelyi, M. et al., 1981;
field of the lamina I neuron, shown at right, consists of inhibitory 1983; 1989). Both inhibitory and excitatory intrala-
inputs (shown with filled symbols) and excitatory inputs (shown minar, as well as extralaminar connections, are
with open symbols). (d) This receptive field is typical for spinal possible (Lu, Y. and Perl, E. R., 2005). A few neurons
cord nociceptive neurons and indicates both of the processes
in lamina II have also been shown to project long
described in (a)(c). The central crosshatched region is the
excitatory receptive field in which noxious mechanical connections to the thalamus (Giesler, G. J., Jr. et al.,
stimulation causes excitation of the spinal neuron. Noxious 1978; Willis, W. D. et al., 1978). Whether these
mechanical stimulation of the region outside the excitatory projections represent ectopic lamina I neurons, or
receptive field causes inhibition and is indicated by minus more projections of this variety have yet to be
signs. This inhibitory field fades out gradually as the distance
detected is unknown.
from the excitatory field increases. Adapted from Light, A. R.
1992. Pain and Headache (Vol. 12): the Initial Processing of Thus, lamina IIo neurons may function both as
Pain and its Descending Control: Spinal and Trigeminal inhibitory neurons helping to create the inhibition
Systems. Karger, reprinted with permission. surround receptive fields described above, and as
320 Spinal Cord Physiology of Nociception

excitatory relay neurons for convergence of receptive predicted by the terminations of single group IV
fields and possible amplification of the excitatory muscle afferents in this layer (Ling, L. J. et al., 2003).
signal (see Figure 8). Potentially then, nociceptive neurons, and possibly,
metaboreceptive neurons should be found in inner
lamina II. Whether these neurons represent special
types of nociceptive inputs (such as inputs sensitive
24.12.1 Inner Lamina II (IIi)
to inflammation) or have nociceptive inputs (such as
The inner division of lamina II has many neurons muscle nociceptive inputs) that have not been found in
that respond to innocuous mechanical stimuli (Light, recording studies thus far remains to be examined.
A. R. et al., 1979; Price, D. D. et al., 1979; Woolf, C. J.
and Fitzgerald, M., 1983; Light, A. R. and Kavookjian, 24.12.1.1 Projections and function of
A. M., 1988; Light, A. R. and Willcockson, H. H., inner lamina II neurons
1999). A detailed analysis of the types of inputs to Like other neurons in lamina II, lamina IIi has sub-
this lamina has not been made. One survey (Light, A. R., stantial axonal projections to other regions of lamina
1992) found that between 50% and 80% of the II (Szentagothai, J., 1964; Matsushita, M., 1969;
neurons responded only to innocuous mechanical Rethelyi, M. and Szentagothai, J., 1969; Rethelyi, M.
stimuli. A very common response was a preferential et al., 1981; 1983; 1989). In addition, at least some of
response to a very slowly moving stimulus (e.g., see these neurons project axons to deeper laminae, where
Figure 9(a)). The response to mechanical stimula- they synapse with other spinal cord neurons (Light,
tion often adapts with very few repetitions of the A. R. and Kavookjian, A. M., 1988; Eckert, W. A., III,
stimulus, sometimes after only a single stimulation. et al., 2003). Basbaums laboratory has also recently
Responses of this type do not relate directly to any proposed that neurons in this region project to lamina
known type of primary afferent input. Intracellular V neurons which subsequently project to higher
analysis of these response properties identified a brain regions, not usually considered in nociceptive
prominent IPSP evoked by innocuous mechanical processing (Braz, J. M. et al., 2005).
stimulation that was capable of inhibiting nearly all The innocuous responses of lamina IIi neurons
action potential generation (Light, A. R. et al., 1979). seems most related to sensuous touch (Olausson, H.
Neurons with these types of inputs seem to domi- et al., 2002) or tickle. As these neurons have not been
nate this sublamina. shown to project in large numbers to the thalamus or
However, in addition to innocuous mechanical other brain regions rostral to the spinal cord, it is
inputs, considerable circumstantial evidence suggests unknown how these inputs are relayed to higher
that nociceptive inputs should also reach inner brain centers. However, it is interesting that some
lamina II. First, a few neurons in this layer have have noted that tickle is abolished following lesions
been found to be activated preferentially by nocicep- of the anterolateral tract (Lahuerta, J. et al., 1990).
tive stimuli. Second, primary afferents labeling with
isolectin B4 (IB4) terminate heavily in inner lamina
24.12.2 Lamina III
II. At least some of the IB4-positive afferents have
been identified as nociceptive, and eliminating IB4- Lamina III mostly contains neurons that receive only
containing primary afferents reduced inflammatory innocuous mechanical inputs (Pomeranz, B. et al., 1968;
pain in rats. vanilloid receptor like 1 (VRL-1 also Price, D. D. and Mayer, D. J., 1974; Wall, P. D. et al.,
called TRPV2)also labels inner lamina II suggesting 1979; Light, A. R. and Durkovic, R. G., 1984). However,
that a population of myelinated nociceptor may at least some nociceptive neurons can be found here.
selectively terminate in this region (Lewinter, R. D. Nociceptive neurons in lamina III may respond only to
et al., 2004). Neurons in lamina IIi show increases in noxious mechanical stimuli, or to noxious thermal and
protein kinase C (PKC) following inflammation chemical stimuli as well as to noxious mechanical
which results in allodynia (Martin, W. J. et al., 1999; stimuli (Brown, A. G. and Fyffe, R. E., 1981; Maxwell,
2001). At least a few nociceptive primary afferent D. J. et al., 1983; Maxwell, D. J. and Koerber, H. R.,
fibers have shown extensive terminations in lamina 1987). Most investigators have reported that neurons
IIi (Sugiura, Y. et al., 1986; Koerber H. R. et al., that do respond to noxious stimuli, also respond to
personal communication, 2004). Finally, either mus- innocuous stimuli, albeit with much lower action
cle nociceptive input to laminae IIi cells, and/or potential frequency. The importance of lamina III in
muscle metaboreceptor input to this layer is nociception is unknown. In cats, many lamina III
 Spinal Cord Physiology of Nociception 321

neurons project to the lateral cervical nucleus. In rats This is partly due to the relative ease of recording the
and primates, at least some of these neurons project to larger neurons located in this layer, and partly due to
the dorsal column nuclei (Rustioni, A. et al., 1979; its known large projection to the thalamus, making it a
Giesler, G. J., Jr. et al., 1984). As with lamina IIi, large contributor to the spinothalamic tract and this
PKC is upregulated in lamina III neurons following tract has long been implicated as a pain transmission
nerve injury and inflammation (Polgar, E. et al., 1999; pathway.
Miletic, V. et al., 2000), suggesting at least inflammatory Lamina V contains many neurons that respond to
inputs, if not nociceptive inputs to this lamina. both noxious and innocuous stimuli, and also neurons
that respond selectively to noxious stimuli. Innocuous
24.12.3 Lamina IV mechanoreceptive neurons are also found in this lamina
(Mendell, L. M. and Wall, P. D., 1965; Hillman, P. and
Lamina IV neurons are also dominated by responses to Wall, P. D., 1969; Light, A. R. and Durkovic, R. G.,
innocuous mechanical stimuli (Armett, C. J. et al., 1962; 1984).
Pomeranz, B. et al., 1968; Price, D. D. and Browe, A. C., The receptive fields of lamina V neurons tend to be
1973). Many other neurons, however, respond more larger than neurons with similar modalities in lamina
vigorously to noxious stimuli (both mechanical and IIV. A common feature of the receptive fields, like
thermal) than to innocuous stimuli (the WDR those in lamina I, is a zone of inhibitory influence
response). In cats, for example, Light A. R. and surrounding the excitatory receptive field (inhibition
Durkovic R. G. (1984) found that 56% of the recorded surround). Typically, the most effective stimulus for
neurons responded only to innocuous mechanical sti- eliciting inhibition is the same as that of the excitatory
muli, while 37% responded to innocuous stimuli, but region (Hillman, P. and Wall, P. D., 1969).
increased their response rate to noxious stimuli. Only The response characteristics of at least some
6% of the neurons responded only to noxious stimuli. lamina V neurons correlate well with human esti-
In cats, a substantial number of lamina IV neurons mates of pain intensity and with reflex response of
project to the lateral cervical nucleus and are, there- animals to noxious stimuli (Dubner, R. et al., 1977;
fore, spinocervical neurons (Brown, A. G., 1968; Brown, 1980; Casey, K. L., 1982; Dubner, R. et al., 1989; e.g.,
A. G. and Franz, D. N., 1969; Bryan, R. N. et al., 1973;
see figure 10). This, along with the fact that many
1974). At least some of these neurons encode some
lamina V neurons project in the anterolateral tract to
aspects of noxious stimuli. The proportion of spinocer-
the thalamus, both of which are implicated in pain
vical neurons in primates appears to be less than in cats
transmission, indicate that lamina V neurons are
(Bryan, R. N. et al., 1974). Again some of these neurons
important in at least some aspects of pain perception.
encode noxious stimuli (Brown, A. G., 1968; Brown, A.
The likely contribution of lamina V neurons to the
G. and Franz, D. N., 1969; Bryan, R. N. et al., 1973;
flexion reflex pathway probably contributes to the
1974; Kajander, K. C. and Giesler, G. J., Jr., 1987).
observations that lamina V neuron responses are
Specific functions for noxious inputs relayed via the
spinocervical tract are unknown, but may represent an similar to nociceptive reflexes (Light, A. R. and
alternative pathway for pain transmission, separate Durkovic, R. G., 1984; Morgan, M. M., 1998).
from the spinothalamic tract (Willis, W. D. and The presence of innocuous activation of many
Westlund, K. N., 1997). Finally, at least some lamina lamina V neurons in addition to nociceptive activation
IV neurons project to the thalamus (Trevino, D. L. requires additional mechanisms to allow discrimina-
et al., 1973; Albe-Fessard, D. et al., 1974; Trevino, D. tion between innocuous and noxious stimuli.
L. et al., 1974; Willis, W. D. et al., 1974; Price, D. D. and
Mayer, D. J., 1975).

24.13 The Ventral Horn

24.12.4 Lamina V
A very large literature exists on the nociceptive prop- The ventral horn contains many neurons that project
erties of neurons in lamina V (Wall, P. D., 1967; to the thalamus. However, the properties of these
Mendell, L. M. and Wall, P. D., 1965; Hillman, P. spinal neurons suggest a role in proprioception,
and Wall, P. D., 1969; Wall, P. D., 1973; Price, D. D. rather than in nociceptive processing (Menetrey, D.
and Dubner, R., 1977; Dubner, R., 1978; Dubner, R. et al., 1984; Ammons, W. S., 1987; Craig, A. D., Jr. et al.,
and Bennett, G. J., 1983; Willis, W. D., 1985; 1988). 1989).
322 Spinal Cord Physiology of Nociception

(a) Behavior (b) WDR1 (N = 6) *

1.6 80
*
*
*
Detection speed (1/sec)

Peak freq - BKGRD

* 60
1.2

40 *
0.8

T1 = 45C 20
T1 = 45C
T1 = 46C
T1 = 46C
0.4 0
0.2 0.4 0.6 0.8 0.2 0.4 0.6 0.8
T2 (C) T2 (C)

(c) WDR2 (N = 6) (d) NS (N = 5)

80 T1 = 45C 80 T1 = 45C
T1 = 46C T1 = 46C
Peak freq - BKGRD

Peak freq - BKGRD

60 60

40 * 40

20 20 *

0 0
0.2 0.4 0.6 0.8 0.2 0.4 0.6 0.8
T2 (C) T2 (C)
Figure 10 (a) Detection speeds in response to 0.4, 0.6, and 0.8  C T2 stimuli from T1s of 45 and 46  C while recording from
wide-dynamic range (WDR)1, WDR2, and nociceptive-specific (NS) neurons. There was a significant increase in detection
speed for each T2 temperature when Tl was 46  C (P < 0.01). Each point represents the average of four trials from each of 17
neurons. (b)(d) Responses of WDRl (b), WDR2(c), and NS (d) neurons to the same stimuli as in (a). Asterisks indicate
significant increases (P < 0.01) in neuronal discharge when Tl was 46  C. Each point represents the average of four trials per
neuron. From Dubner, R., Kenshalo, D.R., Jr., Maixner, W., Bushnell, M. C., and Oliveras, J. L. 1989. The correlation of
monkey medullary dorsal horn neuronal activity and the perceived intensity of noxious heat stimuli. J. Neurophysiol. 62, 450
457, reprinted with permission.

24.14 Somatotopic Organization the segmental body plan. However, the map is dis-
torted in the regions that encode the highly
Submodality is encoded by neurons in different innervated digits and feet with a much greater
laminae. However, spatial encoding of the body sur- representation of these body regions than the neck
face is encoded in the medial-lateral and rostral- and trunk regions. The distortions in this map also
caudal planes (Brown, P. B. and Fuchs, J. L., 1975; represent the density of primary afferent innervation
Koerber, H. R. and Brown, P. B., 1977; Light, A. R. of the skin, with digits being much more densely
and Durkovic, R. G., 1984; Ritz, L. A. et al., 1985; innervated than proximal limb regions and trunk
Brown, P. B. et al., 1997; 2005). The overall map of regions. These distortions are correlated to nocicep-
the body surface is similar in all laminae. Figures 11 tive discrimination of these same body regions with
and 12 show the overall somatotopic map and more greater discrimination possible at the tips of the
detail of the lumbar region. Overall, the map follows digits, and less on the trunk.
 Spinal Cord Physiology of Nociception 323

Cervical Thoracic Lumbar Sacral

7 8
5 6Shoulder 1 2 3 5 1
2
3
4 4 5 6 7 8 9 10 11 12 1 2 3 4 Hip 3 4
1 2 Ankle
Dorsal trunk Foot
1 234 5

Ar m t
Wris
Ventral trunk
1 2 3 45

Figure 11 Somatotopic map of representation of body surface in primal spinal cord gray matter. Cervical, thoracic, lumber, and
sacral segments are indicated. Maps of this sort are somewhat misleading in that receptive fields often encompass more than one
of the body regions indicated on the map. For example, in the lumbar cord, receptive fields notes as hip may be restricted to the
hip, but others may include a strip that includes the entire leg including the toes as well as the hip. From Light, A. R. 1992. Pain and
Headache (Vol. 12): the Initial Processing of Pain and its Descending Control: Spinal and Trigeminal Systems. Karger, reprinted
with permission.

Lateral attaining recordings from the cell bodies. Because

access is easier, whole-cell patch clamp of the spinal
Hip Hip cord neurons can be readily accomplished in slice
preparations. Recordings in this mode are more
Medial ankle Lateral ankle
stable, details of synaptic inputs more easily
Foot
kle
observed, and membrane currents rather than voltage
An changes can be measured.
The major disadvantage, however, is that except
ot

Digit Digit Digit Digit

Fo

2 3 4 5 for very recent recordings (Koerber, personal com-

munication, 2006) the inputs to the spinal cord are
Plantar cushion removed. Thus, the adequate stimulus for the neu-
Surrounding skin rons is unknown in slice recordings. Since neurons
L5 L6 L7 are heterologous with respect to their inputs in all
Medial areas of the spinal cord, the physiological properties
Figure 12 Somatopic organization of the foot obtained from slice preparations can only be related
representation in the spinal cord of cats. Most medial toe to the types of inputs they receive in a broad fashion,
designated as toe 2; most lateral toe designated as toe 5. drawing on what is know about the most common
Map depicts density centers and does not indicate the
types of neurons found in the specific laminae. With
overlapping of cell groups that have different receptive field
locations. From Light, A. R. and Kavookjian, A. M. 1984. these limitations in mind, still, much useful knowl-
Substantia gelatinosa neurons with axons projecting to edge has been obtained from slice preparations.
other dorsal horn laminae. Soc. Neurosci. Abstr. 10, 489, Where possible, we relate the properties observed
reprinted with permission. in slice preparations to those obtained from intact
preparations.
24.15 Physiological Properties of
Spinal Neurons
24.16 Intrinsic Properties of Spinal
Considerable detail of the physiological properties of Dorsal Horn Neurons
the intrinsic neurons of the spinal cord has been
obtained by using recordings from spinal cord slices. In the spinal dorsal horn, using sharp electrode and
These preparations have the advantage of a known a whole-cell patch clamp recording several types of
stable milieu surrounding the neurons. In addition, neurons have been distinguished on the basis of
access to neurons in either the transverse or horizon- their intrinsic firing properties. However, these firing
tal slice is less problematic because less white matter types and the relative numbers of each type vary
tissue must be traversed with a microelectrode before depending on the experimental conditions and
324 Spinal Cord Physiology of Nociception

species. Seven types of firing have been identified. amplitude attenuation during sustained depolariza-
Tonic, burst, delayed onset, gap, phasic, and single tion. (Figures 13 (Aa) and 13 (Ca)). The majority of
spike. These are shown in Figure 13. tonic neurons are either WDR or nociceptive-specific
Tonically firing neurons have repetitive action neurons (Lopez-Garcia, J. A. and King, A. E., 1994).
potentials with little frequency adaptation and little The ion channels expressed by tonic neurons that

Figure 13 Firing patterns of neurons. (A) Five patterns of rat spinal dorsal horn neurons. Firing patterns were obtained in
response to 1-s injections of depolarizing current (25350 pA, 25-pA steps) at resting membrane potential. Bottom traces in
(a)(e), injected currents. (B) Typical firing patterns of lamina I projection neurons. Firing patterns were obtained in response to
depolarizing current injection from hyperpolarized holding potentials. C: Deep dorsal horn neurons show three modes of
firing. A: from Ruscheweyh, R. and Sandkuhler, J. 2002. Lamina-specific membrane and discharge properties of rat spinal
dorsal horn neurones in vitro. J. Physiol. 541, 231244, reprinted with permission; B: from Dahlhaus, A., Ruscheweyh, R., and
Sandkuhler, J. 2005. Synaptic input of rat spinal lamina I projection and unidentified neurones in vitro. J. Physiol. 566(Pt 2),
355368, reprinted with permission; C: from Derjean, D., Bertrand, S., Le Masson, G., Landry, M., Morisset, V., and Nagy, F.
2003. Dynamic balance of metabotropic inputs causes dorsal horn neurons to switch functional states. Nat. Neurosci. 6(3),
274281, reprinted with permission.
 Spinal Cord Physiology of Nociception 325

shape this response pattern are only partly understood. understood. However, some have suggested that this
It has been suggested that a voltage gated Na current pattern could contribute to the distortion of nocicep-
in combination with a pronounced delayed rectifier tive processing in the dorsal horn that is associated
K (KDR) current and absent or weak A-type K (KA) with persistent pathological pain (Herrero, J. F. et al.,
current help to generate the basic tonic firing pattern 2000). An example of an animal model of this is that
(Ruscheweyh, R. and Sandkuhler, J., 2002; Melnick, I. increased burst firing of neurons in the spinal cord
V. et al., 2004b). A Ca2-dependent K (KCa) conduc- was correlated with the induction of recurrent autot-
tance that is sensitive to apamin regulates the omy in nerve injured rats (Asada, H. et al., 1996).
discharge frequency, prolonging interspike intervals,
and stabilizing firing pattern evoked by sustained
24.16.2 Delayed-Onset Firing Neurons
membrane depolarization (Melnick, I. V. et al., 2004a).
Tonic firing neurons in lamina I display pro- Delayed-onset firing neurons demonstrate a slow ramp
longed excitatory postsynaptic potentials (EPSPs). depolarization in response to subthreshold depolariz-
The mechanisms for these prolonged EPSPs appears ing current pulses (Yoshimura, M. and Jessell, T. M.,
not to be a property of the presynaptic release, but 1989; Grudt, T. J. and Perl, E. R., 2002; Prescott, S. A.
are mediated by a persistent sodium current, INa,p and De Koninck, Y., 2002; Ruscheweyh, R. and
that amplifies and prolongs the depolarization caused Sandkuhler, J., 2002). With higher current injections,
by brief stimulation, combined with a persistent a delay between the onset of the depolarizing current
calcium current, ICa.p, that contributes to the prolon- and the first action potentials gradually shortens.
gation, but not the amplification (Prescott, S. A. and De Repetitive firing is less regular than in tonic firing
Koninck, Y., 2005). The net result is that these cur- neurons, and often, the firing rate increases during
rents can increase integration time and encourage the current pulse (Prescott, S. A. and De Koninck, Y.,
temporal summation of inputs, at the expense of 2002; Ruscheweyh, R. and Sandkuhler, J., 2002).
reduced ability to encode the timing of synaptic Yoshimura M. and Jessell T. M. (1989a) and
inputs. Thus, the tonic firing neurons operate as Ruscheweyh R. and Sandkuhler J. (2002) suggested
integrators of synaptic inputs. that the transient outward rectifying current (IA) was
involved in the response of neurons to sudden depo-
larization and served to delay the onset of the first
24.16.1 Burst Firing Neurons
action potentials.
Burst firing neurons exhibit a high-frequency barrage Delayed-onset neurons were found almost entirely
of action potentials that ride on the peak of a low- in lamina I and II, and not in deeper laminae.
threshold Ca2 spike (Figure 13 (Bb)). In the super- Whether these neurons are nociceptive, and the func-
ficial dorsal horn (laminae I and II), the burst firing tional relevance of this firing pattern is not known.
pattern was characterized by a short burst of two to
four action potentials riding on a slow depolarizing
24.16.3 Gap Firing Neurons
wave at the onset of a depolarizing current pulse
(Ruscheweyh, R. et al., 2004; see Figure 13 (Bb)). Gap firing neurons, also called late spiking neurons, are
With stronger current injections, this pattern could characterized by a considerably delayed-onset first
be converted to the tonic pattern (Ruscheweyh, R. spike flowed by tonic firing after week current pulses.
and Sandkuhler, J., 2002). The burst pattern was With stronger current pulses, a long interval occurs
evident only if the neuron was held more negative between the first and second spike apparently due to a
than 60 mV in whole-cell recordings. The current transient hyperpolarization occurring after the onset of
causing the bursting pattern was not affected by the first depolarizing impulse (see Figure 13 (Ba)).
tetrodotoxin, but was partially blocked by Ni2 and Ruscheweyh R. et al. (2004) suggested that this firing
Cd2. Interestingly, the burst firing occurred almost pattern could be evoked only if the neuron was held at
exclusively in projection neurons and was nearly a negative potential more negative than 75 mV. They
absent in nonprojection neurons, and found more also suggested that gap firing and burst firing are the
often in neurons projecting to the periaqueductal most common patterns of projection neurons with less
gray than neurons projecting to the parabrachial excitable membranes having a broad action potential
region (Ruscheweyh, R. et al., 2004). duel to a hump in the falling phase of the action
The potential function of burst firing for dorsal potential. They determined that gap firing neurons
horn nociceptive neurons is not currently preferentially project to the parabrachial region of the
326 Spinal Cord Physiology of Nociception

brainstem, with fewer projecting to the periaqueductal current may be involved in generating the single
gray region. Apparently, a slow IA is required for the spike pattern. 4-aminopyridine (a Kv blocker) con-
appearance of gap firing (Ruscheweyh, R. et al., 2004). verts single-spike firing to tonic firing (Ruscheweyh,
R. and Sandkuhler, J., 2002).

24.16.4 Phasic Firing Neurons

Phasic firing neurons (Figure 13Ac) generate a short 24.17 Relationship between
burst of action potentials during sustained depolariza- Intrinsic Firing Patterns and
tion. They have been reported by several groups Physiological Types of Dorsal
(Thomson, A. M. et al., 1989; Lopez-Garcia, J. A. and Horn Neurons
King, A. E., 1994; Prescott, S. A. and De Koninck, Y.,
2002; Ruscheweyh, R. and Sandkuhler, J., 2002; Very few investigators have attempted to relate the
Melnick, I. V. et al., 2004b). The phasic pattern may firing patterns described above to the types of adequate
be related to the density of Na channels, and their stimuli that normally activate these neurons. Figure 14
voltage dependency, which may be shifted towards demonstrates the data from one such attempt in rats.
more positive potentials. Melnick I. V. et al., (2004a) As this figure indicates Lopez-Garcia J. A. and King
found that phasic neurons were found in the lateral part A. E. (1994) determined that the majority (81%) of
of lamina II. Lopez-Garcia J. A. and King A. E. (1994) WDR neurons responded with tonic firing, while noci-
classified most phasic neurons as nociceptive specific. ceptive-specific neurons were mostly phasic firing
neurons. Low-threshold mechanoreceptor (LTM) neu-
rons were of both the tonic and phasic firing types.
24.16.5 Single-Spike Firing Neurons
Neurons that were not fired by adequate stimuli, but
Single-spike firing neurons (Figure 13(Ad)) typically demonstrated subthreshold inputs were either tonic
generate only one action potential at the onset of a firing or single spike type of neurons. In contrast to
current pulse, regardless of the intensity of the cur- their conclusions, Graham B. A. et al. (2004) using
rent pulse. A transient, voltage-dependent outward whole-cell recordings in adult mice in vivo did not

16

14
Tonic firing
12
Phasic firing
Single spike firing
Number of cells

10
Anomalous rectification
8

0
WDR LT NS INH SUB
Neuronal responses
Figure 14 Distribution of electrophysiological groups from a hemisected slice preparation from rats with hindlimb attached.
The number of physiologically classified wide-dynamic range neurons (WDR), low-threshold neurons (LT), nociceptive-
specific neurons (NS), and neurons displaying inhibitory events (INH) are shown along with the firing type. SUB, subthreshold
for firing action potentials. Adapted from Lopez-Garcia, J. A. and King, A. E. 1994. Membrane properties of physiologically
classified rat dorsal horn neurons in vitro: correlation with cutaneous sensory afferent input. Eur. J. Neurosci. 6, 9981007,
with permission.
 Spinal Cord Physiology of Nociception 327

find a consistent relationship between any of the firing Dorsal horn neurons are modulated by inputs
pattern types of neurons and physiological inputs. from higher brain centers. Inputs from other centers
Lu Y. and Perl E. R. (2003) also found that all can greatly modify the amplitude of signals relayed
lamina II neurons receiving inhibitory inputs from from primary afferent neurons.
other lamina II neurons were of the phasic firing Dorsal horn neurons are quite heterogeneous with
type, indicating at least some functional organization respect to the types of inputs that they encode, and
is related to the firing pattern of lamina II neurons. with respect to their intrinsic membrane properties.
Clearly, the firing patterns described here reflect In at least some cases, it appears that the intrinsic
specific currents mediated by specific ion channels. membrane properties are associated with the types of
These specific ion channels can be altered by a number inputs they receive, and these intrinsic properties
of pharmacologic agents, endogenous neurotransmit- may allow the neurons to better integrate certain
ters and neuromodulators. The specificity of neurons types of inputs.
for these channels could convey specificity of drug
action on particular functional types of neurons.
Some of this is the topic of Chapter 20, and much of
this will be the subject of future investigations into the References
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 25 What is a Wide-Dynamic-Range Cell?
D Le Bars, INSERM U-713, Paris, France
S W Cadden, University of Dundee, Dundee, UK
2009 Elsevier Inc. All rights reserved.

25.1 Introduction 332

25.2 Segmental Excitatory Receptive Fields 332
25.2.1 General 332
25.2.2 Plasticity 333
25.3 Segmental Inhibitory Receptive Field 335
25.3.1 General 335
25.3.2 Plasticity 335
25.4 Perturbance of the Basic Somesthetic Activity by a Painful Focus 335
25.5 Multiple or Vast Painful Foci 336
25.6 Summary and Conclusions 337
References 337

Glossary
allodynia Pain caused by stimuli which would not excitatory receptive field Area of the body (sur-
normally cause pain (i.e., non-noxious stimuli). face or interior) which when stimulated will produce
angina pectoris Chest pain from cardiac excitation of a given neuron.
ischemia. hyperalgesia Increased level of pain felt in
basic somesthetic activity Description of the on- response to a noxious stimulus (cf. allodynia).
going activity in somatosensory pathways in the hypoalgesic Pertaining to a reduction of pain.
absence of any deliberate stimuli. inhibitory receptive field Area of the body (sur-
bistable A mechanism which can essentially be in face or interior) which when stimulated will produce
one of two states (e.g., on or off) inhibitory effects on the activity of a given neuron.
body image Conscious mental representation of microelectrophoretic application Application of
the body. a substance in its ionic form from a microelectrode
body schema Unconscious model of the body in by the application of electrical current.
the brain. monoarthritis Inflammation of a single joint.
bulbo-spinal control A neural control mechanism nociceptive stimulus Stimulus which activates
originating in the brainstem and projecting to the nociceptive afferents (which could be a noxious
spinal cord. stimulus (see below) or an electrical stimulus).
dermatome Area of skin innervated by nerves noxious stimulus Stimulus which causes or
from a given segment of the spinal cord. threatens to cause tissue damage.
diffuse noxious inhibitory controls polyarthritis Inflammation of several joints.
(DNIC) Neural controls triggered by noxious sti- propriospinal mechanism Neural mechanism
mulation of widespread areas of the body which mediated entirely within the spinal cord.
exert an inhibitory influence on wide-dynamic- radiating pain Pain which spreads from the area
range neurons. of stimulation to adjacent areas.
dorsal column stimulation Technique for indu- referred pain Pain which is felt in areas remote to
cing analgesia/hypoalgesia by electrically the area of stimulation (as well as or instead of from
stimulating the dorsal columns of the spinal the area of stimulation).
cord.

331
332 What is a Wide-Dynamic-Range Cell?

supraspinal mechanism Neural mechanism nerves through the skin (i.e., with electrodes placed
which at least in part, involves structures above the on the skin).
spinal cord. trigeminal nucleus caudalis Most caudal part of
transcutaneous electrical nerve the trigeminal sensory nuclear complex (or of the
stimulation Technique for inducing analgesia/ trigeminal spinal nucleus); also sometimes called
hypoalgesia by electrically stimulating peripheral the medullary dorsal horn.

25.1 Introduction the concept that they exhibit whole body receptive
fields. One consequence of this unusual property is
Wide-dynamic-range (WDR) neurons (various other that it necessitates consideration of the output of the
names have been used by different authors for these whole population of these neurons (Le Bars, D.,
neurones, e.g., trigger, lamina V-type, class 2, multi- 2002). The extent of a nociceptive focus can differ
receptive, and convergent neurones) are found in the greatly depending on its cause, and consequently the
spinal dorsal horn (and its medullary homolog, tri- global volume of information sent to the brain will
geminal nucleus caudalis), both in the superficial also vary. Although a priori this volume of informa-
layers and more so in deeper areas centered on tion is determined by the intensity, duration, and area
lamina V. They are activated by both nociceptive being stimulated, it can also be altered profoundly by
and non-nociceptive stimuli and include interneur- past and present experiences.
ons involved in polysynaptic reflexes and neurons
projecting in pathways such as the spinothalamic
and spinoreticular tracts (Willis, W. D. and
25.2 Segmental Excitatory Receptive
Coggeshall, R. E., 1991). For various reasons, these
Fields
neurons are believed to play a key role in pain. For
example, many procedures used to alleviate pain in 25.2.1 General
man (e.g., systemic or intrathecal morphine, transcu- WDR neurons receive information from all three
taneous electrical nerve stimulation, and dorsal groups of cutaneous afferents, that is, A-, A-, and
column stimulation), also result in a reduction in C-fibers (e.g., Wagman, I. H. and Price, D. D., 1969;
the responses of WDR cells to nociceptive stimuli. Menetrey, D. et al., 1977). The excitatory receptive
Indeed, some WDR neurons show patterns of activity fields from which this information arises are fairly
which are more reminiscent of certain types of pain restricted in size (Figure 1). Studies in conscious
than activity in peripheral nociceptive nerves monkeys illustrate that these receptive fields can
(Adriaensen, H. et al., 1984; Cervero, F. et al., 1988). expand or contract depending on the attentional
For example, a WDR neuron may remain active after state of the animal (e.g., Dubner, R. et al., 1981).
removal of a stimulus from its peripheral excitatory Although the receptive fields are usually larger than
field (e.g., Wagman, I. H. and Price, D. D., 1969) those of nociceptive-specific neurons, they can still
while conversely it may remain inactive following provide useful information regarding stimulus loca-
the brief activation of inhibitory mechanisms even if tion. Usually, these receptive fields exhibit a gradient
there is still a stimulus on the excitatory field of sensitivity: in the center even light mechanical
(Cadden, S. W., 1993). Such bistable patterns of activ- stimuli can activate the neuron, while at the periph-
ity might be explained by inherent properties of ery only intense stimuli elicit responses (e.g.,
WDR neurons (e.g., Morisset, V. and Nagy, F., Menetrey, D. et al., 1977).
1996) and/or their involvement in positive feedback When increasing intensities of mechanical or
circuits (see Cadden, S. W., 1993). thermal stimuli are applied to the center of the
WDR neurons also constitute a site where excita- receptive field, the rate and duration of neuronal
tory and inhibitory influences converge. This chapter firing increases (see insert in Figure 1). In this
focuses on the properties of WDR neurons under respect, these neurons seem better than nocicep-
physiological conditions, and most particularly, on tive-specific neurons at encoding intensities of
 What is a Wide-Dynamic-Range Cell? 333
Cutaneous field:
- excitatory field (EF)
200 Hz
- inhibitory field (IF)
1 min
EF IF

Tactile stimulus 41 C 43 C 46 C 48 C

()
(+)
(+)
Brain
Visceral field (+)

A & A fibers
A fibers
A & C fibers
Muscular field

Figure 1 Schematic organization of the segmental receptive field of a wide-dynamic-range (WDR) neuron. The principal
part on the skin is made up of an excitatory (EF) and an inhibitory (IF) field. The center of the EF can be activated by both
innocuous and nociceptive stimuli (blue and red stripes, respectively). The periphery of the EF is activated only by nociceptive
stimuli (red). The inhibitory field (yellow) is activated mainly by non-nociceptive stimuli. As shown, the receptive field may also
include visceral and/or muscular components thus completing a convergence of excitatory () and inhibitory () information
onto a single neuron. A ratemeter recording from a WDR neuron in the rat lumbar cord (insert, top right) shows responses to
stimuli applied to the center of the excitatory field: repetitive innocuous (light pressure) stimuli produced a high level of firing
while applications of radiant heat (indicated by bars with temperatures achieved) evoked responses which increased with
temperature within the noxious range. Adapted from Le Bars, D. and Chitour, D. 1983. Do convergent neurons in the spinal
cord discriminate nociceptive from non-nociceptive information? Pain 17, 119.

noxious stimuli (Hoffman, D. S. et al., 1981). It is often helps explain clinical curiosities such as radiating or
believed that WDR neurons respond more strongly referred pain, for example, to the left arm in angina
to noxious stimulation than to any form of innocuous pectoris. Indeed, WDR neurons can capture all the
stimulation. However, this is not quite true. In fact, information coming from interfaces with both the
the responses evoked in WDR neurons by noxious external environment (the skin) and the internal
heat can be exceeded by repetitively applied innoc- milieu (viscera, muscles, etc.). This information con-
uous mechanical stimuli (insert of Figure 1) (see Le stitutes a basic somesthetic activity. Thus, it is
Bars, D. and Chitour, D., 1983). Thus, firing fre- possible that these neurons are not involved exclu-
quency alone cannot distinguish between non- sively in pain and, by transmitting this basic
noxious and noxious stimuli. How can this fact be somesthetic activity, inform the brain that the organ-
reconciled with WDR neurons having a central role ism is suffering no special perturbation from either
in pain processing? It seems that the meaningful external or internal sources.
signal is the pattern of activity from the whole popu-
lation of WDR cells and not simply the signals from
25.2.2 Plasticity
each individual neuron. As their receptive fields
overlap one another, the organization of spatial con- The convergence onto a single WDR neuron is actu-
vergence from WDR neurons must be taken into ally greater than is observed under normal
account. This is illustrated by the fact that when a conditions. This discrepancy is such that on fringes
tactile stimulus is applied to a given territory, it of the excitatory field, there is a zone which when
activates only the few WDR neurons whose excita- stimulated will trigger a depolarization of the WDR
tory fields have centers in the territory. However, a neuron (Figure 2(c)) which normally will be insuffi-
noxious stimulus applied in the same place, will cient to elicit action potentials. This is consistent
activate not only the same WDR neurons, but also with the observation that cutaneous receptive fields
those whose excitatory fields have peripheries in the of WDR neurons are apparently smaller when
territory (Figure 2). mapped using natural as opposed to electrical stimu-
Moreover, some WDR neurons are activated from lation (e.g., Mendell, L. M., 1984), given that the
more than one type of tissue (e.g., cutaneous and visc- electrical stimuli produce more synchronized affer-
eral or cutaneous and muscular). This convergence ent activity which results in more effective
334 What is a Wide-Dynamic-Range Cell?

(a) (b)

Brain Brain

(c) (d)

Brain Brain

(e) (f)

Brain Brain

Figure 2 (a) When recordings are made from an individual wide-dynamic-range (WDR) neuron (the blue neuron in the
diagram), activation by noxious stimuli applied to any part of its excitatory receptive field can be observed (blue surface on the
skin). Its center (blue dark) is also sensitive to low-intensity stimuli. (b) The excitatory fields of adjacent WDR neurons overlap one
another, so that a noxious stimulus will activate not only the center of some receptive fields (blue area) but also the edges of
others (pale surfaces colored red, purple, and yellow). Thus, when applied to a given surface area (here the center of the blue
field), a noxious stimulus will activate many neurons (four in this case) whereas a non-noxious stimulus will activate far fewer
(here only one). (c) In fact the surface potentially covered by the excitatory receptive field of a WDR neuron is larger than that
observed under physiological conditions. If the neuron (blue) from which recordings are being made is partially depolarized (e.g.,
by microelectrophoretically applying an excitatory amino acid to its membrane), an enlargement of the excitatory receptive field
can be observed. This means that the span of the excitatory receptive fields is potentially much greater than shown in (a) (as
indicated by the broken white lines). This is probably what occurs under inflammatory conditions. (d) Thus, considering four
neurons, it can be seen that a non-noxious stimulus applied to the center of the blue field will now be able to activate the red,
purple, and yellow neurons as well (cf. (b)). (e) At the periphery of their excitatory fields, WDR neurons have an inhibitory field
(white zone). When mechanical stimuli are applied to this area, the neuron is inhibited. (f) The inhibitory fields of a population of
adjacent WDR neurons substantially overlap one another. Thus in the center of the red field, mechanical stimuli activate the red
neuron but inhibit the purple and yellow neurons. When tactile stimuli are applied to a large surface, they will not only activate the
centers of excitatory fields and potentially generate a false nociceptive signal, they will also activate inhibitory fields with a
resulting attenuation of the global response. This is the basis for some physical methods of pain-relief (see text).

summation of excitatory postsynaptic potentials. The Nishikawa, Y., 1979). Under these circumstances,
concept of quiescent synapses (e.g., Wall, P. D., 1977) the receptive fields enlarge and this is also seen
developed with the idea that in some conditions, this after transient electrical stimulation of C-fibers,
fringe of the receptive field can produce significant injury, noxious pinch, intradermal injection of cap-
information transfer, that is, trigger action potentials. saicin, or noxious stimulation of viscera or muscle.
Such synapses can be made more efficacious experi- Such plasticity may be triggered naturally by per-
mentally by partially depolarizing a neuron using ipheral or central pathologies. In general, sensitization
microelectrophoretic application an excitatory of excitatory mechanisms or a deficiency of inhibitory
amino acid (Zieglgansberger, W. and Herz, A., ones will produce both increased activity in the neu-
1971) or pharmacologically blocking inhibitory ronal population associated with a painful focus and a
transmitter mechanisms (e.g., Yokota, T. and growth of this population (Figure 2(d)). This is
 What is a Wide-Dynamic-Range Cell? 335

illustrated by the increased size of excitatory receptive such information could constitute a form of allodynia
fields during acute or chronic inflammation (e.g., such as that triggered by brushing the skin with cotton
Grubb, B. D. et al., 1993). The resulting increased wool, which is characteristic of some neuropathies.
information is transmitted to, and decoded in, the The convergence of excitatory and inhibitory
brain where it constitutes a form of hyperalgesia. influences may also explain the hypoalgesic effects of
This increase in nociceptive information varies with stimulating parts of the body close to a painful focus,
the volume of the body affected by the lesion (i.e., in a for example, by rubbing or transcutaneous electrical
three-dimensional way) and also has a temporal nerve stimulation (TENS). Indeed, the fact that large
dimension. and fine cutaneous afferent fibers interact with this
potential effect has been known for a long time (e.g.,
Head, H. et al., 1905; Noordenbos, W., 1959; Melzack,
25.3 Segmental Inhibitory Receptive R. and Wall, P. D., 1965).
Field
25.3.1 General
25.4 Perturbance of the Basic
Somesthetic Activity by a Painful
WDR neurons frequently exhibit not only excitatory Focus
cutaneous receptive fields, but also inhibitory fields
that are often situated proximally (e.g., Hillman, P. In addition to the segmental inhibitory influences
and Wall, P. D., 1969). When applied to this area, discussed above, WDR neurons are inhibited by
most mechanical stimuli, notably those of low inten- nociceptive stimulation of areas of the body remote
sity, inhibit the activity of WDR neurons. It is from their segmental receptive fields. These effects
often implied that these phenomena are triggered involve propriospinal and supraspinal mechanisms.
only by the activation of A-fibers. However, numer- The former can be observed in the dorsal horn of
ous studies have demonstrated that activation of spinal animals (e.g., Gerhart, K. D. et al., 1981) but the
A-fibers results in more powerful inhibitions (e.g., underlying mechanisms have not been investigated
Chung, J. M. et al., 1984). Thus the activation of a pool in detail. The supraspinal mechanisms are observed
of A-fiber terminals will both activate those WDR in intact but not in spinalized animals and have been
neurons with excitatory receptive fields in this parti- studied more extensively (see Chapter Diffuse
cular area and inhibit those with overlapping Noxious Inhibitory Controls (DNIC)).
inhibitory receptive fields. As described below, the In fact, any noxious stimulus will activate descend-
final output from these neurons constitutes a con- ing bulbo-spinal controls and these will strongly
trasted picture reminiscent of the lateral inhibition depress the activity of neurons in the spinal dorsal
described in other systems (e.g., visual and touch). horn and its trigeminal homolog. In several species,
the vast majority of WDR neurons (and some noci-
ceptive-specific neurons) are profoundly inhibited by
25.3.2 Plasticity
noxious stimulation of any part of the body provided it
It is important to consider the overlap of excitatory is not the neurons own excitatory receptive field. In
and inhibitory fields of neighboring WDR neurons effect, the receptive fields of these neurons are made
(Figures 2(e) and 2(f)). The organization of the inhi- up by the whole body mainly in the form of an
bitory fields offers an explanation for how, at least in inhibitory receptive field except in its own neural
nonpathological conditions, the high levels of activity segment where both inhibitory and excitatory fields
produced in individual neurons by application of mul- exist. These whole body phenomena which have been
tiple non-nociceptive stimuli to large surface areas termed diffuse noxious inhibitory controls (DNIC)
(incorporating numerous centers of excitatory fields) work by hyperpolarizing WDR neurons (see Chapter
do not constitute a nociceptive signal. Such an even- Diffuse Noxious Inhibitory Controls (DNIC)).
tuality is prevented by the concomitant stimulation of In summary, a noxious focus activates a segmental
numerous inhibitory fields which attenuates the over- subset of WDR neurons and inhibits all other WDR
all response of the neuronal population. in contrast, a neurons (Figure 3(b)). This mechanism effectively
deficit of such inhibitory effects will result in a greater improves the signal-to-noise ratio by increasing the
amount of activity being produced by harmless sti- contrast between the activities of the excited neurons
muli. Once transmitted to and decoded by the brain, and the depressed activity of the remainder.
336 What is a Wide-Dynamic-Range Cell?

42 C A
Ba

45 C
Bulbar
reticular
formation

50 C 50 Hz
DNIC Bb
30 s

DLF DLF

Nociceptive
stimulus

Bc

A & C
fibres

Ventro-lateral
quadrant

Figure 3 (A) Triggering of descending inhibitory controls by nociceptive stimuli. One consequence of a painful focus is the
modification of activity of a whole population of WDR neurons. Nociceptive signals pass up the ventrolateral quadrant to
activate structures in the bulbar reticular formation. These structures in turn send signals down the dorsolateral cord to trigger
powerful inhibitory mechanisms in all segments not concerned with the initial stimulus. The insert (top left) shows a recording
from a WDR neuron in the rat lumbar cord whose receptive field on the extremity of a hindpaw, was mechanically stimulated
by a paint brush once every 10 s (indicated by black spots). The resulting activity was inhibited by the application of remote
noxious stimuli (immersion of the tail in 42, 45, or 50  C water). This inhibition increased with the water temperature. (B)
Schematization of the activity of all WDR neurons: (Ba) in the absence of a painful focus, these neurons produce the basic
somaesthetic information, consisting of signals from the body about its immediate environment; (Bb) as a consequence of a
painful focus, some of these neurons are activated (four of them in the center of this diagram), but it is difficult to extract this
information because of the background noise; (Bc) however the inhibition of the other neurons by the mechanisms described
in (a), makes it possible for the brain to extract the information about the painful focus due to the contrast between the
activated and the inhibited neurons. DLF, dorsolateral funiculus; DNFC, diffuse noxious inhibitory control.

Interestingly, DNIC are exacerbated in experimental The size of a painful focus can vary greatly
models of clinical pain (e.g., monoarthritis, polyar- depending on the underlying pathology and conse-
thritis, and peripheral mononeuropathies; e.g., quently the volume of nociceptive information sent
Danziger, N. et al., 2000). to the brain will also vary. How do WDR neurons
behave when noxious stimuli are applied to increas-
ingly large surface areas? In fact, two opposite effects
25.5 Multiple or Vast Painful Foci are observed: up to a particular area of stimulation
approximately twice that of the neurons excitatory
If it is accepted that WDR neurons are important in field there is an accelerating transfer function;
pain, then interactions between nociceptive signals however for larger surface areas, the responses gra-
from separate parts of the body must result in inter- dually diminish (Bouhassira, D. et al., 1995). This is
actions between the corresponding pains. In fact, a consistent with findings in humans that noxious ther-
painful stimulus can decrease or even mask pain mal stimulation of increasingly large areas of skin
arising from another part of the body (see Chapter causes an increasingly painful sensation only over a
Diffuse Noxious Inhibitory Controls (DNIC)). restricted range of surface areas (e.g., Hardy, J. D.
 What is a Wide-Dynamic-Range Cell? 337

et al., 1952) and that painful thermal stimuli evoke relative unconsciousness and the painful focus
pleasant warmth when applied to larger areas becomes pre-eminent and relatively oversized.
(Melzack, R. et al., 1962). The classical poor correla- Spino-bulbo-spinal mechanisms could underlie this
tion between the extent of an injury and the intensity over-representation by providing a filter which
of pain may also involve such mechanisms. imposes the nociceptive signal to the detriment of
Thus it seems that a painful focus activates a information from the rest of the body.
negative feedback loop affecting WDR neurons After all, this is what happens within any large
only when it generates a sufficient volume of noci- gathering: silence is necessary if a speaker is to be
ceptive information. This volume is undoubtedly heard. DNIC establish such silence among spinal
determined by the intensity of the stimulus, the neurons. Conversely, a hubbub does not reduce the
area to which it is applied, and its duration. In this speaker to silence, but drowns his speech to the extent
respect, the role of temporal summation in triggering that he cannot be heard. This is one way in which
inhibitory bulbo-spinal mechanisms must be morphine works and DNIC are extremely sensitive
addressed (Gozariu, M. et al., 1997). to low doses of morphine (see Le Bars, D. et al., 1995).
In summary, WDR neurons do not constitute a
single-channel transmission network. Nociceptive
25.6 Summary and Conclusions information is processed by complex mechanisms
which can modify the gain of the system. The
The global population of WDR neurons is activated modulation of this gain is determined by the three-
continuously but unpredictably by all the non-nox- dimensional characteristics of the painful focus not
ious stimuli provided by the environment. The only its intensity and duration but also its surface
resulting basic somaesthetic activity can constitute area. Beyond a critical level, an increase in the num-
either noise from which the brain may have difficulty ber of activated noci-responsive neurons is associated
extracting a clear signal, or a significant message. with a decrease in their individual responses.
According to modern neuropsychological concepts, A complete analysis of nociceptive processes must
this on-going information from the whole body, consider not only a large population of neurons
along with that from other sensory systems, notably but also their interactions. Pain in clinical practice
the vestibular and visual systems, is integrated to is not punctuate and involves a large number of
generate a permanent, unconscious, representation excitatory receptive fields of peripheral fibers
of the physiological reality of self. At the beginning and central neurons. When several foci coexist, the
of the twentieth century, the great neurologist Henry global signal from WDR neurons may be poly-
Head proposed that our brains contain two schemas morphic with the resulting pain showing
a postural model of ourselves and another schema fluctuations, instability, or even versatility due to
allowing the recognition of the locality of the stimu- the resulting interactions.
lated spot (Head, H. and Holmes, G., 1911).
However, following Schilder P. (1950), the simple
term body schema is usually used to refer to both References
models. Even though the terms are sometimes used
interchangeably, it is advisable to distinguish body Adriaensen, H., Gybels, J., Handwerker, H. O., and Van Hees, J.
schema which is an unconscious cerebral model of 1984. Nociceptor discharges and sensations due to
the body, from body image which is a conscious prolonged noxious mechanical stimulation: a paradox.
Human Neurobiol. 3, 5358.
mental representation. WDR neurons are good can- Bouhassira, D., Gall, O., Chitour, D., and Le Bars, D. 1995.
didates for constructing and endlessly rebuilding the Dorsal horn convergent neurones: negative feed-back
memory of ones physical self (Le Bars, D., 2002). triggered by spatial summation of nociceptive afferents. Pain
62, 195200.
Previous experiences, be they neutral, pleasant, or Cadden, S. W. 1993. The ability of inhibitory controls to switch-
unpleasant (e.g., pain), contribute to the building of off activity in dorsal horn convergent neurones in the rat.
this memory; all will have activated WDR neurons. Brain Res. 628, 6571.
Cervero, F., Handwerker, H. O., and Laird, J. M. 1988.
Intense or long-lasting pains are probably among the Prolonged noxious mechanical stimulation of the rats tail:
physical causes which most commonly disturb the responses and encoding properties of dorsal horn neurones.
body schema: they generate a painful body by focus- J. Physiol. 404, 419436.
Chung, J. M., Fang, Z. R., Hori, Y., Lee, K. H., and Willis, W. D.
ing attention on one part of the body to the detriment 1984. Prolonged inhibition of primate spinothalamic tract
of the others. The body schema then emerges from cells by peripheral nerve stimulation. Pain 19, 259275.
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Danziger, N., Le Bars, D., and Bouhassira, D. 2000. Diffuse Le Bars, D., Bouhassira, D., and Villanueva, L. 1995. Opioids
Noxious Inhibitory Controls and Arthritis in the Rat. In: Pain and and Diffuse Noxious Inhibitory Controls in the Rat. In: Pain
Neuroimmune Interactions (eds. N. E. Saade, A. V. Apkarian, and the Brain: From Nociceptor to Cortical Activity
and S. J. Jabbur), pp. 6978. Kluwer Academic Press. (eds. B. Bromm and J. E. Desmedt), Advances in Pain
Dubner, R., Hoffman, D. S., and Hayes, R. L. 1981. Research and Therapy, Vol. 22, pp. 517539. Raven
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1981. Inhibitory receptive fields of primate spinothalamic tract Mendell, L. M. 1984. Modifiability of sipnal synapses. Physiol.
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Gozariu, M., Bragard, D., Willer, J. C., and Le Bars, D. 1997. Menetrey, D., Giesler, G. J., Jr., and Besson, J. M. 1977. An
Temporal summation of C-fiber afferent inputs: competition analysis of response properties of spinal cord dorsal horn
between facilitatory and inhibitory effects on C-fiber reflex in neurones to nonnoxious and noxious stimuli in the spinal rat.
the rat. J. Neurophysiol. 78, 31653179. Exp. Brain Res. 27, 1533.
Grubb, B. D., Stiller, R. U., and Schaible, H. G. 1993. Dynamic Morisset, V. and Nagy, F. 1996. Modulation of regenerative
changes in the receptive field properties of spinal cord neurons membrane properties by stimulation of metabotropic
with ankle input in rats with chronic unilateral inflammation in glutamate receptors in rat deep dorsal horn neurons. J.
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Hardy, J. D., Woolf, H. G., and Goodel, H. 1952. Pain Noordenbos, W. 1959. Pain. Elsevier.
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Head, H. and Holmes, G. 1911. Sensory disturbances from Body. International Universities Press.
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Head, H., Rivers, W. H. R., and Sherren, J. 1905. The afferent cells of M. Mulatta to cutaneous and sural A and C fiber
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 26 Spinal Cord Mechanisms of Hyperalgesia and Allodynia
T J Coderre, McGill University, Montreal, QC, Canada
2009 Elsevier Inc. All rights reserved.

26.1 Characteristics of Hyperalgesia 341

26.1.1 Definition and Historical Background 341
26.1.2 Precipitating Conditions for Hyperalgesia and Evidence of Central Sensitization 341
26.1.2.1 Cutaneous injury 341
26.1.2.2 Peripheral nerve stimulation 342
26.1.2.3 Inflammation 343
26.1.2.4 Peripheral nerve injury 343
26.1.2.5 Spinal cord injury 344
26.1.2.6 Illness-induced hyperalgesia 345
26.1.2.7 Chronic opiate exposure 345
26.2 Alterations in Spinal Cord Dorsal Horn 345
26.2.1 Alterations in Transmitter or Growth Factor Expression or Release 345
26.2.1.1 Neuropeptides 345
26.2.1.2 Excitatory amino acids 346
26.2.1.3 Other transmitters 349
26.2.1.4 Neurotrophins 349
26.2.1.5 Phenotype switch 350
26.2.2 Expression of Second Messengers and Transcription Factors 350
26.2.2.1 Second messengers 350
26.2.2.2 Transcription factors 353
26.2.3 Alterations in Receptor and Ion Channel Expression, Activity, or Cellular Location 354
26.2.3.1 Neuropeptide receptors 354
26.2.3.2 Excitatory amino acid receptors 354
26.2.3.3 Other receptors 355
26.2.3.4 Scaffolding and adaptor proteins 356
26.2.3.5 Ion channels 356
26.2.4 Alterations in Transporter Activity 358
26.2.4.1 Glutamate transporters 358
26.2.4.2 Cation chloride transporters 358
26.2.5 Glial Cell Alterations 358
26.2.5.1 Microglia 358
26.2.5.2 Astrocytes 359
26.2.5.3 Neuronglial interactions 359
26.2.6 Descending Facilitation 360
26.2.7 Anatomical Changes 360
26.2.7.1 Cell death and apoptotic genes 360
26.2.7.2 Sprouting 361
26.3 Conclusion 362
References 362

Glossary
adaptor protein A protein which is accessory to protein-protein interactions that drive the formation
main proteins in a signal transduction pathway. of protein complexes.
These proteins often lack any intrinsic enzymatic allodynia Pain due to a stimulus that does not
activity themselves, but instead mediate specific normally provoke pain.

339
340 Spinal Cord Mechanisms of Hyperalgesia and Allodynia

antisense oligonucleotides Single strands of across the plasma membrane of all living cells by
DNA or RNA that are complementary to a chosen allowing the flow of ions down their electrochemical
sequence. Antisense RNAs prevent translation of gradient.
complementary RNA strands by binding to them. long-term potentiation (LTP) An increase in the
apoptosis An orchestrated series of biochemical strength of a chemical synapse that lasts from
events leading to cell death. minutes to several days.
carrageenan A linear sulphated polysaccharide neuropeptides A variety of peptides found in
extracted from red seaweeds that is used as an neural tissue.
inflammatory agent. neurotrophins Also called neurotrophic factors,
central sensitization Activity- or injury-depen- are a family of protein which induce the survival of
dent changes in the excitability of central nervous neurons. They belong to a family of growth factors,
system (CNS) neurons, typically in pain pathways, secreted proteins which are capable of signaling
and is reflected by increased spontaneous activity, particular cells to survive, or differentiate, or grow.
reduced thresholds or increased responsiveness to phenotype switch A process by which neurons
afferent inputs, prolonged after- discharges to change their phenotype and begin to produce and
repeated stimulation, and the expansion of the release substances they normally do not produce.
peripheral receptive fields of central neurons. protein kinase Is a kinase enzyme that modifies
complete Freunds adjuvant An antigen solution other proteins by chemically adding phosphate
composed of inactivated and dried Mycobacterium groups to them (phosphorylation).
tuberculosis emulsified in mineral oil, used as an protein tyrosine phophatase (PTP) An enzyme
immunopotentiator (booster of the immune system) that remove phosphate groups from phosphory-
and inflammatory agent. lated tyrosine residues on proteins.
2-deoxy-D-glucose A marker for tissue glucose scaffold proteins Proteins that bring together
use. various other proteins in a signaling pathway and
excitatory amino acids Amino acid neurotrans- allows for their interaction. They recruit down-
mitters that include aspartate and glutamate. stream effectors in a pathway and enhance
Glutamate is the most abundant fast excitatory specificity of the signal.
neurotransmitter in the mammalian nervous secondary messengers Secondary messengers
system. are a component of signal transduction cascades
excitatory postsynaptic potential (EPSP) Is a that utilize receptors on the surface of the plasma
temporary increase in postsynaptic membrane membrane which are generally coupled to a kinase
potential caused by the flow of positively charged on the interior surface of the membrane. The kinase
ions into the postsynaptic cell. then phosphorylates another molecule which car-
glial cells Commonly called neuroglia or simply ries out another action.
glia, are non-neuronal cells that provide support transcription factor A protein that works in con-
and nutrition, maintain homeostasis, form myelin, cert with other proteins to either increase or
and participate in signal transmission in the nervous decrease the transcription of genes.
system. transporters Membrane-bound pumps that move
homosynaptic sensitization Facilitation of activ- biochemicals and other atomic or molecular sub-
ity occuring at the synapses that connect two stances across membranes.
neurons. wide dynamic range A property of spinal cord
hyperalgesia An increased response to a stimulus dorsal horn neurons (typically in lamina V) that
which is normally painful. Primary hyperalgesia occurs exhibit graded responses to non-noxious and nox-
at the site of injury, and secondary hyperalgesia in ious stimuli since they receive convergent input
adjacent, or sometimes remote, uninjured tissue. from non-nociceptive and nociceptive primary
inhibitory postsynaptic potential (IPSP) Is the afferent fibers.
change in membrane voltage of a postsynaptic wind-up Cumulative depolarization, action poten-
neuron which results from synaptic activation of tial bursting and afterdischarges in spinal cord
inhibitory neurotransmitter receptors. dorsal horn neurons following high frequency,
ion channels Pore-forming proteins that help to C-fiber strength electrical stimulation or intensely
establish and control the small voltage gradient noxious natural stimuli.
 Spinal Cord Mechanisms of Hyperalgesia and Allodynia 341

26.1 Characteristics of Hyperalgesia 26.1.2 Precipitating Conditions for

Hyperalgesia and Evidence of Central
26.1.1 Definition and Historical Sensitization
Background
26.1.2.1 Cutaneous injury
The term hyperalgesia was first coined by Gowers 26.1.2.1.(i) Heat injury Considerable evidence
in the 1800s, and is often still used as a term to demonstrates that heat injury of the skin produces a
define what Hardy J. D. (1956) described as a state sensitization of peripheral nociceptors. Peripheral
of increased pain sensation induced by either nox- sensitization occurs both in A and C-fiber nocicep-
ious or ordinarily nonnoxious stimulation of tors (Beitel, R. E. and Dubner, R., 1976; Fitzgerald, M.
peripheral tissue. The International Association and Lynn, B., 1977), and largely accounts for primary
for the Study of Pain (IASP) currently uses sepa- hyperalgesia (Meyer, R. A. and Campbell, J. N., 1981;
rate terms for the two reactions Hardy described. Torebjork, H. E. et al., 1984). In addition, dorsal horn
Thus, hyperalgesia is defined as an increased neurons fire with increasing frequency in response to
response to a stimulus which is normally painful, repeated applications of a noxious heat stimulus
while pain due to a stimulus that does not nor- (Perl, E. R. et al., 1976, Kenshalo, D. R. et al., 1979)
mally provoke pain is known as allodynia (IASP or following thermal injury (Price, D. D. et al., 1978;
website). Over the years, hyperalgesia (not distin- Kenshalo, D. R. et al., 1982), indicative of central
guished from allodynia) has been further separated sensitization. In addition to the sensitization and pro-
into primary and secondary hyperalgesia; primary longed excitation of dorsal horn cells, noxious
hyperalgesia occurs at the site of injury, and sec- stimulation associated with tissue injury also pro-
ondary hyperalgesia in adjacent, or sometimes duces an expansion of the receptive fields of dorsal
remote, uninjured tissue. Primary hyperalgesia is horn neurons. Thus, neurons in the dorsal horn of the
reflected by a hypersensitivity to thermal and spinal cord with receptive fields adjacent to a cuta-
mechanical stimuli, and is driven largely by sensi- neous heat injury expand their receptive fields to
tization of peripheral nociceptors. Secondary incorporate the site of injury (McMahon, S. B. and
hyperalgesia is predominantly characterized by Wall, P. D., 1984).
hypersensitivity to mechanical stimuli, and likely Evidence that central sensitization contributes to
involves central sensitization. Central sensitization secondary hyperalgesia after heat injury was sup-
refers to activity- or injury-dependent changes in ported initially by reports that heat injury-induced
the excitability of central nervous system (CNS) cutaneous hyperalgesia spreads well beyond the area
neurons, typically in pain pathways, and is of nociceptor sensitization. Thalhammer J. G. and
reflected by increased spontaneous activity, Lamotte R. H. (1982) found that a heat injury in
reduced thresholds or increased responsiveness to one half of a cutaneous nociceptors receptive field
afferent inputs, prolonged after-discharges to did not produce heat sensitization in the other half,
repeated stimulation, and the expansion of the despite the fact that hyperalgesia spread into this
peripheral receptive fields of central neurons area. Typically, nociceptor sensitization associated
(Woolf, C. J. et al., 1988; Coderre, T. J. et al., with injury is restricted to about 510 mm of the
1993). Although central sensitization has been site of injury (Fitzgerald, M., 1979) while, in contrast,
demonstrated in neurons throughout the cutaneous hyperalgesia spreads as far as 1020 cm
somatosensory nervous system, the mechanisms beyond the site of injury (Hardy, J. D. et al., 1950).
underlying central sensitization have been largely Although it is commonly reported that there is
generated from studies concentrating on spinal mechanical, but not heat hyperalgesia in the second-
cord dorsal horn neurons. The characteristics and ary zone (Raja, S. N. et al., 1984), secondary heat
underlying mechanisms of central sensitization in hyperalgesia has been reported in some cases
spinal cord dorsal horn neurons will be the main (Pedersen, J. L. and Kehlet, H. 1998). Importantly,
focus of the remaining sections of this chapter, and secondary heat hyperalgesia is significantly reduced
will be a recurring theme in various chapters of when lidocaine is administered i.v. prior to heat
this volume that address alterations in pain proces- injury (Holthusen, H. et al., 2000), suggesting that
sing associated with injury or disease of peripheral central sensitization plays a role in its initiation.
tissue, primary afferent nerves, or CNS structures. Animal behavioral studies support this conclusion,
as the spread of hyperalgesia to the rat hind paw
342 Spinal Cord Mechanisms of Hyperalgesia and Allodynia

contralateral to the paw that received a thermal develops after intradermal injection of capsaicin, is
injury is unaffected by either deafferentation or anes- maintained even after anesthetizing the region where
thetic blocks of the injured hind paw following the capsaicin was injected (Lamotte, R. H. et al., 1991).
injury, but is prevented if deafferentation or anes- However, if the skin region is anesthetized prior to
thetic block precedes the injury (Coderre, T. J. and capsaicin injection, cutaneous hyperalgesia does not
Melzack, R., 1985; 1987). develop. Furthermore, hyperalgesic responses to
capsaicin can be prevented if the area of skin where
26.1.2.1.(ii) Mechanical injury Mechanical injury the injection is made is rendered anesthetic by a
of the skin also produces both primary and secondary proximal anesthetic block of the peripheral nerve
hyperalgesia, as has been demonstrated in the rat which innervates it. Thus, for hyperalgesia to
plantar incision model (Brennan, T. J. et al., 1996), develop it is critical that initial inputs from the injury
as well as following incision of human skin reach the CNS. However, once hyperalgesia is estab-
(Kawamata, M. et al., 2002a). Although both pre- lished, it does not need to be maintained by inputs
and postincisional administration of intrathecal (i.t.) from the injured peripheral tissue. In support of this,
morphine or bupivacaine reduce secondary hyperal- Torebjork H. E. et al. (1992) have shown that pain
gesia in the rat model (Zahn, P. K. and Brennan, T. J., thresholds to intraneural electrical stimulation of
1999), only preincisional administration of s.c. or i.v. afferent fibers are dramatically reduced following
lidocaine reduce secondary hyperalgesia in humans intradermal capsaicin injection in the skin from
(Kawamata, M. et al., 2002a). These results again which the stimulated nerve emanates: neural stimu-
support a role for central sensitization in the initiation lation which was felt as tactile before administration
of secondary hyperalgesia after mechanical skin injury. of capsaicin, was painful after capsaicin. Importantly,
This conclusion is also consistent with the finding that this reduced pain threshold is evident even when the
rat plantar incisions or other mechanical injuries pro- sensory projected field of the afferent nerve is
duce an increase in the responsiveness of spinal wide anesthetized after the capsaicin injection. Again, a
dynamic range (WDR) neurons to innocuous and state of central sensitization is indicated since once
noxious stimuli, as well as increases in their receptive they have established their effects, inputs from the
fields (Cervero, F. et al., 1988; Kawamata, M. et al., injured region are not required to maintain the low-
2002b). ered threshold.

26.1.2.1.(iii) Capsaicin Capsaicin, the active 26.1.2.2 Peripheral nerve stimulation

ingredient of hot chilli peppers, produces hyperalge- Much of the early evidence that spinal mechanisms
sia in both animals (Gilchrist, H. D. et al., 1996) and contribute to hyperalgesia and allodynia came from
humans (Carpenter, S. E. and Lynn, B., 1981) when studies that used peripheral nerve stimulation as a
injected or applied to the skin. Evidence that capsai- conditioning stimulus, and assessed the influence of
cin-induced hyperalgesia depends on central the conditioning on spinal function or the function of
sensitization initially came from animal studies that spinal output neurons. Early studies showed that
showed that intradermal injections of capsaicin pro- repeated high-frequency C-fiber afferent stimulation
duced enhanced excitability of spinal cord dorsal sequentially increases dorsal horn activity resulting
horn neurons in cats (Simone, D. A. et al., 1989) and in a prolonged discharge of the cell, lasting from
monkeys (Dougherty, P. M. and Willis, W. D., 1992). seconds to minutes poststimulation (Mendell, L. M.
In both monkeys (Baumann, T. K. et al., 1991) and and Wall, P. D., 1965; Mendell, L. M., 1966). This
humans (Lamotte, R. H. et al., 1992), capsaicin injec- phenomenon, which has been labeled as windup, results
tions produce extensive, spreading hyperalgesia from a homosynaptic sensitization (Woolf, C. J. et al.,
without producing the same degree of spreading 1988) of dorsal horn neurons, and likely represents a
sensitization of primary afferent nociceptors. short-term sensitization of dorsal horn neurons that
Indeed, capsaicin caused the spread of both brush underlies the intense reactions and lingering after sen-
allodynia and pinprick hyperalgesia beyond the bor- sations to stimuli at the pain tolerance level. Although
ders of the territory of the nerve which innervates the windup is not synonymous with central sensitization
injected skin area (Sang, C. N. et al., 1996), suggesting (Woolf, C. J., 1996), stimulation that causes windup can
that there is extraterritorial pain that is dependent on produce some of the classical characteristics of central
central sensitization. Furthermore, in humans, hyper- sensitization including expansion of dorsal horn neuro-
algesia to punctate mechanical stimuli, which nal receptive fields and enhanced spinal neuronal
 Spinal Cord Mechanisms of Hyperalgesia and Allodynia 343

responses to C-fiber, but not A-fiber electrical stimula- electrical nerve stimulation (Hylden, J. L. K. et al.,
tion (Li, J. et al., 1999). 1989; Ren, K. et al., 1992). After subcutaneous CFA,
In early studies of central mechanisms of hyperal- there is also an increased ability to induce LTP
gesia, it was shown that noxious electrical stimulation (Vikman, K. S. et al., 2003), expanded receptive fields
of cutaneous and muscle afferent nerves (Woolf, C. J., in rat dorsal horn neurons (Hylden, J. L. K. et al., 1989;
1983; Wall, P. D. and Woolf, C. J., 1984), as well as Ren, K. et al., 1992), and an increase in the excitability
cutaneous (Woolf, C. J., 1983) and deep (Woolf, C. J. of flexor efferent neurons (Seybold, V. S. et al., 2003).
and McMahon, S. B., 1985) tissue injury, produce an
increase in the excitability of the ipsilateral and con- 26.1.2.3.(ii) Joint inflammation Injection of kao-
tralateral flexor efferent nerves in response to lin and carrageenan into the rat knee joint produces
noxious mechanical stimulation of the hind paw. limping and guarding of the limb (Sluka, K. A. and
Because the increased excitability in the contralateral Westlund, K. N., 1993), as well as a sensitization of
flexor efferent nerve is maintained even after inputs peripheral nociceptors (Neugebauer, V. and
from the injured paw are blocked by local anesthesia, Schaible, H. G., 1988). The same stimulus produces
the results suggest that central, not peripheral, enhanced responsiveness of spinal cord dorsal horn
changes underlie this effect. neurons to normal movement or tactile stimulation of
Although typically associated with hippocampal the knee joint (Neugebauer, V. and Schaible, H. G.,
neurons, evidence suggests that the phenomenon 1988; 1990), as well as an expansion of the receptive
long-term potentiation (LTP) can also be elicited in field of dorsal horn neurons that respond to joint
spinal cord dorsal horn neurons after stimulation of input (Neugebauer, V. and Schaible, H. G., 1990).
primary afferents in transverse spinal cord slices There is also an expansion of dorsal horn neuron
(Randic, M. et al., 1993; Lozier, A. P. and Kendig, J. J., receptive fields following the induction of polyarthri-
1995), as well as in vivo after electrical nerve stimula- tis in multiple joints three weeks or more following
tion (Svendsen, F. et al., 1997; 1999) or intense natural CFA injections to the rat tail (Menetrey, D. and
noxious stimulation (Rygh, L. J. et al., 1999). Besson, J. M. 1982; Calvino, B. et al., 1987).
Importantly, LTP induced by electrical nerve stimu-
lation or noxious stimulation is maintained after 26.1.2.4 Peripheral nerve injury
lidocaine block distal to the stimulation site in the Traumatic (Bennett, G. J. and Xie, Y. K., 1988; Seltzer,
sciatic nerve (Svendsen, F. et al., 1999) or proximal to Z. et al., 1990; Kim, S. H. and Chung, J. M., 1992;
the site of noxious stimulation (Rygh, L. J. et al., 1999). Decosterd, I. and Woolf, C. J., 2000), inflammatory
(Eliav, E. et al., 1999; Milligan, E. D. et al., 2003), or
26.1.2.3 Inflammation chemical (Tanner, K. D. et al., 1998; Polomano, R. C.
26.1.2.3.(i) Subcutaneous inflammation Sub- et al., 2000) injury of peripheral nerves induces heat
cutaneous injections of inflammatory substances such and/or mechanical hyperalgesia, or mechanical and/
as carrageenan and complete Freunds adjuvant (CFA) or cold allodynia in animal models. While there is an
in the rat hind paw produce thermal hyperalgesia and established contribution of peripheral pathology to
mechanical allodynia (Buritova, J. et al., 1996; Arevalo, hyperalgesia and allodynia in these models (Chen, Y.
M. I. et al., 2003), as well as a sensitization of peripheral and Devor, M., 1998; Liu, X. et al., 1999), it is also clear
nociceptors (Kocher, L. et al., 1987; Andrew, D. and that central sensitization plays a critical role. Thus, 9
Greenspan, J. D., 1999). There is also evidence of 11 days after chronic constriction injury (CCI) of the
central sensitization associated with peripheral inflam- rat sciatic nerve, lumbar spinal cord dorsal horn neu-
mation. Subcutaneous carrageenan injections produce rons exhibit abnormal characteristics including
increased excitability of rat spinal cord neurons as spontaneous activity, exaggerated, and prolonged
shown by increased responses to mechanical and elec- responses to noxious and innocuous mechanical sti-
trical stimulation, enhanced windup, expanded mulation (Laird, J. M. and Bennett, G. J., 1993), with
receptive fields (Hedo, G. et al., 1999; Torsney, C. prolonged after-discharges more prevalent in WDR
and Fitzgerald, M., 2002), and increased flexor efferent neurons, as opposed to nociceptive-specific neurons
neuron responsiveness (Xu, X.-J. et al., 1995). (Sotgiu, M. L. et al., 1995; Pitcher, G. M. and Henry, J.
Subcutaneous injections of CFA in rats also produce L., 2000). Two weeks after L5/L6 spinal nerve ligation
central sensitization, as evidenced by the enhancement (SNL), rat dorsal horn neurons also show increased
of dorsal horn neurons spontaneous activity and spontaneous firing with irregular firing patterns
increased responsiveness to thermal, mechanical and (Suzuki, R. and Dickenson, A. H., 2005).
344 Spinal Cord Mechanisms of Hyperalgesia and Allodynia

Thresholds for excitatory postsynaptic potentials The enhanced spontaneous or evoked activity of
(EPSPs) of lamina II neurons are dramatically lower dorsal horn neurons likely contribute to hyperalgesia
after CCI, and spared nerve injury (Kohno, T. et al., and allodynia since systemic low-dose lidocaine
2003), and there is a long-lasting posttetanic potentia- reduces both spontaneous activity in WDR neurons
tion of sciatic-evoked A-fiber superficial dorsal horn (Sotgiu, M. L. et al., 1992) and the thermal hyperal-
field potentials that is present over the same time gesia (Abram, S. E. and Yaksh, T. L., 1994) in CCI
course as thermal hyperalgesia in CCI rats rats. Mechanical allodynia induced by partial sciatic
(Draganic, P. et al., 2001). Tight ligation of the L5/L6 nerve lesion or SNL is also attenuated by i.t. treat-
spinal nerve also results in a lower threshold for acti- ment with very low doses of lidocaine that are not
vation of C-fiber-driven dorsal horn field potentials, effective when given systemically (Ma, W. et al.,
and for the induction of LTP in dorsal horn neurons 2003a). The hyperalgesia after CCI depends partly
(Xing, G. et al., 2003). on an injury discharge at the time of surgery, since
Increases in spontaneous and evoked activity in the onset of thermal hyperalgesia in CCI rats was
dorsal horn neurons of CCI rats are paralleled by delayed by treating the nerve with bupivacaine
increases in [14C]-2-deoxyglucose metabolic activity 15 min before, but not 15 min after nerve constriction
throughout the lumbar spinal dorsal horn, with (Yamamoto, T. et al., 1993).
greater increases in the medial portion of the ipsilat-
eral dorsal horn (Mao, J. et al., 1992a). Fos protein 26.1.2.5 Spinal cord injury
expression, often used as a measure of neuronal acti- Animal models of central pain have been generated
vation, is also increased in the spinal cord dorsal horn following traumatic, inflammatory, or excitotoxic
of CCI rats (Ro, L. S. et al., 2004). Importantly, Fos injury of the spinal cord (see Vierck, C. J. and Light,
protein expression is elevated throughout the dorsal A. R., 2000 or Yezierski, R. P., 2000 for reviews).
horn (lamina IVI) of the entire lumbar spinal cord, Traumatic (Christensen, M. D. et al., 1996;
and like [14C]-2-deoxyglucose activity, significant Christensen, M. D. and Hulsebosch, C. E., 1997;
Bruce, J. C. et al., 2002), inflammatory (Cahill, C. M.
but lower increases were found in the contralateral
et al., 1998), and excitotoxic (Yezierski, R. P. et al.,
dorsal horn. Thus, for both the [14C]-2-deoxyglucose
1998) injury of spinal cord produces mechanical
and Fos protein expression measures, there is a
and thermal allodynia and/or hyperalgesia in rat
spread of neuronal activity to spinal regions outside
hind paws. Spinal cord injury also results in a sensi-
of the normal central terminations of the sciatic
tization of spinal cord dorsal horn neurons as
nerve. This may explain why abnormal pain sensa-
indicated by increased spontaneous activity,
tions are sometimes exhibited contralaterally, or in
increased evoked responses to mechanical stimuli,
body areas that do not correspond with the termina-
and increased duration of after-discharges and
tions of an injured peripheral nerve (Koltzenburg, M. enhanced windup (Yezierski, R. P. and Park, S. H.,
et al., 1999; Ro, L. S. et al., 2004). 1993; Christensen, M. D. and Hulsebosch, C. E., 1997;
In addition to enhanced spontaneous and evoked Wang, J. et al., 2005; Zhang, H. et al., 2005). It was
activity, spinal cord dorsal horn neurons of CCI rats recently demonstrated that spinal sensitization
are also found to have expanded receptive fields (enhanced spontaneous and evoked activity and
(Pitcher, G. M. and Henry, J. L., 2000). Rats with enhanced windup in dorsal horn neurons), as well
diabetic neuropathy following treatment with as hind paw thermal and mechanical allodynia, are
streptozotocin are also found to have dorsal horn reduced in spinal cord injured rats that are treated
neurons with high level of spontaneous activity and spinally with bupivacaine at the time of the injury
with expanded receptive fields (Chen, S. R. and (Zhang, H. et al., 2005), suggesting that spinal sensiti-
Pan, H. L., 2002). A recent study measured dorsal zation underlies the development of allodynia/
horn neuronal activity in neuropathic pain patients hyperalgesia after spinal cord injury. Spinal treat-
undergoing surgery for ablation of the dorsal root ment with morphine also reduces the enhanced
entry zone (Guenot, M. et al., 2003). These authors mechanically evoked responses of WDR neurons in
found similar alterations in dorsal horn neuron the spinal cord dorsal horn of spinal cord injured rats
activity including bursting activity (in patients (Wang, J. et al., 2005). Consistent with these findings,
with deafferentation pain after brachial plexus it has recently been shown that central pain (sponta-
avulsion) and nonrandom patterns of spontaneous neous pain, mechanical allodynia, and/or mechanical
discharge (in patients with peripheral nerve injury). hyperalgesia) associated with spinal cord injury in
 Spinal Cord Mechanisms of Hyperalgesia and Allodynia 345

rats is reduced by intravenous lidocaine administra- causes the release of substance P (SP) (Go, V. L.
tion (Attal, N. et al., 2000; Finnerup, N. B. et al., 2005). and Yaksh, T. L., 1987; Oku, R. et al., 1987; Duggan,
A. W. et al., 1988), neurokinin (NK) A (Hua, X. Y.
26.1.2.6 Illness-induced hyperalgesia et al., 1986; Duggan, A. W. et al., 1990), somatostatin
Thermal hyperalgesia is also induced by intraperito- (Kuraishi, Y. et al., 1985; Morton, C. R. et al., 1988),
neal (i.p.) injection of agents that induce illness calcitonin gene-related peptide (CGRP) (Saria, A.
including lithium chloride and the bacterial cell wall et al., 1986), dynorphin 117 (Parra, M. C. et al.,
endotoxin lipopolysaccharide (LPS) (Wiertelak, E. P. 2002), and galanin (Morton, C. R. and Hutchison,
et al., 1994; Watkins, L. R. et al., 1994a). Illness-induced W. D., 1990; Colvin, L. A. and Duggan, A. W. et al.,
hyperalgesia has been shown to be dependent on 1998) in spinal cord dorsal horn. Spontaneous and
spinal mechanisms that are activated by a descending noxious stimulus-evoked release of neuropeptide Y
pain facilitatory pathway (see below) that projects (NPY) is enhanced in the dorsal horn of rats with
from the nucleus raphe magnus (NRM) through the CCI of the sciatic nerve (Mark, M. A. et al., 1998).
dorsolateral funiculus to the spinal cord dorsal horn. Furthermore, the iontophoretic application of SP and
The NRM has also been shown to be activated by other NKs produces an excitation of dorsal horn neu-
brain regions that receive inputs from hepatic vagal rons (Henry, J. L., 1976; Willcockson, W. S. et al.,
afferent nerves (Watkins, L. R. et al., 1994b). 1984a), while i.t. treatment produces behavioral hyper-
algesia (Moochhala, S. M. and Sawynok, J., 1984;
26.1.2.7 Chronic opiate exposure Cridland, R. A. and Henry, J. L., 1986) or nociceptive
Thermal and/or mechanical hyperalgesia are behaviors (Hylden, J. L., and Wilcox, G. L., 1981;
observed in rats after repeated high-dose morphine Seybold, V. S. et al., 1982; Gamse, R. and Saria, A.,
treatment (Kayan, S. et al., 1971; Mao, J. and Mayer, D. 1986) in rodents.
J., 2001) or in some cases after administration of low There is a decrease in immunoreactive staining for
doses of morphine (Holtman, J. R., Jr. and Wala, E. P., the neuropeptides SP and CGRP in the superficial
2005). This phenomenon mirrors a trend that has dorsal horn of rats with experimental arthritis (Sluka,
been observed in clinics in which chronic pain K. A. et al., 1992). While the decrease in these neuro-
patients are treated with morphine ( Jacobsen et al., peptides occurs early (48 h) after knee inflammation,
1995). Mao J. et al. (1995a) and Mayer D. J. et al. (1999) substance P and CRGP are increased after 57 days in
have noted a parallel between hyperalgesia that the dorsal root ganglia and spinal cord of rats with
occurs after repeated morphine exposure and that inflammation of the knee (Sluka, K. A. and Westlund,
produced by neuropathic injury, each causing the K. N., 1993) or hind paw (Donnerer, J. et al., 1993).
activation of similar intracellular signaling pathways Conversely, and spinal cord dorsal horn substance P
and the appearance of dark neurons in spinal cord and CGRP are decreased in the weeks after CCI of
dorsal horn. It is also significant that expression of the the sciatic nerve (Cameron, A. A. et al., 1991). There is
transcription factor Fos, which is commonly used as also a decrease in both mRNA and peptide levels of
an indicator of neuronal activation, is upregulated in SP, somatostatin and CGRP in the DRG after per-
the spinal dorsal horn of rats that are morphine ipheral nerve section (Jessell, T. M. et al., 1979;
dependent and have withdrawal precipitated by injec- Nielsch, V. et al., 1987; Noguchi, K. et al., 1989, 1990,
tion of naltrexone (Rohde, D. S. et al., 1996). 1993; Henken, D. B. et al., 1990; Baranowsli, A. P. et al.,
1993), whereas the levels of cholecystokinin (CCK),
galanin, pituitary adenylate cyclase activating peptide
26.2 Alterations in Spinal Cord (PACAP), vasoactive intestinal polypeptide (VIP)
Dorsal Horn and NPY are increased (Shehab, S. A. and Atkinson,
M. E., 1986; Hokfelt, T. et al., 1987; Villar, M. J. et al.,
26.2.1 Alterations in Transmitter or Growth
1989; Noguchi, K. et al., 1989, 1993; Wakisaka, S. et al.,
Factor Expression or Release
1991, 1992; Zhang, X. et al., 1993b, Zhang, Y. Z., 1996).
26.2.1.1 Neuropeptides Dynorphin mRNA is also increased in the spinal
Several lines of evidence suggest that C-fiber neuro- dorsal horn following traumatic spinal cord injury
peptides are involved in triggering CNS plasticity (Tachibana, T. et al., 1998).
following injury or noxious stimulation. A role for For SP, the decreased immunoreactivity may
C-fiber neuropeptides in nociception is suggested as reflect increased neuropeptide release since in vivo
noxious stimulation or peripheral inflammation microdialysis studies show that spinal SP is increased
346 Spinal Cord Mechanisms of Hyperalgesia and Allodynia

following partial sciatic nerve ligation (Wallin, J. 1976; Randic, M. and Miletic, V., 1977). Dorn horn
and Schott, E., 2002). The increased levels of galanin neuronal LTP induced in vivo by high-intensity,
may play an antinociceptive role as galanin overex- high-frequency tetanic afferent nerve stimulation is
pressing mice have attenuated mechanical and blocked by i.v. administration of NK-1) or NK-2
heat hyperalgesia after partial ischemic injury antagonists (Liu, X. and Sandkuhler, J., 1997).
(Hygge-Blakeman, K. et al., 2004) or CCI Knockout of the preprotackykinin-A gene, from
(Eaton, M. J. et al., 1999) of the sciatic nerve. Spinal which SP and NKA are derived, results in a reduc-
administration of galanin also produces an inhibition tion in the time course over which dorsal horn WDR
of dorsal horn neuronal responses to natural and neurons are sensitized following mustard oil applica-
electrical stimuli in rats with SNL (Flatters, S. J. tion to the mice hind paw, as well as a reduction in
et al., 2002). In contrast, galanin knockout mice have the magnitude of poststimulus discharges of WDR
attenuated responses to hind paw formalin injections neurons (Martin, W. J. et al., 2004).
(Kerr, B. J. et al., 2001a), and have reduced thermal The increased excitability in flexor efferents,
hyperalgesia after intraplantar injection of carragee- induced either by C-fiber electrical stimulation or
nan (Kerr, B. J. et al., 2001a) or partial sciatic nerve by the application of chemical irritants, is blocked
transection (Kerr, B. J. et al., 2000). In addition, both by pretreatment of the sciatic and saphenous nerves
windup and the facilitation of spinal reflexes follow- with the C-fiber neurotoxin capsaicin (Woolf, C. J.
ing conditioning stimulation are significantly and Wall, P. D., 1986) or by the SP antagonist spantide
reduced in galanin knockout mice with sciatic nerve II (Wiesenfeld-Hallin, Z. et al., 1990). I.t. application of
transections (Kerr, B. J. et al., 2001b). Upregulated the C-fiber neuropeptides SP (Woolf, C. J. and
CCK may contribute to mechanical hypersensitivity Wiesenfeld-Hallin, Z., 1986), NK A (Xu, X.-J. et al.,
as allodynia induced by spinal ischemic injury is 1991), CGRP (Woolf, C. J. and Wiesenfeld-Hallin, Z.,
reduced following administration of a CCK-B recep- 1986), VIP (Wiesenfeld-Hallin, Z., 1987), somatosta-
tor antagonist (Wiesenfeld-Hallin, Z. et al., 1997). tin (Wiesenfeld-Hallin, Z., 1985), and galanin
Prodynorphin knockout mice or mice receiving (Wiesenfeld-Hallin, Z. et al., 1989) produces pro-
dynorphin A antisera do not develop mechanical longed enhancements in the excitability of the
and thermal hyperalgesia after SNL (Gardell, L. R. flexion reflex.
et al., 2004), suggesting that upregulated dynorphin In addition, the hyperalgesia that develops in the
contributes to nerve injury-induced hyperalgesia. hind paw contralateral to a thermal injury is mimicked
Indeed, elevated spinal dynorphin content associated by i.t. treatment with SP and NKA, and reversed by
with SNL has been demonstrated to be coincident pretreatment with a SP antagonist (Coderre, T. J. and
with the onset of mechanical allodynia and thermal Melzack, R., 1991). Subcutaneous injection of forma-
hyperalgesia (Malan, T. P. et al., 2000). Although lin, which elicits a persistent nociceptive response
the release of both VIP and NPY are elevated in associated with central changes (Coderre, T. J. et al.,
the deep dorsal horn of rats with spinal nerve trans- 1990), evokes an immediate, intense barrage of C-fiber
ection, the increases do not appear to be related to afferent activity (McCall, W. D. et al., 1996), and
neuropathic pain as increases were observed even produces an increase in SP in the cerebrospinal fluid
in rats that did not exhibit behavioral signs of hyper- (Kuraishi, Y. et al., 1989). Furthermore, nociceptive
algesia (Kim, H. J. et al., 2003). responses to formalin are significantly suppressed in
A role of C-fiber neuropeptides in noxious stimulus- rats pretreated with peptide (Murray, C. W. et al.,
induced plasticity is further suggested by several 1991) and nonpeptide (Yamamoto, T. and Yaksh, T.
findings. Repetitive stimulation of dorsal roots elicits a L., 1991; Yashpal, K. et al., 1993) NK-1 receptor
slow depolarization in dorsal horn neurons which is antagonists, as well as the C-fiber neurotoxin capsaicin
mimicked by SP (Murase, K. and Randic, M., 1984), (Dray A. and Dickenson, A. H., 1991). Finally, various
NK A (Murase, K. et al., 1989), CGRP (Ryu, P. D. nonpeptide NK-1 receptor antagonists attenuate allo-
et al., 1988), VIP (Urban, L. and Randic, M., 1984), or dynia and hyperalgesia that is evoked by CCI of the
CCK (Murase, K. et al., 1987), and is blocked by SP sciatic nerve (Cumberbatch, M. J. et al., 1998; Cahill, C.
antagonists or capsaicin applied to the tissue bath M. and Coderre, T. J., 2002).
(Urban, L. and Randic, M., 1984). Iontophoretic
application of neuropeptides, such as SP, produces 26.2.1.2 Excitatory amino acids
enhanced dorsal horn neuron responses to noxious Additional evidence implicates a contribution of
thermal and mechanical stimulation (Henry, J. L., excitatory amino acids (EAAs) to injury-induced
 Spinal Cord Mechanisms of Hyperalgesia and Allodynia 347

neuroplasticity. EAAs have widespread activity flexor efferents (Woolf, C. J. and Wiesenfeld-Hallin, Z.,
in the CNS including the spinal cord (Watkins, J. C. 1986), while competitive or noncompetitve NMDA
and Evans, R. H., 1981; Davies, J. and Watkins, J. C., antagonists reduce the facilitation of flexion reflexes
1983) and thalamus (Eaton, S. A. and Salt, T. E., induced by electrical (C-fiber) stimulation or cutaneous
1990). The role of EAAs in nociception is suggested application of the chemical irritant mustard oil (Woolf,
since noxious stimulation or peripheral inflammation C. J. and Thompson, S. W., 1991). Hyperalgesia that
causes the release of glutamate and aspartate in develops in the hind paw contralateral to a thermal
spinal cord dorsal horn (Skilling, S. R. et al., 1988; injury is both mimicked following i.t. treatment with
Sorkin, L. S. et al., 1992). The dorsal horn synaptoso- NMDA, and reversed by an NMDA receptor antagonist
mal level of EAAs is increased in rats with CCI of the (Coderre, T. J. and Melzack, R. 1991). Studies in humans
sciatic nerve (Somers, D. L. and Clemente, F. R., (Kristensen, J. D. et al., 1992), show that i.t. treatment
2002), and dorsal horn levels of EAAs are increased with the competitive NMDA receptor antagonist 3-
in rats with a plantar incision (Zahn, P. K. et al., 2002) (2-carboxypiperazin-4-yl)-propyl-1-phosphonic acid
or CCI of the sciatic nerve (al-Ghoul, W. M. et al., (CPP) abolished after-discharges and spreading pain
1993; Kawamata, M. and Omote, K., 1996). and hyperalgesia (symptoms proposed to be associated
Furthermore, iontophoretic application of EAAs pro- with windup) in a patient with neuropathic pain.
duces an excitation of dorsal horn neurons (Curtis, D. A contribution of ionotropic glutamate receptors
R. and Watkins, J. C., 1960; Willcockson, W. S. et al., (iGluRs) to nociceptive processing was originally sug-
1984b; Schneider, S. P. and Perl, E. R., 1988), while i.t. gested by the ability of AMPA/kainate, and
treatment produces both behavioral hyperalgesia and particularly NMDA, receptor antagonists to produce
spontaneous nociceptive behaviors (Aanonsen, L. M. analgesic effects in both phasic and tonic nociceptive
and Wilcox, G. L., 1986; 1987). tests in rats (Cahusac, P. M. et al., 1984; Nasstrom, J. et al.,
A role of EAAs in noxious stimulus-induced plas- 1992). In addition, AMPA antagonists have been found
ticity is further suggested by several findings. The to produce an inhibition of miniature endplate poten-
windup of dorsal horn neuron activity after frequency tials in dorsal horn neurons (Hori, Y. and Endo, K., 1992)
C-fiber stimulation is mimicked by the application and dorsal root potentials evoked by single shock C-
of L-glutamate or N-methyl-D-aspartate (NMDA) fiber stimulation (Thompson, S. W. et al., 1992).
(Gerber, G. and Randic, M., 1989), and blocked by Furthermore, tetanic electrical stimulation of the pri-
application of either competitive (Dickenson, A. H. mary afferent fibers in vitro (Randic, M. et al., 1993) or in
and Sullivan, A. F., 1987; Thompson, S. W. et al., vivo (Liu, X. G. and Sandkuhler, J., 1995) induces a LTP
1990) or noncompetitive (Davies, S. N. and Lodge, of the C-fiber-evoked potentials in dorsal horn neurons
D., 1987; Thompson, S. W. et al., 1990) NMDA antago- that is prevented by superfusion with NMDA
nists. Iontophoretic application of EAAs produces antagonists.
receptive field changes in dorsal horn neurons NMDA antagonists have been particularly effec-
(Zieglgansberger, W. and Herz, A., 1971), as well as tive at reducing persistent pain associated with central
enhanced dorsal horn neuron responses to nonnoxious sensitization. The noncompetitive NMDA antagonist
and noxious mechanical stimulation (Aanonsen, L. M. MK-801 reduces the hyperalgesia which develops in
et al., 1990; Dougherty, P. M. and Willis, W. D., 1991). rats with peripheral neuropathy (Davar, G. et al., 1991;
Dorsal horn neurons that are sensitized following per- Mao, J. et al., 1992c) or adjuvant-induced inflamma-
ipheral tissue injury/inflammation show increased tion (Ren, K. et al., 1992) and reduces autotomy
responsiveness to the iontophoretic application of behavior in rats with peripheral nerve sections
EAAs (Dougherty, P. M. and Willis, W. D., 1992; (Seltzer, Z. et al., 1991). Various competitive and non-
Dougherty, P. M. et al., 1992b), and exhibit a reduction competitive NMDA receptor antagonists reduce the
in responsiveness or sensitization following intrave- enhanced after-discharges, windup, and thermal- and
nous administration of ketamine or iontophoretic mechanical-evoked responses of spinal dorsal horn
application of NMDA antagonists (Schaible, H.-G. neurons in rats with SNL (Leem, J. W. et al., 1996;
et al., 1991), or the administration of -amino-3- Suzuki, R. et al., 2001). MK-801 has been found to
hydroxy-5-methylisoxazole-4-propionic acid (AMPA) reduce the increases in intracellular calcium ion
or NMDA antagonists to dorsal horn neurons by (Ca2) concentration in spinal dorsal horn slices of
reverse microdialysis (Dougherty, P. M. et al., 1992a). rats which had CCI of the sciatic nerve (Kawamata,
I.t. administration of the EAAs L-glutamate or M. and Omote, K., 1996), as well as reducing
L-aspartate produces an increase in the excitability of increased Fos protein expression in response to
348 Spinal Cord Mechanisms of Hyperalgesia and Allodynia

nonnoxious mechanical stimulation of the hind paw R. et al., 1994; 1995) or increased evoked responses
of CCI rats (Kosai, K. et al., 2001). MK-801 also (Palecek, J. et al., 1994) in dorsal horn neurons. 1S,3R-
reduces the adjuvant inflammation-induced expan- ACPD also produces a ventral root (VR) depolariza-
sion of the receptive fields of nociceptive neurons in tion in neonatal rat spinal cord in vitro (Boxall, S. J.
spinal cord dorsal horn (Dubner, R. and Ruda, M. A., et al., 1996; 1998). The VR depolarization produced
1992). In humans, ischemic and postoperative pain is by 1S,3R-ACPD and windup after stimulation
suppressed by subanesthetic doses of ketamine of dorsal roots, or capsaicin application, are blocked
(Maurset, A. et al., 1989), while in the rat the by the mGluR antagonist (/)-alpha-methyl-4-
increased activity in dorsal horn in response to ische- carboxphenylglycine (MCPG) (Boxall, S. J.
mia associated with femoral artery occlusion is et al., 1996).
inhibited by i.t. application of an NMDA antagonist Selective group I mGluR antagonists also reduce
(Sher, G. and Mitchell, D., 1990). Subcutaneous rat dorsal horn neuronal activity evoked by hind paw
injection of formalin evokes an increased release of mustard oil (Boxall, S. J. et al., 1996) or capsaicin
glutamate and aspartate in spinal cord dorsal horn (Neugebauer, V. et al., 1999). Furthermore, dorsal
(Skilling, S. R. et al., 1988), while the sustained horn neuronal activity is increased by spinal admin-
responses of spinal nociceptive cells to noxious per- istration of the group I mGluR agonist
ipheral stimulation produced by subcutaneous dihydroxyphenylglycine (DHPG) (Young, M. R.
formalin injection are reduced by i.t. administration et al., 1997; Neugebauer, V. et al., 1999); although
of selective NMDA antagonists (Haley, J. E. et al., see Chen Y. et al. (2000). DHPG may act primarily
1990). Nociceptive responses to formalin are also at mGluR1, as agonists of mGluR5 (trans-ADA,
both enhanced by pretreatment with the EAA CHPG) have either less (Young, M. R. et al., 1997)
agonists, L-glutamate and L-aspartate, as well as or no (Neugebauer, V. et al., 1999) excitatory effects.
combinations of NMDA AMPA or NMDA However, 1S,3R-ACPD, DHPG, and CHPG all pro-
trans-1-aminocyclopentane-1,3-dicarboxylate (ACPD) duce a long-lasting potentiation of polysynaptic
(Coderre, T. J. and Melzack, R., 1992a), and suppressed EPSPs in dorsal horn cells receiving C-fiber afferent
by pretreatment with NMDA antagonists (Murray, C. input (Zhong, J. et al., 2000). The neuronal activation
W. et al., 1991; Coderre, T. J. and Melzack, R., 1992a). evoked by spinal DHPG, or hind paw application of
Evidence suggests that in addition to iGluRs, meta- mustard oil or capsaicin, is also reduced by mGluR1-
botropic glumate receptors (mGluRs) also play a selective antagonists (Young, M. R. et al., 1997;
significant role in nociception. Unlike iGluRs, which Neugebauer, V. et al., 1999). However, a selective
gate cation channels, mGluRs are coupled by G pro- mGluR5 antagonist 2-methyl-6-(phenylethynyl)-
teins to various intracellular messengers (Pin, J. P. and pyridine (MPEP) attenuates shock-induced VR
Duvoisin, R., 1995). Molecular cloning techniques potentials and windup in isolated spinal cords
have identified eight subtypes of mGluRs to date, (Bordi, F. and Ugolini, A., 2000), as well as reducing
some of which include splice variants (Pin, J. P. et al., the enhanced spontaneous and noxious-stimulus-
1992; Hollmann, M. and Heinemann, S. F., 1994). The evoked activity in spinal cord neurons of rats
eight subtypes of mGluRs are divided into three with CCI of the sciatic nerve (Sotgiu, M. L. et al.,
groups based on sequence homology, signal transduc- 2003). Alternatively, group II and III mGluR
tion mechanisms, and ligand selectivites (Nakanishi, agonists depress spinal reflexes or activity in isolated
S., 1992). Group I includes mGluR1 and 5 which spinal cords (Saitoh, T. et al., 1998; Gerber, G.
stimulate phospholipase C (PLC), leading to protein et al. 2000; Krieger, P. and El Manira, A., 2002),
kinase C (PKC) activation, phosphoinositide (PI) and reverse the hypersensitivity of dorsal horn
hydrolysis, and intracellular Ca2 mobilization neurons after skin capsaicin treatment (Neugebauer,
(Houamed, K. M. et al., 1991; Masu, M. et al., 1991; V. et al., 2000) or SNL (Chen, S. R. and Pan, H. L.,
Abe, T. et al., 1992). 2005).
The nonselective mGluR antagonist L-AP3 Behavioral studies also suggest mGluRs influence
attenuates rat dorsal horn neuronal activity asso- nociception. I.t. quisqualate, an mGluR and AMPA
ciated with repeated mustard oil application receptor agonist, produces hyperalgesia (Aanonsen,
(Young, M. R. et al., 1994; 1995) and knee joint L. M. et al., 1990; Kolhekar, R. and Gebhart, G. F.,
inflammation (Neugebauer, V. et al., 1994). Further- 1994), as well as nociceptive behaviors (Sun, X. and
more, the nonselective mGluR agonists trans-ACPD Larson, A. A. 1991) in rodents. The mGluR agonist
or 1S,3R-ACPD produce a depolarization (Young, M. 1S,3R-ACPD (Fisher, K. and Coderre, T. J., 1996a)
 Spinal Cord Mechanisms of Hyperalgesia and Allodynia 349

produces similar behaviors, implicating mGluRs in mGluR agonists (Fisher, K. et al., 2002), and by anti-
these responses. Furthermore, nociceptive behaviors sense oligonucleotides to mGluR1 and mGluR5
and hyperalgesia are also produced by i.t. DHPG (Fundytus, M. E. et al., 2001). Hyperalgesia after
(Fisher, K. and Coderre, T. J., 1996a; 1998; Dolan, S. SNL is reduced by group I mGluR antagonists
and Nolan, A. M., 2000; Hama, A. T., 2003), but (Dogrul, A. et al., 2000; Zhu, C. Z. et al., 2004; Varty,
importantly, not by the selective mGluR5 agonist G. B. et al., 2005) and group II and III agonists
trans-ADA (Fisher, K. and Coderre, T. J., 1996a). (Simmons, R. M. A. et al., 2002; Chen, S. R. and Pan,
This suggests a pronociceptive role of spinal H. L., 2005). Tactile allodynia after spinal cord injury
mGluR1. Conversely, i.t. administration of trans- is also reduced by the mGluR1 antagonist (RS)-1-
ACPD, and a group II mGluR agonist (L-CCG-I), aminoindan-1,5-dicarboxylic acid (AIDA) (Mills, C.
produce increased mechanical withdrawal thresh- D. et al., 2000), as well as group II and III mGluR
olds, that are blocked by a group II mGluR agonists (Mills, C. D. et al., 2002). Group II
antagonist (Chen, S. R. and Pan, H. L., 2005). This (mGluR3,4) and group III (mGluR4, 6, 7, 8) result
suggests an antinociceptive role for group II mGluRs. in reduced production of cyclic adenosine monopho-
Group I mGluRs are also implicated in the devel- sphate (cAMP) (Schoepp, D. D. et al., 1995; Tanabe,
opment of hyperalgesia and persistent pain associated Y. et al., 1993).
with inflammatory or chemical injury. Thus, nonse-
lective and group I selective mGluR antagonists 26.2.1.3 Other transmitters
(Walker, K. et al., 2001; Zhang, L. et al., 2002; Zhu, The expression of a variety of other transmitters are
C. Z. et al., 2004), and group I mGuR antisense affected following inflammatory or nerve injuries
oligonucleotides (Fundytus, M. E. et al., 2002) reduce that produce hyperalgesia and allodynia. Thus,
hyperalgesia in rats with hind paw inflammation. there is a reduction in the number of gamma-amino-
Furthermore, formalin nociception is enhanced butyric acid (GABA)- and glutamate decarboxylase
by spinal i.t. 1S,3R-ACPD (Coderre, T. J. and (GAD)-, the synthetic enzyme for GABA, immunor-
Melzack, R., 1992a) and DHPG (Fisher, K. and eactive cells in the spinal cord of rats with CCI of the
Coderre, T. J., 1996b), and nociception associated sciatic nerve (Eaton, M. J. et al., 1998). There is also a
with hind paw formalin (Fisher, K. and Coderre, T. reduction in the level of GABA in spinal cord micro-
J., 1996b; Varty, G. B. et al., 2005) and i.p. acetic acid dialysates taken form rats with a partial constriction
injections are reduced by group I mGluR antagonists of the sciatic nerve that exhibit allodynia (Stiller, C.
(Chen, Y. et al., 2000; Zhu, C. Z. et al., 2004). Formalin O., et al., 1996). Recent studies also indicate that there
nociception is also reduced by group II and III is a reduction in endomorphin-2 immunoreactivity in
mGluR agonists (Fisher, K. and Coderre, T. J., the superficial dorsal horn of mice with a partial
1996b; Simmons, R. M. A. et al., 2002), and by enhan- ligation of the sciatic nerve, which paralleled the
cing an endogenous group II mGluR agonist development of thermal hyperalgesia (Smith, R. R.
(Yamamoto, T. et al., 2004). Thus, while Group I et al., 2001).
activation is nociceptive, activation of Group II or
III mGluRs is antinociceptive. Indeed, group I 26.2.1.4 Neurotrophins
mGluR-linked intracellular messengers (i.e., PKC, Neurotrophins (NTs) are growth factors which sti-
IP3 (inositol trisphosphate), and iCa2) contribute mulate neuronal growth in embryonic development,
to nociception (Igwe, O. J. and Ning, L., 1994; Mao, and maintain neuronal viability in adult tissues. NTs
J. et al., 1995b), while events linked to group II/III include nerve growth factor (NGF) that acts selec-
mGluR (i.e., decreased cyclic adenosine monopho- tively at tyrosine kinase receptor A (TrkA), brain-
sphate (cAMP) production) produce antinociception derived neurotrophic factor (BDNF), NT-4, and
(Gereau, R. W. and Conn, P. J., 1994; Shen, J. et al., NT-5, which act at TrkB receptors, and NT-3 that
2000). acts at TrkC receptors in spinal cord. An additional
mGluRs also influence hyperalgesia induced NT, glial-derived neurotrophic factor (GDNF),
by nerve injury. Group I mGluR antagonists reduce binds to the receptor GFR-alpha1. Small diameter
hyperalgesia in CCI-injured rats (Fisher, K. et al., unmyelinated primary afferent fibers generally fall
1998). CCI-induced hyperalgesia is also reduced into two groups: one which contains CGRP, SP, and
by selective mGluR1 and mGluR5 antagonists TrkA receptors, and depends on NGF for its devel-
(Fisher, K. et al., 2002; Zhu, C. Z. et al., 2004), as opment; a second contains the lectin IB-4 and TrkB
well as group II (2R,4R-APDC) and III (L-AP4) receptors, and is dependent on GDNF for its
350 Spinal Cord Mechanisms of Hyperalgesia and Allodynia

development (McMahon, S. B. et al., 1994). produce thermal hyperalgesia, while antisense to

Importantly, these neurotrophic factors are not only TrkB attenuates carrageenan-induced thermal hyper-
required for neuronal development; they continue to algesia in rats (Groth, R. and Aanonsen, L. M., 2002).
be produced by numerous types of cells, produce In contrast, antisense oliginucleotides to NT-3 have
excitatory effects on primary afferent fibers, and are been found to attenuate mechanical allodynia in rats
required to maintain normal neuronal function of with partial sciatic nerve ligation (White, D. M., 2000).
primary afferent fibers. BDNF has been found to be Alternatively, rats intrathecally treated with adeno-
expressed in primary afferent fibers (particularly C virus to produce a spinal cord overexpression of
fibers containing CGRP and SP) (Barakat-Walter, I., BDNF (Eaton, M. J. et al., 2002), or transgenic mice
1996), and to act in the spinal cord to enhance neu- that overexpress NT-3 in muscle (Gandhi, R. et al.,
ronal excitability (Kerr, B. J. et al., 1999; Thompson, S. 2004), each exhibit reduced mechanical allodynia and
W. et al., 1999). thermal hyperalgesia, after CCI of the sciatic nerve or
It has been hypothesized that hyperalgesia after acid injection into the gastrocnemius muscle, respec-
inflammatory injury is influenced by an upregulation tively. Protein and mRNA levels of GDNF and
of BDNF in primary afferent fibers that occurs in GFR-1 are significantly increased in DRGs of rats
response to peripheral stimulation of primary affer- with CCI of the sciatic nerve, and thermal hyperalge-
ent fibers with NGF (Thompson, S. W. et al., 1999). sia in CCI rats is significantly enhanced by i.t.
Thus, inflammation leads to increased production of administration of GFR-1 antisense oligonucleotides,
NGF in peripheral tissue, which stimulates TrkA suggesting that GDNF may act at GFR-1 to produce
receptors on the first group of primary afferent fibers antihyperalgesic effects (Dong, Z. Q. et al., 2005).
(peptidergic fibers) described above (Apfel, S. C.,
2000). The stimulation of TrkA receptors results 26.2.1.5 Phenotype switch
in increased BDNF production in this group of C Evidence suggests that specific primary afferent fibers
fibers (Apfel, S. C. et al., 1996; Michael, G. J. et al., may be able to switch their phenotype following
1997), and precipitates enhanced BDNF release, and intense noxious stimulation. Thus, it has been shown
a subsequent sensitization of dorsal horn neurons that after inflammatory injury A fibers begin to pro-
(Thompson, S. W. et al., 1999). Noxious thermal, duce and release SP (which they normally do not
mechanical, and chemical stimuli, but not innocuous contain) (Neumann, S. et al., 1996); a phenomenon
stimuli, also causes activation of spinal TrkB recep- that has important implications for the development
tors as shown by increased Trk phosphorylation and of mechanical allodynia (which is believed to
increased extracellular signal-regulated kinase mediated largely by A fibers). Evidence suggests
(ERK) activation (see below) in dorsal horn, which that growth factors or NTs may play a critical role
is blocked by sequestering BDNF with a TrKBIgG in the development of these injury-induced phenotype
fusion molecule (Pezet, S. et al., 2002). changes (Woolf, C. J., 1996). The NT BDNF itself has
Importantly, BDNF protein and mRNA expres- been shown to undergo a phenotype switch after nerve
sion is increased in rat DRG and spinal dorsal horn injury as there is a shift in the size distribution of rat
following hind paw injection of CFA (Cho, H. J. et al., DRG neurons that contain BDNF after sciatic nerve
1997a; 1997b). BDNF protein levels are increased lesions, so that large neurons, which do not initially
in DRG and spinal cord dorsal after CCI of the express BDNF, start to express more than small neu-
sciatic nerve (Ha, S. O. et al., 2001; Miletic, G. and rons (Zhou, X. F. et al., 1999a). Large DRG neurons
Miletic, V., 2002), while BDNF mRNA and or pro- also exhibit a phenotype switch when they start to
tein is also increased in uninjured L4 DRG and express both BDNF (Ohtori, S. et al., 2002) and
dorsal horn following L5 SNL (Fukuoka, T. et al., CGRP (Ohtori, S. et al., 2001) after CFA-induced
2001; Ha, S. O. et al., 2001). I.t. administration of inflammation of the lumbar facet joint.
antibodies to BDNF attenuate thermal hyperalgesia
in rats with L5 SNL (Fukuoka, T. et al., 2001), while
26.2.2 Expression of Second Messengers
BDNF antisense olgionucleotides and the BDNF-
and Transcription Factors
scavenging protein TrkBIgG attenuates carragee-
nan-induced thermal hyperalgesia (Kerr, B. J. et al., 26.2.2.1 Second messengers
1999; Groth, R. and Aanonsen, L. M., 2002). A growing body of literature suggests that intracel-
Conversely, i.t. administration of BDNF and NT-4/ lular messengers linked with various ionotropic and
5, which act at TrkB, but not the TrkC agonist NT-3, metabotropic receptors contribute to synaptic
 Spinal Cord Mechanisms of Hyperalgesia and Allodynia 351

plasticity in spinal cord dorsal horn. Ca2 is a key that PKC is increased in the rat spinal cord dorsal
intracellular messenger that is elevated in the cyto- horn after CCI of the sciatic nerve (Mao, J. et al.,
plasm of neurons following influx through receptor- 1995b), and the synaptosomal membrane fraction of
operated and voltage-gated Ca2 channels, or both PKC and PKC-II is increased in the spinal
following release from intracellular stores. cord dorsal of CCI rats (Miletic, V. et al., 2000).
Glutamate and aspartate stimulate the influx of Importantly, PKC knockout mice exhibit reduced
Ca2 through NMDA receptor-operated channels mechanical allodynia and thermal hyperalgesia after
(MacDermott, A. B. et al., 1986). SP produces an partial sciatic nerve ligation (Malmberg, A. B. et al.,
elevation in intracellular Ca2 by mobilizing its 1997a). PKC" is also upregulated in rat DRG follow-
release from intracellular stores (Womack, M. D. ing hind paw injections of carrageenan or CFA, in a
et al., 1988), while both SP (Womack, M. D. et al., manner that is correlated with the development of
1989) and CGRP (Oku, R. et al., 1988) increase Ca2 inflammatory hyperalgesia (Zhou, Y. et al., 2003).
influx through voltage-gated Ca2 channels. Activity Increased neuronal Ca2 infux also activates
at either NK-1 receptors by SP (Mantyh, P. W. et al., phospholipase A2 that triggers the production of
1984), or at metabotropic EAA receptors by gluta- arachidonic acid (Dumuis, A. et al., 1988; 1990),
mate and aspartate (Sugiyama, H. et al., 1987), which is metabolized into various eicosanoids,
stimulates the hydrolysis of inositol phospholipids including prostaglandins (PGs). Further increases in
by activating a polyphosphoinositide-specific PLC. arachidonic acid occur following the breakdown of
PLC is an enzyme which catalyzes the hydrolysis of DAG (Gammon, C. M. et al., 1989), which is pro-
polyphosphatidylinositol into the intracellular mes- duced following stimulation of PLC with activity at
sengers IP3 and diacylglycerol (DAG). Following its various metabotropic receptors. Spinal administra-
production, IP3 stimulates the release of Ca2 from tion of PGE2 enhances the responsiveness and
internal stores; on the other hand, DAG stimulates receptive field sizes of rat spinal dorsal horn neurons
the translocation and activation of PKC. (Vasquez, E. et al., 2001), as well as producing
Gerber G. et al. (1989) showed that activators of mechanical allodynia and thermal hyperalgesia in
PKC enhance the basal and evoked release of gluta- mice (Minami, T. et al., 1994a; 1994b). There is an
mate and aspartate in the spinal cord slice, as well as increase in the levels of PGE2 in CSF of rat after knee
the depolarizing responses of dorsal horn neurons to joint injection of carrageenan/kaolin (Yang, L. C.
exogenous glutamate and NMDA. Furthermore, et al., 1996) or i.t. injection of SP (Hua, X. Y. et al.,
Chen L. and Huang L.-Y. M. (1992) demonstrated 1999a). The breakdown of arachidonic acid into PGs
that PKC increases NMDA-activated currents in in inhibited by nonsteroidal anti-inflammatory drugs
isolated trigeminal cells by increasing the probability (NSAIDs), and i.t. administration of NSAIDs pro-
of channel openings and by reducing the voltage- duces analgesia in animals subjected to i.p. acetic
dependent Mg2 block of NMDA receptor channels. acid or hind paw formalin injections (Yaksh, T. L.,
PKC is involved in synaptic plasticity causing a 1982; Malmberg, A. B. and Yaksh, T. L., 1992a), as
phosphorylation of both group I mGluR (Alaluf, S. well as reducing enhanced flexor reflexes in rats with
et al., 1995) and NMDA receptors (Chen, L. and hind paw injections of CFA (Seybold, V. S. et al.,
Huang, L.-Y. M., 1992). Enhanced spinal PKC activ- 2003). I.t. NSAIDs block thermal hyperalgesia
ity is observed following inflammatory (Yashpal, K. induced by i.t. treatment with NMDA or SP
et al., 1995; 2001; Sweitzer, S. M. et al., 2004b) and (Malmberg, A. B. and Yaksh, T. L., 1992b). There is
nerve injury (Mao, J. et al., 1992b; Miletic, V. et al., also an elevation of the enzyme cyclooxygenase-2
2000; Fundytus, M. E. et al., 2001; Yashpal, K. et al., (COX-2) in the spinal CSF of rats with hind paw
2001) and PKC inhibitors reduce pain induced by carrageenan injections (Ibuki, T. et al., 2003) or in the
hind paw injection of noxious chemicals (Yashpal, K. spared nerve injury model (Broom, D. C., et al., 2004).
et al., 1995; 2001; Li, K. C. and Chen, J., 2003; Furthermore, thermal hyperalgesia associated with
Sweitzer, S. M. et al., 2004b) and nerve injury (Hua, carrageenan injection is attenuated by i.t. treatment
X. Y. et al., 1999b; Miletic, V. et al., 2000; Yajima, Y. with a selective inhibitor of COX-2 (Dirig, D. M.
et al., 2003). PKC also enhances NMDA receptor et al., 1998; Ibuki, T. et al., 2003), but not COX-1
activity (Liao, G. Y. et al., 2001; Skeberdis, V. A. (Yaksh, T. L. et al., 2001). I.t. COX-2 inhibitors are
et al., 2001) and NMDA receptor trafficking to neu- found to relieve hyperalgesia in a model of hind paw
ronal plasma membranes (Lan, J. Y. et al., 2001). PKC inflammation, but not in the spared nerve injury
has several different isoforms, and evidence suggests model (Broom et al., 2004).
352 Spinal Cord Mechanisms of Hyperalgesia and Allodynia

Elevation in intracellular Ca2 in neurons also K. A. and Willis, W. D., 1997), and thermal hyper-
leads to increases in additional protein kinases algesia induced by i.t. injections of NMDA or an NO
such as Ca2/calmodulin-dependent protein kinase donor (Tao, Y. X. and Johns, R. A., 2000). I.t. admin-
(CaMK) (Kennedy, M. B. et al., 1983). There is an istration of a PKG activator produces mechanical
increased expression and phosphorylation of CaMK- allodynia which is absent in PKG-I knockout mice,
II in rat spinal cord dorsal horn after hind paw injec- further implicating a role for spinal PKG in allodynia
tion of capsaicin, while i.t. administration of a (Tegeder, I. et al., 2004a).
CaMK-II inhibitor reduces enhanced dorsal horn Activity at various neuropeptide receptors leads
neuronal responsiveness and behavioral responses to increased levels of adenylate cyclase in spinal cord
after hind paw capsaicin (Fang, L. et al., 2002), as neurons (see Millan, M. J., 1999 for a review). The
well as reducing mechanical allodynia and thermal enzyme adenylate cyclase increases cAMP and its
hyperalgesia in CCI mice (Garry, E. M. et al., 2003). associated kinase, protein kinase A (PKA), which
Mice with a mutation of the alpha-CaMK-II gene, causes membrane phosphorylation that enhances
which prevents autophosphorylation of CaMK-II, Na and Ca2 currents, and decreases K currents
also have diminished nociceptive responses to hind in primary afferent fibers (Vanegas, H. and Schaible,
paw formalin injections (Zeitz, K. P. et al., 2004). H. G., 2001). Enhancing the spinal cord levels of PKA
The binding of Ca2 to calmodulin activates nitric leads to a sensitization of dorsal horn neurons to
oxide (NO) synthase, which generates NO from free noxious mechanical stimulation (Lin, Q. et al., 2002),
L-arginine (Garthwaite, J. et al., 1988), which in turn and i.t. administration of PKA inhibitors attenuate
activates soluable guanylate cyclase and increases the mechanical allodynia induced by hind paw injec-
cGMP (Southam, E. et al., 1991). Intracellularly the tion of capsaicin (Sluka, K. A. and Willis, W. D.,
Ca2 binds to calmodulin which activates 1997). There is also a reduction in thermal hyperal-
NO synthase. NO synthase converts L-arginine to gesia associated with hind paw inflammation in mice
NO, and the released NO stimulates the production with a target mutation of the type I regulatory sub-
of cGMP from soluable guanylate cyclase unit of PKA (Malmberg, A. B. et al., 1997b).
(Synder, S. H., 1992). The levels of neuronal NO Activity of growth factors and various hormones
synthase, as well as NADPHd and [3H]citrulline leads to the activation of a mitogen-activated protein
staining (markers of NO activity), are increased in kinase (MAPK) pathway through stimulation of a
the rat DRG and spinal cord dorsal horn after CCI or GTP-binding protein Ras and protein kinases, such
transection of the sciatic nerve (Cizkova, D. et al., as PKC (Davis, R. J., 1993). A growing body of litera-
2002a; Lukacova, N. et al., 2003), SNL (Steel, J. H., ture suggests that MAPK, including ERK, p38
et al., 1994), or hind paw formalin injection (Lam, H. MAPK, and c-Jun N-terminal kinase ( JNK) contri-
H. et al., 1996). There is also increased cGMP in the bute to spinal nociceptive processing. Recent studies
spinal dorsal horn of rats with CCI of the sciatic have shown ERK activity is observed in lamina I and
nerve that parallels the development of mechanical IIo of the ipsilateral dorsal horn after noxious thermal
allodynia and thermal hyperalgesia (Siegan, J. B. et al., stimulation of the hind paw or C-fiber electrical
1996). In addition, NO synthase inhibitors or an nerve stimulation, but not after nonnoxious thermal
inhibitor of soluable guanylate cyclase reduce the or A fiber electrical stimulation ( Ji, R. R. et al., 1999).
thermal hyperalgesia that develops in rats with ERK activity is also observed in rodent DRG or
chronic nerve constriction injury (Meller, S. T. spinal cord dorsal horn in response to hind paw
et al., 1992) or hind paw carrageenan (Meller, S. T. injections of carrageenan (Galan, A. et al., 2002),
et al., 1994; Osborne, M. G. and Coderre, T. J., 1999). CFA ( Ji, R. R. et al., 2002a, Obata, K. et al., 2003),
Mediators such as NO, which increase neuronal formalin ( Ji, R. R. et al., 1999, Karim, F. et al., 2001), or
cGMP, stimulate the production of cAMP-depen- capsaicin ( Ji, R. R. et al., 1999), or to sciatic nerve
dent protein kinase (PKG) (Scott, J. D., 1991). The transection (Obata, K. et al., 2003), CCI (Obata, K.
spinal dorsal horn levels of PKG1 are increased in et al., 2004a) partial sciatic nerve ligation (Ma, W. and
response to hind paw injection of formalin, and i.t. Quirion, R., 2002) or L5 SNL (Obata, K. et al., 2004b).
administration of PKG inhibitors reduces nocicep- Inhibitors of ERK have been found to reduce noci-
tive responses to formalin (Tao, Y. X. et al., 2000; ception induced by formalin ( Ji, R. R. et al., 1999,
Schmidtko, A. et al., 2003). I.t. treatment with a PKG Karim, F. et al., 2001), and hyperalgesia or allodynia
inhibitor also reverses mechanical allodynia observed after hind paw injection of carrageenan (Sammons,
in rats with a hind paw injection of capsaicin (Sluka, M. J. et al., 2000), CFA ( Ji, R. R. et al., 2002a) or
 Spinal Cord Mechanisms of Hyperalgesia and Allodynia 353

capsaicin (Kawasaki, Y. et al., 2004), and in rats with treatment with a selective Src inhibitor delays the
CCI of the sciatic nerve (Ciruela, A. et al., 2003) or L5 onset of mechanical allodynia and hyperalgesia
SNL (Obata, K. et al., 2004b). Recently, it has been induced by hind paw injection of CFA (Guo, W.
shown that stimulation of group I mGluRs leads to et al., 2002), as well as reducing capsaicin-induced
activation of ERK in spinal cord dorsal horn, and that ERK activation in spinal cord slices (Kawasaki, Y.
antagonists of group I mGluRs produce a reduction et al., 2004). In contrast to the effects of PTKs,
in formalin-stimulated ERK activation (Karim, F. NMDA receptors currents have been shown to be
et al., 2001). Adwanikar, H. et al. (2004) has also inhibited by protein tyrosine phosphatases (PTPs)
shown that ERK inhibitors significantly reduce (Wang, Y. T. et al., 1996). Recent studies have
DHPG-induced spontaneous nociceptive behaviors shown that i.t. administration of PTP inhibitors pro-
(SNBs) in mice that have received a prior hind paw duces an enhancement of mechanical allodynia and
injection of CFA. hyperalgesia induced by hind paw injection of cap-
Recent evidence indicates there is an increase in saicin (Zhang, X. et al., 2003). Studies using selective
phosphorylated p38 MAPK in DRG neurons and inhibitors or antisense oligonucleotides have also
microglia of the DRG and spinal cord following implicated various other kinases including: IkappaB
hind paw injection of formalin (Kim, S. Y. et al., kinase (Tegeder, I. et al., 2004b), cyclin-dependent
2002) or capsaicin (Sweitzer, S. M. et al., 2004a; kinase 5 (Cdk5) (Wang, C. H. et al., 2005), and Rho
Mizushima, T. et al., 2005) or CCI of the sciatic kinase (Tatsumi, S. et al., 2005) in nociception or
nerve (Kim, S. Y. et al., 2002; Obata, K. et al., 2004a). hyperalgesia associated with hind paw formalin or
I.t. treatment with a p38 MAPK inhibitor reverses zymosan injection or CCI of the sciatic nerve.
thermal hyperalgesia produced by hind paw
capsaicin injection (Sweitzer, S. M. et al., 2004a; 26.2.2.2 Transcription factors
Mizushima, T. et al., 2005) or in CCI rats (Obata, K. Noxious stimulation leads to the expression of proto-
et al., 2004a). There is also increased p38 MAPK oncogenes and their protein products which act as
activity in neurons of both the injured L5 and the transcription factors. Hunt S. P. et al. (1987) first
uninjured L4 DRG after L5 SNL, and i.t. treatment demonstrated that the c-fos protein product Fos is
with a p38 MAPK inhibitor reverses mechanical expressed in postsynaptic dorsal horn neurons fol-
allodynia and thermal hyperalgesia in L5 SNL rats lowing noxious thermal or chemical stimulation of
(Obata, K. et al., 2004b). In contrast, JNK activity is the skin. The expression of Fos has also been demon-
increased only in L5 DRG neurons after L5 SNL, strated in rat spinal dorsal horn in response to
and i.t. administration of a JNK inhibitor reverses noxious pinch of the hind paws (Bullitt, E., 1989),
only mechanical allodynia in rats with L5 SNL the injection of formalin (Presley, R. W. et al., 1990) or
(Obata, K. et al., 2004b). JNK activity is also increased carrageenan (Draisci, G. and Iadorola, M. J., 1989)
in the spinal cord dorsal horn of rats with partial into a hind paw or sodium urate crystals into joints
sciatic nerve ligation (Ma, W. and Quirion, R., 2002). (Menetrey, D. et al., 1989), the injection of acetic acid
It is now evident that activity of the NMDA into viscera (Menetrey, D. et al., 1989), the induction
receptor is regulated by the protein tyrosine kinases of polyarthritis with CFA (Menetrey, D. et al., 1989).
(PTKs), Src and Fyn, which phosphorylate Fos expression in the rat spinal dorsal horn is also
the NMDA channel and enhance its activity increased after transection (Chi, S. I. et al., 1993), CCI
(Yu, X. M. et al., 1997; Tezuka, T. et al., 1999; Yu, X. (Kajander, K. C. et al., 1996; Catheline, G. et al., 1999)
M. and Salter, M. W., 1999). Evidence that this pro- or partial ligation (Delander, G. E. et al., 1997) of the
cess might contribute to hyperalgesia after nerve sciatic nerve. Importantly, there is a strong correla-
injury is suggested as phosphotyrosine staining is tion between pain behavior and the number of cells
increased in the spinal cord dorsal horn following expressing Fos (Presley, R. W. et al., 1990). The time
sciatic nerve transection (Eckert, W. A. et al., 1994). course of Fos expression after peripheral inflamma-
Furthermore, i.t. administration of nonselective PTK tion (Draisci, G. and Iadorola, M. J., 1989) or nerve
inhibitors, reduce hind paw mechanical and thermal injury (Delander, G. E. et al., 1997) has been found to
hyperalgesia produced by injection of carrageenan be correlated with the development of hyperalgesia.
and kaolin to the base of the rat tail, or following i.t. Moreover, morphine pretreatment produces a dose-
injection of NMDA (Sato, E. et al., 2003), as well as dependent suppression of Fos expression which cor-
reducing thermal hyperalgesia in rats with partial responds with its analgesic effects (Presley, R. W.
sciatic nerve ligation (Yajima, Y. et al., 2002). I.t. et al., 1990; Tolle, T. R. et al., 1990).
354 Spinal Cord Mechanisms of Hyperalgesia and Allodynia

An increase in phosphorylated cyclic AMP by the finding that destruction of neurons with NK-1
response element binding (pCREB) protein is receptors with the toxin saporin bound to SP
observed in the spinal cord dorsal horn of rats with abolishes the development of mechanical and ther-
partial ligation or CCI of the sciatic nerve (Ma, W. mal hyperalgesia following intraplantar capsaicin
and Quirion, R., 2001; Miletic, G. et al., 2002, 2004b) injection (Khasabov, S. G. et al., 2002). NK-1 knock-
or hind paw injections of capsaicin (Kawasaki, Y. out mice also have attenuated dorsal horn neuronal
et al., 2004; Fang, L. et al., 2005). Elevation in spinal responses to suprathreshold stimuli and a reduction
pCREB associated with nerve injury, hind paw cap- of windup of deep dorsal horn neurons after C-fiber
saicin, or electrical nerve stimulation has been electrical nerve stimulation (Suzuki, R. et al., 2003).
found to be dependent on the activation of PKA, As for other neuropeptide receptors, it has been
PKC, ERK, or CaMK (Kawasaki, Y. et al., 2004; shown that sciatic nerve transection produces a
Miletic, G. et al., 2004b; Fang, L. et al., 2005; marked increase if CCK-B receptor mRNA in the
Miyabe, T. and Miletic, V., 2005). I.t. treatment dorsal root ganglia (Zhang, X. et al., 1993a). Although
with antisense oligonucleotides to CREB reduces there is no change in the expression of the VIP/
the mechanical allodynia in rats with partial sciatic PACAP receptor PAC1, there is a decrease in
nerve ligation (Ma, W. et al., 2003b). VPAC1 and an increase in VPAC2 in the dorsal
Noxious stimulation, inflammatory, or nerve horn following CCI of the sciatic nerve.
injury also leads to the spinal cord expression of
other transcription factors, including Fos B, Jun, Jun 26.2.3.2 Excitatory amino acid receptors
B, Jun C, Jun D (Herdegen, T. et al., 1991a; 1991b; The NMDA receptor has two subunits NMDAR1
Delander, G. E. et al., 1997), NGFI-A/Krox 24, (NR1) and NMDAR2 (NR2), with four splice var-
NGFI-B, serum response factor (Herdegen, T. et al., iants of the NR2 subunit (NR2AD). Two of the
1990; Wisden, W. et al., 1990), FRA-1, FRA-2 NR2 subunits are heavily expressed in the spinal
(Munglani, R. and Hunt, S. P., 1995), AP-1, nuclear cord dorsal horn and have been studied in respect
factor kappa B, octamer factors (Chan, C. F. et al., to plasticity associated with nociception. Recent
2000; Pollock, G. et al., 2005), activating transcription evidence indicates that there is enhanced phosphor-
factor 3 (Tsujino, H. et al., 2000), cyclic AMP respon- ylation of the NR1 subunit of rat dorsal horn neurons
sive element modulator (Naranjo, J. R. et al., 1997), after intradermal injection of capsaicin (Zou, X. et al.,
and peroxisome proliferators-activated receptor- 2000) or after noxious heat stimulation of the hind
alpha (Benani, A. et al., 2004). paw (Brenner, G. J., et al., 2004), effects which
depends on the activation of kinases, including PKA
and/or PKC. Elevated levels of phosphorylated NR1
26.2.3 Alterations in Receptor and Ion
is also found in the spinal dorsal horn of rats with an
Channel Expression, Activity, or Cellular
excitotoxic injury of the spinal cord that causes cen-
Location
tral neuropathic pain (Caudle, R. M. et al., 2003), or
26.2.3.1 Neuropeptide receptors following hind paw inflammation with carrageenan
Recent studies show that there is an increase in the (Caudle, R. M. et al., 2005).
internalization of NK-1 receptors in the spinal cord There is also a prolonged increase in the phosphor-
dorsal horn following electrical nerve or noxious ylation of NR2B subunits in rat spinal dorsal horn after
peripheral stimulation, and that this internalization inflammation induced by hind paw injection of CFA,
is enhanced in rats with inflammatory or nerve inju- that is dependent on activation of PKC and Src kinase
ries (Allen, B. J. et al., 1999). There is also an (Guo, W. et al., 2002), although decreases in rat spinal
upregulation of NK-1 receptors in the spinal cord NR2B phosphorylation have also been observed after
dorsal horn in response to hind paw inflammation hind paw inflammation (Caudle, R. M. et al., 2005).
induced by CFA and following sciatic nerve transec- Increased phosphorylation of NR2B in spinal dorsal
tion (Aanonsen, L. M. et al., 1992). Importantly, horn has also been observed in mice with transected
upregulation of dorsal horn NK-1 mRNA after par- sciatic nerves. The increased phosphorylated NR2B
tial sciatic nerve ligation is significantly correlated was attenuated by administration of a NR2B selective
with the development of thermal hyperalgesia antagonist and in mice lacking Fyn kinase, a Src-
(Taylor, B. K. and McCarson, K. E., 2004). The family kinase (Abe, T. et al., 2005). Recently, the
importance of NK-1 receptors to the development C-fiber-stimulated windup of spinal dorsal horn neu-
of hyperalgesia after tissue injury is also highlighted rons was shown to be attenuated by i.v. administration
 Spinal Cord Mechanisms of Hyperalgesia and Allodynia 355

of a selective NR2B antagonist (Kovacs, G. et al., 2004). receptors to the postsynaptic membrane and synapses
The importance of phosphorylation of spinal NR sub- of spinal neurons that normally do not have func-
units to central sensitization is based on now long tional AMPA receptors. The phenomenom of silent
established evidence that such phosphorylation pro- synapses has been demonstrated in cultured hippo-
duces an enhancement of Ca2 currents at NMDA campal neurons exposed to stimuli that produce LTP
receptor channels (Chen, L. and Huang, L.-Y. M., (Liao, D. et al., 1995), and may contribute to hyper-
1992). Thus, PKC activation, initiated by glutamate- excitability of dorsal horn neurons in persistent pain
or aspartate-induced PLC activity and Ca2 influx conditions (Kerchner, G. A. et al., 1999).
through NMDA channels, leads to further influx of Recent studies have demonstrated alterations in
Ca2 through NMDA channels, creating a positive the expression of various mGluRs in the DRG and
feedback loop for glutamate and aspartate neurotrans- dorsal horn of rats following nerve injury or persis-
mission. As described above, a similar mechanism has tent pain. There is a dramatic downregulation of
been demonstrated for pathways that involve Src or mGluR1 mRNA in rat L5 DRG cells following tibial
Fyn kinase (Yu, X. M. et al., 1997; Tezuka, T. et al., nerve axotomy (Hofmann, H. A. et al., 2001). In con-
1999). trast, mGluR5 immunoreactivity is increased in L4
A recent question concerning synaptic activity at and L5 DRGs of rats with L5 SNL (Hudson, L. J.
glutamate synapses is the issue of possible alterations et al., 2002), and in the rat spinal cord dorsal horn
in the trafficking of glutamate receptor to the plasma after hind paw carrageenan injection (Dolan, S. et al.,
membrane and synapses of postsynaptic neurons. In 2003). There is also increased mGluR3 mRNA in the
recent years it has been established that glutamate dorsal horn of rats with their hind paws exposed to
receptors move between the cell cytoplasm and the ultraviolet irradiation (Boxall, S. J. et al., 1998), and an
plasma membrane, altering the number of receptors upregulation of both mGluR3 and mGluR5 mRNA
that are available to respond to glutamate and aspar- in the dorsal horn of sheep with chronic hindlimb
tate. Generally referred to as receptor trafficking, inflammation (Dolan, S. et al., 2003). Upregulations of
glutamate receptors are inserted into and removed mGluR2, 3, and 5 mRNA in the dorsal horn have also
from the plasma membrane by exocytosis and endo- been reported a sheep model of postsurgical pain
cytosis (or internalization), respectively, and diffused (Dolan, S. et al., 2004). Spinal mGluR1, but not
laterally within the plasma membrane (Collingridge, mGluR5, mRNA is also upregulated in response to
G. L. et al., 2004). Recent studies have implicated spinal cord injury, while group II mGluRs are down-
trafficking of AMPA (Malinow, R. R., and Malenka, regulated (Mills, C. D. et al., 2001).
R. C., 2002), kainite ( Jaskolski, F. et al., 2005), and
NMDA (Carroll, R. C. and Zukin, R. S., 2002) recep- 26.2.3.3 Other receptors
tors as key processes in regulating synaptic plasticity. In addition to effects on neuropeptides and EAA
Alterations in the activity of AMPA receptors are receptors, inflammatory and nerve injuries that pro-
implicated in plasticity associated with nociception. duce hyperalgesia are known to produce alterations
AMPA receptors in lamina IIIIV of the dorsal horn in various other transmitter receptors and ion chan-
are upregulated following dorsal rhizotomy (Carlton, nels in neurons in spinal cord dorsal horn or DRG
S. M. et al., 1998). There are four subunits of AMPA neurons. Sciatic nerve transection produces an upre-
receptors, known as GluR14, some of which have gulation of GABA-A and a downregulation of
splice variants, and GluR1 and GluR2 are predomi- GABA-B receptors in the rat spinal cord dorsal
nant in the spinal cord (Nagy, G. G. et al., 2004). It has horn (Castro-Lopes, J. M. et al., 1995). A similar
recently been shown that there is a PKA- and PKC- downregulation of GABA-B receptors occurs after
dependent upregulation of phosphorylated GluR1 in joint inflammation induced by CFA injection in the
the spinal cord dorsal horn of rats that have received rat knee, but no change in GABA-A receptors was
a hind paw injection of capsaicin or noxious stimula- found (Castro-Lopes, J. M. et al., 1995). There is also a
tion (Fang, L. et al., 2003; Nagy, G. G. et al., 2004). downregulation of -opioid receptor immunoreac-
Recent evidence also indicates that GluR1 receptors tivity in the rat and monkey spinal cord dorsal horn
are trafficked to the plasma membrane of dorsal horn following peripheral nerve axotomy (Zhang, X. et al.,
neurons in response to visceral inflammation (Galan, 1998). In addition, there is an increase in phosphory-
A. et al., 2004), via a CaMK-II-dependent mechanism. lated -opioid receptors following sciatic nerve
This raises the possibility that noxious stimulation ligation (Narita, M. et al., 2004). The reduction in
may lead to a recruitment of functional AMPA number of -opioid receptors or the phosphorylation
356 Spinal Cord Mechanisms of Hyperalgesia and Allodynia

of remaining receptors may explain why morphine- the adaptor proteins known as postsynaptic density
induced antinociception (Ossipov, M. H. et al., 1995; proteins 93 and 95 (PSD-93, PSD-95), which are
Narita, M. et al., 2004) and the presynaptic inhibitory abundantly expressed in the spinal cord dorsal horn
effects of -opioids (Kohno, T. et al., 2005) are (Garry, E. M. et al. 2003; Tao, Y. X. et al., 2003).
reduced in mice and rats with peripheral nerve Knockout of PSD-93 reduces surface NR2A and
injury, although see also Suzuki R. et al. (1999). NR2B, lowers NMDA receptor-mediated postsynap-
There is, however, also an increase in the levels of tic currents and potentials, and reduces hyperalgesia
phosphorylated -opioid receptors in the spinal cord and/or allodynia in mice with SNL or hind paw
dorsal horn of mice with a partial sciatic nerve lesion, injection of CFA (Tao, Y. X. et al., 2003). Antisense
an effect that is not shown in prodynorphin knockout knockdown of spinal cord PSD-93 also reduces
mice that have enhanced tactile allodynia and ther- mechanical allodynia and thermal hyperalgesia in
mal hyperalgesia (Xu, M. et al., 2004). These findings rats with hind paw CFA injection or following per-
suggest that the loss of endogenous dynorphin activ- ipheral nerve injury (Zhang, B. et al., 2003). Knockout
ity at -opioid receptors may also contribute to or knockdown of PSD-95 also reduces hyperalgesia
allodynia and hyperalgesia after nerve injury. and allodynia in CCI mice or in rats with L5 SNL,
In contrast to the effects of opioid receptors, CCI respectively (Tao, F. et al., 2001; Garry, E. M. et al.,
of the sciatic nerve leads to a MAPK-dependent 2003).
upregulation of spinal cannabinoid-1 (CB-1) recep- Homer is an intracellular scaffolding protein that
tors (Lim, G. et al., 2003). The upregulation of spinal is implicated in the intracellular retention and traf-
CB-1 was determined to be responsible for the anti- ficking of group I mGluRs (Roche, K. W. et al., 1999;
allodynic and antihyperalgesic effects of a CB-1 Ango, F. et al., 2000). Homer proteins also interact
agonist in CCI rats, as the reduction in mechanical with the postsynaptic density protein Shank, a PDZ
allodynia and thermal allodynia produced by Win domain-containing protein binding to the PSD-95/
55,212-2 was lost in rats pretreated with an ERK NMDA receptor complex (Naisbitt, S. et al., 1999;
inhibitor (PD98059) that blocked the spinal CB-1 Tu, J. C. et al., 1999). There are increases in the levels
upregulation (Lim, G. et al., 2003). CCI of the rat of both Homer and Shank in spinal cord dorsal horn
sciatic nerve also leads to an upregulation of P2X3 in rats with CCI of the sciatic nerve at the time points
receptors in spinal cord dorsal horn (Novakovic, S. D. when they exhibit mechanical allodynia and thermal
et al., 1999). This upregulation may contribute to hyperalgesia (Miletic, G. et al., 2005). Another scaf-
hyperalgesia and allodynia, as an adenosine kinase folding protein, neurofilament light chain (NFL)
inhibitor produces a greater inhibition of post- protein, is downregulated in the DRG and spinal
discharge, windup, and C-fiber-evoked responses in dorsal horn of rats with inflammation after a hind
spinal cord neurons of rat with SNLs than in sham paw injection of zymosan (Kunz, S. et al., 2004), the
controls. Recent studies also indicate that there is an breakdown of NFL protein and inflammatory hyper-
upregulation of 3 and 5 spinal nicotinic acetylcho- algesia were blocked with an inhibitor of the protease
line (nAcH) receptors following rat SNL (Vincler, M. calpain. The cell adhesion molecule E-cadherin is
and Eisenach, J. C., 2004), and an increase in spinal dramatically downregulated in spinal cord dorsal
melanocortin (MC) receptors after CCI of the rat horn in response to sciatic nerve transection (Brock,
sciatic nerve (Vrinten, D. H. et al., 2000). It is J. H. et al., 2004), while the microtubule associated
expected that upregulated 5 nAcH and MC recep- protein 1B is upregulated (Soares, S. et al., 2002). The
tors may contribute to allodynia after nerve injuries, presynaptic vesicle protein synaptophysin is
as an 5 nAcH antisense olgionucleotides and an MC increased in spinal dorsal horn in parallel with ther-
receptor antagonist significantly attenuate mechan- mal hyperalgesia in CCI rats (Chou, A. K. et al., 2002).
ical allodynia in SNL and CCI rats, respectively
(Vrinten, D. H. et al., 2000; Vincler, M. A. and 26.2.3.5 Ion channels
Eisenach, J. C., 2005). Inflammatory and nerve injuries lead to alterations in
sensory neuron ion channel expression, distribution
26.2.3.4 Scaffolding and adaptor proteins and function that contribute to hyperalgesia and
NMDA receptors bind directly to membrane-asso- allodynia. In particular, evidence indicates that var-
ciated guanylate kinases (MAGUKs) that regulate ious sodium (Na) channels subtypes are altered in
surface and synaptic NMDAR trafficking in the association with inflammation or neuropathy. There
CNS (Ponting, C. P. et al., 1997). MAGUKs include is an upregulation of the Na channel subunit
 Spinal Cord Mechanisms of Hyperalgesia and Allodynia 357

NaV1.3 in DRG neurons or spinal cord dorsal horn channels 1B) in both DRG and spinal dorsal horn
neurons following peripheral nerve axotomy after hind paw injection of carrageenan (Yokoyama,
(Waxman, S. G. et al., 1994), traumatic spinal cord K. et al., 2003) or CCI of the sciatic nerve (Cizkova, D.
injury (Hains, B. C. et al., 2003) and CCI of the sciatic et al., 2002b). Many preclinical studies have shown
nerve (Hains, B. C. et al., 2004), as well as hind paw that antagonists of N-type Ca2 channels reduce
injection of carrageenan (Black, J. A. et al., 2004). In allodynia or hyperalgesia associated with inflamma-
the case of CCI, a spinal upregulation likely contri- tory or nerve injuries, and suggest that CaV2.2 is a
butes to hyperalgesia/allodynia, as i.t. administration key target (see Winquist, R. J. et al., 2005 for a review).
of antisense oligonucleotides to NaV1.3 decreased There is also a reduction of tactile allodynia and
expression to normal levels and decreased the thermal hyperalgesia associated with SNL in
enhanced spontaneous activity and evoked responses CaV2.2 knockout mice (Saegusa, H. et al., 2001).
of dorsal horn neurons, as well as reducing thermal Although the P-type Ca2 channel have also been
hyperalgesia and mechanical allodynia in CCI rats implicated in the development of hyperalgesia, the
(Hains, B. C. et al., 2004). effects of their antagonists have been less effective
There is also an upregulation of NaV 1.7 and (Sluka, K. A., 1998; Matthews, E. A. and Dickenson,
NaV1.8, but not NaV1.9, in DRG in response to A. H., 2001), and a selective Q-type antagonist was
hind paw injection of carrageenan (Tanaka, M. et al., ineffective (Nebe, J. et al., 1997). In addition, others
1998; Black, J. A. et al., 2004). Hyperalgesia in have found in response to axotomy or CCI of the
response to hind paw inflammation is reduced or sciatic nerve there is a downregulation of CaV1.2 (L-
delayed in rats treated with NaV1.8 antisense oligo- type 1C), CaV1.3 (L-type 1D), CaV3.2 (T-type
nucleotides (Khasar, S. G. et al., 1998) or in NaV1.8 1H) and CaV3.3 (T-type 1I) mRNA expression
knockout mice (Akopian, A. N. et al., 1999). Although in rat DRG (Kim, D. S. et al., 2001). A role for T-type
NaV1.8 channels are not upregulated in DRG after channels in hyperalgesia is suggested since antisense
nerve injury (Okuse, K. et al., 1997; Novakovic, S. D. knockdown of CaV3.2 reduces both hyperalgesia and
et al., 1998), antisense oligonucleotide knockdown of allodynia in rats with CCI of the sciatic nerve
NaV1.8 attenuates neuropathic pain in rats with SNL (Bourinet, E. et al., 2004).
(Lai, J. et al., 2002; Gold, M. S. et al., 2003). The Recent studies also show there is an upregulation
effectiveness of the knockdown may be explained of 2 Ca2 channel auxillary subunits found in the
by findings that there is a redistribution of NaV1.8 DRG and spinal dorsal horn of rats after SNL and to
channels in injured nerves with an accumulation of a lesser extent CCI of the sciatic nerve (Luo, Z. D.
these channels at the site of the nerve injury et al., 2001; 2002). The upregulation of 2 subunits is
(Novakovic, S. D. et al., 1998; Gold, M. S. et al, 2003). temporally related to the development of tactile allo-
As for NaV1.9, it has been shown that knockdown of dynia after SNL, and i.t. treatment of antisense
its protein has no influence on allodynia or hyperal- oligonucleotide to 2 diminishes the tactile allody-
gesia in SNL rats (Porreca, F. et al., 1999), while nia in SNL rats (Li, C. Y. et al., 2004). The importance
disruption of the SCN11A gene, which encodes of this subunit to hyperalgesia and allodynia is also
NaV1.9, attenuates thermal hyperalgesia and sponta- highlighted by the fact that the 2 subunit is the
neous pain after peripheral inflammation (Priest, B. T. only known binding site for gabapentin and pregaba-
et al., 2005). lin which are effective antihyperalgesic agents for
Ca2 channels also show changes in expression many neuropathic pain patients (Frampton, J. E. and
following nerve injury. Thus, after axotomy or partial Foster, R. H., 2005; Wiffen, P. J. et al., 2005).
nerve ligation of the sciatic nerve there is an upre- There is considerable evidence that sensory neu-
gulation of CaV1.2 (L-type 1C) and CaV2.3 (R-type ron vanilloid receptors, including TRPV1 (VR-1)
1E) (Yang, L. et al., 2004; Dobremez, E. et al., 2005). and TRPV2 (VRL-1), play a critical role in the
Evidence suggests that L-type Ca2 channels contri- transduction of noxious heat (Caterina, M. J. and
bute to tonic nociception (Del Pozo, E. et al., 1987; Julius, D., 2001). An up-regulation of TRPV1 has
Coderre, T. J. and Melzack, R., 1992b), and recent also been found in the spared L4 DRG following
evidence indicates that CaV2.3 knockout mice have L5 SNL (Hudson, L. J. et al., 2001) and CCI
reduced responses to inflammatory pain (Saegusa, H. (Wilson-Gerwing, T. D. et al., 2005). Importantly,
et al., 2000), suggesting that R-type Ca2 channels the upregulation of L4 DRG TRPV1 correlates tem-
may also play a role in hyperalgesia/allodynia. There porally with the development and maintenance of
is also an upregulation of CaV2.2 (N-type Ca2 thermal hyperalgesia after L5 SNL (Fukuoka, T.
358 Spinal Cord Mechanisms of Hyperalgesia and Allodynia

et al., 2002). I.t. administration of NT-3 reduces ther- the uptake of glutamate is significantly reduced in
mal hyperalgesia as well as reducing the elevated the spinal cord dorsal horn of rats with SNL (Binns,
DRG TRPV1 expression in CCI rats (Wilson- B. C. et al., 2005), illustrating the impact of glutamate
Gerwing, T. D. et al., 2005). Systemic treatment transporter suppression.
with a TRPV1 antagonist reduces thermal hyperal-
gesia induced by hindpaw capsaicin, carrageenan or 26.2.4.2 Cation chloride transporters
CFA, as well as mechanical hyperalgesia after partial Recent studies suggest that alterations in the function
sciatic nerve lesion in the guinea pig (Walker, K. M. of cation chloride cotransporters may play a role in
et al., 2003), and carrageenan-induced thermal hyper- hyperalgesia states by modulating the chloride con-
algesia is absent in TRPV1 knock-out mice (Davis, J. centration inside nociceptive cells. Galan A. and
B. et al., 2000). In contrast, i.t. administration of a Cervero F. (2005) report that there is a 50% increase
TRPV1 antagonist is less effective at reducing nox- in the NaClK isoform 1 (NKCC1) in the
ious stimulus-evoked dorsal horn neuronal responses dorsal horn of mice after intracolonic capsaicin injec-
in SNL rats as compared to sham controls (Kelly, S. tion, as well as an increase in the phosphorylation of
and Chapman, V., 2002). Evidence suggests that there NKCC1. In addition, NKCC1 knockout mice have a
is also an up-regulation of TRPV1 in DRG asso- reduction in mechanical allodynia to stroking
ciated with peripheral inflammation, and that the induced by intradermal capsaicin (Laird, J. M. et al.,
upregulation is associated with thermal hyperalgesia 2004). They propose that increased NKCC-1 shifts
and depends on p38 MAPK activation (Ji, R. R. et al., the anion gradient and alters normal presynaptic
2002b). Recent evidence indicates that while the inhibition in dorsal horn by producing an increase
levels of TRPV2 receptors in DRG are not affected in the intracellular concentration of chloride and an
by peripheral nerve axotomy, their levels in sympa- enhanced GABA-mediated primary afferent depolar-
thetic post-ganglionic neurons are upregulated after ization that leads to mechanical allodynia (Galan, A.
nerve sections (Gaudet, A. D. et al., 2004). and Cervero, F., 2005). Thus, GABAergic depolari-
zation in primary afferents will lead to a cross
excitation between low- and high-threshold afferents
26.2.4 Alterations in Transporter Activity
that produce allodynia by reducing GABA-mediated
26.2.4.1 Glutamate transporters presynaptic inhibition (Price, T. J. et al., 2005).
Glutamate transporters are critical for the reuptake Furthermore, Coull J. A. et al. (2003) found a reduc-
of transmitter in glutamate synapses, and the major tion in the potassium chloride exporter (KCC2) in the
ones include vesicular glutamate transporters spinal dorsal horn of CCI rats. This decrease was par-
VGluT1-3 and EAAC1 and glial glutamate transpor- alleled by a shift in the transmembrane anion gradient
ters (GLAST and GLT-1) (Shigeri, Y. et al., 2004). that caused normally inhibitory anion synaptic currents
There is a downregulation of EAAC1, GLAST, and to be excitatory, and an increase in the net excitability
GLT-1 is the spinal cord dorsal horn of CCI rats at of lamina I dorsal horn neurons in CCI rats (Coull, J. A.
time points when they exhibit mechanical allodynia et al., 2003). In addition, blockade or knockdown of
and thermal hyperalgesia (Sung, B. et al., 2003). The spinal KCC2 in intact rats produced mechanical allo-
EAAC1, GLAST, and GLT-1 downregulation in rats dynia, suggesting that disruption of anion homeostatis
with taxol-induced neuropathy is paralleled by could be responsible for allodynia in neuropathic rats
increases in spontaneous activity, prolonged after- (Coull, J. A. et al., 2003). The decrease in spinal KCC2 is
discharges, and windup in dorsal horn neurons proposed then to result in a loss of GABA-/glycinergic
(Cata, J. P. et al., 2005; Weng, H. R. et al., 2005). inhibitory tone and, in some cases, inverting its action
Dorsal horn EAAC1, GLAST, and GLT-1 are also into net excitation that causes allodynia after nerve
all downregulated in chronic morphine-treated rats injury (Price, T. J. et al., 2005).
in a manner that is temporally correlated with the
development of thermal hyperalgesia (Mao, J. et al.,
26.2.5 Glial Cell Alterations
2002a; Lim, G. et al., 2005). I.t. administration of
inhibitors of glutamate transporters has also been 26.2.5.1 Microglia
shown to produce mechanical allodynia and thermal Activation of microglial and the release of proinflam-
hyperalgesia (Mao, J. et al., 2002a; Liaw, W. J. et al., matory cytokines from these cells was originally
2005). Finally, using in vivo voltametry and found to play a significant role in illness-induced
pressure-ejected glutamate, it has been shown that hyperalgesia, including that produced by bacterial
 Spinal Cord Mechanisms of Hyperalgesia and Allodynia 359

endotoxins and viruses such as HIV-1 (see Watkins, subtype of ATP receptors, is also enhanced in rat
L. R. and Maier, S. F., 1999 for a review). There is spinal microglia after SNL, and ATP acting at P2X4
also an increase in activated microglia, as indicated receptor on microglia leads to the release of cytokines
by increased complement receptor C3bi (OX-42) which activate p38 MAPK (Inoue, K., 2006).
immunoreactivity in rat spinal cord dorsal horn fol- Pharmacological blockade or knockdown of P2X4
lowing sciatic nerve transection (Eriksson, N. P. et al., receptors reverse mechanical allodynia after SNL in
1993). There is an increase in immunoreactivity for rats, while i.t. administration of microglia activated by
microglial markers OX-42 and histocompatibility P2X4 receptor stimulation produces mechanical allo-
complex II (OX-6), as well, in the dorsal horn of dynia in nave rats (Tsuda, M. et al., 2003).
rats with partial sciatic nerve ligation (Coyle, D. E.,
1998), with a linear correlation between increased 26.2.5.2 Astrocytes
mechanical allodynia and increased OX-42 staining. There is also an increase in activated astrocytes as
OX-42 is also increased in dorsal horn following indicated by increased staining of glial fibrillary
injury of spinal nerve roots, and is accompanied by acidic protein (GFAP) in rat spinal cord dorsal horn
increases in expression of the interleukin 1 (IL-1) following sciatic nerve transection (Eriksson, N. P.
(Hashizume, H. et al., 2000) and the fractalkine recep- et al., 1993) or partial sciatic nerve ligation (Ma, W.
tor X3CR1 in dorsal horn microglia (Verge, G. M. and Quirion, R., 2002). The degree of thermal hyper-
et al., 2004; Lindia, J. A. et al., 2005). Peripheral inflam- algesia or mechanical allodynia in rats with CCI of
mation induced by hind paw CFA injections also the sciatic nerve (Garrison, C. J. et al., 1991) and
leads to increases in microglial markers, including injury of spinal nerve roots (Hashizume, H. et al.,
OX-42, Mac-1, TLR4, and CD14 in spinal dorsal 2000) or partial sciatic nerve injury (Coyle, D. E.,
horn (Raghavendra, V. et al., 2004). I.t. administration 1998), respectively, is correlated with the magnitude
of an inhibitor of microglial cell activation attenuates of GFAP staining in the spinal cord dorsal horn.
mechanical allodynia in rats with sciatic nerve GFAP is also increased in dorsal horn following
inflammation, as well as reducing the expression of SNL, and is accompanied by increases expression
the proinflammatory cytokines IL-1 and tumor the cytokine fibroblast growth factor-2 (Madiai, F.
necrosis factor alpha (TNF) and their converting et al., 2003) and increases in fractalkine (CX3CL1) in
enzymes in spinal dorsal horn after sciatic nerve astrocytes (Lindia, J. A. et al., 2005). Hind paw injection
inflammation (Milligan, E. D. et al., 2003; Ledeboer, of CFA also produces spinal astrocyte activation as
A. et al., 2005). A glial metabolic inhibitor also reduces indicated by increased GFAP and S100B expression
C-fiber afferent-stimulated LTP in spinal cord dorsal in the dorsal horn (Raghavendra, V. et al., 2004). There
horn neurons (Ikeda, H. and Murase, K., 2004). is also an increase in MAPK activity in astrocytes as
Mechanical allodynia in ipsilateral and contralat- indicated by increases in phosphorylated ERK and
eral hind paws of rats with a unilateral inflammation JNK in cells that colocalize GFAP (Ma, W. and
of the sciatic nerve is also reduced following i.t. Quirion, R., 2002).
treatment with antagonists of the proinflammatory
cytokines IL-1, IL-6, and TNF (Milligan, E. D. 26.2.5.3 Neuronglial interactions
et al., 2003). I.t. administration of the cytokine frac- Many of the factors released by glial cells discussed
talkine produces mechanical allodynia and thermal in the two above sections (particularly cytokines) act
hyperalgesia that is blocked by an inhibitor of micro- to sensitize neurons and represent neuronglial
glial activation, as well as an antagonist of IL-1 and interactions that contribute to hyperalgesia and/or
antibodies to IL-6 (Milligan, E. D. et al., 2004; 2005). allodynia. A recent study by Coull J. A. et al. (2005)
In contrast, a neutralizing antibody of the fractalkine provides an excellent example of a neuronalglial
receptor (CX3CR1) delays the development of interactive pathway that has been implicated in the
mechanical allodynia and thermal hyperalgesia in development of allodynia and hyperalgesia in nerve-
rats with CCI or inflammatory injury of the sciatic injured rats. These authors have shown that ATP-
nerve (Milligan, E. D. et al., 2004). Increased cytokine stimulated microglia cause a shift in the anion
activity in spinal nerve injured rats leads to a phos- reversal potential in spinal lamina I neurons similar
phorylation of p38 MAPK in dorsal horn microglia, to that observe in rats with sciatic nerve injury. The
and a p38 MAPK inhibitor reduces mechanical allo- shift inverts the polarity of GABA currents so that
dynia in neuropathic rats (Inoue, K. et al., 2003; Tsuda, GABA produces excitation rather than the normal
M. et al., 2004). The expression of the P2X4 receptor, a inhibition of lamina I neurons. In addition, they
360 Spinal Cord Mechanisms of Hyperalgesia and Allodynia

showed that applying BDNF mimics the alteration in 26.2.7 Anatomical Changes
anion gradient, that ATP stimulation evokes the
26.2.7.1 Cell death and apoptotic genes
release of BDNF from microglia, and that blocking
26.2.7.1.(i) Cell death Transection of the sciatic
BDNF effects on TrkB receptors reverses allodynia
nerve results in transganglionic degeneration atrophy
and anion gradient shifts that follow nerve injury
and signs of Wallerian degeneration in the monkey
or administration of ATP-stimulated microglia
spinal cord (Knyihar-Csillik, E. et al., 1987). In rats,
(Coull, J. A. et al., 2005). This study illustrates the
there is a dramatic increase of terminal deoxynucleo-
importance of neuronglial interactions to allodynia
tidyl transferase-mediated biotinylated UTP nick end
in neuropathic rats, and indicates that BDNF is a
labeling (TUNEL)-stained interneurons in superfi-
crucial signalling molecule between microglia and
cial (Azkue, J. J. et al., 1998) and deep (Oliveira, A. L.
neurons.
et al., 1997) dorsal horn neurons after sciatic nerve
transection, indicating that apoptotic mechanisms are
26.2.6 Descending Facilitation involved in the cell death process. CCI of the rat
Sensitization of spinal cord dorsal horn neurons is not sciatic nerve also leads to degenerative changes in
only enabled by increased input from primary affer- spinal cord with the appearance of hyperchromatic
ent nociceptors, but also by a decrease in descending or dark neurons (Sugimoto, T. et al., 1990;
inhibitory controls and a phenomenon known as Nachemson, A. K. and Bennett, G. J., 1993) and
descending facilitation. The role of descending facil- TUNEL-labeled neurons (Whiteside, G. T. and
itation was reported by Watkins L. R. et al. (1994b) Munglani, R., 2001; Maione, S. et al., 2002) in the
who found lesions of the NRM reduced illness- dorsal horn. The apoptotic degeneration after nerve
induced hyperalgesia. Pertovaara, A. (1998) further injury appears to depend on EAA-induced excitotoxi-
reported that mustard oil application to the rat hind city since premptive treatment with NMDA or
paw produced a facilitation of evoked nociceptive mGluR5 antagonists reduces TUNEL staining after
responses of dorsal horn neurons that was attenuated transection (Azkue, J. J. et al., 1998) or CCI
by spinal transection or lidocaine block of the ros- (Whiteside, G. T. and Munglani, R., 2001; de
troventromedulla (RVM). Descending facilitation of Novellis, V. et al., 2004) of the sciatic nerve.
spinal neurons has also been observed through a Reducing neuronal apopotosis in spinal dorsal horn
pathway that involves the nucleus reticularis gigan- by administration of erythroprotein also prevents
tocellularis (Zhuo, M. and Gebhart, G. F. 1992). mechanical allodynia in rats with spinal nerve root
Spinal cord transection or RVM blocks, as well as crush injury (Sekiguchi, Y. et al., 2003).
inhibition of RVM NMDA receptors or NO Recent evidence suggest that there is a decrease in
synthase, also prevent the development of secondary GABA-A-mediated inhibitory postsynaptic poten-
hyperalgesia in rats after topical mustard oil, carra- tials (IPSPs) in rats with spared nerve injury or CCI
geenan inflammation and SNL (see Urban, M. O. and (Moore, K. A. et al., 2002), suggesting a reduction in
Gebhart, G. F., 1999). Mechanical allodynia after the presynaptic release of GABA after nerve injury.
SNL in rats is reversed by ipsilateral dorsal column Partial nerve injured was also found to produce a
lesions of by administration of lidocaine into the reduction in the GABA synthesizing enzyme gluta-
ipsilateral nucleus gracilis, suggesting that descend- mic acid decarboxylase (GAD) as well as increased
ing facilitation from supraspinal sites depends on TUNEL staining indicative of apoptosis in dorsal
afferent signals transmitted by large myelinated horn (Moore, K. A. et al., 2002), although it has been
fibers in the dorsal column pathway (Sun, H. et al., recently suggested that most TUNEL staining is in
2001). Importantly, large myelinated fibers projecting microglia and not neurons (Polgar, E. et al., 2005).
to the nucleus gracilis have been found to change Blocking apoptosis with a caspase inhibitor prevents
their phenotype and express SP following sciatic the loss of spinal GABAergic interneurons, the
nerve transection, and NK-1 receptor antagonists reduction in spinal IPSCs, and mechanical and cold
suppress stimulus-induced Fos expression in the allodynia after CCI or partial sciatic nerve injury
gracile nucleus in sciatic nerve lesioned rats (Scholz, J. et al., 2005). Furthermore, rats with SNL
(Noguchi, K. et al., 1995). It has also been suggested have alterations in the normal inhibition of
that a descending facilitatory serotonergic pathway is C-fiber-evoked activity by GABA-A agonists
activated by superficial dorsal horn neurons that (Kontinen, V. K. and Dickenson, A. H., 2000; Sokal,
express NK-1 (Suzuki, R. et al., 2002). D. M. and Chapman, V., 2003), and spinal cord
 Spinal Cord Mechanisms of Hyperalgesia and Allodynia 361

injured rats with allodynia do not exhibit the normal neurotoxicity, reduces the expression of dark neu-
increases in evoked responses of dorsal horn neurons rons and allodynia and hyperalgesia in CCI rats
after ionotophorectic application of GABA-A (Mao, J. et al., 1997). Galectin-1, a member of the
antagonists (Drew, G. M. et al., 2004). In addition, a family of -galactoside-binding animal lectins, has
reduction in the glycinergic inhibitory control of been shown to be a proapoptotic factor, and is
spinal neurons occurs after CFA-induced inflamma- increased in spinal cord dorsal horn after peripheral
tion of the hind paw as reflected by a reduced mean nerve transection (Imbe, H. et al., 2003; McGraw, J.
frequency of glycine-mediated miniature IPSCs in et al., 2005). I.t. treatment with antibodies to galectin-
lamina I neurons (Muller, F. et al., 2003). All these 1 attenuates the mechanical allodynia in the spared
findings point to a potentially important contribution nerve injury model (Imbe, H. et al., 2003).
to hyperalgesia and/or allodynia of cell death of
inhibitory neurons. 26.2.7.2 Sprouting
26.2.7.2.(i) Sympathetic fibers Although contro-
26.2.7.1.(ii) Apoptotic genes Recent research versial, there is long-standing evidence that
suggests that chronic pain produces alterations in alterations in sympathetic nervous system function
apoptotic and antiapoptotic genes either following plays a role in hyperalgesia and allodynia following
gene transcription or by posttranslational mechan- nerve injury (Janig, W. et al., 1996). Sympathetic
isms. CCI of the sciatic nerve leads to alterations of activity may influence inputs to spinal cord dorsal
dorsal horn levels of various members of Bcl-2 gene horn following the development of abnormal con-
family, including an upregulation of the proapoptotic nectivity between sympathetic efferent fibers and
gene Bax and the antiapoptotic genes Bcl-2 and Bcl- primary afferent fibers. McLachlan E. M. et al.
xL, and a downregulation of Bcl-xS (Maione, S. et al., (1993) showed that sympathetic axons sprout into
2002; de Novellis, V. et al., 2004). Importantly, spinal the DRGs of rats with sciatic nerve transection and
neuron apoptosis indicated by TUNEL staining, form basket-like structures around large neurons.
thermal, and mechanical hyperalgesia and the Similar sympathetic fiber sprouting is observed after
alterations in apoptotic genes were all reversed SNL (Chung, K. et al., 1993) and partial ligation
by treatment with an mGluR5 antagonist (Lee, B. H. et al., 1998) or CCI of the sciatic nerve
(de Novellis, V. et al., 2004). There is also an upregu- (Ramer, M. S. and Bisby, M. A., 1997). Mechanical
lation of the proapoptotic protein Bax and caspase 3, and allodynia in SNL rats is abolished after sym-
and a downregulation of the antiapopototic protein pathectomy, which eliminates most sprouting fibers
Bcl-2 in chronic morphine-treated rats that exhibit (Chung, K. et al., 1996). Sympathetic sprouting,
thermal hyperalgesia (Mao, J. et al., 2002b). Neuronal mechanical allodynia, and thermal hyperalgesia
apopotosis in dorsal horn and thermal hyperalgesia in after CCI depends on the occurrence of Wallerian
chronic morphine-treated rats are attenuated by i.t. degeneration (Ramer, M. S. et al., 1997) and an upre-
administration of a nonselective caspase and a cas- gulation of NGF (Ramer, M. S. and Bisby, M. A.,
pase-3-selective inhibitor (Mao, J. et al., 2002b). I.t. 1997) in DRG neurons. There is also and upregula-
quisqualic acid-induced excitotoxic spinal cord tion of NGF and NT-3 in glial cells of the DRG in
injury produces pain-related behaviors and increased SNL rats, and antibodies to NGF, BDNF, and NT-3
expression of CD-95 or Fas ligand and TNF-related reduce sympathetic sprouting in rat DRG and ther-
apoptotis-inducing ligand (TRAIL), and i.t. treat- mal hyperalgesia after SNL (Zhou, X. F. et al., 1999b;
ment with IL-10, an anti-inflammatory cytokine, Deng, Y. S. et al., 2000). Sympathetic sprouting and
reduced pain-related behaviors in spinal injured rats mechanical allodynia after SNL are attenuated in IL-
(Plunkett, J. A. et al., 2001). Furthermore, inhibition of 6 knockout mice, implicating cytokines as key factors
activator (1,2,8, and 9) and effector (3) caspases contributing to these changes (Ramer, M. S. et al.,
attenuates neuropathic pain induced in rats by cancer 1998).
and HIV/AIDS chemotherapeutic agents, as well
as pain-related behavior induced by TNF 26.2.7.2.(ii) A fibers It it difficult to explain the
(Joseph, E. K. and Levine, J. D., 2004). A caspase development of allodynia in response to activity-
inhibitor also reduces mechanical and cold allodynia drive plasticity as A fibers, which transmit low-
in rats with partial nerve injury (Scholz, J. et al., 2005), threshold touch inputs generally do not terminate
while an inhibitor of poly(ADP-ribose) synthetase, an on spinal nociceptive neurons. Evidence generated
enzyme involved in glutamate-induced over the past 15 years suggests, however, that
362 Spinal Cord Mechanisms of Hyperalgesia and Allodynia

allodynia may be partly explained the nerve injury- and astrocytes and the release of mediators such as
induced sprouting of the terminals of A fibers into cytokines and neurotrophins result in neuronglial
dorsal horn lamina I and IIo where they are able to interactions that also contribute to increased excit-
contact spinal nociceptive neurons (Woolf, C. J. et al., ability in the spinal cord. Finally there is evidence of
1992; Shortland, P. and Woolf, C. J., 1993). It has been cell death, trans-synaptic degeneration, and the
suggested that regenerating A fibers exhibit collat- sprouting of sympathetic efferent fibers in DRG
eral sprouting into superficial dorsal horn lamina and A-fiber primary afferent fibers in the dorsal
after the terminal arbors of small A and C fibers horn which contribute to central sensitization and
degenerate following peripheral nerve injury the development and/or maintenance of hyperalge-
(Wilson, P. and Kitchener, P. D., 1996), a theory sia and allodynia.
that is supported by evidence of similar A fiber
sprouting after local application of the C-fiber neu-
rotoxin capsaicin to the sciatic nerve (Mannion, R. J. Acknowledgments
et al., 1996). A fiber sprouting into rat spinal lamina
II has also been observed in rats with SNL or CCI of This work is supported by grants form CIHR,
the sciatic nerve (Shortland, P. et al., 1997; Nakamura, NSERC, and FRSQ to T. J. C.
S. and Myers, R. R., 1999) or even chronic inflamma-
tion of the rat hind paw (Ma, Q. P. and Tian, L.,
2002). Lekan H. A. et al. (1997) also found increased
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 27 Glycine Receptors
H U Zeilhofer, University of Zurich, Zurich, Switzerland
2009 Elsevier Inc. All rights reserved.

5.27.1 Molecular Architecture of Glycine Receptors 381

5.27.2 Glycinergic Innervation in the Spinal Cord Dorsal Horn 381
5.27.3 Inhibitory Glycine Receptors and Spinal Pain Control 381
5.27.3.1 Glycine Receptors in the Spinal Control of Inflammatory Pain 382
5.27.3.2 Glycine Receptors in the Spinal Control of Neuropathic Pain 383
5.27.3.3 Transmitters Facilitating Glycinergic Inhibition 384
References 384

Glossary
GABA Gamma aminobutyric acid; fast inhibitory cyclooxygenase-2 Inducible enzyme mediating
neurotransmitter in the mammalian CNS. the production of prostaglandin precursors.
GABAA receptor Chloride channel-coupled GABA EGFP Enhanced green fluorescent protein
receptor blocked by bicuculline. used, for example to label cells in vitro and
glycine Fast inhibitory neurotransmitter in the in vivo.
spinal cord and brain stem. KCC2 Potassium/chloride exporter protein,
gephyrin Postsynaptic protein mediating the needed for central neurons to reduce their intra-
clustering of GABAA and glycine receptors in the cellular chloride concentration to physiological
postsynaptic membrane. levels.

27.1 Molecular Architecture of life. In most organs GlyR2 homomers are replaced by
Glycine Receptors GlyR1/ heteromers during postnatal maturation.
GlyR3 is another adult glycine receptor subunit
Glycine is the major inhibitory neurotransmitter in but much less abundant than GlyR1. GlyR4
the spinal cord and brain stem. In addition, glycine is is probably not expressed in humans because of a
an obligatory co-agonist at excitatory glutamate recep- premature stop codon in the GLRA4 gene.
tors of the N-methyl-D-aspartate (NMDA) subtype.
This chapter focuses on inhibitory (strychnine-sensi-
tive) glycine receptors. These glycine receptors are 27.2 Glycinergic Innervation in
heteropentameric chloride-permeable ion channels the Spinal Cord Dorsal Horn
composed of  and  subunits (Legendre, P., 2001).
Four different genes (glra1glra4) encode for the gly- Immunofluorescence studies have shown that markers
cine-binding  subunits and one gene (glrb) encodes for of glycinergic innervation are abundant in the spinal
the so-called structural  subunit.  Subunits carry the cord dorsal horn (Todd, A. J. and Sullivan, A. C., 1990).
glycine binding site and are capable of forming func- Glycine receptors are found throughout the dorsal
tional homomeric receptor channels. The  subunit horn. As in other regions of the central nervous system
permits postsynaptic clustering of glycine receptors (CNS), most glycine receptors in the adult spinal cord
through an interaction with the postsynaptic density are 1/ heteromeric receptor channels. However, in
protein gephyrin. Most glycine receptors in the adult the superficial layers of the dorsal horn, where most
are heteromeric receptors composed of 1 and  sub- nociceptive afferents terminate, glycine receptors con-
units. GlyR2 forms homomeric receptor channels tain GlyR3 in addition to GlyR1 and GlyR
during embryonic development and in early postnatal (Figure 1) (Harvey, R. J. et al., 2004). The prominent

381
382 Glycine Receptors

glycine (and also of GABAA) receptors can elicit

and exaggerate nociceptive responses (for a recent
review see Zeilhofer, H. U., 2005). Several groups
have meanwhile demonstrated that relief from inhi-
bition also occurs endogenously during inflammation
and possibly also after peripheral nerve injury.

27.3.1 Glycine Receptors in the Spinal

Control of Inflammatory Pain
Inflammatory pain originates to a large extent from
prostaglandins, which are produced in response to
inflammation and tissue damage mainly by inducible
cyclooxygenase-2 (COX-2). The pivotal role of pros-
GlyR 3
taglandins for pain sensitization is obvious from our
250 m
everyday experience that profound analgesia can be
Figure 1 Expression of GlyR3 in the spinal cord dorsal achieved through inhibition of prostanoid formation
horn. GlyR3 (green) is distinctly expressed in the either by the classical nonspecific cyclooxygenase
superficial layers where nociceptive afferents terminate, (COX) inhibitors (aspirin and related drugs) or by
whereas gephyrin (red), a marker for glycinergic and
the more recently developed COX-2-specific inhibi-
GABAergic synapses, is found throughout the spinal
cord. Adapted from Harvey, R. J., Depner, U. B., Wassle, H., tors (Brune, K. and Zeilhofer, H. U., 2005).
Ahmadi, S., Heindl, C., Reinold, H., Smart, T. G., Harvey, K., For a long time it was widely believed that
Schutz, B., Abo-Salem, O. M., Zimmer, A., Poisbeau, P., prostaglandins sensitize the nociceptive system only
Welzl, H., Wolfer, D. P., Betz, H., Zeilhofer, H. U., and Muller, U. at the level of the peripheral nociceptor. This
2004. GlyR 3: an essential target for spinal inflammatory pain
long-held view has changed significantly since the
sensitization. Science 304, 884887; with permission.
mid-1990s when several groups demonstrated that
inflammation induces COX-2 expression not only
expression of glycine receptors in the spinal cord is
in the peripheral inflamed tissue, but also in the
accompanied by dense glycinergic innervation.
CNS, in particular in the spinal cord dorsal horn. A
Glycinergic axon terminals, characterized by the
central action of prostaglandins and accordingly also
expression of the neuronal glycine transporter isoform
a central mode of action for the analgesic action of
GlyT2, and glycinergic cell bodies, identified by their
COX inhibitors are meanwhile generally accepted
immunoreactivity against glycine, are widely distribu-
(Malmberg, A. B. and Yaksh, T. L., 1992; Samad, T.
ted in the dorsal horn. Recently, mice expressing
A. et al., 2002).
enhanced green fluorescent protein (EGFP) under the
Electrophysiological experiments have shown
transcriptional control of the GlyT2 gene have been
that PGE2 reduces inhibitory glycinergic neurotrans-
generated (Zeilhofer, H. U. et al., 2005). In the dorsal
mission in the spinal cord dorsal horn, but does
horn of these mice, numerous glycinergic (EGFP/
not interfere with GABAergic or glutamatergic neu-
GlyT2-positive) neurons are found in the deeper dorsal
rotransmission (Figure 2) (Ahmadi, S. et al., 2002).
horn (laminae IIIV) and in lamina I, whereas relatively
This inhibition is (at least in adult animals) specific
few glycinergic neurons are seen in lamina II. In addi-
for neurons in the superficial layers of the dorsal horn
tion to these local inhibitory interneurons, descending
and for PGE2 (PGD2, PGI2, and PGF2 are without
antinociceptive glycinergic fiber tracts (Antal, M. et al.,
effect). It involves activation of postsynaptic prosta-
1996; Kato, G. et al., 2006) probably contribute to the
glandin E receptors of the EP2 subtype and a specific
glycinergic innervation of the dorsal horn.
protein kinase A (PKA)-dependent phosphorylation
of GlyR3-containing receptors, which are in the
spinal cord distinctly expressed in the superficial
27.3 Inhibitory Glycine Receptors layers (Harvey, R. J. et al., 2004). Subsequent experi-
and Spinal Pain Control ments using site-directed mutagenesis have shown
that GlyR3 is phosphorylated at serine 346 in the
Plenty of information meanwhile indicates that relief long intracellular loop between transmembrane
from synaptic inhibition by spinal blockade of regions 3 and 4. Figure 3 shows a schematic
 Glycine Receptors 383

(a) (b) (c)

Glycine-IPSCs 50 100
Control PGE2 Wash

% control PSC
40 90

% PSC block
80
30
70
20
60
10 50

100 pA
20 ms 0 0.001 0.01 0.1 1.0 10
Glycine GABA AMPA NMDA
PGE2 (M)

Figure 2 PGE2 selectively inhibits glycinergic neurotransmission onto spinal cord dorsal horn neurons. (a) Averaged
inhibitory postsynaptic current (IPSC) traces recorded from a neuron in the superficial dorsal horn under control conditions, in
the presence of PGE2, and after its removal. (b) Statistical analysis (mean SEM). Significant inhibition by PGE2 occurred only
for glycinergic IPSCs, GABAergic IPSCs, and excitatory postsynaptic currents mediated by glutamate receptors of the alpha-
amino-3-hydroxy-5-methyl-4-isoxazolepropionic acid (AMPA) or N-methyl-D-aspartate (NMDA) type remained unchanged.
(c) Concentrationresponse curve. Significant inhibition of glycinergic IPSC occurred at low nanomolar concentrations.
Adapted from Ahmadi, S., Lippross, S., Neuhuber, W. L., and Zeilhofer, H. U. 2002. PGE2 selectively blocks inhibitory
glycinergic neurotransmission onto rat superficial dorsal horn neurons. Nat. Neurosci. 5, 3440, with permission from Nature
Publishing Group.

COX-2 PGE2 carrying a loss-of-function mutation in the GlyR1

subunit lack a nociceptive phenotype.
PGE synthase
EP2 receptor Mice lacking GlyR3 (or EP2 receptors) not only
exhibit a nearly complete loss of pain sensitization by
Ca2+ spinal PGE2 but also display nearly identical noci-
ceptive phenotypes in models of inflammatory pain
GS
(Figure 4) (Harvey, R. J. et al., 2004, Reinold, H. et al.,
PKA 2005; see also Zeilhofer, H. U., 2005). Both types of
Cl
knockout mice develop normal initial thermal and
GlyR3
Glycine mechanical sensitization, but recover much faster
GABA from hyperalgesia than wild-type mice. Inhibition
Cl of glycinergic neurotransmission by PGE2 thus is a
GlyR1 key event in the generation of inflammatory pain.

Ca2+ 27.3.2 Glycine Receptors in the Spinal

Control of Neuropathic Pain
Figure 3 Schematic representation of the intracellular
pathway leading to inhibition of glycinergic A role for cyclooxygenase products has also been
neurotransmission by PGE2. PGE2 activates E-type proposed for neuropathic pain originating from
prostaglandin receptors of the EP2 subtype, which
peripheral nerve injury. However, a significant con-
subsequently lead to an increase in intracellular camp and
activate protein kinase A (PKA). PKA finally phosphorylates tribution of prostaglandins to this pain form is
and inhibits GlyR3. Adapted from Progress in Pain discussed controversially and other mechanisms
research and Management, vol. 30. leading to a similar relief from inhibition may be
more important (Hosl, K. et al., 2006). Possible
representation of the intracellular pathway that mechanisms include an inhibition of synaptic glycine
underlies inhibition of glycine receptors by PGE2. release, reduced responsiveness of postsynaptic gly-
Unlike GlyR3-containing receptors, homomeric cine receptors, a reduction in the transmembrane
glycine receptors formed by GlyR1 and hetero- chloride gradient rendering the inhibitory tone of
meric channels composed of GlyR1 and GlyR GABAergic and glycinergic synaptic input less effi-
are not inhibited by PKA activation and mice cient, and a loss of inhibitory innervation, for
384 Glycine Receptors

(a) (b)
Zymosan A Zymosan A
(0.06 mg sct.) (0.06 mg sct.)
18 100
Paw withdrawal latency (s)

wt
16 GlyR3/

Reaction score (%)

80 EP2/
14
12 60
10
8 40

6
20
0 1 2 3 4 5 6 7 0 1 2 3 4 5 6 7
Time (days) Time (days)
Figure 4 Inflammatory pain sensitization in mice lacking PGE2-mediated inhibition of glycinergic neurotransmission. An
inflammatory response was induced by subcutaneous injection by a yeast extract (zymosan A) into one hind paw.
Inflammatory hyperalgesia was assessed by stimulating the inflamed paw either with noxious heat or mechanically with
calibrated von Frey filaments. Both mice lacking the EP2 receptor (EP2/ mice) and mice deficient in GlyR3 (GlyR3/
mice) recovered quickly from inflammatory hyperalgesia. Adapted from Zeilhofer, H. U. 2005. The glycinergic control of spinal
pain processing. Cell Mol. Life Sci. 62, 20272035.

example due to a selective death of GABAergic or 27.3.3 Transmitters Facilitating Glycinergic

glycinergic interneurons. Inhibition
There is indeed experimental evidence for all
Given that a relief from glycinergic and/or
three possibilities, but their contribution to neuro-
GABAergic inhibition underlies inflammatory and
pathic pain is not clear. A reduction in the trans-
neuropathic pain, one might speculate that transmit-
membrane chloride gradient in dorsal horn neurons
ters, which facilitate inhibitory transmission might
following peripheral nerve injury has been reported
contribute to endogenous antinociception. Several
by Coull J. A. et al. (2003). Peripheral nerve trauma
transmitters including ATP (Jang, I. S. et al., 2001;
reduced the expression of the potassium chloride
Rhee, J. S. et al., 2000) and norepinephrine (Baba, H.
exporter KCC2 in dorsal horn neurons. This induced
et al., 2000a; 2000b) facilitate glycine release through
a shift of the chloride equilibrium potential to
the activation of P2X receptors and 2 adrenoceptors
more depolarized values and rendered inhibitory
in the spinal cord, respectively. In rat sacral commis-
GABAergic and glycinergic input less efficient.
sural neurons (lamina X) glycinergic membrane
Another extensively discussed report suggests that
currents are potentiated by norepinephrine acting
peripheral nerve injury induces a specific loss of
on 2 adrenoceptors through a postsynaptic mechan-
spinal inhibitory GABAergic neurotransmission in
ism (Nabekura, J. et al., 1999). These and possibly
the dorsal horn of rats in the chronic constriction
other similar processes may contribute to endogen-
injury model and the spared nerve injury model of
ous pain-controlling processes. Unfortunately, only
neuropathic pain (Moore, K. A. et al., 2002). This
very few drugs target glycine receptors and most of
original report has proposed that the loss of
them lack the isoform specificity (Laube, B. et al.,
GABAergic input was due to the selective apoptotic
2002).
death of GABAergic interneurons. Subsequent stu-
dies have however suggested that such a loss may not
be necessary for the development of hyperalgesia in
the chronic nerve injury model of neuropathic pain
References
(Polgar, E. et al., 2003; 2004). Finally, the neuropep-
tide nocistatin is a specific inhibitor of GABA and
Ahmadi, S., Lippross, S., Neuhuber, W. L., and Zeilhofer, H. U.
glycine release in the spinal cord dorsal horn 2002. PGE2 selectively blocks inhibitory glycinergic
(Zeilhofer, H. U. et al., 2000) and it increases noci- neurotransmission onto rat superficial dorsal horn neurons.
ceptive responses in the rat formalin test and in the Nat. Neurosci. 5, 3440.
Antal, M., Petko, M., Polgar, E., Heizmann, C. W., and Storm-
chronic constriction injury of neuropathic pain after Mathisen, J. 1996. Direct evidence of an extensive
intrathecal injection. GABAergic innervation of the spinal dorsal horn by fibres
 Glycine Receptors 385

descending from the rostral ventromedial medulla. glycine response in rat sacral dorsal commissural neurons.
Neuroscience 73, 509518. Neuroscience 9, 2941.
Baba, H., Goldstein, P. A., Okamoto, M., Kohno, T., Ataka, T., Polgar, E., Gray, S., Riddell, J. S., and Todd, A. J. 2004. Lack of
Yoshimura, M., and Shimoji K. 2000a. Norepinephrine evidence for significant neuronal loss in laminae IIII of the
facilitates inhibitory transmission in substantia gelatinosa of spinal dorsal horn of the rat in the chronic constriction injury
adult rat spinal cord (part 2): effects on somatodendritic sites model. Pain 111, 144150.
of GABAergic neurons. Anesthesiology 92, 485492. Polgar, E., Hughes, D. I., Riddell, J. S., Maxwell, D. J.,
Baba, H., Shimoji, K., and Yoshimura, M. 2000b. Puskar, Z., and Todd, A. J. 2003. Selective loss of spinal
Norepinephrine facilitates inhibitory transmission in GABAergic or glycinergic neurons is not necessary for
substantia gelatinosa of adult rat spinal cord (part 1): effects development of thermal hyperalgesia in the chronic
on axon terminals of GABAergic and glycinergic neurons. constriction injury model of neuropathic pain. Pain
Anesthesiology 92, 473484. 104, 229239.
Brune, K. and Zeilhofer, H. U. 2005. Antipyretic Analgesics Reinold, H., Ahmadi, S., Depner, U. B., Layh, B., Heindl, C.,
Basic Aspects. In: Melzack and Walls Textbook of Pain, 5th Hamza, M., Pahl, A., Brune, K., Narumiya, S., Muller, U., and
edn. (eds. S. McMahon and M. Koltzenburg), pp. 459469. Zeilhofer, H. U. 2005. Spinal inflammatory hyperalgesia is
Churchill Livingstone. mediated by prostaglandin E receptors of the EP2 subtype.
Coull, J. A., Boudreau, D., Bachand, K., Prescott, S. A., Nault, F., J. Clin. Invest. 115, 673679.
Sik, A., De Koninck, P., and De Koninck, Y. 2003. Trans- Rhee, J. S., Wang, Z. M., Nabekura, J., Inoue, K., and Akaike, N.
synaptic shift in anion gradient in spinal lamina I neurons as a 2000. ATP facilitates spontaneous glycinergic IPSC
mechanism of neuropathic pain. Nature 424, 938942. frequency at dissociated rat dorsal horn interneuron
Harvey, R. J., Depner, U. B., Wassle, H., Ahmadi, S., Heindl, C., synapses. J. Physiol. 524, 471483.
Reinold, H., Smart, T. G., Harvey, K., Schutz, B., Abo- Samad, T. A., Sapirstein, A., and Woolf, C. J. 2002. Prostanoids
Salem, O. M., Zimmer, A., Poisbeau, P., Welzl, H., and pain: unraveling mechanisms and revealing therapeutic
Wolfer, D. P., Betz, H., Zeilhofer, H. U., and Muller, U. 2004. targets. Trends Mol. Med. 8, 390396.
GlyR 3: an essential target for spinal inflammatory pain Todd, A. J. and Sullivan, A. C. 1990. Light microscope study of
sensitization. Science 304, 884887. the coexistence of GABA-like and glycine-like
Hosl, K., Reinold, H., Harvey, R. J., Muller, U., Narumiya, S., and immunoreactivities in the spinal cord of the rat. J. Comp.
Zeilhofer, H. U. 2006. Spinal prostaglandin E receptors of the Neurol. 296, 496505.
EP2 subtype and the glycine receptor 3 subunit, which Zeilhofer, H. U. 2005. The glycinergic control of spinal pain
mediate central inflammatory hyperalgesia, do not processing. Cell Mol. Life Sci. 62, 20272035.
contribute to pain after peripheral nerve injury or formalin Zeilhofer, H. U., Muth-Selbach, U., Guhring, H., Erb, K., and
injection. Pain 126, 4653. Ahmadi, S. 2000. Selective suppression of inhibitory
Jang, I. S., Rhee, J. S., Kubota, H., Akaike, N., and Akaike, N. synaptic transmission by nocistatin in the rat spinal cord
2001. Developmental changes in P2X purinoceptors on dorsal horn. J. Neurosci. 20, 49224999.
glycinergic presynaptic nerve terminals projecting to rat Zeilhofer, H. U., Studler, B., Arabadzisz, D., Schweizer, C.,
substantia gelatinosa neurones. J. Physiol. 536, 505519. Ahmadi, S., Layh, B., Bosl, M. R., and Fritschy, J. M. 2005.
Kato, G., Yasaka, T., Katafuchi, T., Furue, H., Mizuno, M., Glycinergic neurons expressing enhanced green fluorescent
Iwamoto, Y., and Yoshimura, M. 2006. Direct GABAergic and protein in bacterial artificial chromosome transgenic mice. J.
glycinergic inhibition of the substantia gelatinosa from the Comp. Neurol. 482, 123141.
rostral ventromedial medulla revealed by in vivo patch-clamp
analysis in rats. J. Neurosci. 26, 17871794.
Laube, B., Maksay, G., Schemm, R., and Betz, H. 2002.
Modulation of glycine receptor function: a novel approach
for therapeutic intervention at inhibitory synapses? Trends Further Reading
Pharmacol. Sci. 23, 519527.
Legendre, P. 2001. The glycinergic inhibitory synapse. Cell.
Mol. Life Sci. 58, 760793. Baba, H., Kohno, T., Moore, K. A., and Woolf, C. J. 2001. Direct
Malmberg, A. B. and Yaksh, T. L. 1992. Hyperalgesia mediated activation of rat spinal dorsal horn neurons by prostaglandin
by spinal glutamate or substance P receptor blocked by E2. J. Neurosci. 21, 17501756.
spinal cyclooxygenase inhibition. Science 257, 12761279. Sherman, S. E., Luo, L., and Dostrovsky, J. O. 1997. Spinal
Moore, K. A., Kohno, T., Karchewski, L. A., Scholz, J., Baba, H., strychnine alters response properties of nociceptive-specific
and Woolf, C. J. 2002. Partial peripheral nerve injury neurons in rat medial thalamus. J. Neurophysiol.
promotes a selective loss of GABAergic inhibition in the 78, 628637.
superficial dorsal horn of the spinal cord. J. Neurosci. Sivilotti, L. and Woolf, C. J. 1994. The contribution of GABAA
22, 67246731. and glycine receptors to central sensitization: disinhibition
Nabekura, J., Xu, T. L., Rhee, J. S., Li, J. S., and Akaike, N. and touch-evoked allodynia in the spinal cord. J.
1999. Alpha2-adrenoceptor-mediated enhancement of Neurophysiol. 72, 169179.
This page intentionally left blank
 28 Pain Following Spinal Cord Injury
R P Yezierski, Comprehensive Center for Pain Research and The McKnight Brain Institute, University
of Florida, Gainesville, FL, USA
2009 Elsevier Inc. All rights reserved.

28.1 The Condition of Pain Following Spinal Cord Injury 387

28.2 Clinical Characteristics of Spinal Injury Pain 388
28.3 The Research Challenge of Spinal Injury Pain 389
28.4 Pathophysiology of Spinal Cord Injury 390
28.5 Mechanism(s) of Pain Following Spinal Injury 391
28.5.1 Loss of Inhibitory Tone 392
28.5.2 Pattern Generators of Pain 393
28.5.3 Cell Signaling Pathways: Synaptic Plasticity and Functional State 394
28.5.4 Supraspinal Changes Following Spinal Cord Injury 395
28.6 Future Directions 395
References 396

Glossary
at-level pain Pain condition associated with spinal that ultimately lead to changes in the functional
cord injury. Located in dermatomes associated properties of spinal and supraspinal sensory
with spinal segments immediately adjacent to the neurons.
site of injury. Characteristics typically include ele- central sensitization Change in the functional
vated sensitivity to mechanical and thermal stimuli. state of central neurons secondary to persistent
Spontaneous sensations (i.e., tingling, numbness, input or injury-induced elevations of excitatory
burning) can also be associated with this condition amino acids leading to the activation of cellular
of abnormal sensation. Believed to have a predo- signaling pathways. The end result of this change in
minately spinal mechanism. functional state is an increase in excitability that
below-level pain Pain condition associated with can lead to clinical symptoms of allodynia, hyper-
spinal cord injury. Located in dermatomes asso- algesia, or pain.
ciated with spinal segments below the level of pattern generators of pain Dysfunctional neu-
injury. Pain is typically spontaneous with charac- rons with increased excitability are capable of
teristics of burning, stabbing, and shooting producing abnormal discharges. Collectively the
sensations. Mechanism of below-level pain is discharges generated from a large population of
believed to involve spinal as well as supraspinal neurons can form the substrate responsible for the
mechanisms. perception of pain.
central injury cascade Pathological events trig- secondary injury Following an initial mechanical
gered by injury to the central nervous system. Initial or vascular insult to the brain or spinal cord there is
events include the extracellular release of excita- a series of events that are components of the cen-
tory amino acids and the initiation of excitotoxic tral injury cascade. These events lead to the spread
and inflammatory cascades. Included in this cas- of pathological damage of central neurons beyond
cade are neurochemical and molecular changes the primary insult.

387
388 Pain Following Spinal Cord Injury

28.1 The Condition of Pain Following 28.2 Clinical Characteristics of

Spinal Cord Injury Spinal Injury Pain

Pain following spinal cord injury (SCI) is one of Pain associated with SCI is typically bilateral and
many challenges associated with the life-threatening perceived in anesthetic regions at and below the level
consequences of SCI. Over the past 15 years clinical of injury. Although changes in sensation above the
and preclinical studies related to at- and below-level injury have been reported, there is little evidence
pain have provided an understanding of the spinal supporting the clinical significance of this type of
and supraspinal mechanism(s) responsible for these altered sensation. The more common at- and below-
conditions. In spite of the fact that loss of sensory level conditions are often referred to as deafferentation
and motor function below the level of injury are pain, dysesthetic pain, or central dysesthesia syndrome
regarded as the most significant consequences of (Beric, A. et al., 1988; Bonica, J. J., 1991; Yezierski, R. P.,
SCI, the condition of pain has a direct relationship 2002). A characteristic of SCI pain that has been con-
with the ability of patients to regain an optimal level sistently documented is the detrimental impact across
of activity, return to work, and achieve an accepta- multiple quality of life domains, including life satisfac-
ble quality of life. The impact of spinal injury pain tion, physical health, and overall handicap. Commonly
on the healthcare community is evident from studies used descriptors of SCI pain include burning, tingling,
reporting the incidence of painful sensations at a numbness, aching, and throbbing. Although it is often
rate of 6080% for all SCI patients with nearly difficult to classify different categories of pain, below-
40% reporting severe pain to the extent they level neuropathic pain is the most common and most
would trade any chance of recovering motor func- difficult to treat (Finnerup, N. M. et al., 2002). When
tion for relief of pain (Nepomuceuno, C. et al., 1979; discussing different classifications of pain it is helpful to
Beric, A., 1990; Britell, C. W. and Mariano, A. J., have a standard taxonomy. A Task Force of the
1991; Tasker, R. R. et al., 1991; Beric, A., 1992; International Association for the Study of Pain pro-
Widerstrom-Noga, E. G. et al., 1999a). The func- posed a taxonomy for SCI pain based on the type of
tional impact of pain following SCI is further pain, location, and putative mechanism of pain onset
demonstrated by a report describing the challenge (Siddall, P. J. et al., 2002; Table 1). Having a taxonomy
of dealing with pain is only surpassed by the that is accepted and used by clinical and basic research
decreased ability to walk, loss of sexual function, specialists in the fields of pain and spinal injury is
and bladder and bowel dysfunction (Widerstrom- essential to achieving a better understanding of differ-
Noga, E. G. et al., 1999b). ent pain conditions.

Table 1 International Association for the Study of Pain proposed classification of spinal cord injury pain

Broad type (tier one) Broad system (tier two) Specific structures/pathology (tier three)

Nociceptive Musculoskeletal Bone, joint, muscle trauma, or inflammation

Mechanical instability
Muscle spasm
Secondary overuse syndromes
Visceral For example, Renal calculus, bowel dysfunction
Dysreflexic headache
Neuropathic (above level) Compressive mononeuropathies
Complex regional pain syndromes
(at level) Nerve root compression, cauda equina
Syringomyelia
Spinal cord trauma/ischaemia
Dual level cord and root trauma
(below level) Spinal cord trauma/ischaemia

Adapted from Siddall, P. J., Yezierski, R. P., and Loeser, J. D. 2002. Taxonomy and Epidemiology of Spinal Cord Injury. In:
Spinal Cord Injury Pain: Assessment, Mechanisms, Management (eds. R. P. Yezierski and K. Burchiel), pp. 924. IASP Press,
with permission from International Association for the Study of pain.
 Pain Following Spinal Cord Injury 389

The condition of pain following spinal injury has a nociception that have the potential to reveal changes in
variable onset depending on the type of pain. Siddall excitability of spinal circuits, an important conse-
P. J. et al. (2003) described the prevalence and onset of quence of spinal injury. Enhancement of flexion/
different types of pain in a cohort of 100 patients. withdrawal reflexes as reported in the spastic syn-
Musculoskeletal pain (59%) was the most common drome, however, can be dissociated from chronic
type of pain followed by at-level (41%) and below- below-level pain in individual cases of subtotal SCI
level (34%) neuropathic pain, with visceral pain (Vierck, C. J. et al., 2002a; 2002). These responses do not
(5%) the least common type of pain experienced. represent the component of pain perception that
The mean onset time for any type of pain was 1.6 requires cerebral processing of sensation (Mauderli,
years with at-level and musculoskeletal pain having A. P. et al., 2000) and are more likely to apply to the
the shortest onset times, 1.2 and 1.3 years, respec- study of at-level pain that is thought to depend on a
tively. Below-level pain generally had a longer onset spinal mechanism. A misconception in the preclinical
time (1.8 years) and visceral pain had the longest (4.2 study of SCI pain is that stimulation of peripheral
years). While there has been considerable effort to dermatomes below the lesion to elicit a reflex response
identify physical factors such as completeness and is thought to qualify as the study of below-level pain.
level of injury that correlate with pain onset, few The fact is below-level pain is likely to involve
consistent predictors have been identified (Siddall, supraspinal mechanisms and therefore cannot be stu-
P. J. et al., 1999a; Werhagen, L. et al., 2004). A positive died with reflex-based outcome measures.
relationship between the higher incidence of pain in Another strategy of behavioral assessment is the use
patients with thoroacolumbar and incomplete lesions of operant tests that require discrimination between
has been described (Demirel, G. et al., 1998). nociceptive and non-nociceptive stimulation and the
organization of complex behavioral adaptations to spe-
cific environmental contingencies. This approach is
28.3 The Research Challenge of believed to provide a more accurate assessment of the
Spinal Injury Pain pain experience (Vierck, C. J. et al., 2002). The challenge
of selecting appropriate behavioral measures is espe-
A major challenge in the study of altered sensation cially evident in the study of below-level pain. If it is
following SCI is the development of appropriate injury assumed that this type of pain requires cortical involve-
models that parallel the human condition. The study of ment then to study this type of pain effectively a
SCI pain has seen the use of several different models, behavioral measure that relies on activation of cortical
including mechanical trauma, isolated lesions, com- structures must be used. The only measures to fit these
plete transection, chemical lesions, and ischemic criteria are operant behaviors that rely on cortical pro-
injury, each with pathological components found in cessing for the perception of nociceptive stimuli as well
the human condition (Wiesenfeld-Hallin, Z. et al., as the planning and execution of learned responses.
1994; Siddall, P. J. et al., 1995; Christensen, M. D. and Acceptance of the differences between responses
Hulsebosch, C. E., 1997; Yezierski, R. P. et al., 1998; obtained with operant versus reflex measures represents
Bruce, J. C. et al., 2002; Scheifer, C. et al., 2002). a major challenge in the study of spinal injury pain,
Behavioral outcome measures used to evaluate especially studies related to at- and below-level pain.
injury-induced changes in sensation represents another By far the most significant sensory consequence of
critical factor in the study of spinal and supraspinal spinal injury is below-level pain. This condition has
mechanisms of different pain conditions. The majority been difficult to study in preclinical models as the pain
of behavioral measures used in the study of spinal is typically spontaneous and occurs in parts of the body
injury pain rely on responses to mechanical and ther- that are anesthetic. Below-level pain has been attribu-
mal stimuli to evaluate changes in nociceptive ted to white matter damage, although evidence
sensitivity. These studies have used a variety of noci- supports the presence of abnormal activity within the
ceptive reflexes, such as hindpaw withdrawal, that are gray matter contributing to below-level hyperalgesia
regulated by segmentally organized spinal mechanisms and spontaneous pain behavior. This was demonstrated
and are present in spinalized animals. Lick and guard in experiments with rats and monkeys in which
responses have also been used and these responses to ischemic involvement of spinal gray matter was a
nociceptive input depend on spino-bulbo-spinal cir- critical factor in the development of hyperalgesia
cuits and are present in decerebrate animals (Woolf, C. after interruption of the spinothalamic tract (Vierck,
J., 1984). Reflex responses are important measures of C. J. et al., 2000). Similarly, quisqualic acid injections in
390 Pain Following Spinal Cord Injury

the spinal gray matter produce below-level hyperalge- impact the survivability and functional integrity of
sia and spontaneous pain behavior without damage to spinal neurons (Figure 1). The clinical consequences
surrounding gray matter (Yezierski, R. P. et al., 1998; associated with SCI, including at- and below-level
King, C. D. et al., 2006). Clinically, results of a magnetic pain, are believed to result from the influence of this
resonance imaging study in patients with and without cascade on the functional state of spinal and suprasp-
below-level pain showed the most common pathologi- inal neurons. Therefore to understand the end result
cal feature of patients with below-level pain was the (i.e., pain), it is important to appreciate the basic biol-
presence of lesions that included the central core of ogy of secondary events that precede this condition.
spinal gray matter (Finnerup, N. M. et al., 2003b). The most obvious pathological changes associated
with traumatic or ischemic injury to the spinal cord
include, but are by no means limited to, the loss
28.4 Pathophysiology of Spinal Cord of neurons, damage to surrounding white matter,
Injury astrocytic scarring, syrinx formation, and breakdown
of the spinal bloodbrain barrier (Kakulas, B. A. et al.,
In order to understand the mechanisms responsible for 1990; Bunge, R. P. et al., 1993). Also contributing to the
SCI pain it is important to acknowledge that asso- progression of tissue damage are secondary injury
ciated with spinal injury is a cascade of biochemical, cascades that include excitotoxic and inflammatory
molecular, and anatomical changes that collectively processes (Young, W., 1987; Tator, C. H. and

Spinal injury
(ischemia/trauma)

Neurochemical
AAs (glutamate, GABA); Anatomical
ionic (Na+, Ca2+, Cl); Necrosis, apoptosis,
peptides (dynorphin, Sub P, CGRP); gliosis, demyelination,
2nd messengers (cGMP, NO, cytoskeletal damage,
c-fos, NFB, ELK), kinases deafferentation,
(ERK, PKC), cytokines (TNF, IL-1 ), sprouting
enzymes (calpain, PLA2, caspases)

Excitotoxicity Inflammation
EAAs, Microglia activation,
Ionic gradients, NOS cytokines (TNF, IL-1 ),
free radicals, reactive oxygen peptides (dynorphin),
species, microglia, enzymes (COX-2, iNOS)
depolarization, cell death

Physiological
Sodium channels, excitability, receptive fields, background activity, gain,
after-discharge

Clinical/behavioral
Allodynia, hyperalgesia, pain

Figure 1 Components of the spinal injury cascade that contribute to the mechanism responsible for the development of
pain. Evidence supporting the involvement of this cascade comes from results of clinical and experimental studies involving
mechanical, ischemic, and/or chemical models of spinal injury (see text for details). The four major components of the cascade
(neurochemical, excitotoxicity, anatomical, and inflammation) are represented as interactive and collectively lead to conditions
producing functional changes in spinal and supraspinal neurons. The endpoint of the cascade is the onset of clinical and
behavioral symptoms, e.g., allodynia, hyperalgesia, and pain. AAs, amino acids; cGMP, cyclic guanidine monophosphate;
CGRP, calcium gene-related peptide; COX-2, cyclo-oxygenase-2; EAA, excitatory amino acid; ELK, ets-like gene; ERK,
extracellular signal-regulated kinase; GABA, gamma-aminobutyric acid; IL-1, interleukin-1; iNOS, inducible nitric oxide
synthase; NFkB, nuclear factor kappa B; NO, nitric oxide; PKC, protein kinase C; PLA2, phospholipase A2; Sub P, substance P;
TNF, tumor necrosis factor. Reproduced from Yezierski, R. P. 2006. Pain Following Spinal Cord Injury: Central Mechanisms. In:
Handbook of Clinical Neurology (eds. F. Cervero and T. S. Jensen), Vol. 81, pp. 293307, with permission from Elsevier.
 Pain Following Spinal Cord Injury 391

Fehlings, M. G., 1991; Hsu, C. Y. et al., 1994; Regan, R. associated with the MAPK signaling pathway
and Choi, D. W., 1994; Bethea, J. R. et al., 1999). (Crown, E. D. et al., 2005; Yu, C. G. and Yezierski,
Excitotoxic events following spinal injury are initiated R. P., 2005a), increased NR1 serine phosphorylation
by the well-documented effects of excitatory amino of the NMDA receptor (Caudle, R. M. et al., 2003),
acids (EAAs) that are involved in the secondary changes in galanin immunoreactivity (Zvarova, K.
pathology, including neuronal degeneration (Faden, et al., 2004), and increased expression of c-fos
A. I. and Simon, R. P., 1988; Nag, S. and Riopelle, R. mRNA (Yakovlev, A. G. and Faden, A. I., 1994;
J., 1990; Hao, J. X. et al., 1991; Yezierski, R. P. et al., Siddall, P. J. et al., 1999b; Abraham, K. E. and
1993; Regan, R. and Choi, D. W., 1994; Wrathall, J. R. Brewer, K. L., 2001). Furthermore, analysis of mole-
et al., 1994). For this reason glutamate has been viewed cular events using DNA microarray analysis has
as one of several putative chemical mediators contri- shown that 1 h after spinal trauma there are changes
buting to the central cascade of secondary pathological in mRNA levels for 165 genes (Nesic, O. et al., 2002;
changes following injury (Tator, C. H. and Fehlings, 2005), including those regulating transcription fac-
M. G., 1991). Inflammation is another major contribu- tors, inflammatory processes, cell survival, and
tor to secondary injury (Bethea, J. R. and Dietrich, W. membrane excitability. In spite of the wide range of
D., 2002). With the aid of specific histological and changes associated with spinal injury, the challenge
immunohistological stains the temporal profile of glia remains to determine which ones have a causal rela-
activation (astrocytes, microglial) and the infiltration tionship with the mechanism of pain onset, as
of macrophages and other inflammatory cell types opposed to being secondary events associated with
can be followed. Upregulation of messenger RNA the process of injury.
(mRNA) for c-fos, tumor necrosis factor (TNF)- An important consideration in the progression of
and dynorphin have also been described following pathological changes associated with spinal injury is
SCI (Yakovlev, A. G. and Faden, A. I., 1994). From the longitudinal extent over which spinal tissue is
this discussion it should be clear that the pathological influenced by the central injury cascade. In a study
sequela of SCI is by no means simple neither is it using agents with neuroprotective properties it was
restricted to the site of insult as pathological conse- shown that limiting the spread of injury had a signifi-
quences of SCI have been observed throughout the cant impact on the expression of pain-related
full extent of the neuraxis (Brewer, K. L. et al., 1997; behaviors (Yu, C. G. et al., 2003). This study led to
Jain, N. et al., 1998; Morrow, T. J. et al., 2000). the proposal of a spatial threshold in which a critical
One of the earliest responses to spinal injury is the distance of spinal tissue must be affected by the sec-
breakdown of cellular membranes and subsequent ondary injury cascade in order for pain behaviors to
activation of phospholipases. Within minutes of develop. Therefore, a critical determinant underlying
injury, lipid peroxidation and membrane hydrolysis the expression of pain following SCI may be a combi-
release polyunsaturated fatty acids capable of causing nation of specific injury related events together with
damage, through arachidonic acid and eicosanoid the longitudinal spread of these events within the cord
pathways (Anderson, D. and Hall, E., 1993; Tator, (Yezierski, R. P., 2000; Gorman, A. L. et al., 2001).
C., 1995). Reactive oxygen species (ROS) likewise Identifying the mechanism responsible for the spread
trigger excitotoxic reactions (glutamate and quinoli- of injury may provide valuable insight into the devel-
nate); catecholamine oxidation; mitochondrial opment of different pain conditions. Given the
breakdown; oxidation of extravasated hemoglobin; permissive microenvironment for the development of
neutrophil infiltration; microglial activation; and pain associated with the wide spectrum of molecular,
release of nitric oxide, cytokines, and growth factors. anatomical, neurochemical, and functional changes
Thus, ROS contribute to the development of a per- that occur following spinal injury it is not surprising
missive environment for pain following SCI (Kim, H. that pain is a common clinical finding following injury.
K. et al., 2004; Hains, B. C. and Waxman, S. G., 2006;
Peng, X. M. et al., 2006). Other changes associated
with different models of SCI include: afferent sprout- 28.5 Mechanism(s) of Pain Following
ing in distant segments (Ondarza, A. B. et al., 2003), Spinal Injury
upregulation of vanilloid receptor expression (Zhou,
Y. et al., 2002), changes in expression of metabotropic The precise mechanism(s) underlying different types
glutamate receptors (Mills, C. D. et al., 2001), activa- of neuropathic pain (at- and below-level) associated
tion of protein kinases and transcription factors with spinal injury remain to be determined. While
392 Pain Following Spinal Cord Injury

below-level pain is considered to have a central progression of a secondary injury cascade, (5) reorga-
mechanism involving cortical and subcortical struc- nization of supraspinal structures, and (6) activation of
tures, at-level pain may have peripheral and central the MAPK cell signaling pathway (Figure 2).
components. Unfortunately, little is known about the
potential peripheral contribution to spinal injury
pain. Little is also known about the role of the auto- 28.5.1 Loss of Inhibitory Tone
nomic nervous system (ANS) in the development of A critical factor in the onset of painlike behaviors
different pain conditions following injury. Given the following SCI is the loss of inhibitory influences
likely damage to sympathetic and parasympathetic within the injured spinal cord (Wiesenfeld-Hallin,
neurons following injury and the contribution of the Z., et al. 1994). The loss of inhibitory control allows
ANS to chronic pain states, there is a need for studies for the recruitment of surrounding neurons and
focusing on the role of autonomic dysfunction in the intensification and spread of abnormal sensations,
development of spinal injury pain. including pain. Support for this conclusion comes
Results of experimental studies have provided evi- from four lines of evidence: (1) following ischemic
dence to construct a working model for at- and below- injury there is an increased excitability of WDR
level behavioral effects of SCI. This model includes a neurons that is reversed by the GABAB receptor
combination of six conditions: (1) loss of spinal inhi- agonist baclofen (Hao, J. X. et al., 1992), (2) hypersen-
bitory neurons, (2) emergence of spinal generators sitivity to peripheral stimuli can be produced in
and supraspinal amplifiers of abnormal activity, (3) normal animals by the intrathecal administration of
dependence on a population of NK-1R expressing gamma-aminobutyric acid (GABA) receptor antago-
neurons in the superficial dorsal horn, (4) longitudinal nists (Yaksh, T. L., 1989; Hao, J.-X. et al., 1994),

Deafferentation 1. Loss of spinal

Reorganization inhibitory
Supraspinal Abnormal input mechanisms.
generator Sensitization
amplifier Bursting 2. Spinal and
Hyperactivity supraspinal
generators
and/or amplifiers.

3. NK-1 receptor
expressing neurons
in lamina I.
4. Longitudinal
progression of injury
Anatomical cascade.
Neurochemical
Spinal Excitotoxicity 5. Functional
generator Site of Inflammation reorganization of
injury supraspinal
Sensitization
Hyperactivity structures.

6. Injury-induced
activation of cell
signaling pathways.
Figure 2 Summary of six different injury-induced changes contributing to the development of at- and below-level pain
associated with spinal cord injury. A spinal pain generator evolves due to the collective impact of anatomical, neurochemical,
inflammatory, and excitotoxic changes that result in the increased excitability of spinal sensory neurons. Specific changes
contributing to the development of the spinal generator include loss of inhibitory mechanisms, the longitudinal progression of
the spinal injury cascade, and the injury-induced activation of cell-signaling pathways. At supraspinal levels the presence of a
generator/amplifier mechanism emerges as a result of the loss of afferent input from spinal segments below the level of injury
(deafferentation). The functional impact of this and other injury-induced changes at supraspinal levels (e.g., sprouting,
unmasking of connections) results in the activation of supraspinal regions by input from somatotopically inappropriate regions
of the cord and the referral of pain sensations to dermatomes below the level of injury. Reproduced from Yezierski, R. P. 2006.
Pain Following Spinal Cord Injury: Central Mechanisms. In: Handbook of Clinical Neurology (eds. F. Cervero and T. S.
Jensen), Vol. 81, pp. 293307, with permission from Elsevier.
 Pain Following Spinal Cord Injury 393

(3) decreased numbers of GABA-positive neurons propofol are also effective (Herman, R. M. et al., 1992;
following ischemic spinal injury (Zhang, A.-L. et al., Canavero, S. et al., 1995). The anticonvulsant lamo-
1999), and (4) administration of drugs that prolong trigine that affects sodium channels involved in
the action of inhibitory neurotransmitters (i.e., tricyc- hyperexcitability is also suggested to be effective in
lic antidepressants) are effective in the short-term SCI patients with spontaneous and evoked pain
treatment of spinal injury pain (Tunks, E., 1986; (Finnerup, N. M. et al., 2002).
Lejon, G. and Boivie, J., 1991; Boivie, J., 1994). Consistent with the excitability hypothesis and
Although existing evidence supports the decreased involvement of neurons in the superficial dorsal
inhibitory influence of GABAergic neurotransmission horn in spontaneous and evoked pain behaviors, are
in altering the functional properties of neurons in the clinical reports of abnormal focal hyperactivity
injured cord, not to be overlooked in this process is a within the superficial laminae of the injured cord
decreased influence of supraspinal inhibitory path- (Edgar, R. E. et al., 1993). Microcoagulation of these
ways that may be disrupted following injury. hyperactive areas in the dorsal root entry zone
Alterations in descending serotonergic projections (DREZ) at depths of 12 mm, representing Rexeds
above and below the site of injury have led to the laminae III, results in a significant diminution of
suggestion that loss of inhibitory influences below the pain. A variant of the DREZ technique utilizing
injury may contribute to the increased excitability of intramedullary recordings of C-fiber evoked
neurons while an increase in serotonergic projections responses to guide DREZ lesioning allows for the
above the injury may contribute, through descending somatotopic mapping of the DREZ with regard to
facilitation, to a change in functional state of neurons the generation of central deafferentation pain. In 25
above the level of injury (Oatway, M. A. et al., 2004). patients where this technique was used, 84% received
complete pain relief (Falci, S. et al., 2002). An involve-
ment of this region in the generation of pain is further
28.5.2 Pattern Generators of Pain
supported by a report where elimination of NK-1R-
Another component of the pathophysiological expressing neurons in the superficial dorsal horn
sequela of SCI that is thought to contribute to the resulted in not only the prevention of spontaneous
onset of altered sensations is the emergence of a pain behavior, but also the reversal of the behavior
pattern generating mechanism (spinal and suprasp- once it had been initiated (Yezierski, R. P. et al., 2004).
inal). Several observations are consistent with the Injury-induced changes contributing to increased
pattern generating hypothesis: (1) existence of focal excitability of spinal circuitry has also been attributed
regions of hyperactivity in the spinal cord and thala- to an increased expression of the Nav1.3 sodium chan-
mus of spinal injured patients (Loeser, J. D. et al. 1968; nel in spinal segments L3L5 following contusion
Edgar, R. E. et al., 1993; Lenz, F. A. et al., 2000); injury at T9 (Hains, B. C. et al., 2003a). Upregulation
(2) effectiveness of spinal anesthesia in alleviating of Nav1.3 expression at sites remote from the epicen-
pain when delivered proximal to the site of injury ter of injury was shown to have a significant role
(Pollock, L. J. et al., 1951; Botterell, E. H. et al., 1953; in producing increased excitability of spinal multi-
Loubser, P. G. and Donovan, W. H., 1991); and receptive neurons recorded in L3L5. Efforts to
(3) sensitization and prolonged after-discharges of block these changes after injury showed that respo-
spinal and supraspinal sensory neurons following nses of neurons in animals treated intrathecally with
SCI (Hao, J. X. et al., 1992; Yezierski, R. P. and Park, antisense oligodeoxynucleotides targeting Nav1.3
S. H., 1993; Lenz, F. A. et al., 2000; Drew, G. M. et al., decreased expression of Nav1.3 mRNA and protein,
2001; Hains, B. C. et al., 2003b; Hoheisel, U. et al., reduced hyperexcitability of multireceptive neurons,
2003; Hao, J. X. et al., 2004; Hains, B. C. et al., 2005). and attenuated mechanical allodynia and thermal
Consistent with an involvement of neuronal hyper- hyperalgesia after SCI. Similarly, upregulation of
excitability as a component of the spinal and possibly Nav1.3 sodium channels in thalamic neurons has also
supraspinal mechanism of SCI pain are results of been shown to contribute to changes in the electro-
pharmacological studies. For example, lidocaine and physiological properties of these neurons (Hains, B. C.
ketamine, two drugs that reduce membrane excitabil- et al., 2005). Thus, changes in the functional state of
ity and glutamate receptor activation are effective in sensory neurons secondary to upregulation of sodium
treating SCI pain (Loubser, P. G. and Donovan, W. channels provides the mechanism for a pain generator
H., 1991; Eide, P. K. et al., 1995; Attal, N. et al., 2000). in the injured cord as well as an amplifier of abnormal
Efforts to increase inhibition with either baclofen or activity in thalamus.
394 Pain Following Spinal Cord Injury

The contribution of abnormal spinal input calcium, maintaining events (nitric oxide synthesis,
together with the effects of deafferentation (second- activation of protein kinases), perpetuating forces
ary to loss of spinal projection neurons) and (activation of cyclic AMP response element-binding
upregulation of sodium channels could result in the (CREB), nuclear factor kappa B (NF-kB), and term-
development of abnormal generators or amplifiers of inal results (altered synaptic efficacy). In contrast to
spinal input at supraspinal sites (Vierck, C. J., Jr., cellular events underlying persistent pain of periph-
1991). The transient relief of below-level pain in eral origin, the mechanism underlying long-term
humans following application of local anesthetic to plasticity associated with pain of central origin
the proximal stump of a spinal lesion (Pollock, L. J. remains largely unexplored.
et al., 1951) suggests that the presence of a spinal Important events contributing to the development
generator plays a significant role in maintaining of injury-induced pain are found in a construct of
chronic below-level pain. Thus, a combination of central sensitization and plasticity (Woolf, C. J. and
supraspinal changes secondary to deafferentation, Csotigan, M., 1999; Ji, R. R. and Woolf, C. J., 2001;
and the emergence of a spinal generator of abnormal Ji, R. R. et al., 2003). Hypersensitivity is believed to be
activity are two significant events that may contri- a consequence of early posttranslational changes as
bute to the development and maintenance of below- well as later transcription-dependent changes in
level pain. Recently, the report of elevated blood effector genes. Neuroplasticity resulting from these
flow in thalamic nuclei (possibly reflecting changes events includes activity-dependent changes in
in the functional state of neurons) lend additional neurons and specific signaling molecules in signal
support to the thalamic involvement in the suprasp- transduction pathways. These changes likely contri-
inal response to SCI and to the mechanism of injury- bute to alterations in the basal state of excitability that
induced pain (Morrow, T. J. et al., 2000). ultimately contributes to the development of spinal
pain generators and possibly supraspinal amplifiers.
Examples of cell signaling events following spinal
28.5.3 Cell Signaling Pathways: Synaptic
injury were described in a study of changes in the
Plasticity and Functional State
mitogen-activated protein kinase (MAPK) signaling
Another consideration for the mechanism of chronic pathway following excitotoxic spinal injury including:
pain following spinal injury is synaptic plasticity and (1) increased phosphorylation of ERK1/2, (2) increased
changes in the functional state of spinal neurons. A activation of NF-kB and phosphorylation of ELK-1,
significant response to injury of central or peripheral and (3) increased gene expression for the NK-1 recep-
origin is the cellular response believed to contribute to tor and NR1 and NR-2A subunits of the NMDA
the sensitization of spinal neurons (Woolf, C. J. and receptor (Yu, C. G. and Yezierski, R. P., 2005a).
Csotigan, M., 1999; Woolf, C. J., 1992; Willis, W. D., Blockade of the MAPK cascade with the MEK inhibi-
2002). Synaptic plasticity in the central nervous sys- tor PD98059 inhibited phosphorylation of ELK-1,
tem is thought to be part of central sensitization and activation of NF-kB, and gene expression of NR1,
together with the long-term changes in functional NR-2A, and NK-1R, and prevented the development
state of spinal sensory neurons represents a potential of spontaneous pain behavior. Injury-induced eleva-
mechanism for persistent pain (Ji, R. R. et al., 2003). tions in spinal levels of EAAs thus lead to the
Unfortunately, we are only at the beginning of under- activation of the ERK ! ELK-1 and NF- B signaling
standing the biochemical and molecular events cascades and the transcriptional regulation of receptors
underlying long-term functional plasticity and their important in the development of chronic pain.
role in chronic pain. Included in the events thought to Blockade of these intracellular kinase cascades prevents
be involved in producing long-term changes are: (1) the onset of injury-induced pain behavior. Similar
phosphorylation of regulatory proteins (as well as changes in different components of the MAPK signal-
dephosphorylation of others), (2) positive and negative ing pathway have been described in the diencephalon
regulation of gene transcription, (3) injury-induced following injury (Yu, C.-G. and Yezierski, R. P., 2005b).
synthesis of proteins (as well as reduced synthesis of The persistent upregulation of CREB in spinothalamic
others), (4) strengthening of some and weakening of tract cells is another example of how cell signaling
other connections (synaptic long-term potentiation, events are affected by SCI (Crown, E. D. et al., 2005).
sprouting, and pruning), and (5) the death or rescuing The above results suggest that many of the same
of neurons. These neuronal changes share initiating molecular changes described as activity-dependent
events such as glutamate-induced elevations in following peripheral nerve and tissue injury can also
 Pain Following Spinal Cord Injury 395

be induced following central injury, that is, injury cingulate cortex and PPD in the parietal cortex, both
dependent. This suggests that the mechanism respon- ipsilateral and contralaterally at various timepoints
sible for the increased excitability of spinal neurons following injury. Increases in PPD were significant in
following spinal injury may be similar to the well- animals that developed spontaneous pain behaviors
documented activity-dependent mechanism induced versus those that did not. PPE expression in the ante-
by damage to peripheral tissue and/or nerves, a rior cingulate cortex and PPD expression in the
mechanism resulting in the activation of intracellular contralateral parietal cortex were significantly higher
kinase cascades and ultimately long-term changes in in animals with spontaneous pain behaviors versus
synaptic efficacy and neuronal excitability. The end those without pain behaviors. The important conclu-
result of these events is the emergence of a hyperex- sion from these reports is that following spinal injury
citable pain-generating mechanism. there are significant changes in selected supraspinal
sites including somatosensory structures putatively
involved in pain processing. These results are consis-
28.5.4 Supraspinal Changes Following
tent with a recent report in humans using proton
Spinal Cord Injury
magnetic resonance spectroscopy showing changes in
One cannot discuss SCI without acknowledging the the thalamic concentrations of selected metabolites in
most fundamental of all anatomical facts and that is SCI patients with pain (Pattany, P. M. et al., 2002).
the connection between the brain and spinal cord. Thus there is likely to be a significant supraspinal
Following injury to the cord it has been well docu- component to the mechanism responsible for the
mented that there are multiple changes (e.g., development of below-level SCI pain. Understanding
anatomical, molecular, functional) that occur within the role of these supraspinal changes in the develop-
cortical and subcortical structures. Changes in ment of pain, especially below-level, remains a
supraspinal structures could potentially contribute challenge for future studies.
to the central mechanism responsible for the onset
and progression of injury-induced pain behaviors.
Studies carried out include an evaluation of changes 28.6 Future Directions
in forebrain blood flow (Morrow, T. J. et al., 2000),
and evaluation of opiate transmitters in selected An important realization of clinical and experimental
supraspinal sites (Abraham, K. E. et al., 2001). In the studies directed toward understanding the mechan-
study by Morrow T. J. et al. (2000) significant ism(s) of SCI pain is the importance of identifying the
increases in regional cerebral blood flow were critical events responsible for the onset of this con-
found in seven of 22 supraspinal structures examined, dition. Continued studies focusing on different
including the arcuate nucleus, hindlimb region of S1 components of secondary excitotoxic and inflamma-
cortex, parietal cortex and the thalamic posterior, tory cascades ultimately responsible for the
ventral posterior medial and lateral nuclei. All of dysfunction of spinal and supraspinal neurons are
these structures have well-documented roles in sen- essential to the design of more effective interven-
sory processing. These studies complement clinical tions. The condition of pain associated with spinal
observations showing similar changes in thalamic injury is unlikely to have a simple mechanism that
blood flow following SCI (Ness, T. J. et al., 1998), relies exclusively on any one consequence of injury.
alterations in the chemical profile of ventroposterolat- It is more probable that several components of a
eral thalamus in patients with SCI pain (Pattany, P. M. central injury cascade contribute to the development
et al., 2002), and reports of hyperactive foci of thalamic and maintenance of different pain states. The precise
activity in SCI patients with spontaneous burning pain mechanism is likely to depend on the nature of the
(Lenz, F. A. et al., 2000; Ohara, S. et al., 2002). injury and progression of pathological and biochem-
Additional support for supraspinal changes contri- ical changes along the rostrocaudal axis of the cord,
buting to the mechanism of spinal injury pain is the and include anatomical, molecular, and functional
work from Brewer and colleagues focusing on changes changes occurring in supraspinal structures. The
in peptidergic transmitter systems at supraspinal levels mechanism is likely to include the combination of
following excitotoxic SCI (Abraham, K. E. et al., 2001). deafferentation resulting from spinal lesions com-
Preproenkephalin (PPE) and preprodynorphin (PPD) bined with tonic excitatory input from a generator
expression was shown to increase in cortical regions of abnormal activity in or around the epicenter of
associated with nociceptive function; PPE in the gray matter damage, that is, penumbral region. This
396 Pain Following Spinal Cord Injury

input reaches deafferented supraspinal processing Attal, N., Gaude, V., Brasseur, L., Dupuy, M., Guirimand, F.,
Parker, F., and Bouhassira, D. 2000. Intravenous lidocaine in
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In conclusion, well-established animal models indi- pp. 2736. Karger.
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 29 Long-Term Potentiation in Pain Pathways
J Sandkuhler, Medical University of Vienna, Vienna, Austria
2009 Elsevier Inc. All rights reserved.

29.1 Introduction 401

29.2 Induction of Long-Term Potentiation in Pain Pathways 402
29.3 Signal Transduction Pathways 403
29.3.1 Long-Term Potentiation Induction is Context Sensitive 404
29.4 Prevention of Long-Term Potentiation 404
29.5 Reversal of Long-Term Potentiation 405
29.6 Functional Consequences of Long-Term Potentiation in Pain Pathways 405
29.7 Conclusions 405
References 405

Glossary
synaptic efficacy, syn. synaptic LTP induction Cellular signal transduction path-
strength Magnitude of postsynaptic current or ways which lead to LTP.
potential in response to a presynaptic action LTP maintenance Cellular signal transduction
potential. Synaptic strength may be modified by pathways which are involved in stabilizing estab-
pre- and/or postsynaptic modulation. lished LTP.
long-term potentiation (LTP) Enduring increase spinal field potential Extracelluar electrical field
in synaptic efficacy. which is mainly generated by simultaneous post-
long-term depression (LTD) Enduring decrease synaptic currents in spinal cord neurons in
in synaptic efficacy of a putatively nonpotentiated response to a brief stimulus, similar to evoked
synapse. potentials in cerebral cortex.
depotentiation Enduring decrease of synaptic
efficacy of a previously potentiated synapse, that
is, reversal of LTP.

29.1 Introduction Nicoll, R. A., 1999). LTP has also been shown in pain
pathways at synapses between primary afferent A- or
Long-term potentiation (LTP) is defined as the per- C-fibers and spinal dorsal horn neurons and is one of
sistent increase in synaptic strength (Bliss, T. V. P. and the proposed cellular mechanisms of pain amplification
Collingridge, G. L., 1993; Malenka, R. C. and Nicoll, (Moore, K. A. et al., 2000; Sandkuhler, J., 2000). The
R. A., 1999). Repetitive use of a synapse often leads to potential consequences of LTP in pain pathways are
LTP which may last for a few hours only or for days to manifold: (1) after LTP is established, suprathreshold
months. Synaptic strength may also be reduced for excitatory input will now evoke stronger excitation of
prolonged periods of time which is called long-term nociceptive neurons. Thus, LTP at nociceptive
depression (LTD) when induced in a normal synapse synapses may underlie hyperalgesia; (2) previously
or depotentiation when elicited in a previously poten- subthreshold excitatory input, from the subliminal
tiated synapse, that is, in the maintenance phase of fringe of a neurons receptive field may now elicit
LTP (Huang, C. C. and Hsu, K. S., 2001). action potential firing. This may widen painful areas
LTP has been most intensively studied in hippo- and contribute to hyperalgesia as well; and (3) some
campus where it is considered a fundamental cellular spinal dorsal horn neurons receive subthreshold input
model of learning and memory formation (Bliss, T. V. P. from somatotopically inappropriate body areas, for
and Collingridge, G. L., 1993; Malenka, R. C. and example, the contralateral body half. LTP anywhere

401
402 Long-Term Potentiation in Pain Pathways

along this subliminal input path may cause mirror 1. Electrical nerve stimulation: LTP has been induced
image pain or radiating pain. LTP at nociceptive by conditioning high intensity, high frequency
synapses may, however, not significantly lower pain burst-like stimulation (HFS, typically given as
thresholds and is therefore not considered a likely 50100 Hz bursts for 1 s at C-fiber strength) in
mechanism of allodynia. vitro (Randic, M. et al., 1993; Hamba, M. et al.,
The most direct way to ascertain LTP in pain 2000; Ikeda, H. et al., 2000; Ikeda, H. et al., 2003)
pathways is to measure monosynaptically evoked post- and in vivo (Liu, X. G. and Sandkuhler, J., 1997;
synaptic currents or potentials. So far this has only been Miletic, G. and Miletic, V., 2000; Ma, J. Y. and
done in vitro by recording A- or C-fiber-evoked exci- Zhao, Z. Q., 2001; Zhang, H. M. et al., 2001).
tatory postsynaptic currents or potentials from dorsal Stimulation intensity must be sufficient to recruit
horn neurons in response to electrical stimulation of C-fibers; stimulation at A-fiber intensity is ineffec-
dorsal roots. When postsynaptic currents are elicited tive to induce LTP at C-fiber synapses (Liu, X. G.
synchronously in a sufficient number of neighboring and Sandkuhler, J., 1997). C-fibers do, however,
neurons, electrical field potential are generated which discharge at such high frequencies only rarely.
are strong enough to be detected with extracellular Thus, the pathological relevance of HFS-induced
microelectrodes (Schouenborg, J., 1984). Thus, to eval- LTP can be challenged.
uate changes in synaptic strength, C-fiber-evoked field 2. Recent studies show that not only HFS but also
potentials can be recorded in superficial spinal dorsal conditioning low frequency stimuli at 210 Hz
horn in vivo and in vitro in response to supramaximal which is well within the frequency band of C-
electrical stimulation of a dorsal root or a sensory nerve, fiber discharges during inflammation or tissue
for example, the sciatic nerve. If, however, events injury causes robust LTP, both, in vivo and in
downstream to postsynaptic currents are recorded, vitro (Terman, G. W. et al., 2001; Ikeda, H. et al.,
such as action potential discharges, polysynaptically 2006)/electrical stimulation of sciatic nerve at C-
evoked responses, withdrawal reflexes or pain ratings, fiber intensity (2 Hz, 2 min) induces robust LTP to
LTP can no longer be assessed directly. These mea- 300% of control values which is not reversible
sures are, however, indispensable to evaluate the within the recording periods of up to 18 h
functional consequences of LTP in pain pathways. (Figure 1; Ikeda, H. et al., 2006).
This is discussed in greater detail elsewhere 3. Electrical stimulation of nerves evokes regular and
(Sandkuhler, J., 2007). highly synchronous discharges which is not
typical for naturally occurring, irregular nonsyn-
chronous discharge patterns. Thus, it was
important to show that LTP can also be induced
29.2 Induction of Long-Term by excitation of sensory nerve endings, for exam-
Potentiation in Pain Pathways ple, by subcutaneous injection of either capsaicin
or formalin. In animals with spinal cord and des-
In superficial spinal lamina I, LTP is selectively cending pathways intact, intraplantar, s.c.
induced in a group of nociceptive specific neurons injections of capsaicin or formalin induce slowly
which express the NK1 receptor for substance P and rising LTP (Figure 1; Ikeda, H. et al., 2006).
which project to the brain (see Figure 1; Ikeda, H. et al., 4. Finally, with descending spinal pathways inter-
2003; Ikeda, H. et al., 2006). This group of neurons rupted noxious skin heating and acute nerve
plays a key role in hyperalgesia following inflamma- injury also causes LTP at C-fiber synapses
tion and nerve injury (Mantyh, P. W. et al., 1997). In
(Sandkuhler, J. and Liu, X., 1998). This demon-
spinal cord LTP can be induced in different ways, for
strates that naturally occurring, low frequency,
example, by electrical nerve stimulation , by nerve
irregular, and nonsynchronous afferent barrage
injury, or peripheral noxious events. Stimulation
can cause synaptic plasticity in pain pathways.
within the innervation territory of a sensory nerve,
for example, with chemical irritants, or by inflamma-
tion may also trigger LTP at C-fiber synapses. LTP of C-fiber-evoked spinal field potentials can
All conditioning stimuli tested so far which induce also be induced pharmacologically. In spinalised rats,
LTP at A-synapses (Randic, M. et al., 1993) and/ or topical application of N-methyl-D-aspartate (NMDA),
C-fiber synapses (Liu, X. G. and Sandkuhler, J., 1995) substance P or neurokinin A induces LTP (Liu, X. G.
may also cause hyperalgesia in experimental animals. and Sandkuhler, J., 1998). In intact animals, spinal
 Long-Term Potentiation in Pain Pathways 403

(a) (b)
300 300
Capsaicin/solvent Formalin/solvent
200 200

Area of C-fiber-evoked postsynaptic response (% of control) 100

100

0 0
0 120 240 0 120 240

(c) (d)

300 300 LF S
LFS
200 LFS 200

100 100

MK-801
0 0
0 120 240 0 120 240

(e) (f)
300
400
LFS LF S
200 300
LF S
200
100
100
L-NMMA
0
0 120 240 0 20 40 60
Time (min)
Figure 1 LTP can be induced by natural, low-frequency afferent barrage evoked by inflammation of peripheral tissue in vivo and
by low-frequency stimulation of dorsal root afferents in vitro. Mean time courses of C-fiber-evoked field potentials recorded
extracellulary in superficial spinal dorsal horn in response to electrical stimulation of left sciatic nerve of deeply anaesthetized adult
rats with spinal cords and afferent nerves intact (a)(e). Subcutaneous injections of transient receptor potential vanilloid 1 channel
agonist capsaicin (1%, 100 ml, n 5, (a) or formalin (5%, 100 ml, n 6), (b) into the glabrous skin at the ipsilateral hind paw, within
the innervation territory of the sciatic nerve at time zero (arrows) induced LTP (closed circles), while injections of the respective
solvents (open circles) had no effects (n 3 in each group). Conditioning electrical LFS (2 Hz, 2 min at C-fiber intensity) of sciatic
nerve at time zero (arrow) also induced LTP (n 28), (c) which was prevented by NMDA receptor antagonist MK-801 (3 mg kg 1,
i.v.-infusion over 30 min: horizontal bar, n 5); (d) a second conditioning LFS four hours later (arrow) was partially effective in
inducing LTP. NOS inhibitor L-NMMA (100 mg kg1 h1, i.v., horizontal bar, n 5); (e) also blocked LTP induction. This block was
fully reversible as shown by a second LFS 3 h later (arrow). Similarly, LFS of dorsal roots at C-fibre strength induces robust LTP at
synapses between C-fibre afferents and lamina I neurons with a projection to the midbrain periaqueductal gray (PAG) in a rat spinal
corddorsal root preparation (n 18). Adapted from Ikeda, H., Stark, J., Fischer, H., Wagner, M., Drdla, R., Jager, T., and
Sandkuhler, J. 2006. Synaptic amplifier of inflammatory pain in the spinal dorsal horn. Science 312, 16591662.

application of dopamine D1/D5 or protein kinase A hyperalgesia in behaving animals. At C-fiber synapses,
activators but not D2 receptor agonists selectively LTP induction requires co-activation of ionotropic glu-
induces protein synthesis-dependent late phase of tamate receptors of the NMDA type (Randic, M. et al.,
LTP (Yang, H. W. et al., 2005). 1993; Liu, X. G. and Sandkuhler, J., 1995; Ikeda, H. et al.,
2003; 2006; see, however, Hamba, M. et al. 2000) and
group I but not group II or group III metabotropic
29.3 Signal Transduction Pathways glutamate receptors (mGluRs) (Hamba, M. et al., 2000;
Zhong, J. et al., 2000; Azkue, J. J. et al., 2003). LTP-
The signal transduction pathways which lead to LTP in inducing conditioning stimuli release substance P in
spinal dorsal horn largely overlap with those involved in spinal cord (Afrah, A. W. et al., 2002) and activation of
404 Long-Term Potentiation in Pain Pathways

NK1 receptors for substance P and NK2 receptors for inhibition in spinal cord may be impaired.
neurokinin A are required for LTP induction (Liu, X. G. Conditioning stimulation at 50 Hz produces a short
and Sandkuhler, J., 1997). Opening of T-type voltage lasting potentiation followed by LTD in control ani-
gated calcium channels is also necessary (Ikeda, H. et al., mals but LTP in animals with a chronic constriction
2003; 2006). All this leads to an activity-dependent injury of sciatic nerve (Miletic, G. and Miletic, V.,
rise in intracellular calcium ion concentration and acti- 2000). Topical application of muscimol (10 mg) to
vation of calcium-dependent signal transduction. The spinal cord prevents tetanus-induced LTP of A-
magnitude of HFS-induced LTP of C-fiber-evoked fiber-evoked field potentials in animals with a nerve
responses is linearly correlated with the rise in postsy- injury. In the presence of GABAA receptor agonist
naptic [Ca2]i (Ikeda, H. et al., 2003). Blockade of muscimol the same conditioning stimulation evokes
calcium rise in the postsynaptic neuron or calcium- an LTD rather than an LTP (Miletic, G. et al., 2003).
calmodulin-dependent protein kinase II prevents LTP Pharmacological activation of spinal NMDA
induction (Ikeda, H. et al., 2003; see, however, Hamba, receptors and NK1 or NK2 receptors is insufficient
M. et al., 2000; Yang, H. W. et al., 2004). Activation of to induce LTP in intact animals. When descending
group I mGluRs may activate phospholipase C (PLC) pathways, which are mainly inhibitory in acute pre-
pathway leading to either protein kinase C activation or parations, are cut, then the same receptor agonists
formation of inositol-1,4,5-trisphosphate (IP3). Blockade now trigger LTP (Liu, X. G. and Sandkuhler, J.,
of either PLC, PKC, or IP3 receptors all abolish LTP 1998). This confirms that level of postsynaptic inhi-
induction (Ikeda, H. et al., 2003). bition regulates induction and polarity of synaptic
Activation of NMDA, NK1, or NK2 receptors is plasticity. Some pharmacological induction protocols
not necessary for maintenance of LTP (Liu, X. G. do not require any presynaptic stimulation which
and Sandkuhler, J., 1997). Maintenance of the late suggests that LTP can be induced in the absence of
phase of LTP which starts about 3 h after induction presynaptic activity. This may be relevant when
requires, however, de novo protein synthesis (Hu, N. neuromodulators such as substance P diffuse to dis-
W. et al., 2003). tant, inactive synapses (volume transmission), which
is a potential mechanism of spreading synaptic plas-
ticity in the spinal cord.
29.3.1 Long-Term Potentiation Induction is
Functional blockade of glial cells by i.t. adminis-
Context Sensitive
tration of fluorocitrate changes the direction of HFS
Induction of LTP not only depends upon the activity induced plasticity of C-fiber-evoked potentials.
patterns at the presynaptic terminals, but also on the When HFS is given 1 h, but not 3 h after fluorocitrate
functional state of the postsynaptic neurons, namely LTD rather than LTP is induced (Ma, J. Y. and Zhao,
the level of postsynaptic inhibition. Under conditions Z. Q., 2002). The mechanism of this switch in polarity
of impaired inhibition, previously ineffective presy- of synaptic plasticity is presently unknown but recent
naptic activity may now induce LTP and protocols evidence suggests that glial cell-derived nitric oxide
which normally induce LTD may then cause LTP. In is liberated via activation of type 1 mGluRs and
vitro, both, A-fiber-evoked (Randic, M. et al., 1993) triggers LTP (Ikeda, H. and Murase, K., 2004).
and C-fiber-evoked (Ikeda, H. et al., 2003) responses
are potentiated by conditioning 100 Hz stimulation
of dorsal roots when postsynaptic neurons are mildly
depolarized to 70 to 60 mV. The same condition- 29.4 Prevention of Long-Term
ing stimulation induces, however, LTD of A-fiber- Potentiation
evoked responses if cells are hyperpolarized to
85 mV, suggesting that the direction of synaptic LTP of C-fiber-evoked field potentials in vivo is not
plasticity is voltage dependent (Randic, M. et al., affected by deep (surgical) level of anesthesia with
1993). urethane, isoflurane, or sevoflurane in mature rats
Spinal A-/A-fiber-evoked field potentials are (Benrath, J. et al., 2004). LTP is, however, prevented
depressed by conditioning 50 Hz stimulation of scia- by low-dose intravenous infusion of -opioid recep-
tic nerve. After blockade of GABAA receptors with tor agonist fentanyl (Benrath, J. et al., 2004). Similarly,
bicuculline the same conditioning stimulus now spinal field potentials elicited by stimulation in the
produces LTP rather than LTD (Miletic, G. and tract of Lissauer in spinal cord slices is blocked by
Miletic, V., 2001). In animals with a nerve injury, DAMGO, a more specific agonist at these receptors
 Long-Term Potentiation in Pain Pathways 405

(Terman, G. W. et al., 2001). Spinal application of 29.7 Conclusions

Zn2 also blocked LTP induction in vivo (Ma, J. Y.
and Zhao, Z. Q., 2001). Induction protocols signal transduction pathways and
pharmacology of LTP and some forms of hyperalgesia
are virtually identical, rendering LTP in spinal dorsal
29.5 Reversal of Long-Term horn an attractive cellular model of hyperalgesia.
Potentiation

Brief, high-frequency conditioning of sciatic nerve at

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potentiation of primary afferent neurotransmission at
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Hu, N. W., Zhang, H. M., Hu, X. D., Li, M. T., Zhang, T,
Zhou, L. J., and Liu, X. G. 2003. Protein synthesis inhibition
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fiber-evoked discharges largely overlap, which is in 2003. Synaptic plasticity in spinal lamina I projection neurons
line with the hypothesis that LTP underlies these that mediate hyperalgesia. Science 299, 12371240.
Ikeda, H., Stark, J., Fischer, H., Wagner, M., Drdla, R., Jager, T.,
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Treede, R. D. 2004. Perceptual correlates of nociceptive
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 30 Immune System, Pain and Analgesia
H L Rittner, H Machelska, and C Stein, Charite Universitatsmedizin Berlin, Campus Benjamin Franklin,
Berlin, Germany
2009 Elsevier Inc. All rights reserved.

30.1 Introduction 407

30.2 Proalgesic Mechanisms 408
30.2.1 Proinflammatory Cytokines 408
30.2.1.1 Tumor necrosis factor- 409
30.2.1.2 Interleukin-1 410
30.2.1.3 Interleukin-6 411
30.2.1.4 Interleukin-18 412
30.2.2 Chemokines 412
30.2.3 Nerve Growth Factor 413
30.2.4 Other Mediators: Bradykinin 414
30.2.5 Clinical Implications and Perspectives 415
30.3 Analgesic Mechanisms 415
30.3.1 Anti-Inflammatory Cytokines 415
30.3.2 Opioid Peptides 416
30.3.2.1 Peripheral opioid receptors 416
30.3.2.2 Opioid peptides produced by immune cells 417
30.3.2.3 Migration of opioid-containing immune cells to inflamed tissue 418
30.3.2.4 Release of opioid peptides from immune cells 420
30.3.2.5 Analgesia produced by immune-derived opioid peptides 420
30.3.2.6 Clinical implications 422
30.3.2.7 Perspectives 422
References 423

Glossary
CFA Complete Freunds adjuvant. NMDA N-methyl-D-aspartate.
CCR/CXCR Chemokine receptor. PC Prohormone convertase.
CRF Corticotropin releasing factor. PECAM-1 Platelet-endothelial cell adhesion
COX Cyclooxygenase. molecule-1.
CXCL1 KC keratinocyte-derived chemokine. PENK Proenkephalin.
CXCL2/3 MIP-2 macrophage inflammatory POMC Proopiomelanocortin.
protein. SRIF Somatotropin release inhibiting factor
CXCL8 IL-8. (somatostatin).
DRG Dorsal root ganglion. TNF- Tumor necrosis factor-.
GDNF Glial cell-line-derived neurotrophic factor. TrkA Tyrosine receptor kinase A.
ICAM-1 Intercellular adhesion molecule-1. TRPV1 Vanilloid receptor-1 (transient receptor
IL Interleukin. potential vanilloid 1).
IL-1ra IL-1 receptor antagonist. TTX Tetrodotoxin.
NGF Nerve growth factor.

407
408 Immune System, Pain and Analgesia

30.1 Introduction injection, or peritoneal inflammation using acetic

acid or glycogen.
Tissue destruction or inflammation are accompanied Neuropathic pain can arise following injury of
by an invasion of immune cells and liberation of peripheral nerves, when damaged or neighboring
numerous mediators. While these mediators contribute undamaged nerve fibers are sensitized or fire ectopi-
to the bodys ability to counteract the destruction of cally. It is also characterized by mechanical and
tissue integrity, they also elicit pain by activation of thermal hyperalgesia. In addition, animals with neu-
specialized primary afferent neurons called nociceptors ropathy are sensitive to stimuli that do not evoke
(reviewed in Julius, D. and Basbaum, A. I., 2001). They pain behavior under normal conditions, e.g., to
are defined as receptors preferentially sensitive to a touch, cooling, or warming. This is called allodynia
noxious stimulus or to a stimulus which would become (Merskey, H. and Bogduk, N., 1994). The most com-
noxious if prolonged (definition of the International mon models of neuropathic pain are tight ligation of
Association for the Study of Pain, IASP) (Merskey, H. spinal nerves and tight or loose ligation of the sciatic
and Bogduk, N., 1994). Trigeminal and dorsal root nerve. These models differ in the degree of neuronal
ganglia (DRG) contain nociceptor cell bodies which damage but all resemble human neuropathy resulting
give rise to myelinated A and unmyelinated C fibers. from trauma-induced injury of peripheral nerves (e.g.
A and C fibers transduce and propagate noxious entrapment, chronic nerve compression) (Bennet, G.
stimuli to the dorsal horn of the spinal cord from J., 1994).
where these stimuli are transmitted to the brain. At In the following section the effects of different
the level of the spinal cord and at supraspinal sites, mediators including cytokines, chemokines, nerve
various neurotransmitters come into plays which, growth factor (NGF), and bradykinin are discussed
together with environmental and cognitive factors, in three aspects: injection of the mediator into non-
contribute to the eventual sensation of pain (Scholz, J. inflamed tissue, blocking the mediator in models of
and Woolf, C. J., 2002). Strictly speaking, analgesia is inflammatory pain, and the role of the mediator in
defined as the inhibition of pain in man, while anti- neuropathic pain.
nociception is defined as the inhibition of behavioral
responses to noxious stimuli in animals. Although cen-
tral mechanisms also play a prominent role, the
30.2.1 Proinflammatory Cytokines
following chapter will focus on the peripheral injured
tissue itself. Cytokines are small proteins produced in the
inflamed tissue as well as in lymphoid organs with
stimulatory or inhibitory properties on immune
function. Cytokines act through high-affinity recep-
30.2 Proalgesic Mechanisms tors. Among the many different cytokines, the type I
cytokines have a similar four  helical bundle struc-
Inflammatory pain is characterized by an increased ture. Their receptors correspondingly share
response to mechanical and heat stimuli, which are characteristic features that have led to their descrip-
normally painful (Merskey, H. and Bogduk, N., tion as the cytokine receptor superfamily, or type I
1994). This is called mechanical or thermal hyper- cytokine receptors. Cytokine receptors in general are
algesia, respectively. After tissue injury, local tissue characterized by low abundance and a multisubunit
macrophages and dendritic cells are activated. The structure yielding low- and high-affinity receptor
inflammatory response is amplified by migration of forms. They are integral membrane glycoproteins
leukocytes into the inflamed tissue, by production of with the N-terminal outside and a single mem-
inflammatory mediators as well as tissue acidifica- brane-spanning domain. Unlike growth factors that
tion. Several agents are used experimentally to contain an instrinsic tyrosine kinase in their cytoplas-
induce nonspecific inflammation in experimental mic domain, cytokine receptors induce tyrosine
animals. The most widely used are carrageenan and phosphorylation via additional molecules. Multiple
complete Freunds adjuvant (CFA) injected into the signaling pathways/molecules have been observed
hindpaw of rodents. Carrageenan is used for short- for various cytokines. Collectively, these include
term inflammation, while CFA is used to model the Jak-STAT pathway, the Rasmitogen-activated
longer time courses of tissue injury. Less frequently protein (MAP) kinase pathway, Src and ZAP70 and
used agents include lipopolysaccharide, zymosan related proteins, phosphatidylinositol 3-kinase
 Immune System, Pain and Analgesia 409

(PI3K), insulin receptor substrates 1 and 2 (IRS-1 and dose dependently sensitized nociceptors to mechan-
IRS-2), and phosphatases. ical stimuli. This effect was restricted to A and C
Cytokines are produced on demand and travel fibers and no effect was seen in A mechanoreceptive
only over short distances. In vivo concentrations are fibers. Interestingly, however, higher doses of TNF-
in the range of a few picograms to nanograms per  produced lower responses (U-shaped dose
milliliter. Cytokines are pleiotropic and redundant, response). The authors postulated that this was
they influence mostly immune cells and the inflam- related to TNF--induced release of anti-inflamma-
matory response. Recently, however, cytokines have tory cytokines (Sorkin, L. S. et al., 1997; Junger, H.
been shown to link the immune system and the and Sorkin, L. S., 2000).
nervous system and they seem to modulate pain TNF- exerts its effects through two known
and hyperalgesia. receptors, TNFR1 and TNFR2. Expression of both
was evaluated in DRG from rats (Li, Y. et al., 2004):
30.2.1.1 Tumor necrosis factor- TNFR1 mRNA was expressed in virtually all neu-
The tumor necrosis factor (TNF) recptor superfam- rons and in nonneuronal cells with increased levels in
ily is comprised of at least 19 genes encoding 20 type models of peripheral inflammation. TNFR2 was
II (i.e., intracellular N-terminus, extracellular C-ter- exclusively expressed and regulated in nonneuronal
minus) transmembrane proteins. Included are several cells. In situ hybridization revealed varying levels of
well-known ligands such as TNF-, TNF- (lym- TNFR1 mRNA in virtually all DRG neurons includ-
photoxin-, LT-), Fas ligand, and CD40 ligand, as ing putative nociceptive neurons coding for
well as an increasing number of newly described calcitonin gene-related peptide, substance P, or
mediators. TNF- is the prototypic proinflamma- vanilloid receptor 1 (transient receptor potential
tory cytokine due to its role in initiating a cascade vanilloid 1; TRPV1). Similarly, expression of
of cytokines and growth factors in the inflammatory TNFR1 and TNFR2 protein was seen in rat DRG
response. Intraplantar injection of TNF- into non- neurons and their axons and increased in models of
inflamed paws produced a short-lived dose- neuropathic pain (Shubayev, V. I. and Myers, R. R.,
dependent mechanical hyperalgesia in rats (Cunha, 2001; Schafers, M. et al., 2003c). In contrast, TNFR1
F. Q. et al., 1992). TNF- seems to have indirect as is not expressed in DRG neurons from normal mice
well as direct hyperalgesic properties. Indirect action but is seen in mice with nerve injury (Ohtori, S. et al.,
is supported by the following evidence: inhibitors of 2004). TNFR localization on peripheral endings has
prostaglandin synthesis via cyclooxygenase (COX) not been examined but seems likely because of
inhibitors, and antagonists of adrenergic receptors expression in DRG and its presence in axons.
can attenuate TNF--induced hyperalgesia in rats. Some studies are available regarding the role of
In mice, TNF- action is only dependent on the endogenous TNF- in inflammatory hyperalgesia.
prostaglandin pathway (Cunha, T. M. et al., 2005). In one study, antisera against TNF- could comple-
Interestingly, the actions of TNF- seem to be tely block hyperalgesia induced by intraplantar
longer lasting if TNF- is administered repetitively. carrageenan injection (Cunha, F. Q. et al., 1992).
Such treatment resulted in hyperalgesia lasting up to Similar results are seen in lipopolysaccharide-
30 days after cessation of the daily TNF- injections induced inflammation (Ferreira, S. H. et al., 1993).
in rats (Sachs, D. et al., 2002). TNF--induced hyper- A third study examined TNF- in CFA-induced
algesia in this study was again dependent on release hindpaw inflammation (Woolf, C. J. et al., 1997).
of prostaglandins, as well as sympathetic amines, Here, a single injection of anti-TNF- serum before
because blockade of either reduced hyperalgesia. CFA application significantly delayed the onset of
Other groups have demonstrated a role of NGF in the resultant inflammatory hyperalgesia and reduced
TNF--induced hyperalgesia. Pretreatment of rats interleukin (IL)-1 but not NGF levels measured
with anti-NGF antiserum significantly attenuated 24 h later. A role for TNF- in inflammatory pain
TNF--induced mechanical and thermal hyperalge- has also been confirmed in mice using TNFR1
sia (Woolf, C. J. et al., 1997). A direct action on knockout mice (Cunha, T. M. et al., 2005). The inten-
nociceptors was postulated in electrophysiological sity of hyperalgesia after intraplantar carrageenan
studies (Sorkin, L. S. et al., 1997; Junger, H. and injection was significantly lower in these animals.
Sorkin, L. S., 2000). TNF- applied onto the sciatic Therefore, TNF- seems to be an endogenous med-
nerve or injected subcutaneously into normal paws iator in inflammatory pain in mice and in rats.
produced aberrant electrophysiological activity and Monocytes and tissue macrophages are the primary
410 Immune System, Pain and Analgesia

sources for TNF- synthesis. TNF- synthesis is mechanical allodynia (Schafers, M. et al., 2003d). In
stimulated by a wide variety of agents. In macro- vivo studies have recently been substantiated by elec-
phages, it is induced by biological, chemical, and trophysiological studies. In vitro perfusion of DRG
physical stimuli such as viruses, bacterial and parasitic cells with TNF- elicits neuronal discharges in A
products, tumor cells, complement, cytokines, ische- and C fibers, which are markedly higher and longer
mia, trauma, and irradiation (Figure 1) (Cunha, F. Q. lasting after nerve injury (Schafers, M. et al., 2003b).
and Ferreira, S. H., 2003; Cunha, T. M. et al., 2005). In This demonstrates an increased sensitivity of injured
peripheral tissues of rats, TNF- activity is followed neurons to TNF-. TNF--induced hyperalgesia in
by increases in IL-6 and IL-8, and IL-6 is followed by this model is mediated via TNFR1 (Schafers, M.
IL-1 and then by NGF. All of these finally mediate et al., 2003c). Downstream effectors of TNF-
their effects through prostaglandins or sympathetic include the p38 MAP kinase pathway. Intrathecal
amines. This cascade is also seen with few modifica- infusion of a p38 inhibitor reduced neuropathic pain
tions in mice (Cunha, T. M. et al., 2005). if it was started before but not 7 days after spinal
The role of TNF- has also been studied in nerve ligation. Cessation of therapy resulted in
neuropathic pain. TNF- mRNA was upregulated increased allodynia.
in the sciatic nerve at 2 weeks after its loose ligation In summary, TNF- has hyperalgesic properties
(a chronic constriction injury model) (Okamoto, K. if injected exogenously. It also has endogenous
et al., 2001). Also, TNF- protein was detected in hyperalgesic actions in inflammatory and neuro-
inflammatory cells accumulating at the ligated sciatic pathic pain. The effect seems to be mediated in part
nerve (Cui, J. G. et al., 2000; Schafers, M. et al., 2003a). indirectly via prostaglandins and sympathetic
Treatment of animals with neuropathic injury using amines. Recent data also suggest direct sensitizing
anti-TNF- antibodies and especially with the com- effects on sensory neurons.
bination of anti-TNF- and anti-IL-1 antibodies
30.2.1.2 Interleukin-1
significantly reduced signs of neuropathic pain
IL-1 and its family members are primarily proinflam-
(Schafers, M. et al., 2001). In another model, treatment
matory cytokines. They can stimulate the expression of
starting 2 days before spinal nerve ligation with the
genes associated with inflammation and autoimmune
TNF- receptor antagonist etanercept attenuated
diseases. The most salient and relevant properties of
IL-1 in inflammation are the initiation of COX, type 2
Early inflammation
phospholipase A, and inducible nitric oxide synthase.
Inflammatory
stimulus Hyperalgesic effects of IL-1 have first been demon-
Migrated leucocytes strated by injection into hindpaws of rats without
G and resident cells
M inflammation or nerve injury (Ferreira, S. H. et al.,
TNF- 1988): in contrast to IL-1, IL-1 is able to sensitize
nociceptors. Several mechanisms have been proposed
CXCL8 IL-6 Bradykinin to mediate this indirect action, including prostaglan-
or CXCL1

IL-1
dins, nitric oxide, NGF, and bradykinin. The
hyperalgesic effect of IL-1 is blocked by treatment
NGF
with indomethacin, a nonselective COX inhibitor
Sympathetic amines LTB4 Prostaglandins reducing the formation of prostaglandins (Figure 1)
(Ferreira, S. H. et al., 1988). In mice, IL-1 also induces
Hyperalgesia
prostaglandin formation mediating hyperalgesia
(Cunha, T. M. et al., 2005). Injection of IL-1 into
Figure 1 Hyperalgesic mechanisms in early inflammation. normal tissue induces thermal hyperalgesia and the
An inflammatory agent stimulates the migration of
leukocytes, e.g., granulocytes (G) and monocytes (M), into production of NGF. The thermal hyperalgesia but
the inflamed tissue. In the tissue, these leukocytes as well as not the NGF production can be prevented by pretreat-
resident cells initiate a cascade of cytokines including tumor ment with recombinant IL-1 receptor antagonist (IL-
necrosis factor- (TNF-) and interleukins (ILs), chemokines 1ra). There is also evidence of a direct action of IL-1
(CXCL8, CXCL1), nerve growth factor (NGF), bradykinin,
on nociceptors: intraplantar injection of IL-1 potenti-
and secondary mediators like sympathetic amines,
leukotriene B4 (LTB4), and prostaglandins culminating in ates action potentials in rat DRG neurons in response
hyperalgesia. TNF-, IL-6, IL-1, and bradykinin can also to thermal or mechanical stimuli, and a decrease of the
have direct hyperalgesic effects on nociceptors. mechanical threshold for nerve firing was observed
 Immune System, Pain and Analgesia 411

after local injection of IL-1 (Fukuoka, H. et al., 1994). 2001; Okamoto, K. et al., 2001). Blocking of IL-1R in
In the in vitro skinnerve preparation brief exposure to mice with chronic constriction injury reduces ther-
IL-1 facilitates the heat-induced release of calcitonin mal hyperalgesia, mechanical allodynia, and also
gene-related peptide (a pronociceptive mediator) from immunoreactivity for TNF- (Sommer, C. et al.,
peptidergic sensory neurons (Opree, A. and Kress, M., 1999). However, the mechanisms remain unclear. So
2000). This was confirmed by a follow-up study far, no study has demonstrated IL-1R expression in
(Obreja, O. et al. 2002a). IL-1 potentiated heat-activated neuropathy. In summary, IL-1 seems to have
inward currents and shifted the activation threshold hyperalgesic actions in different models of pain.
towards lower temperature without altering intracel- The mechanism direct or indirect still needs to
lular calcium currents. be clarified.
Intracellular mediators of these effects included
protein tyrosine kinases and protein kinase C. One of 30.2.1.3 Interleukin-6
the IL-1 receptors subtypes, the IL-1RI, was IL-6 is a one of the major proinflammatory cytokines.
expressed in DRG cells as shown by polymerase Its level is below detection under physiological con-
chain reaction and in situ hybridization. The authors ditions but is rapidly and strongly upregulated in
propose that recruitment of tyrosine kinases and early inflammation and can be detected early in the
downstream activation of protein kinase C phosphor- serum. Intraplantar injection of IL-6 induces hype-
ylate TRPV1 to cause heat hyperalgesia. Both ralgesia in normal rats (Poole, S. et al., 1995). An
mRNA and IL-1RI protein are found in DRG neu- indirect mechanism of this effect is supported by
rons and glial cells (Li, M. et al., 2005) of rats with and data showing inhibition of IL-6-induced hyperalgesia
without paw inflammation. However IL-1RI locali- when animals are treated with COX blockers
zation has not been shown on peripheral nerve (Cunha, F. Q. et al., 1992) (Figure 1). However, so
terminals. Using autoradiography, we did not detect far it is not known which cells and receptors are
IL-1 binding on nerves in either inflamed or nonin- involved.
flamed paws, but on immune cells in inflamed paws Direct effects have been proposed by other groups:
(Mousa, S. A. et al., 1996). Therefore, IL-1 receptor In the rat in vitro skinnerve preparation basal and
expression in the periphery requires further studies. heat-evoked release of calcitonin gene-related pep-
IL-1 also has an endogenous hyperalgesic role in tide from nociceptors was measured. IL-6 did not
inflammatory as well as in neuropathic pain. In hind- induce heat sensitization, when applied alone, but
paw inflammation induced by carrageenan or was effective in the presence of soluble IL-6 receptor
lipopolysaccharide IL-1ra can reduce, by not com- (sIL-6R) (Opree, A. and Kress, M., 2000). sIL-6R is
pletely block, hyperalgesia in rats and in mice needed for cells that express only the signaling recep-
(Ferreira, S. H. et al., 1993; Cunha, J. M. et al., 2000; tor unit gp130. The complex of sIL-6R and IL-6 can
Cunha, T. M. et al., 2005). The same effect can be seen then induce a signal to activate cells. This so-called
in CFA-induced hindpaw inflammation or in mice trans-signaling expands the spectrum of cells respon-
after intraperitoneal injection of acetic acid (Safieh- sive to IL-6 to include neurons. IL-6, in combination
Garabedian, B. et al., 1995; Cunha, J. M. et al., 2000). with its soluble receptor, can sensitize nociceptors to
In CFA-induced inflammation, the authors propose heat, providing evidence for the constitutive expres-
that the effect is mediated by NGF, because treatment sion of gp130, but not of the IL-6-membrane-bound
with IL-1ra also reduced the local production of NGF (specific) receptor, in nociceptors (Obreja, O. et al.,
(Safieh-Garabedian, B. et al., 1995). Interestingly, our 2002b). Hyper-IL-6 (HIL-6), a fusion protein of IL-6
group has shown analgesic effects of IL-1 via release and sIL-6R, was designed to mimic the effects of the
of opioid peptides from immune cells in CFA-induced IL-6sIL-6R complex. In vitro exposure of DRG neu-
inflammation (see Analgesic Effects of Corticotropin rons to HIL-6 potentiated heat-activated inward
Releasing Factor, Interleukin-1, and Noradreline for currents and shifted the activation thresholds towards
discussion). lower temperatures without affecting intracellular
In neuropathic pain models, the role of IL-1 has calcium levels (Obreja, O. et al., 2005). This effect
not been thoroughly examined. Expression of IL-1 was reduced by a Janus tyrosine kinase inhibitor, a
is seen in inflammatory cells accumulating at the selective protein kinase C inhibitor as well as a selec-
sciatic nerve in ligation and inflammatory models of tive blocker of the protein kinase C delta isoform, but
neuropathy as assesed by immunohistochemistry and not by the COX inhibitor indomethacin. In situ hybri-
by polymerase chain reaction (Gazda, L. S. et al., dization and reverse transcription-polymerase chain
412 Immune System, Pain and Analgesia

reaction revealed expression of the signal-transducing healing. They can be divided into subfamilies based
beta subunit of gp130 in neuronal somata but not in on their structural motifs (the position of cysteine
satellite cells in DRG from normal rats (Gadient, R. residues located near the N-terminus of the protein:
A. and Otten, U., 1996; Obreja, O. et al., 2005). These CC, CXC, and CX3C). Chemokines bind to a group
data suggest a direct activation of sensory neurons by of chemokine receptors (CCR, CXCR, and CX3CR;
IL-6sIL-6R. seven transmembrane-spanning Gi protein-coupled
As seen for IL-1 and TNF-, endogenous IL-6 is receptors) expressed on leukocytes. Although each
also important for the development of hyperalgesia: receptor subtype typically binds multiple chemo-
Antisera neutralizing IL-6 inhibited lipopolysacchar- kines, the specificity is defined by a chemokine
ide-induced hyperalgesia in rats (Ferreira, S. H. et al., subfamily. Recently, chemokines and their receptors
1993). IL-6 knockout mice were shown to have have been detected on cells in the central and
reduced thermal and mechanical hyperalgesia after peripheral nervous system.
injection of carrageenan into the hindpaw (Xu, X. J. Several observations indicate a role of chemokines
et al., 1997). In this study, however, IL-6 knockout mice in hyperalgesia. Chemokine receptors such as
without inflammation displayed baseline hyperalgesia, CXCR4 and CCR4 are expressed on subpopulations
which could not be explained by the authors. of DRG neurons and their corresponding ligands
In neuropathic mice, IL-6 mRNA levels were can induce calcium influx (Oh, S. B. et al., 2001).
upregulated in DRG and correlated well with the However, in neonates the percentage of neurons
development of nerve injury-induced thermal hype- responding to chemokine stimulation was signifi-
ralgesia and mechanical allodynia (Murphy, P. G. cantly lower than the percentage responding to
et al., 1999). IL-6 knockout mice had reduced thermal bradykinin or capsaicin. Direct intraplantar injection
and mechanical hyperalgesia in the chronic constric- of some chemokines (CCL5, CXCL12, and CCL22)
tion injury model. In summary, IL-6 belongs to the induces hyperalgesia in normal animals (Oh, S. B.
group of proinflammatory hyperalgesic cytokines et al., 2001). In addition, some chemokines (human
with direct as well as indirect effects on neurons, CXCL8 (IL-8) or rat CXCL1 (KC keratinocyte-
similar to IL-1 and TNF-. derived chemokine)) can indirectly cause hyperal-
gesia through release of sympathetic amines
30.2.1.4 Interleukin-18 when applied subcutaneously into the hindpaw
IL-18 is a member of the IL-1 ligand superfamily and (Lorenzetti, B. B. et al., 2002; Cunha, F. Q. and
is expressed in various cell types including macro- Ferreira, S. H., 2003). Hyperalgesic effects of CXCL1
phages and dendritic cells. Its biological functions were further examined in mice (Cunha, T. M. et al.,
include stimulation of activated T cells and enhance- 2005). Intraplantar injection of a small dose of CXCL1
ment of natural killer cell lytic activity. Furthermore, induced hyperalgesia with a peak between 3 and 5 h
IL-18 activates and attracts neutrophils by inducing returning to normal after 24 h. Pretreatment of mice
the production of TNF- and subsequently leuko- with indomethacin, a COX inhibitor, or guanethidine,
triene B4. Injection of IL-18 into normal paws caused a blocker of sympathetic amines, reduced the hyper-
time and dose-dependent mechanical hyperalgesia algesia. An almost complete blockade was achieved if
(Verri, W. A., Jr. et al., 2004) Interestingly, only mor- both agents were combined. Interestingly, CXCL1-
phine and dexamethasone blocked IL-18-induced induced hyperalgesia could also be inhibited by IL-
hyperalgesia, whereas blockade of the prostaglandin 1ra but no change was seen in TNFR1 knockout mice.
or sympathetic pathway had no effect. In fact, the Therefore, IL-1 seems to be downstream of CXCL1,
hyperalgesia elicited by IL-18 seems to be mediated whereas TNF- is upstream. In contrast, our own
via endothelin receptors, because it was abolished by unpublished data suggest that intraplantar injection
blockage of endothelin receptor B (Verri, W. A., Jr. of CXCL1 as well as CXCL2/3 (MIP-2 macrophage
et al., 2004). inflammatory protein) in normal rats does not evoke
thermal or mechanical hyperalgesia for up to 12 h
postinjection despite significant recruitment of granu-
30.2.2 Chemokines
locytes. Differences could be explained by the doses or
Chemokines (CC and CXC) are chemotatic media- the behavioral test used (see below). Doses producing
tors produced in inflamed tissue. They control maximal chemokine-induced granulocyte recruitment
trafficking of leukocytes under physiological and do not induce hyperalgesia in our model Rittner, H. L.,
inflammatory conditions, angiogenesis, and wound et al. 2006.
 Immune System, Pain and Analgesia 413

In a recent study, a connection between chemokines attracting monocytes into inflamed tissue.
and TRPV1 was shown (Zhang, N. et al., 2005). CCL3 Therefore the authors conclude that the decreased
(macrophage inflammatory protein 1) that binds to number of monocytes is responsible for the reduced
CCR1 was tested in vitro and in vivo. Activation of pain in this model. In summary, the contribution of
CCR1 resulted in increases in TRPV1-mediated chemokines to hyperalgesia may depend on their
Ca2 influx and increased sensitivity of TRPV1 to its type, the model of pain, the time frame and the
ligand capsaicin. In vivo the latency in the mouse hot dose of chemokine used. Some of the indirect effects
plate test was reduced for up to 45 min after a local are mediated by the ability of chemokines to attract
injection of CCL3. Thus, CCL3 seems to be capable of monocytes.
enhancing the sensitivity of TRPV1, possibly through
a G protein-dependent signaling pathway.
30.2.3 Nerve Growth Factor
The endogenous role of chemokines was studied
in rats and mice in carrageenan-induced hindpaw NGF belongs to the family of neurotrophic proteins,
inflammation. In studies by Ferreira et al., human together with brain-derived neurotrophic factor
CXCL8 and rat CXCL1 were shown to contribute (BDNF), neurotrophin-3 (NT-3), NT-4/5, and NT-
to carrageenan-induced hyperalgesia in rats (Cunha, 6. NGF governs the innervation of target tissues during
F. Q. et al., 1991; Lorenzetti, B. B. et al., 2002; Sachs, D. development and plays an important role in neuronal
et al., 2002). Similarly, injection of carrageenan eli- survival and maintenance of connectivity. Besides its
cited hyperalgesia, which could be reduced by action on neurons, NGF also has immunomodulatory
pretreatment with anti-CXCL1 antibodies in mice effects in different inflammatory diseases including
(Cunha, T. M. et al., 2005). Therefore, part of the asthma, arthritis, and psoriasis. NGF is constitutively
hyperalgesia seen in carrageenan inflammation is expressed in keratinocytes in normal skin. However,
dependent directly or indirectly on certain chemo- many cells including all subtypes of leukocytes as well
kines. However, our own studies in CFA-induced as sensory neurons can express NGF in inflammation
inflammation suggest an additional role of chemo- (Vega, J. A. et al., 2003). After binding to its high-
kines in counteracting inflammatory pain. In this affinity receptor tyrosine receptor kinase A (trkA) on
model, concomitant injection of CXCL2/3 and peripheral terminals of sensory neurons, NGF is inter-
CFA induced a threefold increase in the number of nalized and retrogradely transported to the somata in
neutrophils in the inflamed paw after 2 h, while the DRG. NGF is a potent regulator of gene expres-
mechanical and thermal hyperalgesia was not chan- sion of neuropeptides like calcitonin gene-related
ged (Brack, A. et al., 2004d). Furthermore, inhibition peptide and substance P, of receptors like TRPV1
of CXCL1 and CXCL2/3 in inflamed paws had no and of ion channels like tetrodotoxin (TTX)-resistant
effect on mechanical hyperalgesia despite significant sodium channels. The role of endogenous NGF has
reduction of the number of infiltrating neutrophils mostly been examined using proteins that block the
after 2 h (Brack, A. et al., 2004c) (see also Analgesic bioactivity of NGF because deletion of NGF or trkA
Effects of Corticotropin Releasing Factor, in mice is lethal within the first week of life.
Interleukin-1, and Noradreline and Endogenous Injection of NGF into the normal rat paw induces a
Opoid Analgesia). The differences between our stu- long-lasting thermal as well as mechanical hyperalgesia
dies and those by Cunha et al., might be related to the (Lewin, G. R. et al., 1994). These effects seem to be
model of inflammation (CFA versus carrageenan) or in part dependent on mast cells and on a central
to the type of behavioral test used. While we deter- component involving N-methyl-D-aspartate (NMDA)
mined the amount of mechanical pressure required receptors. Nociceptor sensitization was also observed in
for the rat to withdraw its paw (modified Randall vitro in a skinnerve preparation (Rueff, A. and
Selitto method), Cunha et al., used a substantially Mendell, L. M., 1996). Infusion of trkAIgG into the
more painful stimulus that induced sympathetic acti- noninflamed rat paw to block local NGF diminished
vation with a freezing reaction and apnea. thermal hyperalgesia and decreased calcitonin gene-
In neuropathic pain the role of chemokines has related peptide content in DRG neurons (McMahon,
not been thoroughly examined. Disruption of the S. B. et al., 1995). The detection of sensitizing properties
gene for CCR2 prevented mechanical allodynia of NGF has been corroborated by the analysis of trans-
in the chronic constriction injury model (Abbadie, genic mice overexpressing NGF in the epidermis
C. et al., 2003). CCR2, activated by CCL2 (monocyte (Stucky, C. L. et al., 1999). These animals developed
chemotrattractant protein-1), is responsible for hyperalgesia without any signs of inflammation of the
414 Immune System, Pain and Analgesia

skin. In electrophysiological studies unmyelinated noci- DRG neurons but not in the spinal cord between
ceptors responded with a fourfold intensity to heat wild-type and NGF- or GDNF-overexpressing
stimuli in comparison to wild-type mice. Moreover, animals. This suggested that the somatosensory sys-
almost every C fiber responded to heat in NGF over- tem is capable of producing normal pain behavior
expressing animals in contrast to only half of the C despite significant modification of nociceptors.
fibers in wild-type animals. No enhanced response to NGF blockade seems to produce analgesia also in
mechanical stimuli was seen. Together, this provides other pain states. In a model of bone cancer pain,
supporting evidence for the heat-sensitizing property of osteosarcoma tumor cells are injected into the
NGF. Importantly, endogenous NGF seems to be intramedullary space of the mouse femur. In a
responsible for maintenance of the sensitivity of noci- recent study, the authors focused on a novel
ceptors as application of trkAIgG fusion molecule to NGF-sequestering antibody and demonstrated that
the innervation territory of the cutaneous saphenous two administrations produce a profound reduction
nerve resulted in a reduction of innervation density of in both ongoing and movement-evoked bone cancer
the epidermis and of the percentage of neurons pain-related behaviors that was greater than
responding to heat or bradykinin (Bennett, D. L. et al., that achieved with acute administration of 10 or
1998). 30 mg kg1 of morphine (Sevcik, M. A. et al., 2005).
Blocking of NGF by trkAIgG in carrageenan- In summary, neutralization of NGF produces analge-
induced inflammation prevents the development of sia in most pain models studied.
thermal hyperalgesia and the sensitization of nocicep-
tors despite normal development of inflammation as
30.2.4 Other Mediators: Bradykinin
measured by tissue edema (McMahon, S. B. et al., 1995;
Koltzenburg, M. et al., 1999). Similarly, injection of anti- Bradykinin is a nonapeptide generated from high-
NGF antibodies blocks the development of thermal molecular-weight kininogen through cleavage by kal-
hyperalgesia in CFA-induced inflammation (Lewin, likrein under inflammatory conditions in plasma. It is
G. R. et al., 1994). Thus, NGF seems to be part of the also released from mast cells during asthma attacks and
inflammatory cascade involving initially inflammatory within the gut as a gastrointestinal vasodilator.
cytokines like IL-1 and TNF- and subsequently Bradykinin is rapidly generated after tissue injury and
NGF (Figure 1) (Cunha, T. M. et al., 2005). seems to modulate most of the events observed during
Neuropathic pain is also influenced by NGF; the inflammatory process including vasodilatation,
however, the data seem to be controversial. increase of vascular permeability, plasma extravasation,
Treatment with anti-NGF in the chronic constric- and cell migration (reviewed in Calixto, J. B. et al.,
tion injury model reduced hyperalgesia but the effect 2000). It is also one of the most potent endogenous
showed a delayed onset, short duration, and depen- proalgesic mediators (Ferreira, S. H. et al., 1993). It
dency on the dosage (Ro, L. S. et al., 1999). In a recent directly activates bradykinin receptors on primary
paper the role of NGF and glial cell-line-derived afferent neurons or sensitizes nociceptors indirectly
neurotrophic factor (GDNF) on nociceptor function through release of prostaglandins, nitric oxide, neuro-
was examined (Zwick, M. et al., 2003). Two lines of kinins, calcitonin gene-related peptide, cytokines, and
transgenic mice that contained an increased number histamine (Figure 1), or via alterations of vanilloid
of either NGF- or GDNF-dependent neurons were receptor channel gating (Chuang, H. H. et al., 2001).
used. These mice were tested in a model of inflam- In fact, lipopolysaccharide- or carrageenan-induced
matory pain (CFA) and neuropathic pain (spinal inflammation triggers the TNF--driven cytokine cas-
nerve ligation model). Contrary to expectations, cade via the release of bradykinin (references in
neither line of transgenic mice became more hype- Cunha, F. Q. and Ferreira, S. H., 2003). Bradykinin is
ralgesic following induction of persistent pain. In fact, released at the beginning of the inflammation because
NGF-overexpressing mice recovered more rapidly delayed treatment with bradykinin receptor antago-
from initial inflammatory hyperalgesia and became nists 2 h after injection of lipopolysaccharide or
hypoalgesic despite extensive paw swelling in the carrageenan has no effect on hyperalgesia or
inflammatory model. In the neuropathic model, inflammation.
only wild-type mice became hyperalgesic. mRNA Bradykinin also plays a role in neuropathic pain.
expression of opioid receptors, NMDA receptors, Ablation of the gene for the bradykinin receptor
metabolic glutamate receptor, sodium channels and 1 resulted in a significant reduction of mechanical
vanilloid receptors showed marked differences in allodynia and thermal hyperalgesia in early stages of
 Immune System, Pain and Analgesia 415

nerve injury (Ferreira, J. et al., 2005). Furthermore, mediators include anti-inflammatory cytokines,
systemic treatment with the bradykinin receptor 1- somatostatin (somatotropin release inhibiting factor;
selective antagonist des-Arg9-[Leu8]-bradykinin SRIF), and the endocannabinoids.
reduced the established mechanical allodynia
observed 728 days after nerve lesion in wild-type
mice. Nonpeptidic orally active bradykinin receptor 30.3.1 Anti-Inflammatory Cytokines
1 and 2 antagonists are currently being developed as
In later stages of inflammation cytokines are pro-
peripheral analgesics (Sawynok, J., 2003; Marceau, F.
duced, which limit inflammation and counteract
and Regoli, D., 2004).
hyperalgesia (references in Cunha, F. Q. and
Ferreira, S. H., 2003). For example, IL-4, IL-10,
30.2.5 Clinical Implications and IL-13, and IL-1ra can produce analgesia (Figure 2)
Perspectives (Cunha, F. Q. et al., 1999; Cunha, J. M. et al., 2000;
Cunha, F. Q. and Ferreira, S. H., 2003). IL-4 and
A myriad of peripheral mediators is involved in
IL-13 are produced by Th2 lymphocytes and mast
inflammatory as well as neuropathic pain. The most
cells and suppress the production of proinflammatory
frequently used pain medications are selective or
mediators such as IL-1, TNF-, interferon-, and
nonselective inhibitors of COX. These nonsteroidal
CXCL8. In addition, inhibition of COX-2 is observed.
anti-inflammatory drugs inhibit the common final
IL-10 is a product of T-lymphocytes and monocytes
pathway of most proalgesic compounds, the produc-
and inhibits the production of Th1 proinflammatory
tion of prostaglandins. However, these drugs have
cytokines including IL-1, IL-6, IL-8, and TNF-.
serious side effects such as gastrointestinal ulcer for-
Besides, it upregulates IL-1ra. These effects are also
mation, bleeding and/or, thromboembolic events.
seen in models of pain. Pretreatment with IL-4, IL-10,
Because of the multitude of proinflammatory and
and IL-13 dose dependently blocked hyperalgesia
proalgesic mediators the development of new analge-
induced by carrageenan, bradykinin, and TNF-
sics is difficult. Inhibiting one pathway may still leave
but did not affect hyperalgesia induced by CXCL8
other mediators available. On the other hand, cyto-
and prostaglandins. Furthermore, the endogenous
kine antagonists such as the TNF- receptor
role of anti-inflammatory cytokines to limit hyperal-
antagonist etanercept have been developed for treat-
gesia was demonstrated by application of antisera
ment of autoimmune disease like rheumatoid
against IL-4, IL-10, and IL-13. They potentiated
arthritis and have been shown to produce analgesia
hyperalgesia induced by carrageenan, bradykinin,
in patients (Sommer, C. et al., 2001). Despite the
almost innumerable number of mediators new treat-
ments are under current investigation including, e.g., Late inflammation Inflammatory
stimulus
the blockade of the bradykinin system (Sawynok, J., Migrated leucocytes
2003; Marceau, F. and Regoli, D., 2004). The deve- M `and resident cells
L
lopment of peripherally acting drugs would have TNF-
advantages such as the avoidance of central side
CXCL8 IL-6
effects. For example, the localized (e.g., topical) or CXCL1
IL-4, IL-10,
administration of drugs can potentially optimize IL-13
drug concentrations at the site of injury without IL-1

systemically active drug levels and fewer adverse NGF IL-1ra

systemic effects, fewer drug interactions, and no
need to titrate doses into a therapeutic range com- Sympathetic amines LTB4 Prostaglandins
pared with systemic administration.

Analgesia

30.3 Analgesic Mechanisms Figure 2 Analgesic cytokines in late inflammation. During

ongoining inflammation leukocytes, e.g., lymphocytes (L)
and monocytes/macrophages (M), start to produce anti-
Leukocytes are the source not only of hyperalgesic
inflammatory cytokines like interleukin-4 (IL-4), IL-10, IL-13,
mediators but also of antinociceptive mediators. The and IL-1 receptor antagonist (IL-1ra). These cytokines
best-characterized and clinically tested compounds inhibit the proinflammatory cytokines such as TNF-, IL-1,
are the endogenous opioid peptides. Other analgesic and IL-6, and block the cascade.
416 Immune System, Pain and Analgesia

and TNF-. This effect was not seen in athymic and characteristics (affinity) of peripheral and central
mast-cell-depleted rats. Thus, it seems that endogen- opioid receptors are similar but the molecular masses
ous sources of IL-4 and IL-13 are mast cells and of peripheral and central -receptors appear to be
lymphocytes, respectively. Analgesic actions of IL-4, different (references in Stein, C. et al., 2003). If these
IL-10, and IL-13 are not only seen in paw inflamma- findings are confirmed, a search for selective ligands at
tion but also in models of peritonitis and knee joint such distinct receptors may be warranted.
incapacitation induced by zymosan. Analgesic prop- It has been suggested that opioid receptors are
erties of anti-inflammatory cytokines are independent also located on sympathetic postganglionic neuron
of endogenous production of opioid peptides (Vale, terminals. However, there are reports arguing
M. L. et al., 2003). In summary, during ongoing inflam- against this notion and studies attempting the direct
mation analgesic cytokines are produced which demonstration of opioid receptor mRNA in sympa-
counteract the effects of the proinflammatory hyper- thetic ganglia have produced negative results. In
algesic cytokines generated in the early stages. addition, thorough morphological investigations
have clearly demonstrated the presence of -recep-
tors on unmyelinated primary afferent neurons and
30.3.2 Opioid Peptides
the absence of such receptors on postganglionic
30.3.2.1 Peripheral opioid receptors sympathetic neurons in skin, lip, and cornea.
30.3.2.1.(i) Localization Opioid peptides are the Moreover, chemical sympathectomy with 6-hydro-
natural ligands at opioid receptors. Three cDNAs xydopamine does not change the expression of
and their genes have been identified, encoding the - opioid receptors in the DRG or the peripheral
, -, and -opioid receptor (MOP, DOP, and KOP), analgesic effects of -, -, and -receptor agonists
respectively (Kieffer, B. L. and Gaveriaux-Ruff, C., in a model of inflammatory pain. Together, these
2002). All three receptors can mediate pain inhibition findings have corroborated the notion that periph-
and they are found throughout the nervous system, eral opioid receptors mediating analgesia are
including somatic and visceral sensory neurons, spinal exclusively localized on primary sensory neurons
cord projection and interneurons, midbrain, and cor- (references in Stein, C. et al., 2001).
tex. Recent interest has focused on the identification of Opioid binding sites and the expression of opioid
opioid receptors on peripheral processes of sensory receptor transcripts have also been demonstrated in
neurons. The cell bodies of these neurons in DRG immune cells. Opioid-mediated modulation of the
express all three mRNAs and receptor proteins proliferation of these cells and of their functions
(Stein, C. et al., 2001). Opioid receptors are intraaxon- (e.g., chemotaxis, superoxide and cytokine produc-
ally transported into the neuronal processes and are tion, mast cell degranulation) has been reported.
detectable on peripheral sensory nerve terminals in These immunomodulatory actions can be stimula-
animals and in humans. Colocalization studies con- tory as well as inhibitory and have been ascribed to
firmed the presence of opioid receptors on neurons the activation of opioid receptors (Sacerdote, P. et al.,
expressing substance P and/or calcitonin gene-related 2003; Sharp, B. M., 2003). However, the significance
peptide in DRG small- and medium-sized neurons of such effects with regard to pain transmission has
and in their central and peripheral terminals (Dado, not been investigated as yet.
R. J. et al., 1993; Zhang, X. et al., 1998; Wenk, H. N. and
Honda, C. N., 1999; Guan, J. S. et al., 2005). Further, 30.3.2.1.(ii) Opioid receptor signaling in sensory
anatomical and electrophysiological studies showed neurons All three types of opioid receptors medi-
expression of opioid receptors on unmyelinated C ate the inhibition of high-voltage-activated calcium
and on myelinated A fibers (i.e., nociceptors) currents in cultured primary afferent neurons. These
(Beland, B. and Fitzgerald, M., 2001; Pare, M. et al., effects are transduced by G proteins (Gi and/or Go)
2001; Suzuki, R. and Dickenson, A. H., 2002) but not (references in Stein, C. et al., 2001; 2003). Although it
on large myelinated A fibers (Arvidsson, U. et al., is well-known that opioids induce membrane hyper-
1995; Beland, B. and Fitzgerald, M., 2001). However, polarization due to increased potassium currents in
after nerve injury morphine depressed responses of A central neurons, this could not be detected in DRG
fibers (Suzuki, R. and Dickenson, A. H., 2002), and neurons so far (Akins, P. T. et al., 1993). Thus, the
expression -receptors on the A fibers in Meissner modulation of calcium channels appears to be the
corpuscules in glaborus skin has been recently primary mechanism for the inhibitory effects of
reported (Pare, M. et al., 2001). The binding opioids on peripheral sensory neurons. In addition,
 Immune System, Pain and Analgesia 417

opioids via inhibition of adenylyl cyclase sup- 2001). In addition, the specific milieu (low pH, pros-
press TTX-resistant sodium- and nonselective cation tanoid release) of inflamed tissue may increase opioid
currents stimulated by the inflammatory agent pros- agonist efficacy by enhanced G protein coupling and
taglandin E2 (Ingram, S. L. and Williams, J. T., 1994; by increased neuronal cyclic adenosine monopho-
Gold, M. S. and Levine, J. D., 1996). Interestingly, sphate levels (Selley, D. E. et al., 1993; Ingram, S. L.
TTX-resistant sodium channels are selectively and Williams, J. T., 1994; Zollner, C. et al., 2003).
expressed in nociceptors, they are important for Inflammation also leads to an increase in the number
impulse initiation and action potential conductance, of sensory nerve terminals (sprouting) and disrupts the
they mediate spontaneous activity in sensitized noci- perineurial barrier, thus facilitating the access of
ceptors and they accumulate at the site of injury in opioid agonists to their receptors (Stein, C. et al.,
damaged nerves leading to ectopic impulse gene- 2001; 2003).
ration (Porreca, F. et al., 1999; Laird, J. M. et al.,
2002). The latter observations may explain the nota- 30.3.2.2 Opioid peptides produced by
ble efficacy of peripheral opioids in inflammatory immune cells
and neuropathic pain (Stein, C. et al., 2001; 2003). Three families of opioid peptides are well characte-
Consistent with their effects on ion channels, opioids rized in the central nervous and neuroendocrine
attenuate the excitability of peripheral nociceptor systems (Akil, H. et al., 1998). Each family derives
terminals, the propagation of action potentials, the from a distinct gene and from one of the
release of excitatory proinflammatory neuropeptides three precursor proteins proopiomelanocortin
(substance P, calcitonin gene-related peptide) from (POMC), proenkephalin (PENK), or prodynorphin,
peripheral sensory nerve endings, and vasodilatation respectively. Appropriate processing yields their
evoked by stimulation of C fibers (references in respective representative opioid peptides, the endor-
Stein, C. et al., 2001). All of these mechanisms result phins, enkephalins, and dynorphins (Table 1). These
in analgesia and/or anti-inflammatory actions. peptides exhibit different affinity and selectivity for
the three opioid receptors  (endorphins, enkepha-
30.3.2.1.(iii) Peripheral opioid receptors and lins),  (enkephalins, endorphins), and (dynorphin).
inflammation Peripheral opioid analgesic effects Two additional endogenous opioid peptides have
are augmented under conditions of tissue injury such been isolated from bovine brain: endomorphin-1
as inflammation, neuropathy, or bone damage (Stein, and endomorphin-2 (Table 1). Both peptides are
C. et al., 2001; 2003). One underlying mechanism is an considered highly selective -receptor ligands.
increased number (upregulation) of peripheral opioid Their precursors are not known yet. All of these
receptors. In neuronal cell cultures, -receptor tran- opioid peptides have been detected in immune cells
scription is upregulated by the cytokine IL-4 through but the POMC and PENK families have been studied
binding of STAT6 transcription factors to the - most extensively.
receptor gene promoter (Kraus, J. et al., 2001). In
DRG, the synthesis and expression of opioid receptors 30.3.2.2.(i) Proopiomelanocortin-derived opioid
can be increased by peripheral tissue inflammation (Ji, peptides POMC-related opioid peptides have
R. R. et al., 1995; Zollner, C. et al., 2003). Subsequently, been found in immune cells of many vertebrates
the axonal transport of opioid receptors is greatly and nonvertebrates (references in Machelska, H.
enhanced leading to their upregulation and to et al., 2002). To determine whether these immune-
enhanced agonist efficacy at peripheral nerve term- competent cells actually synthesize POMC rather
inals ( Jeanjean, A. P. et al., 1995; Mousa, S. A. et al., than simply absorb related peptides from plasma,

Table 1 Structure of endogenous opioid peptides

-Endorphin Tyr-Gly-Gly-Phe-Met-Thr-Ser-Glu-Lys-Ser-Gln-Thr-Pro-Leu-Val-Thr-Leu-Phe-Lys-
Asn-Ala-Ile-Val-Lys-Asn-Ala-His-Lys-Lys-Gly-Gln-OH
Methionine-enkephalin Tyr-Gly-Gly-Phe-Met-OH
Leucine-enkephalin Tyr-Gly-Gly-Phe-Leu-OH
Dynorphin A Tyr-Gly-Gly-Phe-Leu-Arg-Arg-Ile-Arg-Pro-Lys-Leu-Lys-Tyr-Asp-Asn-Gln-OH
Endomorphin-1 Tyr-Pro-Trp-Phe-OH
Endomorphin-2 Tyr-Pro-Phe-Phe-OH
418 Immune System, Pain and Analgesia

mRNA encoding POMC was sought for and demon- In the pituitary, POMC processing begins as the nas-
strated in many of these studies. The pituitary cent polypeptide enters the endoplasmic reticulum
POMC gene is organized into three exons separated directed by the signal peptide, and POMC cleavage
by intervening sequences which are removed during begins in the trans-Golgi network (references in
processing following transcription to produce the Mousa, S. A. et al., 2004). The POMC prohormone is
full-length, 1200-nt transcript. Initially, truncated directed to the regulated secretory pathway at the
POMC transcripts were found in leukocytes (refe- trans-Golgi network by binding to a sorting receptor,
rences in Machelska, H. et al., 2002). Lyons P. D. and membrane-bound carboxypeptidase E. The prohor-
Blalock J. E. (1997) reexamined the question of mone convertases PC1 (also called PC1/3) and PC2
POMC mRNA expression using novel polymerase cleave POMC within the trans-Golgi network. PC1
chain reaction procedures. With this exacting and mediates the initial cleavage into adrenocorticotropic
sensitive methodology, they found expression of and -lipotropic hormones. The inactive pro-PC2 is
full-length transcripts encoding all three POMC bound to 7B2 (a chaperone-like binding protein) and
exons in rat mononuclear leukocytes. This POMC is transported from the endoplasmic reticulum to later
transcript is spliced in the same way as the pituitary compartments of the secretory pathway, where it
transcript and contains the sequence for the signal matures to active PC2; thereafter, PC2 converts -
peptide, which is necessary for the correct routing lipotropic hormone into -melanocyte stimulating
into the regulated secretory pathway. The POMC hormone and -endorphin (references in Mousa, S.
protein is also proteolytically processed in a way A. et al., 2004). Recently, we detected -endorphin and
consistent with the pituitary gland (Lyons, P. D. and POMC alone and colocalized with PC1, PC2, carbox-
Blalock, J. E., 1997; Mousa, S. A. et al., 2004). These ypeptidase E, and 7B2 in leukocytes in the blood and
results unequivocally demonstrate that immune cells within inflamed paw tissue (Mousa, S. A. et al., 2004).
can produce full-length POMC transcripts. This demonstrates that immune cells express the
Apparently, this production is stimulated by various entire machinery required for POMC processing
immune and inflammatory mediators (Smith, E. M., into functionally active -endorphin.
2003). In this CFA model, mRNAs encoding POMC and
PENK and the corresponding opioid peptides
30.3.2.2.(ii) Proenkephalin-derived opioid -endorphin and Met-enkephalin are abundant in
peptides PENK-derived opioid peptides have cells of inflamed but not in noninflamed tissue
also been detected in human and rodent immune (Przewlocki, R. et al., 1992). Histomorphological pro-
cells. Both the mRNA and methionine (Met)-enke- cedures and flow cytometry have identified
phalin protein were detected. Preproenkephalin the opioid-containing cells as T- and B-lympho-
mRNA was found in T- and B-cells, macrophages, cytes, granulocytes, and monocytes/macrophages
and mast cells. In subpopulations of immune cells this (Przewlocki, R. et al., 1992; Rittner, H. L. et al.,
mRNA was shown to be highly homologous to brain 2001). Also, it was shown that -endorphin is present
PENK mRNA, abundant and apparently translated, in activated/memory T-cells within inflamed tissue
as immunoreactive enkephalin is present and/or (Cabot, P. J. et al., 1997; Mousa, S. A. et al., 2001).
released. The appropriate enzymes necessary for Thus, opioid peptides are processed and present both
posttranslational processing of PENK have also in circulating and inflammatory cells infiltrating
been identified in immune cells (references in injured tissue.
Machelska, H. et al., 2002).
30.3.2.3 Migration of opioid-containing
30.3.2.2.(iii) Immune-derived opioid peptides in immune cells to inflamed tissue
inflammation Immune-derived opioid peptides The recruitment of leukocytes from the circulation
apparently play a substantial role in the modulation into areas of inflammation begins with the attach-
of inflammatory pain (Machelska, H. et al., 2002; Stein, ment of these cells to vascular endothelium, followed
C. et al., 2003). POMC mRNA and -endorphin, as by their transmigration into the tissue. This is
well as Met-enkephalin and dynorphin were found in mediated by specific cell adhesion and chemoattrac-
cells derived from lymph nodes, and in leukocytes in tant/activator molecules. Leukocytes are recruited
the blood and within inflamed tissue in CFA model of from the circulation by a well-orchestrated set of
inflammatory pain (Cabot, P. J. et al., 1997; 2001; events. They undergo multiple attachments to and
Machelska, H. et al., 2003; Mousa, S. A. et al., 2004). detachments from the vessels endothelial cells prior
 Immune System, Pain and Analgesia 419

to transendothelial migration. This includes slowing Recent findings indicate that these events can also
and rolling along the endothelial cell wall that is be involved in the endogenous control of inflamma-
mediated predominantly by the interaction of selec- tory pain. In the model of unilateral hind paw
tins expressed on leukocytes (L-selectin) and on inflammation with CFA we have shown that integrin
endothelial cells (P- and E-selectin) with their 4 and the chemokines CXCL2 and CXCL1 are
ligands on endothelium or immune cells, respec- expressed by leukocytes, while adhesion molecules
tively. The rolling leukocytes can then be activated such as P- and E-selectins, ICAM-1, and PECAM-1
by chemoattractants such as chemokines released are upregulated on endothelium in inflamed paw
from inflammatory cells and presented on endothe- tissue (Mousa, S. A. et al., 2000; Machelska, H. et al.,
lium. This leads to the upregulation and increased 2002; Brack, A. et al., 2004; Machelska, H. et al., 2004)
avidity of integrins. These mediate the firm adhesion (Figure 3). Expression of CXCL1 and CXCL2
of leukocytes to endothelial cells via ligands of mRNAs and protein contents significantly increased
the immunoglobulin superfamily, e.g., intercellular in inflamed tissue during the course of inflammation
adhesion molecule-1 (ICAM-1). Finally, the immune (Brack, A. et al., 2004). Importantly, L-selectin, integ-
cells transmigrate through the endothelial wall rins 2, and the CXC chemokine receptor 2
mediated by immunoglobulin superfamily members (CXCR2) are coexpressed by opioid-containing leu-
(e.g., platelet-endothelial adhesion molecule-1; kocytes, which have migrated to inflamed
PECAM-1) and are directed to the sites of inflamma- subcutaneous paw tissue (Mousa, S. A. et al., 2000;
tion to initiate a host defense (reviewed by Brack, A. et al., 2004; Machelska, H. et al., 2004).
Petruzzelli, L. et al., 1999; von Andrian, U. H. and Furthermore, pretreatment of rats with a selectin
Mackay, C. R., 2000). blocker (fucoidin), selective antibodies against

Endothelium
Vessel lumen

Opioid peptide

Adhesion molecules

CRF DRG
Chemokines CRFR
IL-1
IL-1R PAN

PGSN

Opioid receptor

AR
NA Zeichnung C Christine Voigts-Grafik Charit 5/2005

Figure 3 Migration of opioid-producing cells and opioid secretion within inflamed tissue. Adhesion molecules are
upregulated on vascular endothelium and are coexpressed by circulating immune cells producing opioid peptides. These
cells coexpress receptors for chemokines, which are presented on endothelium. Certain adhesion molecules mediate rolling
of opioid-containing leukocytes along the endothelium. The rolling leukocytes can then be activated by chemokines, which
upregulate another class of adhesion molecules mediating adhesion. Finally cells transmigrate through the endothelium.
Once extravasated, these cells can be stimulated by stress or releasing agents such as corticotropin releasing factor (CRF),
interleukin-1 (IL-1), chemokines, and noradrenaline (NA) to secrete opioid peptides. CRF, IL-1, and NA (the latter derived
from postganglionic sympathetic neurons; PGSN) elicit opioid release by activating CRF receptors (CRFR), IL-1 receptors
(IL-1R), and adrenergic receptors (AR) on immune cells, respectively. Opioids bind to peripheral opioid receptors (produced in
dorsal root ganglia (DRG) and transported to peripheral endings of primary afferent neurons; PAN) and lead to analgesia.
420 Immune System, Pain and Analgesia

ICAM-1, integrins 4 and 2, or chemokines CXCL1 nodes and paw tissue in CFA model. Their
and CXCL2 substantially decreases the number of pharmacological characteristics were similar to the
opioid-containing leukocytes infiltrating the high-affinity CRF- and IL-1-binding sites in the
inflamed tissue (Machelska, H. et al., 1998; pituitary (Mousa, S. A. et al., 1996). Furthermore,
Machelska, H. et al., 2002; Brack, A. et al., 2004; the coexpression of CRF-1 and CRF-2 receptors
Machelska, H. et al., 2004) and in consequence with -endorphin in monocytes/macrophages, gran-
abolishes endogenous peripheral opioid analgesia ulocytes and lymphocytes were recently shown in
(see Endogenous Opoid Analgesia). This suggests the blood and in inflamed paw tissue (Mousa, S. A.
that circulating opioid-producing immune cells et al., 2003). In contrast, CRF receptors were not
home to inflamed tissue where they secrete the detected on peripheral sensory nerves. (Mousa, S. A.
opioids to inhibit pain. Afterward, they travel to the et al., 1996; Mousa, S. A. et al., 2003). The activation of
regional lymph nodes, depleted of the opioid pep- CRF and IL-1 receptors on cells from lymph nodes
tides (Cabot, P. J. et al., 1997; 2001). Thus, local results in the secretion of opioid peptides in vitro. In
signals apparently not only stimulate the synthesis those studies -endorphin, Met-enkephalin, and
of opioid peptides in resident inflammatory cells but dynorphin were dose dependently released by CRF,
also attract opioid-containing cells from the circula- while IL-1 released -endorphin and dynorphin
tion to the site of injury to reduce pain. This is but not Met-enkephalin. These effects were specific
controlled by specific chemotactic and adhesive to CRF and IL-1 receptors (Schafer, M. et al., 1994;
mechanisms. Cabot, P. J. et al., 1997; 2001; Mamet, J. et al., 2002).
Moreover, this release of opioid peptides was
30.3.2.4 Release of opioid peptides from calcium-dependent and mimicked by elevated extra-
immune cells cellular concentrations of potassium. This is
Regulated secretion of peptides requires secretory consistent with a regulated pathway of release from
granules deriving from the Golgi network for secretory vesicles, as in neurons and endocrine cells
transport to the cell membrane. As discussed in (Cabot, P. J. et al., 1997; 2001).
Immune-Derived Opoid Peptides in Inflammation Adrenergic receptor agonists have also been
immune cells in the blood and in inflamed tissue shown to secrete -endorphin from human periph-
coexpress the entire machinery required for POMC eral blood mononuclear cells (Kavelaars, A. et al.,
processing into functionally active -endorphin 1990). Similar mechanisms are involved in -endor-
(Mousa, S. A. et al., 2004). Furthermore, our ultra- phin release from leukocytes in the rat CFA model of
structural observations show that -endorphin- inflammatory pain (Binder, W. et al., 2004) (Figure 3).
expressing macrophages, monocytes, granulocytes, Double labeling demonstrated adrenergic 1-, 2-,
and lymphocytes, contain rough endoplasmic reticu- and, to a lesser degree, 2 receptors expressed on
lum and Golgi apparatus, similar to pituitary cells. -endorphin-containing leukocytes in inflamed
Immunostaining of -endorphin was detectable in paw tissue. -endorphin-containing and adrenergic
secretory granules, which were grouped in small or 1- and  2-receptor-expressing immune cells were
large membranous vesicular structures. The smaller localized in close proximity to sympathetic nerve
-endorphin-immunoreactive secretory granules fibers, and chemical ablation of these fibers abolished
were localized within cytoplasm, and the larger intrinsic opioid analgesia. Finally, noradrenaline
ones were arranged at the cell periphery ready for induced adrenergic receptor-specific release of -
the exocytosis, similar to the pituitary (Mousa, S. A. endorphin from immune cells in vitro (Binder, W.
et al., 2004). In the pituitary, -endorphin and other et al., 2004). Taken together, CRF, IL-1, and sym-
POMC-derived peptides are released by corticotro- pathetic neuron-derived noradrenaline can act on
pin releasing factor (CRF) and IL-1 (references in their respective receptors on immune cells resulting
Schafer, M., 2003). Similar mechanisms can trigger in release of opioid peptides (Figure 3).
opioid release within peripheral inflamed tissue
(Figure 3). CRF is present in immune cells, fibro- 30.3.2.5 Analgesia produced by immune-
blasts and vascular endothelium and peripheral CRF derived opioid peptides
expression is enhanced in inflamed synovial and sub- 30.3.2.5.(i) Analgesic effects of corticotropin
cutaneous tissue of animals and humans (Schafer, M. releasing factor, interleukin-1, and noradrenaline
et al., 1996). CRF and IL-1 receptors and their CRF-, IL-1-, and noradrenaline-induced release of
upregulation were demonstrated in inflamed lymph opioids from immune cells also occurs in vivo
 Immune System, Pain and Analgesia 421

(Figure 3). CRF and IL-1 injected into inflamed of CFA inflammation (Woolf, C. J. et al., 1997) or
paws in CFA model produce dose-dependent analge- in short-lasting (30 min to 3 h) inflammation, e.g.,
sia reversible by their respective antagonists (Schafer, induced with carrageenan, lipopolysaccharide, or
M. et al., 1994). CRF can produce analgesia both in acetic acid (Cunha, F. Q. et al., 1992; Ferreira, S. H.
early and late stages of inflammation (2 h and 6 days) et al., 1993; Cunha, J. M. et al., 2000; Cunha, T. M.
and, in accord with anatomical findings (see Release et al., 2005). Thus, the presence or absence of inflam-
of Opoid Peptides from Immune Cells), CRF analge- mation, the duration and/or model of inflammatory
sia involves both CRF-1 and CRF-2 receptors pain are factors to be taken into consideration.
(Machelska, H. et al., 2003; Mousa, S. A. et al., 2003; Noradrenaline administered directly into
Brack, A. et al., 2004c). Intravenous administration of inflamed tissue has been shown to produce analgesia,
these agents in locally effective doses does not reversible by 1-, 2-, and 2-adrenergic receptor
change pain thresholds, demonstrating a peripheral antagonists in CFA model. Further, this effect was
site of action (Schafer, M. et al., 1994; Machelska, H. dose dependently blocked by  and  receptor
et al., 2003; Mousa, S. A. et al., 2003; Brack, A. et al., antagonists and antibody against -endorphin
2004c). Leukocytes apparently are the target for CRF (Binder, W. et al., 2004). These data suggest that this
and IL-1 because immunosuppression with cyclos- catecholamine produces analgesia via opioid pep-
porine A, depletion of granulocytes, blockade of tides activating peripheral opioid receptors. In
chemokines (CXCL1 and CXCL2/3), as well as anti- noninflamed tissue, noradrenaline did not influence
selectin and anti-ICAM-1 treatments result in a pain behavior in this model, consistent with the lack
significant reduction of opioid-containing immune of opioid-containing cells and with the scarcity of
cells and of CRF- or IL-1-induced analgesia adrenergic receptors (Binder, W. et al., 2004).
(Schafer, M. et al., 1994; Machelska, H. et al., 1998; However, the role of peripheral adrenergic receptors
2002; Brack, A. et al., 2004c). Also, cyclosporine A- in nociception appears controversial. In noninflamed
induced attenuation of CRF analgesia could be tissue, noradrenaline has been shown to produce
restored by injection of activated lymphocytes hyperalgesia via 2-adrenergic receptors. Others
(Hermanussen, S. et al., 2004). CRF- and IL-1- found that 2-agonists could also produce peripheral
induced analgesia is blocked by an antibody against analgesia. It has even been postulated that hyperal-
-endorphin, suggesting that this opioid plays a gesia is mediated via 2B and analgesia is mediated
major role. In addition, Met-enkephalin appears to via 2C receptors (references in Binder, W. et al.,
be involved in CRF-, and dynorphin in IL-1- 2004). Thus, different receptor subtypes, receptor
induced analgesia (Schafer, M. et al., 1994). localization, microenvironment, and the presence or
These results are in line with other studies on absence of inflammation are important parameters to
local analgesic effects of CRF (Hargreaves, K. M. be considered.
et al., 1989; McLoon, L. K. et al., 2002) and of the
cytokines IL-6 and TNF- (Czlonkowski, A. et al., 30.3.2.5.(ii) Endogenous opioid analgesia Stress
1993) but are in contrast to hyperalgesia mediated by is a natural stimulus triggering inhibition of pain
IL-1, IL-6, and TNF- discussed in Cunha F. Q. (Willer, J. C. et al., 1981; Terman, G. W. et al., 1984).
and Ferreira S. H. (2003). Importantly, however, the In rats, stress induced by cold water (4  C) swim (for
latter hyperalgesic effects of exogenous cytokines 1 min) elicits potent analgesia in inflamed but not in
were observed after injections into noninflamed tis- noninflamed paws in CFA model (Stein, C. et al., 1990a;
sue (Cunha, F. Q. et al., 1992; Poole, S. et al., 1995; Machelska, H. et al., 2003). In early inflammation (6 h),
Safieh-Garabedian, B. et al., 1995; Woolf, C. J. et al., this swim stress-induced analgesia was only partially
1997; Cunha, T. M. et al., 2005). Apparently, hyper- attenuated by peripherally selective doses of different
algesia in highly inflamed tissue (4 days after CFA) opioid peptide antibodies and receptor antagonists but
has already reached a ceiling effect (Machelska, H. was fully reversed by centrally acting doses of an
et al., 2003) and therefore cannot be further increased. opioid receptor antagonist (Machelska, H. et al., 2003).
Instead, in such tissue opioid-containing immune At later stages of inflammation (46 days), swim stress
cells became the predominant target for cytokines analgesia was completely abolished by peripherally
to produce analgesia. Contribution of endogenous selective doses of antibody against -endorphin and
proinflammatory cytokines to the generation of pain by  and -antagonists (Stein, C. et al., 1990a).
was mostly observed either in early stages (36 h) Together, these data indicate that at early stages of
when cytokine blockade preceded induction CFA-induced inflammation all three families of opioid
422 Immune System, Pain and Analgesia

peptides and receptors are involved, while at later (Mousa, S. A. et al., 2001; Rittner, H. L. et al., 2001;
stages -endorphin acting at - and -receptors dom- Zollner, C. et al., 2003).
inates. Whereas at early stages both peripheral and
central opioid receptors are involved, at later stages 30.3.2.6 Clinical implications
endogenous analgesia is mediated exclusively by per- Peripheral endogenous opioid analgesia has found
ipheral opioid receptors. Thus, peripheral opioid many clinical applications. Opioid receptors have
mechanisms of pain control become more prevalent been demonstrated on peripheral terminals of sen-
with the duration and severity of inflammation. sory nerves in human synovia (Stein, C. et al., 1996)
Endogenous triggers of swim stress-induced analgesia and these receptors mediate analgesia in patients
are locally produced CRF and sympathetic nerve- with various types of pain (e.g., in chronic rheuma-
derived catecholamines because this effect is abolished toid arthritis and osteoarthritis, bone pain, after
by local neutralization of CRF (Schafer, M. et al., 1996; dental, laparoscopic, urinary bladder, and knee
Machelska, H. et al., 2003), and by sympathetic block- surgery) (references in Stein, C. et al., 2003). Opioid
ade (Binder, W. et al., 2004). peptides are found in human synovial lining cells,
Various types of immune cells are the source of mast cells, lymphocytes, and macrophages. The pre-
opioids as demonstrated by the abolishment of stress- vailing peptides are -endorphin and Met-
induced analgesia by immunosuppression with enkephalin, while only minor amounts of dynorphin
cyclosporine A or whole-body irradiation and by are detectable (Stein, C. et al., 1993; 1996). The inter-
depletion of monocytes/macrophages (Stein, C. action of synovial opioids with peripheral opioid
et al., 1990b; Przewlocki, R. et al., 1992; Brack, A. receptors was examined in patients undergoing
et al., 2004a). Moreover, this effect is also extin- knee surgery. Blocking intraarticular opioid recep-
guished by inhibiting the extravasation of - tors by the local administration of naloxone resulted
endorphin-containing immune cells following block- in significantly increased postoperative pain (Stein,
C. et al., 1993). Taken together, these findings suggest
ade of L- and P-selectins, 4 and 2 integrins or of
that in a stressful (e.g., postoperative) situation,
ICAM-1 (Machelska, H. et al., 1998; 2002; 2004).
opioids are tonically released from inflamed tissue
These adhesion molecules apparently regulate the
and activate peripheral opioid receptors to attenuate
migration of opioid-containing immune cells and
clinical pain. Importantly, these endogenous opioids
the subsequent generation of intrinsic pain control
do not interfere with exogenous morphine, i.e.,
in inflamed tissue (Figure 3).
intraarticular morphine is an equally potent analgesic
A future challenge is to identify factors that will
in patients with and without opioid-producing
increase homing of opioid-containing cells to injured
inflammatory synovial cells (Stein, C. et al., 1996).
tissue and will enhance analgesia. To this end, we
This suggests that, in contrast to the rapid develop-
have shown that hematopoetic growth factors ment of tolerance in the central nervous system, the
strongly mobilized granulocytes in the blood but immune cell-derived opioids do not readily produce
resulted only in a minor increase in the number of cross-tolerance to morphine at peripheral opioid
opioid-containing leukocytes in inflamed paws and receptors.
in no change of CRF- and swim stress-induced
analgesia (Brack, A. et al., 2004b). Another approach 30.3.2.7 Perspectives
was to increase the migration of opioid-containing Effective control of inflammatory pain can result from
cells to inflamed tissue with local injections of interactions between leukocyte-derived opioid pep-
CXCL2. However, this did not result in stronger tides and their receptors on peripheral sensory
CRF- or swim stress-induced analgesia either, most neurons. These findings provide new insights into
probably as a result of the small number of peripheral intrinsic mechanisms of pain control and open strate-
opioid receptors at this very early (2 h) stage of gies to develop new drugs and alternative approaches
inflammation (Brack, A. et al., 2004d). Indeed, pre- to treatment of pain. Immunocompromised patients
vious studies had shown that intrinsic analgesia (e.g., in AIDS, cancer, diabetes) frequently suffer from
increases with the duration of inflammation (2 h to painful neuropathies, which can be associated with
4 days), in parallel with the number of opioid-con- intra- and perineural inflammation, with reduced
taining leukocytes, with the number of peripheral intraepidermal nerve fiber density and with low
opioid receptors and with the efficacy of opioid CD4 lymphocyte counts (Polydefkis, M. et al., 2002).
receptorG protein coupling in sensory neurons Thus it may be interesting to investigate the opioid
 Immune System, Pain and Analgesia 423

production/release and the migration of opioid-con- Akil, H., Owens, C., Gutstein, H., Taylor, L., Curran, E., and
Watson, S. 1998. Endogenous opioids: overview and current
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modulation by opioids and cyclic AMP. Neuroscience
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may in fact carry a significant risk to exacerbate pain. It migration. Two sides of the same coin. N. Engl. J. Med.
343, 10201034.
would be highly desirable to identify stimulating fac- Arvidsson, U., Riedl, M., Chakrabarti, S., Lee, J. H.,
tors and strategies that selectively attract opioid- Nakano, A. H., Dado, R. J., Loh, H. H., Law, P. Y.,
producing cells and increase peripheral opioid receptor Wessendorf, M. W., and Elde, R. 1995. Distribution and
targeting of a mu-opioid receptor (MOR1) in brain and spinal
numbers in damaged tissue. Augmenting the synthesis cord. J. Neurosci. 15, 33283341.
and/or secretion of opioid peptides and opioid Beland, B. and Fitzgerald, M. 2001. Mu- and delta-opioid
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Schafer, M. 2004. Sympathetic activation triggers
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Many efforts are currently undertaken to develop versus opioid-mediated peripheral antinociception.
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Brack, A., Rittner, H. L., Machelska, H., Beschmann, K.,
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Stein, C. 2004d. Endogenous peripheral antinociception in
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Further Reading
Zhang, N., Inan, S., Cowan, A., Sun, R., Wang, J. M.,
Rogers, T. J., Caterina, M., and Oppenheim, J. J. 2005. A Machelska, H. and Stein, C. 2002. Immune mechanisms in pain
proinflammatory chemokine, CCL3, sensitizes the heat- and control. Anesth. Analg. 95, 10021008.
capsaicin-gated ion channel TRPV1. Proc. Natl. Acad. Sci.
U. S. A. 102, 45364541.
Zollner, C., Shaqura, M. A., Bopaiah, C. P., Mousa, S. A.,
Stein, C., and Schafer, M. 2003. Painful inflammation induced Relevant Website
increase in  opioid receptor binding and G-protein coupling in
primary afferent neurons. Mol. Pharm. 64, 202210.
Zwick, M., Molliver, D. C., Lindsay, J., Fairbanks, C. A., http://www.iasp.pain.org International Association for
Sengoku, T., Albers, K. M., and Davis, B. M. 2003. the Study of Pain.
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 31 Mechanisms of Glial Activation after Nerve Injury
L R Watkins, E D Milligan, and S F Maier, University of Colorado at Boulder, Boulder, CO, USA
2009 Elsevier Inc. All rights reserved.

31.1 Introduction 429

31.2 Glial Activation in Response to Neurotransmitters 429
31.3 Glial Activation in Response to Neuromodulators 430
31.4 Glial Activation by Unique Neuron-to-Glia Signals 430
31.5 Beyond Neuropathic Pain: Glial Activation in Response to Opioids 431
31.6 Conclusions 432
References 432

31.1 Introduction (Svensson, C. I. et al., 2004). Upon activation, neu-

roexcitatory products that glia release are
It has been known since the early 1970s that periph- importantly involved in the initiation and mainte-
eral nerve injury leads to the activation of glia in nance of neuropathic pain, via their actions on
anatomically linked regions of the brain and spinal neurons of the pain pathway (Watkins, L. R. et al.,
cord (Aldskogius, H. and Kozlova, E. N., 1998). In 2001; Watkins, L. R. and Maier, S. F., 2003). Thus,
these studies, glial activation was inferred by upre- converging lines of evidence utilizing a variety of
gulation of the so-called glial activation markers. techniques all support the conclusion that glia are
These signs that glial activation has occurred can be key players in neuropathic pain.
visualized under the microscope using immunohisto- However, what is known to date regarding glial
chemistry to detect cell-type specific changes (e.g., involvement in neuropathic pain begs a fundamen-
increased expression of glial fibrillary acidic protein tally important question: how do glia know to
by astrocytes; increased expression of complement become activated? That is, what message(s) do glia
type 2 receptors by microglia). Such studies revealed receive that trigger their transition from basal to
that damage of sensory afferent nerves activates glia activated states? This will be discussed first in the
in the region of the afferent terminations in the context of glial activation in response to injury or
central nervous system (CNS). Likewise, damage of inflammation of peripheral nerves, and then more
motor efferents activates glia in the CNS in the broadly in the context of glial activation in response
region of the involved motor neuron cell bodies to analgesics used clinically to treat chronic pain,
(Aldskogius, H. and Kozlova, E. N., 1998). While such as morphine and methadone.
this early literature documented the phenomenon of
glial activation in CNS following peripheral nerve
injury, the mechanisms underlying such activation 31.2 Glial Activation in Response to
remained obscure. Neurotransmitters
Two decades later, glial activation was linked to
neuropathic pain. This link was again initially based Given that (1) glia become activated in response to
on the upregulation of glial activation markers inflammation and damage of peripheral tissues and
(Garrison, C. J. et al., 1991; 1994). Pharmacology, peripheral nerves and (2) this activation occurs within
anatomy, and molecular biology studies that followed the somatotopically appropriate region of spinal cord,
documented that microglia and then astrocytes are this suggests that glia may be activated by neurotrans-
sequentially activated in response to inflammation mitters released by nocisponsive primary afferents.
and damage to peripheral nerves (Raghavendra, V. Indeed, calcitonin gene-related peptide (CGRP)
et al., 2003; Ledeboer, A. et al., 2005). Indeed, studies (Haas, C. A. et al., 1991; Reddington, M. et al., 1995),
of p38 mitogen-activated protein (MAP) kinase acti- adenosine triphosphate (ATP) (Araque, A. et al., 1999;
vation demonstrated that microglial activation occurs Tsuda, M. et al., 2003), glutamate (Araque, A. et al.,
with remarkable rapidity following nerve injury 1999; Tsuda, M. et al., 2003), and substance P

429
430 Mechanisms of Glial Activation after Nerve Injury

(Svensson, C. I. et al., 2003; Marriott, I., 2004) can each not required for glial activation to take place. This
activate microglia and astrocytes. Such neurotransmit- predicts that other signals, beyond neurotransmitters,
ter-driven glial activation can be relevant for pain must exist which can trigger the activation of glia.
facilitation. For example, pain facilitation that is nor-
mally produced upon intrathecal administration of the
excitatory amino acid, N-methyl-D-aspartate 31.3 Glial Activation in Response
(NMDA), is abolished by blocking microglial activa- to Neuromodulators
tion (Hua, X.-Y. et al., 2005).
Regarding substance P, studies of spinal cord glia Neuromodulators, such as prostaglandins and nitric
have demonstrated that it can activate both astrocytes oxide (NO) are well documented to enhance pain
and microglia. Astrocytes isolated from spinal cord (Woolf, C. J. and Salter, M. W., 2000). In addition to
(but not from other brain regions) release prostaglan- direct excitatory effects on neurons, these substances
dins upon exposure to substance P (Marriott, D. R. can also activate glia. Using NO as the example, NO
et al., 1991). Substance P can also stimulate the pro- induces calcium oscillations in astrocytes, a marker of
duction of proinflammatory cytokines in both activation in this cell type. In turn, NO-induced cal-
astrocytes (Martin, F. C. et al., 1992; Palma, C. et al., cium oscillations causes a nondecrementing, rapid
1997; Lieb, K. et al., 1998) and microglia (Martin, F. C. spread of excitation to surrounding astrocytes through
et al., 1993; Luber-Narod, J. et al., 1994). When sub- cell-to-cell low-resistance connections called gap
stance P is injected intrathecally in rats, this activates junctions (Heidemann, A. C. et al., 2004; Sul, J. et al.,
spinal cord microglia as reflected by phosphorylation 2004). Thus NO-excited astrocytes lead to the excita-
(activation) of microglial p38 MAP kinase, a signaling tion of a much larger population of astrocytes. Once
pathway linked to pain facilitation (Svensson, C. I. activated, these glia can then begin releasing neuroex-
et al., 2004). Interestingly, microglia can also produce citatory substances (Araque, A. et al., 1999). In addition,
substance P, suggesting that these glial cells may NO induces the release of glutamate, ATP, and pros-
potentially contribute to substance P-mediated pain taglandins from astrocytes (Molina-Holgado, F. et al.,
signaling (Lai, J. P. et al., 2002). 1995; Bal-Price, A. et al., 2002) and stimulates the
ATP has also recently attracted attention as a neu- production of proinflammatory cytokines in both
rotransmitter that can enhance pain via the activation microglia and astrocytes (Sun, D. et al., 1998).
of glia. Microglia (but not neurons or astrocytes) Regarding actions in spinal cord, NO has recently
express an ionotropic ATP receptor subtype called been demonstrated to drive the production and release
P2X4 whose expression can become dramatically of proinflammatory cytokines in response to intrathe-
upregulated in response to peripheral nerve injury cally administered gp120, a protein expressed on the
(Tsuda, M. et al., 2003). Pharmacological blockade of surface of the acquired immune deficiency syndrome
spinal P2X4 receptors and knockdown of P2X4 recep- (AIDS) virus that activates glia (Holguin, A. et al.,
tors using antisense oligodeoxynucleotides each 2004). As gp120-induced proinflammatory cytokines
reverse neuropathy-induced tactile allodynia (Tsuda, induce mechanical allodynia, it would be predicted
M. et al., 2003). Intriguingly, activation of microglia by that NO synthesis inhibitors would block such pain
ATP is sufficient to cause pain facilitation. That is, changes concomitant with blocking NO-induced pro-
microglia that are first stimulated in vitro by ATP, and duction of these cytokines. Indeed, that was the result
then injected intrathecally in rats, induces mechanical found (Holguin, A. et al., 2004).
allodynia (Tsuda, M. et al., 2003).
While, at first glance, it would seem logical that glia
must become activated in response to neurotransmit- 31.4 Glial Activation by Unique
ters released as a consequence of peripheral nerve Neuron-to-Glia Signals
injury, one must remember the literature of the
1970s. That is, if one damages a pure motor nerve, Beyond neurotransmitters and neuromodulators
glia are activated surrounding the motor neuron cell classically known for their pain-enhancing effects,
bodies within the CNS. In this case, there is no damage glia can also be activated by substances novel to the
to sensory afferent fibers. So, despite there being no pain field. This class includes fractalkine and sub-
neurotransmitter release, glia are still being intensely stances released by damaged cells. Regarding
activated. Thus, while glia do express receptors for fractalkine, it is a protein expressed on the extracel-
neurotransmitters and can respond to them, they are lular surface of both sensory afferent neurons and
 Mechanisms of Glial Activation after Nerve Injury 431

neurons intrinsic to the spinal cord (Verge, G. M. oligodeoxynucleotide to reduce the expression of
et al., 2004; Lindia, J. A. et al., 2005). Fractalkine is spinal TLR4. This treatment also decreased expres-
tethered to the neurons in such a way as to be sion of spinal microglial markers and proinflammatory
enzymatically released under strong neuroexcitatory cytokines, compared to controls (Tanga, F. Y. et al.,
conditions. Upon release, fractalkine acts as a diffu- 2005). While TLR4 is best known as the receptor that
sible signaling molecule (Chapman, G. et al., 2000). recognizes bacterial endotoxin (lipopolysaccharide), it
Intriguingly, under basal conditions, the receptor for also binds endogenous substances associated with cell
fractalkine is expressed predominantly by spinal damage. These include members of the heat shock
microglia (Verge, G. M. et al., 2004; Lindia, J. A. protein family and proteoglycans.
et al., 2005). Thus, upon insult to the body, released The potential link to heat shock proteins is espe-
fractalkine serves as a neuron-to-microglial signal. In cially intriguing. Heat shock proteins are classically
response to peripheral nerve inflammation and thought of as intracellular proteins that aid in cell
injury, there is an increase in fractalkine receptor survival after stress or trauma (Costigan, M. et al.,
expression by microglia in the dorsal horn and micro- 1998). However, heat shock proteins are also released
glia remain the sole apparent source of fractalkine by damaged and dying cells and such extracellular heat
receptor expression (Verge, G. M. et al., 2004; Lindia, shock proteins can activate glia (Kakimura, J. et al.,
J. A. et al., 2005). Under some (but not all) neuropathic 2002; Takata, K. et al., 2003). Indeed, heat shock pro-
conditions, astrocytes may begin producing fractalk- teins are known to be upregulated for months in the
ine, suggestive that astrocytes also stimulate spinal processes of sensory neurons whose peripheral
microglia via the release of fractalkine, so to maintain axons are damaged (Costigan, M. et al., 1998).
microglial activation under such conditions (Verge, Preliminary data suggest that heat shock proteins can
G. M. et al., 2004; Lindia, J. A. et al., 2005). synergize with other glial excitatory stimuli to enhance
Regarding the role of fractalkine in pain facilita- the production of proinflammatory cytokines by dorsal
tion, what is known to date is that fractalkine spinal cord microglia (Wieseler-Frank, J. et al., 2004).
upregulates P2X4 receptors in dorsal spinal cord
microglia (Wieseler-Frank, J. et al., 2004). As
reviewed above, this microglia-specific ATP recep- 31.5 Beyond Neuropathic Pain: Glial
tor is implicated in the induction of mechanical Activation in Response to Opioids
allodynia (Tsuda, M. et al., 2003). When fractalkine
is injected intrathecally, it induces both mechanical While the focus to this point has been on endogenous
allodynia and thermal hyperalgesia, via the release of signals that activate glia, it is also noteworthy that glia
proinflammatory cytokines (Johnston, I. N. et al., are activated by opioids, including morphine (Song, P.
2004; Milligan, E. D. et al., 2004; 2005). Importantly, and Zhao, Z. Q., 2001; Raghavendra, V. et al., 2002;
endogenous fractalkine enhances pain. That is, Johnston, I. N. et al., 2004; Shavit, Y. et al., 2004) and
blocking fractalkine actions both delays and reverses methadone (Hutchinson, M. R. et al., 2005). The results
neuropathic pain (Milligan, E. D. et al., 2004). This for morphine that have been reported across labs to
suggests that peripheral nerve injury causes perse- date are remarkably consistent in demonstrating that
verative release of fractalkine and this release spinal cord glia become progressively more activated
contributes to the induction and maintenance of by morphine upon repeated administration and that
neuropathic pain. morphine-induced glial activation leads to the pro-
Lastly, glia can be activated by substances that duction and release of proinflammatory cytokines.
signal neuronal damage. Microglia, like other immune This opioid-induced proinflammatory cytokine
cells, express evolutionarily conserved receptors for release has been linked, using diverse methodologies,
detecting cell damage, as well as for detecting patho- to an attenuation of acute morphine analgesia, to
gens such as bacteria and viruses. The genes for these the development of morphine tolerance, and to the
receptors are homologous to the Toll gene in expression of withdrawal-induced thermal hyperalge-
Drosophila and have therefore come to be referred to sia and mechanical allodynia (for a review, see
as Toll-like receptors (TLRs). One TLR subtype, Watkins, L. R. et al., 2005). What is important about
TLR4, has been clearly implicated in pain facilitation this line of research is that it documents that glia are
(Tanga, F. Y. et al., 2005). TLR4 knockout mice show not only activated in response to peripheral injury and
reduced neuropathic pain in response to L5 nerve inflammation, but also by first-line opioids used for
transection, as do rats treated with TLR4 antisense clinical pain control. Thus, glial activation becomes a
432 Mechanisms of Glial Activation after Nerve Injury

double-edged sword, occurring both in response to c-fos gene expression in rat astrocyte cultures. Eur.
J. Neurosci. 3, 708712.
situations that induce pathological pain and in Heidemann, A. C., Schipke, C. G., Peters, O., and
response to drugs intended to treat it. Kettenmann, H. 2004. Nitric oxide triggers repetitive calcium
transients in astrocytes in mouse cortical slices. Proc. Soc.
Neurosci. 34, 405407.
Holguin, A., Biedenkapp, J., Campisi, J., Wieseler-Frank, J.,
31.6 Conclusions OConnor, K. A., Milligan, E. D., Maksimova, E.,
Bravmann, C., Hansen, M. K., Martin, D., Fleshner, M.,
Maier, S. F., and Watkins, L. R. 2004. HIV-1 gp120 stimulates
It is clear from the studies reviewed above that spinal proinflammatory cytokine-mediated pathological pain via
cord glia can become activated in response to a wide activation of nitric oxide synthase-I (nNOS). Pain
array of chemical signals. Some are neurotransmitters 110, 517530.
Hua, X.-Y., Svensson, C. I., Matsui, T., Fitzsimmons, B.,
and neuromodulators naturally released in spinal Yaksh, T. L., and Webb, M. 2005. Intrathecal minocycline
cord dorsal horn in response to inflammation and attenuates peripheral inflammation-induced hyperalesia by
damage in the body, others are unique neuron-to- inhibiting p38 MAPK in spinal microglia. Eur. J. Neurosci.
22, 24312440.
glia signals released by activated and damaged neu- Hutchinson, M. R., Milligan, E. D., Maier, S. F., and
rons, and surprisingly some are drugs commonly Watkins, L. R. 2005. Interleukin-1 receptor antagonist (IL1ra)
used for clinical pain control. Regardless of what unmasks analgesia following both R- & S-methadone:
evidence for induction of spinal proinflammatory cytokines
the activating stimulus is, the end result is strikingly (PICs) via non-classical opioid receptors. Proc. Soc.
similar. That is, activated glia begin producing and Neurosci. abstract no. 49.11.
releasing an array of neuroexcitatory substances, Johnston, I. N., Milligan, E. D., Wieseler-Frank, J., Frank, M. G.,
Zapata, V., Campisi, J., Langer, S., Martin, D., Green, P.,
including proinflammatory cytokines. Taken Fleshner, M., Leinwand, L., Maier, S. F., and Watkins, L. R.
together, these studies clearly predict that targeting 2004. A role for proinflammatory cytokines and fractalkine in
spinal cord glia and glial proinflammatory products analgesia, tolerance, and subsequent pain facilitation
induced by chronic intrathecal morphine. J. Neurosci.
should suppress pain facilitation induced by inflam- 24, 73537365.
mation and damage to peripheral tissues and enhance Kakimura, J., Kitamura, Y., Takata, K., Umeki, M., Suzuki, S.,
the clinical efficacy of opioids. Shibagaki, K., Taniguchi, T., Nomura, Y., Gebicke-
Haerter, P. J., Smith, M. A., Perry, G., and Shimohama, S.
2002. Microglial activation and amyloid-beta clearance
induced by exogenous heat-shock proteins. FASEB J.
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 32 Trigeminal Mechanisms of Nociception: Peripheral
and Brainstem Organization
D A Bereiter, University of Minnesota, Minneapolis, MN, USA
K M Hargreaves, University of Texas Health Science Center, San Antonio, TX, USA
J W Hu, University of Toronto, Toronto, ON, Canada
2009 Elsevier Inc. All rights reserved.

32.1 Introduction 435

32.2 The Peripheral Trigeminal Nerve System 438
32.2.1 General Features 438
32.2.2 Neurochemical and Molecular Properties of Trigeminal Ganglion Neurons 439
32.3 Central Aspects of Trigeminal Organization 441
32.3.1 Somatotopy 442
32.3.2 Intersubnuclear Connections 443
32.3.3 Relationship to the Autonomic Nervous System 443
32.3.4 Neurochemical Markers 444
32.3.5 Efferent Projections 446
32.4 Functional Considerations 447
32.4.1 Ocular Pain Processing 448
32.5 Chronic Craniofacial Pain 448
References 450

32.1 Introduction mammals (see Bryant, B. and Silver, W. L., 2000). The
Vn also supplies all specialized receptor regions of the
At first glance the trigeminal nerve (Vn), the largest head including the fungiform taste papillas
cranial nerve, serves many of the same functions as (Whitehead, M. C. et al., 1999), cochlea (Vass, Z.
spinal nerves for lower parts of the body by relaying et al., 1997) and the eye (Belmonte, C. et al., 1997). In
innocuous and noxious mechanical, thermal and che- some mammals and lower vertebrates the Vn relays
mical sensory information from all tissues of the head sensory information unrelated to nociception from
and oral cavity to the brain (Figure 1). Although there unique peripheral structures over distinct central
are many similarities, peripheral and central aspects of pathways to meet environmental demands. For exam-
the Vn system are organized quite differently from the ple, Vn afferents that supply the pit organ of crotaline
spinal system (see Table 1). A primary function of the snakes, a highly sensitive thermal receptor with broad
Vn system is to detect damaging or potentially dama- spectral tuning that complements visual input, do not
ging environmental challenges to the head and oral respond to capsaicin (Moiseenkova, V. et al., 2003) and
cavity and one obvious distinction from spinal systems project to separate regions of the trigeminal brainstem
is the diversity of specialized craniofacial tissues complex (Molenaar, G. J., 1978). The skin of the bill of
innervated by the Vn that provide such detection. the platypus contains some 30 00040 000 mucous
Tissues such as the dental pulp, cornea, and dura sensory glands innervated by small myelinated
lack appreciable innervation by large diameter affer- trigeminal afferent fibers that detect electromagnetic
ent fibers and generally evoke only pain sensation to radiation emitted by prey and used for underwater
natural stimuli (Feindel, W. et al., 1960; Kenshalo, D. R., navigation (Proske, U. et al., 1998). The whisking of
1960; Beuerman, R. W. and Tanelian, D. L., 1979; mystacial vibrissas on the face of rodents provides
Trowbridge, H. O. et al., 1980). Sensory afferents that essential textural and perceptual information for
innervate mucosal tissues of the eye, nose, and oral exploration and food-gathering in a frequency-depen-
cavity have a high sensitivity to airborne chemicals dent manner (Petersen, C. C., 2003; Moore, C. I.,
(Cometto-Muniz, J. E. and Cain, W. S., 1995) and are 2004), whereas the trigeminal innervation of the
thought to underlie the common chemical sense in snout of the coati mundi (Barker, D. J. and Welker,

435
436 Trigeminal Mechanisms of Nociception: Peripheral and Brainstem Organization

Meninges Vg

Cornea
Vp

Vo
Vtr
Nasal/oral Vi
mucosa

Vc

Dental
pulp

TMJ TSNC

Figure 1 Unique craniofacial tissues supplied by the Trigeminal Nerve.

W. I., 1969) and Eimers organ of the appendages of is organized in unusual patterns (e.g., teeth) and not
the star-nosed mole (Catania, K. C., 1999) relay highly always proportional to innervation density (e.g., ocular
refined tactile information and appear more analogous surface). In lower mammals, peripheral and central
to somatosensory receptors of the hands than face of specializations can lead to remarkable magnification
primates. Thus, a significant feature of the peripheral of the cortical representation of orofacial tissues such
Vn is its association with highly specialized sensory as the nasal appendages of the star-nosed mole which
functions of craniofacial and oral tissues. accounts for over 50% of the area of primary somato-
The organization of the central Vn system also dis- sensory cortex (SmI; Catania, K. C. and Kaas, J. H.,
plays unusual features distinct from spinal systems that 1997). Similarly, the representation of the enlarged
may be relevant to pain processing. At the level of the incisors of the naked mole rat used for tunneling and
initial synapse, the trigeminal sensory brainstem underground navigation accounts for over 30% of SmI
nuclear complex (TSNC), craniofacial tissues are (Catania, K. C. and Remple, M. S., 2002). In the albino
represented somatotopically at multiple, and in some rat the face occupies more than 60% of SmI with a
cases discontinuous levels, while spinal nerves project large area devoted to the cortical barrels and mystacial
to a few dominant contiguous segments (see Renehan, vibrissae (Welker, C., 1971), whereas a significant por-
W. E. and Jacquin, M. F., 1993; Bereiter, D. A. et al., tion of the lateral somatosensory cortex is devoted to
2000; Sessle, B. J., 2000). The importance of multiple the teeth (Remple, M. S. et al., 2003). Cortical repre-
representation remains uncertain; however, based on sentation of craniofacial tissues in primates is much
encoding properties and efferent projections, this fea- reduced compared to lower mammals; however, in
ture may reflect the segregation of neuronal select regions such as area 3b, the representation of
populations that serve different aspects of trigeminal the face and oral cavity remains unexpectedly large,
function. Although the representation of craniofacial though curiously, the ocular surface with its high inner-
tissues accounts for a relatively large area of sensory vation density is represented only weakly (Manger, P.
thalamus and somatosensory cortex, a feature that has R. et al., 1995; Jain, N. et al., 2001). Beyond the biological
long been appreciated since the classic studies of importance of protecting the head and oral cavity from
Penfield W. and Rasmussen T. (1950), the cortical damaging stimuli, the Vn is critical for preserving nor-
representation of some specialized craniofacial tissues mal eating and sleeping habits, while the emotional
 Trigeminal Mechanisms of Nociception: Peripheral and Brainstem Organization 437

Table 1 General distinctive features of the trigeminal sensory system

Factor Details References

Peripheral trigeminal system

Sensory innervation Cavernous sinus, cornea/conjunctiva, dental Rozsa A. J. and Beuerman R. W. (1982), Anggard
of specialized pulp, lacrimal gland, nasal mucosa, pineal A. et al. (1983), Byers M. R. (1984), Kido M. A.
tissues gland, salivary glands, temporomandibular et al. (1995), Bleys R. L. et al. (1996), Reuss S.
joint, tongue, vibrissae (1999), Cheng S. B. et al. (2000), Muller L. J.
et al. (2003), Kawabata A. et al. (2004)
Somatotopy within TG neurons from major Vn branches Kerr F. W. L. (1963), Marfurt C. F. (1981),
TG (ophthalmic, maxillary and mandibular) Arvidsson J. et al. (1992), Aigner M. et al.
distributed from anteromedial to (2000)
posterolateral, respectively
Mesencephalic Vmes within the CNS; jaw muscle spindle and Cody F. W. et al. (1972), Linden R. W. (1978)
nucleus of V periodontal ligament afferents only
(Vmes)
Muscle innervation Sparse or misshaped muscle spindles in lip, Lennartsson B. (1979), Bruenech J. R. and
extraocular and jaw opening muscles of Ruskell G. L. (2001)
humans
Embryonic origin Ophthalmic division of TG from neurogenic Noden D. M. (1991), Artinger K. B. et al. (1998),
placodes maxillary, mandibular divisions of TG Baker C. V. et al. (2002)
from neural crest
Sensory axon % A- fibers > % C-fibers Young R. F. and King R. B. (1973), Holland G. R.
diameter and Robinson P. P. (1992)
Sensory fiber Descending branch of Vtr: % C-fibers > A-- Tashiro T. et al. (1984)
bifurcation fibers
Sensory fiber Direct sensory fiber projection to autonomic Jacquin M. F. et al. (1983), Marfurt C. F. and
termination relay nuclei (e.g., NTS) Rajchert D. M. (1991), Panneton W. M. et al.
(1994)
Linkage to Cutaneous vasodilatation, hypotension and Kumada M. et al. (1977), Drummond P. D. (1992),
parasympathetic bradycardia to TG stimuli Ramien M. et al. (2004)
outflow
Sympathetic fibers Vn << spinal nerves Hoffmann K. D. and Matthews M. A. (1990)
Central trigeminal system
Facial dermatomes Representation of orofacial tissues at multiple Marfurt C. F. (1981), Panneton W. M. and Burton
levels of TSNC H. (1981), Shigenaga Y. et al. (1986b), Ma P.
M. (1991)
Onion skin Perioral and midline tissues represented more Jacquin M. F. et al. (1986), Shigenaga Y. et al.
organization rostral than lateral tissues (1986a)
Sensory Termination of cranial nerves V, VII, IX, X, and Kerr F. W. L. (1961), Beckstead R. M. and
convergence upper cervical rootlets in caudal Vc Norgren R. (1979), Hu J. W. et al. (2005)
Intersubnuclear Lissauers tract equivalent plus deep bundles Gobel S. and Purvis M. B. (1972), Kruger L. et al.
connections (1977), Ikeda M. et al. (1984), Jacquin M. F.
et al. (1990)

CNS, central nervous system; NTS, nucleus tractus solitarius; TG, trigeminal ganglion; TSNC, trigeminal sensory brainstem nuclear
complex.

significance of protecting ones face and self-image orofacial pain such as headache, temporomandibular
suggests that the Vn system has evolved features that disorders (TMD), odontalgia, and dry eye are difficult
set it apart from spinal systems. to treat clinically and may be associated with lower
Chronic orofacial pain is a significant public health pain thresholds elsewhere in the body (Burstein, R.
concern that affects more than 20% of the population et al., 2000; Sarlani, E. and Greenspan, J. D., 2003; Van
in western societies (Macfarlane, T. V. et al., 2002b). Bijsterveld, O. P., et al., 2003). Moreover, epidemiolo-
Persistent orofacial pain has profound effects on eating gical studies indicate that many chronic craniofacial
and sleeping patterns, a strong association with pain conditions are more prevalent and occur with
depression, and a reduced sense of wellbeing greater intensity in women than men (Lipton, J. A.
(Korszun, A., 2002). Many common forms of chronic et al., 1993; Sandstedt, P. and Sorensen, S., 1995;
438 Trigeminal Mechanisms of Nociception: Peripheral and Brainstem Organization

LeResche, L., 1997; Rasmussen, B. K., 2001; Wolf, E. Specialized transduction processes also are associated
et al., 2001). This review highlights several neurobio- with these unique tissues such as the proposed hydro-
logical aspects of the Vn system that differ from spinal dynamic movement of fluid as the basis for dentinal
systems and may contribute to the pattern and magni- pain (Brannstrom, M., 1986).
tude of craniofacial pain. Peripheral Vn afferents are subject to frequent injury
due to trauma, infection, or iatrogenic damage follow-
ing surgery that can lead to peripheral sensitization.
32.2 The Peripheral Trigeminal Indeed, one of the first demonstrations of enhanced
Nerve System peripheral C-fiber activity due to injury was reported
for facial afferents in the monkey (Beitel, R. E. and
32.2.1 General Features
Dubner, R., 1976). Since peripheral sensitization of
The sensory portion of the Vn consists of three main small myelinated A- versus unmyelinated C-fibers
nerve branches: ophthalmic, maxillary, and mandibu- does not occur uniformly, but rather varies for different
lar. Proprioceptive afferents from select craniofacial tissue types and depends on the nature of the injury (see
muscles and periodontal ligaments travel within the Meyer, R. A. et al., 2006), it may be significant that the
Vn, however; the cell bodies are located in the mesen- sensory root of the Vn has a greater percentage of A-
cephalic nucleus of V (Vme) in the rostral pons (Cody, fibers than C-fibers compared to spinal nerves as deter-
F. W. et al., 1972; Linden, R. W., 1978). Unlike the mined by counts of axons (Young, R. F. and King, R. B.,
spinal dorsal root ganglion (DRG) that has no apparent 1973; Holland, G. R. and Robinson, P. P., 1992) or lectin
intraganglionic organization, the somata of neurons staining for C-fibers (Ambalavanar, R. and Morris. R.,
giving rise to the three branches of Vn are somatoto- 1992; Wang, H. et al., 1994). Correspondingly, corneal
pically arranged within the trigeminal ganglion (TG; (Belmonte, C. et al., 1997) and dural afferents (Levy, D.
Kerr, F. W. L., 1963; Marfurt, C. F., 1981; Arvidsson, J. and Strassman, A. M., 2002) conducting in the A-
et al., 1992; Aigner, M. et al., 2000). Table 1 lists several range display greater sensitization than more slowly
features of the peripheral Vn system that differ from conducting C-fibers.
the spinal system. Differences relevant to pain proces- Complex regional pain syndrome (CRSP), for-
sing include the innervation of specialized tissues, merly referred to as causalgia, is a neuropathic pain
composition, and central termination of afferents; rela- condition with sympathetic involvement, often
tionship of sensory nerves to sympathetic efferent resulting from trauma or bone fracture to the distal
outflow; and neurochemical markers in ganglion cells. limbs (see Wasner, G. et al., 1998). The incidence of
The Vn innervates a heterogeneous and unique group CRSP-like symptoms following trauma to craniofa-
of craniofacial tissues well associated with pain sensa- cial tissues is relatively low compared to injury of
tion. The cornea is the most densely innervated tissue other tissues (Matthews, B., 1989). The exact reason
of the body, supplied exclusively by small diameter for this observation is not certain; however, fewer
fibers (Rozsa, A. J. and Beuerman, R. W., 1982); is the sympathetic efferent fibers course within sensory
only tissue in which nerve fibers penetrate the outer branches of Vn compared to spinal nerves, and
epithelial layers (Hoyes, A. D. and Barber, P., 1976; instead these fibers travel along the arteries that
Muller, L. J. et al., 1996); and responds to very small supply the head and oral cavity (Hoffmann, K. D.
changes in ocular surface temperature, moisture status, and Matthews, M. A., 1990; Maklad, A. et al., 2001).
or foreign bodies (Belmonte, C. et al., 1997). The oral Spinal nerve injury induces sprouting of noradrener-
mucosal epithelium also is densely innervated by poly- gic fibers into the DRG (McLachlan, E. M. et al.,
modal small diameter Vn afferents with relatively large 1993), whereas comparable injury to Vn produces
receptive fields (RFs) compared to cutaneous receptors no such sprouting into the TG (Bongenhielm, U.
(Toda, K. et al., 1997) that provide proprioceptive et al., 1999; Benoliel, R. et al., 2001). Furthermore,
information on lip/tongue position necessary for eating cervical sympathectomy does not alter the develop-
and speech as well as detecting noxious stimuli within ment of ectopic discharge or increased mechanical
the mouth (Trulsson, M. and Johansson, R. S., 2002). By sensitivity after inferior alveolar nerve transection
contrast, the dental pulp is well insulated from the (Bongenhielm, U. et al., 1998). Since the basis for
exterior environment and, while pain is the predomi- sprouting into the DRG following nerve injury is
nant sensation relayed by pulpal afferents, dental thought to involve neurotrophins (Davis, B. M. et al.,
afferents respond best to natural stimuli after injury 1994), it is interesting to note that the number of
(Byers, M. R., 1984; Hildebrand, C. et al., 1995). trkA-positive corneal and dental sensory TG
 Trigeminal Mechanisms of Nociception: Peripheral and Brainstem Organization 439

neurons is significantly less (< 10%, Mosconi, T. receptor subtypes (Collo, G. et al., 1996; Cook, S. P.
et al., 2001) than for spinal DRG neurons (40%, et al., 1997; Xiang, Z. et al., 1998). However, substantial
Molliver, D. C. and Snider, W. D., 1997). Failure to differences also are observed that include: the per-
observe sympathetic sprouting may be specific for centage of neurons stained for substance P and
nerve injury, since central administration of nerve somatostatin (TG > DRG; Kai-Kai, M. A., 1989),
growth factor (NGF) induces significant sprouting trkA receptor (DRG > TG; Mosconi, T. et al., 2001),
into the TG and spinal DRG (Nauta, H. J. et al., 1999). galectin-1 (DRG >> TG; Akazawa, C. et al., 2004),
m- and -opioid receptors (DRG > TG; Buzas, B. and
Cox, B. M., 1997), CCK (DRG > TG; Ghilardi, J. R.
32.2.2 Neurochemical and Molecular
et al., 1992), cytokeratin (TG >> DRG; Okabe, H.
Properties of Trigeminal Ganglion Neurons
et al., 1997), and neuropeptide Y (NPY)-binding
Although this review emphasizes comparisons of the sites (TG > DRG; Mantyh, P. W. et al., 1994). In
TG and DRG systems in the adult, significant devel- addition, TG and DRG systems display a differential
opmental differences also exist. While the functional sensitivity to ganglion cell labeling by selected ana-
significance of developmental differences (e.g., tomical tracers in which the TG system has a greater
neurogenic placode vs. neural crest) may not be uptake of Fluoro-Gold than DRG but not of Fast
obvious in the healthy adult, many responses to Blue (Yoshimura, N. et al., 1994).
injury recapitulate developmental differences and Differences in cellular properties between the
thus may have implications in the injured adult TG and DRG systems under naive conditions may
nervous system. Studies reporting similarities in contribute to differential responses to tissue injury.
development of the TG and DRG systems have For example, sprouting of sympathetic nerve term-
included: P2X3 receptor expression (Ruan, H. Z. inals into the DRG (McLachlan, E. M. et al., 1993),
et al., 2004); phenotype expression in trkB knockout but not the TG after nerve injury (Bongenhielm, U.
mice (Klein, R. et al., 1993; Gonzalez-Martinez, T. et al., 1999; Benoliel, R. et al., 2001) is consistent with
et al., 2004); vesicular monoamine transporter levels findings that trkA-positive neurons are more
(Hansson, S. R. et al., 1998); the apoptotic enzyme, numerous in the DRG than TG (Mosconi, T.
CPP32/apopain (Mukasa, T. et al., 1997); neuronal et al., 2001). Comparison of nerve injury-induced
tyrosine hydroxylase (Son et al., 1996; Kim, S. J. et al., changes for a majority of neuropeptides associated
1997); six 4/ARECC3 mRNA (Esteve, P. and nociception such as substance P, TRPV1, P2X3, and
Bovolenta, P., 1999); the transcription factor, reti- NPY appear similar for TG and DRG systems
noid-X receptor  (Georgiades, P. et al., 1998); (Zhang, X. et al., 1996; Okuse, K. et al., 1997;
guanosine triphosphate (GTP)-binding protein G- Eriksson, J. et al., 1998; Elcock, C. et al., 2001;
z (Kelleher, K. L. et al., 1998); the fetal plasma glyco- Tsuzuki, K. et al., 2001; Stenholm, E. et al., 2002;
protein, fetuin (Kitchener, P. D. et al., 1997); FGF Tsuzuki, K. et al., 2003). However, species differ-
receptor mRNA (Wanaka, A. et al., 1991); and the ences have been reported such as a decrease in
large zinc finger protein, KRC (Hicar, M. D. et al., galanin in TG of ferret (Elcock, C. et al., 2001)
2002). By contrast, studies reporting significant compared to an increase in rat (Zhang, X. et al.,
differences in the developing TG and DRG systems 1996) after Vn injury, similar to the increase in
have included: expression of trkC at day 12.5 (DRG: galanin in rat DRG after spinal nerve injury
yes, TG: no; Elkabes, S. et al., 1994); requirement for (Villar, M. J. et al., 1989). Nerve injury causes higher
NGF at day E16.5 (DRG > TG; Goedert, M. et al., spontaneous discharge rates and greater rhythmic
1984); and sensitivity to cadmium-induced neuro- firing patterns in DRGs than TGs (Tal, M. and
toxicity (TG > DRG; Arvidson, B., 1983). Thus, Devor, M., 1992), effects often associated with
developmental biology indicates some similarities sodium channel activity (see Wood, J. N. et al.,
with notable differences in the responsiveness to 2004). However, the basis for this difference is not
certain neurotrophic factors. certain since changes in the expression of the tetro-
Table 2 summarizes results from studies compar- dotoxin (TTX)-resistant sodium channel, NaV1.8,
ing adult TG and DRG systems under basal (naive) appear comparable for DRG and TG systems
conditions. These studies reveal many similarities (Dib-Hajj, S., et al., 1996; Bongenhielm, U. et al.,
among markers associated with nociception such 2000). Susceptibility to infection also may differ
as the percentage of TRPV1-positive neurons between TG and DRG systems, since injection of
(Ambalavanar, R. et al., 2005) and expression of P2X herpes simplex virus (HSV) to the left ear pinna in
440 Trigeminal Mechanisms of Nociception: Peripheral and Brainstem Organization

Table 2 Comparison of trigeminal (TG) and dorsal root ganglion (DRG) systems under naive conditions

Factor Details References

Substance P TG  DRG Kai-Kai M. A. (1989)

Somatostatin TG  DRG Kai-Kai M. A. (1989)
P2X1, P2X2, P2X3, P2X4, P2X5, P2X6 TG  DRG; no P2X2, P2X3 in Vme Collo G. et al. (1996), Cook S. P. et al.
mRNA (1997) Xiang Z. et al. (1998)
P2X3-isolectin B4 co-expression DRG >> TG Ambalavanar R. et al. (2005)
TRPV1 TG  DRG Guo A. et al. (1999)
trkA DRG > TG;  40% DRG versus Mosconi T. et al. (2001)
1015% of TG neurons innervating
pulp or cornea
Arginine vasopressin DRG > TG About 40% AVP is in Kai-Kai M. A. and Che Y. M. (1995)
capsaicin-sensitive neurons in
both ganglia
5-HT1d TG  DRG Potrebic S. et al. (2003)
Oxytocin TG > DRG Kai-Kai M. A. (1989)
NADPH-diaphorase DRG: T5L1 >> C1-T4 L2S TG Aimi Y. et al. (1991)
CCK mRNA in monkey DRG 20%; TG 10% of neurons Verge V. M. et al. (1993)
CCK(B) receptor Rat, rabbit: TG  DRG Ghilardi J. R. et al. (1992)
Monkey: DRG > TG (nondetectable)
PYY binding sites (NPY receptor) TG  DRG Mantyh P. W. et al. (1994)
Galectin-1 mRNA DRG >> TG (nondetectable) Akazawa C. et al. (2004)
Oncostatin M (OSM-) DRG > TG (OSMr- coexpressed in Tamura S. et al. (2003)
TRPV1-positive neurons)
glycogen phosphorylase DRG > TG Pfeiffer B. et al. (1995)
Cytokeratin (AE1 and CAM5.2) TG >> DRG Okabe H. et al. (1997)
Glucocorticoid receptor (GR) TG: GR expressed in substance P, DeLeon M. et al. (1994)
CGRP, but not galanin-positive
neurons
DRG: GR expressed in substance P,
CGRP, and galanin-positive neurons
MOR mRNA DRG: lumbar > thoracic  cervical > TG Buzas B. and Cox B. M. (1997)
DOR mRNA DRG: lumbar  thoracic  cervical > TG Buzas B. and Cox B. M. (1997)
TGF- mitogenic effect in vitro DRG: Yes Chalazonitis A. et al. (1992)
TG: No
Parvalbumin TG smaller size than DRG, but both Ichikawa H. et al. (1994)
populations have high expression of
carbonic anhydrase and low
expression of CGRP
Osteocalcin parvalbumin TG: 25% of neurons (31% express Ichikawa H. et al. (1999)
parvalbumin)
Vme: 63% of neurons (>90% express
parvalbumin)
DRG: 16% of neurons (>90% express
parvalbumin)
Osteocalcin and TRPV1 coexpression TG 14% Ichikawa H. and Sugimoto T. (2002)
DRG none
Calretinin TG: neurons mostly < 800 mm2 and Ichikawa H. et al. (1993)
34% positive for tachykinin
DRG: neurons mostly > 800 mm2 and
7% positive for tachykinin
S100 calcium-binding protein TG: 59% (> 90% coexpress Ichikawa H. et al. (1997)
parvalbumin and calbindin D-28k)
DRG: 44% (> 90% coexpress
parvalbumin, none with calbindin
D-28k)

(Continued )
 Trigeminal Mechanisms of Nociception: Peripheral and Brainstem Organization 441

Table 2 (Continued)

Factor Details References

tr7kC at day E12.5 of development DRG: Yes Elkabes S. et al. (1994)

TG: No
NT-3 overexpression TG > DRG for responsiveness Albers K. M. et al. (1996)
Neonatal anti-NGF antisera at E16.5 DRG > TG for loss of substance P- and Goedert M. et al. (1984)
somatostatin-positive neurons
Uptake of fluorescent dyes TG  DRG uptake of Fast Blue Yoshimura N. et al. (1994)
TG > DRG uptake of Fluoro-Gold

5-HT1d, serotonin receptor subtype; CCK, cholecystokinin; CGRP, calcitonin gene-related peptide; DOR, -opioid receptor; DRG, dorsal
root ganglion; MOR, m-opioid receptor, messenger RNA; NADPH, nicotinamide adenine dinucleotide phosphate hydrogen; ; NGF, nerve
growth factor; NPY, neuropeptide Y; P2X, ATP receptor; PYY, polypeptide Y; TG, trigeminal ganglion; TGF, tumor growth factor; trkA,
tyrosine kinase A receptor subtype; TRPV1, vanilloid receptor.

mice produces 100% HSV infection in both the dorsal to Vp, an elongated spinal nucleus (Vsp)
ipsilateral TG and cervical DRG. Interestingly, extending from the pons to the upper cervical spinal
70% of the TGs contralateral to injection, while cord, and the interstitial islands or the paratrigem-
only 10% of contralateral DRGs were infected inal region (Pa5) embedded within the spinal
(Thackray, A. M. and Field, H. J., 1996). trigeminal tract, dorsal and lateral to the caudal
Interhemispheric neural communication likely con- Vsp (Figure 2; see Kruger, L. and Young, R. F.,
tributes to the progression of joint-related pain in 1981; Renehan, W. E. and Jacquin, M. F., 1993).
trigeminal (see Bereiter, D. A. et al., 2005b) as well The Vsp is further subdivided, from rostral to cau-
as spinal systems (Levine, J. D. et al., 1985; Shenker, dal, into subnucleus oralis (Vo), subnucleus
N. et al., 2003). The responsiveness to viral vectors interpolaris (Vi), and a laminated trigeminal subnu-
in the Vn system has prompted recent efforts to cleus caudalis (Vc) as described originally by
deliver targeted transgene-derived products for Olszewski J. (1950). Although nociceptive neurons
control of trigeminal neuropathic pain (Meunier, in the caudal laminated portion of the TSNC, Vc,
A. et al., 2005). Collectively, these studies reveal display properties similar to those at spinal levels
significant differences in peripheral Vn and spinal (Price, D. D. et al., 1976; Dubner, R. and Bennett, G.
systems under nave and injured conditions, differ- J., 1983) consistent with a prominent role in noci-
ences that could not be predicted on the basis of ceptive processing (see also Bereiter, D. A. et al.,
results from spinal sensory systems alone. 2000; Sessle, B. J., 2000), the contribution of rostral
portions of the TSNC to orofacial pain is less
certain.
32.3 Central Aspects of Trigeminal
Organization
Vp
Noxious sensory information is relayed from Vn
Vo
afferents to second-order neurons in the TSNC Vi
Vmo
and the upper cervical spinal cord. The TSNC is Vc
the initial site of synaptic integration for sensory Rostral
Vme
input from the head and oral cavity and shares this
feature with the spinal dorsal horn and dorsal col-
umn nuclei that receive sensory input from the rest
of the body (Figure 1). However, unlike the spinal NTS
cord, the TSNC is comprised of several cell groups
with distinct cytological and organizational features
(see Darian-Smith, I., 1973; Kruger, L. and Young,
R. F., 1981) yet each cell group receives direct
primary afferent projections from specific craniofa- Caudal
cial tissues. The TSNC consists of: the principal
nucleus (Vp), supratrigeminal region (Vsup) lying Figure 2 Trigeminal brainstem sensory complex.
442 Trigeminal Mechanisms of Nociception: Peripheral and Brainstem Organization

32.3.1 Somatotopy that rostral portions of TSNC play a more prominent

role in dental and intraoral pain than caudal portions
Somatotopy is a key feature of the trigeminal system
(Young, R. F. and Perryman, K. M., 1984; see Sessle,
and is seen within the TG as well as the TSNC
B. J., 2000); however, the importance of multiple
distinct from the spinal cord. Craniofacial tissues
representation of extraoral tissues for other forms of
are represented at multiple levels of the TSNC,
craniofacial pain is not well understood. One reason
while sensory afferents from other body loci termi-
for this uncertainty may be due to the fact that many
nate at several contiguous spinal segments of the craniofacial tissues are represented in a discontinuous
spinal dorsal horn. Also, at caudal levels of the manner along the rostrocaudal extent of the TSNC as
TSNC craniofacial tissues are represented in a series summarized in Table 3. Although a small percentage of
of semicircular bands that converge at the rostral Vn afferents projects to both rostral and caudal portions
midline of the face, often referred to as an onion- of the TSNC (Silverman, J. D. and Kruger, L., 1985; Li,
skin arrangement and by a medial-lateral representa- Y. Q. et al., 1992), most fibers either ascend in a short
tion in which the head is inverted (Jacquin, M. F. sensory root to terminate in Vp or descend to give off
et al., 1986; Shigenaga, Y. et al., 1986a). Although branches to Vo, Vi, Vc, and the upper cervical dorsal
somatotopy along the mediolateral axis is preserved horn. Several aspects of the afferent input pattern to
at all levels of the TSNC, the onion-skin arrange- TSNC are notable. First, the superficial laminas in Vc
ment is most apparent in Vc. The implications of this receive substantial input from all specialized tissues
organization for facial pain have been debated since well associated with craniofacial pain conditions (e.g.,
an early report by Sjoqvist O. (1938) that trigeminal cornea, dura, teeth, temporomandibular joint (TMJ)),
tractotomy at the level of rostral Vc reduced the pain while afferents from structures with no known rela-
of trigeminal neuralgia, while preserving the sense of tionship to pain perception (vibrissas) do not project
temperature and touch on the face. It is understood to this region. Second, dental pulp afferents are the

Table 3 Summary of the relative density of trigeminal primary afferent terminals within different portions of the sensory
trigeminal sensory nuclear complex.

Vp Vo Vc

dm vl dm vl Vi dPa5 Vi/Vc III IIIIV V References

Cornea      Panneton W. M. and Burton H.

(1981), Marfurt C. F. and del
Toro D. R. (1987), Marfurt C.
F. and Echtenkamp S. F.
(1988)
Nasal   Anton F. and Peppel P. (1991);
mucosa Panneton W. M. (1991)
Dura       Arbab M. A. et al. (1988), Liu Y.
et al. (2004)
Teeth   Marfurt C. F. and Turner D. F.
(1984), Shigenaga Y. et al.
(1986c); Takemura M. et al.
(1993)
Masseter     Nishimori T. et al. (1986),
muscle Shigenaga Y. et al. (1988),
Arvidsson J. and Raappana P.
(1989)
TMJ     Jacquin M. F. et al. (1983);
Shigenaga Y. et al. (1986a),
(1986b), Capra N. F. (1987),
Takemura M. et al. (1987)
Vibrissae       Jacquin M. F. et al. (1986),
Nomura S. et al. (1986),
Arvidsson J. (1982)

, very few or none; ,,, weak, moderate and dense terminal distribution; dPa5, dorsal paratrigeminal region; TMJ,
temporomandibular joint; Vc, subnucleus caudalis; Vi, subnucleus interpolaris; Vo, subnucleus oralis; Vp, principal sensory nucleus.
 Trigeminal Mechanisms of Nociception: Peripheral and Brainstem Organization 443

only group of putative nociceptive fibers with a sub- (Greenwood, L. F. and Sessle, B. J., 1976; Chiang, C.
stantial projection to rostral portions of the TSNC, Y. et al., 2002), dura (Davis, K. D. and Dostrovsky, J. O.,
namely, to the dorsomedial portions of Vp and Vo, 1988), and cornea (Hirata, H. et al., 2003). By contrast,
thus supporting the role for these regions in intraoral innocuous sensory input from facial skin (Greenwood,
pain (Marfurt, C. F. and Turner, D. F., 1984; L. F. and Sessle, B. J., 1976) and vibrissae (Hallas, B. H.
Shigenaga, Y. et al., 1986c). Also, compared to other and Jacquin, M. F., 1990) often was enhanced. Fewer
orofacial tissues associated with pain sensation, the studies have assessed the influence of rostral TSNC
termination pattern of dental afferents is particularly regions on caudal Vc neural activity, though in a recent
widespread along the rostrocaudal extent of the study, muscimol blockade of the Vi/Vc transition
TSNC. Third, input from structures supplied by region facilitated cornea-responsive neurons in laminas
the ophthalmic branch of the Vn (e.g., cornea, nasal III of caudal Vc (Hirata, H. et al., 2003). These results
dura) project only sparsely to rostral portions of the suggest that ascending as well as descending connec-
TSNC suggesting that these regions play a lesser role tions within the TSNC contribute to the integration of
than Vc in mediating pain due to uveitis, dry eye, sensory inputs relevant for craniofacial pain.
sinusitis, or headache. Fourth, discontinuous repre-
sentation is not unique to tissues supplied by the
32.3.3 Relationship to the Autonomic
ophthalmic division (e.g., cornea, dura, nasal cavity)
Nervous System
since the auriculotemporal nerve, a major source of
innervation for the TMJ region also displays an The trigeminal system is closely linked to brain regions
uneven terminal distribution in the TSNC that control autonomic outflow, especially parasympa-
(Shigenaga, Y. et al., 1986a; 1986b). In rodents vibris- thetic outflow and vagus nerve activity. This linkage
sae afferents project to most levels of the TSNC likely contributes to craniofacial pain conditions such as
(Arvidsson, J., 1982; Nomura, S. et al., 1986); however, primary headache (Edvinsson, L. and Uddman, R.,
the architectonic representation of the vibrissae 2005) and dry eye (Hocevar, A. et al., 2003), sudden
fields, the so-called barrelettes, are well delineated bradycardia and asystole during maxillofacial surgery
in Vp, Vi, and Vc, but not Vo (Ma, P. M., 1991), (Schaller, B., 2004), and the so-called diving reflex in
supporting the notion that different levels of the infant humans (Goksor, E. et al., 2002) and aquatic
TSNC mediate different aspects of sensory proces- mammals (Butler, P. J. and Jones, D. R., 1997). Even
sing of innocuous as well as noxious inputs. facial skin differs from other cutaneous regions in that it
is well supplied by parasympathetic fibers (Ramien, M.
et al., 2004). Under experimental conditions noxious
32.3.2 Intersubnuclear Connections
stimulation of craniofacial tissues in humans evokes
Rostral and caudal portions of the TSNC are con- long-lasting vasodilatation in orofacial regions
nected by a rich longitudinal fiber network coursing (Drummond, P. D., 1992; Izumi, H., 1999) that differs
within spinal trigeminal tract and through the deep from responses evoked by stimulation of other body
bundles that extend from Vp to the upper cervical regions consistent with the existence of specialized
spinal cord (Gobel, S. and Purvis, M. B., 1972; trigeminal vasodilator reflex mechanisms
Kruger, L. et al., 1977; Ikeda, M. et al., 1984; Jacquin, (Kemppainen, P. et al., 2001). Two aspects of the rela-
M. F. et al., 1990). Although propriospinal-like connec- tionship between autonomic nerves and the trigeminal
tions of the TSNC share anatomical similarities with system are distinct from spinal cord and deserve special
those at lower spinal levels, intersubnuclear connec- mention. Unlike at spinal levels where nearly all sen-
tions in the TSNC link spatially distinct brainstem sory nerves relay initially in the dorsal horn or dorsal
regions with common somatotopic representation of column nuclei, many Vn afferents, especially those
facial fields. The implications of this organization for from the mandibular branch, project directly to brain-
facial pain remain to be determined; however, results stem nuclei that control autonomic outflow such as the
from numerous animal studies support the clinical nucleus tractus solitarius (NTS), parabrachial complex,
findings of Sjoqvist O. (1938) and indicate that ascend- and ventrolateral medulla (Kerr, F. W. L., 1961; Jacquin,
ing connections from caudal Vc generally facilitate the M. F. et al., 1983; Marfurt, C. F. and Rajchert, D. M.,
activity of neurons in more rostral portions within the 1991; Panneton, W. M., 1991; Panneton, W. M. et al.,
TSNC. Lesion or chemical blockade of caudal Vc 1994). Also, unlike lower portions of the spinal cord,
reduced the excitability of rostral trigeminal neurons there is an extensive convergence of Vn, facial, glosso-
responsive to noxious stimulation of tooth pulp pharyngeal, and vagal afferents to common laminae of
444 Trigeminal Mechanisms of Nociception: Peripheral and Brainstem Organization

the lower TSNC and upper cervical dorsal horn 32.3.4 Neurochemical Markers
(Denny-Brown, D. and Yanagisawa, N., 1973;
The role of central neurons in mediating the various
Beckstead, R. M. and Norgren, R., 1979; Contreras, R.
aspects of pain (e.g., sensation, autonomic control,
J. et al., 1982; Altschuler, S. M. et al., 1989; McNeill, D. L.
motor reflexes) can be predicted, in part, on the basis
et al., 1991). The dorsal paratrigeminal region (dPa5)
of key factors such as the nature of the sensory input,
also receives inputs from multiple cranial nerves and
encoding properties, response to analgesic agents, effer-
upper cervical rootlets, has widespread connections to
ent projections (Price, D. D. and Dubner, R., 1977;
central autonomic pathways and other regions of the
Price, D. D. et al., 2003), and, more recently, the dis-
TSNC, and may be a key site of somaticautonomic
tribution of a growing list of neurochemical markers
integration for cutaneous and visceral sensory input
associated with nociceptive processing (Woolf, C. J. and
and control of homeostasis (Panneton, W. M. and
Burton, H., 1985; Saxon, D. W. and Hopkins, D. A., Salter, M. W., 2000; Julius, D. and Basbaum, A. I., 2001;
1998; Caous, C. A. et al., 2001). Considerable evidence Lewin, G. R. et al., 2004). The data provided in Tables 4
suggests that the Vc/upper cervical cord (Vc/C2) junc- (neurochemical markers) and 5 (efferent projections)
tion region differs from lower spinal cord. In addition to are consistent with the notion that different portions
receiving convergent input from multiple cranial of the TSNC contribute to different aspects of cranio-
nerves and upper cervical rootlets (Pfaller, K. and facial pain. However, these data derive from results in
Arvidsson, J., 1988; Neuhuber, W. L. and Zenker, W., nave animals and do not specifically indicate which
1989), second-order Vc/C2 neurons have widespread regions respond with phenotypic or long-term struc-
ascending connections to the hypothalamus (Burstein, tural changes during chronic pain (see Basbaum, A. I.,
R. et al., 1990) and periaqueductal gray (PAG; Keay, K. 1999; Hunt, S. P. and Mantyh, P. W., 2001; Scholz, J.
A. et al., 1997), brainstem regions well associated with and Woolf, C. J., 2002), changes that likely occur
control of autonomic outflow, and endogenous pain unequally in different portions of the TSNC.
modulation circuits. The Vc/C2 junction also sends Table 4 summarizes the pattern of distribution of
long-range descending projections to the lower spinal selected neurochemical markers with known associa-
cord and is a critical region for visceral sensory, parti- tion to nociceptive processing and/or its modulation.
cularly vagus nerve, modulation of somatic input to The most striking aspect of these data is the dense and
lower spinal segments (Chandler, M. J. et al., 2002). almost universal distribution of all markers within the
Although vagus nerve stimulation generally is asso- superficial laminae of Vc, while only weak labeling is
ciated with antinociception in humans (Kirchner, A. seen in laminae not associated with nociceptive proces-
et al., 2000) and animals (Randich, A. and Gebhart, G. sing (laminae IIIIV). In rostral portions of the TSNC
F., 1992; Khasar, S. G. et al., 1998), the relationship the density of different markers is more varied than in
between vagus nerve activity and facial pain may be Vc. For example, the dorsomedial portions of Vp and
more complex. For example, increased vagal afferent Vo, regions that have a high density of afferent term-
activity has been suggested as one source of facial pain inals from tooth pulp nerves, also display intense
referred from the lung in cancer patients (see Sarlani, labeling for calcitonin gene-related peptide (CGRP)
E. et al., 2003), whereas in animal studies increased and trkA, whereas labeling for substance P and inositol
vagal activity inhibits painlike behavior and c-fos 1,4,5 triphosphate (IP3) receptor are relatively weak
expression after formalin injection into facial skin and that for MOR1, the m-opioid receptor, is absent.
(Bohotin, C. et al., 2003) and tooth pulp-evoked activ- Interestingly, selective agonists for the 5-HT1B recep-
ity of Vc/C2 dorsal horn neurons (Tanimoto, T. et al., tor, a serotonergic receptor subtype that binds
2002). The effects of vagus nerve stimulation have sumatriptan, an effective therapeutic agent for
been tested mainly on neurons in the caudal portions migraine, displays moderate density in the dorsomedial
of the TSNC; however, vagal stimulation also inhibits portions of Vp and Vo, yet neither region receives
tooth pulp-evoked digastric reflexes (Bossut, D. F. significant input from meningeal afferents. Although
et al., 1992) suggesting that modulation of neurons in the laminar distribution of most neurochemical markers
rostral portions of the TSNC is possible. Given the in Vc and the spinal dorsal horn are similar, significant
extensive convergence and efferent projections of sec- differences have been reported for IB4 (Sugimoto, T.
ond-order neurons at the dPa5 and Vc/C2 regions, it et al., 1997a) and TRPV1 (Bae, Y. C. et al., 2004), where
is tempting to speculate that trigeminalvagal interac- spinal lamina IIi contains a greater expression of both
tions play a significant role in modulating pain above markers than lamina IIo, while the reverse is seen in Vc.
as well as below the neck. Also, the distribution of CGRP and substance P appears
 Trigeminal Mechanisms of Nociception: Peripheral and Brainstem Organization 445

Table 4 Summary of the distribution of neurochemical markers associated with nociceptive processing in different
portions of the trigeminal sensory nuclear complex

Vp Vo Vc

dm vl dm vl Vi dPa5 Vi/Vc III IIIIV V References

IB4      Sugimoto T. et al. (1997a)

TRPV1     Bae Y. C. et al. (2004)
SP    Boissonade F. M. et al.
(1993), Sugimoto T. et al.
(1997b)
NK1   Nakaya Y. et al. (1994)
CGRP    Henry M. A. et al. (1996),
Kruger L. et al. (1988),
Sugimoto T. et al. (1997b)
P2X2 Kanjhan R. et al. (1999)
trkA  Pioro E. P. and Cuello A. C.
(1990), Sobreviela T. et al.
(1994)
BDNF     Connor J. M. et al. (1997)
trkB Yan Q. et al. (1997)
EP3          Nakamura K. et al. (2000)
ChAT       Tatehata T. et al. (1987)
nNOS    Dohrn C. S. et al. (1994),
Rodrigo J. et al. (1994)
NR1 Petralia R. S. et al. (1994)
mGluR2 Ohishi H. et al. (1998)
GABA Ginestal E. and Matute C.
(1993)
GABAaR Fritschy J. M. and Mohler H.
(1995), Pirker S. et al.
(2000)
GABAbR Margeta-Mitrovic M. et al.
(1999)
NE/DA   Kitahama K. et al. (2000),
Levitt P. and Moore R. Y.
(1979)
AR/ Talley E. M. et al. (1996)
5-HT Harding A. et al. (2004),
Steinbusch H. W. M.
(1981)
5-HT1B/1D   Potrebic S. et al. (2003, Thor
K. B. et al. (1992)
IP3R      Rodrigo J. et al. (1993)
Calcineurin     Strack S. et al. (1996)
Osteocalcin    Ichikawa H. and Sugimoto
T. (2002)
Endo2      MartinSchild S. et al. (1999)
MOR1        Bereiter D. A. and Bereiter D.
F. (2000), Ding Y. Q. et al.
(1996)
ER        Bereiter D. A. et al. (2005a)

, very weak or no staining; ,,, weak, moderate and dense staining; 5-HT, serotonin; 5HT1/2, serotonin receptor subtypes;
AR/, adrenergic receptor subtypes; BDNF, brain-derived neurotrophic factor; CGRP, calcitonin gene-related peptide; ChAT, choline
acetyltransferase; dPa5, dorsal paratrigeminal region; Endo2, endomorphin 2; EP3, prostaglandin receptor; ER, estrogen receptor
alpha subtype; GABA, gamma-aminobutyric acid; GABAaR, GABA receptor  subtype, b2/3 subunit; GABAbR, GABA receptor 
subtype, R1a/b subunit; IB4, isolectin B4; IP3R, inositol triphosphate receptor; MOR1, m-opioid receptor; NE/DA, norepinephrine/
dopamine; NK1, neurokinin 1 receptor; nNOS, neuronal nitric oxide synthase; NR1, N-methyl-D-aspartic acid (NMDA) receptor
subunit; P2X2, ATP receptor; SP, substance P; trkA, tyrosine kinase A receptor subtype; trkB, tyrosine kinase B receptor subtype;
TRPV1, vanilloid receptor; Vc, subnucleus caudalis; Vi, subnucleus interpolaris; Vo, subnucleus oralis; Vp, principal sensory nucleus.
446 Trigeminal Mechanisms of Nociception: Peripheral and Brainstem Organization

more widely distributed across laminae IIII in Vc than poor localization and spreading of pain for many cra-
in spinal dorsal horn where both neuropeptides are niofacial pain conditions.
restricted to laminae I and IIo. The functional signifi-
cance of these differences are not certain; however, the
32.3.5 Efferent Projections
high degree of convergence of afferents from multiple
sensory ganglion sources (e.g., trigeminal, nodose, cer- Table 5 summarizes major efferent projection targets
vical dorsal root) to the caudal Vc may underlie the of trigeminal neurons in different portions of the

Table 5 Summary of efferent projections from different portions of the trigeminal sensory nuclear complex to thalamic,
pontine and medullary targets associated with various aspects of nociception

Vp Vo Vc

dm vl dm vl Vi dPa5 Vi/Vc III IIIIV IIIIV References

Thalamus
VPM Shigenaga Y. et al. (1983),
Bruce L. L. et al. (1987);
Mantle-St. John L. A.
and Tracey D. J. (1987)
PO    Dado R. J. and Giesler G.
J. (1990), Guy N. et al.
(2005)
SM        Craig A. D. and Burton H.
(1981), Dado R. J. and
Giesler G. J. (1990),
Yoshida A. et al. (1991)
Pf      Krout K. E. et al. (2002)
Hypothalamus
VMH    Malick A. and Burstein R.
(1998)
LH   Malick A. and Burstein R.
(1998), Ikeda T. et al.
(2003)
Tectum
APT       Yoshida A. et al. (1992)
SC     Bruce L. L. et al. (1987),
Ndiaye A. et al. (2002)
PAG         Beitz A. J. (1982), Mantyh
P. W. (1982), Wiberg M
et al. (1986), Keay K. A.
et al. (1997)
PBA   Panneton W. M. et al.
(1994), Feil K. and
Herbert H. (1995), Allen
G. V. et al. (1996)
NTS    Menetrey D. and Basbaum
A. I. (1987), Zerari-Mailly
F. et al. (2005)
ION      Huerta M. F. et al. (1985),
Van Ham J. J. and Yeo
C. H. (1992), Yatim N.
et al. (1996)

, very weak or no staining; ,,, weak, moderate, and dense staining; APT, anterior pretectal nucleus; dPa5, dorsal
paratrigeminal region; ION, inferior olivary nucleus; LH, lateral hypothalamic area; NTS, nucleus tractus solitarius; PAG, periaqueductal gray
region; PBA, parabrachial region; Pf, medial and lateral parafascicular nuclei; PO, posterior thalamic nucleus; SC, superior colliculus; SM,
nucleus submedius of thalamus; Vc, subnucleus caudalis; VMH, ventromedial hypothalamic area; Vi, subnucleus interpolaris; Vo,
subnucleus oralis; Vp, principal sensory nucleus; VPM, ventroposteromedial nucleus of thalamus.
 Trigeminal Mechanisms of Nociception: Peripheral and Brainstem Organization 447

TSNC independent of the functional class (i.e., noci- (Sessle, B. J. et al., 1981; Chiang, C. Y. et al., 1994)
ceptive versus innocuous sensory encoding). These and Vo (Chiang, C. Y. et al., 1989) are markedly
data derive largely from studies in rodents; however, inhibited by direct stimulation of PAG or rostral
the qualitative pattern of efferent projections from ventromedial medulla (RVM), the afferent pathways
the TSNC is similar across species, although there from second-order TSNC neurons to these endogen-
are significant quantitative species differences for ous pain control regions are not well defined.
some targets. Efferent projections from second- Compared to the significant input from upper cervi-
order neurons to the sensory thalamus have com- cal levels of spinal cord (Keay, K. A. et al., 1997)
manded considerable attention. The main sources of projections from TSNC to PAG are sparse (Beitz,
projections from the TSNC to the ventral posterior A. J., 1982). These results add substantial support to
medial nucleus (VPM) arise from Vp and Vi, which the notion that laminae III of Vc are critical regions
in the case of rodents and most carnivores, is due to a for processing nociceptive information relevant for
heavy projection from vibrissae-driven rather than multiple aspects of craniofacial pain. Behavioral evi-
nociceptive neurons (Veinante, P. et al., 2000). dence, though less extensively tested compared to
Nociceptive neurons in Vc that project to VPM spinal pain models, indicates that the Vc is necessary
originate mainly in laminae I and V (Ikeda, M. et al., for opioid modulation of cutaneous facial pain
2003) and moreover, the majority of projection neu- (Oliveras, J. L. et al., 1986) and that levels of attention
rons are found in rostral Vc rather than near the Vc/ markedly influence Vc nociceptive neurons and
C2 junction region (Guy, N. et al., 2005). Lamina I of behavioral responsiveness (Hayes, R. L. et al., 1981).
Vc projects heavily to the posterior thalamus (PO);
however, ventrolateral portions of Vo also provide a
significant input to PO (Guy, N. et al., 2005). Both Vc 32.4 Functional Considerations
and spinal dorsal horn lamina I cells project to similar
though adjacent regions of PO (Gauriau, C. and Recent developments in methods that assess neural
Bernard, J. F., 2004). Projections to parafascicular activity and encoding properties provide the stron-
thalamic nuclei from Vc are sparse compared to gest evidence regarding the functional role
spinal dorsal horn (Craig, A. D., 2004; Gauriau, C. of different regions of the TSNC in craniofacial
and Bernard, J. F., 2004). Trigeminal projections to pain. Advances in neuroimaging can distinguish
thalamic nucleus submedius (SM) reveal a unique somatotopic and simultaneous activation of brain-
pattern that differs markedly from spinal cord and stem and cortical responses to trigeminal stimuli in
exhibits significant species differences. In the rat the conscious humans (DaSilva, A. F. et al., 2002), though
majority of SM projections arise from the ventrolat- resolution is not yet sufficient to discern the relative
eral portion of the Vi/Vc transition region with only activation of different portions of the TSNC.
weak input from lamina I and V of caudal Vc, while Nociceptive neurons in the TSNC have been iden-
spinal projections to SM originate mainly from lami- tified and their properties determined mainly on the
nae VVII (Dado, R. J. and Giesler, G. J., 1990; basis of electrophysiological recording and, more
Yoshida, A. et al., 1991). By contrast, in cat and mon- recently, immediate early gene expression such as
key lamina I cells in Vc and spinal cord provide a c-fos. Immunostaining for Fos, the protein product
significant direct input to SM with Vc displaying a of c-fos, is a reliable method to identify populations of
somewhat more extensive projection (Craig A. D. and nociceptive central neurons at the single cell level
Burton, H., 1981). Trigeminal projections to auto- (see Bullitt, E., 1990), an advantage not readily
nomic control regions such as hypothalamus, achieved by electrophysiology. Although there are
parabrachial area, and NTS derive mainly from the examples of mismatches, properly designed c-fos stu-
dPa5, the Vi/Vc transition and lamina I of Vc, while dies generally complement electrophysiological
more rostral regions of the TSNC provide relatively results and have shed new light on long-standing
sparse input. This pattern is consistent with the dPa5 controversies in trigeminal physiology. For example,
and lamina I of Vc receiving direct input from the a role for Vo in dental pain is suggested by: a dense
vagus nerve. Spinal lamina I cells also project heavily terminal pattern in dorsomedial Vo from tooth pulp
to similar autonomic control regions of the brainstem afferents (Marfurt, C. F. and Turner, D. F., 1984), a
(Cechetto, D. F. et al., 1985; Menetrey, D. and moderate-to-high density of CGRP staining
Basbaum, A. I., 1987; Westlund, K. N. and Craig, A. (Sugimoto, T. et al., 1997b), and behavioral studies
D., 1996). Although nociceptive neurons in Vc revealing preservation of dental pain after trigeminal
448 Trigeminal Mechanisms of Nociception: Peripheral and Brainstem Organization

tractotomy at the level of caudal Vi (Young, R. F. and region is critical for reflex lacrimation (Hirata, H.
Perryman, K. M., 1984). By contrast, few Vo neurons et al., 2004).
can be driven by natural stimulation of the tooth pulp Repeated ocular surface stimulation evokes a wind-
compared to Vc (Hu, J. W. and Sessle, B. J., 1984) and up like response among caudal Vc units, while Vi/Vc
few Fos-positive neurons are found in Vo after acute cells rapidly become desensitized (Meng, I. D. et al.,
thermal stimulation of teeth (Chattipakorn, S. C. et al., 1997). In a model for endotoxin-induced uveitis,
1999). Thus, despite the fact that many Vo cells can at 7days after ocular inflammation convergent
be classified as nociceptive on the basis of cutaneous cutaneous RF areas become enlarged and responsive-
RF properties (Dallel, R. et al., 1990), display wind-up ness to ocular surface stimulation is enhanced among
to repeated cutaneous stimulation (Dallel, R. et al., caudal Vc neurons, while Vi/Vc transition cells display
1999), and are inhibited by systemic morphine no evidence of hyperalgesia (Bereiter, D. A. et al.,
(Dallel, R. et al., 1996), the role of Vo in acute dental 2005c).
pain remains uncertain. It is possible that the Vo acts Systemic morphine inhibits all ocular cells at the
as a silent pain relay in the TSNC and becomes caudal Vc, while nearly 30% of Vi/Vc cells are
active only after persistent tissue damage since Fos- enhanced; an effect that can be produced by micro-
positive cells first appear in Vo only several days injection of m-opioid receptor agonists directly into
after molar tooth pulp exposure (Byers, M. R. et al., the caudal Vc (Meng, I. D. et al., 1998; Hirata, H. et al.,
2000) and Vo neurons display sensitization provided 2000).
input from Vc remains intact (Chiang, C. Y. et al., The modality of ocular units at the Vi/Vc and
2002; Hu, B. et al., 2002). Alternatively, rostral TSNC caudal Vc predicts, in part, the efferent projections to
contributions to intraoral sensation and homeostasis PO or salivatory nucleus in the brainstem (Hirata, H.
may involve nonpulpal tissues (e.g., periodontal, et al., 2000). The Vi/Vc transition is unique among
muscosal receptors) and mediate select aspects of TSNC regions and is the main source of ascending
craniofacial pain (e.g., somatomotor reflexes). projections to SM (Yoshida, A. et al., 1991; Ikeda, M.
et al., 2003). These data suggest that the caudal Vc
underlies the sensory-discriminative aspects of ocu-
lar pain and modulation of ocular cells in more
32.4.1 Ocular Pain Processing
rostral regions via intersubnuclear connections. By
Ascribing a role for different portions of the TSNC in contrast, the Vi/Vc transition appears to play a sig-
ocular pain appears more straightforward than for nificant role in mediating ocular-specific reflexes
dental pain. (e.g., lacrimation, eye blink). Projections to SM,
Corneal afferents terminate mainly in ventrolat- coupled with the finding that many Vi/Vc neurons
eral portions of caudal Vi and Vc with few fibers display enhanced responsiveness after morphine,
projecting to more rostral regions (Panneton, W. M. suggests that this region may be part of the neural
and Burton, H., 1981; Marfurt, C. F. and del Toro, D. circuit that recruits endogenous pain controls in
R., 1987; Marfurt, C. F. and Echtenkamp, S. F., 1988). response to craniofacial tissue injury. Since a high
Acute stimulation of the ocular surface in the rat percentage of ocular cells at each region also respond
evokes a high density of Fos-positive cells at the to meningeal stimulation (Strassman, A. M. et al.,
Vi/Vc transition and caudal Vc and none in rostral 1994; Burstein, R. et al., 1998; Schepelmann, K. et al.,
regions of TSNC (Lu, J. et al., 1993; Strassman, A. M. 1999), it is proposed that rostral and caudal portions
and Vos, B. P., 1993; Bereiter, D. A. et al., 1994; Meng, of Vc mediate different aspects of headache as well as
I. D. and Bereiter, D. A., 1996). Converging lines of ocular pain.
evidence support the notion that the Vi/Vc transition
and caudal Vc serve different aspects of ocular pain.
All ocular cells in laminae III at the Vc/C2 32.5 Chronic Craniofacial Pain
junction are classified as nociceptive (wide-dynamic
range (WDR), nociceptive specific (NS)), while Chronic pain involving craniofacial tissues is a sig-
many cells at the Vi/Vc transition have no cutaneous nificant public health concern and a recognized
RF (Meng, I. D. et al., 1997; Hirata, H. et al., 1999). research priority for the National Institutes of
Many cells at the Vi/Vc transition are sensitive to the Health (NIH; e.g., PA 03-173: Neurobiology of
moisture status of the ocular surface, while few such Persistent Pain Mediated by the Trigeminal Nerve).
neurons are found in caudal Vc, suggesting that this The classification, diagnosis, and management of
 Trigeminal Mechanisms of Nociception: Peripheral and Brainstem Organization 449

chronic craniofacial pain remain difficult since the Sorensen, S., 1995). Among TMD (Isselee, H. et al.,
mechanisms for many of these conditions are not well 2002; LeResche, L. et al., 2003) and migraine patients
understood and animal models, though instructive, (Rasmussen, B. K., 1993; MacGregor, E. A. and
do not mimic the clinical state (Vos, B. P. et al., 1994; Hackshaw, A., 2004) pain severity and symptoms
Roveroni, R. C. et al., 2001). Considerable progress vary over the menstrual cycle suggesting a significant
has been in delineating the long-term changes in interaction with factors related to sex hormone status.
peripheral and central neural circuits that mediate Correspondingly, recent anatomical (Amandusson,
pain sensation following tissue injury (Treede, R. D. A. et al., 1996; Pajot, J. et al., 2003; Bereiter, D. A.
et al., 1992; Woolf, C. J. and Salter, M. W., 2000; Hunt, et al., 2005a; 2005b; Puri, V. et al., 2005) and electro-
S. P. and Mantyh, P. W., 2001; Julius, D. and physiological evidence (Okamoto, K. et al., 2003;
Basbaum, A. I., 2001; Lewin, G. R. et al., 2004). Flake, N. M. et al., 2005) from animal models support
Although similar cellular and molecular mechanisms the notion that estrogen preferentially enhances the
may contribute to chronic pain due to Vn damage excitability of trigeminal neurons that contribute to
(Lavigne, G. et al., 2005), many chronic craniofacial craniofacial pain. Furthermore, within the TSNC,
pain conditions such as TMD, migraine, or chronic estrogen receptor-positive neurons are found almost
daily headache and trigeminal neuralgia present with exclusively within the superficial laminae of Vc and
no overt signs of tissue injury. Indeed, it has long not in more rostral portions of the complex (Bereiter,
been appreciated that the correlation between tissue D. A. et al., 2005a) suggesting that this region plays a
injury and magnitude of pain sensation may be weak key role in differential processing of orofacial sensory
(Wall, P. D., 1979). This has lead to proposals of information under different sex hormone conditions.
emotional or neuropsychological (Tenenbaum, H. Second, spreading and referral of pain and sensiti-
C. et al., 2001; Korszun, A., 2002) and genetic factors zation evoked from outside the affected dermatomal
as significant determinants of some forms of chronic region are common features of many chronic craniofa-
craniofacial pain (TMD, Diatchenko, L. et al., 2005; cial pain conditions. Sensory disturbances occurring
migraine, Wessman, M. et al., 2004; trigeminal neur- outside the affected region have been well documented
algia, Duff, J. M., et al., 1999; dry eye in Sjogrens in clinical studies of TMD (Maixner, W. et al., 1998;
syndrome, Takei, M. et al., 2005). Chronic craniofa- Sarlani, E. and Greenspan, J. D., 2003; 2005), migraine
cial pain can be broadly classified according to the (Burstein, R. et al., 2000; Katsarava, Z., et al., 2002;
pattern and origin of pain episodes: chronic/recur- Goadsby, P. J., 2005), trigeminal neuralgia (Dubner,
rent (TMD, migraine headache, trigeminal R. et al., 1987; Nurmikko, T. J. and Eldridge, P. R.,
neuralgia), chronic/persistent (burning mouth, dry 2001; Devor, M. et al., 2002), and burning mouth syn-
eye syndromes), and chronic deafferentation pain drome (Svensson, P. et al., 1993; Ito, M. et al., 2002).
(postherpetic neuralgia, posttraumatic neuralgia, Patients with postherpetic neuralgia involving trigem-
phantom tooth). The mechanisms that underlie inal dermatomes had lower thermal warm and cool
these diverse conditions involve markedly different thresholds, while those with infection of spinal derma-
tissues and are likely quite heterogeneous. However, tomes had elevated thresholds (Pappagallo, M. et al.,
three features of many chronic craniofacial pain con- 2000) suggesting different underlying pathologies for
ditions are notable and provide evidence of neuropathic pain in trigeminal and spinal systems.
commonality. Mechanical allodynia and increased temporal summa-
First, the prevalence of most chronic craniofacial tion are consistent with the notion that central neural
pain conditions is higher in women than men (Dao, mechanisms maintain chronic craniofacial pain, while
T. T. and LeResche, L., 2000; Macfarlane, T. V. et al., peripheral mechanisms are required mainly for initia-
2002a). This is especially apparent for the chronic/ tion of the pain state.
recurrent conditions of TMD and migraine headache Third, many chronic craniofacial pain conditions
(LeResche, L., 1997; Rasmussen, B. K., 200l) and are accompanied by significant disturbances of the
somewhat less so for trigeminal neuralgia (Kitt, C. autonomic and/or endocrine systems. Chronic TMD
A. et al., 2000; Manzoni, G. C. and Torelli, P., 2005). patients display altered secretion of stress hormones
Women also are more likely to develop burning (Jones, D. A. et al., 1997; Korszun, A. et al., 2002) and
mouth (Bergdahl, M. and Bergdahl, J., 1999; elevated levels of neuropeptides and proinflamma-
Grushka, M. et al., 2003) and dry eye syndromes tory cytokines that could affect blood flow to joints
(Yazdani, C. et al., 2001) and report greater sensory (Kopp, S., 2001). The relationship between vascular
disturbances after Vn damage (Sandstedt, P. and reactivity and migraine has long been considered a
450 Trigeminal Mechanisms of Nociception: Peripheral and Brainstem Organization

critical variable (Janig, W., 2003; Goadsby, P. J., Allen, G. V., Barbrick, B., and Esser, M. J. 1996. Trigeminal-
parabrachial connections: possible pathway for nociception-
2005), whereas stimulus-evoked oral mucosal blood induced cardiovascular reflex responses. Brain Res.
flow is greater in patients with burning mouth syn- 715, 125135.
drome (Heckmann, S. M. et al., 2001). Evidence from Altschuler, S. M., Bao, X., Bieger, D., Hopkins, D. A., and
Miselis, R. R. 1989. Viscerotopic representation of the upper
animal models indicate that contributions from vagus alimentary tract in the rat: sensory ganglia and nuclei of the
nerve activity (Khasar, S. G. et al., 2001) and adrenal solitary and spinal trigeminal tracts. J. Comp. Neurol.
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Amandusson, A., Hermanson, O., and Blomqvist, A. 1996.
marked sex-related effects on cutaneous pain beha- Colocalization of oestrogen receptor immunoreactivity and
vior. Although treatments to reduce sympathetically preproenkephalin mRNA expression to neurons in the
maintained pain generally have a poor outcome for superficial laminae of the spinal and medullary dorsal horn of
rats. Eur. J. Neurosci. 8, 24402445.
posttraumatic trigeminal neuralgic patients (Gregg, J. Ambalavanar, R. and Morris, R. 1992. The distribution of binding
M., 1990), the influence of the autonomic nervous by isolectin I-B4 from Griffonia simplicifolia in the trigeminal
system on other forms chronic craniofacial pain has ganglion and brainstem trigeminal nuclei in the rat.
Neuroscience 47, 421429.
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 33 Migraine A Disorder Involving Trigeminal
Brainstem Mechanisms
P J Goadsby, University of California, San Francisco, CA, USA
2009 Elsevier Inc. All rights reserved.

33.1 Introduction 461

33.2 Migraine Explaining the Clinical Features 461
33.3 Genetics of Migraine 462
33.3.1 Genetic Epidemiology 462
33.3.2 Familial Hemiplegic Migraine 462
33.4 Migraine Aura 462
33.5 Headache Anatomy 463
33.5.1 The Trigeminal Innervation of Pain-Producing Intracranial Structures 463
33.6 Headache Physiology Peripheral Connections 463
33.6.1 Plasma Protein Extravasation 463
33.6.2 Sensitization and Migraine 463
33.6.3 Neuropeptide Studies 464
33.7 Headache Physiology Central Connections 464
33.7.1 The Trigeminocervical Complex 464
33.7.2 Higher-Order Processing 466
33.7.3.1 Thalamus 466
33.7.3.2 Activation of modulatory regions 466
33.8 Central Modulation of Trigeminal Pain 466
33.8.1 Brain Imaging in Humans 466
33.8.2 Animal Experimental Studies of Sensory Modulation 467
33.8.3 Electrophysiology of Migraine in Humans 467
33.9 What is Migraine? 467
References 468

33.1 Introduction 33.2 Migraine Explaining

the Clinical Features
Headache in general, and in particular migraine
(Goadsby, P. J. et al., 2002) and cluster headache Migraine is in essence a familial episodic disorder
(Goadsby, P. J., 2002), is better understood now than whose key marker is headache with certain associated
has been the case for the last four millennia (Lance, J. features (Table 1). It is these features that give clues
W. and Goadsby, P. J., 2005). Migraine is a common, to its pathophysiology, and ultimately will provide
disabling recurrent disorder of the central nervous insights leading to new treatments.
system with a core manifestation involving activation, The essential elements to be considered are:
or the perception of activation, of trigeminal nocicep- Genetics of migraine;
tive afferents (Olesen, J. et al., 2005). Here how studies Physiological basis for the aura;
of the anatomy and physiology of the pain-producing Anatomy of head pain, particularly that of the
innervation of the dura mater and large cranial ves- trigeminovascular system;
sels, the trigeminovascular system, has contributed to Physiology and pharmacology of activation of the
our current understanding of one of the most common peripheral branches of ophthalmic branch of the tri-
maladies of humans will be explored. geminal nerve;

461
462 Migraine A Disorder Involving Trigeminal Brainstem Mechanisms

Table 1 International Headache Society features of 33.3.2 Familial Hemiplegic Migraine

migraine (Headache Classification Committee of the
International Headache Society, 2004) In approximately 50% of the reported families,
familial hemiplegic migraine (FHM) has been
Repeated episodic headache (472 h) with the following
assigned to chromosome 19p13. Few clinical differ-
features:
Any two of: Any one of: ences have been found between chromosome 19-
unilateral nausea/vomiting linked and unlinked FHM families. Indeed, the clin-
throbbing photophobia and phonophobia ical phenotype does not associate particularly with
worsened by movement the known mutations. The most striking exception is
moderate or severe
cerebellar ataxia, which occurs in approximately
50% of the chromosome 19-linked, but in none of
the unlinked families. Another less striking difference
includes the fact that patients from chromosome 19-
Physiology and pharmacology of the trigeminal linked families are more likely to have attacks that
nucleus, in particular its caudal most part, the trige- can be triggered by minor head trauma or are that
minocervical complex (TCC); associated by coma.
Brainstem and diencephalic modulatory systems The biological basis for the linkage to chromo-
that influence trigeminal pain transmission and some 19 is mutations (Ophoff, R. A. et al., 1996)
other sensory modality processing. involving the Cav2.1 (P/Q) type voltage-gated cal-
cium channel CACNA1A gene. Now known as
FHM-I, this mutation is responsible for about
50% of identified families. Mutations in the
33.3 Genetics of Migraine ATP1A2 gene have been identified to be responsi-
ble for about 20% of FHM families. Interestingly,
One of the most important aspects of the pathophy- the phenotype of some FHM-II involves epilepsy,
siology of migraine is the inherited nature of the while it has also been suggested that alternating
disorder. It is clear from clinical practice that many hemiplegia of childhood can be due to ATP1A2
patients have first-degree relatives who also suffer mutations. The latter cases are most unusual for
from migraine (Lance, J. W. and Goadsby, P. J., migraine. Most recently mutations in the neuronal
2005). Transmission of migraine from parents to voltage-gated sodium channel SCN1A have been
children has been reported as early as the seven- identified as the cause of FHM-III, thus continuing
teenth century, and numerous published studies the ionopathic theme.
have reported a positive family history. Taken together, the known mutations suggest that
migraine, or at least the neurological manifestations
currently called the aura, are caused by an ionopathy.
Linking the channel disturbance for the first time to
33.3.1 Genetic Epidemiology the aura process has demonstrated that human muta-
Studies of twin pairs are the classical method to tions expressed in a knockin mouse produce a
investigate the relative importance of genetic and reduced threshold for cortical spreading depression
environmental factors. A Danish study included (CSD), which has some profound implications for
1013 monozygotic and 1667 dizygotic twin pairs of understanding that process.
the same gender, obtained from a population-based
twin register. The pairwise concordance rate was
significantly higher among monozygotic than dizy- 33.4 Migraine Aura
gotic twin pairs (P < 0.05). Several studies have
attempted to analyze the possible mode of inheri- Migraine aura is defined as a focal neurological
tance in migraine families and conflicting results disturbance manifest as visual, sensory, or motor
have been obtained. Both twin studies and popula- symptoms (Headache Classification Committee of
tion-based epidemiological surveys strongly suggest the International Headache Society, 2004). It is
that migraine without aura is a multifactorial disor- seen in about 30% of patients, and it is clearly
der, caused by a combination of genetic and neurally driven. The case for the aura being the
environmental factors. human equivalent of the CSD of Leao has been
 Migraine A Disorder Involving Trigeminal Brainstem Mechanisms 463

well made (Lauritzen, M., 1994). In humans, visual 33.6 Headache Physiology
aura has been described as affecting the visual Peripheral Connections
field, suggesting the visual cortex, and it starts at
33.6.1 Plasma Protein Extravasation
the center of the visual field and propagates to the
periphery at a speed of 3 mm min1. This is very Moskowitz M. A. (1990) has provided a series of
similar to spreading depression described in rabbits. experiments to suggest that the pain of migraine
Blood flow studies in patients have also shown may be a form of sterile neurogenic inflammation.
that a focal hyperemia tends to precede the spread- Although this seems clinically implausible, the
ing oligemia, and again this is similar to what model system has been helpful in understanding
would be expected with spreading depression. some aspects of trigeminovascular physiology.
After this passage of oligemia, the cerebrovascular Neurogenic plasma extravasation can be seen during
response to hypercapnia in patients is blunted electrical stimulation of the trigeminal ganglion in
while autoregulation remains intact. Again this pat- the rat. Plasma extravasation can be blocked by ergot
tern is repeated with experimental spreading alkaloids, indomethacin, acetylsalicylic acid, and the
depression. Human observations have rendered serotonin-5-HT1B/1D agonist, sumatriptan. The
the arguments reasonably sound that human aura pharmacology of abortive antimigraine drugs has
has as its equivalent in animals in CSD. An area of been reviewed in detail. In addition there are struc-
controversy surrounds whether aura triggers the tural changes in the dura mater that are observed
rest of the attack, and is indeed painful. Based on after trigeminal ganglion stimulation. These include
the available experimental and clinical data this mast cell degranulation and changes in postcapillary
author is not at all convinced that aura is painful venules including platelet aggregation. While it is
per se, but this does not diminish its interest or the generally accepted that such changes, and particu-
importance of understanding it. Indeed therapeutic larly the initiation of a sterile inflammatory response,
would cause pain, it is not clear whether this is
developments may shed further light on these rela-
sufficient of itself, or requires other stimulators, or
tionships, and studies are required to understand
promoters. Preclinical studies suggest that CSD may
how in some patients aura is clearly not a sufficient
be a sufficient stimulus to activate trigeminal neu-
trigger to pain.
rons, although this has been a controversial area.
Although plasma extravasation in the retina,
which is blocked by sumatriptan, can be seen after
trigeminal ganglion stimulation in experimental ani-
33.5 Headache Anatomy
mals, no changes are seen with retinal angiography
33.5.1 The Trigeminal Innervation of Pain- during acute attacks of migraine or cluster headache.
Producing Intracranial Structures A limitation of this study was the probable sampling
of both retina and choroids elements in rats, given
Surrounding the large cerebral vessels, pial vessels,
that choroidal vessels have fenestrated capillaries.
large venous sinuses, and dura mater is a plexus of
Clearly, however, blockade of neurogenic plasma
largely unmyelinated fibers that arise from the
protein extravasation is not completely predictive of
ophthalmic division of the trigeminal ganglion and
antimigraine efficacy in humans as evidenced by the
in the posterior fossa from the upper cervical dorsal
failure in clinical trials of substance P, neurokinin-1
roots. Trigeminal fibers innervating cerebral vessels antagonists, specific plasma protein extravasation
arise from neurons in the trigeminal ganglion that (PPE) blockers, CP122,288 and 4991w93, an
contain substance P and calcitonin gene-related pep- endothelin antagonist, and a neurosteroid. The
tide (CGRP), both of which can be released when the implications of these data have been recently
trigeminal ganglion is stimulated either in humans or reviewed (Peroutka, S. J., 2005).
cats (Goadsby, P. J. et al., 1988). Stimulation of the
cranial vessels, such as the superior sagittal sinus
(SSS), is certainly painful in humans (Wolff, H. G.,
1948). Human dural nerves that innervate the cranial 33.6.2 Sensitization and Migraine
vessels largely consist of small diameter myelinated While it is highly doubtful that there is a significant
and unmyelinated fibers that almost certainly sub- sterile inflammatory response in the dura mater during
serve a nociceptive function. migraine, it is clear that some form of sensitization takes
464 Migraine A Disorder Involving Trigeminal Brainstem Mechanisms

place during migraine, since allodynia is common. interest in this regard that compounds that have not
About two-thirds of patients complain of pain from shown activity in migraine (Peroutka, S. J., 2005),
non-noxious stimuli, allodynia (Selby, G. and Lance, J. notably the conformationally restricted analogue of
W., 1960). A particularly interesting aspect is the sumatriptan, CP122,288, and the conformationally
demonstration of allodynia in the upper limbs ipsilateral restricted analog of zolmitriptan, 4991w93, were
and contralateral to the pain. This finding is consistent both ineffective inhibitors of CGRP release after
with at least third-order neuronal sensitization, such as superior sagittal sinus in cats. The recent develop-
sensitization of thalamic neurons, and firmly places the ment of nonpeptide highly specific CGRP
pathophysiology within the central nervous system. antagonists, and the announcement of proof-of-
Sensitization in migraine may be peripheral with local concept for a CGRP antagonist in acute migraine
release of inflammatory markers, which would certainly (Olesen, J. et al., 2004), firmly establishes this as a
activate trigeminal nociceptors. More likely in migraine novel and important new emerging principle for
is a form of central sensitization, which may be classical acute migraine. At the same time the lack of any
central sensitization, or a form of disinhibitory sensitiza- effect of CGRP blockers on plasma protein extrava-
tion with dysfunction of descending modulatory sation, explains in some part why that model has
pathways (Knight, Y. E. et al., 2002). Just as dihydroer- proved inadequate at translation into human thera-
gotamine (DHE) can block trigeminovascular peutic approaches (Peroutka, S. J., 2005).
nociceptive transmission, probably at least by a local
effect in the TCC, DHE can block central sensitization
associated with dural stimulation by an inflammatory 33.7 Headache Physiology Central
soup, as can cyclo-oxygenase block both sensitization Connections
and trigeminocervical transmission.
33.7.1 The Trigeminocervical Complex
Fos immunohistochemistry is a method for looking at
33.6.3 Neuropeptide Studies
activated cells by plotting the expression of Fos pro-
Electrical stimulation of the trigeminal ganglion in tein. After meningeal irritation with blood Fos
both humans and cats leads to increases in extracer- expression is noted in the trigeminal nucleus cauda-
ebral blood flow and local release of both CGRP and lis, while after stimulation of the superior sagittal
SP (Goadsby, P. J. et al., 1988). In the cat trigeminal sinus Fos-like immunoreactivity is seen in the tri-
ganglion stimulation also increases cerebral blood geminal nucleus caudalis and in the dorsal horn at the
flow by a pathway traversing the greater superficial C1 and C2 levels in cats and monkey. These latter
petrosal branch of the facial nerve (Goadsby, P. J. and findings are in accord with similar data using
Duckworth, J. W., 1987) again releasing a powerful 2-deoxyglucose measurements with superior sagittal
vasodilator peptide, vasoactive intestinal polypeptide sinus stimulation. Similarly, stimulation of a branch
(VIP; May, A. and Goadsby, P. J., 1999). Interestingly, of C2, the greater occipital nerve, increases metabolic
the VIPergic innervation of the cerebral vessels is activity in the same regions, i.e., trigeminal nucleus
predominantly anterior rather than posterior, and caudalis and C1/2 dorsal horn, and fos expression can
this may contribute to this regions vulnerability to be elicited by injection of mustard oil into the occi-
spreading depression, explaining why the aura is so pital muscles. In experimental animals one can
very often seen to commence posteriorly. record directly from trigeminal neurons with both
Stimulation of the more specifically vascular pain- supratentorial trigeminal input and input from the
producing superior sagittal sinus increases cerebral greater occipital nerve, a branch of the C2 dorsal root
blood flow and jugular vein CGRP levels. Human (Bartsch, T. and Goadsby, P. J., 2002). Stimulation of
evidence that CGRP is elevated in the headache the greater occipital nerve for 5 min results in sub-
phase of migraine (Goadsby, P. J. et al., 1990), sup- stantial increases in responses to supratentorial dural
porting the view that the trigeminovascular system stimulation, which can last for over 1 h. Conversely,
may be activated in a protective role in these condi- stimulation of the middle meningeal artery dura
tions. Moreover, nitric oxide (NO)-donor-triggered mater with the C-fiber irritant mustard oil sensitizes
migraine, which is in essence typical migraine, also responses to occipital muscle stimulation. Taken
results in increases in CGRP that are blocked by together these data suggest convergence of cervical
sumatriptan, just as in spontaneous migraine and ophthalmic inputs at the level of the second-
(Goadsby, P. J. and Edvinsson, L., 1993). It is of order neuron. Moreover, stimulation of a lateralized
 Migraine A Disorder Involving Trigeminal Brainstem Mechanisms 465

structure, the middle meningeal artery, produces Fos second-order neurons that reduce cell activity and
expression bilaterally in both cat and monkey brains. suggest a further possible site for therapeutic interven-
This group of neurons from the superficial laminas of tion in migraine. This action can be dissected out to
trigeminal nucleus caudalis and C1/2 dorsal horns involve each of the 5-HT1B, 5-HT1D, and 5-HT1F
should be regarded functionally as the TCC. receptor subtypes, and are consistent with the localiza-
These data demonstrate that trigeminovascular tion of these receptors on peptidergic nociceptors.
nociceptive information comes by way of the most Interestingly, triptans also influence the CGRP promo-
caudal cells. This concept provides an anatomical ter, and regulate CGRP secretion from neurons in
explanation for the referral of pain to the back of the culture, as well as perhaps require cell surface expres-
head in migraine. Moreover, experimental pharmaco- sion for their effect. Furthermore, the demonstration
logical evidence suggests that some abortive that some part of this action is postsynaptic with either
antimigraine drugs, such as, ergot derivatives, acetyl- 5-HT1B or 5-HT1D receptors located nonpresynati-
salicylic acid, sumatriptan, eletriptan, naratriptan, cally offers a prospect of highly anatomically localized
rizatriptan, and zolmitriptan can have actions at these treatment options (Figure 1).

Ventroposteromedial
thalamus
Posterior hypothalamus

Periaqueductal gray matter

Dural vasculature (PAG) PAG

Locus coeruleus
(LC)
LC
Trigeminal ganglion

Cervical
muscle
and
joints Cervical dorsal root ganglion Trigeminocervical
complex

Midline

Figure 1 Illustration of the some elements of migraine biology. Patients inherit a dysfunction in brain control systems for
pain and other afferent stimuli, which can be triggered and are in turn capable of activating the trigeminovascular system as
the initiating event in a positive feedback of neurally driven vasodilatation. Nociceptive afferents from the cervical region
terminate in the trigeminocervical complex (illustrated by Fos protein expression in the superficial laminas) and this accounts
for the nontrigeminal distribution of pain in many patients. These afferents project to the thalamus, including
ventroposteromedial thalamus, and are at least influenced by neurons in the posterior hypothalamic gray, the periaqueductal
gray (PAG), and probably by neurons of the nucleus locus coeruleus in the pons. Functional brain imaging suggests that the
brainstem, notably the pons as illustrated after Bahra, A., Matharu, M. S., Buchel, C., Frackowiak, R. S. J., and Goadsby, P. J.
2001. Brainstem activation specific to migraine headache. Lancet 357, 10161017, is a pivotal region in the migraine process.
466 Migraine A Disorder Involving Trigeminal Brainstem Mechanisms

Table 2 Neuroanatomical processing of vascular head pain

Structure Comments

Target innervation: Ophthalmic branch of trigeminal nerve

Cranial vessels
Dura mater

First Trigeminal ganglion Middle cranial fossa

Second Trigeminal nucleus (quintothalamic tract) Trigeminal nucleus caudalis and C1/C2
dorsal horns
Third Thalamus Ventrobasal complex
Medial nucleus of posterior group
Intralaminar complex
Modulatory Midbrain Periaqueductal gray matter
Hypothalamus ?
Final Cortex Insulas
Frontal cortex
Anterior cingulate cortex
Basal ganglia

33.7.2 Higher-Order Processing the ventrolateral periaqueductal gray matter (PAG).

PAG activation in turn feeds back to the TCC with
Following transmission in the caudal brainstem and
an inhibitory influence. PAG is clearly included in
high cervical spinal cord information is relayed ros-
the area of activation seen in positron emission tomo-
trally (Table 2).
graphy (PET) studies in migraineurs. This typical
negative feedback system will be further considered
33.7.3.1 Thalamus below as a possible mechanism for the symptomatic
Processing of vascular nociceptive signals in the tha- manifestations of migraine.
lamus occurs in the ventroposteromedial (VPM) Another potentially modulatory region activated
thalamus, medial nucleus of the posterior complex, by stimulation of nociceptive trigeminovascular
and in the intralaminar thalamus. It has been shown input is the posterior hypothalamic gray. This area
by application of capsaicin to the superior sagittal is crucially involved in several primary headaches,
sinus that trigeminal projections with a high degree notably cluster headache (Goadsby, P. J., 2002),
of nociceptive input are processed in neurons parti- Short-lasting unilateral neuralgiform headache
cularly in the ventroposteromedial thalamus and in attacks with conjunctival injection and tearing
its ventral periphery. These neurons in the VPM can (SUNCT), paroxysmal hemicrania and hemicrania
be modulated by activation of gamma-aminobutyric continua. Moreover, the clinical features of the pre-
acid (GABA)A inhibitory receptors, and perhaps of monitory phase, and other features of the disorder,
more direct clinical relevance by propranolol though suggest dopamine neuron involvement. Orexinergic
a 1-adrenoceptor mechanism (Shields, K. G. and neurons in the posterior hypothalamus can be both
Goadsby, P. J., 2005). Remarkably, triptans through pro- and antinociceptive, offering a further possible
5-HT1B/1D mechanisms can also inhibit VPM neu- region whose dysfunction might involve the percep-
rons locally, as demonstrated by microiontophoretic tion of head pain.
application, suggesting a hitherto unconsidered locus
of action for triptans in acute migraine. Human ima-
ging studies have confirmed activation of thalamus 33.8 Central Modulation of
contralateral to pain in acute migraine (Bahra, A. Trigeminal Pain
et al., 2001). 33.8.1 Brain Imaging in Humans
Functional brain imaging with PET has demon-
33.7.3.2 Activation of modulatory regions strated activation of the dorsal midbrain, including
Stimulation of nociceptive afferents by stimulation of the PAG, and in the dorsal pons, near the locus
the superior sagittal sinus in cats activates neurons in coeruleus, in studies during migraine without aura.
 Migraine A Disorder Involving Trigeminal Brainstem Mechanisms 467

Dorsolateral pontine activation is seen with PET in compared to controls. Changes in CNV predict
spontaneous episodic and chronic migraine, and with attacks and preventive therapies alter, normalize,
nitrogylcerin-triggered attacks (Bahra, A. et al., 2001; such changes. Attempts to correlate clinical pheno-
Afridi, S. et al., 2005). These areas are active imme- types with electrophysiological changes, may
diately after successful treatment of the headache but enhance further studies in this area.
are not active interictally. The activation corre-
sponds with the brain region that Raskin initially
reported, and confirmed, to cause migrainelike head-
ache when stimulated in patients with electrodes 33.9 What is Migraine?
implanted for pain control. Similarly, excess iron in
the PAG of patients with episodic and chronic Migraine is an inherited, episodic disorder invol-
migraine, and chronic migraine can develop after a ving sensory sensitivity. Patients complain of pain
bleed into a cavernoma in the region of the PAG, or in the head that is throbbing, but there is no
with a lesion of the pons. What could dysfunction of reliable relationship between vessel diameter and
these brain areas lead to? the pain, or its treatment. They complain of dis-
comfort from normal lights and the unpleasantness
of routine sounds. Some mention otherwise plea-
33.8.2 Animal Experimental Studies of
sant odors are unpleasant. The anatomical
Sensory Modulation
connections of, for example, the pain pathways
It has been shown in experimental animals that sti- are clear, the ophthalmic division of the trigeminal
mulation of nucleus locus coeruleus, the main central nerve subserves sensation within the cranium and
noradrenergic nucleus, reduces cerebral blood flow explains why the top of the head is headache, and
in a frequency-dependent manner (Goadsby, P. J. the maxillary division is facial pain. The conver-
et al., 1982) through an 2-adrenoceptor-linked gence of cervical and trigeminal afferents explains
mechanism. This reduction is maximal in the occipi- why neck stiffness or pain is so common in pri-
tal cortex. While a 25% overall reduction in cerebral mary headache. The genetics of channelopathies is
blood flow is seen, extracerebral vasodilatation opening up a plausible way to think about the
occurs in parallel (Goadsby, P. J. et al., 1982). episodic nature of migraine. However, where is
In addition, the main serotonin-containing nucleus the lesion, what is actually the pathology?
in the brainstem, the midbrain dorsal raphe nucleus, Migraine aura cannot be the trigger alone, there
can increase cerebral blood flow when activated. is no evidence at all after 4000 years that it occurs
Furthermore, stimulation of PAG will inhibit sagittal in more than 30% of migraine patients; aura can
sinus-evoked trigeminal neuronal activity in cats, be experienced without pain at all, and is seen in
while blockade of P/Q-type voltage-gated Ca2 the other primary headaches. Perhaps electrophy-
channels in the PAG facilitates trigeminovascular siological changes in the brain have been
nociceptive processing (Knight, Y. E. et al., 2002) mislabeled as hyperexcitability whereas dyshabitua-
with the local GABAergic system in the PAG still tion might be a simpler explanation. If migraine
intact. was basically an attentional problem with changes
in cortical synchronization (Niebur, E. et al., 2002),
hypersynchronization, all its manifestations could
33.8.3 Electrophysiology of Migraine in
be accounted for in a single over-arching patho-
Humans
physiological hypothesis of a disturbance of
Studies of evoked potentials and event-related subcortical sensory modulation systems. While it
potentials provide some link between animal studies seems likely that the trigeminovascular system,
and human functional imaging. Authors have shown and its cranial autonomic reflex connections, the
changes in neurophysiological measures of brain acti- trigeminal-autonomic reflex (May, A. and Goadsby,
vation but there is much discussion as to how to P. J., 1999), act as a feed-forward system to facil-
interpret such changes (Schoenen, J. et al., 2003). itate the acute attack, the fundamental problem in
Perhaps the most reliable theme is that the migrai- migraine is in the brain. Unraveling its basis will
nous brain does not habituate to signals in a normal deliver great benefits to patients and considerable
way. Similarly, contingent negative variation (CNV), understanding of some very fundamental neurobio-
an event related potential, is abnormal in migraineurs logical processes.
468 Migraine A Disorder Involving Trigeminal Brainstem Mechanisms

Acknowledgment Knight, Y. E., Bartsch, T., Kaube, H., and Goadsby, P. J. 2002.
P/Q-type calcium channel blockade in the PAG facilitates
trigeminal nociception: a functional genetic link for migraine?
The work of the author has been supported by the J. Neurosci. 22, 16.
Migrane Trust. Lance, J. W. and Goadsby, P. J. 2005. Mechanism and
Management of Headache. Elsevier.
Lauritzen, M. 1994. Pathophysiology of the migraine aura. The
spreading depression theory. Brain 117, 199210.
References May, A. and Goadsby, P. J. 1999. The trigeminovascular system
in humans: pathophysiological implications for primary
headache syndromes of the neural influences on the cerebral
Afridi, S., Matharu, M. S., Lee, L., Kaube, H., Friston, K. J., circulation. J. Cerebr. Blood Flow Metabol. 19, 115127.
Frackowiak, R. S. J., and Goadsby, P. J. 2005. A PET study Moskowitz, M. A. 1990. Basic mechanisms in vascular
exploring the laterality of brainstem activation in migraine headache. Neurolog. Clin. 8, 801815.
using glyceryl trinitrate. Brain 128, 932939. Niebur, E., Hsiao, S. S., and Johnson, K. O. 2002. Synchrony:
Bahra, A., Matharu, M. S., Buchel, C., Frackowiak, R. S. J., and a neural mechanism for attentional selection? Curr. Opin.
Goadsby, P. J. 2001. Brainstem activation specific to Neurobiol. 12, 190194.
migraine headache. Lancet 357, 10161017. Olesen, J., Diener, H. C., Husstedt, I. W., Goadsby, P. J.,
Bartsch, T. and Goadsby, P. J. 2002. Stimulation of the greater Hall, D., Meier, U., Pollentier, S., and Lesko, L. M. 2004.
occipital nerve induces increased central excitability of dural Calcitonin gene-related peptide (CGRP) receptor antagonist
afferent input. Brain 125, 14961509. BIBN4096BS is effective in the treatment of migraine
Goadsby, P. J. 2002. Pathophysiology of cluster headache: a attacks. New Engl. J. Med. 350, 11041110.
trigeminal autonomic cephalgia. Lancet Neurol. 1, 3743. Olesen, J., Tfelt-Hansen, P., Ramadan, N., Goadsby, P. J., and
Goadsby, P. J. and Duckworth, J. W. 1987. Effect of stimulation Welch, K. M. A. 2005. The Headaches. Lippincott, Williams &
of trigeminal ganglion on regional cerebral blood flow in cats. Wilkins.
Am. J. Physiol. 253, R270R274. Ophoff, R. A., Terwindt, G. M., Vergouwe, M. N., van Eijk, R.,
Goadsby, P. J. and Edvinsson, L. 1993. The trigeminovascular Oefner, P. J., Hoffman, S. M. G., Lamerdin, J. E,,
system and migraine: studies characterizing Mohrenweiser, H. W., Bulman, D. E., Ferrari, M., Haan, J.,
cerebrovascular and neuropeptide changes seen in humans Lindhout, D., van Ommen, G. J., Hofker, M. H., Ferrari, M. D.,
and cats. Ann. Neurol. 33, 4856. and Frants, R. R. 1996. Familial hemiplegic migraine and
Goadsby, P. J., Edvinsson, L., and Ekman, R. 1988. Release of episodic ataxia type-2 are caused by mutations in the Ca2
vasoactive peptides in the extracerebral circulation of man channel gene CACNL1A4. Cell 87, 543552.
and the cat during activation of the trigeminovascular Peroutka, S. J. 2005. Neurogenic inflammation and migraine:
system. Ann. Neurol. 23, 193196. implications for therapeutics. Mol. Intervent. 5, 306313.
Goadsby, P. J., Edvinsson, L., and Ekman, R. 1990. Vasoactive Schoenen, J., Ambrosini, A., Sandor, P. S., and Maertens de
peptide release in the extracerebral circulation of humans Noordhout, A. 2003. Evoked potentials and transcranial
during migraine headache. Ann. Neurol. 28, 183187. magnetic stimulation in migraine: published data and
Goadsby, P. J., Lambert, G. A., and Lance, J. W. 1982. viewpoint on their pathophysiologic significance. Clin.
Differential effects on the internal and external carotid Neurophysiol. 114, 955972.
circulation of the monkey evoked by locus coeruleus Selby, G. and Lance, J. W. 1960. Observations on 500 cases of
stimulation. Brain Res. 249, 247254. migraine and allied vascular headache. J. Neurol. Neurosurg.
Goadsby, P. J., Lipton, R. B., and Ferrari, M. D. 2002. Migraine Psychiatry 23, 2332.
current understanding and treatment. New Engl. J. Med. Shields, K. G. and Goadsby, P. J. 2005. Propranolol modulates
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1160. University Press.
 34 Tooth Pain
M R Byers, University of Washington, Seattle WA, USA
2009 Elsevier Inc. All rights reserved.

34.1 Introduction 470

34.2 Dental Sensory Mechanisms 470
34.2.1 Normal Teeth/Acute Pain 470
34.2.2 Inflammatory Tooth Pain 472
34.2.3 Dental Neuropathic Pain 472
34.3 Tooth Pain: Diagnosis and Management 473
34.4 Conclusions 474
References 474

Glossary
ASIC receptors Acid-sensing ion channels that odontalgia Tooth pain.
detect low pH, a typical condition of pulpitis. odontoblast Neural crest-derived cells that
atypical odontalgia A condition in which tooth make dentin and regulate the pulpdentin
pain derives from neuropathic or referred mechan- barrier.
isms. Tooth extractions or root canals do not relieve P2 receptors Purinergic nucleotide receptors that
the pain. respond to adenosine triphosphate (ATP).
BK receptors Activated by the inflammatory periapex Region at base of root socket that sur-
mediator, bradykinin. rounds the root apex, and includes periodontal
convergence Many sensory afferents from multi- ligament, neurovascular bundles and endings, and
ple tissues project to individual central trigeminal alveolar bone.
neurons. prepain The first sensation (tingling, vibration, or
dentin Specialized, calcified, collagenous matrix touch) elicited by electrical stimulation of teeth. It
that surrounds the pulp. changes to sharp tooth pain at stronger levels of
hot tooth An inflamed tooth that resists regional stimulation.
anesthesia and remains sensitive when neighbor- pulpitis Inflamed tooth pulp that can be healed
ing teeth are numb. locally (reversible), that can consume the pulp
hypersensitive dentin Sharp pain elicited from and spread into periapex (irreversible), or that
light touch to exposed dentin. can be undetected (silent) until it reaches the
ionotropic receptors Activated by specific periapex.
ligands causing ion flux through receptor pores. receptive field Patch of tissue that activates a
mesencephalic trigeminal nucleus Location of primary afferent or central neuron.
cell bodies of primary sensory neurons that inner- referred pain Pain that is felt at a different site
vate stretch receptors in periodontal ligament, from the neural activity that causes it.
sutures, or mastication muscles. Ruffini mechanoreceptors Complex stretch
metabotropic receptors Interaction with ligand mechanoreceptors located in periodontal
activates G-protein intracellular signaling. ligament.
NK receptors Receptors for neurokinins such as Trk receptors Tyrosine kinase receptors that
substance P or neurokinin A. respond to neurotrophin factors.
nucleus caudalis Caudal region of spinal trigem- TRP receptors Transient receptor potential chan-
inal subnuclei, specialized for processing, relaying nels (vanilloid receptor family) that are activated by
and modulating orofacial pain. capsaicin, heat, or low pH.

469
470 Tooth Pain

34.1 Introduction 2002; Hu, J. W., 2004; Lavigne, G. et al., 2004;

Truelove, E., 2004; Sessle, B. J., 2005; Henry, M. A.
Tooth pain is the most common type of orofacial and Hargreaves, K. M., 2007).
pain (Lipton, J. et al., 1993; Hargreaves, K. M., 2002).
Dental nerve fibers branch centrally to activate many
neurons in the trigeminal brainstem complex or
34.2 Dental Sensory Mechanisms
extratrigeminal relay sites, and those central neurons
also receive extensive convergent input from other 34.2.1 Normal Teeth/Acute Pain
orofacial tissues, making location of tooth pain diffi-
A-fibers respond especially well to acute stimuli that
cult. However, most of the time our teeth do not hurt,
move fluid in dentin (Brannstrom, M and Astrom, A.,
and most dental neural activity is unperceived. The 1972) and C-fibers respond to acute heat stimuli or
sensory functions of tooth nerves are presented here, pulp damage (Narhi, M. V. O. et al., 1996; Narhi, M.
along with unusual features of central processing, V. O., 2005). Different factors affect axonal conduc-
mechanisms, perception, diagnosis, and treatment of tion in trigeminal nerves, ganglion, central tracts, and
tooth pain, which can be sharp or dull, focused or synaptic termination regions in the brainstem
diffuse, episodic or relentless, referred or neuropathic (Figure 1). Dental afferents project to low threshold
(for reviews see: Narhi, M. V. O. et al., 1996; Olgart, mechanoreceptive, nociceptive-specific, and wide-
L., 1996; Byers, M. R. and Narhi, M. V. O., 1999; dynamic-range central neurons, all of which receive
Dionne, R. A. and Berthold, C. W., 2001; Byers, M. a major input from other tissues, often from more
R. and Narhi, M. V. O., 2002; Hargreaves, K. M., than one trigeminal division (Sessle, B. J. et al., 1986).

Figure 1 Interactions affecting dental neuronal function. ATP, adenosine triphosphate; LTM, low-threshold mechanoreceptor;
NS, nociceptive specific; WDR, wide-dynamic neuron. Target Neurons: reprinted from Pain, 27, Sessele, B. J., Hu J. W., Amano,
N., and Zhong, G., Convergence of cutaneous, tooth pulp, visceral, neck and muscle afferents onto nociceptive and non-
nociceptive neurons in trigeminal subnucleus caudalis (medullary dorsal horn) and its implications for referred pain, 219235,
Copyright 1986, with permission from The International Association for the Study of Pain.
 Tooth Pain 471

Most tooth pain perceptions are acute, and they neuropeptides, neurotrophins, and opioid peptides
derive from activation of neurons in nucleus caudalis, that sensitize, inhibit, or modulate sensory neurons
although rostral trigeminal nuclei are also involved, (Hu, J. W., 2004), and even immune regulators
and there are other connection sites such as the (Wadachi, R. and Hargreaves, K. M., 2006). The regula-
paratrigeminal nucleus and reticular formation tion of the pulpal milieu and the quality of tooth pain
(Sessle, B. J., 2000; 2005). vary in relation to those activating and modulating
Human studies show three kinds of evoked sensa- systems.
tion from dental nerves: prepain, sharp pain, and dull Many dental nerve endings form close appositions
ache. The first two depend on activation of fast A- with the odontoblasts (Figures 2(a) and 2(b)), while
and A--fibers and the latter involves polymodal cap- others end freely in pulp and dentin. Neuro-odonto-
saicin-sensitive slow A- and C-fibers (Narhi, M. V. O. blast interactions are not fully understood, and may
et al., 1996; Ikeda, H. et al., 1997), some of which express involve odontoblast support for the free sensory end-
neuropeptide receptors for autocrine modulation ings, modulation of sensory activity, and/or specific
(Suzuki, H. et al., 2002). Each of our teeth is innervated sensory activity. The lack of synaptic contacts or gap
by many hundreds of highly branched trigeminal neu-
rons, and the density of sensory nerve endings in 0 10 20 30 40 50
coronal pulp and inner dentin is enormous (Figure 2).
However, most intradental neural activity involves a
unperceived neural efferent functions or reflexes,
such as vasodilatation by neuropeptides from sensory
b
fibers that is counterbalanced by sympathetic-mediated
vasoconstriction (Figure 3; Olgart, L. 1996; Fristad, I.
et al., 1997; Berggreen, E. and Heyeraas, K. J., 2000). c
Touch sensations during chewing come from Ruffini
mechanoreceptors in the periodontal ligament outside
Control Sympathetic block or cut
the roots, while unconscious aspects of jaw reflexes
involve intradental mechanoreceptive A-fibers and Figure 3 Laser Doppler demonstration of sensory nerve-
the periodontal endings of mesencephalic trigeminal mediated increased blood flow in rat incisor pulp after
-adrenergic block (b) or sympathectomy (c) compared to
neurons (Dong, W. K. et al., 1993). Dental neurons can intact blood flow (a). All teeth received brief bipolar electrical
express a variety of ionotropic receptors (e.g., TRP-V1, stimulation of the intact tooth crown. Reproduced from
TRP-V2, P2X3, ASIC), metabotropic receptors (BK-1, Olgart, L. and Kerezoudis, N. P. 1994. Nervepulp
BK-2, NK1-3, TrkA), ion channels, receptors for interactions. Arch. Oral Biol. 39, 47S54S, with permission.

(a) (b) (c)

OD
OD

N
D RD
OD OD
A

A
P
50 m 0.5 m 10 m

Figure 2 (a) Sensory endings in pulp (P) and dentin (D) (thin arrows) are shown by autoradiography of axonally transported
3
H-protein in adult rat molars. They avoid reparative dentin (RD) but come close to surface of tooth (wide arrow). Reprinted
from Byers, M.R. 1984. Dental sensory receptors. Int. Rev. Neurobiol. 25, 3994. (b) Electron microscopic autoradiography
showed that transported 3H-proteins (black coiled silver grain) are confined to sensory endings (N). Odontoblasts (OD) are
connected by numerous gap junctions (arrowheads). A special apposition separates OD and N (white arrow). Reproduced
from Byers, M. R. 1977. Fine structure of trigeminal receptors in rat molars. In: Pain in the Trigeminal Region (eds. D. J.
Anderson and B. Matthews), pp. 1324, with permission from Elsevier. (c) Mouse molar root nerves include A- fibers that
exceed 6 mm in diameter and several sizes of A- axons.
472 Tooth Pain

junctions between odontoblasts and nerves suggests a (a) Rt

supportive role. However, demonstrations of neural-
like ion channels (Guo, L. and Davidson, R. M., 1998)
and TREK-1, a mechanosensitive potassium channel,
in odontoblasts (Magloire, H. et al., 2003) show that
they are excitable and mechanosensitive. They also
attract sensory nerves (Maurin, J. C. et al., 2004) and
express neurotrophin factors and receptors in devel-
oping, adult, and injured teeth (Fried, K. et al., 2000;
Woodnutt, D. A. et al., 2000). It is still not clear 0.1 mm
whether odontoblast excitability directly affects
(b) (c)
tooth pain or just allows those cells to monitor denti- Rt
nogenic requirements.

34.2.2 Inflammatory Tooth Pain

When pulp or periapex are inflamed, there are local
cellular changes, nerve sprouting, peripheral and 0.1 mm 50 m
central sensitization, and neurochemical plasticity
that alter the quality of tooth pain perceptions Figure 4 (a, b) Several weeks after tooth injury the lesion
has destroyed the pulp and emerged from the root (Rt) into
(Hargreaves, K. M., 2002; Hu, J. W., 2004). Many of periapical tissue, where there is intense sprouting and
the events in inflamed teeth are typical of any inflam- immunoreactivity for calcitonin gene-related peptide
mation, but there are unusual nerve-sprouting (CGRP) nerve fibers compared to a normal molar (b). Arrows
reactions near the pulpitis, and those either subside indicate sensory nerve in alveolar canal. (c) Satellite cells
if healing occurs, or they persist for months or years surrounding trigeminal cell bodies are shown by
immunocytochemistry to express glial fibrillary acidic
when lesions escape into the periapical region protein (black rings) after molar injury in adult rats. (a, b)
(Figures 4(a) and 4(b); Byers, M. R. and Narhi, M. V. Reproduced from Kimberly, C. L. and Byers, M. R. 1988.
O., 1999). Important changes occur in dental neuronal Inflammation of rat molar pulp and periodontium causes
satellite cell glia (Figure 4(c)) when inflammation increased calcitonin gene-related peptide and axonal
continues in teeth, such as expression of glial fibrillary sprouting. Anat. Rec. 222, 289300, with permission.
acidic protein (Stephenson, J. L. and Byers, M. R., 1995).
Specialized vascular reactions also occur (Olgart, L.,
1996). Recording from individual fibers shows that
there are expansions of receptive fields of the A-fiber (a) Controls (b) Inflamed 1 week
afferents after 12 weeks of inflammation (Figure 5),
that would further affect sensitization in the central
neurons.

34.2.3 Dental Neuropathic Pain

Teeth are major players in referred orofacial pain,
either as the source or the referral site, and that
situation can lead to unnecessary multiple extrac-
tions. Teeth can also exhibit neuropathic symptoms
such as hyperalgesia, allodynia, and spontaneous pain
(Truelove, E., 2004; Lavigne, G. et al., 2004). A prin- Figure 5 Receptive fields (black patches) for individual
cipal mechanism for atypical odontalgia or dental A-fibers were larger in dog teeth after 1 week of induced
neuropathic pain is the extensive convergence of pulpitis compared to control teeth. Reproduced from Narhi,
M. V. O., Yamamoto, H., and Ngassapa, D. 1996. Function
inputs onto brainstem neurons from a wide area,
of Intradental Nociceptors in Normal and Inflamed Teeth. In:
often involving multiple trigeminal divisions and a Dentin/Pulp Complex (eds. M. Shimono, T. Maeda, H. Suda,
variety of tissues (Sessle, B. J., 2000; 2005), and glial and K. Takahashi), pp. 136140. Quintessence Publishing
actions also affect pain quality (Xie, Y. F. et al., 2006). Co, Inc., with permission.
 Tooth Pain 473

34.3 Tooth Pain: Diagnosis and 2001). However, pain can persist weeks, months, or
Management years, especially in individuals who have had long-
term tooth pain or other chronic pain (Bender, I. B.,
Dentists routinely use evoked acute pain to diagnose 2000; Truelove, E., 2004). Tooth pain can expand to a
tooth pathology, pulp vitality, and treatment (Bender, wide area, making it difficult to locate the pathology.
I. B., 2000). Dental procedures such as orthodontia or Competition from a second pain source can narrow
root canal treatment cause transient pain that usually the site to just one tooth, at least for a few minutes
decreases within a few days, as the local inflammation to aid diagnosis (Figure 6). This diffuse noxious
subsides, especially with nonsteroidal anti-inflamma- inhibition implies substantial changes in central
tory treatment (Dionne, R. A. and Berthold, C. W., physiology for the widely dispersed tooth pains

Before the tourniquet At ischemic tolerance 5 min past ischemic

procedure tolerance point

50
Individual responses
Mean response (n = 10) Very intolerable

40 * p < 0.05
Relative magnitude

Intolerable
30

20
Very distressing
(SEM)

Very annoying
10 Distressing

Annoying

Slightly annoying
0

Before tourniquet 5 min past IPT

At ischemic pain tolerance

Figure 6 Prior to tourniquet-evoked arm pain, the area of dental pain was large. After maximal arm pain, the tooth pain area
was reduced to an individual tooth. By 5 min later, the pain was still focused although the pain intensity (red lines) was
returning to prestimulus levels. IPT, ischemic pain tolerance. Reprinted from Pain, 57, Sigurdsson, A. and Maxiner, W., Effects
of experimental and clinical noxious counterirritants on pain perception, 265275, Copyright 1994, with permission from The
International Association for the Study of Pain.
474 Tooth Pain

(Sessle, B. J., 2005), though important changes in Dionne, R. A. and Berthold, C. W. 2001. Therapeutic uses of
non-steroidal anti-inflammatory drugs in dentistry. Crit. Rev.
neural sodium channels occur in painful teeth Oral Biol. Med. 12, 315330.
(Renton, T. et al., 2005; Henry, M. A. and Dong, W. K., Shiwaku, T., Kawakami, Y., and Chudler, E. H.
Hargreaves, K. M., 2007) offering possible pharma- 1993. Static and dynamic responses of periodontal ligament
mechanoreceptors and intradental mechanoreceptors. J.
cologic therapeutic targets. Neurophysiol. 69, 15671582.
Fried, K., Nosrat, C., Lillesaar, C., and Hildebrand, C. 2000.
Molecular signaling and pulpal nerve development. Crit. Rev.
Oral Biol. Med. 11, 318332.
Fristad, I., Kvinnsland, I. H., Johnsson, R., and Heyeraas, K. J.
34.4 Conclusions 1997. Effect of intermittent long-lasting electrical tooth
stimulation on pulpal blood flow and immunocompetent
Unusual features of tooth pain include: (1) difficulty cells: a hemodynamic and immunohistochemical study in
young rat molars. Exp. Neurol. 146, 230239.
locating the source of pain; (2) intense pain from Guo, L. and Davidson, R. M. 1998. Potassium and chloride
stimulation of dentin; (3) hypersensitive dentin after channels in freshly isolated odontoblasts. J. Dent. Res.
loss of its protective enamel or periodontal covering, 77, 341350.
Hargreaves, K. M. 2002. Pain Mechanisms of the Pulpodentin
with concomitant pulpitis; (4) special neuro-pulpal Complex. In: Seltzer and Benders Dental Pulp
interactions; (5) silent pulpal inflammation that only (eds. K. M. Hargreaves and H. E. Goodis), pp. 181203.
becomes painful when it invades periapical tissues; (6) Quintessence.
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perhaps, surprising that prolonged tooth pain is unu- pp. 141162. Quintessence.
sual, given the huge number of dental procedures Ikeda, H., Tokita, Y., and Suda, H. 1997. Capsaicin-sensitive A-
every day that can cause tooth, nerve, or bone damage delta fibers in cat tooth pulp. J. Dent. Res. 76, 13411349.
Kimberly, C. L. and Byers, M. R. 1988. Inflammation of rat molar
(i.e., dental implants that screw into the jaw, third pulp and periodontium causes increased calcitonin gene-
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prevalence and distribution of reported orofacial pain in the
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2005; Henry, M. A. and Hargreaves, K. M., 2007). Bleicher, F. 2003. Expression and localization of TREK-1
K channels in human odontoblasts. J. Dent. Res.
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Maurin, J. C., Couble, M. L., Didier-Bazes, M., Brisson, C.,
Magloire, H., and Bleicher, F. 2004. Expression and
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Bender, I. B. 2000. Pulp pain diagnosis a review. J. Endodon. Function of Intradental Nociceptors in Normal and
26, 175179. Inflamed Teeth. In: Dentin/Pulp Complex (eds. M. Shimono,
Berggreen, E. and Heyeraas, K. J. 2000. Effect of the sensory T. Maeda, H. Suda, and K. Takahashi), pp. 136140.
neuropeptide antagonists h-CGRP (8-37) and SR 140.33 on Quintessence.
pulpal and gingival blood flow in ferrets. Arch. Oral Biol. Narhi, M. V. O. 2006. Nociceptors in the Dental Pulp. In: Pain
45, 537542. Encyclopedia (eds. R. F. Schmidt and W. D. Willis), Springer.
Brannstrom, M and Astrom, A. 1972. The hydrodynamics of Olgart, L. 1996. Neural control of pulpal blood flow. Crit. Rev.
dentine: its possible relationship to dentinal pain. Int. Dent. J. Oral Biol. Med. 7, 159171.
22, 219227. Olgart, L. and Kerezoudis, N. P. 1994. Nervepulp interactions.
Byers, M. R. 1978. Fine Structure of Trigeminal Receptors in Rat Arch. Oral Biol. 39, 47S54S.
Molars. In: Pain in the Trigeminal Region (eds. D. J. Anderson Renton, T., Yiangou, Y., Plumpton, C., Tate, S., Bountra, C., and
and B. Matthews), pp. 1324, Elsevier. Anand, P. 2005. Sodium channel Nav1.8 immunoreactivity in
Byers, M. R. and Narhi, M. V. O. 1999. Dental injury models: painful human dental pulp. BMC Oral Health 5, 5.
experimental tools for understanding neuro-inflammatory Sessle, B. J. 2000. Acute and chronic craniofacial pain:
interactions and polymodal nociceptor functions. Crit. Rev. brainstem mechanisms of nociceptive transmission and
Oral Biol. Med. 10, 439. neuroplasticity, and their clinical correlates. Crit. Rev. Oral
Byers, M. R. and Narhi, M. V. O. 2002. Nerve Supply of the Biol. Med. 11, 5791.
Pulpodentin Complex and Responses to Injury. In: Seltzer Sessle, B. J. 2005. Orofacial Pain. In: The Paths of Pain
and Benders Dental Pulp (eds. K. M. Hargreaves and (eds. H. Merskey, J. D. Lowser, and R. Dubner), pp. 131150.
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Sessle, B. J., Hu, J. W., Amano, N., and Zhong, G. 1986. Truelove, E. 2004. Management issues of neuropathic
Convergence of cutaneous, tooth pulp, visceral, neck and trigeminal pain from a dental perspective. J. Orofac. Pain
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Stephenson, J. L. and Byers, M. R. 1995. GFAP pain due to infection. J. Dent. Res. 85, 4953.
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Sigurdsson, A. and Maixner, W. 1994. Effects of experimental growth factor are differentially expressed in adjacent
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 35 Ascending Pathways: Anatomy and Physiology
D Lima, Universidade do Porto, Porto, Portugal
2009 Elsevier Inc. All rights reserved.

35.1 Introduction 479

35.1.1 Defining Nociceptive Ascending Pathways 479
35.1.2 The Spinothalamic System 480
35.2 Spinocervical Pathway 481
35.2.1 Spinal Laminae of Origin and Sites of Termination 481
35.2.2 Structural Types of Neurons Involved 482
35.2.3 Spinal Location of Ascending Fibers 482
35.2.4 Response properties 482
35.2.5 Pathways Driven at the Target 483
35.3 Spinobulbar Pathways 483
35.3.1 Ventrolateral Reticular Formation 483
35.3.1.1 Spinal laminae of origin and sites of termination 483
35.3.1.2 Structural types of neurons involved 484
35.3.1.3 Spinal location of ascending fibers 485
35.3.1.4 Surface receptors 485
35.3.1.5 Neurotransmitters 485
35.3.1.6 Response properties 486
35.3.1.7 Pathways driven at the target 486
35.3.2 Dorsal Reticular Nucleus 487
35.3.2.1 Spinal laminae of origin and sites of termination 487
35.3.2.2 Structural types of neurons involved 488
35.3.2.3 Spinal location of ascending fibers 488
35.3.2.4 Surface receptors 488
35.3.2.5 Neurotransmitters 488
35.3.2.6 Response properties 488
35.3.2.7 Pathways driven at the target 489
35.3.3 Nucleus Tractus Solitarii 489
35.3.3.1 Spinal laminae of origin and sites of termination 489
35.3.3.2 Structural types of neurons involved 489
35.3.3.3 Spinal location of ascending fibers 491
35.3.3.4 Neurotransmitters 491
35.3.3.5 Response properties 491
35.3.3.6 Pathways driven at the target 491
35.3.4 Rostral Ventromedial Medulla 492
35.3.4.1 Spinal laminae of origin and sites of termination 492
35.3.4.2 Structural types of neurons involved 493
35.3.4.3 Spinal location of ascending fibers 493
35.3.4.4 Neurotransmitters 493
35.3.4.5 Response properties 493
35.3.4.6 Pathways driven at the target 493
35.4 Spinopontine Pathways 494
35.4.1 Parabrachial Nuclei 494
35.4.1.1 Spinal laminae of origin and sites of termination 494
35.4.1.2 Structural types of neurons involved 495
35.4.1.3 Spinal location of ascending fibers 495
35.4.1.4 Surface receptors 496

477
478 Ascending Pathways: Anatomy and Physiology

35.4.1.5 Neurotransmitters 496

35.4.1.6 Response properties 496
35.4.1.7 Pathways driven at the target 497
35.5 Spinomesencephalic Pathways 497
35.5.1 Periaqueductal Gray 497
35.5.1.1 Spinal laminae of origin and sites of termination 497
35.5.1.2 Structural types of neurons involved 500
35.5.1.3 Spinal location of ascending fibers 500
35.5.1.4 Surface receptors 500
35.5.1.5 Neurotransmitters 500
35.5.1.6 Response properties 500
35.5.1.7 Pathways driven at the target 501
35.6 Spinodiencephalic Pathways 501
35.6.1 Lateral Thalamus 502
35.6.1.1 Spinal laminae of origin and sites of termination 502
35.6.1.2 Structural types of neurons involved 504
35.6.1.3 Spinal location of ascending fibers 504
35.6.1.4 Surface receptors 505
35.6.1.5 Neurotransmitters 505
35.6.1.6 Response properties 505
35.6.1.7 Pathways driven at the target 507
35.6.2 Medial Thalamus 507
35.6.2.1 Spinal laminae of origin and sites of termination 507
35.6.2.2 Structural types of neurons involved 508
35.6.2.3 Spinal location of ascending fibers 509
35.6.2.4 Neurotransmitters 509
35.6.2.5 Response properties 509
35.6.2.6 Pathways driven at the target 509
35.6.3 Hypothalamus 510
35.6.3.1 Spinal laminae of origin and sites of termination 510
35.6.3.2 Spinal location of ascending fibers 511
35.6.3.3 Surface receptors 512
35.6.3.4 Response properties 512
35.6.3.5 Pathways driven at the target 512
35.7 Spinothelencephalic Pathways 512
35.7.1 Thelencephalic Targets of Spinal Ascending Fibers 512
35.7.2 Spinal Laminae of Origin 513
35.8 Discussion 514
35.8.1 Multiple Parallel Ascending Pathways 514
35.8.2 Spinal Neuronal Populations at the Origin of Nociceptive Ascending Pathways 516
35.8.3 Stimulus Discrimination 517
35.8.4 Nociceptive Ascending Pathways as Part of a Complex Nociceptive Integration
System 517
References 518

Glossary
anterograde tracing Staining of axonal terminal antidromic activation Evoking neuronal spikes by
arborizations with a substance (tracer) picked up by electric activation of the axon terminal field.
the neuronal soma and dendrites and transported antinociceptive action A neuronal effect
through the axon up to its terminal structures. that results in decrease of responses to pain
 Ascending Pathways: Anatomy and Physiology 479

and inhibiton of neurons driven by nociceptive nociresponsive Driven by the arrival of nocicep-
input. tive input.
ascending pathway Neuronal tract that conveys noxious stimulation Presentation of stimuli that
input from caudal to rostral areas along the spinal represent a potential or effective aggression to a
cord and brain. peripheral tissue.
contralateral pathway Pathway connecting gray pain inhibitory center Region in the central ner-
matter regions (nuclei) located in opposite sides of vous system whose stimulation results in pain
the spinal cord or brain. depression.
dendrites The branch units of a dendritic arbor. primary sensory neurons Neurons that convey
dendritic arbor The receptive area of a neuron, input from the periphery to the central nervous
organized from the perikarya as the ramifying system.
branches of a tree. receptive field of a neuron The peripheral area
dendritic spines Small protrusions of the dendritic whose stimulation elicits neuronal responses.
surface that more often appear as a knob con- retrograde tracing Staining of the neuronal soma
nected to the dendritic shaft by a short pedicle. and dendritic arbors with a substance (tracer)
electrophysiological recording Recording of picked up by the respective axonal boutons and
changes in the membrane potential or current in a transported back through the axon.
neuron. second-order neurons Neurons that transmit
facilitatory loop Neuronal circuit playing a positive input from primary sensory neurons to higher order
feedback action so that neuronal activity is processing centers.
enhanced. somatotopy Structural arrangement that corre-
fiber decussation Crossing of axons from one lates topographic organization at different sites,
side to the other side of the spinal cord or the brain. either at the periphery or in the central nervous
high-threshold neurons Neurons responsive system.
solely to stimuli of high intensity (noxious). spinofugal transmission Input transmission away
ipsilateral pathway Pathway connecting gray from the spinal cord.
matter regions (nuclei) located in the same side of target Area where the axon terminal arborization
the spinal cord or brain. of a neuron is distributed.
low-threshold neurons Neurons responsive transmitter receptors Plasma membrane mole-
solely to stimuli of low intensity (innocuous). cules which, upon activation by a ligand
neuronal soma The central area of a neuron (neurotransmitter), open ionic channels inducing
where the nucleus and most organelles are located. alteration in the membrane potential.
neurotransmitters Molecules that functionally wide dynamic-range neurons Neurons respon-
connect neurons at synapses by being delivered sive to stimuli of graded intensities, from innocuous
by the presynaptic element upon depolarization to noxious, but more intensively at the noxious range.
and acting upon ligand-gated receptors at the
postsynaptic element.

35.1 Introduction processing centers so that pain is ultimately perceived

in its multiple dimensions and adequate adaptive
35.1.1 Defining Nociceptive Ascending
responses are generated. These ascending pathways
Pathways
are believed to terminate in the cortex with one or
Nociceptive information traveling from the periph- several relay stations in their way, although for most
ery in primary sensory neurons is transmitted to of the tracts, studies demonstrating that the interven-
second-order neurons located at the spinal cord and ing neurons are indeed serially connected from the
cranial sensory nuclei. From this first relay, various spinal cord to the cortex are missing. The term noci-
pathways distribute nociceptive input through higher ceptive ascending pathways is thus normally used in
480 Ascending Pathways: Anatomy and Physiology

a restricted sense, to designate the neuronal tracts pain, and a medial pathway responsible for diffuse,
that connect the spinal cord with supraspinal regions, poorly localized persisting pain (Figure 1). From the
each pathway being named from the brain area at spinal cord, nociceptive input is conveyed both to the
which it terminates. posterior lateral sensory nuclei of the thalamus, in the
The nociceptive nature of a pathway is classically lateral or neospinothalamic pathway (Bowsher, D.
demonstrated by recording responses to noxious sti- 1957; Mehler, W. R. 1957; Willis, W. D. et al., 1974;
muli from spinal neurons antidromically activated Giesler, G. J. et al., 1976) and to medial thalamic
from the target (Perl, E. R. and Whitlock, D. G. nuclei, in the medial or paleospinothalamic pathway
1961; Dilly, P. N. et al., 1968). More recently, detec- (Mehler, W. R. et al., 1956; Bowsher, D. 1957; Mehler,
tion of molecular markers of nociceptive activation
(Hunt, S. P. et al., 1987) in conjunction with retro-
grade tracing has been largely used. By revealing
large populations of putative nociceptive neurons,
such a procedure allows an easy characterization of
the location and morphology of neurons projecting in
each pathway, and is particularly valuable to uncover
varying population activation patterns as a function
of stimulation conditions (Lima, D., 1998).
The structural features that best describe a path-
way are the topographic and morphofunctional
characteristics of the spinal neurons involved and
the area of termination of their axons. Studies addres-
sing the morphology of spinal projecting neurons are
particularly few and mainly related to lamina I. One
of the reasons for the restricted use of this kind of
evaluation is the lack of systematic models of classi-
fication of spinal neurons apart from laminae I
(Gobel, S. 1978a; Lima, D. and Coimbra, A., 1983;
1986) and II (Gobel, S., 1978b). Recent studies, how-
ever, call our attention to the importance of neuronal
morphology, in particular, dendritic geometry, in
defining the signal-processing properties of neurons
(Prescott, S. A. and De Koninck, Y. 2002; Szucs, P.
et al., 2003; Mainen, Z. F. and Sejnowski, T. J. 2006),
which justify taking these aspects into account when
addressing the anatamophysiology of a pathway.
The multitude of ascending nociceptive path-
ways, together with the subtleness of the anatomical
and physiological features that separate them as to
their origin at the spinal cord, makes it difficult to
attribute a particular functional meaning to each one.
A tentative way of unraveling the role of each path-
way in nociceptive processing has been the Medial spinothalamic system
elucidation of the connections established by the
target. Lateral spinothalamic system

Figure 1 Diagrammatic representation of the areas of

35.1.2 The Spinothalamic System termination of the lateral spinothalamic pathway and the
medial spinothalamic pathway, and the spinoreticular
The classical view of the ascending nociceptive sys-
pathways that serve as relays in the medial spinothalamic
tem puts particular emphasis on the dual distribution system. Brain and spinal cord photomicrographs were
of nociceptive input through a lateral pathway adapted from Paxinos, G. and Watson, C. 1998. The Rat
responsible for sharp, well-localized short lasting Brain in Stereotaxic Coordinates, 4th edn. Academic Press.
 Ascending Pathways: Anatomy and Physiology 481

W. R. 1957; Giesler, G. J. et al., 1981b). While the first involved in the spinofugal transmission of nociceptive
is monosynaptic, made up of spinal neurons projecting input will be referred following a caudorostral
directly to the thalamus (Trevino, D. L. and Carstens, sequence.
E. 1975; Willis, W. D. et al., 1979; Giesler, G. J. et al.,
1979a), the second is either monosynaptic or polysy-
naptic with a variable number of relays along the 35.2 Spinocervical Pathway
brainstem (Johnson, F. H. 1954; Mehler, W. R. et al.,
1956; Rossi, G. F. and Brodal, A. 1957; Bowsher, D. The spinocervical pathway is one of the ascending
1957; Carstens, E. and Trevino, D. L. 1978b; Willis, W. putative nociceptive pathways uncovered the ear-
D. et al., 1979; Giesler, G. J. et al., 1979a). liest, based on its capacity to activate cortical
The lateral spinothalamic pathway was proposed sensory areas (Catalano, J. V. and Lamarche, G.,
to be involved in the discriminative aspects of pain 1957; Mark, R. F. and Steiner, J., 1958) and on the
(Bowsher, D. 1957; Melzack, R. and Casey, K. L. finding that neurons of the lateral cervical nucleus
1968) based on early studies on the perception defi- (LCN) project to the ventrobasal complex of the
cits resulting from lesions of lateral thalamic nuclei thalamus (Ha, H. and Liu, C. N., 1966; Craig, A. D.
(Dejerine, J. and Roussy, G. 1906; Melzack, R. and and Burton, H., 1979; Baker, M. L. and Giesler, G. J.,
Casey, K. L. 1968). This was later supported by 1985; Giesler, G. J., et al., 1987). It has been claimed,
studies showing that both spinal and thalamic neu- however, to be mostly dedicated to the processing of
rons of the lateral spinothalamic pathway present tactile, hair movement input (Lundberg, A. and
small receptive fields (Giesler, G. J. et al., 1981b; Oscarsson, O., 1961; Taub, A. and Bishop, P. O.,
Willis, W. D. 1988), are capable of encoding the 1965) and be mainly represented in carnivores
extent of the stimulated area and the intensity of (Flink, R. and Westman, J., 1986). The identification
the stimulus(Kenshalo, D. R., Jr. et al., 1979; of a neuronal column continuing the lateral cervical
Peschanski, M. et al., 1980; Guilbaud, G. et al., 1985; nucleus caudally along the entire length of the cord
Willis, W. D. 1988; Guilbaud, G. and Kayser, V. (the lateral spinal nucleus), together with the finding
1988), and terminate following a somatotopic pattern that their neurons also send projections to the thala-
in the thalamus and cortex respectively (Whitsel, B. mus as well as to many other brain areas (see ahead ),
L. et al., 1978; Boivie, J. 1979). The medial spinotha- raised the possibility that the lateral cervical nucleus
lamic pathway was claimed to deal with the affective is part of a more extensive spinal neuronal aggrega-
and volitive aspects of pain (Bowsher, D. 1957; tion lying within the dorsolateral fasciculus, which
Melzack, R. and Casey, K. L. 1968) based on similar serves as a relay in various ascending nociceptive
behavioral studies on human and experimental ani- pathways. Nonetheless, differences in the morphol-
mals suffering from lesions of various brainstem or ogy and response properties of lateral cervical and
thalamic areas (Dejerine, J. and Roussy, G. 1906; lateral spinal neurons (Giesler, G. J. et al., 1979b;
Walker, A. E. 1942b; Hecaen, H. et al., 1949). This Menetrey, D. et al., 1980) point to a specific role of
view was again supported by studies revealing the former in conveying ascending input to the lat-
that spinal and thalamic neurons of the medial path- eral thalamus.
way present large receptive fields, responses
unrelated to stimulus intensity (Giesler, G. J. et al.,
35.2.1 Spinal Laminae of Origin and Sites of
1981b; Guilbaud, G. et al., 1985) and no topographical
Termination
arrangement of their axonal terminal arborizations in
both the thalamus (Boivie, J. 1979) and cortex Neurons projecting to the lateral cervical nucleus are
(Morison, R. S. and Dempsey, E. W. 1942; Jones, E. mainly located in the ipsilateral spinal cord at lamina
G. and Leavitt, R. Y. 1974). IV (Figure 2) (Bryan, R. N. et al., 1973; Craig, A. D.,
In the last decades, the use of very sensitive electro- 1976; Brown, A. G. et al., 1976; Cervero, F. et al., 1977;
physiological and tracing techniques revealed a wide Brown, A. G. et al., 1977; Craig, A. D., 1978; Brown, A.
variety of brainstem loci capable of contributing as G. et al., 1980a; Baker, M. L. and Giesler, G. J., 1984),
relays to the medial spinothalamic system (reviewed where they amount to 60% of the entire population,
by (Lima, D., 1997), and uncovered ascending path- followed by lamina III (10%) (Brown, A. G. et al.,
ways that bypass the thalamus to terminate directly on 1980a). Scattered neurons are present in other lami-
telencephalic areas, including the cortex (Cliffer, K. D. nae such as laminae I, V, VI, and VII (Craig, A. D.,
et al., 1991). In this chapter, the various pathways 1976; Craig, A. D., 1978; Brown, A. G. et al., 1980a;
482 Ascending Pathways: Anatomy and Physiology

1984), but neurons with large round soma have also

been observed in lamina IV (Craig, A. D., 1978).
They present dorsally oriented dendrites (Brown,
C1
A. G. et al., 1976; Jankowska, E. et al., 1976; Brown,
A. G. et al., 1977; Craig, A. D., 1978; Brown, A. G. et al.,
1980b), which extend rostrocaudally for up to
2000 mm without penetrating lamina II (Brown, A. G.
et al., 1977; Brown, A. G. et al., 1980b).

35.2.3 Spinal Location of Ascending

Fibers
Spinal axons targeting the lateral cervical nucleus
course ipsilaterally in the dorsal part of the lateral
C5 funiculus (Figure 2) (Brown, A. G. et al., 1977; Baker, M.
L. and Giesler, G. J., 1984; Giesler, G. J. et al., 1988).
According to Ha and Liu (Ha, H. and Liu, C. N., 1966),
they are often collaterals of fibers coursing to more
rostral levels.

35.2.4 Response properties

L5
Nociceptive neurons at the origin of the spinocervi-
Spino-LCN cal pathway present background activity in bursts
(Brown, A. G. and Franz, D. N., 1969). They are
activated by A, A, and C primary afferent fibers
(Taub, A. and Bishop, P. O., 1965), A fibers exciting
Figure 2 Diagram representing the spinal laminae of the entire neuronal population either alone (29%) or
origin, ascendingb course in the spinal cord, and areas of
in convergence with C fibers (71%) (Brown, A. G.
termination of the spino-LCN pathway. Note the
somatotopic arrangement of the axon terminal fields et al., 1975). They belong in the low-threshold (LT),
according to their rostrocaudal origin. (in this figure and wide-dynamic range (WDR) and high-threshold
figures 3, 7, 10, 12, 13, 15, 17, 19, 21 and 24, the left (NS) classes (Brown, A. G. and Franz, D. N., 1969;
side is ipsilateral to the side of arrival of peripheral Bryan, R. N. et al., 1973; Bryan, R. N. et al., 1974;
input) Brain and spinal cord photomicrographs were
Cervero, F. et al., 1977; Downie, J. W. et al., 1988).
adapted from Paxinos, G. and Watson, C. 1998. The
Rat Brain in Stereotaxic Coordinates, 4th edn. Academic Neurons receiving high-threshold input were
Press. reported to make up between 60% and 86% of the
entire projecting population (Cervero, F. et al., 1977;
Baker, M. L. and Giesler, G. J., 1984). Axonal term- Kajander, K. C. and Giesler, G. J., 1987). High-
inals are somatotopically organized, fibers originated threshold input is generated either by pressure,
in caudal levels terminating in the dorsolateral aspect pinch, heat, or cold (Brown, A. G. and Franz, D. N.,
of the caudal portion of the lateral cervical nucleus, 1969; Bryan, R. N. et al., 1973; Bryan, R. N. et al., 1974;
and fibers originated in rostral levels in the medial Cervero, F. et al., 1977; Downie, J. W. et al., 1988),
aspect of its rostral portion (Figure 2) (Svensson, B. A. while low-threshold input is mainly originated in
et al., 1985). hair follicle afferent receptors (Brown, A. G. and
Franz, D. N., 1969). Receptive fields are small and
located in hairy as well as glabrous skin (Bryan, R. N.
et al., 1974; Kunze, W. A. A. et al., 1987; Downie, J. W.
35.2.2 Structural Types of Neurons
et al., 1988). They are organized somatotopically so
Involved
that cells located more laterally have receptive fields
Neurons targeting the lateral cervical nucleus are in the dorsal surface of the body and cells located
commonly described as small-sized (1020 mm) more medially in the ventral surface (Bryan, R. N.
(Craig, A. D., 1978; Baker, M. L. and Giesler, G. J., et al., 1973; Bryan, R. N. et al., 1974; Brown, A. G. et al.,
 Ascending Pathways: Anatomy and Physiology 483

1980a). Inhibitory receptive fields were described similarities with the rat, pointing to a high degree of
adjacent to the excitatory receptive field (Brown, phylogenetic conservation.
A. G. et al., 1987; Short, A. D. et al., 1990) or in
the contralateral limb (Brown, A. G. and Franz,
D. N., 1969). Natural stimuli causing neuronal inhi- 35.3.1 Ventrolateral Reticular Formation
bition include hair movement, pressure, and
squeezing (Brown, A. G. and Franz, D. N., 1969; 35.3.1.1 Spinal laminae of origin and sites
Brown, A. G. et al., 1987; Short, A. D. et al., 1990). of termination
Convergence of cutaneous and deep tissues input Spinal cells projecting to the caudal ventrolateral
has been reported (Kniffki, K. D. et al., 1977; reticular formation (VLM) are distributed through
Hamann, W. C. et al., 1978; Harrison, P. J. and laminae I, II, IVVII, VIII, X, and the lateral spinal
Jankowska, E., 1984). Only cells responsive to both nucleus (Figure 3) (Menetrey, D. et al., 1983;
hair movement and skin pressure were shown to Menetrey, D. et al., 1984; Leah, J. et al., 1988; Lima,
receive group III and IV muscle afferent input D. et al., 1991; Galhardo, V. et al., 2000). Projections
(Hamann, W. C. et al. 1978). Muscle and joint primary were first described to be mainly contralateral
afferent activation can also elicit neuronal inhibition (Menetrey, D. et al., 1983; Menetrey, D. et al., 1984;
(Hamann, W. C. et al. 1978; Harrison, P. J. and Thies, R., 1985), but an important ipsilateral compo-
Jankowska, E. 1984). nent was revealed by the use of more sensitive
retrograde tracers (Menetrey, D. et al., 1982; Lima,
D. et al., 1991; Menetrey, D. et al., 1992b; Galhardo, V.
35.2.5 Pathways Driven at the Target et al., 2000). There is, however, a large variability in
the proportion of cells labeled in each spinal side,
Axons from neurons in the lateral cervical nucleus
particularly with respect to cells located in the super-
join the medial lemniscus contralaterally and termi-
ficial dorsal horn. Both ipsilateral and contralateral
nate in the midbrain, the ventral posterial
predominance have been observed, in a few cases in
lateral nucleus (VPL), and the posterior complex
the same animal at different rostrocaudal levels
(PO) of the thalamus (Ha, H. and Liu, C. N.,
(Lima, D. et al., 1991). The occurrence of subtle
1966). In the midbrain, the lateral part of the peria-
differences in the area of termination of each spinal
queductal gray (PAG) receives afferents from
side along the cord length was proposed as a tentative
the lateral two-thirds of the lateral cervical nucleus
explanation, but experiments designed to clarify a
(Mouton, L. J. and Holstege, G., 2000; Mouton, L. J.
putative somatotopic arrangement are missing.
et al., 2004). In the VPL, fibers ascending from
the lateral cervical nucleus are topographically
arranged so that those originated in its dorsolateral
portion terminate in the VPL, pars lateralis, and those
originated in its ventromedial portion terminate
in the VPL, pars medialis (Craig, A. D. and Burton,
H., 1979).

35.3 Spinobulbar Pathways

Most spinobulbar pathways were uncovered rela- Spino-VLMlat

tively recently and are not as thoroughly studied as Spino-LRT (lateral)

the more rostrally terminating pathways such as the Spino-LRT (medial)
spinomesencephalic and the spinothalamic. With the
exception of the rostral ventromedial medulla Figure 3 Diagram representing the spinal laminae of
(RVM), the majority of the studies dealing with spi- origin, ascending course in the spinal cord, and areas of
termination of the spino-VLM pathway. Note the
nobulbar pathways were performed in the rat. An
contribution of lamina II neurons. Brain and spinal cord
anecdotic study addressing the spinal pathways ter- photomicrographs were adapted from Paxinos, G. and
minating in the caudal medulla of the pigeon Watson, C. 1998. The Rat Brain in Stereotaxic Coordinates,
(Galhardo, V. et al., 2000), however, reveals remarkable 4th edn. Academic Press.
484 Ascending Pathways: Anatomy and Physiology

Spino-VLM axons appear to give axonal collaterals

to various sites within the brainstem (Thies, R.,
1985), such as the parabrachial nuclei (PBN) and
the PAG (Spike, R. C. et al., 2003).
The use of discrete injections, which became pos-
sible with the advent of particularly sensitive tracing
methods, clearly demonstrated that the spino-VLM
system is composed of three pathways originating in
three distinct spinal regions (Figure 3).
The pathway originating in the superficial dorsal
horn (laminae IIII) and lateral spinal nucleus was
observed in the rat (Lima, D. and Coimbra, A., 1991;
Tavares, I. et al., 1993), cat, and monkey (Craig, A. D.,
1995) to terminate in the lateral-most portion of the
VLM (VLMlat), in between the spinal trigeminal
nucleus and the ventral tip of the lateral reticular
nucleus (LRt). The pathways originating in the
deep dorsal horn (laminae IVVI) and intermedi-
ate/ventral horn (laminae VII and X) terminate,
respectively, in the lateral half and the medial half
of the LRt (Lima, D. et al., 1991).
A curious aspect of the spino-VLMlat projection
is the participation of a large amount of cells of the
substantia gelatinosa, or lamina II (Figure 4) (Lima,
D. et al., 1991; Lima, D. and Coimbra, A., 1991), an
area that was not shown to participate significantly in
any other ascending pathway and is therefore usually
taken as involved in nociceptive modulatory local or
propriospinal circuits (see Chapter Spinal Cord
Physiology of Nociception). Lamina II neurons pro-
ject, together with neurons in lamina I and the lateral
spinal nucleus, to the VLMlat (Figure 3), contribut-
ing to almost one-third of the entire spinal-VLMlat
projection in the cervical enlargement, and to one-
fifth in the lumbar enlargement.
(Lima, D. and Coimbra, A., 1991). There are still
no clues on the physiological meaning of this unique
lamina II projection to the VLMlat. Studies aimed at
clarifying whether these cells participate in inhibi-
tory circuits similar to those described in the
superficial dorsal horn failed to reveal GABA content
in these neurons (Tavares, I. and Lima, D., 2002).
Figure 4 Superficial dorsal horn neurons labeled
35.3.1.2 Structural types of neurons retrogradely with cholera toxin subunit B (CTb) from the
involved VLMlat. (a) Lamina I neuron of the fusiform type, subtype B, in
Lamina I neurons projecting in the spino-VLM path- horizontal view. (b) Lamina I fusiforn neurons and lamina II
way were characterized in the rat, based on the neurons in transverse view. Note the ventrally oriented
dendrites of fusiform B neurons (large arrows in (b)). DF
comparison of the structural features of neurons retro- Dorsal funiculus. Scale bar 30 mm. Adapted from figure 3 of
gradely labeled with CTb at various spinal cord levels Lima, D., Mendes-Ribeiro, J. A., and Coimbra, A. 1991. The
in the three anatomical planes (Lima, D. et al., 1991). In spino-latero-reticular system of the rat: projections from the
the rat, as in the pigeon (Galhardo, V. et al., 2000), superficial dorsal horn and structural characterization of
spino-VLM neurons belong in three (fusiform, marginal neurons involved. Neuroscience 45, 137152.
 Ascending Pathways: Anatomy and Physiology 485

pyramidal, and flattened) of the four structural 35.3.1.3 Spinal location of ascending
neuronal groups present in lamina I. The majority fibers
(around 80%) are of the fusiform type, a neuronal According to the intense fiber staining occurring in
group whose main characteristic is the strict longitu- the dorsolateral fasciculus after retrograde tracer
dinal spiny dendritic arbor. Some fusiform neurons injections in theVLMlat (Lima, D. et al., 1991),
present a few dendrites oriented ventrally and pene- VLM projections from laminae IIII course in the
trating the entire width of lamina II (Figure 4). These dorsal portion of the lateral funiculus (Figure 3), as is
neurons, classified as fusiform B by Lima and Coimbra the case of the spinomesencephalic (McMahon, S. B.
(Lima, D. and Coimbra, A., 1986), represent a large and Wall, P. D., 1985; Hylden, J. L. K. et al., 1986b)
fraction of VLMlat-projecting fusiform cells (20%) and spinothalamic (Apkarian, A. V. et al., 1985;
when compared to the small contribution of this neu- Apkarian, A. V. and Hodge, C. J., 1989c) pathways.
ronal subtype (6%) to the entire lamina I fusiform
neuronal population (Lima, D. and Coimbra, A., 35.3.1.4 Surface receptors
1986). Curiously, fusiform B neurons could not be Large numbers of lamina I neurons projecting to the
observed in other spinofugal pathways. This finding VLM express neurokinin I (NK1) (Figure 5) (Todd,
is particularly interesting in the light of the contribu- A. J. et al., 2000; Spike, R. C. et al., 2003; Castro, A. R.
tion of lamina II neurons to this pathway (Figure 4). It et al., 2006) and GABAB receptors (Castro, A. R. et al.,
may indicate that neurons of the fusiform B subtype 2006), the co-localization of both being relatively
cooperate with lamina II neurons in transmitting to the frequent (Castro, A. R. et al., 2006).
VLMlat primary input arriving at lamina II. The In addition, 90% of the large deep dorsal horn
remaining 20% VLM-projecting lamina I neurons neurons with dendrites entering superficial laminae
belong to the flattened and pyramidal types in similar and exhibiting the NK1 receptor project to the VLM
amounts. These two cell groups have in common long (Todd, A. J. et al., 2000). Appositions of serotonin
practically aspiny dendrites coursing horizontally, par- (5HT) and noradrenalin-containing axonal boutons
allel to the dorsal surface of lamina I. Pyramidal upon VLM-projecting lamina I neurons are common
neurons have, in addition, dendrites that ramify inside (Tavares, I. et al., 1996a; Polgar, E. et al., 2002).
the white matter overlying lamina I. Serotoninergic boutons establish symmetrical synap-
VLM-projecting lamina II neurons present ovoid, tic contacts and are more abundant upon neurons
rostrocaudally elongated dendritic arbors that extend expressing the NK1 receptor (Polgar, E. et al., 2002).
as parasagittal sheets through the entire width of
lamina II (Lima, D. and Coimbra, A., 1991), resem- 35.3.1.5 Neurotransmitters
bling the central and the limiting cells of Ramon y As in most ascending pathways conveying nociceptive
Cajal (Ramon y Cajal, S., 1909). input, studies addressing the neurochemical content of

(a) (b)

(c) (d)

Figure 5 Flattened (a, b) and pyramidal (c, d) lamina I neurons, in horizontal view, retrogradely labeled with cholera toxin
subunit B (CTb) from the VLM (red) and immunoreactive for the NK1 receptor (green). Scale bar 50 mm. Adapted from
figure 9 of Spike, R. C., Puskar, Z., Andrew, D., and Todd, A. J. 2003. A quantitative and morphological study of projection
neurons in lamina I of the rat lumbar spinal cord. Eur. J. Neurosci. 18, 24332448.
486 Ascending Pathways: Anatomy and Physiology

the neurons of origin of the spino-VLM pathway have the VLM region that establishes connections with
focused on neuropeptides. The exhaustive study of other supraspinal pain inhibitory centers (Tavares, I.
Leah and collaborators (Leah, J. et al., 1988) revealed et al., 1996b; Cobos, A. et al., 2003), pointing to a role of
a relatively large amount of vasoactive intestinal pep- the superficial dorsal horn-VLMlat pathway in driving
tide (VIP), bombesin, dynorphin, and substance P- the potent descending inhibition that can be elicited
immunoreactive VLM-projecting neurons in the lat- upon stimulation of VLM (Gebhart, G. F. and Ossipov,
eral spinal nucleus. A few enkephalin immunoreactive M. H., 1986). At the RVM and the pontine A5 nora-
neurons were observed in lamina X. Calbindin is pre- drenergic group, terminal arborizations of VLM axons
sent in particularly large numbers of lumbosacral cells appose spinal-projecting neurons (Figure 6), indicating
projecting to the VLM-bilaterally, especially within that pain control actions from those areas are indeed
lamina I and the lateral spinal nucleus, but also in under the control of the VLM (Tavares, I. et al., 1996b).
lamina X (Menetrey, D. et al., 1992b). Although fusi-
form neurons in lamina I (Lima, D. et al., 1993) and
neurons in lamina II (Todd, A. J. and Spike, R. C., 1993)
are known to contain GABA, GABA-immunostaining
(a)
of neurons retrogradely labeled from the VLM could
not be observed (Tavares, I. and Lima, D., 2002).

35.3.1.6 Response properties

The response properties of neurons projecting to the
ventrolateral reticular formation were studied in the
rat (Menetrey, D. et al., 1984) and cat (Thies, R.,
1985). A high proportion of VLM-projecting neurons
are spontaneously active (Menetrey, D. et al., 1984;
Thies, R., 1985). NS neurons make up about half of
the entire population of VLM-projecting neurons
(Menetrey, D. et al., 1984; Thies, R., 1985). The
remaining are either WDR or LT/proprioceptive
neurons (Menetrey, D. et al., 1984; Thies, R., 1985).
They often present bilateral symmetrical receptive (b)
fields as well as cutaneous inhibitory receptive fields
(Menetrey, D. et al., 1984). Convergence of cuta-
neous, visceral, and muscle input is frequently
observed (Thies, R., 1985). By monitoring noxious-
evoked c-fos induction (Tavares, I. et al., 1993), 10%
to 20% of VLM-projecting neurons were shown to
be activated by heat or mechanical stimulation in
laminae I and IIo. In lamina IIi, neurons were acti-
vated in fewer numbers and only after thermal
stimulation.

35.3.1.7 Pathways driven at the target

The VLM projects to several brain areas also receiving Figure 6 RVM (a) and A5 (b) neurons retrogradely
nociceptive input from the spinal cord and involved in labeled with cholera toxin subunit B (CTb) from the spinal
cord (brown granules) and receiving appositions (arrows)
pain control as well as cardiovascular, endocrine, or from axonal boutons anterogradely labeled with
limbic functioning. Among these areas stand the RVM; biotinilated dextran amine (BDA) from the VLM. Neuron in
the dorsal reticular nucleus (DRt); the A5, A6, and A7 (b) is immunoreactive for dopamine--Hydroxylase (DBH;
pontine noradrenergic groups (Tavares, I. et al., 1996b); blue). Scale bar 20 mm. (Adapted from figure 3 of
Tavares, I., Lima, D., and Coimbra, A. 1996b. The
the hypothalamus (Calaresu, F. R. et al., 1984; Malick,
ventrolateral medulla of the rat is connected with the
A. et al., 2000); and the central nucleus of the amygdala spinal cord dorsal horn by an indirect descending
(Zardettosmith, A. M. and Gray, T. S., 1995). Studies pathway relayed in the A5 noradrenergic cell group. J.
focused on the brainstem showed that the VLMlat is Comp. Neurol. 374, 8495).
 Ascending Pathways: Anatomy and Physiology 487

In the A5, spinal-projecting neurons contacted by 35.3.2 Dorsal Reticular Nucleus

VLMlat fibers are noradrenergic (Figure 6) and post-
35.3.2.1 Spinal laminae of origin and sites
synaptic in asymmetric, putative excitatory synaptic
of termination
contacts (Tavares, I. et al., 1996b). This was taken as
The medullary dorsal reticular nucleus (DRt) was
suggestive that the spinal 2 adrenoreceptor-
first shown in the rat to be the site of termination of
mediated antinociceptive action triggered in the
an important, mainly ipsilateral, pathway ascending
VLM is dependent on VLMlat activation and relayed
from the spinal cord (Lima, D. and Coimbra, A., 1985;
in the A5 group.
Lima, D., 1990; Villanueva, L. et al., 1991). By the
The VLM also sends direct descending projec-
tions to the spinal cord (Tavares, I. and Lima, D., same time, neurons in the DRt were shown to be
1994). Projections to both the superficial and deep exclusively or preferentially activated by noxious
dorsal horn originate in the VLMlat (Tavares, I. and stimuli from the skin and viscera (Villanueva, L.
Lima, D., 1994). Data from several studies indicate et al., 1988). The spino-DRt pathway was later uncov-
that noradrenaline is not used in the direct VLM- ered in other species such as the cat, monkey (Craig,
spinal pathway (Westlund, K. N. et al., 1981; A. D., 1995), and pigeon (Galhardo, V. et al., 2000). It
Westlund, K. N. et al., 1983; Tavares, I. et al., 1996b). is constituted by a dorsal and ventral component
VLMlat axons targeting lamina I make up a recipro- differing in the area of termination within the DRt
cal closed VLM-spino-VLM loop which is entirely (Figure 7).
excitatory at the VLM level, and both excitatory and The dorsal pathway terminates at the dorsal-most
inhibitory at the spinal level (Tavares, I. et al., 1998; portion of the DRt, immediately above the level of
Tavares, I. and Lima, D., 2002). the central canal and surrounding the ventral border
The cerebellar connections of the LRt (Cledenin, of the cuneate nucleus (Almeida, A. et al., 1995;
M. et al., 1974; Parenti, R. et al., 1996), together with Almeida, A. et al., 2000). It originates from the med-
the fact that, contrary to the VLMlat, the LRt does ial-most part of laminae IIII ipsilaterally, with a
not contribute descending projections to the spinal marked predominance of lamina I, and from lamina
cord dorsal horn (Tavares, I. and Lima, D., 1994), X, bilaterally (Figure 7) (Lima, D., 1990). The ventral
point to a role of the deep dorsal horn/ventral horn- pathway terminates ventrally to the area of termina-
LRt pathways in the control of motor activity in tion of the dorsal pathway, within both sides of the
response to pain. DRt (Almeida, A. et al., 1995; Almeida, A. et al., 2000).

Spinodorsal DRt

Spinoventral DRt

Figure 7 Diagram representing the spinal laminae of origin, ascending course in the spinal cord, and areas of termination
of the spino-DRt pathway. Note the preferential medial location of dorsal horn neurons. Fibers of the spinodorsal DRt
pathway course in the dorsal funiculus. Brain and spinal cord photomicrographs were adapted from Paxinos, G. and Watson,
C. 1998. The Rat Brain in Stereotaxic Coordinates, 4th edn. Academic Press.
488 Ascending Pathways: Anatomy and Physiology

Its cells of origin prevail ipsilaterally in the medial was completely abolished in ipsilateral laminae IIII
portion of laminae IVVI, with additional bilateral and markedly diminished in the deep dorsal horn
participation from laminae VII and X (Figure 7) at segments caudal to the lesion. Anterograde tracing
(Lima, D., 1990). Interestingly, retrograde labeling later revealed that numerous fibers are labeled in
from the superficial or deep dorsal horn demon- the dorsal funiculus after injections in superficial
strated descending pathways arising, respectively, laminae, and in the dorsolateral fasciculus after
from the ipsilateral dorsal-most part of the DRt or injections in deep dorsal horn laminae (Almeida, A.
from both sides of the ventral DRt (Tavares, I. and et al., 1995). Accordingly, injections in the dorsal
Lima, D., 1994). The DRt is thus part of two distinct funiculus resulted in DRt labeling restricted to
reciprocal loops connecting its dorsal part with the its ipsilateral dorsal part, while injections in the
superficial dorsal horn ipsilaterally, and ventral part dorsolateral fasciculus produced ipsilateral ventral
with the deep dorsal horn bilaterally (Almeida, A. DRt labeling (Almeida, A. et al., 1995). These find-
et al., 1993; Almeida, A. et al., 2000). ings not only confirm a dual ascending tract for the
dorsal and ventral DRt pathways, but also indicate
35.3.2.2 Structural types of neurons that fiber decussation in the ventral pathway takes
involved place near the segment of origin, as in most other
Lamina I neurons projecting to the dorsal DRt were pathways.
structurally characterized in the rat based on obser-
vations of neurons retrogradely filled with CTb in 35.3.2.4 Surface receptors
the three anatomical planes (Lima, D. and Coimbra, A significant amount of lamina I neurons projecting to
A., 1990). As to other spinal laminae, no data the DRt express the NK1 receptor alone (Todd, A. J.
have been collected so far. In the rat, about 30% of et al., 2000; Castro, A. R. et al., 2006) or together with
lamina I neurons projecting to the DRt belong in the the GABAB receptor (Castro, A. R. et al., 2006). DRt-
pyramidal and flattened types in similar amounts. projecting neurons expressing only the GABAB recep-
The remaining 70% are of the multipolar type, tor are, however, much more numerous (Castro, A. R.
a finding particularly curious taking into account et al., 2006). In laminae IIIIV, about 20% of NK1-
that multipolar neurons were not seen to project in expressing neurons project to the DRt (Todd, A. J.
other ascending pathways. These cells are rather et al., 2000).
peculiar due to the typical lamina II crossing of the
dendritic arbor, the bush pattern of the highly rami- 35.3.2.5 Neurotransmitters
fied proximal dendritic branches and the profusion So far, there are no studies addressing the possible
and large variety of dendritic spines (Lima, D. and neurotransmitters used in the spino-DRt pathway.
Coimbra, A., 1986). Another interesting feature of Nevertheless, it is interesting to note that, although
multipolar cells is their preferential location in the immunoreactions for GABA revealed immunostain-
medial third of lamina I in the rat (Lima, D. and ing of lamina I neurons of the multipolar type, (Lima,
Coimbra, A., 1983; Lima, D. and Coimbra, A., 1986). D. et al., 1993), even in the early 2000s, GABA could
The same structural types of lamina I neurons were not be detected in projecting spinal neurons
seen to project to the DRt in the pigeon, although (Gamboa-Esteves, F. O. et al., 2001b; Tavares, I. and
flattened and pyramidal neurons were relatively more Lima, D., 2002).
abundant in this species (Galhardo, V. et al., 2000).
35.3.2.6 Response properties
35.3.2.3 Spinal location of ascending The few data regarding the response characteristics
fibers of spino-DRt neurons rely on the induction of the
An interesting aspect of the spino-DRt pathway, c-fos proto-oncogene as a marker of activation of
apparently only shared with the spino-NTS pathway spinal neurons following noxious stimulation (Hunt,
(Gamboa-Esteves, F. O. et al., 2001c), is the course S. P. et al., 1987). This kind of approach only per-
of their axons in the dorsal funiculus. This was mitted to conclude on the activation by various kinds
first suggested by the dorsal orientation of axonal of cutaneous and visceral noxious stimulation of
processes of lamina I neurons labeled retrogradely DRt-projecting lamina I neurons of all the three
from the DRt and by comparing retrograde labeling structural groups involved, namely multipolar, flat-
rostrally and caudally to lesions of the dorsal funi- tened, and pyramidal (Almeida, A. and Lima, D.,
culus (Lima, D., 1990). Retrograde labeling 1997). The rate of activation of DRt-projecting
 Ascending Pathways: Anatomy and Physiology 489

lamina I neurons varied from 20% to 80% depending (Lima, D., 1998). A ventral DRt-deep dorsal horn
on the kind of stimulation applied and the neuronal pain facilitatory loop is also likely to be established
cell group (Almeida, A. and Lima, D., 1997). In the as indicated by the increased responsiveness of WDR
deep dorsal horn, although high levels of c-fos deep dorsal horn neurons upon DRt stimulation
expression were observed, the amount of activation (Dugast, C. et al., 2003).
of projecting neurons did not exceed 5%.
Electrophysiological studies are absolutely needed
35.3.3 Nucleus Tractus Solitarii
to further characterize the response properties of
spino-DRt neurons and to clarify the involvement 35.3.3.1 Spinal laminae of origin and sites
of the deep dorsal horn in the transmission of noci- of termination
ceptive input to the DRt. The spino-NTS pathway was demonstrated in the
rat (Menetrey, D. and Basbaum, A. I., 1987; Leah, J.
35.3.2.7 Pathways driven at the target et al., 1988; Esteves, F. et al., 1993) and pigeon
Projections from the DRt reach the parafascicular, (Galhardo, V. et al., 2000). It originates in laminae I,
ventromedial, and reunions thalamic nuclei (Figure 8) IVVI, VII, X, and the lateral spinal nucleus, mainly
in the rat (Villanueva, L. et al., 1998), which in turn are contralaterally, but with an important ipsilateral
connected with telencephalic areas involved in emo- component (Figure 10) (Esteves, F. et al., 1993).
tional/affective and cognitive control. A putative In the rat, the thoracic and sacral autono-
spinoreticulothalamocortical projection relayed at the mic cell columns were also shown to participate
DRt has been proposed (Desbois, C. and Villanueva, L., (Menetrey, D. and Basbaum, A. I., 1987; Menetrey, D.
2001). Widespread projections to other brain targets of and DePommery, J., 1991). In addition, an important
nociceptive ascending pathways have also been contribution from the superficial laminae of the spinal
described. These include the VLM; the NTS; the trigeminal nucleus, pars caudalis, was revealed
rostral ventromedial medulla; the pontine noradre- (Menetrey, D. and Basbaum, A. I., 1987). The spino-
nergic cell groups A5, A6, and A7 (PBN); the PAG; NTS pathway terminates in the caudal part of
the posterior thalamus; the hypothalamus; the septal the NTS, the general visceral zone (Figure 10)
nuclei; the globus pallidus; and the amygdala (Loewy, A. D., 1990). Based on restricted retrograde
(Figure 8) (Bernard, J. F. and Besson, J. M., 1990; tracer injections, cells projecting to the lateral subnu-
Bernard, J. F. et al., 1990; Villanueva, L. et al., 1998; cleus are fewer than those targeting the medial NTS,
Leite-Almeida, H. et al., 2006). Important projections and do not include lamina I cells (Figure 10) (Esteves,
to the orofacial motor nuclei (Bernard, J. F. et al., F. et al., 1993). Anterograde tracing confirmed this
1990; Leite-Almeida, H. et al., 2006) as well as to the finding by showing that fibers originating in the super-
deep cerebellar nuclei (Leite-Almeida, H. et al., 2006) ficial dorsal horn terminate bilaterally in the medial
favor an important role in the organization of facial part of the commissural subnucleus, while fibers origi-
expressions and vocalization triggered by noxious nating in the deep dorsal horn terminate ipsilaterally in
stimulation. the lateral subnucleus, with a few fibers distributed to
The DRt also projects to the spinal cord super- the dorsomedial subnucleus (Gamboa-Esteves, F. O.
ficial (Tavares, I. and Lima, D., 1994) and deep dorsal et al., 2001c).
horn (Tavares, I. and Lima, D., 1994; Villanueva, L.
et al., 1995) as well as to the intermediate/ventral 35.3.3.2 Structural types of neurons
horn (Villanueva, L. et al., 1995). DRt axons terminat- involved
ing in lamina I participate in a closed reciprocal Again, studies referring to the structural characteris-
spinodorsal DRt-spinal loop which, based on the tics of spino-NTS neurons only addressed lamina I.
asymmetric structure of synapses, is likely to be Similar to what was observed for the VLM, lamina I
excitatory at both the spinal and medullary levels neurons projecting to the NTS belong in the fusiform,
(Figure 9) (Almeida, A. et al., 1993; Almeida, A. et al., pyramidal, and flattened groups. However, contrary to
2000; Lima, D. and Almeida, A., 2002). Nociceptive the VLM, fusiform neurons contribute a small fraction.
input arriving at the DRt is thus thought to drive a In spite of constituting half of the lamina I neuronal
reverberating, lamina I centered pain facilitatory cir- population (Lima, D. and Coimbra, A., 1983; Lima, D.
cuit, which is in accordance with the high proportion and Coimbra, A., 1986), fusiform neurons amount
of c-fos activated spino-DRt neurons when compared only to 25% of NTS-projecting lamina I neurons,
to that of activated cells projecting to other targets whereas flattened and pyramidal neurons make up
490 Ascending Pathways: Anatomy and Physiology

(a) (f)

(b) (g)

(c) (h)

(i)
(d)

(e) (j)

Figure 8 Distribution of DRt efferents anterogradely labeled with phaseolus vulgaris leucoagglutinin (PHA-L) in the
diencephalon and telencephalon, in horizontal view. Adapted from figure 6 of Villanueva, L., Desbois, C., Le Bars, D., and
Bernard, J. F. 1998. Organization of diencephalic projections from the medullary subnucleus reticularis dorsalis and the
adjacent cuneate nucleus: a retrograde and anterograde tracer study in the rat. J. Comp. Neurol. 390, 133160.

about 3540% each. Similar relative amounts were of flattened cells in the spino-NTS pathway deserves
observed in the pigeon, although pyramidal cells particular attention since it may be related to the
prevailed followed by flattened and fusiform cells specific role of this pathway in pain processing.
(Galhardo, V. et al., 2000). The abundant participation Flattened cells make up 10% of the entire lamina I
 Ascending Pathways: Anatomy and Physiology 491

(a) (b)
nucleus, and bombesin in lamina X (Leah, J. et al.,
1988). Calbindin was immunodetected in spinal
NTS-projecting neurons located mainly in lamina I
(Figure 11) and the lateral spinal nucleus (Menetrey, D.
et al., 1992b; Gamboa-Esteves, F. O. et al., 2001a), as well
as in lamina I of the spinal trigeminal nucleus, pars
caudalis (Menetrey, D. et al., 1992a). In lamina I, about
40% of fusiform NTS-projecting neurons contain cal-
bindin. Pyramidal and flattened neurons also exhibit
calbindin (Figure 11), but in smaller fractions
(Gamboa-Esteves, F. O. et al., 2001a). Glutamate also
occurs mainly in fusiform neurons followed by pyra-
Figure 9 Anterogradely labeled axonal bouton () in the midal neurons, and co-localizes extensively with
DRt (a) and superficial dorsal horn (b) establishing calbindin (Gamboa-Esteves, F. O. et al., 2001a). Nitric
asymmetric synaptic contacts (arrows) upon retrogradely oxide synthase is present in fewer cells, almost all of
labeled dendrites after injecting cholera toxin subunit B
them the fusiform type (Gamboa-Esteves, F. O. et al.,
(CTb) in the superficial dorsal horn (a) or horseradish
peroxidase (HRP) in the dorsal DRt (b). In (b), tracer 2001a). Calbindin-immunoreactive NTS-projecting
deposits are pointed by cuved arrows. Scale bar 0.3 mm (a) neurons are c-fos activated by visceral and cutaneous
Adapted from figure 3 of Almeida, A., Tavares, I., and Lima, stimulation (Gamboa-Esteves, F. O. et al., 2001b).
D. 2000. Reciprocal connections between the medullary Glutamate-positive and nitric oxide synthase-positive
dorsal reticular nucleus and the spinal dorsal horn in the rat.
neurons are c-fos activated only by visceral stimulation
Eur. J. Pain. 4, 373387. (b) Adapted from figure 3 of
Almeida, A., Tavares, I., Lima, D., and Coimbra, A. 1993. (Gamboa-Esteves, F. O. et al., 2001b). About 5% neu-
Descending projections from the medullary dorsal rons of NTS-projecting neurons of the pyramidal
reticular nucleus make synaptic contacts with spinal cord group are immunoreactive for substance P (Gamboa-
lamina I cells projecting to that nucleus: an electron Esteves, F. O. et al., 2001a).
microscopic tracer study in the rat. Neuroscience 55,
10931106.
35.3.3.5 Response properties
By the use of the c-fos approach, spino-NTS neurons
located in lamina I were shown to be activated by
neuronal population and participate in a similar pro-
cutaneous and visceral noxious stimulation (Menetrey,
portion in the spino-VLM (Lima, D. et al., 1991) and
D. and DePommery, J., 1991; Lima, D. et al., 1994;
the spino-DRt (Lima, D. and Coimbra, A., 1990) path-
Gamboa-Esteves, F. O. et al., 2001b). They belong in
ways. Their relative amount in the NTS system is,
the three neuronal groups that participate in the path-
however, particularly high surpassing the one observed
way (Esteves, F. et al., 1993) irrespective of the kind of
in the lateral spinothalamic system (2530% at the
cutaneous or visceral noxious stimulation employed
spinal enlargements).
(Lima, D. et al., 1994). However, cells activated by
visceral input prevail over those activated by cutaneous
35.3.3.3 Spinal location of ascending input, and neurochemical differences between cuta-
fibers neous- and visceral-activated cells were found
By comparing anterograde tracing produced by (Gamboa-Esteves, F. O. et al., 2001b).
injections centered in the dorsal funiculus or the
dorsolateral fasciculus with that obtained from super- 35.3.3.6 Pathways driven at the target
ficial or deep dorsal horn laminae, it was concluded The caudal NTS projects to several brain areas
that fibers from lamina I neurons course in the dorsal involved in pain processing such as the caudal ventro-
funiculus and fibers from deep dorsal horn neurons in lateral reticular formation (Cobos, A. et al., 2003), the
the dorsolateral fasciculus (Figure 10) (Gamboa- dorsal reticular nucleus (Almeida, A. et al., 2002), the
Esteves, F. O. et al., 2001c). rostral ventromedial medulla (Sim, L. J. and Joseph,
S. A., 1994), the PBN (Cechetto, D. F. et al., 1985), the
35.3.3.4 Neurotransmitters PAG (Bandler, R. and Tork, I., 1987; Herbert, H. and
A small fraction of the neurons projecting to the NTS Saper, C. B., 1992), and the hypothalamus (Reis, L. C.
was found, in immunocytochemical studies, to con- et al., 2000). The NTS is also connected with the
tain dynorphin in lamina I, VIP in the lateral spinal medullary vasopressor (Agarwal, S. K. and Calaresu,
492 Ascending Pathways: Anatomy and Physiology

Spino-NTS (commissural subnucleus)

Spino-NTS (lateral subnucleus)

Figure 10 Diagram representing the spinal laminae of origin, ascending course in the spinal cord, and areas of termination
of the spino-NTS pathway. Note the deep dorsal horn location of neurons terminating in the lateral subnucleus. Brain and
spinal cord photomicrographs were adapted from Paxinos, G. and Watson, C. 1998. The Rat Brain in Stereotaxic
Coordinates, 4th edn. Academic Press.

F. R., 1990) and vasodepressor (VLM) areas (Cobos, A. Nahin, R. L. et al., 1986), cat (Abols, I. A. and
et al., 2003). Although the NTSVLM pathway is Basbaum, A. I., 1981; Ammons, W. S., 1987), and mon-
potentially involved in nociceptive/cardiovascular key (Haber, L. H. et al., 1982; Kevetter, G. A. et al., 1982).
integration (Tavares, I. et al., 1997), different VLM The pathway is mainly contralateral, although a
neurons are likely to mediate the antinociceptive and large number of ipsilaterally and bilaterally project-
vasodepressive effects (Lima, D. et al., 2001). ing cells have also been reported (Figure 12) (Haber,
The caudal NTS also gives rise to projections des- L. H. et al., 1982; Thies, R. and Foreman, R. D., 1983;
cending directly to the spinal cord (Loewy, A. D. and Foreman, R. D. et al., 1984; Ammons, W. S., 1987).
Burton, H., 1978; Mtui, E. P. et al., 1993; Tavares, I. and Cells are distributed throughout the entire length of
Lima, D., 1994). NTS fibers targeting the dorsal horn the spinal cord, but are much more numerous at the
originate in the commissural subnucleus and terminate upper cervical level due to an important contribution
in superficial laminae (Tavares, I. and Lima, D., 1994). of the ipsilateral ventral horn (Kevetter, G. A. and
Willis, W. D., 1983). Scattered neurons were
observed in lamina I (Foreman, R. D. et al., 1984;
35.3.4 Rostral Ventromedial Medulla Leah, J. et al., 1988) and the lateral spinal nucleus in
35.3.4.1 Spinal laminae of origin and sites the rat (Leah, J. et al., 1988). The axonal termination
of termination domain distributes through the nucleus reticularis
Spinal cells projecting in the spino-RVM pathway gigantocellularis and the nucleus paragigantocellu-
predominate in laminae V, VII, VIII, and X laris (Figure 12) (Bowsher, D. and Westman, J., 1970;
(Figure 12) in the rat (Maunz, R. A. et al., 1978; Kerr, F. W. L., 1975; Peschanski, M. and Besson, J. M.,
Kevetter, G. A. and Willis, W. D., 1982; Kevetter, G. 1984). In the cat and monkey, fibers originating in
A. et al., 1982; Kevetter, G. A. and Willis, W. D., 1983; lamina I were seen to extend medially through the
Chaouch, A. et al., 1983; Peschanski, M. and Besson, J. reticular formation to terminate in nucleus raphe
M., 1984; Nahin, R. L. and Micevych, P. E., 1986; magnus (Craig, A. D., 1995).
 Ascending Pathways: Anatomy and Physiology 493

(a) (b) Neurons in lamina X were also shown to contain

cholecystokinin (CCK), somatostatin and, in fewer
numbers, bombesin (Leah, J. et al., 1988). In the lateral
spinal nucleus, a relatively large neuronal population
is immunoreactive to VIP, while bombesin and
dynorphin are expressed in just a few neurons
(c) (d)
(Leah, J. et al., 1988). Scarce somatostatin-immunor-
eactive neurons occur in lamina V (Leah, J. et al.,
1988).

35.3.4.5 Response properties

Spino-RVM neurons are excited by stimulation of A
(e) (f)
and C fibers from the skin, muscles, and viscera
(Maunz, R. A. et al., 1978; Thies, R. and Foreman, R.
D., 1983; Foreman, R. D. et al., 1984; Ammons, W. S.,
1987). Both noxious and innocuous stimulation are
effective, but the majority of RVM-projecting neu-
Figure 11 Lamina I neurons of the flattened (a, b), rons are nociceptive (Maunz, R. A. et al., 1978; Haber,
pyramidal (c, d), and fusiform (e, f ) types retrogradely L. H. et al., 1982; Thies, R. and Foreman, R. D., 1983;
labeled with cholera toxin subunit B (CTb) from the NTS, in
Foreman, R. D. et al., 1984). Neurons responding
parasagittal view. In (b, d, and f ), retrogradely labeled
neurons present immunoreactivity for calbindin (green). only to cutaneous input belong in the LT, WDR,
Scale bar 50 mm. (Adapted from figures 3 and 5 of and NS classes (Fields, H. L. et al., 1977). Receptive
Gamboa-Esteves, F. O., Kaye, J. C., McWilliam, P. N., Lima, fields vary from limited (Fields, H. L., et al., 1977) to
D., and Batten, T. F. 2001a. Immunohistochemical profiles large and complex, often including inhibitory
of spinal lamina I neurons retrogradely labelled from the
regions (Fields, H. L. et al., 1977; Maunz, R. A. et al.,
nucleus tractus solitarii in rat suggest excitatory projections.
Neuroscience 104, 523538). 1978; Cervero, F. and Wolstencroft, J. H., 1984).
Neurons responding to stimulation of deep struct-
ures are particularly numerous. They are mostly
35.3.4.2 Structural types of neurons activated by deep noxious stimulation (Cervero, F.
involved and Wolstencroft, J. H., 1984) and may receive
Spino-RVM neurons located in lamina VII in the convergent noxious or innocuous input from the
monkey were structurally relatively small as compared skin (Fields, H. L. et al., 1977; Cervero, F. and
to spinothalamic neurons, with multipolar or, less Wolstencroft, J. H., 1984). Neurons responsive to
frequently, fusiform or round perikarya and long den- cutaneous, deep, and visceral noxious stimulation
drites across the lateromedial axis (Kevetter, G. A. have also been reported (Foreman, R. D. et al., 1984;
et al., 1982). Blair, R. W. et al., 1984a, 1984b; Ammons, W. S.,
1987). The majority of these cells belong in the
35.3.4.3 Spinal location of ascending high-threshold class, the remaining being WDR neu-
fibers rons (Thies, R. and Foreman, R. D., 1983; Foreman,
Spino-RVM axons course in the contralateral ven- R. D. et al., 1984). Similar to pure somatic neurons,
trolateral quadrant in the spinal cord (Figure 12) and spinoreticular neurons with visceral input present
follow ventrolaterally in the caudal medulla oblon- either well-delimited or complex receptive fields,
gata before they turn medially to reach their sites of the latter including areas resulting in innocuous- or
termination (Rossi, G. F. and Brodal, A., 1957; noxious-evoked inhibition (Thies, R. and Foreman,
Anderson, F. D. and Berry, C. M., 1959; Mehler, W. R. D., 1983; Foreman, R. D. et al., 1984). Ten percent
R. et al., 1960; Kerr, F. W. L., 1975; Nahin, R. L. et al., to 20% of the neurons projecting to the RVM also
1986). target the lateral thalamus (Haber, L. H. et al., 1982;
Foreman, R. D. et al., 1984).
35.3.4.4 Neurotransmitters
Immunocytochemical staining for enkephalin was 35.3.4.6 Pathways driven at the target
observed in spino-RVM neurons located in laminae The RVM was shown to project to the medial thala-
VII and X (Nahin, R. L. and Micevych, P. E., 1986). mus, including the intralaminar complex (Giesler, G.
494 Ascending Pathways: Anatomy and Physiology

Spino-RVM

Figure 12 Diagram representing the spinal laminae of origin, ascending course in the spinal cord, and areas of termination
of the spino-RVM pathway. Brain and spinal cord photomicrographs were adapted from Paxinos, G. and Watson, C. 1998.
The Rat Brain in Stereotaxic Coordinates, 4th edn. Academic Press.

J. et al., 1981b; Peschanski, M. and Besson, J. M., 1984), such putative nociceptive pathways were not thor-
and hence was proposed to function as a relay station oughly investigated. This section will, therefore, deal
in a spinoreticulothalamic pathway involved in motor only with the spino-PBN system.
responses to noxious stimulation. It is also at the
origin of an important pain control descending path-
way that terminates in the superficial and deep dorsal 35.4.1 Parabrachial Nuclei
horn and includes pain inhibitory and facilitatory
neurons (Fields, H. L. et al., 1995; Fields, H. L., 2000). 35.4.1.1 Spinal laminae of origin and sites
of termination
Injections centered in the PBN in both the rat
(Cechetto, D. F. et al., 1985; Lima, D. and Coimbra,
35.4 Spinopontine Pathways A., 1989; Hylden, J. L. et al., 1989; Menetrey, D. and
DePommery, J., 1991; Traub, R. J. and Murphy, A.,
The search for spinal fibers terminating in the brain- 2002) and cat (Panneton, W. M. and Burton, H., 1985;
stem (Craig, A. D., 1995), in particular in Hylden, J. L. K. et al., 1986a; 1986b) produce dense
catecholaminergic nuclei (Westlund, K. N. and bilateral retrograde labeling in lamina I, mainly near
Craig, A. D., 1996), revealed various pontine spinal the dorsal root entry zone, as well as in the lateral
targets, which include the ventrolateral pons (A5), reticular portion of lamina V and laminae VIII and X
the locus coeruleus (A6), the subcoerulear region, (Figure 13). Additional labeling was observed in the
the KollikerFuse nucleus, and the PBN intermediolateral column at thoracic levels, and in the
(Figure 13). These studies were, however, focused parasympathetic column at sacral levels (Menetrey, D.
on lamina I ascending fibers, and, except for the PBN, and DePommery, J. 1991). Spino-PBN neurons with
 Ascending Pathways: Anatomy and Physiology 495

Spino-PBN

Figure 13 Diagram representing the spinal laminae of origin, ascending course in the spinal cord, and areas of termination
of the spino-PBN pathway. Note the preferential lateral location of dorsal horn neurons. Spinal axonal termination areas in
the locus coeruleus (LC), nucleus subcoeruleus (subCA) and the A5 noradrenergic group are also represented. Brain and
spinal cord photomicrographs were adapted from Paxinos, G. and Watson, C. 1998. The Rat Brain in Stereotaxic
Coordinates, 4th edn. Academic Press.

axonal collaterals to the lateral thalamus were (although extending to the cuneiform nucleus)
observed in all spinal areas with a marked prevalence belonged in the fusiform (6570%) and pyramidal
in lamina I (Hylden, J. L. K. et al., 1985; Hylden, J. L. (3035%) groups (Lima, D. and Coimbra, A., 1989).
et al., 1989). A large percentage of spino-PBN neurons In the cat, lamina I neurons antidromically activated
also project to the VLM (Spike, R. C. et al., 2003). from mesencephalic sites with similar location and
The areas of termination of spinal fibers in the intracellularly stained resembled fusiform neurons
PBN are distributed bilaterally through the dorsal (Figure 14) both from the description of their long-
part of the lateral parabrachial nucleus, namely the itudinally extended spiny dendritic arbors and from
dorsal, central, internal, and superior lateral subnuclei their camera lucida drawings (Hylden, J. L. K. et al.,
and the KollikerFuse (Figure 13) (Cechetto, D. F. 1986a; see Figure 5).
et al., 1985; Blomqvist, A. et al., 1989; Slugg, R. M. and
Light, A. R., 1994). The medial and ventral lateral
35.4.1.3 Spinal location of ascending
subnuclei are not targeted by spinal axons (Cechetto,
fibers
D. F. et al., 1985). No topographical arrangement has
been disclosed (Blomqvist, A. et al., 1989). In the cat, spino-PBN fibers course bilaterally in the
dorsolateral fasciculus and ipsilaterally in the ven-
trolateral and ventral funiculi (Figure 13) (Hylden, J.
35.4.1.2 Structural types of neurons L. K. et al., 1986b; Hylden, J. L. et al.. 1989). Fibers
involved originated in lamina I were shown to ascend through
Lamina I neurons labeled retrogradely in the rat the dorsal aspect of the dorsolateral fasciculus
following CTb injections centered in the PBN (Hylden, J. L. K. et al., 1986b). About one-fifth of
496 Ascending Pathways: Anatomy and Physiology

(a) (b) et al., 2003), and most lamina I neurons expressing the
NK1 receptor and receiving synaptic contacts from
TRPV1 immunoreactive primary afferent fibers pro-
ject to the PBN (Hwang, S. J. et al., 2003). Lamina I
PBN-projecting Giant cells (lamina I cells three
times larger than the remaining, amounting to about
5% in each lamina I structural group) (Lima, D. and
Coimbra, A., 1983, 1986) of the pyramidal type were
shown to lack the NK1 receptor and exhibit instead
the glycine receptor-associated protein gephyrin
(Puskar, Z. et al., 2001). These cells are apposed by
nitric oxide synthase and GABA-containing axonal
boutons (Puskar, Z. et al., 2001). In the deep dorsal
horn, about 60% of NK1-immunoreacive neurons
project to the PBN (Todd, A. J. et al., 2000).

35.4.1.5 Neurotransmitters
Around half of the lamina I neuronal population
projecting to the PBN immunostains for either
dynorphin or enkephalin. Staining of sequential sec-
tions did not reveal co-localization of the two
peptides (Standaert, D. et al., 1986). According to
Lima and colleagues (Lima, D. and Coimbra, A.,
1989; Lima, D. et al., 1993), lamina I enkephalinergic
PBN-projecting cells should belong in the pyramidal
group, whereas dynorphinergic cells could be either
pyramidal or fusiform. Lumbosacral neurons project-
ing both ipsi- and contralaterally, mainly from lamina
I and the lateral spinal nucleus, are immunoreactive
for calbindin (Menetrey, D. et al., 1992b).

35.4.1.6 Response properties

Figure 14 Lamina I PBN-projecting neurons retrogradely Neurons antidomically activated from the PBN in
labeled with CTb (a) or intracellularly stained during lamina I of the lumbar spinal cord of the rat (Bester,
antidromic activation (b), in horizontal (a) and parasagittal (b)
H. et al., 2000) and cat (Hylden, J. L. K. et al., 1985;
views. In (a), arrows point to thin distal dendritic branches. In
(b), the arrow points to the axon and the open arrow to the Hylden, J. L. K. et al., 1986a) belong mostly in the NS
cell body. Scale bars 30 mm. (a) Adapted from figure 4 of class (7590%), the remaining being WDR neurons.
Lima, D. and Coimbra, A. 1989. Morphological types of They present extremely low spontaneous activity
spinomesencephalic neurons in the marginal zone (lamina and small receptive fields, respond to stimulation of
I) of the rat spinal cord, as shown after retrograde
A and C primary afferent fibers and conduct in the
labeling with cholera toxin subunit B. J. Comp. Neurol.
279, 327339. (b) Adapted from figure 5 of Hylden, J. L. K., CA range (Hylden, J. L. K. et al., 1986a; Bester, H.
Hayashi, H., Dubner, R., and Bennett, G. J. 1986a. et al., 2000). The large majority respond to both
Physiology and morphology of the lamina I mechanical and heat-noxious stimulation and a few
spinomesencephalic projections. J. Comp. Neurol. 247, also to noxious cold stimulation. C-fos studies identi-
505515).
fied thoracolumbar spino-PBN neurons located
preferentially in the superficial dorsal horn that
lamina I neurons project bilaterally along the spinal were activated by visceral input (Menetrey, D. and
cord (Hylden, J. L. K. et al., 1986a). DePommery, J., 1991; Traub, R. J. and Murphy, A.,
2002). In whole-cell patch-clamp recordings, most
35.4.1.4 Surface receptors lamina I neurons projecting to the PBN present a
Most spino-PBN neurons located in lamina I express gap firing pattern, with a voltage-dependent delay in
the NK1 receptor (Todd, A. J. et al., 2000; Spike, R. C. action potential firing, which was only shared by part
 Ascending Pathways: Anatomy and Physiology 497

of the neurons projecting to the PAG and not by are likely to deal with different aspects of pain
neurons that were not labeled from these two sites processing.
(Ruscheweyh, R. et al., 2004).
35.5.1 Periaqueductal Gray
35.4.1.7 Pathways driven at the target
By combining antidromic activation from the thala- 35.5.1.1 Spinal laminae of origin and sites
mus with orthodromic activation from the periphery, of termination
Bourgeais and coworkers (Bourgeais, L. et al., 2001b) Neurons of origin of the spino-PAG pathway are
demonstrated that neurons in the parabrachial inter- located in lamina I, the reticular region of laminae
nal lateral nucleus responding exclusively, with IVV, laminae VIVIII, lamina X, and the lateral
sustained firing, to noxious stimulation of large spinal nucleus (Figure 15) in the rat (Menetrey, D.
receptive fields, project to the paracentral thalamic et al., 1982; Beitz, A. J., 1982; Liu, R. P., 1983; Swett, J.
nucleus. This spino-PBN-paracentral thalamic path- E. et al., 1985; Yezierski, R. P., 1988; Lima, D. and
way was claimed to be responsible for triggering the Coimbra, A., 1989; Yezierski, R. P. and Broton, J. G.,
aversive reactions to pain at the prefrontal cortex. By 1991; Yezierski, R. P. and Mendez, C. M., 1991), cat
the use of a similar approach as well as by anatomical (Wiberg, M. and Blomqvist, A., 1984; Yezierski, R. P.,
tracing, a spino-PBN-amygdaloid pathway with 1988), and monkey (Trevino, D. L., 1976; Mantyh, P.
relay neurons in the external pontine parabrachial W., 1982; Wiberg, M. et al., 1987; Yezierski, R. P.,
area and terminating in the lateral capsular division 1988; Zhang, D. et al., 1990). In the cat, cells located
of the central nucleus of the amygdala was demon- in lamina I were found to account for the majority of
strated (Ma, W. and Peschanski, M., 1988; Bernard, J. spino-PAG neurons in the cervical and lumbar enlar-
F. and Besson, J. M., 1990). This pathway was con- gements, but only to around 30% in the remaining
firmed by retrograde transneuronal tracing from the spinal segments (Mouton, L. J. et al., 2001). In the rat,
amygdala with pseudorabies virus, and found to ori- an additional important projection, apparently exclu-
ginate mainly in lamina I neurons (Jasmin, L. et al., sive of this pathway, originates from neurons located
1997). inside the white matter overlying lamina I, at the
dorsal funiculus (Lima, D. and Coimbra, A., 1989).
Projections originated in the dorsal horn are
mainly contralateral, especially from lamina I, the
35.5 Spinomesencephalic Pathways lateral spinal nucleus, and the dorsal funiculus
(Trevino, D. L., 1976; Wiberg, M. et al., 1987; Lima,
Spinomesencephalic pathways target a multitude of D. and Coimbra, A., 1989), although a significant
regions located close to each other, which include the ipsilateral projection from lumbosacral spinal seg-
PAG, the intercollicular nucleus, the superior collicu- ments has been reported (Menetrey, D. et al.,
lus, the cuneiform nuclei, the posterior and anterior 1992b). Projections originated in lamina X and the
pretectal nuclei, and the nucleus of Darkschewitsch ventral horn are bilateral (Figure 15) (Trevino, D. L.,
(Wiberg, M. and Blomqvist, A., 1984; Bjorkeland, M. 1976; Wiberg, M. et al., 1987; Lima, D. and Coimbra,
and Boivie, J., 1984; Yezierski, R. P., 1988). The A., 1989). The upper cervical cord makes a major
PAG itself, the major target of spinofugal mesence- additional bilateral contribution both from the ven-
phalic pathways, has its spinal afferents distributed tral horn and the lateral cervical nucleus (Yezierski,
through several areas, each one playing particular R. P. and Mendez, C. M., 1991; Mouton, L. J. and
integrative roles (Yezierski, R. P., 1988). Most retro- Holstege, G., 2000). Spino-PAG neurons were shown
grade tracing studies that refer to the spino-PAG to leave axonal collaterals in the DRt, RVM, and
pathway are based on injections that encompass dif- locus coeruleus in the rat (McMahon, S. B. and
ferent areas of the PAG as well as part of the above Wall, P. D., 1985; Pechura, C. and Liu, R., 1986),
referred neighbor regions. Since the PAG is the and to collateralize a lot within the mesencephalon
principal site of termination of the spinomesencepha- (Hylden, J. L. K. et al., 1985; Yezierski, R. P. and
lic tract, this chapter, will focus on the spino-PAG Schwartz, R. H., 1986). Projections to both the
pathway without separating the various mesencepha- mesencephalon and thalamus were reported in the
lic targets, as a large number of studies addressing rat (Harmann, P. A. et al., 1988; Yezierski, R. P. and
this ascending system do. However, it should be kept Mendez, C. M., 1991), cat (Hylden, J. L. K. et al.,
in mind that it comprises several parallel systems that 1986a; Yezierski, R. P. and Broton, J. G., 1991) and
498 Ascending Pathways: Anatomy and Physiology

Spinomesencephalic

Figure 15 Diagram representing the spinal laminae of origin, ascending course in the spinal cord, and areas of termination
of the spinomesencephalc pathway. (Only the caudal termination area is represented) Brain and spinal cord photomicro-
graphs were adapted from Paxinos, G. and Watson, C. 1998. The Rat Brain in Stereotaxic Coordinates, 4th edn. Academic
Press.

monkey (Price, D. D. et al., 1978; Yezierski, R. P. et al., Bjorkeland, M. and Boivie, J., 1984; Wiberg, M. et al.,
1987; Zhang, D. et al., 1990). Thalamic sites of termi- 1987; Yezierski, R. P., 1988), and monkey (Kerr, F. W.
nation are mainly located in the ventrobasal complex, L., 1975; Wiberg, M. et al., 1987; Yezierski, R. P., 1988).
but collateralization to the posterior complex and The termination pattern is very similar in the three
medial thalamic nuclei has also been observed species (Figure 15). With the exception of the nucleus
(Yezierski, R. P. and Mendez, C. M., 1991). of Darkschewitsch, terminal arborizations are sparse
The mesencephalic sites of termination of the in the most rostral part of the mesencephalon
fibers ascending from the spinal cord and the spinal (Yezierski, R. P., 1988; Lima, D. and Coimbra, A.,
trigeminal nucleus were depicted in the rat (Yezierski, 1989). Fibers are mainly distributed through the mid-
R. P., 1988), cat (Wiberg, M. and Blomqvist, A., 1984; dle and caudal part of the PAG, nucleus cuneiformis,
 Ascending Pathways: Anatomy and Physiology 499

deep and intermediate gray layers of the superior 1988), with additional labeling dorsally to the aque-
colliculus, and intercollicular nucleus (Wiberg, M. duct (Yezierski, R. P., 1988).
et al., 1987; Yezierski, R. P., 1988). In the caudal- This is an interesting finding in the light of data
most PAG of the monkey, but not in the rat and cat showing that the dorsolateral/lateral PAG is
(Figure 16), spinal afferents contribute to two distinct involved in aversive/defense behavior and vasopres-
dorsolateral and ventrolateral dense arborizations, sor responses, and the vantrolateral PAG in
while immediately rostrally, in the intercollicular immobility, positive reinforcing, and vasodepression
region, they concentrate in a sole domain located (Lovick, T. A., 1993).
laterally (Wiberg, M. et al., 1987; Yezierski, R. P., The possibility that the spino-PAG pathway is a
composite of multiple pathways subserving the var-
ious functions in which the PAG is involved is
(a)
supported by studies showing that different regions
IC in the PAG receive afferents from distinct spinal
neuronal populations (Keay, K. A. and Bandler, R.,
1992; VanderHorst, V. G. J. M. et al., 1996; Mouton, L.
AQ
J. and Holstege, G.., 2000). Neurons at spinal seg-
ments C1C3 that project to the lateral part of the
PAG are mainly located in lamina I, whereas those
projecting to the ventrolateral part of the PAG pre-
vail in laminae VIIVIII (Keay, K. A. and Bandler, R.,
1992). In the lumbosacral spinal cord, neurons in
medial lamina VII and lamina VIII terminate in the
(b) lateral part of the lateral PAG and adjacent tegmen-
IC
tum, whereas neurons distributed thoughout laminae
CG I and V terminate diffusely in the dorsal and lateral
PAG (VanderHorst, V. G. J. M. et al., 1996). In the cat,
Mouton and Holstege (Mouton, L. J. and Holstege,
G., 2000) described five distinct spinal neuronal
AQ groups based on their clustering pattern in the spinal
cord and termination pattern in the PAG: (1) neurons
located in laminae I and V along the entire length of
the spinal cord and terminating in all parts of the
intermediate and caudal PAG; (2) neurons located
bilaterally in lateral laminae VIVII and dorsolateral
(c) lamina VIII of segments C1C3 and terminating in
IC the ventrolateral and lateral part of the entire PAG
and deep tectum; (3) neurons located in lamina X of
the thoracic and upper lumbar cord and terminating
in the ventrolateral and lateral PAG and deep tec-
tum; (4) neurons located in medial laminae VIVII of
AQ segments L5S3 and terminating in the lateral and
ventrolateral intermediate and caudal PAG; and (5)
neurons located laterally in lamina I of segments L6
S2 and laminae VVII and X of segments S1S3 and
terminating in the medial part of the ventrolateral
Figure 16 Anterograde labeling in the caudal midbrain intermediate and caudal PAG. According to electro-
following injection of wheat grem agglutininhorseradish physiologcal studies using antidromic activation
peroxidase (WGA-HRP) in the lumbosacral spinal cord of (Yezierski, R. P. and Schwartz, R. H., 1986), spinal
the rat (a), cat (b), and monkey (c). AQ, Cerebral aqueduct;
cells projecting to the rostral-most part of the PAG
CG , Periaqueductal gray; IC, Inferior colliculus. Adapted
from figure 3 of Yezierski, R. P. 1988. Spinomesencephalic
are located more ventrally, in laminae VVII, than
tract: projections from the lumbosacral spinal cord of the those projecting to the intercollicular and caudal
rat, cat, and monkey. J. Comp. Neurol. 267, 131146. levels, to where lamina I neurons project.
500 Ascending Pathways: Anatomy and Physiology

35.5.1.2 Structural types of neurons 35.5.1.5 Neurotransmitters

involved In the lateral spinal nucleus and lamina X, neurons
Spinomesencephalic neurons were characterized as containing various neuropeptides and projecting to a
to the size and shape of the soma both in the rat and mesencephalic area centered in the PAG (but
cat (Menetrey, D. et al., 1980; VanderHorst, V. G. J. extending to the parabrachial nuclei) were observed
M. et al., 1996). Neurons in lamina I are smaller than (Leah, J. et al., 1988). Neurons in the lateral spinal
in other laminae and present oval to fusiform soma in nucleus were immunoreactive for VIP, bombesin,
transverse view (VanderHorst, V. G. J. M. et al., 1996). and substance P, while those located in lamina X
Neurons in the lateral spinal nucleus and lamina X were immunoreactive for bombesin and enkephalin.
present oval to fusiform soma of variable sizes. Deep A few VIP-immunoreactive neurons were located in
dorsal horn and ventral horn neurons are large and lamina I. At lumbosacral spinal levels, calbindin
multipolar (Menetrey, D. et al., 1980; VanderHorst, V. immunoreactive PAG-projecting neurons were
G. J. M. et al., 1996). observed bilaterally in all spinal areas of origin of
A detailed structural characterization was obtained the pathway, with a particularly high concentration
for lamina I neurons retrogradely labeled with CTb in in lamina I and the lateral spinal nucleus (Menetrey,
the rat (Lima, D. and Coimbra, A., 1989). Large D. et al., 1992b).
numbers of fusiform and pyramidal neurons were
shown to participate. However, all injections were 35.5.1.6 Response properties
directed to the ventrolateral caudal PAG and encom- The response properties of spinal neurons projecting
passed part of the PBN. Injections hitting mainly the to the PAG were recorded in the rat (Menetrey, D.
PBN failed to stain as many pyramidal neurons as et al., 1980), cat (Yezierski, R. P. and Schwartz, R. H.,
those targeting the ventrolateral PAG, while still 1986; Yezierski, R. P. and Broton, J. G., 1991), and
labeling a relatively large number of fusiform neu- monkey (Yezierski, R. P. et al., 1987). PAG-projecting
rons. Pyramidal neurons were therefore taken as neurons belong in the LT, WDR, and NS classes.
projecting mainly to the ventrolateral PAG and fusi- Both WDR and NS neurons respond to mechanical
form neurons to the PBN (Lima, D. and Coimbra, A., and heat stimuli at the noxious range. WDR neurons
largely prevail over the other neuronal classes, repre-
1989). Some fusiform neurons projecting to the PAG
senting about half of the population recorded. Many
were shown to have myelinated axons and give
WDR cells were found to respond to both cutaneous
off collaterals inside lamina I (Hylden, J. L. K. et al.,
and visceral/deep tissue stimulation (Yezierski, R. P.
1986a).
and Schwartz, R. H., 1986; Yezierski, R. P. et al., 1987).
In the rat, NS neurons are predominant in lamina I,
WDR neurons are distributed through both lamina I
35.5.1.3 Spinal location of ascending
and the deep dorsal horn, and LT cells prevail in the
fibers
deep dorsal horn (Menetrey, D. et al., 1980). In the cat
Fibers of the spinomesencephalic pathway are classi-
and monkey, neurons are distributed evenly through
cally considered to travel in the ventrolateral
the dorsal horn and around the central canal irre-
quadrant of the spinal white matter (Mehler, W. R.
spective of the physiological class they belong to
et al., 1960; Kerr, F. W. L., 1975). More recent data
(Yezierski, R. P. and Schwartz, R. H., 1986;
using antidromic activation revealed that fibers aris-
Yezierski, R. P. et al., 1987).
ing from lamina I decussate near their level of origin C-fos studies (Clement, C. I. et al., 2000; Keay, K.
and course in the dorsal part of the dorsolateral A. et al., 2002) revealed neurons at the thoracic spinal
fasciculus (McMahon, S. B. and Wall, P. D., 1985; cord that project to the rostral ventrolateral PAG to
Hylden, J. L. K. et al., 1986b). be activated by noxious visceral stimulation. Neurons
at both the lumbosacral and upper cervical spinal
cord and projecting to the caudal ventrolateral PAG
35.5.1.4 Surface receptors were activated by hind limb muscle noxious stimula-
The majority of spino-PAG neurons in lamina I, but tion. Lamina I neurons projecting to the caudal
not in the deep dorsal horn, express the NK1 recep- ventrolateral PAG at the lumbar enlargement and
tor (Todd, A. J. et al., 2000). Their amounts are, expressing c-fos following either mechanical, ther-
however, smaller than those of neurons projecting mal, or chemical noxious stimulation of the skin or
to the VLM or the PBN (Spike, R. C. et al., 2003). noxious stimulation of the urinary bladder belong in
 Ascending Pathways: Anatomy and Physiology 501

both the fusiform and pyramidal groups (Lima, D. taken as indicative of a PAG role in motor responses,
et al., 1992; Lima, D., 1998). escape/avoidance, aversive versus positive reinfor-
Most spino-PAG neurons, including those few cing, and neuroendocrine and autonomic responses
cells projecting to both the PAG and the ventrobasal to pain (Lovick, T. A., 1993).
complex of the thalamus, present small excitatory A dense descending projection connects the ven-
receptive fields confined to a single limb (Yezierski, trolateral portion of the caudal PAG with the
R. P. et al., 1987). However, neurons with extensive ipsilateral nucleus raphe magnus (NRM) and adja-
and complex receptive fields have also been cent reticular formation, locus coeruleus (LC),
observed, in particular in the upper cervical cord nucleus subcoeruleus, and the ventral reticular for-
and in deep spinal laminae, including lamina X mation of the medulla (Meller, S. T. and Dennis, B. J.,
(Menetrey, D. et al., 1980; Yezierski, R. P. and 1991). Sparse fibers originated in the dorsal PAG and
Schwartz, R. H., 1986; Yezierski, R. P., 1990; superior colliculus also terminate in the locus coer-
Yezierski, R. P. and Broton, J. G., 1991). Both groups uleus/subcoeruleus area (Cowie, R. J. and Holstege,
present complex inhibitory receptive fields and G., 1992). Since PAG projections to the spinal cord
include NS and WDR neurons (Yezierski, R. P. and are limited and restricted to laminae VIIVIII
Schwartz, R. H., 1986; Yezierski, R. P. et al., 1987; (Behbehani, M. M., 1995), the analgesic effects eli-
Yezierski, R. P. and Broton, J. G., 1991). cited from PAG stimulation are likely to be mediated
Spino-PAG lamina I neurons have slow conduct- by these PAG-pontine and PAG-medullary path-
ing velocities, at the A range, while those in the deep ways (Lovick, T. A., 1993).
dorsal horn and ventral horn conduct at the low A
range (Yezierski, R. P. et al., 1987). Neurons in the
lateral spinal nucleus present particularly slow axons,
which belong in the unmyelinated and thin myeli- 35.6 Spinodiencephalic Pathways
nated classes (Menetrey, D. et al., 1980). A recent
study using whole-cell patch-clamp in spinal slices Of the pathways terminating in the diencephalon, the
showed that spino-PAG neurons present either gap- spinothalamic are by far those known for longer and
firing or burst-firing patterns, contrary to neurons therefore more thoroughly investigated. Although a
that were not labeled from either the PAG or the large proportion of the studies dealing with the spi-
PBN (Ruscheweyh, R. et al., 2004). nothalamic system address together the lateral and
medial pathways, it turned clear from clinical
35.5.1.7 Pathways driven at the target (Dejerine, J. and Roussy, G., 1906; Walker, A. E.,
The spinomesencephalic pathway was first uncov- 1942a; Hecaen, H. et al., 1949), electrophysiological
ered as a relay station of the medial spinothalamic (Kenshalo, D. R., Jr. et al., 1979; Giesler, G. J. et al.,
tract. Early anatomical studies revealed a projection 1981b), and anatomical (Boivie, J., 1979) studies that
from the mesencephalon to the intralaminar nuclei of each pathway is engaged in particular aspects of
the thalamus (Bowsher, D., 1957). Later anterograde nociceptive processing. The medial pathway has
tracing studies confirmed this connection as well as been implicated in arousal, motivational, affective,
important projections to the hypothalamus, striatum, and motor responses to pain, and the lateral pathway
and amygdala (Eberhart, J. A. et al., 1985; Meller, S. T. in stimulus discrimination. Accordingly, in this chap-
and Dennis, B. J., 1991). Discrete injections confined ter, the two pathways will be dealt with separately in
to different portions of the PAG in the rabbit showed spite of the difficulties raised by being often assessed
that the ventral portion is the main source of afferent as a whole, particularly in retrograde tracing studies.
systems (Meller, S. T. and Dennis, B. J., 1991). Two There are, nonetheless, common aspects that will be
distinct ascending systems were recognized: a peri- more thoroughly described in the context of the
ventricular system terminating in intralaminar and lateral spinothalamic pathway. Also, a relatively
midline thalamic nuclei and along the hypothalamus, large percentage of spinothalamic neurons (around
and a ventrolateral system terminating in the ventral 15%) of various species and different spinal laminae
tegmental area, ventral thalamus, zona incerta, amyg- projects to both the lateral and medial thalamus
dala, substantia innonimata, lateral preopric nucleus, (Giesler, G. J. et al., 1981b; Kevetter, G. A. and
diagonal band of Broca, and the lateral septal nucleus. Willis, W. D., 1983; Stevens, R. T. et al., 1989; Craig,
This multitude of pathways is likely to reflect the A. D. et al., 1989). These neurons share, however, all
morphofunctional complexity of the PAG and is the properties of lateral spinothalamic neurons
502 Ascending Pathways: Anatomy and Physiology

(Giesler, G. J. et al., 1981b) and are therefore consid- L., 1936). Only much later, with the advent of neu-
ered as part of the lateral spinothalamic pathway. rophysiology and tracing techniques, this pathway
was revealed in detail. Nevertheless, early clinical
studies (Dejerine, J. and Roussy, G., 1906; Melzack,
35.6.1 Lateral Thalamus R. and Casey, K. L., 1968) correlated the lateral
The lateral spinothalamic pathway was the first noci- spinothalamic pathway with the discriminative pro-
ceptive spinofugal pathway described (Edinger, L., cessing of nociceptive input.
1890). Its identification, at the turning of the nine-
teenth century, was based on the observation in 35.6.1.1 Spinal laminae of origin and sites
necropsia tissue from humans, of degenerating pro- of termination
files at the lateral sensory thalamus after ventrolateral The lateral spinothalamic pathway takes origin on
cordotomy that disrupted pain sensation (Quensel, the contralateral spinal cord in laminae I, IVVI,
F., 1898; Kohnstamm, O., 1900; Thiele, F. H. and VIIVIII, X, and the lateral spinal nucleus (Figure 17)
Horsley, V., 1901; Collier, J. and Buzzard, E. F., in the rat, cat, and monkey (Trevino, D. L. and
1903; Foerster, O. and Gagel, O., 1932; Clark, W. E. Carstens, E., 1975; Carstens, E. and Trevino, D. L.,

Lateral spinothalamic

Figure 17 Diagram representing the spinal laminae of origin, ascending course in the spinal cord and areas of termination
of the lateral spinothalamic pathway. Note the cluster appearance of axon termination in the VPL. Brain and spinal cord
photomicrographs were adapted from Paxinos, G. and Watson, C. 1998. The Rat Brain in Stereotaxic Coordinates, 4th edn.
Academic Press.
 Ascending Pathways: Anatomy and Physiology 503

1978b; Willis, W. D. et al., 1979; Giesler, G. J. et al., CTb disclosed an important ipsilateral component
1979a; Berkley, K. J., 1980; Leah, J. et al., 1988; (Lima.D. and Coimbra, A., 1988). Ipsilateral neurons
Lima.D. and Coimbra, A., 1988; Burstein, R. et al., amounted to about half the neurons labeled in the
1990b). contralateral side in the lateral spinal nucleus and the
In laminae I and IV, a somatotopic arrangement deep dorsal horn in almost all the spinal segments
has been described, neurons receiving input from examined. In the ventral horn, ipsilateral neurons
extensor surfaces being located more laterally than equalized in number contralateral neurons, except
neurons receiving input from flexor surfaces (Willis, in the cervical enlargement, where they were more
W. D. et al., 1974). The amount of spinothalamic abundant. Only in lamina I and the intermediate
neurons varies considerably along the rostrocaudal basilar nucleus of Cajal, contralateral neurons largely
extent of the spinal cord due to additional intense surpassed ipsilateral neurons. The need of using a
labeling in particular areas at various spinal levels. very sensitive tracing technique to reveal the ipsilat-
This is the case of the deep dorsal horn contralater- eral neuronal population suggests that these neurons
ally, and the intermediate/ventral horn, bilaterally, may actually be contralaterally projecting neurons
in spinal segments C1C2 (Carstens, E. and Trevino, that send axonal collaterals to the ipsilateral thala-
D. L., 1978a; Carstens, E. and Trevino, D. L., 1978b; mus. However, there is evidence from ventral spinal
Giesler, G. J. et al., 1979a; Lima.D. and Coimbra, A., lesions in primates that nociceptive input is also
1988; Burstein, R. et al., 1990b), and the intermediate conveyed supraspinally in the ipsilateral anterolat-
basilar nucleus of Cajal, contralaterally, in the rat eral quadrant (Vierck, C. J. and Luck, M. M., 1979).
(Giesler, G. J. et al., 1979a; Lima.D. and Coimbra, A., The areas of spinal axon arborization in the lateral
1988; Burstein, R. et al., 1990b). Due to this regional thalamus (Figure 17) were thoroughly studied in
variability, together with the fact that the multiple primates, including the monkey (Mehler, W. R.
spinal groups projecting to the lateral thalamus also et al., 1960; Bowsher, D., 1961; Mehler, W. R., 1966;
project to many other supraspinal targets, the erro- Mehler, W. R., 1969; Kerr, F. W. L. and Lippman, H.
neous assumption that thalamic projections from H., 1974; Boivie, J., 1979; Berkley, K. J., 1980; Mantyh,
certain areas, such as lamina I, to the main lateral P. W., 1983a; Apkarian, A. V. and Hodge, C. J., 1989a)
spinal target, the VPL, are not sufficiently relevant and humans (Mehler, W. R., 1962; Mehler, W. R.,
gained credit (Blomqvist, A. et al., 2000; Craig, A. D. 1974). In these species, as in the rat (Lund, R. D. and
et al., 2002; Klop, E. M. et al., 2004). However, Webster, K. E., 1967; Mehler, W. R., 1969; Zemlan, F.
although the relative participation of lamina I neu- P. et al., 1978; Peschanski, M. et al., 1983; Cliffer, K. D.
rons is below 10% in the cat (Klop, E. M. et al., 2004) et al., 1991), the VPL is the major recipient of spinal
and rat (Burstein, R. et al., 1990b) when the spinal fibers. In the VPL, spinal afferents are somatotopi-
cord is considered as a whole, small relative amounts cally arranged in rostrocaudally oriented clusters so
are only found in segments where additional labeling that axons arriving from the lumbosacral spinal cord
occurs in particular spinal groups, as is the case of the terminate in the lateral part of the nucleus and axons
upper cervical and lumbar cord (Burstein, R. et al., from the cervical enlargement terminate in the med-
1990b). Notably, in the cervical enlargement of the ial part (Boivie, J., 1979; Mantyh, P. W., 1983a). Such
rat, numbers of lamina I spinothalamic neurons equal a somatotopic arrangement supports the ability of the
those in the deep dorsal horn (Burstein, R. et al., lateral spinothalamic pathway to process spatial dis-
1990b). Moreover, many lamina I neurons in the crimination. Other important lateral thalamic areas of
contralateral spinal and medullary dorsal horn of spinal termination are the posterior complex (PO),
the monkey project to the ventrobasal complex of the ventral posteroinferior nucleus (VPI) and the
the thalamus, amounting to one-third of the entire zona incerta (ZI) ((Mehler, W. R., 1974; Boivie, J.,
dorsal horn labeled population (Willis, W. D. et al., 1979; Apkarian, A. V. and Hodge, C. J., 1989a; Cliffer,
2001). K. D. et al., 1991). In the cat, fibers in the lateral
Although the spinothalamic lateral pathway is spinothalamic pathway appear to be fewer and ter-
classically considered to project contralaterally minate in the ZI, the posterior complex, and in a shell
except for the ventral horn in segments C1C2 area surrounding the VPL ventrolaterally (Boivie, J.,
(Trevino, D. L. and Carstens, E., 1975; Carstens, E. 1971; Jones, E. G. and Burton, H., 1974; Berkley, K. J.,
and Trevino, D. L., 1978a; Carstens, E. and Trevino, 1980; Mantyh, P. W., 1983b; Craig, A. D. and Burton,
D. L., 1978b; Willis, W. D. et al., 1979; Giesler, G. J. H., 1985). Recently, Craig and colleagues claimed
et al., 1979a), the use of very sensitive tracers such as that a region located posteromedially to the VPL,
504 Ascending Pathways: Anatomy and Physiology

which they called VMpo, is the site of termination of C1 and C2 (40%) (Lima.D. and Coimbra, A., 1988). In
the lamina I spinothalamic fibers (Craig, A. D. et al., the cat (Zhang, E. T. et al., 1996) and monkey (Zhang, E.
1994, 2002; Blomqvist, A. et al., 2000). However, T. and Craig, A. D., 1997), large tracer injections filling
besides the fact that the so-called VMpo was most both the lateral and medial thalamus, resulted in label-
probably included in the area of termination of spinal ing of fusiform neurons, beyond pyramidal and
and trigeminal thalamic afferents described by flattened neurons. Although the authors explained
Mehler (Mehler, W. R., 1966) in humans, numerous their labeling of fusiform neurons by putative species
retrograde and anterograde studies have proved that differences, fusiform neurons were most probably
lamina I neurons project to many other areas in labeled from the medial thalamus (see further). It
the thalamus (references given earlier), including a should be emphasized that, in the studies by Craig
study by Craig (Craig, A. D., 2003) using anterograde and co-workers (Zhang, E. T. et al., 1996; Zhang, E. T.
tracing with phaseolus vulgaris leucoagglutinin. and Craig, A. D., 1997), flattened neurons were desig-
Moreover, this assumption was based on the dense nated multipolar due to their appearance in horizontal
calbindin-staining observed in the VMpo, but neither view, on which the authors based their observations.
is the calbindin-immunoreactive region restricted to However, this designation is misleading and should be
the VMpo lying within the medial aspect of the avoided since a neuronal group completely distinct in
ventral posterior medial nucleus (Graziano, A. and dendritic geometry and specializations was previously
Jones, E. G., 2004), nor are calbindin-immonoreactive designated multipolar in the rat (Lima, D. and
projecting cells exclusively located in lamina I Coimbra, A., 1986), and subsequently observed in the
(Menetrey, D. et al., 1992b). cat (Galhardo, V. and Lima, D., 1999), monkey (Lima, D.
et al., 2002), and pigeon (Galhardo, V. et al., 2000) as
35.6.1.2 Structural types of neurons well. When compared to flattened neurons, multipolar
involved neurons have ventrally oriented rather than horizontal
Spinothalamic cells were shown, in retrograde label- dendritic arbors and highly spiny rather than smooth
ing studies, to have cell bodies that vary in shape dendritic branches. This kind of misuse of nomencla-
from roundish or flattened to polygonal (Willis, W. ture already led some authors to disregard the
D. et al., 1979). They are mainly small in lamina I and occurrence of flattened neurons as an independent
include cells with fusiform, pyriform, and triangular group in the cat and monkey, based on the results of
shapes in transverse view, beyond the classic retrograde labeling and immunostaining (Yu, X. H.
Waldeyer cells. In deep dorsal horn as well as the et al., 1999), despite the fact that flattened neurons
ventral horn, they are medium to large sized and were clearly identified in both species by the use of
polygonal in shape. Similar data were obtained by Golgi impregnation (Galhardo, V. and Lima, D., 1999;
Apkarian and Hodge (Apkarian, A. V. and Hodge, C. Lima, D. et al., 2002).
J., 1989d), although in this case, tracer injections
included the medial thalamus. Spinothalamic cells 35.6.1.3 Spinal location of ascending
intracellularly stained in laminae IVVIII of the cat fibers
(Meyers, D. E. R. and Snow, P. J., 1982) and monkey Axons of the lateral spinothalamic tract travel in the
(Surmeier, D. J. et al., 1988) presented long dendritic ventral, ventrolateral, and dorsolateral funiculi after
branches that could reach the lateral funiculus, decussating the spinal cord within a short distance
lamina I and lamina X. In the intermediomedial from the cell body (Applebaum, A. E. et al., 1975;
gray matter, spinothalamic cells presented spheroidal Willis, W. D. et al., 1979; Giesler, G. J. et al., 1981a;
cell bodies and narrow dendrites (Milne, R. J. et al., Jones, M. W. et al., 1985; Surmeier, D. J. et al., 1988;
1982). Around the central canal, neuropeptide-con- Stevens, R. T. et al., 1989; Apkarian, A. V. and Hodge,
taining spinothalamic cells had oval cell bodies and C. J., 1989a,b). Spinothalamic axons from lamina I
dendrites oriented transversely, reaching the central cells were shown to project through the dorsolateral
canal medially (Leah, J. et al., 1988). fasciculus in the cat (Apkarian, A. V. et al., 1985;
Injections of CTb confined to the VPL in the rat Stevens, R. T. et al., 1989) and monkey (Apkarian,
revealed that VPL-projecting lamina I neurons belong A. V. and Hodge, C. J., 1989a,b,c). Neurons located in
in the pyramidal and flattened groups (Lima.D. and the deep dorsal horn and ventral horn project
Coimbra, A., 1988). Pyramidal cells prevailed over through the ventrolateral fasciculus and ventral funi-
flattened cells in the cervical and lumbar enlargements culus (Stevens, R. T. et al., 1989; Apkarian, A. V. and
(70% and 77%, respectively), the reverse occurring at Hodge, C. J., 1989b; Zhang, X. J. et al., 2000). In the
 Ascending Pathways: Anatomy and Physiology 505

ventrolateral quadrant of the spinal cord white mat- 1988) were observed in the lateral spinal nucleus at
ter, axons are arranged somatotopically so that those the lumbar cord. In lamina X, neurons immunoreac-
originating in more caudal levels are located dorso- tive for bombesin (Leah, J. et al., 1988), CCK (Ju, G.
laterally to the more rostral ones (Horrax, G., 1929; et al., 1987; Leah, J. et al., 1988), and galanin (Ju, G.
Foerster, O. and Gagel, O., 1932; Hyndman, O. R. and et al., 1987) were described. Galanin and CCK were
Van Epps, C., 1939; Walker, A. E., 1940; Applebaum, seen to co-localize in lamina X neurons projecting
A. E. et al., 1975). The termination sites of the dorso- to the VPL (Ju, G. et al., 1987). Neurons containing
lateral and ventrolateral fibers are equally distributed glutamate or glutaminase were described in the
in the lateral thalamus, except for VPI and the ZI lateral trigeminothalamic system in areas where
whose spinal afferents course mainly in the dorsolat- WDR and low-threshold neurons predominate
eral fasciculus and the ventral spinal quadrant, (Magnusson, K. R. et al., 1987). Calbindin was claimed
respectively (Apkarian, A. V. and Hodge, C. J., to be present in the majority of lamina I spinothala-
1989a). mic neurons of all structural groups projecting to the
thalamus (Craig, A. D. et al., 2002), although antero-
35.6.1.4 Surface receptors grade tracing combined with immunocytochemical
Enkephalin immunoreactive varicosities were shown staining failed to reveal calbindin-immunostaining
to establish asymmetric synaptic contacts upon in lamina I axons terminating in the thalamus
medullary and spinal neurons retrogradely labeled (Graziano, A. and Jones, E. G., 2004).
from large HRP injections centered in the lateral
thalamus of the cat and monkey (Ruda, M. A. et al., 35.6.1.6 Response properties
1984). These neurons make up 30% of lamina I and Spinothalamic cells were shown to present back-
50% of lamina V labeled neurons. In transverse ground activity at variable firing rates depending on
sections, neurons present bipolar configuration in the species and their laminar location in the spinal
lamina I (equivalent to flattened neurons of cord. Only few lamina I cells present background
Lima, D. and Coimbra, A., 1986) and multipolar activity (Craig, A. D. and Kniffki, K. D., 1985) in the
configuration in lamina V (Ruda, M. A. et al., 1984). cat as compared to lamina I cells in the monkey
Spinothalamic neurons exhibiting immunostaining (Ferrington, D. G. et al., 1987) and to laminae IVV
for the NMDA receptor (Zou, X. Y. et al., 2000) and cells in both species (Giesler, G. J. et al., 1981b;
metabotropic glutamate receptor subtype 1 Ferrington, D. G. et al., 1986). Most spinothalamic
(mGluR1) (Millis, C. D. and Hulsebosch, C. E., cells respond to stimulation of C primary afferent
2002) have been described in studies. A study focused fibers (Chung, J. M. et al., 1979), but many of them
on lamina I neurons retrogradely labeled from large respond to volleys in A and A fibers of somatic
injections comprising both the lateral and medial nerves as well (Foreman, R. D. et al., 1975; Beall, J. E.
thalamus revealed NK1 receptors in flattened (called et al., 1977; Chung, J. M. et al., 1979). Spinothalamic
multipolar by the authors) and pyramidal neurons, cells also respond to A and C-fiber volleys in visceral
the latter being relatively few, however (Yu, X. H. nerves (Foreman, R. D. and Weber, R. N., 1980; Blair,
et al., 1999). This was taken as supporting the non- R. W. et al., 1981; Foreman, R. D. et al., 1981, 1984;
nociceptive nature of pyramidal neurons, although Rucker, H. K. and Holloway, J. A., 1982; Ammons, W.
large amounts of pyramidal neurons expressing the S., 1987) and to group II, III, and IV muscle afferents
NK1 receptor were observed by other authors (Foreman, R. D. et al., 1979). Lateral spinothalamic
(Todd, A. J. et al., 2002), and pyramidal neurons are neurons were found to be activated by mechanical
c-fos-activated following various kinds of noxious and/or thermal noxious stimulation of the skin in the
stimulation (Lima, D., 1998). rat (Giesler, G. J. et al., 1976), cat (Fox, R. E. et al., 1980;
Ferrington, D. G. et al., 1986), and monkey (Willis, W.
35.6.1.5 Neurotransmitters D. et al., 1974, 1979; Applebaum, A. E. et al., 1975;
Most studies addressing the neurochemical nature of Price, D. D. et al., 1978; Kenshalo, D. R., Jr. et al.,
spinothalamic neurons looked for the presence of 1979; Giesler, G. J. et al., 1981b; Surmeier, D. J. et al.,
neuropeptides in the rat. Neuropeptide-containing 1986a,b; Ferrington, D. G. et al.. 1987) as well as by
lateral spinothalamic neurons were preferentially noxious chemical stimulation (Simone, D. A. et al.,
observed in the lateral spinal nucleus and around 1991) and low-threshold mechanical (Willis, W. D.
the central canal (including lamina X). Both VIP et al., 1974; Applebaum, A. E. et al., 1975; Giesler, G. J.
(Nahin, R. L., 1988) and bombesin (Leah, J. et al., et al., 1976; Price, D. D. et al., 1978) cooling
506 Ascending Pathways: Anatomy and Physiology

(Dostrovsky, J. O. and Craig, A. D., 1996; Craig, A. D. pathway (Foreman, R. D. and Weber, R. N., 1980;
et al., 2001) and warming (Andrew, D. and Craig, A. Blair, R. W. et al., 1981; Foreman, R. D. et al., 1984;
D., 2001) stimulation. Nociceptive neurons belong in Ammons, W. S. et al.. 1984; Ammons, W. S., 1987; Al-
both the NS and WDR classes (Willis, W. D. et al., Chaer, E. D. et al., 1999; Chandler, M. J. et al., 2000).
1974; Giesler, G. J. et al., 1981b; Ferrington, D. G. et al., Curiously, in lamina X, neurons of the postsynaptic
1986). NS neurons are equally distributed throughout dorsal column tract were shown to receive visceroso-
laminae I and IVV, while WDR neurons predomi- matic input and reported to be more numerous than
nate in laminae IVV (Willis, W. D. et al., 1974) and those of the spinothalamic tract (Al-Chaer, E. D. et al.,
neurons responsive to cooling are located in lamina I 1999; Dorsal Columns and Visceral Pain).These neu-
(Craig, A. D. et al., 2001). Thermal-responsive neurons rons belong in the WDR, NS, or high-threshold
belonging in either the NS or the WDR neuronal inhibitory classes, and present cutaneous receptive
classes were shown to be capable of encoding noxious fields that occupy regions to which pain is frequently
heat intensity (Figure 18) irrespective of their loca- referred (Foreman, R. D. and Weber, R. N., 1980;
tion in lamina I or the deep dorsal horn (Kenshalo, D. Foreman, R. D. et al., 1984; Ammons, W. S. et al.,
R., Jr. et al., 1979; Ferrington, D. G. et al., 1986; 1984). Some of these neurons were shown to leave
Surmeier, D. J. et al., 1986a, b). axonal collaterals in the medial medullary reticular
Neurons with convergent input from the skin and formation (Foreman, R. D. et al., 1984). Some spi-
viscera also contribute to the lateral spinothalamic nothalamic neurons receive convergent input from
the skin and deep tissues (Willis, W. D. et al., 1974;
Giesler, G. J. et al., 1981b; Ferrington, D. G. et al.,
1986). These neurons are thought to have a proprio-
50 ceptive function (Milne, R. J. et al., 1982). They are
43 mainly located in the lateral part of lamina V and
30 intermediomedial gray matter (Stillings nucleus)
and respond to either weak or intense cutaneous
10
stimulation (Willis, W. D. et al., 1974).
Lateral spinothalamic neurons normally present
50 ipsilateral receptive fields that vary from very small
47
(less than one digit) to moderate (the entire limb)
30
(Willis, W. D. et al., 1974; Giesler, G. J. et al., 1981b), as
10 well as additional inhibitory receptive fields
Spikes/bin

(Gerhart, K. D. et al., 1981; Giesler, G. J. et al.,

1981b; Ammons, W. S., 1987). In a few cells, conver-
50
51 gent inhibitory cutaneous or visceral receptive fields
30 were reported (Willis, W. D. et al., 1974; Blair, R. W.
et al., 1981; Milne, R. J. et al., 1982). Receptive fields
10 tend to be smaller in high-threshold neurons and in
lamina I neurons (Applebaum, A. E. et al., 1975;
Giesler, G. J. et al., 1981b; Ferrington, D. G. et al.,
50
55 1987). The relatively small size of the receptive fields
30 of lateral spinothalamic neurons favors a role in dis-
criminating the size of the stimulated area (Giesler,
10 G. J. et al., 1981b).
C-fos induction after noxious cutaneous or visc-
18 54 90
eral stimulation was observed in spinothalamic
neurons in laminae I, IIIVII, and X (Palecek, J.
Time (s)
et al., 2003). Curiously, neurons projecting in the
Figure 18 Responses of a lateral spinothalamic neuron to postsynaptic dorsal column pathway were activated
heat stimulation of increasing intensities. Adapted from
in similar proportions by the noxious cutaneous sti-
figure 1 of Surmeier, D. J., Honda, C. N., and Willis, W. D.
1986a. Temporal features of the responses of primate muli and in even higher proportions by the visceral
spinothalamic neurons to noxious thermal stimulation of stimuli (Palecek, J. et al., 2003). In lamina I, both
hairy and glabrous skin. J. Neurophysiol. 56, 351369. flattened and pyramidal neurons projecting to the
 Ascending Pathways: Anatomy and Physiology 507

VPL were c-fos-activated following cutaneous poorly localized unpleasant feeling was abolished
mechanical, thermal, and chemical noxious stimula- (Hecaen, H. et al., 1949).
tion, and, in smaller amounts, following visceral
chemical stimulation (Lima, D. et al., 1992; Lima, D., 35.6.2.1 Spinal laminae of origin and sites
1998). of termination
Spinal neurons projecting to medial thalamic nuclei
35.6.1.7 Pathways driven at the target prevail in the contralateral intermediate and ventral
The VPL is long-known for sending nociceptive gray matter (Figure 19) in the rat (Giesler, G. J. et al.,
input to the parietal somatosensory cortex (Burton, 1979a), cat (Carstens, E. and Trevino, D. L., 1978b),
H. and Jones, E. G., 1976; Whitsel, B. L. et al., 1978; and monkey (Willis, W. D. et al., 1979; Giesler, G. J.
Kenshalo, D. R., Jr. et al., 1980). Combined antero- et al., 1981b). In the cat, although a predominant
grade tracing from the spinal cord and retrograde location in laminae VII and VIII has been described
tracing from the somatosensory cortex in the monkey by some authors (Carstens, E. and Trevino, D. L.,
revealed that overlapping between spinal thalamic 1978b; Comans, P. E. and Snow, P. J., 1981), others
afferents and thalamic neurons projecting to cortical point to a laminar distribution similar to that
areas SI or SII occurs in the VPL, but also in the VPI observed for the lateral spinothalamic pathway,
and PO, the VPL being the area where the number of namely laminae I, IVVI, and VII to X (Stevens, R.
overlapping neurons is smaller (Stevens, R. T. et al., T. et al., 1989; Craig, A. D. et al., 1989). Lamina I was
1993; Shi, T. and Apkarian, A. V., 1995). VPL afferents also shown to be the source of spinal afferents to the
in cortical areas SI and SII are arranged in a somato- nucleus submedius in the cat and monkey (Craig, A.
topic fashion (Burton, H. and Jones, E. G., 1976; D. and Burton, H., 1981; Stevens, R. T. et al., 1989)
Whitsel, B. L. et al., 1978) so that the more posterior and to contribute to the spinal projections to the
the thalamic source of afferents, the more posterior the intralaminar complex in the monkey (Albe-Fessard,
cortical termination sites. Sparse cells in the lateral part D. et al., 1975; Ammons, W. S. et al., 1985). In the rat,
of the VPL and the ventral part of the VPI target the projections from laminae IVVII were recently
cingulated cortex (Apkarian, A. V. and Shi, T., 1998). shown to target the central lateral nucleus, whereas
The PO projects to the granular insular and retro- the medial dorsal nucleus was found to be mainly
insular cortex in primates (Burton, H. and Jones, E. G., innervated bilaterally by the lateral spinal nucleus
1976) and was therefore proposed to be involved in and, to a lesser extent, by contralateral lamina I
nociceptive visceral processing (Cechetto, D. F. and (Gauriau, C. and Bernard.J.F., 2004).
Saper, C. B., 1987). PO neurons projecting to the Spinal afferents ascending to the medial thalamus
anterior insula were seen to clearly overlap with spi- appear to be fewer than those reaching the lateral
nothalamic axonal arborizations (Apkarian, A. V. and thalamus (Mehler, W. R. et al., 1960; Apkarian, A. V.
Shi, T., 1998). Neurons in the VPI also show insular and Hodge, C. J., 1989a). They target similar medial
projections, but with no overlapping. Moreover, neu- thalamic regions in the rat (Lund, R. D. and Webster,
rons projecting to the insula do not overlap with K. E., 1967; Mehler, W. R., 1969; Zemlan, F. P. et al.,
neurons projecting to SI (Apkarian, A. V. and Shi, T., 1978; Cliffer, K. D. et al., 1991), cat (Boivie, J., 1971;
1998). Berkley, K. J., 1980; Mantyh, P. W., 1983a; Craig, A.
D. and Burton, H., 1985), monkey (Mehler, W. R.
et al., 1960; Boivie, J., 1979; Mantyh, P. W., 1983a;
35.6.2 Medial Thalamus
Apkarian, A. V. and Hodge, C. J., 1989a), and humans
The medial spinothalamic pathway was described at (Mehler, W. R., 1962; Mehler, W. R., 1974). These
the middle 1960s as an ascending nociceptive system include the medial dorsal and paraventricular nuclei
especially devoted to the processing of the affective and the intralaminar complex, namely the central
and motivational aspects of pain (Melzack, R. and lateral, center median, paracentral, and parafascicular
Casey, K. L., 1968). While anatomical degeneration nuclei (Figure 19). In the monkey, the medial dorsal
studies demonstrated the termination in medial tha- nucleus (except for its dorsomedial part) and the
lamic nuclei of spinal axons ascending in the central lateral nucleus are the major sites of termina-
ventrolateral quadrant of the spinal cord (Bowsher, tion of spinal fibers (Apkarian, A. V. and Hodge, C. J.,
D., 1957, 1961; Mehler, W. R. et al., 1960; Boivie, J., 1989a). The nucleus submedius is also a consistent
1971), clinical studies revealed that, in patients with site of spinal axon arborization in the cat (Boivie, J.,
lesions centered in the medial thalamus, the painful 1971; Craig, A. D. and Burton, H., 1981; Mantyh, P.
508 Ascending Pathways: Anatomy and Physiology

Medial spinothalamic

Figure 19 Diagram representing the spinal laminae of origin, ascending course in the spinal cord and areas of termination
of the medial spinothalamic pathway. Brain and spinal cord photomicrographs were adapted from Paxinos, G. and
Watson, C. 1998. The Rat Brain in Stereotaxic Coordinates, 4th edn. Academic Press.

W., 1983b; Craig, A. D. and Burton, H., 1985) and 35.6.2.2 Structural types of neurons
monkey (Boivie, J., 1979; Mantyh, P. W., 1983a; involved
Apkarian, A. V. and Hodge, C. J., 1989a), but in Although there are no descriptions of the morphol-
relatively small amounts (Mantyh, P. W., 1983a; ogy of spinal neurons labeled from injections
Apkarian, A. V. and Hodge, C. J., 1989a). restricted to medial thalamic nuclei, data on laminae
Contrary to the lateral thalamus, no somatotopic VIX spinothalamic cells revealed medium to large-
arrangement of spinal terminals could be detected in sized soma of polygonal or occasionally flattened
the intralaminar complex (Boivie, J., 1979), which configuration (Willis, W. D. et al., 1979). Following
agrees with electrophysiological studies (Giesler, G. injections filling both the lateral and medial thalamus
J. et al., 1981b; Guilbaud, G. et al., 1985) as to the in the cat and monkey (Zhang, E. T. et al., 1996;
inadequacy of the medial spinothalamic system to Zhang, E. T. and Craig, A. D., 1997), lamina I neurons
process information related to stimulus discrimination. belonging in the fusiform, pyramidal, and flattened
In the nucleus submedius, however, a somatotopic groups were labeled. As to pyramidal and flattened
organization was described, with fibers from more neurons, the possibility that labeled neurons picked
rostral spinal levels terminating more rostrally in the up the tracer in the lateral thalamus can not be ruled
nucleus (Craig, A. D. and Burton, H., 1985). out, since they were shown to project to the VPL in
 Ascending Pathways: Anatomy and Physiology 509

the rat (Lima, D. and Coimbra, A., 1988). Fusiform

neurons, however, most probably project to medial
thalamic nuclei as they could not be labeled from the
VPL in the rat (Lima, D. and Coimbra, A., 1988).

35.6.2.3 Spinal location of ascending

fibers
Spinal axons targeting the medial thalamus were
early shown to course in the ventral quadrant of the
spinal cord (Mehler, W. R. et al., 1960; Bowsher, D.,
1961; Mehler, W. R., 1969). In the rat this appears to
be the way taken by all spinothalamic axons (Giesler,
G. J. et al., 1981a), whereas in the monkey axons were
shown to distribute through the lateral funiculus also Figure 20 Receptive fields of spinal cells projecting to the
(Giesler, G. J. et al., 1981b; Apkarian, A. V. and medial thalamus in the monkey. Adapted from figure 3 of
Giesler, G. J., Yezierski, R. P., Gerhart, K. D., and Willis, W.
Hodge, C. J., 1989a). Studies using anterograde tra-
D. 1981b. Spinothalamic tract neurons that project to
cing after lesioning either the contralateral spinal medial and/or lateral thalamic nuclei: evidence for a
ventral quadrant or the contralateral dorsolateral fas- physiologically novel population of spinal cord neurons. J.
ciculus (Apkarian, A. V. and Hodge, C. J., 1989a) Neurophysiol. 46, 12851308.
revealed that the termination pattern of fibers cour-
sing in both pathways is similar except for the fibers third in the lateral spinothalamic pathway). Some are
targeting the dorsolateral region of the medial dorsal WDR neurons and a few respond to deep tissue
nucleus, which course mostly in the ventral quadrant. stimulation (Giesler, G. J. et al., 1981b). They nor-
The ventral quadrant also contributes with particu- mally present very large, frequently complex,
larly large amounts of fibers to the central lateral receptive fields that may encompass the entire sur-
nucleus afferents, mainly in its more anterior portion. face of the body (Figure 20).
Inhibitory receptive fields are rare (Giesler, G. J.
et al., 1981b). Viscerosomatic convergence has been
35.6.2.4 Neurotransmitters
reported (Rucker, H. K. and Holloway, J. A., 1982).
Enkephalin (Coffield, J. A. and Mileti, V., 1987;
Nahin, R. L., 1988) and dynorphin (Nahin, R. L.,
1988) were immunodetected in deep dorsal horn and 35.6.2.6 Pathways driven at the target
intermediate gray spinal neurons projecting to the Intralaminar nuclei, in particular the central lateral
medial thalamus. Enkephalin-immunoreactive neurons nucleus, project to widely distributed areas of the
amount to 10% of the entire spinothalamic population cerebral cortex, including sensorimotor areas, and to
(Coffield, J. A. and Mileti, V., 1987). Spinothalamic the basal ganglia (Jones, E. G. and Leavitt, R. Y.,
lamina X neurons immunoreactive for CCK (Ju, G. 1974). Ascending pathways terminating in the intra-
et al., 1987; Leah, J. et al., 1988) and galanin (Ju, G. et al., laminar nuclei have hence been proposed to be
1987) are likely to terminate in the parafascicular involved in motor and arousal nociceptive responses.
nucleus, which was shown to contain fibers immunor- Nociceptive pathways from the medial dorsal
eactive to both neuropeptides (Ju, G. et al., 1987). nucleus and the central lateral nucleus terminate,
respectively, in the anterior cingulated cortex and
35.6.2.5 Response properties frontal motor cortex (Wang, C. C. and Shyu, B. C.,
Neurons projecting to the medial thalamus respond, 2004). The posterior cingulated cortex also receives
although weakly, to activation of A ( to ) and C projections from restricted areas of both, the medial
primary afferent fibers with sustained after- dorsal and the central lateral nuclei (Apkarian, A. V.
discharges (Giesler, G. J. et al., 1981b). Their back- and Shi, T., 1998). These connections are consonant
ground activity is practically nil. They conduct at with the role of the medial thalamic pathway in the
relatively low velocities, which amount to about half emotional aspects of nociception (Wang, C. C. and
those of lateral spinothalamic neurons (Giesler, G. J. Shyu, B. C., 2004). The medial dorsal nucleus and the
et al., 1981b). Most medial spinothalamic neurons nucleus submedius, which are similar in the response
belong in the NS class (around two-third vs. one- properties of their nociceptive neurons (Dostrovsky,
510 Ascending Pathways: Anatomy and Physiology

J. O. et al., 1987), project to adjacent regions in the of the spinal cord (Figure 22) (Burstein, R. et al., 1987,
orbital cortex (Krettek, J. E. and Price, J. L., 1977; 1990a; 1990b) as well as from the spinal trigeminal
Craig, A. D. et al., 1982; Yoshida, A. et al., 1992). nucleus, mainly the pars caudalis (Malick, A. and
Burstein, R., 1998). Neurons in the deep dorsal horn
make up the largest population of spinohypothalamic
35.6.3 Hypothalamus neurons, especially at upper cervical segments
35.6.3.1 Spinal laminae of origin and sites (around 50%), followed by the lateral spinal nucleus
of termination (around 30%), laminae I and X (around 10% each),
A direct spinohypothalamic pathway (Figure 21) was and the intermediate ventral horn (Burstein, R. et al.,
uncovered in the rat (Burstein, R. et al., 1987; Burstein, 1990a).
R. et al., 1990a; Burstein, R. et al., 1990b; Menetrey, D. An additional contribution from the parasympa-
and DePommery, J., 1991) and cat (Katter, J. T. et al., thetic cell column was described in the rat (Burstein,
1991) by the use of both electrophysiologic and tra- R. et al., 1990a; Menetrey, D. and DePommery, J.,
cing methods. It originates bilaterally, although with a 1991). In the cat, the spinohypothalamic tract appears
slight contralateral prevalence, from the entire extent to be much smaller and to contain fewer lamina I

Spinohypothalamic

Figure 21 Diagram representing the spinal laminae of origin, ascending course in the spinal cord, and areas of termination
of the spinohypothalamic pathway. Note that ipsilaterally terminating axons course contralaterally in the spinal cord. Brain
and spinal cord photomicrographs were adapted from Paxinos, G. and Watson, C. 1998. The Rat Brain in Stereotaxic
Coordinates, 4th edn. Academic Press.
 Ascending Pathways: Anatomy and Physiology 511

Nuc. T4
caud. L1 3

C1 2 T6

L4 5

C4 5 T8

C7 8 T12
L6 S2

Figure 22 Distribution in the spinal cord and spinal trigeminal nucleus, pars caudalis, of neurons retrogradely labeled from
injection of fluorogold encompassing the lateral and medial hypothalamus. (Adapted from figure 4 of Burstein, R., Cliffer, K.
D., and Geisler, G. J. 1990b. Cells of origin of the spinohypothalamic tract in the rat. J. Comp. Neurol. 291, 329344).

neurons than in the rat (Katter, J. T. et al., 1991). No dorsomedial, paraventricular and suprachiasmatic
differences in spinal distribution were found between nuclei, and the preoptic area (Cliffer, K. D. et al.,
the medial and lateral hypothalamic pathways, 1991). Anterograde tracer injections restricted to the
except for the sacral parasympathetic nucleus, superficial or deep dorsal horn confirmed that the
which appears to project mainly to medial nuclei latter contributes much more fibers to the spinohy-
(Burstein, R. et al., 1990a). According to a recent pothalamic pathway, the main areas of termination
study by Braz and coworkers (Braz, J. M. et al., 2005) being the posterior hypothalamic area, the posterior
in which transgenic mice expressing a transneuronal part of the lateral hypothalamic area, and the ventral
tracer in a subset of nociceptors were used, nocicep- part of the paraventricular hypothalamic nucleus
tive input conveyed by nonpeptidergic primary (Gauriau, C. and Bernard. J. F., 2004).
afferent neurons is relayed to lamina V neurons pro-
jecting to the ventromedial hypothalamus by spinal 35.6.3.2 Spinal location of ascending
neurons located in the dorsal part of lamina II. fibers
Consonant with retrograde and antidromic stimu- A few studies based on degeneration following spinal
lation studies, terminal arborizations of spinal axons lesions revealed the termination in the lateral
were shown to distribute massively through the lateral (Anderson, F. D. and Berry, C. M., 1959; Ring, G.
hypothalamus, although important labeling was also and Ganchrow, D., 1983) and medial (Kerr, F. W. L.,
observed in the medial hypothalamus (Figure 21) 1975) hypothalamus of spinal fibers coursing in the
(Burstein, R. et al., 1987; Cliffer, K. D. et al., 1991). In ventral funiculus. However, electrophysiological stu-
the lateral hypothalamus, fibers terminate bilaterally dies using antidromic activation from the supraoptic
along its rostrocaudal extent, and throughout the decussation showed that only 5% of the spinohy-
course of the supraoptic decussation. In the medial pothalamic axons course in the ventral funiculus
hypothalamus, fibers terminate mainly contralaterally (Burstein, R. et al., 1991). The remaining travel
in the posterior and dorsal hypothalamic areas, the through the lateral funiculus, mainly in the
512 Ascending Pathways: Anatomy and Physiology

dorsolateral fasciculus (57%), irrespective of their projecting to the NTS are concentrated in the poster-
origin in the superficial or deep dorsal horn olateral hypothalamus and the paraventricular
(Figure 21) (Burstein, R. et al., 1991). Although spinal nucleus. Projections connecting the paraventricular
and trigeminal neurons project to the hypothalamus nucleus with the ventrolateral medulla were also
bilaterally, their axons ascend contralaterally in the reported (Hardy, S. G. P., 2001).
spinal cord (Burstein, R. et al., 1991) and brainstem
(Kostarczyk, E. et al., 1997). Those terminating ipsi-
laterally cross the midline within the supraoptic 35.7 Spinothelencephalic Pathways
decussation (Burstein, R. et al., 1991). Extensive col-
lateralization along the entire brainstem has been During the last decade, evidence accumulated about
reported (Kostarczyk, E. et al., 1997). the existence of ascending nociceptive pathways that
connect the spinal cord directly with various tele-
35.6.3.3 Surface receptors ncephalic regions in the rat. Although data are mainly
A particularly high number of spinal neurons based on anatomic retrograde and anterograde tra-
projecting to the hypothalamus are apposed by pro- cing studies, both the location of the spinal cells of
files immunoreactive to nitric oxide synthase or to origin, the demonstration of cells responding to nox-
interferon- receptor in the lateral spinal nucleus ious stimulation in areas such as the amygdala
(Kayalioglu, G. et al., 1999). (Miyagama, T. et al., 1986), and a recent study using
genetic-controlled transneuronal tracing initiated in
35.6.3.4 Response properties IB4-positive putative nociceptive primary afferent
The majority of the neurons antidromically activated fibers (Braz, J. M. et al., 2005) suggest that some of
from the hypothalamus both in the spinal cord and these areas are spinal targets of nociceptive input.
the spinal trigeminal nucleus belong in the WDR and The data on spinotelencephalic pathways col-
NS classes and also respond to noxious heat or cool- lected till the early years of 2000 are still few and
ing (Burstein, R. et al., 1987, 1991; Malick, A. et al., each study deals with several systems. Therefore,
2000; Zhang, X. J. et al., 2002). Incremental responses notwithstanding their probable functional indivi-
to increasingly intense noxious heat stimulation were duality, they will be described together although
observed (Burstein, R. et al., 1987). Low-threshold tentatively grouped according to their putative
neurons make up 20% of the trigeminohypothalamic functions.
neurons (Malick, A. et al. 2000) and only 4% of the
spinohypothalamic neurons (Burstein, R. et al., 1991).
35.7.1 Thelencephalic Targets of Spinal
The cutaneous receptive fields are particularly small
Ascending Fibers
indicating that this pathway may convey relatively
precise information about the stimulated area Studies using very sensitive anterograde tracers, such
(Burstein, R. et al., 1991; Malick, A. et al., 2000). as phaseolus vulgaris leucoagglutinin or dextran
About half of the spinohypothalamic neurons (Burstein, R. et al., 1987; Cliffer, K. D. et al., 1991;
recorded in the thoracic spinal cord were activated Gauriau, C. and Bernard.J. F., 2004), revealed axonal
by visceral distension with responses that increased terminal arborizations in various regions of the basal
with increasing stimulus intensities (Zhang, X. J. et al., forebrain and cortex (Figure 23), which can be
2002). Eight percent of lumbar spinohypothalamic grouped as areas involved in motor control and
neurons respond to deep low-threshold input areas of the limbic system.
(Burstein, R. et al., 1991). The first group includes the globus pallidus, sub-
stantia nigra, and nucleus accumbens, in particular its
35.6.3.5 Pathways driven at the target medial part. A participation in the striatopallidal
Projections from the hypothalamus, namely the dor- system as well as a role in innate motor patterns
somedial nucleus, descend through a dorsal pathway triggered by noxious stimuli has been proposed for
to the PAG, and through a ventral smaller pathway to the spinalglobus pallidus projection (Braz, J. M. et al.,
the NTS (Thompson, R. H. et al., 1996). According to 2005). Limbic spinal targets include nuclei of the
c-fos studies (Snowball, R. K. et al., 2000), neurons septal complex, thought to be involved in motivation
projecting to the ventrolateral PAG receive visceral and emotion, but also in attention, arousal, learning,
input in the lateral hypothalamus, presumably in and memory (Burstein, R. et al., 1987; Cliffer, K. D.
convergence with somatic input, and neurons et al., 1991). Positive and negative reinforcement
 Ascending Pathways: Anatomy and Physiology 513

innonimata and stria terminalis as well as the posterior

(a) (d) (g)
hypothalamus (Bourgeais, L. et al., 2001a).

35.7.2 Spinal Laminae of Origin

(b) (e) (h) Based on anterograde labeling from restricted injec-
tions in various spinal laminae, Gauriau and Bernard
(Gauriau, C. and Bernard. J. F., 2004) concluded
that cells of origin of spinopallidal fibers are located
in the deep dorsal horn, laminae VII and X and, in
smaller amounts, in the superficial dorsal
(c) (f) (i) horn (Table 1). Marked labeling was obtained in the
lateral aspect of the globus pallidus from neurons
located in lamina V, as revealed by transneuronal
tracing with wheat germ agglutinin synthesized by
IB4-positive primary afferents in mice (Braz, J. M.
et al., 2005).
Figure 23 Distribution of spinal fibers labeled
anterogradely with phaseolus vulgaris leucoagglutinin in the
Spinal fibers projecting to the nucleus accumbens
diencephalon and telencephalon, in horizontal view. and the septal nuclei have similar bilateral origins at
Adapted from figure 3 of Cliffer, K. D., Burstein, R., and the reticulated portion of the deep dorsal horn, the
Giesler, G. J. 1991. Distributions of spinothalamic, lateral spinal nucleus, and lamina X throughout the
spinohypothalamic, and spinotelencephalic fibers revealed entire length of the spinal cord (Burstein, R. and
by anterograde transport of PHA-L in rats. J. Neurosci. 11,
Giesler, G. J., 1989). Deep dorsal horn neurons
852868.
account to half of the entire projection population,
followed by the lateral spinal nucleus, and lamina X
associated with learning trials during nociceptive
(about 15% each). Neurons in lamina I and the
processing have been claimed to be accomplished
intermediate/ventral horn are very few, but the first
by this pathway (Cliffer, K. D. et al., 1991).
are slightly more numerous in the spinoseptal path-
The central nucleus of the amygdala together with
way (Table 1).
what has been called the extended amygdala,
Spinal neurons projecting to the amygdala present
namely the substantia innonimata and the Bed similar laminar distribution (Menetrey, D. and
Nucleus of the stria terminalis, also receive direct DePommery, J., 1991; Burstein, R. and Potrebic, S.,
projections from the spinal cord (Cliffer, K. D. et al., 1993). Neurons in the reticulated region of the deep
1991; Gauriau, C. and Bernard.J.F., 2004), as do the dorsal horn also make up about half of the entire
medial orbital and infralimbic cortices (Cliffer, K. D. spinoamygdala population, but they are located in
et al., 1991). Projections to the orbital cortex may its dorsal portion in cervical segments and ventral
affect autonomic, endocrine, and behavioral func- portion in thoracolumbar segments. The lateral
tions in relation to pain (Burstein, R. and Potrebic, spinal nucleus contributes to 25% of the projection,
S., 1993). The horizontal and vertical limbs of the lamina X to 13% (mainly at upper lumbar segments),
diagonal band of Broca are also targeted by spinal and the intermediate and ventral horn to 10%, at
axons (Burstein, R. et al., 1987; Cliffer, K. D. et al., upper cervical and lumbar segments (Table 1).
1991). Neurons in lamina V receiving primary afferent
These direct spinotelencephalic projections appear input through lamina II neurons activated by
to be paralleled by ascending polysynaptic pathways IB4-positive primary afferents also send projections
relaying not only in the brainstem, as is the case of the to the amygdala, as well as to the bed nucleus of
spinoparabrachialamygdaloid pathway (Ma, W. and stria terminalis (Braz, J. M. et al., 2005). A projection
Peschanski, M., 1988; Bernard, J. F. and Besson, J. M., to the dorsal part of substantia innonimata from
1990; Jasmin, L. et al., 1997), but also within the tele- the superficial and deep dorsal horn, contralaterally,
ncephalon. Anterograde tracer injections in the central and from the lateral spinal nucleus, ipsilaterally,
and basolateral anterior nuclei of the amygdala has been described (Gauriau, C. and Bernard. J.F.,
revealed projections from there to the substantia 2004).
514 Ascending Pathways: Anatomy and Physiology

Table 1 Relative participation of spinal cord laminae in the various nociceptive ascending pathways

LSN I II III IV V VI VII VIII X

LCN & & & & & &

(10%) (60%)
VLMlat & & & & &
LRt & & & & & &
DRt dorsal & & & &
(Medial) (Medial) (Medial)
DRt ventral & & & & &
(Medial) (Medial) (Medial)
NTS & & & & & & &
RVM & & & &
PBN & & & &
(Lateral) (Lateral)
PAG & & & & & & & &
(Lateral) (Lateral)
Lateral thalamus & & & & & & & &
Medial thalamus & & & &
Hypothalamus & & & & & &
(30%) (10%) (- - - - - - - - - - 50%- - - - - - - - - - ) (10%)
N. Pallidus & & & & & &
N. Accumbens & & & & &
(25%) (- - - - - - - - - - 50%- - - - - - - - - - ) (10%)
Septal nucleus & & & & & &
(15%) (- - - - - - - - - - 50%- - - - - - - - - - ) (15%)
Amygdala & & & & & &
(25%) (- - - - - 50%- - - - - ) (10%) (10%)
Subst. inonimata & & &
Orbital cortex & & & & & &
(15%) (- - - - - 62%- - - - - ) (13%) (10%)

Relative amounts refer to each pathway and do not allow comparisons between pathways. Whenever quantified, the relative contribution to
each pathway is referred between brackets. LSN, lateral spinal nucleus; LCN, lateral cervical nucleus; VLMlat, caudal ventrolateral reticular
formation, lateral portion; LRt, lateral reticular nucleus; DRt dorsal, dorsal reticular nucleus, dorsal portion; DRt ventral, dorsal reticular
nucleus ventral portion; NTS, nucleus tractus solitarii; RVM, rostral ventromedial medulla; PBN, parabrachial nuclei; PAG, periaqueductal
gray; IX, spinal laminae.

The orbital cortex receives its major spinal pro- variety of supraspinal regions that are targeted
jections from the contralateral reticulated area of the (Figure 24). This has been interpreted as the anato-
deep dorsal horn (62%) at the cervical level or its mical substrate for the triggering of a multitude of
ventromedial aspect at the thoracic and lumbar responses to the noxious event, from autonomic and
levels. The lateral spinal nucleus contributes to motor reactions to affective and cognitive behaviors.
15% of the projection, but mainly at lumbar seg- Nevertheless, it is curious to note that, contrary to
ments, while the intermediate/ventral horn and what was thought in the middle of the twentieth
lamina X contribute to 13% and 10%, respectively, century, nociceptive input does not necessarily arrive
both at the upper cervical and lower thoracic/upper to high processing motor, affective, and
lumbar segments (Table 1) (Burstein, R. and cognitive centers in the thelencephalon through mul-
Potrebic, S., 1993). tisynaptic chains capable of filtering information at
various successive levels, but can reach those areas
through direct spinofugal pathways.
35.8 Discussion Another aspect that emerges is that multiple poly-
synaptic pathways seem to parallel monosynaptic
35.8.1 Multiple Parallel Ascending connections between the spinal cord and each suprasp-
Pathways inal target. So far, this organization pattern was
The main feature that stands out is the multiplicity of demonstrated only for a few systems, such as the med-
the ascending nociceptive system in terms of the ial paracentral spinothalamic and the spinoamygdaloid
 Ascending Pathways: Anatomy and Physiology 515

Spinopallidus/accumbens

Spinolimbic

Spinohypothalamic
Lateral spinothalamic
Medial spinothalamic

Spinomesencephalic

Spinopontine

Spino-RVM

Spino-NTS
Spino-DRt
Spino-VLM

Spino-LCN

Figure 24 Diagram illustrating the termination areas of the various ascending nociceptive pathways. The laterality of the
ascending tracts and termination fields with respect to the side of arrival of primary afferent input is represented, the left side
being ipsilateral and the right side contralateral. Brain and spinal cord photomicrographs were adapted from Paxinos, G. and
Watson, C. 1998. The Rat Brain in Stereotaxic Coordinates, 4th edn. Academic Press.
516 Ascending Pathways: Anatomy and Physiology

systems, served by a direct pathway and a disynaptic Table 2 Relative participation of low threshold (LT),
pathway with a relay in the PBN (Ma, W. and wide-dynamic-range (WDR) and nociceptive specific (NS)
spinal cord neurons in the various nociceptive ascending
Peschanski, M., 1988; Bernard, J. F. and Besson, J. M.,
pathways
1990; Jasmin, L. et al., 1997). However, the fact that
most spinal targets send projections to other brain areas LT WDR NS
that are also targeted by spinal fibers strongly suggests
LCN Mainly hair 6086%
a similar architecture for most systems, with the parti- movement
cipation of parallel monosynaptic and multisynaptic VLM 25% 25% 50%
chains of different lengths. Such an arrangement may NTS The majority
imply that the responses to the noxious event gener- RVM The majority
PBN  (75
ated at each site evolve along time according to
90% in
postarriving of noxious-evoked input generated in lamina I)
other pain-processing centers. Notwithstanding the PAG (DDH) 50% (lamina I)
extensive anatomical and electrophysiological data (Lamina
that are still needed to corroborate this hypothesis, it I DDH
Lateral 35%
is worth to take it into consideration in future
thalamus
investigation. Medial 70%
thalamus
Hypothalamus 4% The majority
35.8.2 Spinal Neuronal Populations at
the Origin of Nociceptive Ascending DDH, deep dorsal horn. Other abbreviations as in Table 1.
Pathways
When facing such a variety of nociceptive ascending amount of projecting neurons they involve have
pathways, it is tempting to assume that they differ by
also been pointed out (Apkarian, A. V. and Hodge,
either channeling different sensory modalities to
C. J., 1989d; Burstein, R. et al., 1990b; Mouton, L. J.
brain regions specifically dedicated to their proces-
and Holstege, G., 1998; Willis, W. D. et al., 2001) but a
sing, or by the responses they induce to whatever
complete picture of the quantitative variations is still
stimulus through activation of a particular suprasp-
hard to attain due to the use of tracers of variable
inal region. For the first assumption to be correct, the
sensitivity. As to the possibility that each pathway
spinal source of input should differ between different
identity relies on a particular neurochemical archi-
pathways. The overview of the ascending nociceptive
tecture at the spinal relay, data are too scarce to allow
system here presented clearly shows that this is not
the case. On the contrary, if one compares the con- any sort of considerations.
tribution of the various spinal laminae to each The difficulty in concealing this anatomofunc-
pathway (Table 1), the first emerging picture is that tional organization with the well-known capacity of
of a strong similarity, which favors the second modality discrimination during acute physiological
assumption that the functional properties of a path- pain led some authors to ascribe discrimination capa-
way depend on the functional engagement of its city to a particular spinal region, leaving the
target. However, in spite of a large overlap, there remaining spinal cord with a secondary, largely
are subtle dissimilarities between pathways consist- unknown but eventually not important role in pain
ing of a preponderance of some laminae over others processing. The high concentration of nociceptive
or differences in the location of the projecting specific neurons in lamina I, together with the con-
neurons in each lamina (Table 1). Also the electro- vergence of input of various nature and peripheral
physiological response properties of spinal neurons origin to this lamina and the easy separation of its
participating in the various pathways overlap consid- structural neuronal groups (see Chapter Spinal Cord
erably (Table 2): all pathways including nociceptive Physiology of Nociception) brought it into focus
specific, wide-dynamic-range and low-threshold during the last decade. However, lamina I does not
neurons, as well as neurons responding to cutaneous, contribute to all spinofugal nociceptive pathways
visceral, and deep noxious stimuli. Again, subtle dif- while participating similarly in many others
ferences are likely to occur about the proportion of (Table 1). Moreover, an appraisal of the participation
neurons of each kind that take part in each pathway of the various lamina I structural cell groups in a
(Table 2). Differences between pathways over the sample of nociceptive ascending pathways revealed
 Ascending Pathways: Anatomy and Physiology 517

Table 3 Relative amounts of supraspinally projecting lamina I neurons

Fusiform Flattened Pyramidal Multipolar

VLM 80% 10% 10%

DRt 1030% 520% 6085%
NTS 25% 40% 35%
PBN 70% 30%
PAG 30% 70%
Thalamus VBC 2530% (enlargements) 7075% (enlargements)

VBC, ventrobasal complex of the thalamus.

large superposition, although again slight differences neurons projecting to brainstem regions connected
as to their relative amount in each pathway and the to the medial thalamus do not necessarily share the
specific involvement or noninvolvement of certain same properties of the medial spinothalamic neurons.
groups were detected (Table 3). What is noteworthy Receptive fields of spino-RVM and spino-PAG neu-
is that when taken together, the various electrophy- rons vary from small, confined to a sole limb, to very
siological studies on the response properties of large and complex. PBN-projecting spinal neurons
lamina I neurons do not support a clear-cut struc- were shown to present small receptive fields (Bester,
turalfunctional correlation based on stimulus- H. et al., 2000), contrary to PBN neurons projecting to
modality processing (see discussion in Galhardo, V. the paracentral nucleus of the thalamus, which have
et al., 2000). The possibility that stimulus character- large receptive fields (Bourgeais, L. et al., 2001b). Also
ization at the central nervous system depends on a noteworthy in this respect is the fact that neurons in
combinatory activation process rather than on the the RVM and PAG send axonal collaterals to the
activation of specific channels should be addressed lateral thalamus, while neurons projecting through
in the future. the lateral spinothalamic or the spinocervical path-
As a working hypothesis, it could be postulated at ways send collaterals to the medial thalamus. As a
this point that each pathway has particular character- whole, the data suggest that, although the lateral
istics that depend on both, the kind of input it spinothalamic pathway as well as the spinocervical
transmits (defined by the relative contribution of and the spinohypothalamic pathways appear to
the various spinal neurons) and the functional prop- be morphofuntionally organized to allow stimulus
erties of the target. Ultimately, for each noxious location and intensity discrimination, an extensive
event, ascending transmission of nociceptive input cross-talk between the various nociceptive ascending
would be the result of the relative activation of the pathways is likely to take place.
various pathways.

35.8.4 Nociceptive Ascending Pathways as

35.8.3 Stimulus Discrimination Part of a Complex Nociceptive Integration
System
A clear separation between the lateral and the medial
spinothalamic nociceptive pathways as to the ability Although the ascending transmission system and the
of the former to discriminate between stimulus loca- descending endogenous pain control system (see
tion and intensity has been established, based mainly Chapter Descending Control Mechanisms) are nor-
on the electrophysiological properties of neurons mally dealt with separately, evidence has been
projecting in each pathway. Amongst the supporting accumulated to prove that they are both part of a
data stand out the relatively small size of the recep- sole nociceptive system buildup in such a way that
tive fields, the stimulus intensity-encoding capacity, information is treated at different brain levels in
and the somatotopic organization of the spinal, tha- order to integrate nociception and various brain
lamic, and cortical neurons in the lateral pathway, as functions. The brain areas of termination of the
opposed to the medial pathway. It should be noted, ascending nociceptive pathways are those and the
however, that early studies pointed out that the med- same from where pain-control actions are elicited
ial spinothalamic system conveys spinal input either upon local stimulation (Jones, S. L., 1992). These
directly or through a brainstem relay, but spinal areas are intimately connected with each other, and
518 Ascending Pathways: Anatomy and Physiology

each of them with the spinal cord dorsal horn, in most Almeida, A., Tavares, I., Lima, D., and Coimbra, A. 1993.
Descending projections from the medullary dorsal reticular
cases through direct descending projections that very nucleus make synaptic contacts with spinal cord lamina I
often participate in spino-brain-spinal reciprocal cells projecting to that nucleus: an electron microscopic
loops (Lima, D. et al., 1998). Descending pain-control tracer study in the rat. Neuroscience 55, 10931106.
Ammons, W. S. 1987. Characteristics of spinoreticular and
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Lima, D., 2002). T5 spinothalamic neurons projecting to medial thalamus with
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Res. 291, 118. phosphorylation of NMDA receptor 1 subunits in spinal cord
Wiberg, M., Westman, J., and Blomqvist, A. 1987. Somatosensory dorsal horn and spinothalamic tract neurons after
projection to the mesencephalon: an anatomical study in the intradermal injection of capsaicin in rats. J. Neurosci.
monkey. J. Comp. Neurol. 264, 92117. 20, 69896997.
 36 Dorsal Columns and Visceral Pain
W D Willis Jr. and K N Westlund, University of Texas Medical Branch, Galveston, TX, USA
2009 Elsevier Inc. All rights reserved.

36.1 Clinical Evidence Concerning Spinal Cord Pathways That Signal Visceral Pain 527
36.1.1 Spinothalamic Tract 527
36.1.1.1 Anterolateral cordotomy 527
36.1.1.2 Commissural myelotomy 529
36.1.2 Posterior Column 529
36.1.2.1 Hitchcock procedure (stereotactic C1 central myelotomy) 529
36.1.2.2 Limited midline myelotomy 529
36.2 Basic Science Evidence Concerning Spinal Cord Pathways That Signal Visceral Pain 530
36.2.1 Spinothalamic Tract 530
36.2.2 Spinoreticular, Spinoparabrachial, Spinoamygdalar, and Spinohypothalamic Tracts 531
36.2.3 Postsynaptic Dorsal Column Path 531
36.2.3.1 Historical evidence for a visceral projection in the dorsal column 531
36.2.3.2 Effects of interruption of the dorsal column or a lesion of dorsal column nuclei on
responses of brainstem and thalamic neurons to noxious visceral stimuli 531
36.2.3.3 Effects of a dorsal column lesion on behavioral responses 533
36.2.3.4 Effects of a dorsal column lesion on the regional cerebral blood flow changes that result
from colorectal distention in monkeys 534
36.2.3.5 Blockade of synaptic relay in sacral cord by morphine or 6-cyano-7-nitroquinoxaline-
2,3-dione 534
36.2.3.6 Projections of the postsynaptic dorsal column pathway 534
36.2.3.7 Upregulation of NK1 receptors in PSDC neurons after colon inflammation 536
36.2.3.8 Fos expression in PSDC neurons after noxious visceral stimulation 537
36.3 Descending Facilitation 537
References 539

Glossary
activity box Apparatus used to determine the NMDA N-methyl-D-aspartic acid.
amount and time course of exploratory activity of pain referral Projection of the source of pain to an
an animal, such as a rat. area of the body distant to the actual area of injury.
central neuropathic pain pain that develops fol- paresthesias Unusual sensations, such as tingling
lowing injury to the central nervous system. or burning.
CNQX 6-cyano-7-nitroquinoxaline-2,3-dione, a rhizotomy Transaction of one or more spinal
non-N-methyl-D-aspartic acid receptor antagonist. roots.
kainic acid lesion damage produced by injection viscerospecific responses Neuronal activity
of kainic acid, a substance that produces evoked by sensory input from a particular visceral
excitotoxicity. organ.

527
528 Dorsal Columns and Visceral Pain

36.1 Clinical Evidence Concerning (a) (b)

Spinal Cord Pathways That Signal
Visceral Pain
36.1.1 Spinothalamic Tract
36.1.1.1 Anterolateral cordotomy
It is well known that n is relieved (at least tempora-
rily) and thermal sense is lost over the appropriate
part of the contralateral body following an anterolat- (c)
eral cordotomy (Spiller, W. G., 1905; Spiller, W. G.
and Martin, E., 1912; Foerster, O. and Gagel, G.,
1932; reviewed in Gybels, J. M. and Sweet, W. H.,
1989). The signals are transmitted from one side of
the body to the contralateral thalamus (Kenshalo, D.
R. et al., 1980; Chung, J. M. et al., 1986; Bushnell, M. C.
et al., 1993; Lee, J. I. et al., 1999; 2005) and from there
to the appropriate regions of the cerebral cortex Figure 1 (a) Drawing of a cross section of the human
spinal cord at the third cervical level. The dotted area shows
(see Casey, K. L. and Bushnell, M. C., 2000). It has the region that needs to be sectioned by a cordotomy in
been presumed that the pain signals are conveyed order to relieve contralateral pain completely. Note that the
chiefly by the spinothalamic tract, which decussates lesion must extend to a level that is posterior to the
at the spinal cord level (Mehler, W. R., 1962), expected location of the denticulate ligament and
anteromedially across the lateral funicuclus and into the
although other pathways that accompany the
anterior funiculus. (b) Drawing of a cross section of the
spinothalamic tract are also likely to contribute. human spinal cord at a mid-thoracic level showing the
These include the spinoreticular, spinoparabrachial, location of a limited midline myelotomy that was used to
spinohypothalamic, and other tracts (Willis, W. D. relieve completely the pain of colon cancer for the duration
and Westlund, K. N.,1997; Willis, W. D. and of the 3-month survival period. No opioid analgesics were
required after a tapering-off period. (c) Photomicrograph of
Coggeshall, R. E., 2004). Innocuous thermal signals a cross section of the human spinal cord at the level of a
are attributed just to the spinothalamic tract. limited midline myelotomy that was placed at a mid-thoracic
Anterolateral cordotomy can relieve superficial level in a patient with a painful presacral sarcoma. The
and deep somatic pain, as well as visceral pain (and lesion (shown by the area of demyelination near the midline
of the dorsal columns) eliminated the pain and the need for
can block thermal sense), provided that the lesion is
narcotics for the duration of the patients survival time,
extensive enough (Gybels, J. M. and Sweet, W. H., which was 3 years postsurgery. (a) From Nathan, P. W.,
1989; Nathan, P. W. et al., 2001). For complete pain Smith, M., and Deacon, P. 2001. The crossing of the
relief on the contralateral side of the body, a cordot- spinothalamic tract. Brain 124, 793803. (b) From
omy at an upper cervical level needs to extend from Hirshberg, R. M., Al-Chaer, E. D., Lawand, N. B., Westlund,
K. N., and Willis, W. D. 1996. Is there a pathway in the
just posterior to the denticulate ligament, across posterior funiculus that signals visceral pain? Pain 47,
the remainder of the lateral funiculus and well into 291305.
the anterior funiculus (Figure 1(a); Nathan, P. W.
et al., 2001).
However, even after an initially successful cordot- tended to limit the use of this procedure to patients
omy, pain relief may not persist more than a few with pain from a terminal disease, usually cancer, or
months to a year or so, although in some cases the to abandon the procedure altogether in favor of
pain relief is of very long duration (Gybels, J. M. and pharmacotherapy.
Sweet, W. H., 1989). It is unclear why the pain can Visceral pain can be relieved by cordotomy, pro-
recur. Some suggestions are that other pathways now vided that the lesion extends into the anterior
convey the pain signals, the disease advances (e.g., funciculus medially to the spinal cord gray matter
metastatic cancer may activate spinothalamic (Gybels, J. M. and Sweet, W. H., 1989). A unilateral
neurons whose axons were not interrupted by the cordotomy can be effective if the visceral pain is
cordotomy), or central neuropathic pain develops. restricted to one side. However, if the pain is
Because of the recurrence of pain after a large bilateral, a bilateral cordotomy may be needed.
proportion of cordotomies, neurosurgeons have Unfortunately, this can lead to undesired side effects,
 Dorsal Columns and Visceral Pain 529

such as incontinence because of interruption of des- and by the changes in the sensory examination pro-
cending pathways that control bowel and bladder duced by a lesion. Stimulation in the central cord at
function. If the cordotomy is done at a high cervical C1 at 50 Hz typically resulted in a burning sensation
level for pain originating from the upper part of the in the chest or abdomen (Hitchcock, E., 1970; see
body, there is danger that the respiratory control Gybels, J. M. and Sweet, W. H., 1989). A lesion in
pathways may be interrupted, leading to severe the same location produced a change in pinprick
(potentially lethal) difficulty with breathing. sensation, either a loss of the ability to distinguish
sharp from blunt or loss of a sensation of pain in
36.1.1.2 Commissural myelotomy response to pinprick. The analgesia was bilateral.
An alternative procedure called commissural myelot- Cancer pain was relieved in most cases, and the relief
omy is directed at interrupting the axons of persisted until death or for over 5 years. Some
spinothalamic tract neurons of both sides as they patients (20%) failed to have a sensory loss and also
cross the midline (Armour, D., 1927; Mansuy, L. did not experience pain relief.
et al., 1944; 1976; see Gybels, J. M. and Sweet, W. H., In the surgical series done by Schvarcz J. R. (1977;
1989). A lesion is made near the midline of the spinal 1978), lesions were made at a depth of 5 mm from the
cord and extending rostrocaudally for as many posterior surface of the spinal cord, reaching the base
segments as needed to interrupt the appropriate of the posterior column. Electrical stimulation pro-
projections. The lesion should be sufficiently deep to duced paresthesias in the feet, legs, or trunk (and also
ensure that the crossing spinothalamic axons in the in the trigeminal distribution). Relief of cancer pain
anterior white commissure are cut. Of course, the persisted in 78% of cases for at least 0.52 years (most
crossing axons of a number of tracts that accompany of the patients died by this time). Other types of pain,
the spinothalamic tract, such as spinoreticular axons, including peripheral and central neuropathic pain,
are also interrupted, as are many axons in the posterior were relieved for 0.54 years. No neurological defi-
funciculi. According to the analysis of Dargent, M. cits were seen. Several other clinical studies reported
et al. (1963), this procedure is apparently particularly similar results (Papo, L. and Luongo, A., 1976; Eiras,
effective for vaginal and visceral pain. Interestingly, J. et al., 1980; Sourek, K., 1985).
commissural myelotomy often relieves clinical pain
even if on testing there is no hypalgesia. 36.1.2.2 Limited midline myelotomy
Furthermore, pain and temperature sensation may be Instead of a lesion at an upper cervical level,
lost over a much greater extent of the body than could Gildenberg P. L. and Hirshberg R. M. (1984) made
be predicted from the location and dimensions of the a midline lesion at T10 in patients with pelvic cancer
commissural myelotomy (Gybels, J. M. and Sweet, W. pain. The location was chosen to be just above the
H., 1989). Based on evidence reviewed in the follow- level of entrance of primary afferent fibers from the
ing, it now seems likely that the effect of a pelvic viscera into the spinal cord. The procedure
commissural myelotomy depends at least in part on was termed a limited midline myelotomy. Eight cases
the interruption of a visceral pain pathway that done by Hirshberg, using a mechanical probe, were
ascends in the posterior funiculi. described further and a postmortem specimen show-
ing the spinal cord lesion in one patient was
illustrated (Figure 1(b)) in Hirshberg R. M. et al.
36.1.2 Posterior Column
(1996). The lesion interrupted the medial parts of
36.1.2.1 Hitchcock procedure the posterior funiculi but did not appear to intrude
(stereotactic C1 central myelotomy) into the central gray matter or anterior white com-
Hitchcock developed a completely different missure. The results of the lesions in the eight cases
approach to lesioning the spinal cord to relieve pain were generally quite favorable, with pain relief that
(Hitchcock, E., 1970; 1974; 1977; see also Papo, L. and lasted as long as the patients survived and with little
Luongo, A., 1976; Eiras, J. et al., 1980; Schvarcz, J. R., or no need for strong analgesic drugs.
1977; 1978; Sourek, K., 1985). He placed a lesion in The chief of neurosurgery at our institution, Dr.
the central part of the upper cervical spinal cord H. J. W. Nauta, made lesions using a technique simi-
using a stereotactic approach. The location of the lar to that described by Hirshberg in a series of
lesion was sometimes determined by recordings of patients who had pain from pelvic or other cancers.
evoked potentials but more often by the sensory The results in each case were again generally favor-
effects of electrical stimulation through the electrode able and the need for strong analgesics greatly
530 Dorsal Columns and Visceral Pain

reduced. In one case, a midline myelotomy was done (a) Spinothalamic tract
in a patient with inflammatory bowel disease second-
ary to irradiation that had successfully eradicated a
cervical cancer (Nauta, H. J. W. et al., 1997; 2000).
The patient had severe ongoing pain, and the pain
Skin
intensified during bowel movements. As a conse-
quence of the pain, the patient had lost a substantial
fraction of her body weight, and it was judged that
she would not survive much more weight loss. After
midline myelotomy, this patient lived for almost 5 Viscus Anterolateral
years with no further pelvic pain (although she did tract axon
experience subdiaphragmatic pain because of perito-
nitis). She died from complications of diabetes. (b) Visceral PSDC pathway
In another of the cases reported by Nauta H. J. W.
Gr
et al. (1997; 2000), pain was reduced from 10 to 23 on
the visual analog scale, and narcotic medication was Cu
tapered from 30 mg of intravenously administered
morphine per hour preoperatively to 5 mg per hour
within 5 days postoperatively. Another patient had
unbearable pain from a presacral sarcoma, despite
several efforts to resect the tumor. A midline mye-
lotomy was done in the area shown in Figure 1(c).
The lesion resulted in dramatic pain relief for the
C1
remainder of the patients life. He survived the sur-
gery by 36 months, and for most of this time he did
not require narcotic medication. In general, only in
some cases were even minor and transient sensory
side effects observed. Pelvic cancer pain was consis- Midthoracic
tently reduced following midline myelotomy, and
narcotic usage could be decreased on average by
83%.
Several other groups around the world have had
L6-S1
similar experiences with midline myelotomies
(Becker, R. et al., 1999; Kim, Y. S. and Kwon, S. J.,
2000; Filho, O. V. et al., 2001; Hu, J. S. and Li, Y. J., Figure 2 (a) Convergence of visceral and somatic afferent
inputs on an anterolateral tract neuron. Such
2002; Hwang, S. L. et al., 2004), and the results have viscerosomatic convergence is thought to be responsible
been reviewed by Becker, R. et al., (2002). for referral of visceral pain to somatic structures. (b) Course
followed by the axonal projections of postsynaptic dorsal
column neurons found in the central region of the spinal
36.2 Basic Science Evidence cord gray matter at sacral and mid-thoracic levels. The
anterograde tracer, phaseolus vulgaris leucoagglutinin
Concerning Spinal Cord Pathways (PHA-L), was injected and then several days later traced in
That Signal Visceral Pain axons of the dorsal column to the brainstem. The projection
from the sacral cord ascended at the midline, whereas that
36.2.1 Spinothalamic Tract from the mid-thoracic cord ascended more laterally, just
The spinothalamic tract conveys somatic but also ventral to the dorsal intermediate sulcus. Information about
visceral nociception most likely travels with other sensory
some visceral nociceptive information to the thala- input to the thalamus in the medial lemniscus. Data from
mus (Figure 2(a)). Studies in which recordings were Wang, C. C., Willis, W. D., and Westlund, K. N. 1999.
made from antidromically activated spinothalamic Ascending projections from the central, visceral processing
tract neurons have demonstrated that many of these region of the spinal cord: a PHA-L study in rats. J. Comp.
cells can be activated (or in some instances inhibited) Neurol. 415, 341367; illustration from Willis, W. D., Al-
Chaer, E. D., Quast M. J., and Westlund, K. N. 1999. A
following stimulation of viscera. For example, visceral pain pathway in the dorsal column of the spinal
Foreman and his group have shown that feline and cord. Proc. Natl. Acad. Sci. U. S. A 96, 76757679.
 Dorsal Columns and Visceral Pain 531

primate spinothalamic tract cells can be excited by 1996; Zhang, X. et al., 2002). A cordotomy or a com-
electrical stimulation of cardiopulmonary visceral missural myelotomy is likely to interrupt these
afferent fibers, occlusion of a coronary artery, or pathways, and thus the relief of visceral pain by
injection of bradykinin into the coronary circulation such lesions is likely to be at least partially explained
(see review by Foreman, R. D., 1989). Similarly, by this, as well as by interruption of the spinothala-
spinothalamic tract cells have been shown to respond mic tract.
to electrical stimulation of the greater splanchnic
nerve (Hancock, M. B. et al., 1975; Foreman, R. D.
et al., 1981), distension of any of several hollow vis-
36.2.3 Postsynaptic Dorsal Column Path
cera, including the gall bladder (Ammons, W. S. et al.,
1984), kidney (Ammons, W. S., 1987), ureter 36.2.3.1 Historical evidence for a visceral
(Ammons, W. S., 1989), urinary bladder (Milne, R. J. projection in the dorsal column
et al., 1981), and colon (Al-Chaer, E. D. et al., 1999) or Recordings from the human posterior funiculus have
noxious stimulation of the testicle (Milne, R. J. et al., revealed the presence of visceral afferents that
1981). In general, spinothalamic tract and other neu- respond to distention of the urinary bladder
rons that are activated by visceral stimulation also (Puletti, F. and Blomqvist, A. J., 1967). Responses at
respond to stimulation of somatic tissue. This con- various levels of the dorsal column-medial lemniscus
vergence of visceral and somatic input onto spinal pathway have also been reported in animal experi-
cord neurons, including nociceptive neurons that ments following electrical stimulation of visceral
project to the brain, is thought to help account for nerves (Amassian, V. E., 1951; Aidar, O. et al., 1952;
the referral of visceral pain to the body wall in many Rigamonti, D. D. and Hancock, M. B., 1974; 1978).
clinical conditions (Figure 2(a); Head, H., 1893).
Examples of the somatic receptive fields of viscero- 36.2.3.2 Effects of interruption of the
sensitive spinothalamic cells that correspond to the dorsal column or a lesion of dorsal column
distribution of pain referral are noted in Foreman nuclei on responses of brainstem and
(Foreman, R. D., 1989) and Milne (Milne, R. J. et al., thalamic neurons to noxious visceral
1981). Only rarely can a spinal neuron be found that stimuli
seems to respond to just visceral stimuli and not to To determine if there is a visceral nociceptive path-
somatic ones. However, when the search stimulus is way in the dorsal columns of experimental animals,
colon distension, rather than antidromic activation recordings were made from viscero-responsive neu-
from the lateral thalamus, viscerospecific responses rons in the ventral posterior lateral nucleus of the
can be recorded from cells located rostrally in the thalamus in rats before and after surgical interruption
central lateral nucleus of the intralaminar complex of of either the dorsal columns bilaterally or the ipsilat-
the thalamus (Ren, Y. et al., 2006). This region has eral ventrolateral column, which contains the
been shown to receive direct spinal innervation from spinothalamic and associated nociceptive pathways
lamina X neurons (Wang, C. C. et al., 1999) and is rich (Al-Chaer, E. D. et al., 1996a). The dorsal column
in opiates (Sar, M. et al., 1978). The spinothalamic lesion reduced the responses of the thalamic neurons
projection from lamina X is situated at the medial- to colorectal distension on average by about 80%
most edge of the spinothalamic tract and shifts later- (Figure 3(Ae), cf. upper and middle rows of records).
ally as it ascends with the spinothalamic tract. The remaining response was eliminated by the lesion
of the ventrolateral quadrant of the spinal cord
(Figure 3(Ae), lower row of records). Responses to
36.2.2 Spinoreticular, Spinoparabrachial,
weak mechanical stimuli applied to the skin were
Spinoamygdalar, and Spinohypothalamic
abolished by the dorsal column lesion, whereas the
Tracts
responses to the strongest mechanical stimuli were
Several studies have demonstrated the effectiveness affected only by the ventrolateral quadrant lesion
of visceral stimulation in exciting (or inhibiting) spi- (Figure 3(Ad)). Similar effects were seen when the
noreticular tract neurons (Blair, R. W. et al. 1984; noxious visceral stimulus was inflammation of the
Hobbs, S. F. et al., 1990), spinoparabrachial and colon by intraluminal injection of mustard oil (not
spinoamygdalar neurons (Menetrey, D. and De illustrated). The mustard oil produced a progressive
Pommery, J., 1991; Bernard, J. F. et al., 1994), and increase in the background activity of the viscero-
spinohypothalamic tract neurons (Katter, J. T. et al., sensitive thalamic neurons. A dorsal column lesion
532 Dorsal Columns and Visceral Pain

was more effective in reducing the background activ- column placed at the midline failed to reduce the
ity than was a ventrolateral quadrant lesion. responses (Figure 3(Bb)). However, bilateral lesions
Neurons in the rat ventral posterior lateral thala- just ventral to the dorsal intermediate sulcus at an
mic nucleus could also be found that responded to upper cervical level were effective (Figure 3(Bc)).
distention of the duodenum (Figure 3(Ba); Feng, Y. This shift in the location of the dorsal column lesion
et al., 1998). In these experiments, a lesion of the dorsal needed to interrupt nociceptive responses in the

(A) (a) (b) (c)

(d) (e) Control (f)

500 BR PR PI
200

0 0

DC lesion
Rate (spikes s1)

500 200

0 0

DC + VLC lesions
500 200

0 0

10 s
20 40 60 80 mm Hg

(B) (a)
0.2 ml 0.3 ml 0.4 ml 0.5 ml
30

0
(b)
Rate (spikes s1)

30

0
(c)
30

0
0 100 0 100 0 100 0 100
Time (s)
(d) (e) (f)

VPM
VPL
 Dorsal Columns and Visceral Pain 533

thalamus relates to the fact that the visceral input (cf., Chandler, M. J. et al., 1992; Bruggemann, J. et al.,
originated from an abdominal, rather than a pelvic 1994), as in rats. The effects of a dorsal column lesion
organ (see Figure 2(b), discussion below, and Wang, on such responses in monkeys are consistent with those
C. C. et al., 1999). obtained in rats (Al-Chaer, E. D. et al., 1998).
The off-midline part of the dorsal column path- The thalamic responses to visceral input are
way that carries visceral input from the thoracic level relayed through the dorsal column nuclei. Visceral
of the spinal cord was investigated in behavioral (see responses of neurons in the rat ventral posterior
Figure 2(b) and discussion below) and electrophysio- lateral thalamic nucleus could be reduced by small
logical studies after noxious chemical irritation of the electrolytic or kainic acid lesions of the nucleus gra-
pancreas (Houghton, A. K. et al., 1997; 2001). Noxious cilis (Al-Chaer, E. D. et al., 1997). Responses of cells in
stimulation of the pancreas by applications of brady- the nucleus gracilis have been recorded after noxious
kinin to its exposed surface in anesthetized rats stimulation of either the colon or the pancreas
resulted in excitation of neurons in the ventral pos- (Al-Chaer, E. D. et al., 1996b; 1997; Houghton, A. K.
terior nucleus of the thalamus (Houghton, A. K. et al., et al., 2001).
2001). The mean firing rate of thalamic neurons was
increased 412 120% above baseline during the first
40 s after bradykinin application to the pancreas and 36.2.3.3 Effects of a dorsal column lesion
was sustained for 36 min. Large dorsal column on behavioral responses
lesions effectively eliminated the increased responses Changes observed in behavioral experiments were
of thalamic neurons to applications of bradykinin on consistent with the electrophysiological findings.
the pancreas and baseline activity remained constant. For example, when a rat is put into an unfamiliar
Enhanced responses to skin pinch after pancreatic plastic activity box, it moves freely around in the box
application of bradykinin dropped after a dorsal col- until it adapts to this new environment. The move-
umn lesion from 242% over baseline back to 100%. ments can be tracked automatically by computer.
The effects of noxious stimulation of the pancreas Beams of ultraviolet light cross the plastic box, and
with bradykinin involved a spinal synaptic relay whenever a beam is blocked by the animal, this is
rather than a vagal relay since thalamic activation counted as a movement. Under normal conditions,
was prevented by intrathecal administration of mor- the exploratory activity gradually decreases over
phine. This action of morphine could be antagonized about 45 min. However, if the animal experiences
by naloxone. pain, for example, because of the presence of
Visceral responses can also be recorded from neu- allodynia or hyperalgesia, the exploratory activity
rons in the ventral posterior lateral nucleus in monkeys decreases more rapidly.

Figure 3 (A) shows the effects of sequential lesions of the dorsal columns (DC) and of the ventrolateral column (VLC) on one
side on the responses of a neuron in the rat ventral posterior thalamic nucleus to a graded series of mechanical and visceral
stimuli. The filled circle in (a) shows the recording site; the hatched area in (b) the somatic receptive field, and the black areas
on a drawing of a transverse section of the spinal cord in (c) the extent of the lesions. Responses to brushing the skin (BR),
application of pressure to a fold of skin with an arterial clip (PR) and pinching the skin with a stronger arterial clip (PI) are seen
in (d), before and after the lesions. The responses to colorectal distentions of 20, 40, 60, and 80 mmHg are shown in (e), before
and after the lesions. The action potential of the thalamic neuron at different times during the experiment is illustrated in (f) to
indicate the stability of the recording. (B) Shows the effects of lesions of the dorsal columns placed in the upper cervical spinal
cord at the midline or bilaterally just ventral to the dorsal intermediate sulci on the responses of a neuron in the rat ventral
posterior lateral nucleus. In (a), upper row of records, are shown the responses of the neuron to graded distensions of the
duodenum. The volumes of fluid injected into a balloon at the end of a catheter inserted into the duodenum are indicated in ml.
The middle row of records (b) was taken after a midline lesion was made in the dorsal columns. The lesion had no clear effect.
The lower row of records (c) shows that bilateral lesions of the dorsal columns ventral to the dorsal intermediate sulci
substantially reduced the responses. The action potential of the thalamic neuron at different times during the experiment is
shown in (b) to indicate the stability of the recording. The midline lesion is indicated by the hatched area and the more laterally
placed lesions by the black area in (d). The receptive field of the neuron is shown in (e) and the recording site in (f) (A) From Al-
Chaer, E. D., Lawand, N. B., Westlund, K. N., and Willis, W. D. 1996a. Visceral nociceptive input into the ventral posterolateral
nucleus of the thalamus: a new function for the dorsal column pathway. J. Neurophysiol. 76, 26612674. (B) From Feng, Y.,
Cui, M., Al-Chaer, E. D., and Willis, W. D. 1998. Epigastric antinociception by cervical dorsal column lesions in rats.
Anesthesiology 89, 411420.
534 Dorsal Columns and Visceral Pain

Exploratory activity is also reduced in rats with of the posterior thalamus were compared, before
pancreatitis (Houghton, A. K. et al., 1997), and this and after surgery (Figure 5). In one animal, sham
reduction is counteracted following a lesion of the surgery was done over the dorsal column, but no
dorsal columns. Similarly, infusion of bradykinin into lesion was made (Figure 5, left-most images). In the
the common bile and pancreatic duct in rats results in other three animals, the dorsal columns were
a reduction in rearing behavior, and this vertical interrupted at a mid-thoracic level. At various times
exploratory behavior is increased after a dorsal col- after recovery from the surgery, the animals were
umn lesion (Houghton, A. K. et al., 2001). Since the re-anesthetized and fMRIs repeated at several
pancreas is an intra-abdominal organ, rather than a intervals up to 4 months after the surgery. The sham
pelvic organ, the dorsal column had to be interrupted surgery had no obvious effect on the regional cerebral
bilaterally and it included the areas ventral to the blood flow produced by noxious colorectal distention.
dorsal intermediate sulci (see below and Wang, C. C. However, the lesion of the dorsal columns completely
et al., 1999). eliminated the blood flow changes in all three animals,
Feng Y. et al. (1998) inserted a balloon into the and this effect of the lesion persisted for as long as the
duodenum through a catheter. Distention of the bal- animals were followed (Figure 5).
loon resulted in contractions of the abdominal
muscles that were graded in force depending on the 36.2.3.5 Blockade of synaptic relay in
amount of balloon distention. Interruption of the sacral cord by morphine or 6-cyano-7-
dorsal columns resulted in a reduction in these nitroquinoxaline-2,3-dione
abdominal muscle contractions. The experiments described above indicate that the
The role of the dorsal column versus that of the dorsal columns contain axons that signal visceral
spinothalamic tract and associated pathways was pain. However, a lesion of the dorsal columns inter-
tested by Palecek J. et al. (2002). Exploratory activity rupts not only the ascending branches of dorsal root
of rats was shown to be reduced following either an ganglion cells that project to the dorsal column nuclei
intradermal injection of capsaicin (Figure 4(a)) or by but also axons belonging to the postsynaptic dorsal
inflammation of the colon with mustard oil, coupled column pathway (reviewed in Willis, W. D. and
with a mild degree of colorectal distention Coggeshall, R. E, 2004). An experiment was therefore
(Figure 4(c)). The effects of a capsaicin injection on designed to determine if noxious visceral signals are
exploratory activity could be reversed by dorsal rhi- conveyed by the direct dorsal column projection or by
zotomies on the side of the injection or by a lesion of postsynaptic dorsal column neurons (Al-Chaer, E. D.
the ventrolateral column of the spinal cord on the et al., 1996b). The experimental arrangement is shown
side opposite to the injection, but not by a bilateral in Figure 6.
lesion of the dorsal column (Figure 4(b)). By contrast, Recordings were made from the nucleus gracilis,
the effects of colon inflammation and distention on rather than from the thalamus, to avoid the
exploratory activity were eliminated by a bilateral complication of visceral signals conveyed by the spi-
lesion of the dorsal columns, and this change per- nothalamic and accompanying pathways, as well as
sisted for at least 90 days (Figure 4(d)). by the dorsal columns. The visceral stimulus was
graded colorectal distention. The location of the
36.2.3.4 Effects of a dorsal column lesion part of the spinal cord that relayed the information
on the regional cerebral blood flow to the medulla was restricted to the sacral segments
changes that result from colorectal by transecting the hypogastric nerves bilaterally. A
distention in monkeys microdialysis fiber was inserted across the sacral cord
In each of four monkeys anesthetized with isoflurane, in order to introduce drugs into the dorsal horn that
it was possible to survey the changes in regional could block synaptic transmission but that would not
cerebral blood flow that were produced by colorectal interfere with direct nerve impulse transmission. The
distention, using a 4.7 T magnet for functional drugs included morphine and the non-N-methyl-
magnetic resonance imaging (Willis, W. D. et al., D-aspartic acid (NMDA) glutamate receptor antago-
1999). The images were averaged over a standard nist, 6-cyano-7-nitroquinoxaline-2,3-dione (CNQX).
period of time with or without visceral distention. The prediction was that these drugs would prevent
The intensity of the colorectal distention was to the responses of neurons of the gracile nucleus to
80 mm of Hg, well into the noxious range. Images colorectal distention if postsynaptic dorsal column
through a coronal section of the brain at the level neurons were responsible for the responses, but not
 Dorsal Columns and Visceral Pain 535

(a) (b)
400 600
Entries CAP Entries RHIZ-I
400
200
200
0 0
600
Distance traveled Entries RHIZ-C
1000 400
500 200
0 0
600
3000 Entries STT-C
400
2000
200
1000 Resting 0
0 10 20 30 40 50 600
Entries STT-I
Time (min.) 400
200
0
CONTROL 600
Entries DC
SURG. CONTROL 400
CAP 0.5h 200
CAP 24h
0
0 10 20 30 40 50
Time (min.)
(c) (d)
600 600
Entries COLON
Entries DC
400 400
200 200
0
0
2000 600
Distance traveled Entries DC 90
400
1000
200

0 0
3000 600
Entries DC 180
2000 400
1000 200
Resting
0 0
0 10 20 30 40 50 0 10 20 30 40 50
Time (min.) Time (min.)
CONTROL SURG. CON. COLON 0.5h COLON 24h

Figure 4 Time course of exploratory activity as monitored by a computer after introduction of rats into an activity box. The
level of exploratory activity is determined by the number of interruptions of infrared light beams over a period of 45 min after
the rat was placed in the activity box. The parameters included number of entries into a different zone, total distance traveled,
and resting time. In (a) and (b), an intradermal injection of capsaicin was given, whereas in (c) and (d) mustard oil was injected
into the colon and a latex balloon inserted and inflated to a pressure of 30 mmHg. At 30 min following either stimulus,
exploratory activity was tested. Note that in (a), the capsaicin (CAP) injection resulted in a significant reduction in exploratory
activity (and increased resting time). In (b), this was prevented by a prior extensive dorsal rhizotomy on the side ipsilateral to
the injection (RHIZ-I), but was unaffected by a contralateral dorsal rhizotomy (RHIZ-C). A lesion that interrupted the
spinothalamic tract contralateral to the injection (STT-C) also eliminated the reduction in exploratory activity following CAP,
whereas an ipsilateral cordotomy (STT-I) had no effect; nor did a bilateral dorsal column (DC) lesion. In (c), colon inflammation
and distention reduced the exploratory activity, and in (d) this change was eliminated by a dorsal column (DC) lesion. The
effect lasted for 90 (DC 90) and 180 (DC 180) days. From Palecek, J., Paleckova, V., and Willis, W. D. 2002. The roles of
pathways in the spinal cord lateral and dorsal funiculi in signaling nociceptive somatic and visceral stimuli in rats. Pain 96,
297307.
536 Dorsal Columns and Visceral Pain

Figure 5 Functional magnetic resonance imaging showing the changes in regional cerebral blood flow in the brains of four
different monkeys in coronal slices made at the level of the posterior thalamus. The blood flow changes were evoked by
repeated colorectal distention to 80 mmHg under isoflurane anesthesia. The images in the upper row were made prior to
surgery. Those in the lower row were from the same animals but after surgery. The animal whose images are shown at the left
was subjected to sham surgery: the dorsal column was exposed at a mid-thoracic level, but no lesion was made. In the other
three animals, the dorsal columns were interrupted at a mid-thoracic level. There was no obvious difference in regional blood
flow changes in the animal with sham surgery, but the blood flow changes in the other animals were completely eliminated
(previously unpublished data.).

if the direct dorsal column pathway were involved. made in this region from a number of postsynaptic
This was important, since it was conceivable that dorsal column neurons that responded to noxious
noxious visceral signals could be transmitted to the visceral stimuli (Al Chaer, E. D. et al., 1996a; 1996b).
dorsal column nuclei by unmyelinated, peptidergic Many cells around the central canal have ascending
branches of dorsal root ganglion neurons. Such fibers projections in the dorsal columns that are labeled
have been demonstrated, although the functions of following injections of retrograde tracer into the
these axons are still unknown (see Willis, W. D. and dorsal column nuclei or the dorsal column itself
Coggeshall, R. E., 2004). The results of this experi- (Hirshberg, R. M. et al., 1996). The nociceptive visc-
ment indicated that the postsynaptic dorsal column eral information is likely to be relayed from the
pathway is responsible for the visceral pain signals dorsal column nuclei to the lateral thalamus by
that ascend in the dorsal columns. Both morphine and way of the medial lemnsicus.
CNQX blocked visceral nociceptive transmission However, recent observations indicate that many
from the colon to the gracile nucleus, and the effects postsynaptic dorsal column neurons that are acti-
of morphine were reversed by naloxone, indicating vated by noxious visceral stimuli are also located in
that the morphine acted on opiate receptors the nucleus proprius of the spinal cord dorsal horn
(Figure 7). The CNQX must have exerted its effect (Palecek, J. et al., 2003b), presumably adjacent to
by blocking a critical synaptic relay that involves postsynaptic dorsal column neurons that lack visceral
non-NMDA glutamate receptors. input (Figure 8).
Postsynaptic dorsal column neurons in lamina X of
36.2.3.6 Projections of the postsynaptic the sacral cord were found to project their axons
dorsal column pathway through the medial part of the fasciculi gracilis to
The course of the ascending axons of postsynaptic the nuclei gracilis (Figure 2(b); Wang, C. C. et al.,
dorsal column neurons in the sacral spinal cord was 1999). Thus, the projections of these postsynaptic
traced using the Phaseolus vulgaris leukoagglutinin dorsal column cells are in exactly the position in the
(PHA-L) anterograde tracing technique and com- medial posterior columns that was targeted by the
pared with the projections from postsynaptic dorsal Hitchcock procedure (Hitchcock, E. 1970; 1974;
column neurons in the mid-thoracic spinal cord 1977) and by limited midline myelotomy (see
(Wang, C. C. et al., 1999). Injections of PHA-L Hirshberg, R. M. et al., 1996; Nauta, H. J. W. et al.,
were made into the central region of the spinal 1997; 2000). These axons would also be interrupted by
cord for several reasons. This area is known to a commissural myelotomy. However, axons that
receive visceral afferent input (Honda, C. N., 1985; ascend from postsynaptic dorsal column neurons in
Sugiura, Y. et al., 1989), and recordings had been the mid-thoracic cord to the lateral part of the gracilis
 Dorsal Columns and Visceral Pain 537

NG

DCT10

h.n.

S1 Microdialysis fiber

p.n.

Figure 6 Drawing of the experimental arrangement. Recordings were made of the responses of neurons in the nucleus
gracilis (NG, upper right) in response to colorectal distension. This avoided the problem that would have been
encountered with thalamic recordings, since neurons of the ventral posterior lateral nucleus could be affected by either
the dorsal column pathway or the spinothalamic tract. The input from the colon to the spinal cord was limited to just that
traversing the pelvic nerve and ending in the sacral spinal cord. This was done by sectioning the hypogastric nerves prior
to the experiment. A microdialysis fiber was placed across the spinal cord at S1 so that drugs that are likely to block
synaptic transmission in the dorsal horn could be administered locally. Access to the dorsal column (DC) at T10
permitted a test of the effect of a dorsal column lesion. From Al-Chaer, E. D., Lawand, N. B., Westlund, K. N., and Willis,
W. D. 1996a. Visceral nociceptive input into the ventral posterolateral nucleus of the thalamus: a new function for the
dorsal column pathway. J. Neurophysiol. 76, 26612674.

nucleus and the medial part of the cuneate nucleus do failed to influence the responses. However, lesions
so near the dorsal intermediate septum (Figure 2(b); lateral to the midline in the area of the dorsal inter-
Wang, C. C. et al., 1999). Thus, there appears to be a mediate septum were effective. Clinical studies from a
viscerotopic organization of this visceral pathway group in Korea (Kim, Y. S. and Kwon, S. J., 2000)
similar to that of the somatosensory part of the post- report favorable results in patients experiencing sto-
synaptic dorsal column pathway (see Willis, W. D. and mach cancer pain when lesions were made near the
Coggeshall, R. E., 2004). The results of the studies of incoming thoracic dorsal root fibers rather than at the
stimulation of internal organs of the abdomen midline. This indicates that the viscerotopic organiza-
described above by Houghton A. K. et al. (1997; tion is similar in humans and animals and that off-
2001) and Feng Y. et al.(1998) are consistent with this midline myelotomies will offer better relief of thoracic
observation. In these studies, distention or chemical or upper abdominal visceral pain than do midline
irritation of abdominal viscera was used to activate myelotomies.
neurons in the rat ventral posterior lateral nucleus.
Lesions placed in the dorsal column at the midline
538 Dorsal Columns and Visceral Pain

(a) (b)

(c)
Control
BR PR PI
(d) (e)
100
75
50
25
0

Morphine
100
75
50
Rate (spikes s1)

25
0

Naloxone
100
75
50
25
0

CNQX
100
75
50
25
0
0 10 20 30 40 50 60 70 0 20 40 60 0 20 40 60 0 20 40 60 0 20 40 60
Time (s) Time (s)

20 40 60 80 mm Hg

Figure 7 Effects of morphine and CNQX administered into the sacral cord by microdialysis on the responses of a neuron in
the gracile nucleus to graded somatic and visceral stimuli. (a) Shows the recording site in the gracile nucleus. (b) Indicates the
receptive field of the gracile neuron. In (c) are the responses of the gracile neuron to BR, PR, and PI stimuli applied to the
cutaneous receptive field. (d) Shows the responses to graded colorectal distention to 20, 40, 60, and 80 mmHg. The upper
row of recordings in (c) and (d) are the control responses taken before administration of drugs. The second row of recordings
shows the effects of morphine given by microdialysis. There was little, if any, change in the responses to the somatic stimuli,
but the responses to colorectal distention were eliminated. The third row of recordings shows that systemically administered
naloxone counteracted the effects of morphine. The lowest row of recordings shows the effect of CNQX in blocking the
responses to colorectal distention. In (e) are action potentials recorded at different times during the experiment to show the
stability of the recordings. From Al-Chaer, E. D., Lawand, N. B., Westlund, K. N., and Willis, W. D. 1996b. Pelvic visceral input
into the nucleus gracilis is largely mediated by the postsynaptic dorsal column pathway. J. Neurophysiol. 76, 26752690.

36.2.3.7 Upregulation of NK1 receptors in postsynaptic dorsal column pathway that express Fos,
PSDC neurons after colon inflammation the product of the immediate/early gene, c-fos, follow-
The cells of origin of several nociceptive pathways that ing either an intradermal injection of capsaicin or
ascend from the spinal cord to the brain are known to distention of the ureter (see Figure 8; Palecek, J. et al.,
contain neurokinin-1 (NK-1) receptors (Todd, A. J. 2003b). The spinothalamic neurons were retrogradely
et al., 2000). These pathways include the spinothalamic labeled from the contralateral thalamus and the postsy-
and spinoreticular tracts. However, postsynaptic dorsal naptic dorsal column neurons from the ipsilateral
horn neurons do not contain NK-1 receptors under nucleus gracilis. Ureter distention was accomplished by
normal conditions, although an up-regulation of these tightening a ligature that had previously been placed
receptors does occur in some of these neurons follow- around the ureter. The ureter was distended proximal
ing visceral inflammation (Palecek, J.et al., 2003a). to the site where the ligature blocked the flow of urine.
Fos-labeled neurons were distributed over a consider-
36.2.3.8 Fos expression in PSDC neurons able length of the spinal cord, from T11 to L6. This study
after noxious visceral stimulation confirmed that spinothalamic tract cells can be activated
A comparison was made of the numbers and locations of by a noxious visceral stimulus and that postsynaptic
neurons belonging to the spinothalamic tract and to the dorsal column neurons are also activated by such stimuli.
 Dorsal Columns and Visceral Pain 539

PSDC Ureter STT

A B C D
Fos Fos+ Fos+ Fos
Th4

Th11

Th12

Th13

L1

L2

L3

L4

L5

L6

Figure 8 Effect of distention of the ureter in evoking expression of Fos protein in identified projection neurons in rats.
Postsynaptic dorsal column neurons were labeled retrogradely following injection of a tracer into the nucleus gracilis (left-
most column), and spinothalamic neurons were identified following injection of a different retrograde tracer into the ventral
posterior lateral nucleus of the thalamus (right-most column). The middle columns show neurons identified by retrograde
labeling that were also labeled for Fos protein following distention of the ureter. From Palecek, J., Paleckova, V., and Willis, W.
D. 2003b. Fos expression in spinothalamic and postsynaptic dorsal column neurons following noxious visceral and cutaneous
stimuli. Pain 104, 249257.

36.3 Descending Facilitation enhance the responses of spinal cord neurons to

noxious visceral stimuli.
It has been shown that the responses of spinal cord Palecek J. and Willis W. D. (2003) confirmed that a
neurons to visceral stimulation depend in large part dorsal column lesion did not affect visceromotor
on a spino-bulbo-spinal circuit (Cervero, F. and reflex responses under normal conditions (cf., Ness,
Wolstencroft, J. H., 1984; Tattersall, J. E. et al., 1986; T. J. and Gebhart, G. F., 1988). However, the
Zhuo, M. and Gebhart, G. F., 2002) that when acti- enhanced visceromotor responses to graded colorectal
vated facilitates visceral responses at the spinal cord distention that followed colon inflammation with
level. Our anterograde tract tracing studies of the mustard oil were reduced or prevented by bilateral
projections from lamina X demonstrate an ascending lesions of the dorsal columns. This suggests that the
component that joins the medial edge of the spi- visceral reflex is enhanced because of activity induced
nothalamic tract (Wang, C. C. et al., 1999). This by the colon inflammation and conveyed to the brain-
ascending pathway has terminations in the nucleus stem by way of the dorsal column. A lesion of the
raphe magnus, rostral ventromedial medulla, and ventrolateral spinal cord eliminated the visceromotor
medial reticular formation. This pathway is the reflex, presumably by interrupting a facilitatory path-
only reported direct input to the nucleus raphe mag- way that descends from the brainstem and/or the
nus from the spinal cord and is likely to be important ascending spinothalamic projection from lamina X.
for the activation of brainstem facilitatory neurons It is our working hypothesis that interruption of the
with axons that descend into the spinal cord to postsynaptic dorsal column pathway by a lesion placed
540 Dorsal Columns and Visceral Pain

in the dorsal columns reduces visceral pain not only Multimodal and discriminative properties of thermosensitive
neurons. J. Neurophysiol. 69, 739752.
because such a lesion prevents visceral pain signals from Casey, K. L. and Bushnell, M.C. (eds.) (2000) Pain Imaging. IASP
reaching the thalamus by way of the dorsal column Press.
nuclei and medial lemniscus, but also because of a Cervero, F. and Wolstencroft, J. H. 1984. A positive feedback loop
between spinal cord nociceptive pathways and antinociceptive
reduction of descending facilitation affecting visceral areas of the cats brain stem. Pain 20, 125138.
nociceptive neurons in the spinal cord, including post- Chandler, M. J., Hobbs, S. F., Qing-Gong, Fu., Kenshalo, D. R.,
synaptic dorsal column and spinothalamic tract neurons. Blair, R. W., and Foreman, R. D. 1992. Responses of neurons
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Chung, J. M., Lee, K. H., Surmeier, D. J., Sorkin, L. S., Kim, J.,
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 37 Visceral Pain
G F Gebhart and K Bielefeldt, University of Pittsburgh, Pittsburgh, PA, USA
2009 Elsevier Inc. All rights reserved.

37.1 Introduction 544

37.2 Visceral Sensation 544
37.2.1 Organization of Visceral Sensory Innervation 545
37.2.2 Visceral Nociceptors and Sensory Endings 545
37.2.3 Chemical Character of Visceral Sensory Neurons 547
37.2.4 Central Organization of Visceral Pain 549
37.2.4.1 Vagus nerves 549
37.2.4.2 Spinal nerves 549
37.2.4.3 Spinal pathways and supraspinal terminations 550
37.3 Functional Basis of Visceral Pain 551
37.3.1 Visceral Mechanoreceptors and Mechano-Nociceptors 552
37.3.1.1 Afferent fiber recordings 552
37.3.1.2 Information acquired in vitro 553
37.3.1.3 Gene deletions, visceral pain, and mechanosensation 556
37.3.2 Sleeping (Silent) Nociceptors 557
37.3.3 Visceral Chemo-Nociception 558
37.4 Visceral Hypersensitivity 559
37.4.1 Sensitization and Excitability of Visceral Nociceptors 560
37.4.2 Central Sensitization 561
37.4.3 Central Modulation of Visceral Pain 563
37.5 Summary 564
References 565

Glossary (stretch/tension, ischemia, inflammatory media-

referred visceral pain Visceral pain is not gener- tors), commonly give rise to sensations of
ally felt at the source, but rather is perceived as discomfort and pain.
arising from other tissues (skin and/or muscle) visceral hypersensitivity Increased sensitivity or
innervated by non-visceral nerves having input onto response to a visceral stimulus, which can arise
second-order spinal neurons that converges with from sensitization of visceral sensory endings
input from visceral organs onto the same spinal (e.g., visceral mechanoreceptors), sensitization of
neurons. Accordingly, visceral pain is commonly central nervous system neurons receiving
referred (or transferred, an older terminology) to increased input from sensitized visceral sensory
non-visceral tissues. endings, or a combination of both. Clinically,
nociceptor sensitization Sensitization of noci- visceral hypersensitivity is associated with dis-
ceptors reflects a change in their excitability comfort or pain produced by normally non-pain-
expressed as an increase in response magnitude to producing stimuli (e.g., ingestion of food, normal
a noxious intensity of stimulation accompanied by a bladder fillling) and commonly includes increased
reduction in threshold stimulus intensity required to sensitivity to probing in the areas of referred visc-
activate the nociceptor. eral sensation, which moreover are increased in
visceral nociceptors Sensory endings in viscera size (area).
which, when activated by an adequate stimulus

543
544 Visceral Pain

37.1 Introduction Section 37.3. We are generally unaware of movement

of air in and out of the lungs, blood flowing through
Of all possible sources of pain within the body, pain vessels, beating of the heart, or food in the stomach
arising from internal organs commonly generates the despite the fact that sensory receptors in the bron-
greatest autonomic and emotional responses. This is chial tree, lungs, vessels, heart, esophagus, and
linked to several characteristics of visceral pain, the stomach are activated by these events. Information
most apparent being that the source of pain is neither from activation of visceral sensory neurons is faith-
visible nor typically felt at the source. Visceral fully transmitted to the central nervous system, but
pain also is commonly diffuse in character and rarely is perceived. Rather, activation of most visceral
poorly localized. In addition, it is also referred (or sensory receptors plays important roles in the regu-
transferred) to nonvisceral, somatic structures (like lation and maintenance of many essential functions
skin and muscle), and there is considerable overlap in (e.g., respiration, blood flow, food digestion), which
areas of referred pain from adjacent organs. For requires rapid feedback to adjust to changes in
example, sites of referral from thoracic and abdom- demand due to exertion, food intake or other activ-
inal organs (esophagus, gallbladder, and heart) ities. While autonomic functions associated with the
overlap, making it difficult for both physicians and visceral afferent innervation can influence affective
patients to determine the source of pain. That diffuse and cognitive processes and are, in turn, influenced
substernal chest pain has high emotional valence is by affective and cognitive processes, most of the
readily appreciated, given the potential significance afferent input from the viscera never reaches
of such pain. Similarly, pelvic genitourinary organs consciousness.
have overlapping patterns of referral with the distal Visceral events, of course, do lead to conscious
colon. It is also characteristic of visceral pain that sensations. Under physiological conditions, humans
the areas of referred sensation increase in size and perceive a sense of fullness related to organs involved
sensitivity in organ inflammation and disease. For in food intake (i.e., stomach) or waste elimination
example, repetitive episodes of angina lead to (rectum and bladder). Aside from these sensations
increased tenderness (hyperalgesia) of chest and related to functions that trigger intentional changes
shoulder skin and abdominal tenderness spreads and in behavior, most consciously perceived input from
increases in area in interstitial cystitis and irritable the viscera carries negative connotations, such as
bowel syndrome (IBS). nausea, palpitation, dyspnea, discomfort (e.g., over-
Distinct from cutaneous, muscle, or joint pain, filling, gas, bloating), and pain. Normally, visceral
tissue-damaging stimuli do not commonly produce pain is infrequent, but in cases of functional gastro-
pain when applied to the viscera. Tissue crushing, intestinal disorders (e.g., nonulcer dyspepsia, IBS (see
cutting, or burning stimuli applied to visceral Chapter Irritable Bowel Syndrome), noncardiac
tissue generally produce little conscious sensation chest pain, functional abdominal pain syndrome),
and rarely produce pain. Instead, stretch of the interstitial cystitis/painful bladder syndrome, inflam-
smooth muscle layers of hollow organs (e.g., by matory visceral disorders (e.g., pancreatitis,
distension), traction on the mesentery, ischemia, inflammatory bowel disease), neoplasias, etc., dis-
and inflammation are more reliably uncomfortable comfort and pain are common, persistent, and
or pain-producing in viscera. The bases of the above typically difficult to treat.
attributes of visceral pain and unique features that The so-called functional visceral disorders are
distinguish visceral from nonvisceral pain are the characterized by pain and discomfort and enhanced
focus of this chapter. sensitivity to stimuli in the absence of a demonstrable
organic cause (i.e., there are no apparent structural,
biochemical, or inflammatory conditions to explain
the symptoms). Such functional disorders constitute a
37.2 Visceral Sensation significant socioeconomic burden worldwide; the
estimated prevalence of abdominal pain syndromes
The viscera are invested with a wide range of sensory can range widely for different specific syndromes, but
receptors, although when activated in healthy organs prevalence rates typically range between 22% and 28%
they do not generally give rise to conscious sensation. of the adult population (Halder, S. L. and Locke, G. R.,
This raises the issue of whether these neurons are 2007). Characteristically, these and other visceral dis-
afferent or sensory, which is considered below in orders are characterized by hypersensitivity, meaning
 Visceral Pain 545

that normally non-pain-producing visceral events are and, thus, traverse pre- and paravertebral ganglia en
perceived as uncomfortable or painful, which is likely route to the spinal cord (Figure 1). In prevertebral
contributed to by both peripheral and central nervous ganglia, some afferent axons give rise to collateral
system mechanisms. branches that synapse on intraganglionic neurons and
influence organ function (e.g., motility, secretion).
37.2.1 Organization of Visceral Sensory
Innervation 37.2.2 Visceral Nociceptors and Sensory
Endings
The viscera are unique among all tissues in the body.
Each internal organ is innervated by two nerves, which Because tissue-damaging crushing, cutting, or burn-
share some functions, but are not physiologically ing stimuli that generate pain from skin do not
redundant and include important functional distinc- commonly produce pain when applied to the viscera,
tions. Anatomically, the visceral sensory innervation is it was long argued that the viscera were not inner-
physically associated with the sympathetic and para- vated, and when visceral innervation was established,
sympathetic divisions of the autonomic nervous it was argued that the viscera were insensate. It is now
system. Correspondingly, Langley J. N. (1921) termed well established that stimuli adequate for activation
visceral afferent fibers associated with sympathetic of visceral nociceptors differ from those adequate for
nerves afferent sympathetic fibers. This sometimes activation of cutaneous nociceptors. They include
confusing nomenclature has been replaced by use of distension of hollow organs, traction on the mesen-
the name of the nerve (e.g., inferior cardiac, lumbar tery, ischemia, and inflammation. For example,
splanchnic, pelvic), which in addition avoids assigning experimental hollow organ distension reproduces in
putative function(s) to the visceral innervation. Given patients the localization, intensity, and quality of
the physical association of the visceral innervation with sensations associated with their visceral disorder.
the autonomic nervous system, visceral afferent fibers Because distension of hollow organs, as well as
are thus contained in nerves that terminate in the spinal traction on the mesentery, are mechanical events
cord (spinal nerves) or in the brainstem (vagus nerves). (stretch), it is assumed that many visceral nociceptors
The bilateral vagus nerves are perhaps the most are mechanoreceptors with receptive endings in visc-
widely distributed nerves in the body. They inner- eral smooth muscle layers or serosa where they
vate most internal organs, including all of the transduce the mechanical energy of tension/stretch
thoracic viscera (proximal esophagus, heart, bronch- into electrical activity (nerve membrane depolariza-
opulmonary system), most of the abdominal viscera tion, action potentials). There are chemo-nociceptors
(distal esophagus, stomach, small and proximal large in epithelial and subepithelial layers of visceral
intestines, liver, pancreas, etc.), and some of the pel- organs, but they have not been studied as extensively
vic viscera. The vagus nerves are mixed and contain as have mechanoreceptors and their receptive end-
both efferent and afferent axons, but most axons in ings have not been identified as they have for some
the vagus nerves (80%) are sensory (afferent) fibers mechanoreceptors.
and the remainder parasympathetic efferents. The Virtually all visceral sensory neuron axons are
nodose (primarily) and smaller, rostrally located slowly conducting, thinly myelinated A- or unmye-
jugular ganglia contain the cell bodies of vagal affer- linated C-fibers. Because mechanical events are
ent fibers, most of which terminate bilaterally with common in most organs, visceral mechanosensation
some viscerotopic organization in the medullary and endings in muscle layers of hollow organs that
brainstem nuclei of the solitary tract (NTS). Some respond to tension/stretch have been the focus of
vagal afferents, at least in nonhuman primates and most studies. Unfortunately, the morphology of
rats, project to the upper cervical spinal cord (C12), spinal nerve peripheral terminals in organs is vir-
where they may contribute to referred sensations and tually unknown; they are assumed to be free
modulate nociceptive processing within the spinal without structural specialization. However, two mor-
cord (Foreman, R., 1999). phologically distinct endings associated principally
The cell bodies of spinal afferents innervating the with vagal afferent endings have been described.
viscera are located bilaterally in dorsal root ganglia Intraganglionic laminar endings (IGLEs), which
(DRG) from the cervical to sacral spinal cord. As have been shown to be mechanosensitive, and intra-
mentioned above, their axons travel with sympa- muscular arrays (IMAs) are located in the upper
thetic efferent fibers (except for the pelvic nerves) gastrointestinal tract (Zagorodnyuk, V. P. and
546 Visceral Pain

Nodose ganglion
NTS
Paravertebral
ganglia
Vagus nerve
SCG
Cervical
MCG
StG

Prevertebral
ganglia
Thoracic
CG

SMG

IMG Lumbar

HgN
Sacral
MPG Pelvic
nerve

Figure 1 Cartoon illustrating innervation of the viscera. The vagus nerve innervates organs in the thoracic and abdominal
cavities. Visceral afferent nerves that terminate in the spinal cord also innervate organs in the thoracic and abdominal cavities
as well as those in the pelvic floor, including the genitalia (not shown). The cell bodies of vagal sensory neurons are contained
in the nodose and jugular (not shown) ganglia with central terminations bilaterally in the nucleus tractus solitarii (NTS). The cell
bodies of spinal visceral sensory neurons are contained in dorsal root ganglia interposed between the paravertebral ganglia
(the sympathetic chain) and the spinal cord and are not illustrated in this cartoon. CG, coeliac ganglion; HgN, hypogastric
nerve; IMG and SMG, inferior and superior mesenteric ganglia, respectively; MPG, major pelvic ganglion. SCG, superior
cervical ganglia; MCG, middle cervical ganglia; StG, stellate cervical ganglia.

Brookes, S. J. H., 2000; Zagorodnyuk, V. P. et al., been described for visceral spinal nerves, an
2003). As their name indicates, vagal IGLEs appear IGLE-like low threshold, slowly adapting mechanor-
as branching endings surrounding neurons within a eceptor has been reported in the colonic and rectal
ganglion of the myenteric plexus (Phillips, R. J. and innervation of the guinea-pig (Lynn, P. A. et al., 2003).
Powley, T. L., 2000; Powley, T. L. and Phillips, R. J., Whether activation of either IGLEs or IMAs give
2002). IGLEs associated with vagal afferent terminals rise to visceral pain is uncertain because it would
are present throughout much of the gastrointestinal appear, based on their essentially proximal and distal
tract, but are most densely distributed in the esopha- distribution within the gastrointestinal tract and low
gus, stomach, and duodenum (Wang, F. B. and response threshold, that neither are particularly well
Powley, T. L., 2000). Like IGLEs, IMAs are also suited to that function. Powley T. L. and Phillips R. J.
thought to be mechanosensitive, but differ in mor- (2002) have speculated that IGLEs detect rhythmic
phology and distribution from IGLEs. IMAs are long motor activity, which requires response to muscle/
arrays of terminals, typically running parallel with organ tension; IMAs also respond to muscle/organ
each other and parallel to the orientation of smooth stretch. However, the association of IMAs with
muscle cells. They bridge between the parallel term- sphincters, and because IGLEs and IMAs have
inals within one of the muscle layers of an organ and heretofore been best described and characterized
form a lattice-like network. IMA distribution within in association with vagal afferent endings, suggests
the gastrointestinal tract is more restricted than that of a role for these mechanosensors in normal physiolo-
IGLEs; IMAs associated with vagal afferent terminals gical processes (e.g., food intake, defecation) rather
are present in the stomach (cardia) and sphincters of than nociception. Recent studies suggest that vari-
the upper gastrointestinal tract and are uncommon in cose branch points of spinal afferents within the
the intestines. Although terminal end organs have not serosa or deeper layers of the gastrointestinal tract
 Visceral Pain 547

may correspond to fibers activated by high-intensity, threshold mechanosensitive fibers are rarely found.
noxious mechanical stimuli (Blackshaw, L. A. et al., Thus, peptide expression is a poor predictor of
2007). physiological properties of visceral sensory neurons,
arguing against the use of neurochemical content as
a surrogate indicator of function.
37.2.3 Chemical Character of Visceral
Subsequent studies have established by immuno-
Sensory Neurons
histochemistry, in situ hybridization, and/or current
During ontogeny, sensory neurons develop different or voltage response to ligands applied directly to cell
phenotypes dependent on their differential sensitiv- soma that some visceral sensory neurons also contain
ity and exposure to growth factors. As mentioned other peptides (e.g., vasoactive intestinal polypeptide,
above, essentially all visceral sensory neurons cholecystokinin) and express ligand-gated channels
have unmyelinated or thinly myelinated axons. In and receptors (e.g., different members of the family of
DRG, this population of sensory neurons can be transient receptor potential channels, including
separated into two largely distinct groups based on TRPV1, acid-sensing ion channels (ASICs), both
the expression of growth factor receptors or other ionotropic and metabotropic glutamate, purinergic,
neurochemical markers (Molliver, D. C. et al., 1997). serotoninergic, -aminobutyric acid (GABA)-ergic,
Small-diameter peptide-containing DRG neurons and opioid receptors). As information continues to
express the high-affinity receptor for nerve growth accumulate, it is apparent that there is no marker at
factor trkA, the temperature-, proton-, and capsaicin- present that distinguishes visceral sensory neurons
sensitive transient receptor potential vanilloid ion from nonvisceral sensory neurons or visceral noci-
channel (TRPV)1 and the neuropeptides substance ceptors from nonnociceptors.
P and/or calcitonin gene-related peptide (CGRP). It could be argued that visceral sensory neuron
The second group of small-diameter DRG cells terminals are generally exposed to a richer chemical
depends on glial cell line-derived neurotrophic factor milieu than other sensory neurons (and thus express a
(GDNF) during embryonic development, binds the wider array of receptors and channels, although this
plant lectin isolectin B4 (IB4) and releases neuro- has not been quantified). The environments in which
transmitters such as adenosine triphosphate (ATP) visceral sensory endings reside include exposure to
rather than peptides as transmitters. Immuno- histo- acids, digestive enzymes, hormones, nutrients, toxins
chemical studies have consistently shown that about (ingested or secreted by gut flora, for example), meta-
80% of visceral sensory neurons within the DRG are bolites of xenobiotics, and bioactive chemicals
peptidergic and contain CGRP or substance P. released from nearby enteric nervous system neu-
Visceral sensory neurons in the placode-derived rons, sympathetic nerve terminals, epithelial cells
nodose ganglia, however, depend on different growth like enterochromaffin cells in the gut, and urothelial
factors during development and show a different cells in the urinary bladder and mast cells. Visceral
distribution of neurochemical markers. While the sensory neurons have been shown to respond to
expression of neuropeptides CGRP and/or substance products released in these environments, including
P is closely associated with the presence of TRPV1 protons, norepinephrine, serotonin, histamine, tryp-
channels in DRG neurons, this does not hold for tase, ghrelin, cholecystokinin, and other gastric and
nodose ganglion neurons; less than 10% express neu- duodenal secretions that initiate digestion and
ropeptides CGRP or substance P, whereas about motility.
50% of nodose neurons exhibit TRPV1-like In the gastrointestinal tract, serotonin plays an
immunoreactivity. especially important role as a signaling molecule.
In studies of cutaneous or muscle sensory neurons, More than 90% of the bodys serotonin is found
expression of TRPV1, CGRP, or the presence of within the intestine with most stored in secretory
IB4 binding are often used as surrogate markers granules on the basolateral side of enteroendocrine
to classify neurons as nociceptive. As discussed cells. These enteroendocrine cells are specialized
below, 7080% of mechanosensitive spinal afferents epithelial cells that can release into the lamina pro-
innervating abdominal or pelvic viscera are activated pria a variety of mediators, including serotonin, upon
by innocuous intensities of stimulation (i.e., are mechanical or chemical stimulation, which seconda-
low-threshold mechanosensors). Most studies of rily activate extrinsic and intrinsic sensory neurons.
mechanosensitive vagal afferents, however, reveal Interestingly, the number of enteroendocrine cells
an even more homogeneous response profile; high- increases after intestinal inflammation and remains
548 Visceral Pain

elevated in patients with continuing complaints despite ultrastructural and neurochemical characteristics
resolution of the inciting infection (Spiller, R. C. with large, dense core vesicles and neuropeptide
et al., 2000), suggesting that serotonin release from expression, they are referred to as neuroendocrine
enteroendocrine cells is important in the pathogenesis cells or neuroendocrine bodies when clustered in
of functional bowel disorders. small groups (Cutz, E., 1982). Histological studies
An example illustrating the importance of luminal combining immunohistochemistry, retrograde label-
factors within the gastrointestinal tract is the interac- ing, and/or vagotomy reveal a complex innervation
tion between the PAR2 receptor and the pancreatic with vagal and spinal afferents projecting to these
enzyme trypsin, which is especially relevant under cells or cell clusters, where nerve terminals branch
pathologic conditions. The PAR2 receptor is a mem- with formation of varicosities at branch points
ber of the proteinase-activated receptor (PAR) (Adriaensen, D. I. et al., 2003). Neuroepithelial bodies
family, a group of G-protein-coupled receptors char- likely function as oxygen sensors within the airways as
acterized by a tethered peptideligand that is part of hypoxia inhibits potassium currents (Fu, X. W. et al.,
the molecule and can activate the receptor once 2000). However, acute allergic inflammation triggers
cleaved from the molecule by specific proteases. depletion of peptides from airway neuroepithelial cells,
The pancreatic protease trypsin is one of the endo- suggesting a role in broader signaling under physio-
genous proteases that cleaves the ligand and thus logic and pathophysiologic conditions (Dakhama, A.
activates the receptor. Pancreatic acinar cells secrete et al., 2002).
precursor forms of proteolytic enzymes, such as tryp- While the expression of one or another receptor
sin, that require activation within the lumen of the or ligand-gated channel of a particular subunit com-
small intestine. During pancreatitis, however, acti- position may be determined at some future time to
vated trypsin is also present in the pancreatic duct identify all or a subset of visceral sensory neurons,
and parenchyma. Experimental infusion of trypsin one feature of visceral sensory neurons may distin-
into the pancreatic duct activates pancreatic sensory guish them from nonvisceral counterparts, and that is
neurons, most of which express PAR2 immunoreac- cell size. Cutaneous nociceptors are on the whole
tivity, and triggers aversive behavior in animals. associated with thinly myelinated and unmyelinated
Selective experimental activation of PAR2 receptors axons and it is generally the case that these nocicep-
mimics this effect, suggesting an important role of tors have small-diameter cell bodies (e.g., 15
PAR2 in pain during acute pancreatitis (Hoogerwerf, 20 mm). Virtually all visceral sensory neurons have
W. A. et al., 2001; 2004). Similarly, mouse colon thinly myelinated or unmyelinated axons, yet their
sensory neurons contain immunoreactivity for the cell bodies tend to be medium in size (e.g., 25 mm in
PAR2 receptor and PAR2 agonists depolarize those diameter), at least those associated with the gastro-
neurons and cause a sustained hyperexcitability intestinal tract and urinary bladder. This is of
(Ahmed, K. et al., 2007). Relevant to IBS, biopsy importance when nociceptors are studied and identi-
samples from IBS patients release greater amounts fied principally on the basis of cell diameter because
of proteases than do samples from controls (Cenac, visceral nociceptors are not generally contained in
N. et al., 2007). Supernatants from colonic biopsies of the population of small-diameter sensory neurons
IBS patients produce visceral hypersensitivity when assumed to be nociceptors solely because of small
given intracolonically in control, but not PAR2 diameter.
knockout mice. Mast cells, which are closely asso- The foregoing addresses, at least indirectly, iden-
ciated with nerves in the human gastrointestinal tification of and assignment of function of sensory
mucosa (Stead, R. H. et al., 1989), are also a source neurons, including visceral sensory neurons, based on
of tryptase as well as other potential mediators and different properties of the cells: size (diameter), cell
modulators of visceral sensation (Bueno, L. et al., content (e.g., peptides), expression of one or another
1997; Barbara, G. et al., 2006). Mucosal biopsies channel or receptor, dependence on exposure during
from IBS patients contained mediators that increased development to different growth factors. Assignment
activity in rat mesenteric afferents, an effect inhibited of function based upon one or more of these proper-
by histamine H1 receptor blockade and serine pro- ties is unfortunately relatively common in the
tease inactivation (Barbara, G. et al., 2007). literature, and is both confusing to nondiscriminating
In the airway system, similarly specialized cells readers and misleading. It is not possible at present to
have been identified within the epithelium of the assign function (e.g., nociceptor) to a sensory neuron
intrapulmonary bronchial tree. Based on their based on any of the above properties. As discussed
 Visceral Pain 549

below, nociceptors, including visceral nociceptors, prevertebral ganglia often provided pain relief (at
are defined by response to an adequate stimulus, least for a limited period of time), the spinal visceral
and even then assignment of function can be difficult, nerves (sympathetic afferents) were inferred to be the
if not contentious, if the quality of the stimulus is conveyors of nociceptive information from the
uncertain, unknown or of limited physiologic rele- organs to the spinal cord. Their termination in the
vance, such as punctate stimulation of viscera. spinal cord is noteworthy on several counts. Firstly,
spinal visceral afferent fibers terminate in a pattern
that largely overlaps with terminations of cutaneous
37.2.4 Central Organization of Visceral nociceptors: superficial layers of the spinal dorsal
Pain horn, deeper in lamina V and dorsal to the central
37.2.4.1 Vagus nerves canal, an area often referred to as lamina X. Visceral
Although they extensively innervate the viscera, the afferent fibers also terminate within the interomedio-
vagus nerves are generally considered not to play a lateral cell column/sacral parasympathetic nucleus
role in nociception. However, they may contribute to where afferent input influences efferent output back
the complex sensory experience of visceral pain, to the same as well as to other organs (Figure 2).
which includes strong autonomic reactions, including Secondly, spinal visceral afferents represent less
nausea or dyspnea. Vagal afferents project to the than 10%, and probably closer to 5%, of the total
bilaterally located nuclei of the solitary tract in the afferent input into the spinal cord from all tissues. In
dorsal medulla, an important relay station for visceral compensation for this relatively sparse input, the
input. Vagal input reaches more rostral brain struc- central projection of a single visceral afferent fiber
tures involved in autonomic regulation, such as the bifurcates at its spinal segment of entry in the dorsal
hypothalamus, supraoptic nucleus, and via the root into caudally and rostrally directed main
parabrachial nucleus and ventro-medial thalamus branches that can extend in either the dorsal funicu-
the insular cortex, anterior cingulate cortex, and lus or Lissauers tract for two or three spinal segments
amygdala, regions that play a role the regulation of before penetrating the spinal dorsal horn. In addition,
affective responses to different stimuli including pain. during their longitudinal journey these main
branches give off multiple collateral branches into
37.2.4.2 Spinal nerves the spinal dorsal horn (superficial laminae and lami-
Because vagotomy was found to be generally ineffec- nae V and X, including contralateral laminae V and
tive in relieving visceral pain, whereas spinal nerve X) where, moreover, their number of terminal swel-
transaction or destruction of sympathetic lings are greater than found on cutaneous C-fiber

Figure 2 Distribution of visceral afferent terminals in the thoracic and sacral spinal cord. Photomicrographs illustrate
internalization of the substance P receptor (yellow) in the superficial dorsal horn and area dorsal to the central canal in rat T13 and
S1 spinal cord sections. Internalization of the substance P receptor was produced by distension of the colon in the rat (Honore, P.,
Kamp, E. H., Rogers, S. D., Gebhart, G. F., and Mantyh, P. W. 2002. Activation of lamina I spinal cord neurons that express the
substance p receptor in visceral nociception and hyperalgesia. J. Pain 3, 311). SPN, sacral parasympathetic nucleus.
550 Visceral Pain

Brain consciousness and triggers responses/behavior.

DRG
Interestingly, thalamic and cortical neurons that
receive a visceral input also exhibit viscerosomatic
and viscerovisceral convergence (e.g., Apkarian, A.
et al., 1995) (Figure 4). The ascending pathways in
spinal cord white matter that convey visceral infor-
mation to the brain are contained principally in the
anterolateral quadrants of the spinal cord and in the
dorsal columns. Thus, there are spinomedullary, spi-
nopontine, spinomesencephalic, spinohypothalamic,
and spinothalamic tracts that ascend in the anterolat-
eral quadrant of the spinal cord, predominantly
contralateral to the side of input. The postsynaptic
Figure 3 Illustration of viscerosomatic and viscerovisceral
convergence of inputs onto a second-order spinal neuron. dorsal column paths are ipsilateral to the side of input
Illustrated is input from the abdominal skin, urinary bladder, and ascend to the nuclei gracilis and cuneatus in the
and distal colon onto the same spinal neuron. Activation of medulla, where they cross and ascend to the contral-
the second-order neuron is illustrated here as being ateral ventrobasal thalamus (e.g., Palecek, J. et al.,
conveyed to the brain via the anterolateral ascending
2002). In addition to these principal pathways that
pathway; not illustrated is a postsynaptic dorsal column
pathway, which also conveys visceral sensory information convey information to supraspinal sites, there are
to the brain. DRG, dorsal root ganglion. intraspinal propriospinal pathways that are less well
understood and presumably have modulatory func-
tions and another pathway from lamina I neurons
terminations within the spinal dorsal horn (which are to pontine parabrachial nuclei and thence to the
typically limited to the spinal segment of entry). amygdala, principally the central nucleus. This latter
These anatomic characteristics of visceral afferent pathway is believed to specifically convey visceral
spinal terminations surely contribute to the diffuse information of a noxious character to the amygdala,
nature of visceral pain and difficulty in localizing its which importantly is associated with anxiety and
source. affective dimensions of pain (Phelps, E. A. and
A final notable and defining characteristic of LeDoux, J, E., 2005).
spinal visceral input that also contributes to poor Functional imaging studies have complemented
localization is convergence. It is a feature of virtually and expanded our appreciation of the central repre-
all second-order spinal neurons upon which visceral sentation of visceral inputs. Although the current
afferent fibers terminate that convergent inputs from spatial resolution of imaging methods has limited
somatic and/or other visceral organs are also ability to study the initial processing of visceral
received. Typically, a single dorsal horn neuron input in the brainstem, reports to date suggest that
that receives a visceral input (e.g., from colon) has a cutaneous and visceral stimuli activate similar brain
convergent cutaneous receptive field and also regions. Pain-induced anxiety, which tends to be
receives input from another viscus (e.g., urinary blad- more significant during visceral stimulation, is asso-
der, uterus) (Figure 3). Thus, viscerosomatic and ciated with stronger activation of the midbrain
viscerovisceral convergence upon second-order periaqueductal gray (PAG) (Dunckley, P. et al.,
spinal neurons is the general rule (rather than the 2005). Similarly, activation of cortical structures
exception), and further compromises localization of does not show a pattern during visceral pain distinct
visceral inputs. from somatic pain, with the anterior cingulate cortex
(ACC), the anterior insular cortex and less consis-
37.2.4.3 Spinal pathways and supraspinal tently the primary and secondary sensory cortices
terminations activated by both visceral and somatic pain (Hobson,
Spinal visceral afferent input is further transmitted A. R. and Aziz, Q., 2004). Despite the overall similar-
rostrally and distributed widely in the brainstem ity between brain patterns of activation produced by
(largely associated with reflex functions; e.g., micturi- cutaneous and visceral stimuli, subtle differences
tion), hypothalamus, and thalamus and then to areas have emerged. Compared to noxious cutaneous sti-
of the cerebral cortex where discriminative and affec- mulation, visceral pain tends to cause a stronger
tive components of visceral information assumes activation of the ACC and typically activates the
 Visceral Pain 551

Response to Cutaneous
visceral stimulation receptive fields

+CRD, oED

WDR
+CRD, ED 3V
WDR
+CRD, oED

+CRD, +ED
VL LT
+CRD, oED
VM
VPL WDR

+ Excitation VPM

Inhibition 1 mm LT
o No response

Figure 4 Illustration of viscerosomatic and viscerovisceral convergence onto neurons recorded in the rat thalamus. The
vertical tract recording electrode through the ventral posteriolateral (VPL) thalamus illustrates, from top to bottom, cutaneous
receptive fields on the right for five sequential thalamic neurons and responses of the same neurons to colorectal distension
(CRD) and esophageal distension (ED) on the left. All neurons studied responded to both CRD and either innocuous
cutaneous stimulation (identified as low threshold, LT) or both innocuous and noxious cutaneous stimulation (identified as
wide dynamic range, WDR). Some neurons were also inhibited, excited or did not respond to ED. From Danzebrink R. M. and
Gebhart G. F. (unpublished).

perigenual portion of the ACC, which has been stimuli that reliably generate sensations of discomfort
linked to the generally stronger emotional reactions or pain include, as stated previously, hollow organ
associated with visceral pain (Strigo, I. A. et al., 2003). overdistension, traction on the mesentery, organ
Interestingly, only the intensity and not the pattern ischemia, and organ inflammation, the latter two per-
of activation differs between perceived and unper- haps acting indirectly to alter the chemical milieu at
ceived (subliminal) visceral stimulus intensities an afferent receptive ending to increase neuron excit-
(Sidhu, H. et al., 2004). ability to other stimuli. Accordingly, tension/stretch
(produced by muscle contraction or organ distension,
respectively) is an adequate acute noxious visceral
37.3 Functional Basis of Visceral stimulus at intensities (e.g., balloon pressures) above
Pain threshold for activation of visceral mechano-nocicep-
tors. In experimental studies with humans, verbal
Nociceptors are defined by response to a noxious reports or various scaling procedures identify distend-
stimulus and their ability to encode suprathreshold ing pressures that are uncomfortable or painful.
stimulus intensities. Sherrington C. S. (1906) defined Concomitant recording of contractile phenomena
noxious stimuli as those that damage or threaten allows differentiation between stretch (activation of
damage to skin, and this definition, which has been receptor in parallel with muscle cells) and tension
applied widely to all tissues, is problematic with (activation of receptors in series with muscle cells).
respect to the viscera because tissue-damaging visceral In nonhuman animal experiments, whether the inten-
insult is not reliably pain-producing. Instead, visceral sity of visceral stimulation is noxious has to be inferred
552 Visceral Pain

from autonomic responses, which generally are pro- mechanoreceptors when activated within the physio-
duced at intensities of stimulation below the noxious logical range of intensities generally produce no
threshold, or animal behavior. It is also noteworthy conscious sensation, but rather contribute to auto-
that innervation of the viscera, relative to skin, is nomic regulation of organ function. It has been
sparse and thus spatial summation is important to suggested consequently that these receptors and
activation of visceral mechanoreceptors and the pro- associated axons are not sensory, but rather afferent.
duction of discomfort or pain. Thus, in nonhuman However, low-threshold visceral mechanoreceptors
animals it is necessary to use a distending balloon of possess the uncommon attribute of continuing to
sufficient length to produce a change in behavior that respond, if not also to encode, stimulus intensity
can be interpreted as aversive or painful. In the well into the noxious range. Thus, their activation
absence of behavioral evidence, the quality of mechan- at some (high) intensity of stimulation can lead to
ical stimulation is unknown and often difficult to sensations perceived as uncomfortable or painful.
establish as relevant to activation of mechano-noci- The afferentsensory function of visceral nerves
ceptors and visceral pain. was appreciated by Sherrington C. S. (1900), who
commented that activation of visceral nerves rarely
resulted in conscious sensations (i.e., an afferent func-
37.3.1 Visceral Mechanoreceptors
tion), but when sensation was produced it was
and Mechano-Nociceptors
typically pain (i.e., a sensory function). In all visceral
37.3.1.1 Afferent fiber recordings nerves studied, including gastric vagal afferent fibers,
Obviously, the stomach and temporary storage organs low-threshold mechanosensitive afferents, when
like the urinary bladder and distal colon/rectum tested with high-intensity stimuli, respond vigor-
experience periodic filling (and emptying), which dis- ously and typically at a rate (frequency) greater
tends these organs, commonly without generating than their high-threshold mechano-nociceptor coun-
either discomfort or pain. Mechanosensors in these terparts (Figure 5). Clearly, as stimulus intensity
organs are sensitive to and respond to filling within increases, high-threshold mechano-nociceptors are
the physiological range, eventually leading, for exam- also activated and it can be argued that uncomforta-
ple, to an urge to evacuate the bladder or bowel. ble or painful sensation arises from their activation.
Consistent with these sensations and functions pro- Their contribution is undeniable in acute visceral
duced by organ filling, most mechanosensitive afferent pain, but low-threshold mechanoreceptors (like
fibers contained in the nerves innervating these organs their high-threshold counterparts) also possess the
have low thresholds for response. The proportion of ability to be sensitized (see below and Figure 5), a
low-threshold mechanosensitive endings in spinal quality commonly associated only with nociceptors.
nerves innervating the stomach, esophagus, and gall- Accordingly, it is likely that both low- and high-
bladder (splanchnic nerves) and the urinary bladder, threshold mechanoreceptors in hollow organs contri-
uterus, and distal colon (pelvic nerves) is 7080%, and bute to sensations of discomfort and pain in functional
this has been a generally consistent finding across gastrointestinal (nonulcer dyspepsia, IBS) and
species (cat, chicken, ferret, guinea-pig, mouse, opos- urinary bladder (interstitial cystitis/painful bladder
sum, and rat). The remaining 2030% of afferent syndrome) disorders as well as in ischemia, inflamma-
fibers have high thresholds for response, typically tion, or neoplastic diseases associated with visceral
30 mm Hg distending pressure, which corresponds pain. Consistent with this interpretation, psychophy-
very well to colon distending pressures in rats that sical experiments in human volunteers have
produce behavior leading to avoidance of the stimulus. demonstrated that pharmacologic modulation of sen-
Because high-threshold mechanoreceptors in the vis- sory function similarly shift thresholds for nonpainful
cera respond only to intensities of stimulation in the and painful sensations (Vandenberghe, J. et al., 2005).
noxious range and encode the intensity of suprathres- There are two additional features of low- and high-
hold stimuli, they fulfill requirements as nociceptors threshold mechanosensitive endings in viscera that
and are considered to be mechano-nociceptors that suggest their functions are not always or easily separ-
signal acute visceral pain. able. First, both respond to either or both chemical and
Unlike low-threshold cutaneous mechanorecep- thermal stimuli in addition to mechanical stimulation
tors, which when activated give rise to sensation (the usual experimental search stimulus). That is, they
(e.g., movement of a hair follicle, movement of a are polymodal and virtually all mechanosensitive end-
cotton wisp across the skin), low-threshold visceral ings when tested have been found to respond also to a
 Visceral Pain 553

Total no. indirectly through enteroendocrine cells). Whether

impulses such chemosensitive endings, typically studied in the
50 mucosal surfaces of the gastrointestinal tract, are also
Pre
Post mechano- or thermosensitive has not been system-
40
atically examined, although when examined
mechanosensitivity is commonly noted. If specific
30
and selective visceral chemosensors without mechan-
20 osensitivity exist, which is very likely the case, the
argument that both low- and high-threshold polymo-
Response to organ distension

10 dal mechanosensors can contribute to discomfort and

pain is strengthened. Secondly, low- and high-thresh-
0 old mechanoreceptors cannot be distinguished on the
0 10 20 30 40 50 60 70 80 basis of fiber type (A or C), size or content of cell
Total no. soma, or pharmacologically (e.g., sensitivity to chemi-
impulses cal or inflammatory mediators).
50 Pre
Post 37.3.1.2 Information acquired in vitro
40
In addition to mechanoreceptor characteristics
30 described above, in which afferent fibers were
studied while distending intact hollow organs or
20 occluding vessels, in vitro studies of visceral organs,
typically opened and pinned flat in a dish with nerves
10 attached, provide different information (Figure 6).
For example, receptive endings in the organ can be
0 readily localized by response to applied mechanical
0 10 20 30 40 50 60 70 80 stimuli and the endings can be further isolated
Distension pressure (mm Hg)
experimentally and subjected directly to chemical
Figure 5 Stimulusresponse functions for low-threshold and thermal stimuli. Esophagus, stomach, small
(top) and high-threshold (bottom) mechanosensitive pelvic bowel, urinary bladder, uterus, ureter, testes, and
nerve fibers innervating the urinary bladder of the rat. Before
colonrectum all have been studied in this fashion.
bladder irritation, high-threshold fibers first responded at
30 mm Hg bladder distension and then encoded distending As above, mechanical search stimuli typically have
pressure throughout the range of pressures tested. been used and include brushing/stoking the mucosal
In contrast, low-threshold fibers begin to respond at surface of the organ to locate receptive endings func-
physiologic distending pressures, but also encoded tionally identified as mucosal, punctuate pressure to
distending pressures throughout the ranges of pressures
tested. Note that response magnitude of low-threshold fibers
identify serosal receptors, and circumferential stretch
is greater than for high-threshold fibers at all intensities of of the organ to identify muscular stretch receptors.
bladder distension. Note also that response magnitude Single-fiber studies in all organs examined to date
increases significantly for both low- and high-threshold fibers reveal typical patterns of activation, allowing infer-
after bladder irritation and response threshold for high- ential assignment of mucosal, muscular, and serosal
threshold fibers decreases significantly into the physiologic
receptive endings. Mucosal receptors respond only to
range (i.e., both low- and high-threshold mechanosensitive
fibers sensitize). Reproduced from Su, X., Sengupta, J. N., stroking of the mucosal surface and not to stretch/
and Gebhart, G. F. 1997. Effects of opioids on tension, whereas muscular receptors respond to
mechanosensitive pelvic nerve afferent fibers innervating the stretch/tension and not stroking the mucosal surface.
urinary bladder of the rat. J. Neurophysiol. 77, 15661580, Finally, only intense punctate stimuli activate serosal
used with permission from the American Physiological
receptors. In some organs (e.g., stomach, urinary
Society.
bladder, and colon) receptive endings that respond
to both mucosal stroking and stretch/tension have
chemical and/or to a thermal stimulus. A considerable been found (termed muscularmucosal endings;
literature has established the sensitivity of visceral Figure 6). It should be noted that these descriptors
afferents in both the vagal and spinal innervations to of receptive endings have not been histologically
chemical stimuli (principally hormones and putative confirmed and are based solely on responses to
mediators as well as nutrients that may act directly or mechanical stimuli. Nonetheless, use of in vitro
554 Visceral Pain

Mesenteric that the vagus nerves are involved in chemo-nocicep-

Serosal tion (see below), there is also evidence for a role of vagal
Muscular Mucosal Circumferential afferents in esophageal mechano-nociception (Yu, S.
Mucosal stroking (10 g) stretch (4 g)
et al., 2005). Nonnociceptive esophageal tension recep-
Muscular/mucosal
tors exhibited saturable responses to esophageal
Muscular distension, whereas nociceptive A- and C-fiber eso-
5s phageal vagal afferents discriminated noxious
intensities of esophageal distension (and also responded
10 s
to activators of TRPV1).
Mucosal
Significantly, studies of the two nerves innervat-
ing the same organ have been carried out for both the
mouse colon and mouse urinary bladder. Although
Muscular is has long been appreciated that the two nerves
mucosal innervating an organ execute different functions,
comparison of the properties of the two innervations
provides new information that could, for example,
Lumbar splanchnic inform development of therapeutic strategies. In a
Pelvic nerve
nerve comparison between the lumbar splanchnic nerve
and pelvic nerve innervations of the mouse colon
IMA
(Brierley, S. M. et al., 2004), most of the colonic
receptive endings in the lumbar splanchnic nerve
IMG responded to blunt probing and were located along
LSN the mesenteric attachment and the serosa (Figure 6).
PN In contrast, no pelvic nerve receptive endings were
MPG found in the mesenteric attachment and significantly
more receptive endings responded to stretch than in
the lumbar splanchnic nerve, including muscular
Figure 6 Responses of different types of the mucosal endings (which were not found in the lum-
mechanosensitive endings innervating the mouse colon. bar splanchnic innervation). Analysis of response
Muscular receptors are illustrated topmost as responding to properties of the same receptive endings in the dif-
circumferential stretch of the colon but not stroking of the
ferent innervations revealed further differences.
mucosal surface. Mucosal endings respond to mucosal
stroking, but not to stretch, and musclemucosal fibers Responses to the same intensities of stroking and
respond to both stroking of the mucosa and circumferential stretch stimuli were significantly greater in pelvic
stretch. The application of strokes across the mucosal nerve endings than in lumbar splanchnic counter-
receptive ending and the duration of stretch are illustrated parts. Overall, the pelvic nerve innervation of the
beneath the recordings. The distribution of receptive
mouse colon contained greater proportions of
endings for the lumbar splanchnic nerve and pelvic nerve
innervations of the mouse colon are illustrated bottommost, stretch/tension and mucosal receptors, which more-
showing receptive endings located in the mesentery and over generated greater magnitude responses, than the
serosa as well as those associated with muscle, mucosa lumbar splanchnic innervation. Similarly, compari-
and muscle-mucosa. IMA, inferior mesenteric artery; IMG, son of the lumbar splanchnic nerve and pelvic nerve
inferior mesenteric ganglion; LSN, lumbar splanchnic nerve;
innervations of the mouse urinary bladder reveals
MPG, major pelvic ganglion; PN, pelvic nerve. Adapted from
Brierley, S. M., Jones, R. C. W., III, Gebhart, G. F., and significant differences between the proportions and
Blackshaw, L. A. 2004. Splanchnic and pelvic distribution of types of mechanosensitive receptive
mechanosensory afferents signal different qualities of endings. As found for the colon, the distribution of
colonic stimuli in mice. Gastroenterology 127, 166178. lumbar splanchnic receptive endings in urinary blad-
der was more circumscribed than the distribution of
pelvic nerve receptive endings, which again con-
organnerve preparations has confirmed many of the tained an ending (muscularurothelial) not found in
findings of earlier work and, importantly, has notably the lumbar splanchnic innervation.
expanded our understanding of the contributions of A final means of study of visceral sensory neurons
mechanosensitive endings in viscera to presumed func- is to study cell soma using patch clamp methodology.
tions. For example, in addition to increasing recognition These studies do not, of course, permit the type of
 Visceral Pain 555

functional evaluation derived from single-fiber stu- express different members of the ASIC family of ion
dies, and instead address aspects of mechanisms and channels as well as the vanilloid receptor TRPV1.
changes in visceral neuron excitability, specifically The preferential expression of TRPV1 in small-dia-
the differential roles of voltage- or ligand-gated ion meter, presumably nociceptive neurons raised the
channels. Such studies have been carried out in rat question whether it may play a role in visceral pain.
stomach (vagal and splanchnic innervations), mouse A high percentage (6080%) of mouse and rat colon
intestine, mouse colon, rat colon, and rat urinary sensory neurons express TRPV1 (Robinson, D. R.
bladder (lumbar splanchnic and pelvic innervations). et al., 2004; Christianson, J. A. et al., 2006) and
Because of low innervation density, the study of TRPV1 is the principal detector in mouse colon of
visceral sensory neurons requires the injection of extracellular acidosis (Sugiura, T. et al., 2007).
retrograde labels into muscle layers of the organ of Serotonin acutely alters TRPV1 function in colon
interest. The uptake and retrograde transport of the sensory neurons, shifting the temperature-activation
label into the soma allows identification of neurons curve of this ion channel to normal body tempera-
projecting to the organ of interest. Consistent with tures (Sugiura, T. et al., 2004). About 90% of the
results obtained in vivo, isolated visceral sensory bodys serotonin is found within the gastrointestinal
neurons do not generate spontaneous activity. tract, primarily in enterochromaffin cells, and seroto-
However, cells obtained from animals with experi- nin is released in high concentrations during mucosa
mentally induced inflammation show significant stimulation. Studies of patients with functional bowel
oscillation of the resting membrane potential, result- diseases, such as IBS, suggest changes in serotonin
ing in spontaneous action potential discharge, release and/or reuptake within the intestinal mucosa,
demonstrating an increase in excitability. This raising the question whether modulation of TRPV1
increase in excitability is associated with an increase function may contribute to the enhanced responses to
in voltage-sensitive sodium currents, primarily a physiologic stimuli seen in these patients. In addition
tetrodotoxin (TTX)-resistant sodium current, and a to modulation by G-protein-coupled receptors, changes
decrease in voltage-sensitive potassium currents, pri- in TRPV1 expression have been examined under
marily the transient or A-type current (Yoshimura, experimental and clinical conditions. Animal models
N. and deGroat, W. C., 1999; Stewart, T. M. et al., of visceral inflammation as well as distinct human
2003; Dang, K. et al., 2004; Beyak, M. J. and Vanner, S., disorders from gastroesophageal reflux disease, painful
2005; Xu, G. et al., 2006). bladder syndrome to rectal urgency are associated with
In addition to voltage-sensitive ion channels, the increases in TRPV1 expression (e.g., Chan, C. L. H.
expression and properties of ligand-gated channels et al., 2003; Bhat, Y. and Bielefeldt, K., 2006; Mukerji, G.
also changes in animal models of visceral pain. et al., 2006).
Ischemia is the most common cause of cardiac pain, Purines, which include ATP and its breakdown
experienced by millions of Americans suffering from products, are released during cell injury and may
coronary artery disease. Tissue acidosis due to accu- thus play a role in nociception. Among the families
mulation of lactate can activate cardiac afferents and of purinergic receptors, the ligand-gated P2X recep-
likely triggers ischemic pain. Perhaps consistent with tors have attracted special attention because of their
the importance of ischemia as a trigger of visceral role in visceral sensation. As described Chapter
pain, a comparison between cardiac sensory neurons Urothelium as a Pain Organ, the bladder epithelium
and randomly chosen neurons in the same DRG (urothelium) releases ATP during physiologic stimu-
showed that a higher fraction of visceral sensory lation, such as stretch. Bladder sensory neurons
neurons express ASICs (Benson, C. J. et al., 1999). express P2X receptors and respond to purinergic
Experimental injury, such as gastric ulceration, agonists (Yoshimura, N. et al., 2003; Dang, K. et al.,
increases the current density of these acid-sensitive 2005). Cystitis significantly increases the peak cur-
currents in visceral sensory neurons (Sugiura, T. rent triggered by purinergic agonists, consistent with
et al., 2005), demonstrating that sensitization does a potential role of this signaling pathway in causing
not only affect excitability (i.e., the generation of discomfort and increased micturition frequency,
action potentials in response to a given stimulus), associated with bladder inflammation. Similarly,
but also the signal transduction process (i.e., the colon distension in the rat releases ATP in a stimu-
depolarization triggered by a mechanical, chemical, lus-dependent manner, and the release of ATP is
or thermal stimulus). Based on the properties of pro- increased further after experimental induction of
ton-evoked currents, most visceral sensory neurons colon inflammation (Wynn, G. et al., 2003; 2004).
556 Visceral Pain

Responses to colon distension in the colitis model biophysical studies have not demonstrated a direct
were attenuated by a purinergic receptor antagonist, activation of TRPV1 by stretch. Thus, the channel
suggesting a role for ATP in colon mechanosensory likely alters cellular excitability and enhances
transduction. responses to different stimulus modalities, including
stretch.
37.3.1.3 Gene deletions, visceral pain, As discussed separately in more detail (Chapter
and mechanosensation Urothelium as a Pain Organ), purinergic receptors
The availability of mice with selected gene deletions play an important role in bladder function. Bladder
(knockout mice) has provided the opportunity to filling triggers ATP release from urothelial cells,
examine the potential role of several molecules in which activates bladder sensory neurons and may
visceral pain. To date, voltage-gated sodium channel trigger reflex micturition. Consistent with this
(NaV1.8), TRPV1, purinergic receptor (P2X), and model, P2X3 knockout mice exhibit bladder hypore-
ASIC knockout mice have been studied and each flexia (Cockayne, D. A. et al., 2000).
molecule has been found to contribute at least in A more complex picture emerges from studies
part to responses to noxious visceral stimuli or of examining the role of different members of the
visceral mechanoreceptors. As mentioned above, the ASIC family in visceral sensation. Genetic deletion
TTX-resistant sodium current is increased in neu- of ASIC3 blunts responses to colorectal distension
rons obtained from animals with gastrointestinal or and mechanical stimulation of gastroesophageal
bladder inflammation. In sensory neurons, TTX- afferents in mice, suggesting a role in visceral
resistant current is largely due to activation of the mechanosensation (Jones, R. C. W. et al., 2005;
voltage-gated sodium channel NaV1.8. Genetic dele- Page, A. J. et al., 2005). In contrast, deletion of ASIC1a
tion of this ion channel in mice is associated with a increased mechanosensitivity of gastroesophageal
decrease in aversive behavior in response to intraper- and colonic afferents without affecting cutaneous
itoneal injection of acetic acid (Laird, J. M. et al., afferents and deletion of ASIC2 blunted responses
2002), supporting a potential role of NaV1.8 in visc- of gastroesophageal, but enhanced those of mesen-
eral pain. The pain behavior in this model is also teric afferents in the distal colon (Page, A. J. et al.,
likely contributed to by activation of nonvisceral 2005). ASICs form heteromultimeric channels, lead-
afferents within the parietal peritoneum. More direct ing to speculation that the subunit composition has
evidence was provided by other studies, one employ- significant effects on the mechanosensitive properties
ing transient suppression of TTX-resistant sodium of visceral sensory neurons (Wemmie, J. et al., 2006).
currents in bladder sensory neurons with antisense Thus, genetic studies showed that as is true for
oligonucleotides, which abolished the increase in cutaneous pain no single molecule explains the
micturition frequency seen after acute bladder irrita- complex phenomenon of pain. Yet, some differences
tion (Yoshimura, N. et al., 2001), and the other between cutaneous and visceral sensation has
employing NaV1.8 and NaV1.9 knockout mice, emerged, most notably the potential role of TRPV1
which reported that neuron hyperexcitability pro- and P2X receptors in mechanosensation.
duced by jejunitis was absent in NaV1.8, but not The foregoing studies focused on the initial events
NaV1.9 knockout mice (Hillsley, K. et al., 2006). triggered by noxious visceral stimulation, namely
Genetic deletion of functional TRPV1 receptors signal transduction and cellular excitability in pri-
also leads to changes in visceral function, suggesting a mary afferent neurons. Consistent with a role of
role of TRPV1 in visceral mechanosensation. substance P and neurokinin receptors in synaptic
TRPV1 knockout mice have a higher frequency of transmission of visceral afferent input within the
nonvoiding bladder contractions (Birder, A. L. et al., spinal cord, genetic deletion of neurokinin receptor
2001). Nerve recordings from mesenteric nerve bun- 1 blunted responses to intraperitoneal injection of
dles innervating the small bowel and colonic afferents acetic acid in mice (Laird, J. M. A. et al., 2000).
within the pelvic nerve revealed blunted mechano- Despite these findings and preclinical studies with
sensation in TRPV1 knockout mice compared with different neurokinin receptor blockers in rodent
wild-type controls (Rong, W. et al., 2004; Jones, R. C. W. models of visceral pain, neurokinin antagonists do
et al., 2005). Deletion of TRPV1 also altered not appear to be effective when used to treat visceral
responses to inflammatory mediators, consistent pain in humans.
with a potential role of this ion channel in peripheral The genetic basis for visceral pain has also been
sensitization (Jones, R. C. W. et al., 2005). However, addressed in humans. Considering the high
 Visceral Pain 557

prevalence of functional gastrointestinal disorders in stimulation, but not by the usual high-intensity
the general population and the expression of pain and mechanical stimuli tested. During the minutes and
discomfort as the primary symptoms, most studies hours after experimental inflammation of the knee
have focused on IBS or nonulcer dyspepsia. As men- joint, however, these nociceptors became progres-
tioned above, serotonin plays an important role in sively sensitive to mechanical stimuli, including to
afferent signaling within the gastrointestinal tract. nonnoxious movement of the joint, and developed
Mechanical or chemical stimuli can trigger serotonin spontaneous activity. It has subsequently been
release from enteroendocrine cells into the lamina established that sleeping nociceptors do respond
propria, where high tissue concentrations activate to mechanical stimulation in the absence of inflam-
terminals of both intrinsic and extrinsic primary mation, but only at intensities that are potentially
afferent neurons within the vicinity. Reuptake of damaging to tissue, and hence they are not truly silent.
serotonin by the high-affinity transporter serotonin Microneurographic studies suggest that between 15%
transporter (SERT) expressed by epithelial cells ter- and 20% of C-fiber nociceptors in human skin are of
minates this signal. This led to the hypothesis that the sleeping variety (Torebjork, H. E. et al., 1996), and
differences in the properties of SERT may contribute sleeping nociceptors also have been documented in the
to the pathogenesis of functional gastrointestinal dis- visceral innervation (Figure 7), although estimates of
orders. Consistent with this assumption, deletion of their number vary widely from 30% to >80%.
SERT in mice triggers changes in bowel habits with Studies of sleeping visceral nociceptors have focused
alternating diarrhea and constipation, a common principally on the pelvic nerve innervation of the rat or
clinical manifestation in patients with IBS cat urinary bladder and colon. Whether the afferents
(Gershon, M. D. and Tack, J., 2007). Two allelic studied were actually sleeping is uncertain, and it has
variants of the serotonin transporter have been iden- to be appreciated that assignment to this category is
tified in humans. These alleles have different kinetics typically based on the absence of a response to organ
of serotonin uptake, leading to the hypothesis that distension in the noxious range. Given the filling,
they may contribute to the pathogenesis of functional storage, and emptying functions of the bladder and
bowel disorders. While initially supported, several colon, which are supported by the high proportion
studies have failed to show a linkage between one (70%) of pelvic nerve afferents with low thresholds
of the alleles and IBS (Camilleri, M., 2004). for mechanical activation, that a similar large
Holtmann G. et al. (2004) examined the link between
a G-protein  subunit and functional dyspepsia. Prior
10 Hz]
studies had shown that the expression of the two Preinflammation
allelic forms of this molecule affected intracellular
signaling in neurons and was linked to affective dis-
30 min after inflammation
orders. As was true for SERT, a subsequent study did
not confirm the linkage reported in this initial obser-
vation (Andresen, V. et al., 2006). These conflicting 1 h after inflammation
results likely reflect the fact that a complex trait, such
as enhanced sensitivity to visceral stimulation, can
typically not be explained by a single gene defect, but 5 10 20 30 40 60 80 100
rather a complex interaction between multiple Pressure (mm Hg)
genetic and environmental factors. Thus, contribu- Figure 7 Illustration of a mechanically insensitive, silent
tions of any single candidate gene to the variance of (sleeping) visceral nociceptor. Responses to balloon
phenotypic expression will be limited, requiring very distension of the colon (5100 mm Hg, 20 s) are shown
large studies to more conclusively address their role before experimental organ inflammation (the fiber did not
respond to colon distension or bladder distension, not
in the development of visceral pain syndromes. shown), 30 and 60 min after intracolonic instillation of
zymosan. Spontaneous activity is apparent after colon
inflammation as are responses to colon distension. Adapted
37.3.2 Sleeping (Silent) Nociceptors from Coutinho, S., Su, X., Sengupta, J. N., and Gebhart, G.
F. 2000. Role of Sensitized Pelvic Nerve Afferents from the
In the mid-1980s Schmidt and coworkers (Schmidt,
Inflamed Rat Colon in The Maintenance of Visceral
R. F., 1996) characterized a novel nociceptor in the Hyperalgesia. In: Nervous System Plasticity and Chronic
knee joint of the cat they dubbed silent or sleeping. Pain, Progress in Brain Research (eds. J. Sandkuher, B.
These nociceptors could be activated by electrical Bromm, and G. F. Gebhart), Vol. 129, pp. 375387. Elsevier.
558 Visceral Pain

proportion of pelvic nerve afferents are of the sleeping interaction between different stimulus modalities
variety seems disproportionate. It is clear, however, (Kang, Y. et al., 2004) (Figure 8).
that the proportion of sleeping nociceptors in the visc- Coronary artery occlusion in animals or myocar-
eral innervation is greater than has been reported in dial ischemia in humans leads to the release and
skin, but the proportion and contributions of sleeping accumulation of a variety of substances in sinus
nociceptors to visceral pain states remains to be estab- blood (e.g., bradykinin, adenosine, serotonin, prosta-
lished. Some of these mechanically insensitive afferents glandins; see Meller, S. T. and Gebhart, G. F., 1992
may have been chemosensitive and would have for a review) that have been shown to activate cardiac
responded to a chemical stimulus not tested. A poten- afferent fibers in the spinal innervation. Experimental
tial role of chemical signaling is supported by coronary artery occlusion also triggers a rapid
recruitment of silent vagal gastric afferents after expo- decrease in myocardial pH, and both protons and
sure to bile acids (Page, A. J. et al., 2002). lactic acid (which accumulates during tissue hypoxia)
have been shown to activate ischemia-sensitive
cardiac afferent fibers in the spinal splanchnic inner-
37.3.3 Visceral Chemo-Nociception
vation (Pan, H. L. et al., 1999). Many ischemia-
Most studies of visceral afferents focus on mechano- sensitive cardiac (Pan, H. and Chen, S. R., 2002)
sensation, as mechanical stimuli can trigger and abdominal splanchnic afferents are also respon-
reproducible responses without inducing tissue sive to bradykinin, serotonin, and prostaglandins
damage. However, visceral afferents are also exposed (see Longhurst, J. C., 1995 for review) as well as
to potentially noxious chemical signals, such as acidity mechanical probing. As in other organs, myocardial
in the proximal gastrointestinal tract, and products ischemia has been reported to recruit mechanically
that may accumulate during ischemia, such as lactate, insensitive (sleeping) cardiac splanchnic afferent
protons, and bradykinin, and thus contribute to visc- fibers (Pan, H. and Chen, S. R., 2002).
eral discomfort and pain. Gastro-esophageal reflux In healthy subjects, the afferent innervation of the
disease (GERD, gastric acid reflux into the lower lower airways does not normally contribute to con-
esophagus), myocardial ischemia, and lower airway scious sensations. However, dyspnea, chronic cough,
irritation are all associated with discomfort and/or and chronic obstructive pulmonary disease, in addi-
pain. Although experimental evidence obtained in tion to inhalation of irritant chemicals, are associated
animal studies is limited, both spinal and vagal affer- with discomfort and/or pain, and it appears that the
ents are involved in chemo-nociception. vagal afferent innervation plays a dominant role. As
Using the expression of the immediate early gene indicated previously, the cell bodies of vagal sensory
c-fos as a surrogate marker for neuronal activation fibers are contained in the nodose and smaller jugular
after acid exposure of the stomach, noxious chemical ganglia, which are derived from placode and neural
stimuli activated vagal but not spinal pathways crest cells, respectively. Undem, B. J. and coworkers
(Schuligoi, R. et al., 1998). This is consistent with have provided evidence that the vagal ganglionic
behavioral responses to intragastric acid administra- source of axons supplying the airways is associated
tion, which were blunted after vagotomy (Lamb, K. with the neurochemical and physiological phenotype
et al., 2003). In studies of mouse vagal gastro-esopha- of the afferent fiber (Riccio, M. M. et al., 1996).
geal afferent fibers, more than 50% of fibers that Nodose ganglion A-fibers have low thresholds for
responded to circular tension also responded to bile mechanical activation, adapt rapidly to mechanical
acid (a component of gastric acid reflux into the stimulation, and are not activated by capsaicin. In
esophagus); significantly, a population of mechani- contrast, jugular ganglion A-fibers have higher
cally insensitive (sleeping) afferents was activated by thresholds for mechanical activation, adapt slowly
bile acid, suggesting the presence of specific chemo- to mechanical stimulation, and are typically respon-
sensors (which did to respond to other chemicals sive to capsaicin. C-fibers associated with somata in
studied) (Page, A. J. et al., 2002). Additional support both the nodose and jugular ganglia are also
for vagal involvement in chemo-nociception is pro- mechanically sensitive and also respond to capsaicin
vided by results showing that independent of the and bradykinin (Undem, B. J. et al., 2004); in addition,
acute response to protons (glycocholic acid and C-fibers innervating the lung from the nodose gang-
hydrochloric acid), acid exposure enhanced subse- lion express P2X receptors and respond to purinergic
quent responses of gastric vagal afferents to agonists (jugular C-fibers do not). Because propor-
mechanical stimulation, suggesting a sensitizing tions of these mechanically sensitive nodose and
 Visceral Pain 559

Baseline Distension

20 mV
pH 7

1s

pH 4

2.5
Spike frequency (Hz)

Baseline cm H2O
2.0 pH 4
30
1.5
1.0
0.5 0
0.0
0 10 20 30 40
Pressure (cm H2O)

Figure 8 Intracellular recordings from a nodose ganglion neuron innervating the mouse stomach show low baseline activity
(top left) and an increase (response) in activity (top right) during luminal distension (030 cm H2O, 10 s, shown bottom right).
When the luminal proton concentration was raised to pH 4, baseline activity and the response to distension increased. The
stimulus response function for this neuron is summarized on the bottom left and demonstrates a significant shift to the left in
the presence of an increase in luminal pH to 4. From Bielefeldt K. (unpublished).

jugular ganglia afferents are sensitive to acidic solu- normally unattended afferent input from the viscera
tions and other algogenic chemical stimuli, they are to the level of consciousness in the form of discomfort
considered to function as airway and lung nocicep- and pain. By far, however, most persistent visceral
tors. In support, airway inflammation increases the pain conditions are characterized by hypersensitivity
mechanical sensitivity of the rapidly adapting popu- in the absence of inflammation or other cause.
lation of vagal afferents and triggers an increase in Accordingly, visceral hypersensitivity has been sug-
the synthesis of neuropeptides substance P and gested to represent a biomarker for functional
CGRP (Undem, B. J. et al., 2002). gastrointestinal and bladder disorders, which in the
case of IBS can be experimentally established by
balloon distension of the rectum. These functional
37.4 Visceral Hypersensitivity disorders are relatively common, persistent, and gen-
erally inexplicable clinically. The characteristic
Like other tissues in the body, an increase in the discomfort and pain is often precipitated by ingestion
excitability of sensory neurons innervating an organ of food, but also can be disassociated from events that
can lead to exaggerated responses to applied stimuli, logically would be expected to exacerbate symptoms.
including intensities of stimulation in the physiologic Typically, nonnoxious intensities of visceral stimula-
range that normally do not lead to conscious tion (e.g., ingestion of food or beverage, low pressures
sensation. Inflammation of visceral tissues, like of gastric or rectal distension, bladder filling) trigger
inflammation of skin, muscle, or joints, typically painful sensations, which some have inappropriately
increases visceral neuron excitability and raises and inaccurately termed visceral allodynia.
560 Visceral Pain

There is an association (30%) between IBS and gastric vagal mechanoreceptors, which are generally
previous (and resolved) gastrointestinal infection, considered not to contribute to visceral pain, do
but antecedent or contemporaneous events that sensitize and significantly increase input into the
lead to development of functional disorders are central nervous system (nucleus of the solitary
otherwise unknown. Abuse in childhood, including tract) which must have consequences, if not to
sexual abuse, has been advanced as a contributing interpretation or awareness of gastric sensation,
factor in development of functional disorders as then at least (or in addition) to efferent outflow to
have stress and anxiety, which are commonly the stomach via the dorsal vagal motor nucleus.
reported by patients and can clearly exacerbate Second, that innocuous gastric stimuli can produce
symptoms. Given that the cause of such disorders a time-limited sensitization of responses of gastric
is unknown, there has been considerable attention vagal mechanoreceptors reveals considerable neural
paid to peripheral versus central nervous system plasticity, suggesting that normal visceral events
contributions to initiation and maintenance of dis- can modulate visceral sensory neuron excitability
comfort and pain in functional disorders. Both even in healthy persons.
visceral sensory neuron excitability and sensitization Because sensitization reveals an underlying change
and central nervous system dysregulation are con- in neuron excitability, changes in neuron ion channels
sidered below. must be responsible for increases in response magni-
tude and decreases in response threshold, at least
37.4.1 Sensitization and Excitability during the initial period of sensitization (see above
of Visceral Nociceptors and Gold, M. S., 2006). Experimental models of visc-
eral hypersensitivity are associated with an increase in
Sensitization is a defining characteristic of nocicep- neuron input resistance and decrease in resting mem-
tors first described by Perl for cutaneous nociceptors brane potential. While the exact mechanisms have not
(see Perl, E., 1996 for an overview) and which has
been studied in visceral sensory neurons, changes in
since been established for nociceptors innervating
the expression and/or function of two pore domain
other tissues. Sensitization is an increase in response
potassium channels and hyperpolarization-activated
magnitude to suprathreshold stimuli and/or a
IH likely also contribute. In addition, both voltage-
decrease in response threshold; not uncommonly,
gated sodium (NaV) and voltage-gated potassium
both response magnitude increases and response
(KV) channels have been documented to change in
threshold decreases (and spontaneous activity can
visceral sensory neurons as predicted based on under-
develop) (see Figure 5 for examples of sensitization
standing of general biophysical principles. In
of responses of low- and high-threshold mechano-
sensitive pelvic nerve afferent fibers to bladder experimental models of gastric ulceration, gastritis,
distension). Experimentally, sensitization is com- ileitis, colitis, and cystitis, peak inward sodium cur-
monly produced following tissue inflammation or rents significantly increase after visceral insult, and the
insult, and an extensive literature has identified med- increase is largely contributed to by the TTX-resis-
iators and mechanisms of inflammation-produced tant component of the current (e.g., NaV1.8), although
sensitization, principally in nonvisceral tissues (e.g., precisely which sodium channel has not been estab-
McMahon, S. B. et al., 2006). Clinically, in functional lished. In addition to changes in threshold, the
disorders, sensitization of visceral sensory neurons is sensitization is characterized by an increase in action
present in the absence of gross inflammation or tissue potentially frequency during stimulation. Again vol-
insult, but visceral sensory neurons can sensitize after tage-sensitive sodium channels play an important role
exposure to noninjurious stimuli. For example, gas- in this context. As already mentioned, current density
tric vagal mechanosensitive afferents uniformly have increases, the voltage dependence of channel activa-
low thresholds for response to balloon distension of tion shifts and recovery from inactivation is
the stomach, but also encode stimulus intensity into accelerated, and all of these factors clearly facilitate
the noxious range. Response magnitude to gastric repetitive firing. Conversely, the outward potassium
distension can be affected not only by intragastric current (the A type current) is reduced and the voltage
instillation of acids or by gastric ulceration, but also dependence of activation shifts to more positive
by transient intragastric exposure to stimuli that do potentials in visceral sensory neurons after visceral
not injure the tissue (e.g., see Kang, Y. et al., 2004). insult, which is also consistent with an increase in
The significance of these findings is twofold. First, neuron excitability.
 Visceral Pain 561

37.4.2 Central Sensitization insult causes an increase in the indicator studied as

well as an anatomical expansion of indicator beyond
In addition to increased responses and decreased
the normal, resting range of visceral input to the
response thresholds, sensitization of visceral nocicep-
spinal cord. Figure 9 illustrates this point for the
tors importantly leads to changes in excitability of
central nervous system neurons upon which they
terminate, a consequence termed central sen-
Control
sitization. Nociceptor sensitization and central 100
Inflamed
sensitization are considered to underlie, respectively,
the expression of primary and secondary hypersensi- 75

% internalized
sP receptors
tivity. The most readily apparent reflection of central
sensitization in the clinical setting is expansion of the 50
area of referred sensation, which is accompanied by
tenderness to palpation (a reflection of peripheral 25
sensitization). The spinal cord has been the principal
focus of study with respect to central sensitization, 0
but neurons in supraspinal sites also undergo changes T12L1 L25 L6S1
in excitability and, as discussed below, supraspinal
sites can contribute significantly to the maintenance Inflamed
of central sensitization and may play a critical role in
the maintenance of chronic pain disorders, including
functional visceral disorders.
The sensitization of central neurons, which has
been studied extensively in the spinal cord, results
Control
from increased release of neuroactive substances (e.g.,
glutamate, neurokinins like substance P) from visceral
sensory nerve terminals in the spinal cord and from
second-order spinal interneurons upon which they Lumbar Pelvic nerve
may terminate. Because supraspinal sites are also splanchnic
nerve MPG
engaged by persistent nociceptive input, another IMG
source of neuroactive substances (e.g., serotonin, nor-
epinephrine, and GABA) is derived from projections to
the spinal cord from the brainstem. Finally, non-
neuronal glia become activated in persistent pain Colon
conditions, which has been widely documented in
Figure 9 Internalization of substance P (sP) receptors by
nonhuman animal somatic inflammatory models, and colon distension in the absence (control) and 3 h following
can contribute additional neuroactive substances (e.g., intracolonic instillation of zymosan (inflamed). In the
cytokines, chemokines) and modulators to the complex absence of colon insult (control), noxious colon distension
chemical milieu in spinal cord. Contributions from the (80 mm Hg, 20 s) leads to internalization of the sP receptor in
superficial spinal laminae in an anatomically appropriate
latter sources (supraspinal sites and glia) likely become
distribution (i.e., T12L1 and L6S1 spinal segments via the
more important over time, and their influence may lumbar splanchnic nerve and pelvic nerve innervations of
persist and contribute to altered sensations from the the rat colon there is no receptor internalization in lumbar
viscera in the absence of sustained visceral nociceptor spinal segments 25). Three hours after intracolonic
sensitization (see below). instillation of an inflamogen, zymosan (inflamed), a time at
which responses to colon distension are significantly
Changes within the spinal cord and brain stem increased (i.e., colon hypersensitivity is present
areas of visceral input following experimental organ behaviorally), the amount of sP receptor internalized
inflammation or noxious stimulation have been (quantified at top) is significantly increased and now
assessed by following expression of the immediate includes L25. IMG, inferior mesenteric ganglia; MPG, major
early gene, c-fos, upregulation of the neuronal iso- pelvic ganglion. Adapted from Honore , P., Kamp, E. H.,
Rogers, S. D., Gebhart, G. F., and Mantyh, P. W. 2002.
form of nitric oxide synthase, and internalization of Activation of lamina I spinal cord neurons that express the
the receptor for substance P, the neurokinin 1 recep- substance p receptor in visceral nociception and
tor. These experimental approaches show that organ hyperalgesia. J. Pain 3, 311.
562 Visceral Pain

spinal cord, showing that internalization of the sub- (Pezzone, M. A. et al., 2005) and, conversely, that
stance P receptor produced by colon distension cystitis produces colon hypersensitivity (Lamb, K.
increases significantly after colon insult and, more- et al., 2006; Winnard, K. P. et al., 2006).
over, now includes an area of the spinal cord that Animal and human studies have clearly estab-
showed no receptor internalization after colon dis- lished the contribution of supraspinal sites to
tension in the absence of insult (Honore, P. et al., sensitization to visceral input. In susceptible rodent
2002). This representation of central sensitization strains, stress enhances behavioral responses to visc-
occurred within hours after colonic insult, revealing eral distension (Coutinho, S. V. et al., 2002; Bradesi, S.
that mechanosensitive colon afferents sensitize, et al., 2005). While multiple mechanisms may contri-
sleeping afferents awaken, and the visceral innerva- bute to this modulation of visceral input, descending
tion and second-order neurons upon which the inhibitory or facilitatory pathways likely play a role
afferents terminate are rapidly and easily modified (see below), as shown by the effects of electrical
(i.e., are highly plastic). Other studies in rats show stimulation within the rostral ventromedial medulla
rapid changes in supraspinal sites as well, suggesting on responses to noxious colorectal distension (Zhuo, M.
their involvement in states of increased central excit- and Gebhart, G. F., 2002) and of selective abolition
ability (e.g., Coutinho, S. et al., 1998). Central of descending facilitatory influences in a model of
sensitization also affects somatic and visceral inputs rodent pancreatitis (Vera-Portocarrero, L. P. et al.,
that converge on spinal neurons, as shown by 2007). These reports are consistent with data
reduced thresholds for hindpaw withdrawal after obtained in human volunteers. Activation of des-
bladder or colon insult (Jaggar, S. I. et al., 1999; cending inhibitory mechanisms through heterotopic
Miranda, A. et al., 2004; Lamb, K. et al., 2006). Such stimulation blunted perception of rectal distension in
data obtained in animals are consistent with results healthy volunteers (Song, G. H. et al., 2006).
from human studies. For example, repetitive color-
Conversely, uncomfortable visceral input triggered
ectal distension increases pain perception and the
by acid infusion into the duodenum lowered the pain
area of cutaneous pain referral in normals (Ness, T. J.
threshold to proximal esophageal stimulation
et al., 1990) as well as patients with functional bowel
(Hobson, A. R. et al., 2004). These experiments also
disease (Munakata, J. et al., 1997). Similarly, infusion
support a rapid involvement (minutes to hours) of
of 0.15 M hydrochloric acid into the distal esophagus
supraspinal sites in central sensitization. Longer-term
not only reduces stimulus threshold in the distal
evidence of supraspinal contributions to central sen-
esophagus, but also in the nonacid-exposed esopha-
sitization is provided by both clinical reports of
gus as well as on the chest wall area of referral within
an hour after acid exposure (Sarkar, S. et al., 2000; esophageal or gastric hypersensitivity in patients
2001), again revealing that central sensitization pro- with IBS (or somatic disorders like fibromyalgia)
duced by a visceral input is also apparent in the area (Price, D. D. et al., 2007a; 2007b; Sharma, A. and
of somatic referral. Aziz, Q., 2007) and by various manipulations limited
Another expression of central sensitization, which to the neonatal period that lead either to visceral
can reflect either spinal or supraspinal involvement, hypersensitivity or increased susceptibility to organ
is sensitization that develops between organs with insult in adult animals. In rats, neonatal colon irrita-
inputs onto spinal dorsal horn neurons within the tion (intracolonic mustard oil) or colon distension
same spinal segments, an expression of viscero-visc- (Al Chaer, E. D. et al., 2000), urinary bladder inflam-
eral convergence (e.g., urinary bladder and colon), or mation (intravesical zymosan, Randich, A. et al.,
between organs that do not share patterns of spinal 2006), maternal separation (a stressor, Coutinho, S.
segmental overlap. The former are easy to under- V. et al., 2002), or neonatal immune challenge with
stand anatomically, and while not rigorously studied the bacterial endotoxin lipopolysaccharide (Spencer,
in the human population, patients with functional S. J. et al., 2007) are all associated with increased
bowel disorders often also complain about pelvic organ sensitivity in the adult animal. Because these
pain or symptoms consistent with interstitial cystitis experimental neonatal manipulations are not asso-
(Whitehead, W. et al., 2002). Conversely, many ciated with adult organ inflammation, the adult
patients with interstitial cystitis also suffer from func- visceral hypersensitivity in these models is consid-
tional bowel disorders (Alagiri, M. et al., 1997). In ered to be maintained by central nervous system
nonhuman animals, it has been established that mechanisms, including dysregulation or reorganiza-
colon inflammation triggers bladder hyperactivity tion (discussed in Section 37.4.3).
 Visceral Pain 563

Mechanistically, both human and nonhuman ani- Normally, descending modulation represents a bal-
mal studies have established an important role for ance between inhibitory and facilitatory influences,
peripheral, spinal (and supraspinal) N-methyl-D- with inhibition typically dominant. Facilitatory influ-
aspartate (NMDA) receptors in central sensitization ences, however, have been shown to predominate
following visceral insult (e.g., McRoberts, J. A. et al., in some experimental conditions and have been
2001; Willert, R. P. et al., 2004; see Woolf, C. J. and hypothesized to contribute to maintenance of persis-
Thompson, S. W., 1991 for a general overview), tent pain states (Porreca, F. et al., 2002; Gebhart, G. F.,
although the relevant spinal neurons and signal trans- 2004). With respect to functional visceral disorders
duction pathways in visceral sensory pathways are that persist in the absence of a defined pathobiology,
not revealed by these findings. The process of central and many of which are comorbidly expressed with
sensitization is linked to long-term potentiation, other generalized disorders (e.g., fibromyalgia), the
which has been extensively studied in the hippocam- concept of a dysregulated modulatory system has
pus in relation to learning and memory, and in spinal gained both currency and experimental support.
cord exhibits similar characteristics that involve tran- Most nonhuman animal studies have examined
scriptional and other events that lead to changes in nonvisceral pain, but several studies have confirmed
excitability that can outlast the peripheral input. for visceral inputs what has been more widely docu-
Although not focused on visceral inputs, these studies mented for nonvisceral inputs. Activation of
are based on features common to visceral nociceptive brainstem sites (PAG and rostral ventromedial
input: recordings from lamina I spinal dorsal horn medulla) has been shown to facilitate visceromotor
neurons that project to the pontine parabrachial area reflexes (Zhuo, M. and Gebhart, G. F., 2002) and
and coactivation of substance P and NMDA recep- spinal dorsal horn neuron responses (Zhuo, M. et al.,
tors (Ikeda, H. et al., 2003; 2006). Study of this 2002) to noxious colon distension in the rat.
population of spinal neurons, upon which visceral Similarly, an experimental pancreatitis in rats has
afferents likely terminate (along with nonvisceral been shown to be maintained by facilitatory influ-
nociceptive afferents), reveals synaptic plasticity ences descending from the rostral ventromedial
dependent upon neuroactive substances contained medulla (Vera-Portocarrero, L. P. et al., 2007).
and released from visceral afferent terminals in the These studies confirm that modulation of spinal
spinal cord and a cellular process that likely contri- nociceptive visceral transmission does not differ in
butes to induction of central sensitization. character from modulation of nonvisceral inputs.
Importantly, studies in humans suggest that facilita-
tory modulatory influences can contribute to
37.4.3 Central Modulation of Visceral Pain
persistent pain states, including visceral pain states.
One consequence of peripheral events that lead to Such studies rely on interpretation of changes in
central sensitization is activation of endogenous pain patterns or intensity of brain imaging or on localiza-
modulatory systems and there is growing apprecia- tion, wave form, and magnitude of event-related
tion that visceral disorders characterized by cortical potentials.
persistent discomfort and pain reveal, at least in For example, Dimcevski G. et al. (2007) recently
part, a dysregulated central nervous system. The reported functional brain reorganization in chronic
concept that there exist endogenous mechanisms pancreatitis patients. They electrically stimulated at
that modulate pain arose from studies showing that different sites from above the gastro-esophageal junc-
electrical stimulation or chemical activation (e.g., tion to the horizontal part of the duodenum and
glutamate to activate cells or drugs like opioids act- recorded event-related brain potentials from surface
ing at their cognate receptors) in the brainstem PAG electrodes on the scalp and also assessed sensation.
produced powerful descending inhibitory effects on They found that neuronal sources in the insula were
spinal nociceptive inputs. The initial focus of such most consistently changed in chronic pancreatitis
studies was on inhibitory inputs descending from the patients relative to healthy controls and concluded
brainstem and on modulation of noxious inputs. that reorganization in this part of the brain is a con-
Presently, it is appreciated that descending influ- tributing mechanism to pain in chronic pancreatitis.
ences require a relay in the rostral ventromedial This and other reports (e.g., Price, D. D. et al., 2007a;
medulla between the PAG and spinal cord, are not 2007b) clearly reveal that central mechanisms are
restricted to modulation of only noxious inputs, and important to the maintenance of persistent visceral
importantly not limited to only inhibitory influences. pain conditions. Mechanisms by which the
564 Visceral Pain

excitability of neurons in these or other brain areas influence organ function, as well as paravertebral
might increase or decrease and be sustained are ganglia. Vagal afferent fibers terminate centrally
unknown. Stress and other external factors, as well in the brainstem nucleus of the solitary tract.
as cognitive and emotional events, can contribute to (4) Cell bodies of the vagus nerves are contained in
activity in supraspinal sites and influence the inter- the nodose ganglia (primarily) and cell bodies of
pretation of information arriving there from internal all other visceral nerves are contained in DRG.
organs. The matrix of brain areas activated by painful Generally, visceral sensory neuron cell bodies
visceral stimuli is similar to that seen during antici- are larger in size than would be expected given
pation of such a painful stimulus, demonstrating the that virtually all visceral sensory neurons have
importance of cognitive influences (Yaguez, L. et al., thinly myelinated or unmyelinated axons, which
2005). Stress activates the hypothalamicpituitary in the somatic realm are associated with small-
adrenal axis, resulting in the release of various neu- diameter cell bodies.
roendocrine mediators. Studies in humans have (5) Spinal visceral nerves terminate in laminae I, II,
demonstrated different neuroendocrine responses in V, and X, but importantly branch significantly
patients with chronic visceral pain syndromes within the spinal cord and extend both rostrally
(Posserud, I. et al., 2004). These mediators may and caudally from the spinal segment of entry.
directly or indirectly affect visceral sensory function Significantly, some terminations reach the con-
(e.g., Tache, Y. et al., 2004) and may function as tralateral side of the spinal cord.
targets for pharmacologic interventions. (6) Most mechanoreceptors in viscera have low
thresholds for response to stretch/tension (e.g.,
balloon distension), but a significant proportion
37.5 Summary (2030%) have high thresholds for response.
The latter group fulfills criteria for nociceptors:
their response threshold is in the noxious range
(1) The principal conscious sensations that arise (generally 30 mm Hg distending stimulus)
from the viscera are discomfort and pain. and they encode stimulus intensity in the nox-
Typically, visceral pain is diffuse and difficult ious range, but do not encode innocuous
to localize, not distinctly felt at the source and
intensities of stimulation.
instead referred to other tissues.
(7) Low-threshold visceral mechanoreceptors
(2) Most experimental work in humans and nonhu-
respond in the physiologic range, but in addi-
man animals has utilized hollow organ disten-
tion possess properties that suggest involvement
sion, an adequate (in the Sherringtonian context)
in visceral discomfort and pain. Unlike low-
noxious visceral stimulus. Visceral inflammation
threshold cutaneous mechanoreceptors, low-
and ischemia are also considered to be adequate
threshold visceral mechanoreceptors respond
stimuli, but they are more difficult to control
vigorously to noxious intensities of stimulation,
experimentally and thus have been less widely
continue to encode stimulus intensity into the
used. It is generally held that pain cannot be
noxious range, and sensitize (a defining charac-
produced from all viscera (e.g., the parenchyma
of solid organs like the liver) or from organs that teristic of nociceptors). These properties are not
have only an afferent innervation (and not a unlike some low-threshold sensory neurons that
sensory innervation), but new knowledge has innervate another deep tissue, muscle.
expanded our understanding of visceral sensa- (8) Low-threshold visceral mechanoreceptors are
tions and suggest that these and other issues of normally and principally involved in afferent
visceral insensitivity should be reevaluated. and reflex functions, but in conjunction with
(3) The innervation of the viscera is unique. Each high-threshold visceral mechanoreceptors can
organ is innervated by two nerves, either the contribute to intense acute visceral pain. In
vagus nerve and a spinal visceral nerve or a functional, inflammatory or ischemic visceral
splanchnic nerve and the pelvic nerve, and the disorders, where sensitization develops and
function(s) of each nerve differ. The route from sleeping visceral nociceptors become active,
the organ to the spinal cord of spinal visceral we believe that all mechanosensitive visceral
nerves involves transit of prevertebral ganglia, endings can contribute to uncomfortable and
where axon collaterals of visceral nerves can painful visceral sensation.
 Visceral Pain 565

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Vandenberghe, J., Vos, R., Persoons, P., Demyttenaere, K., 2003. Mechanotransduction by intraganglionic laminar
Janssens, J., and Tack, J. 2005. Dyspeptic patients with endings of vagal tension receptors in the guinea-pig
visceral hypersensitivity: sensitisation of pain specific or oesophagus. J. Physiol. 553, 575587.
multimodal pathways? Gut 54, 914919. Zhuo, M. and Gebhart, G. F. 2002. Facilitation and attenuation
Vera-Portocarrero, L. P., Yie, J., Kowal, J., Ossipov, M., King, T., of a visceral nociceptive reflex from the rostroventral medulla
and Porreca, F. 2006. Descending facilitation from the in the rat. Gastroenterology 122, 10071019.
rostral ventromedial medulla maintains visceral pain in rats Zhuo, M., Sengupta, J. N., and Gebhart, G. F. 2002. Biphasic
with experimental pancreatitis. Gastroenterology modulation of spinal visceral nociceptive transmission from
130, 21552164. the rostroventral medial medulla in the rat. J. Neurophysiol.
Wang, F. B. and Powley, T. L. 2000. Topographic inventories of 87, 22252236.
vagal afferents in gastrointestinal muscle. J. Comp. Neurol.
421, 302324.
Wemmie, J., Price, M., and Welsh, M. 2006. Acid-sensing ion
channels: advances, questions and therapeutic
opportunities. Trends Neurosci. 29, 578586.
Whitehead, W., Palsson, O., and Jones, K. 2002. Systematic Further Reading
review of the comorbidity of irritable bowel syndrome with
other disorders: What are the causes and implications? Berkley, K. J., Robbins, A., and Sato, Y. 1988. Afferent fibers
Gastroenterology 122, 11401156. supplying the uterus in the rat. J. Neurophysiol. 59, 142163.
Willert, R. P., Woolf, C. J., Hobson, A. R., Delaney, C., Brune, K. and Handwerker, H. O. (eds.) 2004. Hyperalgesia:
Thompson, D. G., and Aziz, Q. 2004. The development and Molecular Mechanisms and Clinical Implications, Progress in
maintenance of human visceral pain hypersensitivity is Pain Research and Management,, Vol. 30. IASP Press.
dependent on the N-methyl-D-aspartate receptor. Carr, M. J. and Undem, B. J. 2003. Bronchopulmonary afferent
Gastroenterology 126, 683692. nerves. Respirology 8, 291301.
Winnard, K. P., Dmitrieva, N., and Berkley, K. 2006. Cross-organ Cervero, F. 1994. Sensory innervation of the viscera: peripheral
interactions between reproductive, gastrointestinal, and basis of visceral pain. Physiol. Rev. 74, 95138.
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Francis, C. Y., Duffy, J. N., Whorwell, P. J., and Morris, J. 1997. and 5-HT in the rat jejunum. Am. J. Physiol.
High prevalence of irritable bowel syndrome in patients 283, G612G617.
attending urological outpatient departments. Dig. Dis. Sci. Linden, D. R., Chen, J.-X., Gershon, M. D., Sharkey, K. A., and
42, 404407. Mawe, G. M. 2003. Serotonin availability is increased in
Gebhart, G. F. (ed.) 1995. Visceral Pain, Progress in Pain mucosa of guinea pigs with TNBS-induced colitis. Am. J.
Research and Management, , Vol. 5. IASP Press. Physiol. 285, G207G216.
Janig, W. 2006. The Integrative Action of the Autonomic Ness, T. J. and Gebhart, G. F. 1990. Visceral pain: a review of
Nervous System. Cambridge University Press. experimental studies. Pain 41, 167234.
Kollarik, M. and Undem, B. J. 2002. Mechanisms of Pan, H. L. and Longhurst, J. C. 1996. Ischaemia-sensitive
acid-induced activation of airway afferent nerve fibres in sympathetic afferents innervating the gastrointestinal
guinea-pig. J. Physiol. 543, 591600. tract function as nociceptors in cats. J. Physiol.
Kollarik, M. and Undem, B. J. 2003. Activation of 492, 841850.
bronchopulmonary vagal afferent nerves with bradykinin, Pasricha, P. J., Willis, W. D.,, and Gebhart, G. F. (eds.)
acid and vanilloid receptor agonists in wild-type and 2007. Chronic Abdominal and Visceral Pain., Informa
TRPV1/ mice. J. Physiol. 555, 115123. Healthcare.
Kreis, M. E., Jiang, W., Kirkup, A. J., and Grundy, D. 2002. Spiller, R. and Grundy, D. (eds.) 2004. Pathophysiology of the
Cosensitivity of vagal mucosal afferents to histamine Enteric Nervous System., Blackwell Publishing.
This page intentionally left blank
 38 Irritable Bowel Syndrome
S Bradesi and E A Mayer, University of California, Los Angeles, CA, USA
I Schwetz, Medical University, Graz, Austria
2009 Elsevier Inc. All rights reserved.

38.1 Introduction 571

38.2 Clinical Presentation and Epidemiology 571
5.38.3 The Bidirectional BrainGut Axis Model 572
5.38.4 Visceral Hypersensitivity 572
5.38.4.1 Peripheral Up-Regulation of Visceral Afferent Sensitivity 573
5.38.4.2 Spinal and Supraspinal Up-Regulation of Visceral Afferent Sensitivity 574
5.38.5 Treatment Options 575
5.38.6 Summary and Conclusions 575
References 575

38.1 Introduction hypothesis of hyperresponsiveness of central stress

circuits has been proposed (Mayer, E. A. and
Recurrent abdominal pain or discomfort in the Collins, S. M., 2002).
absence of detectable structural or biochemical
abnormalities, associated with alterations in bowel
habits, are the principal symptoms of irritable bowel
38.2 Clinical Presentation and
syndrome (IBS) (Drossman, D. A. et al., 2002). Due to
Epidemiology
the likely heterogeneity of the syndrome (Mayer, E. A.
and Collins, S. M., 2002) and the lack of reliable
IBS is one of the most common and most thoroughly
organic markers, the development of a unifying, and
studied functional disorders of the gastrointestinal
generally agreed upon, hypothesis of the pathophy-
tract (Drossman, D. A. et al., 2002). In additional to
siology of IBS has remained an elusive goal. Many
chronic abdominal pain, the clinical presentation of
investigators in the field agree that an enhanced IBS typically also includes nonpainful abdominal
perception of physiologically occurring, or experi- discomfort (sensations of urgency, bloating, fullness,
mentally generated visceral events (visceral gas and constipation) (Lembo, T. J. and Fink, R. N.,
hypersensitivity) (Mayer, E. A. and Gebhart, G. F., 2002) and visible abdominal distension. These gas-
1994; see Vagal Afferent Neurons and Pain and trointestinal symptoms are frequently associated with
Visceral Pain) in conjunction with alterations in gas- extraintestinal symptoms such as fatigue, decreased
trointestinal motility and secretory function, are key energy level, impaired sleep, depression, and anxiety
pathophysiological mechanisms underlying the car- (Zimmerman, J., 2003). In the absence of generally
dinal clinical features of IBS. In contrast, many other agreed upon reliable biological markers, the diagno-
alterations reported in IBS patients over the past sis of IBS remains one based on the presence of the
decades, including altered mucus production and so-called symptom criteria (Thompson, W. G. et al.,
altered gastrointestinal motility, have turned out to 1999). The most widely accepted diagnostic criteria
be epiphenomena which are unlikely to be essential are the Rome II Criteria that evolved from the Rome
for symptom generation. Considerable evidence sup- I and the Manning criteria initially defined about a
ports the role of psychosocial (Bennett, E. J. et al., decade ago (Longstreth, G. F., 2005). Different IBS
1998; Collins, S. M., 2002) and physical (Gwee, K. A. patient subtypes have been identified based on their
et al., 1996) (i.e., acute gastroenteric infections) stres- bowel habit predominance and have been classified
sors as central and peripheral triggers of first as: constipation, diarrhea, or alternating periods of
symptom onset or symptom exacerbation of long- both. In a large US sample, approximately 50% of
standing IBS (Mayer, E. A. et al., 2001), and an IBS IBS patients present alternating bowel habit (IBS-A),

571
572 Irritable Bowel Syndrome

30% diarrhea (IBS-D), and 20% constipation (IBS- Therefore, every conceptual model of IBS has to
C) (Tillisch, K. et al., 2005). take into account that neither the central nervous
Large population based studies in the US have system nor the gastrointestinal tract function in iso-
found a prevalence of IBS of about 14% with a lation, but that both systems interact with each other
greater proportion of women affected (female-to- under normal conditions and particularly during per-
male ratio approximately 3:1 to 3:2). The socioeco- turbations of homeostasis. Afferent signals arising
nomic consequences of IBS are considerable with a from the lumen of the gut are transmitted via various
large impact on work productivity and absenteeism visceral afferent pathways (enteric, spinal, vagal) to
(Dean, B. B. et al., 2005). In addition, a substantial the central nervous system. Homeostatic reflexes,
reduction in health-related quality of life (HRQoL) which generate appropriate gut responses to physio-
has been observed in IBS patients with moderate to logical as well as pathological visceral stimuli, occur
severe symptoms who are seen in a referral setting at the level of the enteric nervous system, the spinal
compared with healthy controls (Spiegel, B. M. et al., cord, and pontomedullary nuclei and limbic regions.
2004). Interestingly, bowel habit alterations contrib- Vagal visceral afferent inputs may also play an
uted little to the HRQoL impairment in this study, important role in such diverse functions as modula-
while extraintestinal manifestations such as loss of tion of emotion, pain sensitivity, satiety, and immune
energy, fatigue, and excessive worry were important response ( Janig, W. and Habler, H.-J., 2000).
determinants. The annual cost of IBS in the US has Whereas the reflex circuits within the enteric ner-
been estimated to be between $1.7 billion and $10 vous system in principle can regulate and
billion in direct medical cost (repetitive use of multi- synchronize all basic gastrointestinal functions (moti-
ple healthcare resources) and $20 billion for indirect lity, secretion, blood flow), coordination of gut
costs (work absenteeism and impaired productivity) functions with the overall homeostatic state of the
representing a high socioeconomic burden on society organism requires continuous communication
(Inadomi, J. M. et al., 2003). between the central nervous system and the gastro-
While research over the past few years has pro- intestinal tract (Mayer, E. A. and Collins, S. M., 2002).
vided significant advances in the understanding of Descending cortico-limbic influences can set the gain
IBS pathophysiology, the precise mechanism(s) and responsiveness of these reflexes, or impose distinct
underlying symptom generation remains incomple- patterns of motor responses on lower circuits, should
tely understood, generating considerable controversy the overall condition of the body make it necessary.
among investigators. As a result, the development of Such top-down override of local reflex function occurs
effective IBS therapies has been slow and disappoint- during sleep, during the stress response, or during
ing (Bradesi, S. et al., 2006). IBS pathophysiology is strong emotions such as fear and anger (Ito, M., 2002;
often viewed within the so-called biopsychosocial Mayer, E. A., 2000a; Tache, Y. et al., 2001; Welgan, P.
model in which altered physiology (gastrointestinal et al., 1988). While the great majority of homeostatic
motility and secretion, enhanced perception of afferent input from the gut (as well as other viscera) to
visceral stimuli (visceral hypersensitivity) and psy- the central nervous system is not consciously per-
chosocial factors interact and determine the clinical ceived, there are both peripheral and central
expression of IBS (Schwetz, I. et al., 2004). From a adaptive mechanisms which can result in enhanced
biological perspective, IBS symptomatology can be perception and altered reflex responses to visceral
viewed as the manifestation of alterations in the stimuli (Mayer, E. A. and Gebhart, G. F., 1994).
braingut axis, specifically as a dysregulation in the
complex interplay between events occurring within
the gut, the enteric nervous system, and the central 38.4 Visceral Hypersensitivity
nervous system (Mulak, A. and Bonaz, B., 2004).
The initial clinical observations that led to the hypoth-
esis that IBS patients exhibit visceral hypersensitivity
38.3 The Bidirectional BrainGut Axis include recurring abdominal pain, tenderness during
Model palpation of the sigmoid colon during physical exam-
ination, and excessive pain during endoscopic
Brain-gut interactions play a prominent role in the evaluation of the sigmoid colon (Mayer, E. A. and
modulation of gut function in health and disease Gebhart, G. F., 1994). Multiple human experimental
(Mayer, E. A. et al., 2001; Tache, Y. et al., 2001). studies using barostat-controlled balloon distension
 Irritable Bowel Syndrome 573

paradigms have reported lowered colorectal percep- Lamb, K. et al., 2006); (3) preliminary evidence that
tual thresholds, increased sensory ratings and supernatants from human mucosal biopsies (taken from
viscerosomatic referral areas in IBS patients com- IBS patients) have a unique effect on visceral afferent
pared to healthy individuals (Bouin, M. et al., 2002; function in rodent bioassays (Barbara, G. et al., 2005a;
Chang, L. et al., 2000a; Mertz, H. et al., 1995). Despite Barbara, G. et al., 2005b); (4) the development of IBS-
the uncertainty about the underlying mechanism(s), like symptoms in a small number of individuals fol-
this kind of experimentally induced visceral hyper- lowing an episode of acute gastroenteritis (Gwee, K.
sensitivity has been considered a pathophysiological A. et al., 1998; Spiller, R. C., 2005) and in a subset of
hallmark of the disease. Within the framework of patients with inflammatory bowel disease (Bernstein,
homeostatic afferents, the finding of chronic, con- C. N. et al., 1996); (5) evidence for alteration in the
scious awareness of unpleasant visceral sensations, normal gut flora (including bacterial overgrowth)
together with the presence of altered reflex regula- (Balsari, A. et al., 1982; OLeary, C. and Quigley, E.
tion in the gut, could be explained by several M., 2003; Swidsinski, A. et al., 1999); and (6) thera-
different mechanisms, including the following: (1) a peutic responses to treatment with antibiotics and
peripheral up-regulation of the sensitivity of visceral probiotic treatments (Madden, J. A. and Hunter, J.
afferent pathways which may be related to alterations O., 2002; Pimentel, M. et al., 2000, 2003). Without
in the activity of various effector cells within the gut discussing this large body of evidence supporting
(enteric nerves, enterochromaffin cells, immune each of these points (Barbara, G. et al., 2006;
cells); (2) a spinal or brainstem alteration in the Crowell, M. D. et al., 2005; Schwetz, I. et al., 2003;
sensitivity to incoming visceral afferents which Spiller, R. C., 2005), it is assumed, by many investi-
could be a consequence of primary peripheral or gators focusing on peripheral etiologies of visceral
central inputs; or (3) a primary central amplification hypersensitivity, that the mucosal immune activation
of perceptual and reflex responses to incoming visc- plays some role in maintaining a chronic state of
eral afferent signals. In addition, the frequent visceral afferent sensitization. Based on recent pre-
presence of compromised vital functions such as clinical evidence, it is conceivable that transient
sleep, mood, sexual drive, and affect, suggest a pos- peripheral sensitization may result in long-lasting
sible involvement of other homeostatic mechanisms, up-regulation of spino-bulbo-spinal pain amplifica-
such as tonic serotonergic pontomedullary systems tion mechanisms (Suzuki, R. et al., 2005). While such
(Mason, P., 2005). It remains to be determined if peripheral mechanisms (in particular an increase in
these various abnormalities occur in distinct subsets mucosal mast cells and intestinal enterochromaffin
of patients, at different stages of the disorder, or if cells) may play an important role in a subset of IBS
various combinations of them generate a heteroge- patients, or in the mediation of certain types of IBS
neous patient population. symptoms, a series of observations argues against a
simple relationship between peripheral mucosal
events and IBS symptoms. For example, it has long
38.4.1 Peripheral Up-Regulation of Visceral
been known that some patients in remission from
Afferent Sensitivity
inflammatory bowel disease, especially ulcerative
The concept that gut mucosal alterations may play a colitis, report symptoms similar to those of IBS
role in the pathophysiology of IBS has been patients (Isgar, B. et al., 1983; Simren, M. et al., 2000).
prompted by several lines of clinical and preclinical In contrast, inflammatory bowel disease patients
evidence including: (1) alterations in the number of (without associated IBS features) who have chroni-
immune cells, particularly mast cells, within intest- cally recurring gut inflammation do not exhibit
inal biopsies of patients meeting diagnostic criteria visceral hypersensitivity (Chang, L. et al., 2000b) or
for IBS (Barbara, G. et al., 2004; OSullivan, M. et al., enhanced central nervous system responses to visceral
2000; Weston, A. P. et al., 1993), and the demonstra- distension during periods of remission (Loennig-
tion in rodent studies that mucosal mast cells play a Baucke, V. et al., 1989; Rao, S. S. C. et al., 1987). In
prominent role of transducers within the braingut axis addition, clinical states characterized by chronic
(Santos, J. et al., 2005; Soderholm, J. D. et al., 2002; inflammation of the esophagus (gastroesophageal
Wilson, L. M. and Baldwin, A. L., 1999); (2) demonstra- reflux disease) and stomach (Helicobacter pylori chronic
tion of visceral hypersensitivity in different animal gastritis) are also not associated with visceral mechan-
models of gut immune system activation (Barbara, G. ical hyperalgesia (Fass, R. et al., 1998; Mertz, H. et al.,
et al., 1997; Barreau, F. et al., 2004; La, J. H. et al., 2003; 1998). On the contrary, it has been reported that
574 Irritable Bowel Syndrome

Crohns patients with isolated inflammation in the impairments in adult life (Whitehead, M. and
small bowel have elevated discomfort thresholds to Holland, P., 2003). Long-term consequences of
controlled distension of the rectum (Bernstein, C. N. early aversive events also include an increased vul-
et al., 1996), and that patients with ulcerative colitis do nerability for stress-sensitive disorders such as IBS
not show sensitization following repetitive noxious and posttraumatic stress disorder (Lowman, B. C.
distension of the sigmoid colon (Chang, L. et al., et al., 1987). Stress hyperresponsiveness is related in
2000b). Taken together, these data are consistent part to permanent, stress-induced hypersecretion of
with the interpretation that chronic mucosal inflam- CRF (Heim, C. et al., 2000). A number of preclinical
matory changes in the esophagus, stomach, or colon by studies support the concept of centrally mediated
themselves do not necessarily result in mechanical alteration of visceral perception. Different stress
visceral hypersensitivity. Chronic intestinal inflamma- paradigms were found to lead to enhanced visceral
tion in inflammatory bowel disease seems to be response to colonic distension (Bradesi, S. et al., 2002;
associated with the activation of counterregulatory Gue, M. et al., 1997; Schwetz, I. et al., 2005), which can
antinociceptive systems, inhibition of pain facilitatory be mimicked or abolished by central administration
pathways or both, resulting in a reduced perception of of pharmacological agents. For example, central CRF
visceral afferent information (Chang, L. et al., 2000b). injection was found to mimic the effect of stress
One may speculate that genetic polymorphisms on visceral sensitivity, whereas stress-induced visc-
recently identified as being associated with greater eral hyperalgesia may be reduced by central injection
pain sensitivity may be related to such compromised of a CRF receptor subtype 1 antagonist (Tache, Y.
endogenous pain modulation systems (Diatchenko, L. et al., 2004). Similarly, a neurokinin-1 receptor
et al., 2005). antagonist injected into the spinal cord in stress-sen-
sitized guinea pigs (Greenwood-Van Meerveld, B.
et al., 2003) or rats (Bradesi, S. et al., 2005) abolished
38.4.2 Spinal and Supraspinal Up-
the stress-induced visceral hyperalgesia to colorectal
Regulation of Visceral Afferent Sensitivity
distension. Together, the data available suggest the
A growing body of literature supports the concept of role of central stress circuits in the alteration of
an enhanced stress responsiveness playing a role in the visceral nociceptive response in animal models
development of IBS symptoms in a subset of patients for IBS. In contrast to the well-known analgesic
(reviewed in Mayer, E. A., 2000b; Mayer, E. A. et al., response to severe acute stressors (stress-induced
2001). An individuals response to stress (perturbation somatic analgesia), prolonged and milder stressors
of homeostasis) is generated by a central network that are associated with anxiety-like states are
comprised of integrative brain structures, referred commonly associated with hyperalgesic responses
to as the emotional motor system (EMS) (for a (Boccalon, S. et al., 2006; Vendruscolo, L. F. et al.,
detailed review see Mayer, E. A., 2000b); Mayer, E. 2004).
A. et al., 2001). The main output systems of the EMS Suggestive evidence for an alteration in central
are ascending monoaminergic pathways, the auto- pain modulation mechanisms in IBS patients comes
nomic nervous system, the hypothalamicpituitary from a series of functional brain-imaging studies. In
adrenal axis and endogenous pain modulatory response to rectal balloon distension, IBS patients
systems. The neuropeptide, corticotropin-releasing have shown increased activation of subregions of
factor (CRF), plays a prominent role in integrating the dorsal anterior cingulate cortex (ACC)
these various outputs in response to physical and (Naliboff, B. D. et al., 2001b; Porreca, F. et al., 2002;
psychological stressors (Bale, T. L., 2005). Verne, G. N. et al., 2003), a brain region involved in
The responsiveness of the EMS and its various attentional and emotional modulation of stimulus
output pathways is under partial genetic control perception (Petrovic, P. and Ingvar, M., 2002).
(Pezawas, L. et al., 2005) and is programmed by Dorsal ACC subregions provide input to subcortical
prenatal (Matthews, S. G., 2002) and postnatal endogenous pain inhibitory circuits (Petrovic, P. and
(Ladd, C. O. et al., 2000) aversive events and by Ingvar, M., 2002), as well as to subcortical pain facil-
certain types of pathological stress (Fuchs, E. and itatory circuits (Zhuo, M. and Gebhart, G. F., 2002).
Fluegge, G., 1995). In humans, early (pre- and post- The relative balance between these simultaneously
natal) life adverse experiences can lead to long- activated pain modulation systems determines the
lasting stress hyperresponsiveness, which in turn overall modulation of perception (Porreca, F. et al.,
has been associated with a wide range of health 2002). Thus, the finding of greater dorsal ACC
 Irritable Bowel Syndrome 575

activation by a visceral stimulus in IBS may be 38.5 Treatment Options

related to greater attention and possibly associated
activation of pain facilitation circuits to a visceral Despite its high prevalence and considerable impact
stimulus in IBS. On the other hand, the lesser activa- on patients HRQoL, treatment options for IBS con-
tion of the periaqueductal gray (PAG) reported in tinue to be limited and there are few well designed
some studies (Naliboff, B. D. et al., 2001b) is consistent studies to support the effectiveness of some of the
with possible deficiencies in the cortical activation of most commonly used therapies (for review see
endogenous pain inhibition systems. Several recent Camilleri, M., 2004). Traditional treatment algo-
studies (Chang, L., 2005) provide further support for rithms have primarily been aimed at peripheral
the hypothesis of altered endogenous pain modula- targets and are largely symptom-based (e.g., laxa-
tion in IBS patients. In one study (Mayer, E. A. et al., tives, antidiarrheals, prokinetics). In addition,
2005), IBS patients were compared to patients with centrally targeted therapies include various forms of
ulcerative colitis and with healthy control subjects. cognitive behavioral therapy (Hutton, J., 2005), low-
IBS patients showed consistently greater activation of dose tricyclic antidepressants (Drossman, D. A. et al.,
limbic/paralimbic brain regions (amygdala, hypotha- 2003) and, in patients with co-morbid psychiatric
lamus, ventral/rostral ACC, dorsomedial prefrontal conditions, full dose serotonin reuptake inhibitors
cortex) suggestive of increased activation of arousal (Creed, F. et al., 2003). A series of compounds are
circuits by a visceral stimulus. In addition, the results currently in development, which is targeted at cen-
showed activation in the ulcerative colitis and control tral circuits involved in stress responsiveness and
subjects, but not in IBS patients, in the lateral pre- pain modulation (Bradesi, S. et al., 2006).
frontal regions and a midbrain region including the
PAG. A connectivity analysis using structural equa-
tion modeling supported these regions acting as part 38.6 Summary and Conclusions
of a pain inhibition network that involves lateral and
medial prefrontal influences on the PAG. Another Despite its high prevalence, and considerable burden
study provided evidence for the abnormal activation of illness, treatment of IBS remains unsatisfactory.
of diffuse noxious inhibitory control systems in However, considerable progress has been made to
response to a noxious stimulus in IBS patients identify alterations at different levels of the brain
(Wilder-Smith, C. H. et al., 2004). gut axis which may contribute to characteristic
Several lines of evidence indicate that patients symptoms. In the absence of generally agreed upon
with IBS and other functional disorders have hyper- animal models of the syndrome, functional brain-
vigilance for symptom relevant sensations (Berman, imaging techniques in well defined human patient
S. M. et al., 2002). In a recent longitudinal study of IBS populations has a high promise to identify central
patients exposed to six sessions of rectal inflations abnormalities related to altered stress responsiveness
over a 1-year period, regional cerebral blood flow to and associated pain modulation circuits. A number of
the inflations and anticipation of inflations novel treatment approaches aimed at these central
abnormalities are currently under development.
using H15 2 O positron emission tomography (PET) at
the first and last session were evaluated (Naliboff, B.
D. et al., 2001a). Subjective ratings of the rectal infla-
tions normalized over the course of the study References
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 39 Pain in Childbirth
U Wesselmann, The Johns Hopkins University School of Medicine, Baltimore, MD, USA
2009 Elsevier Inc. All rights reserved.

39.1 Introduction 579

39.2 Physiological Aspects of Labor Pain 579
39.2.1 Variables Associated with the Severity of Labor Pain 580
39.2.2 Treatment of Pain in Childbirth 581
39.2.2.1 Analgesia for Labor and Delivery 581
39.2.2.2 Nonpharmacological, complementary and alternative therapies for relief of pain in
childbirth 582
39.3 Pain during Pregnancy 582
39.4 Postpartum Pain 582
39.5 Future Aspects 582
References 583

Glossary
dysmenorrhea Painful cramping of the lower childbirth but the length of the period is not well
abdomen occurring before or during menses pri- defined.
marily as a result of endogenous prostaglandins, referred pain Pain perceived in a region different
often accompanied by other symptoms such as from the injured area during the time of injury.
sweating, tachycardia, headaches, nausea, vomit- stages of labor pain There are three stages of
ing, diarrhea, and tremulousness. labor pain. Stage 1 begins with regular uterine
dyspareunia Painful intercourse. contractions and ends with complete cervical dila-
epidural analgesia Regional analgesia produced tation at 10 cm. Stage 2 begins once the cervix has
by injection of local anesthetic solution into the completely dilated and ends with delivery of the
peridural space. fetus. Stage 3 lasts from the delivery of the fetus
postpartum pain Pain after childbirth; the post- until the delivery of the placenta.
partum period starts immediately following

39.1 Introduction (Melzack, R., 1984). Different from other acute

and chronic pain experiences, pain during child-
The experience of pain during childbirth is a com- birth is not associated with a pathological process.
plex, multidimensional response to sensory stimuli It is surprising that this physiological process asso-
generated during labor and delivery. Pain in child- ciated with the most basic and fundamental life
birth occurs in the context of an individual experience causes severe pain, and this has been
womans physiology and psychology, as well as in the subject of many philosophic and religious
the context of the sociology of the culture and the debates. The International Association for the
health care system and its providers surrounding Study of Pain (IASP) defines pain as an unpleasant
her. For the majority of women in all societies and sensory and emotional experience associated with
cultures, natural childbirth is likely to be one of actual or potential tissue damage or described in
the most painful events in their lifetime. Average terms of such damage (Merskey, H., 1979).
pain scores for labor pain are exceeded only by Traditionally, different approaches to pain man-
those for causalgia in chronic-pain patients and agement of the pain in childbirth have addressed
amputation of a digit in acute-pain patients either the sensory or the affective dimension of

579
580 Pain in Childbirth

pain, while more recently a multidimensional considered to be due to distension and traction on
approach has been advocated, addressing both pelvic structures surrounding the vaginal vault and
dimensions of labor pain (Lowe, N. K., 2002; from distension of the pelvic floor and perineum.
Melzack, R., 1984). The mean intensity of pain in childbirth has been
reported to be positively correlated with the inten-
sity, duration, and frequency of uterine contractions
39.2 Physiological Aspects of Labor and with the degree of cervical distension.
Pain Pain associated with childbirth provokes a gener-
alized stress response, which has widespread
Compared to many other pain conditions, research physiological and potentially adverse effects on the
on the neurophysiological mechanisms of pain in progress of labor and the well-being of the mother
childbirth has been sparse, due to the difficulty of and the fetus (Brownridge, P., 1995; Beilin, Y., 2002).
developing animal models that could be studied Respiratory effects include hyperventilation, which
adequately using electrophysiological and neuroana- might lead to maternal hypocarbia and respiratory
tomical techniques. Studies examining the influence alkalosis, and rise in cardiac output, peripheral
of pregnancy on somatosensory responses in animals resistance, and blood pressure. Pain associated with
and humans have shown hypoalgesia in late preg- uterine contraction results in stimulation of the
nancy prior to the onset of labor and this chance in release of stress-related hormones from the adrenal
nociceptive threshold is at least in part opioid- sympathetic axis and the hypophysealpituitary
mediated (Bajaj, P. et al., 2002; Jarvis, S. et al., 1997). axis. Labor pain promotes maternal and fetal acido-
There are three distinct stages of labor pain related to sis, which is due to a catecholamine-induced shift
uterine contractions, cervical stretching, and disten- toward lipolytic metabolism, hyperventilation, phy-
sion of the vaginal canal during fetal descend (for sical exertion, starvation, and diminished buffering
reviews see: McDonald, J. S., 2001; Rowlands, S. and capacity secondary to respiratory alkalosis. While
Permezel, M., 1998). The first stage begins with reg- these effects may be largely innocuous during the
ular uterine contractions and ends with complete course of uncomplicated labor, they present a great
cervical dilatation at 10 cm. Stage 1 has been further risk in the presence of certain medical and obstetric
subdivided into an earlier latent phase and an ensu- complications and in situations where fetal compro-
ing active phase, which begins at about 34 cm of mise already exists.
cervical dilatation and heralds a period of more
rapid cervical dilation. Once the cervix has comple-
39.2.1 Variables Associated with
tely dilated, stage 2 of labor has begun. It ends with
the Severity of Labor Pain
delivery of the fetus. Stage 3 of labor lasts from the
delivery of the fetus until the delivery of the placenta. There is a very high level of individual variability
Pain during the first stage of labor (dilatation phase) in the severity of labor pain and this has been
is thought to be due to nociceptive stimuli arising correlated with several factors (Melzack, R., 1984;
from mechanical distension of the lower uterine seg- Brownridge, P., 1995). Primiparas and younger
ment and cervical dilatation. In addition, high- women report more pain than multiparas and
threshold mechanoreceptors in the myometrium older women. Women of higher socioeconomic sta-
may be activated in response to uterine contractions. tus report less pain than women of lower
Several chemical nociceptive mediators have been socioeconomic status. In addition physical factors
suggested, including bradykinines, leukotrienes, might play a role in increased pain ratings during
prostaglandins, serotonin, lactic acid, and Substance labor, including increased fetal size and increased
P. Pain of the first stage of labor is predominantly maternal body weight. Labor pain is influenced by
mediated by neural pathways involving the T10 to maternal positions women experience more pain
L1 spinal cord segments. Similar to other types of when delivering in the horizontal position as com-
visceral pain, labor pain may present with referred pared to the upright position. Childbirth at night
pain to somatic structures in corresponding myo- has been reported to be less painful than childbirth
tomes and dermatomes, including the abdominal during the day. Women with a history of dysmenor-
wall, lumbosacral region, iliac crests, gluteal areas rhea report higher pain levels during labor as
and thighs. Pain in the second and third stages of compared to women, who do not have a history of
labor involves spinal cord segments S2 to S4, and is menstrual pain. In contrast, a previous history of
 Pain in Childbirth 581

nongynecological pain is correlated with decreased women, but are more likely to give rise to positive
labor pain. consequences related to coping, self-efficacy and
Severe fear of pain associated with childbirth self-esteem (Niven, C. A. and Murphy-Black, T.,
occurs in 610% of parturients and is highly corre- 2000).
lated to pain levels reported during the first stage of It is important to emphasize that labor pain,
labor (for review see Saisto, T. and Halmesmaki, E., although being a very prominent aspect of childbirth,
2003). It is not an isolated variable, but associated is just one aspect of the childbirth experience. A
with the womans personal characteristics including recent systematic review of the literature on pain
general anxiety and fear of pain in general, low self- and womens satisfaction with the experience of
esteem, depression, dissatisfaction with her partner- childbirth demonstrated that the influences of pain,
ship, and lack of support. Fear of labor pain is pain relief, and intra-partum medical interventions
strongly associated with fear of pain in general, inde- on subsequent satisfaction are not as powerful as the
pendent of parity and is one of the most common personal expectations, the amount of support from
reasons for requesting a cesarean section. Fear of caregivers, the quality of the caregiver-patient rela-
childbirth has been reported to complicate about tionship, and involvement of decision making
20% of pregnancies in developed countries. (Hodnett, E. D., 2002).
Although it is often assumed that culture and
ethnicity have a significant influence on the intensity
of labor pain, numerous studies have documented
that there is no difference in self-report pain intensity 39.2.2 Treatment of Pain in Childbirth
ratings (see Lowe, N. K., 2002 for review). However, 39.2.2.1 Analgesia for Labor and Delivery
pain behavior is significantly influenced by culture Analgesia for labor and delivery is now safer than
and ethnicity, due to learned values and attitudes to ever and can be offered during all stages of labor,
the perception and expression of acute pain. targeted to the individual needs and wishes of the
The environmental influences on pain perception pregnant woman, without compromising the safety of
have been explored in two prospective studies from her or her unborn child (for reviews see Beilin, Y.,
Scandinavia comparing low-risk women delivering at 2002; Caton, D. et al., 2002; Nystedt, A. et al., 2004).
birth centers and at standard obstetrical hospital- Anesthesia related maternal mortality has decreased
based units (Skibsted, L. and Lange, A. P., 1992; from 4.3 million live births during the years 197981
Waldenstrom, U. and Nilsson, C. A., 1994). These
to 1.7 per million live births during the years 1988
studies suggest that the environment provided may
90. The increased use of regional anesthesia techni-
affect a womans ability to cope with pain. While
ques is partially responsible for this decrease in
women delivering in birth centers reported signifi-
mortality. The most popular method for analgesia
cantly higher pain intensities than women delivering
during labor and delivery is epidural analgesia
in hospital settings, there were no differences
using a combination of local anesthetics and opioids.
between the two groups with respect to satisfaction
Its popularity is related to its efficacy and safety.
with the quality of the birth experience. These results
Drugs can be applied as continuous epidural infu-
emphasize the importance of differentiation between
pain intensity and the attitude toward the pain sions or as patient-controlled epidural analgesia.
experienced in labor and delivery. During the early stages of labor dilute solutions of
It has been postulated that the memory for pain local anesthetic can be used to achieve analgesia. As
associated with disease, trauma, or surgical and med- labor progresses, more concentrated solutions of local
ical procedures can be more damaging than its initial anesthetics can be used and opioids can be added.
experience (Song, S.-O. and Carr, D. B., 1999). Typically the epidural catheter is inserted to main-
Although pain in childbirth is one of the most intense tain a low dermatomal level of analgesia for vaginal
pains many women experience in their lifetime, delivery (T10 to L1). If a cesarean section is required,
childbirth is one of the most positive events of life the dermatomal level can be raised to T4. Combined
for most women. Review of the literature on memory spinal epidural techniques offer the advantage of a
for labor pain shows that while memory of the events very rapid onset of analgesia with minimal motor
of childbirth is very accurate, the accuracy of recalled block. The future of obstetric anesthesia lies in refin-
labor pain remains in question. Memories of labor ing currently available drugs and techniques to make
pain can evoke intense negative reactions in a few obstetric anesthesia even safer and more efficacious.
582 Pain in Childbirth

39.2.2.2 Nonpharmacological, 39.4 Postpartum Pain

complementary and alternative therapies
for relief of pain in childbirth While most of the discussion of adverse sequelae of
Many nonpharmacological, complementary and labor and delivery has focused on urinary and fecal
alternative medicine methods exist to relieve incontinence, there is now increasing awareness that
labor pain (for reviews see Simkin, P. P. and pain associated with childbirth is not only related to
OHara M., 2002; Smith, C. A. et al., 2003; the process of labor and delivery. Women report
Cluett, E. R. et al., 2004; Cyna, A. M. et al., significant pains in many sites of the body after
2004; Huntley, A. L. et al., 2004; Lee, H. and delivery, a phenomenon which has been defined as
Ernst, E., 2004). These methods appeal to postpartum pain. Postpartum pains may include uro-
women and caregivers who are interested to genital, pelvic, back and breast pains as well as
reduce labor pain without creating potentially headaches, persisting for months to years after labor
serious side-effects and high costs. In addition and delivery (Audit Commission, 1997). It has been
many women appreciate the simplicity of these hypothesized that anesthetic techniques during labor
approaches and the sense of control they gain and delivery and gonadal hormonal changes may
from actively managing their pain. However, few play a role in the etiology of these pain complaints.
of these therapies have been subjected to proper Forty-nine percent of women report significant dys-
scientific study. Meta-analysis of randomized pareunia when resuming sexual intercourse after
controlled trials has indicated that acupuncture childbirth (Buhling, K. J. et al., 2005). Women with a
and hypnosis may be beneficial for the manage- history of operative vaginal delivery have the highest
ment of pain in childbirth. There is evidence that prevalence of severe perineal pain when resuming
water immersion during the first stage of labor sexual intercourse. The persistence of dyspareunia
reduces the use of analgesia and reported pain for longer than 6 months after delivery ranges from
intensity, without adverse outcomes on labor 3.4% to 14% based on mode of delivery. Treatment
duration, operative delivery, or neonatal out- approaches for early intervention to prevent persis-
comes. The effects of water immersion during tent perineal pain after childbirth have not been
the later stages of labor are not clear. No differ- explored in detail. A meta-analysis assessing the
ences were observed for women receiving effectiveness of topically applied anesthetics to the
aromatherapy, music, or audio analgesia. perineal region in the early postpartum period
showed no compelling evidence of pain reduction
(Hedayati, H. et al., 2005). However, there has been
no evaluation of the long-term effects of topically
39.3 Pain during Pregnancy applied local anesthetics.

While the focus of this review is on pain associated

with childbirth, it is important to note that pain is
also a significant issue during pregnancy (Stuge, B. 39.5 Future Aspects
et al., 2003). Approximately 50% of women experi-
ence back pain and/or pelvic pain during their Pain in childbirth is a complex, multidimensional
pregnancy, and in up to 15% of the cases the pain experience. While the focus of this chapter is on the
is rated as severe. Several hypotheses have been pain aspect of childbirth, it is important to keep in
suggested to explain the etiology of this pain, mind that pain is an important aspect, but not
including increased weight, decreased stability of the only aspect of the childbirth experience. The
the pelvic girdle due to hormonal changes and acknowledgment of the existence of pain associated
referred visceral pain mechanisms. The role of the with labor and delivery and the recognition of the
hormone relaxin and pelvic pain in pregnancy is severity of this pain by the medical community over
controversial. Physical therapy is often recom- the last 50 years has resulted in a broad spectrum of
mended for the prevention and treatment of these pharmacological and nonpharmacological pain man-
pregnancy-related pains, but the effects of these agement options that can be offered to the parturient
interventions have not been systematically studied today. There have been tremendous advancements in
and thus it is not clear if they are of any benefit to the pharmacological treatment of pain in labor and
the pregnant women. delivery over the last 25 years, mainly due to
 Pain in Childbirth 583

improved regional anesthesia techniques. The future Sakala, C., Simkin, P., and Young, D. 2002. The nature and
management of labor pain: executive summary. Am. J.
of obstetric anesthesia lies in refining currently Obstet. Gynecol. 186, S115.
available drugs and techniques and in developing Cluett, E. R., Nikodem, V. C., McCandlish, R. E., and
new drugs targeted at specific pathophysiological Burns, E. E. 2004. Immersion in water in pregnancy, labor
and birth. Cochrane Database Syst. Rev. CD000111.
mechanisms to make obstetric anesthesia even safer Cyna, A. M., McAuliffe, G. L., and Andrew, M. I. 2004. Hypnosis
and more efficacious. This will require translational for pain relief in labor and childbirth: a systematic review. Br.
research efforts ranging from basic science to clinical J. Anaesth. 93, 505511.
Hedayati, H., Parsons, J., and Crowther, C. A. 2005. Topically
research. As a first step an animal model has recently applied anaesthetics for treating perineal pain after
been developed in the rat to study the neurophysio- childbirth. Cochrane Database Syst. Rev. CD004223.
logical and neuropharmacological mechanisms of Hodnett, E. D. 2002. Pain and womens satisfaction with the
experience of childbirth: a systematic review. Am. J. Obstet.
pain associated with uterine cervical distension Gynecol. 186, S160172.
(Liu, B. et al., 2005). Neuroanatomical studies of the Huntley, A. L., Coon, J. T., and Ernst, E. 2004. Complementary
sensory pathways from the uterine cervix have shown and alternative medicine for labor pain: a systematic review.
Am. J. Obstet. Gynecol. 191, 3644.
that P2X3-receptor-expressing sensory neurons Jarvis, S., McLean, K. A., Chirnside, J., Deans, L. A.,
might play a role during birthing and signal nocicep- Calvert, S. K., Molony, V., and Lawrence, A. B. 1997. Opioid-
tive information such as labor pain (Papka, R. E. et al., mediated changes in nociceptive threshold during
pregnancy and parturition in the sow. Pain 72, 153159.
2005). Numerous nonpharmacological and alterna- Lee, H. and Ernst, E. 2004. Acupuncture for labor pain
tive treatments are available for the treatment of management: A systematic review. Am. J. Obstet. Gynecol.
pain in childbirth. While there is experience with 191, 15731579.
Liu, B., Eisenach, J. C., and Tong, C. 2005. Chronic estrogen
some of those approaches already for centuries, sensitizes a subset of mechanosensitive afferents innervating
many of these treatments have not been subjected the uterine cervix. J. Neurophysiol. 93, 21672173.
to proper scientific study. Clinical studies assessing Lowe, N. K. 2002. The nature of labor pain. Am. J. Obstet.
Gynecol. 186, S1624.
the effects of these interventions on pain in childbirth McDonald, J. S. 2001. Pain of Childbirth. In: Bonicas Management
and on a womans satisfaction with the childbirth of Pain, 3rd edn. (ed. J. D. Loeser), pp. 13881414. Lippincott
experience, as well as studies assessing the effects Williams & Wilkins.
Melzack, R. 1984. The myth of painless childbirth (the John J.
on the safety for the pregnant woman and her child Bonica lecture). Pain 19, 321337.
are urgently needed. Merskey, H. 1979. Pain terms: a list with definitions and a note
on usage. Recommended by the IASP Subcommittee on
Taxonomy. Pain 6, 249252.
Niven, C. A. and Murphy-Black, T. 2000. Memory for labor pain:
Acknowledgments a review of the literature. Birth 27, 244253.
Nystedt, A., Edvardsson, D., and Willman, A. 2004. Epidural
analgesia for pain relief in labor and childbirth a review with
Ursula Wesselmann is supported by NIH grants a systematic approach. J. Clin. Nurs. 13, 455466.
DK57315 (NIDDK), DK066641 (NIDDK), and Papka, R. E., Hafemeister, J., and Storey-Workley, M. 2005.
HD39699 (NICHD, Office of Research for P2X receptors in the rat uterine cervix, lumbosacral dorsal
root ganglia, and spinal cord during pregnancy. Cell Tissue
Womens Health). Res. 321, 3544.
Rowlands, S. and Permezel, M. 1998. Physiology of pain in
labor. Baillieres Clin. Obstet. Gynaecol. 12, 347362.
Saisto, T. and Halmesmaki, E. 2003. Fear of childbirth: a
References neglected dilemma. Acta Obstet. Gynecol. Scand.
82, 201208.
Audit Commission 1997. First Class Delivery: improving Simkin, P. P. and OHara, M. 2002. Nonpharmacologic relief of
Maternity Services in England and Wales. Audit Commission. pain during labor: systematic reviews of five methods. Am. J.
Bajaj, P., Madsen, H., Moller, M., and Arendt-Nielsen, L. 2002. Obstet. Gynecol. 186, S131159.
Antenatal women with or without pelvic pain can be Skibsted, L. and Lange, A. P. 1992. The need for pain relief in
characterized by generalized or segmental hypoalgesia in uncomplicated deliveries in an alternative birth center
late pregnancy. J. Pain 3, 451460. compared to an obstetric delivery ward. Pain 48, 183186.
Beilin, Y. 2002. Advances in labor analgesia. Mt. Sinai J. Med. Smith, C. A., Collins, C. T., Cyna, A. M., and Crowther, C. A. 2003.
69, 3844. Complementary and alternative therapies for pain management
Brownridge, P. 1995. The nature and consequences of in labor. Cochrane Database Syst. Rev. CD003521.
childbirth pain. Eur. J. Obstet. Gynecol. Reprod. Biol. Song, S-O. and Carr, D. B. 1999. Pain and memory. IASP Clin.
59 Suppl, S915. Updates 7, 14.
Buhling, K. J., Schmidt, S., Robinson, J. N., Klapp, C., Stuge, B., Hilde, G., and Vollestad, N. 2003. Physical therapy for
Siebert, G., and Dudenhausen, J. W. 2005. Rate of pregnancy-related low back and pelvic pain: a systematic
dyspareunia after delivery in primiparae according to mode review. Acta Obstet. Gynecol. Scand. 82, 983990.
of delivery. Eur. J. Obstet. Gynecol. Reprod. Biol.124, 4246. Waldenstrom, U. and Nilsson, C. A. 1994. Experience of
Caton, D., Corry, M. P., Frigoletto, F. D., Hopkins, D. P., childbirth in birth center care. A randomized controlled
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This page intentionally left blank
 40 Urothelium as a Pain Organ
L A Birder, University of Pittsburgh School of Medicine, Pittsburgh, PA, USA
2009 Elsevier Inc. All rights reserved.

40.1 Urothelial Cells: Detectors of Mechanical, Chemical, and Thermal Stimuli 585
40.1.1 Sensor Molecules Expressed in Urothelium Which Could Contribute to Bladder Pain 585
40.1.2 Response to Stimuli: Transducer Function of Urothelial Cells 587
40.2 How Might Urothelial Cells Influence Pain Processes? 588
40.3 Potential Clinical Implications: Urothelial Receptors/Release Mechanisms as Targets
for Drug Treatment 589
References 589

40.1 Urothelial Cells: Detectors of Birder, L. et al., 1998; 2004; Smith, P. R. et al., 1998;
Mechanical, Chemical, and Thermal Birder, L. et al., 2001; 2002a; 2002b; Chess-Williams,
Stimuli R., 2002; Beckel, J. et al., 2004; Birder, L., et al., 2004;
Chess-Williams, R., 2004; Burnstock, G. and Knight,
The urothelium is a specialized lining of the urinary G. E., 2004; Murray, E. et al., 2004; Tempest, H. V.
tract, extending from the renal pelvis to the urethra. et al., 2004; Chopra, B. et al., 2005; Birder, L. et al., 2007)
The urothelium is composed of at least three layers: a and that both types of cells use diverse signal trans-
basal cell layer attached to a basement membrane, an duction mechanisms to detect physiological stimuli.
intermediate layer, and a superficial apical layer with
large hexagonal cells (diameters of 25250 mm) which
are also termed umbrella cells (Lewis, S. A., 2000; 40.1.1 Sensor Molecules Expressed in
Apodaca, G., 2004). The umbrella cells, and, perhaps, Urothelium Which Could Contribute to
intermediate cells may have projections to the base- Bladder Pain
ment membrane (Martin, B.F., 1972; Hicks M., 1975; One example of a urothelial sensor molecule is the
Apodaca, G., 2004). Basal cells, which are thought to TRP channel, TRPV1, known to play a prominent
be precursors for other cell types, normally exhibit a role in nociception and in urinary bladder function
low (36 months) turnover rate; however, accelerated (Szallasi, A., 2001). It is well established that painful
proliferation can occur in pathology. For example, sensations induced by capsaicin, the pungent sub-
using a model (protamine sulfate) that selectively stance in hot peppers, are caused by stimulation of
damages the umbrella cell layer has shown that the TRPV1, an ion channel protein (Caterina, M. J. et al.,
urothelium rapidly undergoes both functional and 1997; Caterina, M. J., 2001) which is activated by
structural changes in order to restore the barrier capsaicin as well as by moderate heat, protons, and
following injury (Lavelle, J. et al., 2002). lipid metabolites such as anandamide (endogenous
While the urothelium has been historically ligand of both cannabinoid and vanilloid receptors).
viewed primarily as a barrier, it is becoming increas- TRPV1 is expressed throughout the afferent limb of
ingly appreciated as a responsive structure capable of the micturition reflex pathway, Figure 1, including
detecting physiological and chemical stimuli, and urinary bladder unmyelinated (C-fiber) nerves that
releasing a number of signaling molecules. A number detect bladder distension or the presence of irritant
of investigators have described the release of diffusi- chemicals (Chancellor M. B. and de Groat, W. C.,
ble mediators from the urothelium which could 1999). In the urinary bladder, TRPV1 is not only
influence urinary bladder function (Ferguson, D.R. expressed by afferent nerves or myofibroblasts that
et al., 1997; Hawthorn, M.H. et al., 2000; Burnstock, G., form close contact with urothelial cells, but also by
2001; Chess-Williams, R., 2002). There is now abun- the urothelial cells themselves (Birder, L. et al., 2001).
dant evidence which indicates that urothelial cells Urothelial TRPV1 receptor expression correlates
display a number of properties similar to sensory with the sensitivity to vanilloid compounds, as exo-
neurons (nociceptors/mechanoreceptors) (see genous application of capsaicin or resiniferatoxin to
Table 1; Lewis, S. A. and Hanrahan, J. W., 1985; cultured cells increases intracellular calcium and

585
586 Urothelium as a Pain Organ

Table 1 Examples of sensor molecules (i.e., receptors/ion channels) associated with neurons that have been identified in
urothelial cells

Sensor function/stimuli Urothelial sensor molecules Neuronal sensor molecules

ATP P2X/P2Y P2X/P2Y

Capsaicin resiniferatoxin TRPV1 TRPV1
Heat TRPV1; TRPV2; TRPV4 TRPV1; TRPV2; TRPV3; TRPV4
Cold TRPM8; TRPA1 TRPM8; TRPA1
H TRPV1; ? TRPV1; ASIC; DRASIC
Osmolarity In part TRPV4 In part TRPV4
Bradykinin B1; B2 B1; B2
Acetylcholine Nicotinic/muscarinic Nicotinic/muscarinic
Norepinephrine / subtypes / subtypes
Nerve growth factor p75/trkA p75/trkA
Mechanosensitivity Amiloride sensitive Na channels Amiloride sensitive Na channels

(c) (d)

Small-medium Superficial
diameter DRG spinal cord
neurons dorsal horn

DRG

Urinary bladder Spinal cord

(a) (e)
(b)

Submucosal bladder
nerves
Urothelial cells
Afferent terminals near
preganglionic neurons
Figure 1 TRPV1 is expressed throughout the afferent limb of the micturition reflex pathway. TRPV1-immunoreactivity
(cy-3, red) in basal epithelial cells (cyt 17, FITC green) (a); in nerve fibers (cy-3, red) located in close proximity to basal cells
(FITC, green) (b) (punctate TRPV1 staining in urothelial cells was electronically subtracted to facilitate imaging of the TRPV1-
immunoreactive nerve fiber); in small to medium diameter dorsal root ganglion (DRG) neurons (c); and in superficial regions of
the spinal cord dorsal horn (d) (staining indicate TRPV1 nerve fibers). (e) TRPV1-positive afferent terminals (in black) are
localized in close proximity to pre-ganglionic neurons (PGN neurons labeled by injection of a tracer dye, fast blue, into the
major pelvic ganglion). Reproduced from Birder, L. A. 2005. More than just a barrier: Urothelium as a drug target for urinary
bladder pain. Am. J. Physiol. Renal. Physiol. 289(3): F489F495, used with permission from the American Physiological
Society.
 Urothelium as a Pain Organ 587

evokes transmitter (NO, nitric oxide; ATP, adeno- families of purinergic receptors have been identified,
sine triphosphate) release (Birder, L. et al., 2001; P2X and P2Y, both of which are expressed in urothe-
2002a). In neurons, TRPV1 is thought to integrate/ lial cells (Lee, H.Y. et al., 2000; Birder, L. et al.,
amplify the response to various stimuli playing an 2004; Tempest, H. V. et al., 2004). Although the func-
essential role in the development of inflammation- tion of purinergic receptors in nonexcitable cells is
induced hyperalgesia (Ghuang, H. H. et al., 2001; less clear than in afferent neurons, the presence of
Holzer, P., 2004). Thus, it seems likely that urothelial such receptors may be associated with cell prolifera-
TRPV1 might participate in a similar manner, in the tion, apoptosis, secretion, and sensory transduction
detection of irritant stimuli following bladder inflam- (Coutinho-Silva R. et al., 2005; Greig, A. V. et al.,
mation or infection. 2003). In the urinary bladder, recent studies have
Anatomically normal, TRPV1-null mice exhib- shown that urothelial-derived ATP release can act
ited a number of alterations in bladder urothelial as a trigger for exocytosis, in part, via autocrine
cell function including a reduction of in vitro, activation of urothelial purinergic (P2X; P2Y) recep-
stretch-evoked ATP release and membrane capaci- tors (Wang, E. C. et al., 2005). This type of signaling
tance as well as a decrease in hypotonic-evoked ATP may be similar to that in airway epithelium, where
release (Birder, L. et al., 2002a). These findings nucleotides released to the epithelial surface may act
demonstrate that the functional significance of in a paracrine/autocrine manner to regulate ion trans-
TRPV1 in the bladder extends beyond pain sensation port and/or other functions via interactions with
to include participation in normal bladder function, luminal epithelial purinergic receptors (Poulsen, A.
and is essential for normal mechanically evoked pur- N. et al., 2005).
inergic signaling by the urothelium.
In addition to TRPV1, urothelial cells also express
additional TRP channels, including TRPV2,
TRPV4, TRPM8, and TRPA1. In contrast to 40.1.2 Response to Stimuli: Transducer
TRPV1, TRPV2, and TRPV4, which are detectors Function of Urothelial Cells
of warm temperatures (TRPV4 can also be gated by Release of chemical mediators (NO, ATP, acetylcho-
hypotonic stimuli) (Liedtke, W. et al., 2000;
line (ACh); substance P; prostaglandins (PG))
Alessandri-Haber, N. et al., 2003; Chung, M. K. et al.,
(Ferguson, D. R. et al., 1997; Birder, L. et al., 1998;
2003), TRPM8 and TRPA1 have been shown to be
Burnstock, G., 2001; Birder, L. et al., 2003; Chess-
activated by cold (2528  C) temperatures as well as
Williams, R., 2004) from urothelial cells suggests
by cooling agents (menthol, icilin) and are expressed
that these cells exhibit specialized sensory and
in a subset of sensory neurons as well as in non-
signaling properties that could allow reciprocal com-
neural cells. This expression suggests that these
munication with neighboring urothelial cells as well
cells express a range of thermoreceptors underlying
as nerves or other cells (i.e., immune, myofibroblasts,
both cold and heat stimuli (Stein, R. J. et al., 2004).
While the functional role of these thermosensitive inflammatory) in the bladder wall. Recent studies
channels in urothelium remains to be clarified, it have shown that both afferent as well as autonomic
seems likely that a primary role for these proteins axons are located in close proximity to the urothe-
may be to recognize noxious stimuli in the bladder. lium (Wayabayashi, Y. et al., 1995; Birder, L. et al.,
However, the diversity of stimuli which can activate 2001). For example, peptide and TRPV1 immunor-
these proteins suggests a much broader sensory and/ eactive nerve fibers have been found localized
or cellular role. For example, TRPM8 expression has throughout the urinary bladder musculature and in
been shown to be increased in some epithelia in a plexus beneath and extending into the urothelium.
malignant disorders (prostate tumors), suggesting a Confocal microscopy revealed that TRPV1 immu-
role in proliferating cells (Tsavaler, L. et al., 2001). noreactive nerve fibers are in close association with
Thus, further studies are needed to fully elucidate basal urothelial cells such that their fluorescent sig-
the role of TRP channels in urothelium and their nals overlapped within 0.5 mm optical sections. This
influence on bladder function. type of communication suggests that these cells may
Purinergic receptors, which are activated by ATP be targets for transmitters released from bladder
and related nucleotides, are known to play an impor- nerves or other cells, or that chemicals released by
tant role in bladder function and chronic pain urothelial cells may also alter the excitability of
(Burnstock, G., 2001; North, R. A., 2004). Two bladder nerves.
588 Urothelium as a Pain Organ

Studies have demonstrated that ATP released neuronal cells (urothelial cells, fibroblasts, mast cells)
from urothelium during bladder stretch could con- located near the bladder luminal surface. An impor-
tribute to activation of bladder afferents. tant component of the inflammatory response is ATP
That ATP released from urothelial cells during release from various cell types including urothelium,
stretch can activate a population of suburothelial which can initiate painful sensations by exciting pur-
bladder afferents expressing P2X3 receptors, signal- inergic (P2X) receptors on sensory fibers (Cockayne,
ing changes in bladder fullness and pain supports this D. A. et al., 2000; Burnstock, G., 2001). Recently, it has
idea (Ferguson, D. R. et al., 1997; Burnstock, G., 2001). been shown in sensory neurons that ATP can
Accordingly, P2X3 null mice exhibit a urinary blad- potentiate the response of vanilloids by lowering
der hyporeflexia, suggesting that this receptor and the threshold for protons, capsaicin, and heat
neuralepithelial interactions are essential for normal (Tominaga, M. et al., 2001). This represents a novel
bladder function (Cockayne, D. A. et al., 2000). This mechanism by which large amounts of ATP released
type of regulation may be similar to epithelial- from damaged or sensitized cells in response to injury
dependent secretion of mediators in airway epithelial or inflammation may trigger the sensation of pain.
cells which is thought to modulate submucosal These findings have clinical significance and suggest
nerves and bronchial smooth muscle tone and may that alterations in afferents or epithelial cells in
play an important role in inflammation (Homolya, L. pelvic viscera may contribute to the sensory abnorm-
et al., 2000; Jallat-Daloz, I. et al., 2001). Thus, it is alities in a number of pelvic disorders, such as IC,
possible that activation of bladder nerves and urothe- which is consistent with augmented release of ATP
lial cells can modulate bladder function directly or in urothelial cells from some patients with IC (Sun, Y.
indirectly via the release of chemical factors in the
et al., 2001). A comparable disease in cats is termed
urothelial layer.
feline interstitial cystitis (FIC) (Buffington, C. A.
et al., 1999; 2001), which is also accompanied by
alterations in stretch-evoked release of urothelially
40.2 How Might Urothelial Cells
derived ATP (Birder, L. et al., 2003).
Influence Pain Processes?
Though the urothelium maintains a tight barrier
to ion and solute flux, a number of factors such as
Recent evidence has demonstrated that urothelial
tissue pH, mechanical or chemical trauma, or bacter-
cells exhibit plasticity whereby inflammation or
ial infection can modulate this barrier function of the
injury can alter the expression and/or sensitivity of
urothelium (Hicks, M., 1975; Anderson, G. et al.,
a number of urothelial-sensor molecules. Examples
2003). When this function is compromised during
include changes in urothelial expression of various
receptors including tyrosine kinase (trk), low-affinity injury or inflammation, it can result in the passage
nerve growth factor (p75), bradykinin, TRPV1, pro- of toxic substances into the underlying tissue
tease-activated receptors (PPARs), and purinergic (neural/muscle layers) resulting in urgency, fre-
receptor (P2X and P2Y) subtypes in animal models quency, and pain during bladder distension. For
as well as in patients diagnosed with a number of example, inflammation, injury (spinal cord transec-
bladder disorders including neurogenic bladder and tion), or IC, all of which increase endogenously
interstitial cystitis (IC) a chronic clinical disease generated levels of NO, increase permeability to
characterized by urgency, frequency, and bladder water/urea in addition to producing ultrastructural
pain upon filling (an innocuous stimulus) (Murray, changes in the apical layer (Lavelle, J. et al., 2000;
E. et al., 2004; North, R. A., 2004; Tempest, H. V. et al., Apodaca, G. et al., 2003). Although the mechanism is
2004; Chopra, B. et al., 2005; Dattilio, A. and Vizzard, unknown, these findings may be similar to that in
M. A., 2005). Such changes could contribute to pain other epithelia where excess production of NO has
and hypersensitivity exhibited in these painful syn- been linked to changes in epithelial integrity (Han, X.
dromes. Because urothelial cells appear to exhibit et al., 2004).
sensory function, it is possible that plasticity in Disruption of epithelial integrity may also be due
urothelial receptors is then linked with pain in var- to substances such as antiproliferative factor (APF),
ious bladder syndromes. which has been shown to be secreted by bladder
Sensitization can be triggered by various media- epithelial cells from IC patients and can inhibit
tors (ATP, NO, nerve growth factor (NGF), PGE2) epithelial proliferation thereby adversely affecting
which may be released by both neuronal and non- barrier function (Keay, S. et al., 1999; 2004).
 Urothelium as a Pain Organ 589

40.3 Potential Clinical Implications: References

Urothelial Receptors/Release
Mechanisms as Targets for Drug Alessandri-Haber, N., Yeh, J. J., Boyd, A. E., Parada, C. A.,
Chen, X., Reichling, D. B., and Levine, J. D. 2003.
Treatment Hypotonicity induces TRPV4-mediated nociception in rat.
Neuron 39, 497511.
It is conceivable that the effectiveness of some agents Anderson, G., Palermo, J., Schilling, J., Roth, R., Heuser, J., and
Hultgren, S. 2003. Intracellular bacterial biofilm-like pods in
currently used in the treatment of bladder disorders urinary tract infections. Science 301, 105107.
may involve urothelial receptors and/or release Apodaca, G. 2004. The uroepithelium: not just a passive barrier.
Traffic 5, 112.
mechanisms. For example, intravesical instillation of
Apodaca, G., Kiss, S., Ruiz, W. G., Meyers, S., Zeidel, M. L., and
vanilloids (capsaicin or resiniferatoxin) improves uro- Birder, L. 2003. Disruption of barrier epithelium function after
dynamic parameters in patients with neurogenic spinal cord injury. Am. J. Physiol. 284, F966F970.
Barrick, S. R., Lee, H., Meyers, S., Caterina, M. J., Kanai, A. J.,
detrusor overactivity and reduces bladder pain in Zeidel, M. L., Chopra, B., de Groat, W. C., and Birder, L.
patients with hypersensitivity disorders, presumably 2003 Expression and function of TRPV4 in urinary bladder
by desensitizing bladder nerves (Szallasi, A. and urothelium. Soc. Neurosci. Abstr. 26, 608.6.
Beckel, J., Barrick, S. R., Keast, J.R., Meyers, S., Kanai, A. J., de
Fowler, C. J., 2002; Kim, J. H. et al., 2003). This treat- Groat, W. C., Zeidel, M. L., and Birder, L. 2004. Expression
ment could also target TRPV1 on urothelial cells, and function of urothelial muscarinic receptors and
whereby a persistent activation might lead to receptor interactions with bladder nerves. Soc. Neurosci. 26, 846.23.
Birder, L., Apodaca, G., de Groat, W. C., and Kanai, A. J. 1998.
desensitization or depletion of urothelial transmitters. Adrenergic- and capsaicin-evoked nitric oxide release from
Recent studies have demonstrated that intradetrusor urothelium and afferent nerves in urinary bladder. Am. J.
injection with botulinum neurotoxin type A (BoNTA) Physiol. 275, F226F229.
Birder, L., Kanai, A. J., de Groat, W. C., Kiss, S., Nealen, M. L.,
is an effective treatment for bladder hypersensitivity Burke, N. E., Dineley, K. E., Watkins, S., Reynolds, I. J., and
disorders including neurogenic detrusor overactivity Caterina, M. J. 2001. Vanilloid receptor expression suggests
a sensory role for urinary bladder epithelial cells. Proc. Natl.
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cholinergic nerve terminals and cleaves the protein, Kanai, A., and Caterina, M. 2007. Activation of Urothelial-
TRPV4 by 4-PDD contributes to altered bladder reflexes in
SNAP25, necessary for exocytosis and release of acet- the rat. JPET (in press).
ylcholine (Harper M. et al., 2004). There is evidence Birder, L., Nakamura, Y., Kiss, S., Nealen, M. L., Barrick, S. R.,
the BoNTA can suppress the release of a number of Kanai, A. J., Wang, E., Ruiz, W. G., de Groat, W. C.,
Apodaca, G., Watkins, S., and Caterina, M. J. 2002a. Altered
mediators (acetylcholine, ATP, and neuropeptides) urinary bladder function in mice lacking the vanilloid receptor
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et al., 2001). Suppression of neurotransmitter release Birder, L., Nealen, M. L., Kiss, S., de Groat, W. C.,
Caterina, M. J., Wang, E., Apodaca, G., and Kanai, A. J.
from urothelium would serve to blunt afferent activity 2002b. Beta-adrenoceptor agonists stimulate endothelial
driven by urothelial-derived release of mediators in a nitric oxide synthase in rat urinary bladder urothelial cells. J.
Neurosci. 22, 80638070.
number of lower urinary tract dysfunctions. These Birder, L., Barrick, S. R., Roppolo, J. R., Kanai, A. J., de
findings suggest that urothelial cells exhibit specia- Groat, W. C., Kiss, S., and Buffington, C. A. 2003. Feline
lized sensory and signaling properties that could interstitial cystitis results in mechanical hypersensitivity and
altered ATP release from bladder urothelium. American J.
allow them to respond to their chemical and physical Physiol. 285, F423F429.
environments and to engage in reciprocal communi- Birder, L., Ruan, H. Z., Chopra, B., Xiang, Z., Barrick, S. R.,
cation with neighboring urothelial cells as well as Buffington, C. A., Roppolo, J. R., Ford, A. P., de Groat, W. C.,
and Burnstock, G. 2004. Alterations in P2X and P2Y
nerves within the bladder well. Taken together, phar- purinergic receptor expression in urinary bladder from
macologic interventions aimed at targeting urothelial normal cats and cats with interstitial cystitis. Am. J. Physiol.
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Buffington, C. A. 2001. Visceral pain in humans: lessons from
isms may provide a new strategy for the clinical animals. Curr. Pain Headache Rep. 5, 4451.
management of bladder disorders. Buffington, C. A., Chew, D. J., and Woodworth, B. E. 1999.
Feline interstitial cystitis. J. Am. Vet. Med. Assoc.
215, 682687.
Burnstock, G. 2001. Purine-mediated signalling in pain and
visceral perception. Trends Pharmacol. Sci. 22, 182188.
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NIH (RO1 DK54824 and RO1 DK57284). to the pain pathway. Ann. Rev. Neurosci. 24, 602607.
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 41 The Brainstem and Nociceptive Modulation
M M Heinricher, Oregon Health & Science University, Portland, OR, USA
S L Ingram, Washington State University, Vancouver, WA, USA
2009 Elsevier Inc. All rights reserved.

41.1 The Periaqueductal GrayRostral Ventromedial Medulla Pain-Modulating System 594

41.1.1 Functional Characterization of the Periaqueductal GrayRostral
Ventromedial Medulla Pain-Modulating System 594
41.1.2 Pain Modulation as Part of Adaptive Responses to Behavioral and
Physiological Challenges 595
41.2 The Periaqueductal Gray 596
41.2.1 The Periaqueductal Gray and Facilitation of Nociception 597
41.2.2 Afferents to the Periaqueductal Gray 597
41.2.3 Efferents from the Periaqueductal Gray 598
41.2.4 Columnar Organization of the Periaqueductal Gray 598
41.2.5 Intrinsic Circuitry and Neurotransmitters in the Periaqueductal Gray 599
41.2.5.1 Endogenous opioids 599
41.2.5.2 Norepinephrine 603
41.2.5.3 Substance P 603
41.2.5.4 Cannabinoids 604
41.3 The Rostral Ventromedial Medulla 604
41.3.1 Connections of the Rostral Ventromedial Medulla 604
41.3.2 The Rostral Ventromedial Medulla and Facilitation of Nociception 604
41.3.3 The Neural Basis for Bidirectional Control from the Rostral Ventromedial Medulla:
On- and Off- Cells 605
41.3.3.1 Physiological classification of rostral ventromedial medulla neurons
based on reflex-related activity 605
41.3.3.2 Role of on- and off-cells in pain modulation 607
41.3.3.3 Role of neutral cells 608
41.3.4 Pharmacology of Nociceptive Modulation in the Rostral Ventromedial Medulla 609
41.3.4.1 Gamma-aminobutyric acid and glutamate: the off-cell pause and on-cell burst 609
41.3.4.2 Opioid actions in the rostral ventromedial medulla 610
41.3.4.3 Norepinephrine 611
41.3.4.4 Cannabinoids 612
41.3.4.5 Cholecystokinin 612
41.3.4.6 Neurotensin 612
41.3.4.7 Nociceptin/orphanin FQ 612
41.3.5 Physiological Activation of Nociceptive Modulatory Neurons in the
Rostral Ventromedial Medulla 613
41.3.5.1 Response of RVM neurons to noxious stimulation: does pain inhibit pain? 613
41.3.5.2 Behavioral state control: anesthesia and sleep/waking cycle 613
41.3.5.3 Micturition 614
41.3.5.4 Environmental analgesia 614
41.4 Conclusion 615
References 616

593
594 The Brainstem and Nociceptive Modulation

Glossary
cyclooxygenase An enzymeprotein complex that 12-lipoxygenase An enzyme that catalyzes the
catalyzes the production of prostaglandins from conversion of arachidonic acid to the hydroxyei-
arachidonic acid. Cyclooxygenase activity is inhib- cosenoic acid (12-HETE) structure.
ited by nonsteroidal anti-inflammatory drugs, such NSAIDs Nonsteroidal anti-inflammatory drugs.
as aspirin and ibuprofen. Drugs that inhibit cyclooxygenases and the pro-
GIRK G-protein-activated inwardly rectifying duction of prostaglandins involved in the
potassium channel. A potassium-selective channel inflammatory response (e.g., aspirin, ibuprofen,
that is gated by activated betagamma subunits of naproxen).
Gi/Go G proteins. phospholipase A2 An enzyme that catalyes the
Kv channel Voltage-gated potassium channel. formation of arachidonic acid from phospholipids.

41.1 The Periaqueductal 41.1.1 Functional Characterization of

GrayRostral Ventromedial Medulla the Periaqueductal GrayRostral
Pain-Modulating System Ventromedial Medulla Pain-Modulating
System
The idea that there are well-defined central systems
The PAG is a cell-rich region surrounding the
that selectively modulate nociception is usually traced
cerebral aqueduct in the midbrain. The RVM is
to the demonstration that electrical stimulation in the
defined functionally, rather than cytoarchitectu-
periaqueductal gray (PAG) region of rats produced
rally, and includes the nucleus raphe magnus and
potent analgesia. This phenomenon came to be called
adjacent reticular formation. Numerous behavioral
stimulation-produced analgesia when subsequent
studies have demonstrated that nonselective activa-
work confirmed this finding using quantitative tests
tion of neurons within the PAG or RVM produces
of nociception in animals, and when such stimulation a potent antinociception. Activation of PAGRVM
was shown to produce clinical analgesia in humans. output neurons must mediate this antinociception,
This was an important advance in documenting as direct excitation of either PAG or RVM neurons
that the brain itself could regulate the processing of produces antinociception and inhibition of noxious-
nociceptive information. Inspired by stimulation-pro- evoked activity of dorsal horn neurons, whereas
duced analgesia, a significant research effort led to the inactivation of either the PAG or RVM does not.
definition of a brainstem pain modulatory network The PAGRVM modulatory network is also an
with critical links in the rostral ventromedial medulla important substrate for opioid analgesia. Focal
(RVM) as well as the PAG (Figure 1). The antinoci- application of morphine or mu-opioid agonists at
ception resulting from stimulation in these structures either site is sufficient to produce antinociception
is in large part due to regulation of nociceptive pro- comparable to that resulting from systemic mor-
cessing at the level of the spinal cord. The PAGRVM phine administration (see Fields, H. L. et al., 2005
system mediates the analgesic actions of opioids, and it or Heinricher, M. M. and Morgan, M. M., 1999 for
is recruited by internal and environmental challenges. reviews).
Accumulating evidence from neuroimaging studies Along with the evidence that this system med-
supports a role for this system in top-down modula- iates the analgesic actions of exogenous and
tion of pain in humans, such as that produced by endogenous opioids, the fact that the net behavioral
placebo or shifts in attention. An overview of descend- effect of nonselective experimental activation of the
ing control, including that mediated by the PAG PAG or RVM is antinociception led to a general
RVM system, is provided in Chapter Descending view of the PAGRVM system as an analgesia
Control Mechanisms. The purpose of this chapter is system. This system is indeed activated by acute
to review the properties of pain-modulating neurons stress and opioid analgesics to inhibit spinal noci-
within the PAG and RVM, and to place this circuitry ceptive processing. However, there is now
in a behavioral context. increasing evidence that the system, especially the
 The Brainstem and Nociceptive Modulation 595

41.1.2 Pain Modulation as Part of

Adaptive Responses to Behavioral and
Physiological Challenges
One of the earliest hints that stimulation-produced
PAG
analgesia was not an experimental curiosity was the
demonstration of environmental or stress-induced
analgesia. Stress-induced analgesia refers to the fact
that any number of situations or experimental pro-
cedures that could be characterized as stressful
induce behaviorally measurable, and in some cases
quite potent, analgesia. For example, stress-induced
analgesia can be produced by electric shock, forced
RVM swim, and centrifugal rotation as well as biologically
Pain relevant threat stimuli such as odors from stressed
animals of the same species or exposure to a predator.
Analgesia is also elicited as a conditioned response to
cues that have been paired previously with noxious
or aversive events. This analgesia can be opioid or
nonopioid in nature and has been shown by a number
+ of investigators to be mediated by the PAGRVM

system (Bodnar, R. J. et al., 1980; Lewis, J. W. et al.
1980; Watkins, L. R. et al. 1982; Fanselow, M. S. 1986).
In addition to antinociception, electrical or chemical
stimulation of the PAG also evokes autonomic changes
Figure 1 Functional organization of brainstem pain- commonly associated with cardiovascular aspects of
modulating system with links in midbrain periaqueductal defense, including hypertension (in anesthetized ani-
gray (PAG) and rostral ventromedial medulla (RVM). The mals) and altered heart rate (Lovick, T. A., 1993;
PAG projects to the RVM. The RVM in turn regulates spinal
Bandler, R. et al., 2000; Morgan, M. M. and Carrive, P.,
nociceptive circuitry via a projection through the
dorsolateral funiculus to the dorsal horn. This system exerts 2001). Patients in whom deep brain electrodes were
bidirectional control, and separate populations of RVM implanted in the rostral midbrain/PAG region for relief
neurons mediate descending inhibition and descending of intractable pain often found the stimulation to be
facilitation. The PAG is reciprocally linked with forebrain distinctly disquieting, with reports of a feeling of
structures including prefrontal cortex, amygdala, and
impending doom or a desire to flee when the stimula-
hypothalamus. These substantial interconnections provide
an anatomical substrate through which emotional and tion was activated (Nashold, B. S. et al., 1969). Consistent
cognitive variables could influence nociception via the with these reports in humans, rats will work to terminate
PAGRVM system. electrical stimulation in the dorsal PAG (Kiser, R. S.
et al., 1978). Moreover, stimulation-produced antinoci-
ception in rats is accompanied by species-specific
RVM, also facilitates nociception. The RVM defense behaviors, which include immobility and
has been implicated in hyperalgesia and allodynia escape or attack (Bandler, R. and DePaulis, A., 1991;
associated with inflammation, nerve injury, acute Fanselow, M. S. 1991; Morgan, M. M. and Carrive, P.
opioid withdrawal, chronic opioid admini- 2001; Carrive, P. and Morgan, M. M. 2004).
stration, and the sickness response (see Sections Two factors contributed to the idea that antinocicep-
41.2.1 and 41.3.2, also Porreca, F. et al., 2002 and tion is recruited as part of defense behaviors: the
Heinricher, M. M. et al., 2003 for recent reviews). fact that antinociception is readily evoked by learned
The PAGRVM circuit should therefore be viewed or innate danger signals and the observation that
not specifically as an analgesia system, but more stress induces analgesia through activation of the
generally, as a pain-modulation system. From this PAGRVM system. Pain behaviors must sometimes
perspective, the system has the potential for graded be inhibited in order to give higher precedence to
enhancement or inhibition of nociception under more pressing needs such as escaping from an aggressor
different conditions. or avoiding detection by a predator. However, a more
596 The Brainstem and Nociceptive Modulation

general view of pain modulation has now developed. Distraction or more global variables such as positive
Pain inhibition is currently viewed as one component of or negative mood also alter pain (see Villemure, C. and
a number of organized responses that allow an organism Bushnell, M. C., 2002 for a review). Similarly, expecta-
to prioritize nociceptive behaviors relative to other tion that a particular manipulation will relieve pain can
internal and external demands. In addition to antinoci- lead to inhibition of pain (Price, D. D. et al., 1999).
ception, such responses typically include autonomic, Understanding the neural mechanisms through which
endocrine, and motor elements. One example of such these and other higher psychological factors modulate
an organized defense response would be antinocicep- pain has been challenging, as these variables are at best
tion as part of preparation for fight or flight when difficult to study in nonhuman subjects. Moreover,
confronted by a predator. Although the circuitry some modulation of pain perception undoubtedly
through which the PAGRVM system is brought into occurs within thalamocortical circuits. However, ima-
play in response to threat remains to be fully elucidated, ging studies in humans have now demonstrated that
inputs from the amygdala and hypothalamus are likely the PAG is activated in placebo and distraction para-
to be critical (Helmstetter, F. J., 1992; Lovick, T. A., digms. There are also changes in activation of this
1993; Lumb, B. M. et al., 2002). system and associated structures when subjects antici-
There is also evidence that the PAGRVM sys- pate pain, and in functional pain disorders (see Tracey,
tem is engaged as part of the response to general I., in press for a recent review). Studies of opioid-
immune challenge, as occurs with systemic bacterial dependent placebo have been most compelling.
infection. Thus, systemic administration of lipopoly- Petrovic P. et al. (2002) found that a placebo procedure
saccharide (LPS), a well-accepted model for bacterial (infusion of an inactive vehicle in subjects who had
infection and sickness, results in changes in nocicep- been given the powerful opiate remifentanil in a sepa-
tion mediated at least in part by the RVM (Watkins, rate trial) resulted in reduced pain reports and
L. R. et al., 1994; Romanovsky, A. A. et al., 1996). Just activation of the PAG. This region had previously
been activated by the exogenous opiate, suggesting
as antinociception is viewed as an adaptive response
that the placebo manipulation recruited endogenous
to external threat, hyperalgesia associated with sick-
opioid circuitry within the PAG. Wager T. D. et al.
ness may promote recuperative behaviors and
(2004) saw similar activation of the anterior cingulate
facilitate healing. As with the defense response to
and PAG in a conditioned placebo paradigm. These
external threat, the system is engaged during sickness
two studies suggest that recruitment of the PAG con-
to prioritize processing of nociceptive information in
tributes to the pain inhibition produced by placebo.
accord with other physiological and behavioral goals.
Similar studies investigating the effects of attention
It is likely that the PAGRVM system also med-
on pain responses also suggest that activation of the
iates the more subtle shifts in pain processing that
PAGRVM system contributes to pain suppression
occur in the absence of extreme challenge. For exam-
when attention is redirected from the noxious stimulus
ple, reflexes or other more highly organized behaviors to inputs that have greater behavioral significance
evoked by noxious stimuli would be expected to (Tracey, I. et al., 2002; Valet, M. et al., 2004). Although
interfere with feeding or other homeostatic behaviors. correlative, these imaging studies suggest that the
Nociceptive threshold is increased in hungry cats PAGRVM system plays a role in pain modulation
given access to food (Casey, K. L. and Morrow, T. J. produced by placebo, shifts in attention and presum-
1988; 1989). Moreover, there appears to be an equili- ably other cognitive and affective variables.
brium between responding to noxious inputs and the
need to maintain energy balance. Feeding is sup-
pressed in favor of pain behaviors during the first 41.2 The Periaqueductal Gray
phase of the formalin response, generally thought to
represent a relatively intense sensation. By contrast, The PAG comprises heterogeneous cell populations
pain behaviors are reduced in favor of feeding during surrounding the cerebral aqueduct. It extends rostrally
the second, less intense, phase of the formalin from the pericoerulear area of the pons to the opening
response (LaGraize, S. C. et al., 2004a). Similarly, of the third ventricle. As described above, electrical
paw withdrawal to noxious heat is attenuated during stimulation or focal application of opioids in the PAG
micturition, presumably allowing complete emptying produces behaviorally measurable antinociception.
of the bladder without interruption by noxious- However, in addition to pain modulation, it integrates
evoked movements (Baez, M. A. et al. 2005). a variety of complex functions, including
 The Brainstem and Nociceptive Modulation 597

cardiovascular and other aspects of defense, general ventrolateral and lateral PAG (Menetrey, D. et al.,
autonomic control, reproductive behaviors, and voca- 1982; Yezierski, R. P. and Mendez, C. M. 1991).
lization (Bandler, R. and Shipley, M. T., 1994). These afferents convey innocuous and noxious infor-
mation from cutaneous, musculoskeletal, and visceral
structures (Wiberg, M. et al., 1987; Yezierski, R. P.,
41.2.1 The Periaqueductal Gray and
1988; Yezierski, R. P. and Mendez, C. M., 1991;
Facilitation of Nociception
Clement, C. I. et al., 2000). Inputs from the lumbosa-
The idea that the PAGRVM system exerts bidirec- cral cord arise from a number of regions, including
tional control is now well accepted (Porreca, F. et al., those receiving afferents from pelvic and pudendal
2002; Heinricher, M. M. et al., 2003). Although the nerves (Vanderhorst, V. G. et al., 1996). These latter
focus of functional studies of descending facilitation afferents are proposed to provide important sensory
has been primarily on the RVM (see below, Section information contributing to PAG functions in sexual
41.3.2), there is some evidence that the PAG also behavior and micturition. Approximately half of the
contributes to hyperalgesia under some conditions. spinal input to the PAG is from the upper cervical
First, the PAG contributes to normal levels of spinal cord, but the significance of this cervical pre-
responsiveness on the formalin test and to enhanced dominance has not been determined (Vanderhorst,
nociceptive behaviors after foot shock, suggesting a V. G. et al., 2002).
net facilitating output from this region in these para- The most substantial afferent inputs to the PAG
digms (McLemore, S. et al., 1999; Berrino, L. et al., are from forebrain, including prefrontal and agranu-
2001). Dorsolateral PAG may be important in a gen- lar insular cortices as well as the amygdala and
eralized sensory sensitization induced by footshock hypothalamus (Bandler, R. and Shipley, M. T.,
(Crown, E. D. et al., 2004), a facilitation that is 1994; An, X. et al., 1998; Floyd, N. S. et al., 2000).
recruited in parallel with the well-documented des- Connections from medial prefrontal cortex are part
cending inhibition evoked by footshock. Second, of a medial prefrontal network associated with
focal application of capsaicin or bradykinin into the visceromotor control through links with the hypotha-
PAG has a pronociceptive effect (Burdin, T. A. et al., lamus (Ongur, D. and Price, J. L., 2000). Stimulation
1992; McGaraughty, S. et al., 2003). Third, prosta- in the anterior cingulate has been reported by
glandin release within the PAG apparently recruits different laboratories to facilitate or suppress noci-
descending facilitation, possibly under conditions of ceptive responses (Hardy, S. G., 1985; Fuchs, P. N.
inflammation or systemic infection (Vanegas, H. and et al., 1996; Calejesan, A. A. et al., 2000). Lesions of the
Tortorici, V. 2002). Thus, cyclooxygenase is consti- anterior cingulate are generally agreed to reduce noci-
tutively expressed in PAG, and direct microinjection ception in animal models (Pastoriza, L. N. et al., 1996;
of cyclooxygenase inhibitors in this region reduces Donahue, R. R. et al., 2001; LaGraize, S. C. et al., 2004b;
responding on cutaneous and visceral nociceptive in press), consistent with reports in human patients
tests. Moreover, direct application of prostaglandin (Foltz, E. L. and Lowell, E. W. 1962; Davis, K. D.
E2 in the PAG produces behavioral hyperalgesia and et al., 1994; Talbot, J. D. et al., 1995). However, the
activates RVM neurons that facilitate nociception antinociceptive effect of lesions is presumably due to
(Heinricher, M. M. et al., 2004a). Taken together, interference with cortical processing, rather than to
these findings document the ability of the PAG to activation of descending control. Effects of experi-
facilitate nociception. Consistent with this idea, a mental manipulation of the ventrolateral orbital
recent imaging study demonstrated that stimulation cortex (VLO) on nociception are similarly complex.
of an area of capsaicin-induced secondary hyperal- Stimulation of the VLO has been reported by one
gesia in human subjects was associated with group of investigators to facilitate nociception
activation of the PAG (Zambreanu, L. et al., 2005). (Hutchison, W. D. et al., 1996) and another to suppress
nociception (Zhang, Y. Q. et al., 1997). The antinoci-
ceptive effect of VLO stimulation was blocked by lesion
41.2.2 Afferents to the Periaqueductal
of the PAG, indicating that the connections from this
Gray
cortex to the PAG are relevant to nociceptive modula-
The PAG integrates information from all levels of tion (Zhang, Y. Q. et al., 1997).
the central nervous system. The spinal cord sends Hypothalamic afferents to the PAG are predomi-
direct, somatotopically organized projections from nantly from the lateral hypothalamus and the
the dorsal horn and the intermediate gray to the anterior hypothalamus/medial preoptic area (MPO)
598 The Brainstem and Nociceptive Modulation

(Shipley, M. T. et al., 1991; Semenenko, F. M. and 41.2.3 Efferents from the Periaqueductal
Lumb, B. M. 1992; Behbehani, M. M. and Da Costa Gray
Gomez, T. M., 1996). At least some of the anterior
The PAG has only a sparse projection to the spinal
hypothalamic neurons projecting to the PAG are
cord, but a dense projection to the RVM, which in turn
nociceptive (Lumb, B. M. et al., 2002; Parry, D. M.
projects to the dorsal horn via the dorsolateral funiculus.
et al., 2002). The MPO has important roles in auto- Inactivation of the RVM prevents the antinociceptive
nomic regulation including thermoregulation and effects of PAG manipulations, indicating that the
fever, as well as in sleep, mating, and maternal beha- connection from the PAG to the RVM is the neuro-
viors. Electrical stimulation of the MPO produces anatomical basis for descending modulation of
c-fos labeling throughout the PAG (Rizvi, T. A. nociception by the PAG. PAGRVM projection neu-
et al., 1996) and activates PAGRVM output neurons rons express neuropeptides, excitatory amino acids, and
(Semenenko, F. M. and Lumb, B. M., 1999). Direct serotonin. Functional studies implicate excitatory
application of prostaglandin E2 in the MPO at low amino acids, serotonin, and endogenous opioids in
doses produces hyperalgesia that is likely mediated recruitment of the RVM by the PAG to produce anti-
by the PAGRVM system (Hosoi, M. et al., 1997; nociception. A second relay through which the PAG is
Abe, M. et al., 2001; Heinricher, M. M. et al., 2004b). likely to influence spinal nociceptive processing is pon-
The central nucleus of the amygdala has massive tine catecholaminergic cell groups, as ultrastructural
reciprocal connections with the PAG (Rizvi, T. A. studies demonstrate that these neurons receive inputs
et al., 1991; da Costa Gomez, T. M. and Behbehani, from the ventrolateral PAG (see Fields, H. L. et al., 2005
M. M., 1995). The central nucleus receives inputs from for a recent review).
the basolateral nucleus, which is a major target of
cortical afferents to the amygdala. The central nucleus
of the amygdala also receives nociceptive input from 41.2.4 Columnar Organization of
the spinal cord, both directly and indirectly via the the Periaqueductal Gray
parabrachial nucleus (Burstein, R. and Potrebic, S.,
Based on both connectivity and function, the PAG
1993; Gauriau, C. and Bernard, J. F., 2004). PAG
has been subdivided into rostrocaudaully oriented
neurons respond to stimulation in the amygdala (da
columns, designated as dorsomedial (or dorsal), dor-
Costa Gomez, T. M. et al., 1996), and both stimulation
solateral, lateral, and ventrolateral (Carrive, P. and
and injection of morphine into the amygdala result in
Morgan, M. M., 2004). These columns roughly cor-
antinociception that is dependent on activation of the
respond with subdivisions based on cytoarchitecture
descending pathway from PAG to RVM (Pavlovic, Z.
that were proposed previously by Beitz A. J. (1985)
W. et al., 1996; Helmstetter, F. J. et al., 1998; and Beitz A. J. and Shepard R. D. (1985). Cortical,
McGaraughty, S. and Heinricher, M. M., 2002; 2004). hypothalamic, and spinal afferents show a preferen-
Afferents to the PAG from the brainstem arise tial distribution to different columns, although there
from the medulla, including the RVM. Other brain- is substantial spread across columnar boundaries
stem sources of inputs to the PAG include the (Rizvi, T. A. et al., 1992; An, X. et al., 1998; Floyd, N.
nucleus tractus solitarius, adjacent nucleus cuneifor- S. et al., 2000). Projections to the RVM arise in the
mis, pontine reticular formation, and the locus dorsomedial, lateral, and ventrolateral columns, but
coeruleus and other catecholaminergic nuclei not the dorsolateral column (Abols, I. A. and
(Beitz, A. J., 1982; Herbert, H. and Saper, C. B., 1992). Basbaum, A. I., 1981; Van Bockstaele, E. J. et al., 1991).
The strong input to the PAG from forebrain and Pain modulation is not constrained by columnar
hypothalamus provides an anatomical substrate boundaries, and electrical stimulation or microinjec-
through which emotional and cognitive variables tion of mu-opioid agonists or neuroexcitant agents
could influence pain via the PAG. Information throughout the dorsoventral extent of the PAG pro-
related to motivation and emotion thus converges duces analgesia, particularly at more caudal levels
with direct and indirect visceral and somatic afferent (Waters, A. J. and Lumb, B. M., 1997 Carrive, P. and
input in the PAG. This convergence is presumably Morgan, M. M., 2004; Morgan, M. M. and Clayton, C.
important to pain modulation in allowing the organ- C. 2005). Antinociception resulting from stimulation in
ism to regulate nociceptive processing in accord with the ventrolateral PAG must be mediated by an endo-
current needs and motivational state, particularly in genous opioid connection, as it is blocked by naloxone.
response to environmental or internal challenge. By contrast, the antinociception resulting from
 The Brainstem and Nociceptive Modulation 599

electrical stimulation more dorsally is not mediated by interspersed with periods of inactivity during anti-
endogenous opioids (Morgan, M. M. 1991). nociception evoked from the ventrolateral column
Antinociception evoked by stimulation of the dor- (Vianna, D. M. et al., 2001; Morgan, M. M. and
sal and lateral columns is often associated with Clayton, C. C., 2005). As in the lateral/dorsolateral
escape-like behaviors or defensive posturing, hyper- columns, the current needed to elicit freezing with
tension, and tachycardia. By contrast, antinociception stimulation in the ventrolateral PAG is lower than
produced by stimulation of the ventrolateral PAG is that required to elicit jumping (Vianna, D. M. et al.,
often accompanied by immobility, bradycardia, and 2001). Moreover, although lesions of the ventrolat-
in anesthetized animals, hypotension (Depaulis, A. eral PAG reduce freezing and release running
et al., 1994; Morgan, M. M. and Carrive, P., 2001). behavior in response to threat (Walker, P. and
These findings gave rise to the proposal that the Carrive, P., 2003, Farook, J. M. et al., 2004), lesions
lateral/dorsolateral columns function specifically in of the dorsolateral PAG have no effect on either
active coping, particularly when confronted with an freezing or running evoked by exposure to a cat or
external threat. By contrast, the ventrolateral column to a context associated with footshock. In addition,
was hypothesized to be important in passive coping changes in heart rate and activity in this paradigm are
and recuperation, for example, in response to visceral seen only when the animal is denied access to a safe
pain or some other challenge that cannot be con- hiding place (Dielenberg, R. A. et al., 2004; Farook,
trolled or escaped (Keay, K. A. and Bandler, R., 2001). J. M. et al., 2004; Leman, S. et al., 2003). Thus, neither
Two aspects of this framework for PAG organiza- active fight or flight behaviors nor immobility
tion have been controversial. The first issue is the appears to be a specific output of a particular column.
interpretation of immobility evoked by stimulation in These observations suggest that although the idea
the ventrolateral column. It is not possible to deter- that the lateral/dorsolateral and ventral columns of
mine from lack of movement alone whether an the PAG represent centers for active and passive
animal should be viewed as engaging in recuperative coping has had great heuristic value, a full under-
behaviors or whether it is instead freezing, that is, standing of the functional significance of the
exhibiting an active cryptic defense that reduces the PAG columns is likely to require sophisticated ana-
likelihood of detection by a predator (Blanchard, R. J. lyses of the neural circuitry of these regions in
et al. 1986; Fanselow, M. S., 1991; Bittencourt, A. S. relevant behavioral paradigms.
et al., 2004). The argument that immobility represents
passive coping rather than a defensive response is
41.2.5 Intrinsic Circuitry and
based largely on observations that stimulation in the
Neurotransmitters in the Periaqueductal
ventrolateral column in anesthetized animals pro-
Gray
duce hypotension (Keay, K. A. and Bandler, R.,
2001). However, hypotension is not evoked by such 41.2.5.1 Endogenous opioids
stimulation in awake behaving animals (Morgan, M. All three opioid receptors, mu (MOR), delta (DOR),
M. and Carrive, P., 2001). Furthermore, hypotension and kappa (KOR) opioid receptors, are moderately
may not imply a quiescence or recuperative state. For to densely expressed in the PAG (Mansour, A. et al.,
example, although the freezing in a conditioned fear 1995; Gutstein H. B. et al., 1998; Kalyuzhny, A. E. and
paradigm is associated with hypertension (Carrive, P. Wessendorf, M. W., 1998). The PAG is also rich
2000), defensive freezing to various threat stimuli can in endogenous opioids (Mansour, A. et al., 1995).
be associated with profound hypotension in various Enkephalin-like immunoreactivity is most dense in
species of wild-trapped rodents (Hofer, M. A., 1970). the ventrolateral PAG and enkephalin-containing
A second contentious issue is whether active terminals are found apposed to both gamma amino-
offensive and escape behaviors should be associated butyric acid (GABA)ergic and non-GABAergic
specifically with the lateral/dorsolateral columns, dendrites, including those of a small percentage of
whereas immobility is a function of the ventrolateral PAGRVM neurons (Williams, F. G. and Beitz, A. J.,
column. Immobility as well as escape can be evoked 1990). Endomorphin-2 (Tyr-Pro-Phe-Phe-NH2), an
with stimulation of the lateral/dorsolateral columns. endogenous peptide with high selectivity for the
Notably, the current needed to induce immobility is MOR (Zadina, J. E. et al., 1997), is concentrated in
lower than that required for active flight behaviors the PAG as well as the dorsal horn of the spinal cord
with stimulation in this column (Bittencourt, A. S. (Schreff, M. et al., 1998). Although the endomorphins
et al., 2004). Conversely, bursts of running are often have a high affinity for the MOR, it appears that these
600 The Brainstem and Nociceptive Modulation

peptides are only partial agonists for the MOR in the A. et al., 1987). Also consistent with this hypothesis
PAG (Narita, M. et al., 2000). Another endogenous are observations that MOR1 is frequently expressed
opioid found in the PAG is -endorphin. Discrete in GABAergic neurons within the PAG. However, a
populations of -endorphin-containing neurons in substantial subset of PAGRVM projection neurons
the ventromedial hypothalamus project to the PAG do express MOR1 (Kalyuzhny, A. E. and Wessendorf,
and have been implicated in analgesia produced by M. W., 1998; Commons, K. G. et al., 2000). These
electrical stimulation and stress (Millan, M. J., 2002). MOR1-positive neurons may target nociceptive
MOR agonists produce potent antinociception facilitating neurons in the RVM (Vanegas, H. et al.,
when applied directly in the PAG (Heinricher, M. 1984; Cheng, Z. F. et al., 1986; Morgan, M. M. et al.,
M. and Fields, H. L., 2003). The behavioral antinoci- 1992;). If so, these MOR1-positive output neurons
ception produced by these agents is mediated by would be involved in descending facilitation rather
activation of output neurons projecting to the RVM than descending inhibition.
(Sandkuhler, J. and Gebhart, G. F., 1984; Tortorici, V. Postsynaptic effects of MOR agonists on PAG
and Morgan, M. M., 2002). However, as the direct neurons include hyperpolarization (via activation of
postsynaptic action of MOR agonists on PAG neu- a postsynaptic G-protein-activated inwardly rectify-
rons is hyperpolarization (Chieng, B. and Christie, M. ing potassium conductance, GIRK) and inhibition of
J., 1994a; Osborne, P. B. et al., 1996), activation of the calcium channels (Connor, M. and Christie, M. J.,
output neurons must be via disinhibition. MOR ago- 1998) (Figure 2). MOR agonists also have presynaptic
nists are thought to act presynaptically to block effects, inhibiting GABA and glutamate release from
GABAergic inhibition of PAG output neurons. terminals within the ventrolateral PAG (Chieng, B.
Consistent with this hypothesis, blockade of GABA and Christie, M. J., 1994b; Vaughan, C. W. and
transmission within the PAG by microinjection of a Christie, M. J. 1997; Vaughan, C. W. et al., 1997).
GABAA receptor antagonist produces antinocicep- Presynaptic inhibition of GABAergic neurotransmis-
tion (Moreau, J. L. and Fields, H. L., 1986; Depaulis, sion is through activation of the arachidonic acid

Figure 2 Cellular mechanisms of opioid action within the periaqueductal gray (PAG). Enkephalin-containing synapses are
apposed to cell bodies as well as to GABA- and glutamate-containing terminals. The postsynaptic mu-uopoid receptor (MOR)
activates G-protein-activated inwardly rectifying potassium channels (GIRKs) and inhibits voltage-gated Ca2 channels to
hyperpolarize cells and decrease cell activity. Presynaptic MORs inhibit both GABA and glutamate release on ventrolateral
PAG neurons, and apparently use different signal transduction pathways in the terminals. MORs localized to GABA terminals
are coupled to voltage-gated potassium channels via activation of the arachidonic acid/12-lipoxygenase (12-LOX) second
messenger pathway. Hyperpolarization of the terminal decreases GABA release. The signal transduction pathway for MOR
inhibition of glutamate release is currently unknown. MORs are found on GABA containing interneurons in the PAG, as well as
on PAG output neurons projecting to the rostral ventromedial medulla (RVM). Activation of the descending antinociceptive
pathway by opioids occurs via disinhibition. 12-HETE, 12-hydroxyeicosenoic acid; Kv, voltage-gated potassium; PLA2,
phospholipase A2.
 The Brainstem and Nociceptive Modulation 601

phospholipase A2 second messenger pathway. to test the behavioral effects of DOR- and KOR-
Stimulation of this pathway results in activation of selective agonists in the PAG in the mouse.
voltage-gated potassium channels (Kv channels) by
metabolites of 12-lipoxygenase (Vaughan, C. W. and 41.2.5.1.(i) Opioid/nonsteroidal anti-inflammatory
Christie, M. J., 1997; Vaughan, C. W. et al., 1997). This drugs interactions Microinjections of nonsteroidal
pathway is independent of adenylyl cyclase, protein anti-inflammatory drugs (NSAIDs) into the PAG
kinase A or protein kinase C activity (Vaughan, C. W. produce analgesia (Tortorici, V. and Vanegas, H.,
et al., 1997). Further research is needed to determine 1995). This antinociception is apparently mediated
the relevance of the various presynaptic versus post- at least in part by an endogenous opioid peptide, as
synaptic opioid actions to the nociceptive modulatory the analgesia produced by NSAIDs microinjected
function of the PAG. As already noted, there is good into the PAG and RVM is attenuated by the opioid
evidence that the behavioral antinociception pro- antagonist naloxone (Vanegas, H. and Tortorici, V.,
duced by local application of MOR agonists involves 2002).
activation of the PAGRVM output neurons via dis- In addition to stimulating release of endogenous
inhibition. Whether postsynaptic inhibition or the opioids, recent evidence at the cellular level suggests
suppression of glutamatergic transmission plays a that NSAIDs may also augment the signaling pathway
role in antinociception is unknown. However, at used by opiates, potentiating the actions of exogenous
least in the PAG slice, the effect of presynaptic inhi- opioids (Figure 3). Coapplication of NSAIDs potenti-
bition of GABAergic transmission apparently ates the inhibition of GABA release by the MOR
predominates, because bath application of the MOR partial agonist morphine, although NSAIDs have no
agonist [D-Ala2, N-Met-Phe4, Gly-015] enkephalin effect on GABA release in the absence of morphine
(Vaughan, C. W. et al., 1997). NSAIDs primarily inhibit
(DAMGO) results in activation of neurons that are
cyclooxygenases (COX-1 and COX-2), one of three
also directly hyperpolarized by the opioid (Chiou, L.
types of enzymes (cyclooxygenases, 5-lipoxygenases,
C. and Huang, L. Y., 1999). Thus, under in vitro
and 12-lipoxygenases) that metabolize arachidonic
conditions, disinhibition has a relatively large impact,
acid. One hypothesis proposed to explain the mechan-
and the net effect of MOR agonist administration is
ism of increased analgesia with coapplication of
neuronal activation. However, it is not known
opioids and NSAIDs is that blockade of COX-1 shunts
whether, or under what conditions, endogenous
arachidonic acid metabolism through the lipoxygenase
opioids are released to act simultaneously on presy-
pathways to increase the potency of opioid receptor
naptic and postsynaptic receptors. agonists (Vaughan, C. W. et al., 1997; Vaughan, C. W.
In general, activation of DOR and KORs in the 1998; Christie, M. J. et al., 1999). The fact that inhibi-
PAG does not result in significant antinociception, at tors of 5-lipoxygenases also appear to potentiate the
least in rats. A number of groups have found no effects of opioid agonists in the PAG adds further
analgesic effect of microinjection of DOR agonists weight to this proposal (Vaughan, C. W. et al., 1997;
in the PAG in rats (Bodnar, R. J. et al., 1988; Smith, Christie, M. J. et al., 1999). These results are important
D. J. et al., 1988; Ossipov, M. H. et al., 1995), although in that the combined administration of NSAIDs and
there is one report that microinjection of the delta2 opiates may allow lower doses of morphine to be used
agonist deltorphin into the PAG produces a modest to provide adequate analgesia while reducing the
increase in latency of the tail flick response (Rossi, G. probability of the development of tolerance and side
C. et al., 1994). KOR agonists are also ineffective in effects (such as respiratory depression) associated with
producing analgesia in the PAG (Fang, F. G. et al., high doses of opiates. Functional studies of NSAID/
1989). At the cellular level, DOR and KOR agonists opioid interactions in the PAG would therefore be of
have no effect on presynaptic GABA release and in great interest.
rat do not activate postsynaptic potassium channels
(Vaughan, C. W. and Christie, M. J., 1997). However, 41.2.5.1.(ii) Opioid tolerance and dependence
in mice, KOR agonists inhibit presynaptic GABA The PAG has been implicated in opioid tolerance
release in the PAG, and agonists of all three opioid as well as dependence. Opioid tolerance is the dimin-
receptor subtypes activate postsynaptic potassium ished responsiveness to the antinociceptive actions of
channels (Vaughan, C. W. et al., 2003). The functions opioids with chronic administration. Dependence
of the three receptors may therefore be different in refers to the occurrence of withdrawal signs and/or
the two species, and it would obviously be of interest rebound responses upon removal of the opioid or
602 The Brainstem and Nociceptive Modulation

GABA terminal

Acute morphine Chronic morphine

MOR
MOR

AC
+
AC
AC
AC AC
2
A

NSAIDs AA
PL

12-LOX

+ cAMP
COX + PKA
+

Kv

K+

MOR agonists GABA release MOR agonists MOR potent

GABA release

Figure 3 Mu-opioid receptor (MOR) coupling in presynaptic GABA terminals changes with chronic morphine
administration. Acute administration of MOR agonists activates MORs coupled to phospholipase A2 (PLA2). Activation of
PLA2 increases production of arachidonic acid, which is further metabolized by 12-lipoxygenase (12-LOX). Lipoxygenase
metabolites such as 12-HETE activate voltage-gated potassium channels (Kv channels) to hyperpolarize and decrease GABA
release from the terminals. Nonsteroidal anti-inflammatory drugs (NSAIDs) potentiate this action of opioids by inhibiting
cyclooxygenase (COX)-mediated arachidonic acid metabolism, thereby shunting arachidonic acid to the 12-LOX pathway.
Activation of MORs presumably also acutely inhibits adenylyl cyclase (AC) activity in these terminals. Chronic morphine
administration upregulates AC and protein kinase A (PKA) activity. After chronic, but not acute, opioid treatment, GABA
release is enhanced by increased PKA activity. MOR agonists are more potent inhibitors of this PKA-dependent release, so
that opioid removal or blockade of MORs by antagonists results in a rebound increase in GABA release. This increased GABA
release may contribute to withdrawal behaviors mediated by the PAG.

administration of an opioid antagonist. Animals exhi- (Jacquet, Y. F. and Lajtha, A., 1976; Siuciak, J. A.
bit tolerance to the antinociceptive actions of MOR and Advokat, C., 1987). Moreover, blockade of PAG
agonists when morphine or MOR agonists are opioid receptors using local microinjection of the
applied directly in the PAG, especially in the caudal opioid antagonist naltrexone prevents the develop-
ventrolateral aspect (Morgan, M. M. et al., 2005). This ment of tolerance to systemically administered
tolerance is not due to associative mechanisms, as morphine (Lane, D. A. et al., 2005). Although RVM
continuous administration via an implanted pump neurons lose their responsiveness to PAG morphine
leads to tolerance in the absence of cues associated administration in tolerant animals (Tortorici, V. et al.,
with drug administration (Lane, D. A. et al., 2004). 2001), tolerance does not develop as readily when
The PAG is apparently critical for the development morphine is microinjected into the RVM itself
of antinociceptive tolerance when opioids are given (Morgan, M. M. et al., 2005), and blockade of the
systemically because animals chronically treated RVM does not interfere with the development of
with morphine microinjected into the PAG display tolerance when morphine is applied directly in the
cross-tolerance with morphine given systemically PAG (Lane, D. A. et al., 2005). In addition, animals do
 The Brainstem and Nociceptive Modulation 603

not develop behavioral tolerance to direct activation be the mechanism of the increased firing rate of these
of PAG output neurons that mediate antinociception, neurons during withdrawal (Bagley, E. E. et al., 2005).
for example by microinjecting kainic acid (Morgan, These latter observations would argue that uncou-
M. M. et al., 2003). These latter two findings demon- pling of MOR from pre- or postsynaptic effectors
strate that tolerance is not a result of adaptations does not underlie antinociceptive tolerance in the
downstream from the opioid-sensitive PAG neuron, PAG. Rather, compensatory mechanisms may mask
and indicate that the search for molecular and cellu- an underlying increase in the coupling of presynaptic
lar mechanisms of opioid tolerance should focus on MOR to its effectors that is revealed upon removal of
these opioid-sensitive neurons. the opioid, leading to withdrawal behaviors. Further
The idea that the PAG has a role in opioid depen- work will be needed to bridge the gap between the
dence derives primarily from observations that cellular and behavioral analyses of tolerance and
blockade of PAG opioid receptors produces a num- dependence.
ber of withdrawal signs in tolerant animals, and that
chronic administration of morphine in the PAG leads 41.2.5.2 Norepinephrine
to the development of dependence (Bozarth, M. A. The PAG has reciprocal connections with pontomed-
1994). In addition, precipitated withdrawal is asso- ullary catecholaminergic cell groups (Herbert, H. and
ciated with increased expression of c-fos throughout Saper, C. B. 1992; Bajic, D. et al., 2001).
the PAG, especially in the ventrolateral and lateral Norepinephrine has multiple effects on PAG neu-
aspects (Chieng, B. et al., 1995). A substantial number rons that are mediated by 1- and 2-adrenergic
of neurons expressing c-fos after precipitated with- receptors. Activation of 1-receptors depolarizes all
drawal are GABAergic (Chieng, B. et al., 2005), and PAG neurons, whereas 2-receptor activation hyper-
neurons positive for c-fos do not project to the RVM polarizes all neurons, suggesting that both receptors
(Bellchambers, C. E. et al., 1998). Thus, as was the are colocalized on PAG neurons (Vaughan, C. W.
case with tolerance, the locus for withdrawal signs in et al., 1996). However, most neurons preferentially
the PAG appears to be upstream from the output display either a hyperpolarizing or depolarizing
neuron projecting to the RVM. response to norepinephrine itself. Neurons in ven-
Studies of the membrane properties of PAG neurons trolateral PAG were more likely to be depolarized
following chronic opioid treatment and withdrawal (85%), whereas lateral PAG neurons were split
document a host of changes that could contribute to equally in exhibiting depolarizing and hyperpolariz-
tolerance and/or dependence (Williams, J. T. et al., ing responses. The 2-mediated hyperpolarization is
2001; Bailey, C. P. and Connor, M., 2005). These stu- due to activation of a potassium conductance while
dies have emphasized altered effects of opioids on the 1-mediated depolarization depends on inhibi-
GABAergic inhibition and potassium channels. tion of a potassium conductance and an unidentified
Opioid tolerance is widely assumed to involve a norepinephrine-sensitive conductance (Pan, Z. Z.
functional uncoupling between opioid receptors and et al., 1994; Vaughan, C. W. et al., 1996).
their effectors. The decreased ability of opioids to The 2-adrenoceptor agonist clonidine also sup-
inhibit the firing of PAG neurons after chronic mor- presses GABAergic synaptic transmission, and this
phine treatment is consistent with this idea (Bagley E. suppression is enhanced after chronic morphine
E. et al., 2005; Chieng, B. and Christie, M. D., 1996). treatment (Ingram, S. L. et al., 1998). This action of
However, MOR activation of GIRK channels in clonidine may be related to its ability to suppress
PAG is not reduced in animals subjected to repeated many of the signs of the opioid withdrawal syndrome
intermittent morphine administration that is suffi- (Christie, M. J. et al., 1997).
cient to induce antinociceptive tolerance (Ingram, S.
L. et al., 2007), and the ability of MOR agonists to 41.2.5.3 Substance P
inhibit GABA release is potentiated rather than NK1 receptors are found in the PAG (Commons, K.
reduced in slices from animals treated with chronic G. and Valentino, R. J., 2002), and substance P acti-
morphine. These enhanced actions of MOR agonists vates both opioid-sensitive and opioid-insensitive
are mediated by upregulation of the protein kinase A neurons in this region (Ogawa, S. et al., 1992; Drew,
pathway (Ingram S. L. et al., 1998; Hack, S. P. et al., G. M. et al., 2005). In primary afferents and the dorsal
2003). Moreover, recent findings demonstrate that horn, substance P has long been associated with
induction of a GABA transporter-mediated cation nociceptive transmission, and noxious stimulation
current in opioid-sensitive neurons in the PAG may also increases the release of substance P in the
604 The Brainstem and Nociceptive Modulation

PAG. This observation would suggest that substance Forebrain structures and the hypothalamus/MPO
P plays a role in nociceptive transmission in the PAG thus influence the RVM through the PAG, as well
as in the dorsal horn. However, local morphine as by means of less dense connections directly to the
administration also increases substance P release in RVM itself. In addition, RVM neurons are likely to
the PAG, and microinjection of exogenous substance receive spinothalamic inputs, either through direct
P in this region produces significant antinociception connections to their widespread dendritic arbors or
(Xin, L. et al., 1997; Rosen, A. et al., 2004). The role of relayed via other brainstem regions such as nucleus
substance P in nociceptive transmission or modula- reticularis gigantocellularis or the PAG (see Fields,
tion thus remains to be determined. Substance P has H. L. et al., 2005, for a review.)
also been implicated in defensive behaviors and In addition to a direct projection to the dorsal
aggression, and it produces a conditioned place aver- horn, the RVM can apparently influence spinal noci-
sion when microinjected into the dorsolateral PAG ceptive processing via catecholamine cell groups in
(Aguiar, M. S. and Brandao, M. L., 1994). These the pons, particularly the A7 cell group. Although
results point to a more general role for this peptide there are no noradrenergic neurons within the RVM,
in defense. Thus substance P may apparently influ- antinociception produced by electrical or chemical
ence a number of functions in the PAG, and stimulation in this region is frequently attenuated
functional studies using antagonists are needed to or blocked by intrathecal administration of noradren-
determine the role of endogenous substance P in ergic antagonists (Hammond, D. L. and Yaksh, T. L.,
nociceptive processing and modulation. 1984; Proudfit, H. K., 1992). This indicates that
part of the antinociceptive effect of RVM stimulation
41.2.5.4 Cannabinoids is relayed through one of the descending catechola-
Cannabinoids have been used for centuries to provide minergic pathways, most likely the A7 cell group
pain relief (Walker, J. M. and Huang S. M., 2002a) but in the mesopontine tegmentum. The RVM sends
are of limited utility today because of the significant connections to the A7 region (Clark, F. M. and
psychoactive side effects and illegal status in the United Proudfit, H. K., 1991), and these axons originate
States. CB1 receptors are dense throughout the PAG from a population of neurons distinct from that pro-
(Herkenham, M. et al., 1991; Tsou, K. et al., 1998). jecting to the dorsal horn (Buhler, A. V. et al., 2004).
Cannabinoids, like opioids, can mediate antinociception This connection to A7 allows the RVM to engage
by activating the PAGRVM descending pathway (see spinally projecting noradrenergic neurons, which
Iversen, L. and Chapman, V., 2002 and Walker, J. M. and would parallel the direct projections from the RVM
Huang, S. M., 2002b for reviews). Endocannabinoids, to the dorsal horn.
like endogenous opioids, contribute to the antinocicep- The primary outputs from the RVM are thus
tive effect of activating the PAGRVM system (Walker, descending projections to the dorsal horn, both
J. M. et al., 1999; Hough, L. B. et al., 2002), and endocan- directly and via the mesopontine tegmentum.
nabinoids in the PAG were recently shown to Ascending connections have also been demonstrated
contribute to stress-induced analgesia (Hohmann, A. anatomically (Zagon, A. et al., 1994; Hermann, D. M.
G. et al., 2005). Activation of CB1 receptors inhibits et al., 1996), but whether these rostral projections play
both GABA and glutamate release via presynaptic a role in nociceptive modulation is unknown.
mechanisms. However, cannabinoids have negligible
postsynaptic actions on PAG neurons (Vaughan, C. W.
41.3.2 The Rostral Ventromedial Medulla
et al., 2000).
and Facilitation of Nociception
Although the most robust effect of nonselective
41.3 The Rostral Ventromedial experimental activation of the RVM is antinocicep-
Medulla tion, low-intensity electrical stimulation of the RVM
and infusions of neuropeptides and N-methyl-
41.3.1 Connections of the Rostral
D-aspartic acid (NMDA) in this region have been
Ventromedial Medulla
shown by various groups to facilitate nociceptive
The RVM can be viewed as the brainstem output processing (Zhuo, M. and Gebhart, G. F., 1992;
from the PAGRVM system, receiving a dense Urban, M. O. and Smith, D. J., 1993; Smith, D. J.
innervation from the PAG and projecting to the et al., 1997; Zhuo, M. and Gebhart, G. F., 1997;
dorsal horn through the dorsolateral funiculus. Kovelowski, C. J. et al., 2000; Friedrich, A. E. and
 The Brainstem and Nociceptive Modulation 605

Gebhart, G. F., 2003; Heinricher, M. M. and Neubert, some inhibited, and some unresponsive. The
M. J., 2004; Neubert, M. J. et al., 2004). Inactivation emphasis of most investigators at that time was on
studies demonstrate that hyperalgesia produced by the subset of neurons excited by noxious inputs, at
electrical stimulation is not merely an experimental least in part because of the idea that pain inhibits
curiosity: lesions implicate the RVM in a variety of pain. However, the notion of recruitment of the pain-
models of hyperalgesia and persistent pain, including inhibiting output from the RVM by noxious stimula-
sickness, acute opiate withdrawal, chronic opioid- tion was difficult to reconcile with the observation
induced hyperalgesia, Freunds adjuvant, and mus- that noxious-evoked firing of these neurons was sup-
tard oil inflammatory hyperalgesia, following pressed by opioids given in doses thought sufficient
noxious stimulation of a remote body part, and in to produce analgesia (Anderson, S. D. et al., 1977;
neuropathic pain models (Kaplan, H. and Fields, H. Gebhart, G. F., 1982; Heinricher, M. M. and
L., 1991; Morgan, M. M. et al., 1994; Watkins, L. R. Rosenfeld, J. P., 1985). The on/off/neutral cell clas-
et al., 1994; Pertovaara, A. et al., 1996; Ren, K. and sification, introduced by Fields, H. L. et al. (1983a),
Dubner, R., 1996; Urban, M. O. et al., 1996; Mansikka, was a significant advance in that it provided a defined
H. and Pertovaara, A., 1997; Wiertelak, E. P. et al., framework for hypotheses relating RVM neuronal
1997; Terayama, R. et al., 2000; Porreca, F. et al., 2001; firing to the pain-modulating function of the region.
Vanderah, T. W. et al., 2001a; 2001b). This approach to RVM physiology was based on
These inactivation studies clearly demonstrate a the recognition that pain-modulating neurons should
role for descending facilitation in a variety of discharge in relationship to variables implicated in
enhanced pain states, including inflammation. pain modulation, rather than in response to noxious
Nevertheless, there is evidence for an increase in stimulation. Thus, pain-inhibiting neurons should
descending inhibition from the RVM during inflam- be activated by analgesic drugs, and during periods
mation (Ren, K. and Dubner, R., 2002), with of suppressed nociceptive responsiveness, such as
potentiation of the analgesic effectiveness of opioids that induced by intense stress. Conversely, firing of
in the RVM (Hurley, R. W. and Hammond, D. L., pain-facilitating neurons should be suppressed
2000; 2001). One intriguing possibility is that inflam- during analgesia, and enhanced during hyperalgesia
mation increases the number of RVM neurons (see Fields, H. M. and Heinricher, M. M., 1985 for an
expressing MOR (Zhang, L. et al., 2004). Given both early discussion of the implications of the on/off/
the clear demonstration that descending facili- neutral cell classification).
tation contributes to the hyperalgesia associated In the on/off/neutral cell framework, RVM neu-
with inflammation and the apparently contradictory rons are recorded in lightly anesthetized rats so that
evidence for increased descending inhibition, an cell firing can be related to changes in nociceptive
important question is whether different aspects of responding, as measured by spinal nocifensor
nociceptive transmission are differentially affected reflexes. Off-cells are characterized by a cessation of
by inhibition and facilitation during inflammation firing during nociceptive reflexes (Figure 4). On-cells
(Vanegas, H., 2004; Vanegas, H. and Schaible, are defined by a burst of activity during nociceptive
H.-G., 2004). reflexes (Figure 4). Onset of the on-cell reflex-related
burst and off-cell pause precede the reflex itself, as
well as the thalamic response to the noxious stimulus
41.3.3 The Neural Basis for Bidirectional
(Hernandez, N. et al., 1989). Because nociceptive
Control from the Rostral Ventromedial
reflexes are generally suppressed when RVM neu-
Medulla: On- and Off- Cells
rons are experimentally activated using electrical
41.3.3.1 Physiological classification of stimulation or glutamate microinjection, the fact
rostral ventromedial medulla neurons that off-cell activity ceases abruptly just prior to the
based on reflex-related activity execution of nociceptive reflexes suggested that off-
Initial electrophysiological approaches to the func- cells are the antinociceptive output neurons of the
tion of the RVM focused on the responses of RVM RVM, and that the pause in firing permits responses
neurons to noxious stimulation (Anderson, S. D. et al., to occur. Conversely, because on-cells are active
1977; Behbehani, M. M. and Pomeroy, S. L., 1978; during the animals response to a noxious stimulus,
Guilbaud, G. et al., 1980; Heinricher, M. M. and it seemed unlikely that they exert a significant
Rosenfeld, J. P., 1985). In these experiments in deeply inhibitory effect on nociception. Rather, the reflex-
anesthetized animals, some neurons were excited, related burst suggested that these neurons have
606 The Brainstem and Nociceptive Modulation

Off-cell On-cell

Heat
Computer
controller

Figure 4 Two cell populations in the rostral ventromedial medulla (RVM), on- and off-cells, explain descending inhibition
and facilitation from the RVM. Bottom: Experimental setup shows simultaneous brainstem recording and reflex testing. Top:
Single oscilloscope sweeps show activity of a representative off- and on-cell during a tail heat trial. Upper trace is cell
discharge, lower trace shows tail position, with arrowhead indicating tail movement. Heat is applied starting at the beginning
of the trace. Each trace is 10 s.

a pronociceptive role. Both hypotheses have subse- stimulus at just threshold intensity (Fields, H. L.
quently received ample confirmation in experiments et al., 1983a; Jinks, S. L. et al., 2004a), whereas pinch
using selective pharmacological manipulation of the stimuli in work published to date have not been
different cell classes (see Section 41.3.3.2). At least limited to withdrawal threshold in intensity, and
some cells of both classes project to the dorsal horn have not been linked to a withdrawal reflex (Leung,
(Fields, H. L. et al., 1995). C. G. and Mason, P., 1998; Ellrich, J. et al., 2000; 2001).
The remaining cells in the RVM are classified as On- and off-cell firing is not constant over time. In
neutral cells because they show no change in activity animals lightly anesthetized with barbiturates, on-
associated with nociceptive reflexes. Whether neutral and off-cells frequently show alternations between
cells have a role in pain modulation remains to be periods of silence and spontaneous discharge.
determined (see Section 41.3.3.3). Simultaneous recordings from pairs of on- and off-
Noxious pinch typically activates on-cells and cells show that excitability within each class varies
suppresses off-cell firing. Cutaneous application of across the population, but that firing within each class
mustard oil, which produces an acute inflammation, is in phase. Consequently, a subset of either the on-
evokes a strong activation of on-cells and suppresses or off-cell class is active at any given time (Barbaro,
off-cell firing, leaving neutral cells unaffected N. M. et al., 1989), but the two populations are not
(Kincaid, W. et al., in press). Occasional discrepancies active simultaneously. By contrast, serotonergic neu-
between responses to heat and pinch are most likely rons (which can be considered a specific division of
explained by the fact that the heat-related classifica- neutral cells (Gao, K. et al., 1997; Mason, P., 1997;
tion is based on the reflex response evoked by a Gao, K. et al., 1998)) display regular, more or less
 The Brainstem and Nociceptive Modulation 607

constant activity. Nonserotonergic neutral cells vary body region are correlated with on-cell activation
in their firing, but are often continuously active for (Bederson, J. B. et al., 1990; Morgan, M. M. and
prolonged periods. If a withdrawal reflex is evoked Fields, H. L., 1994). On-cells also facilitate escape
when on-cells are in an active period, there is little or responses to intense noxious stimulation, and probably
no discernible increase in firing (Barbaro N. M. et al., contribute to secondary hyperalgesia during acute
1986). Because on-cell firing apparently approaches inflammation ( Jinks, S. L. et al., 2004b; Kincaid, W.
this ceiling during active phases, definitive classifica- et al., in press). Finally, on-cells have been implicated
tion of a neuron as a neutral cell rather than an on- in hyperalgesia associated with the sickness response.
cell requires extensive characterization, with reflex Hyperalgesia produced by systemic administration of
trials delivered during periods of low spontaneous bacterial endotoxin is blocked by lesion of the RVM
activity. Indeed, Barbaro N. M. et al. (1986) noted (Watkins, L. R. et al., 1994; Wiertelak, E. P. et al., 1997).
that earlier work from the Fields group had likely Although on-cells have not been studied in animals
misclassified a number of on-cells as neutral cells. challenged with endotoxin, these neurons are acti-
On-, off-, and neutral cell classes can be identified vated by direct administration of prostaglandin E2 in
in awake behaving animals (Oliveras, J. L. et al., 1990; the MPO (Heinricher, M. M. et al., 2004b), which can
1991a; 1991b; Leung, C. G. and Mason, P., 1999; Foo, be considered a model for central components of the
H. and Mason, P., 2005). However, on-cells are more sickness response (Oka, T. et al., 1994; Hosoi, M. et al.,
active during waking than sleep, firing in association 1997).
with the animals movements. Neurons of this class The obvious limitation of these observations is
are also generally more responsive when animals are that they are correlative. The RVM plays an impor-
awake, and more easily activated by innocuous stim- tant role in a number of functions other than pain
ulation than when the animals are anesthetized or modulation, and many neurons in this region are
asleep. Off-cells are more active when the animal is known to exhibit correlations with various physiolo-
sleeping, and are inhibited in association with move- gical parameters including EEG, blood pressure, and
ment in awake animals (Leung, C. G. and Mason, P., body temperature. Causal inferences relating on- or
1999; Foo, H. and Mason, P., 2005). off-cell discharge to altered nociceptive processing
thus require selective experimental manipulation of
41.3.3.2 Role of on- and off-cells in each cell class as a whole, for example, by microinjec-
pain modulation tion of drugs that differentially alter the firing of the
Initial functional investigations revealed a number of different classes. If altered firing of a particular class
correlations consistent with the idea that off-cells sup- leads to modified nociceptive responses, one can
press, while on-cells facilitate, nociception. Both on- conclude that the observed change in cell activity
and off-cell classes respond to experimental manipu- mediated the resulting change in behavior.
lations of the PAG that produce behavioral analgesia A series of studies using the above approach of
(Vanegas, H. et al., 1984; Hutchison, W. D. et al., 1996; manipulating RVM circuitry directly have demon-
Tortorici, V. and Morgan, M. M., 2002). Withdrawal strated that activation of off-cells is sufficient to
reflexes can generally be elicited at a lower threshold produce analgesia, and required for the antinocicep-
or with shorter latency if on-cells are active and off- tive actions of systemically administered mor-
cells inactive (Heinricher, M. M. et al., 1989; Ramirez, phine (Heinricher, M. M. and Tortorici, V., 1994;
F. and Vanegas, H., 1989; Bederson, J. B. et al., 1990; Heinricher, M. M. et al., 1994; 1997; 1999; 2001a;
Foo, H. and Mason, P., 2003). Antinociception pro- 2001b; Meng, I. D. and Johansen, J. P., 2004;
duced by administration of morphine systemically or Neubert, M. J. et al., 2004; Meng, I. D. et al., 2005).
within the PAG is associated with a uniform activation Similar approaches have recently been extended to
of off-cells and inhibition of on-cells. Importantly, in testing the effects of off-cell activation. Such studies
animals in which opioid administration fails to pro- show that on-cells do not contribute significantly to
duce behavioral analgesia, changes in cell firing are nociceptive threshold under basal conditions as sup-
inconsistent. Thus, changes in on- and off-cell firing pression of the reflex-related firing of on-cells does
are related to antinociception rather than to opioid not by itself reduce responding (Heinricher, M. M.
administration as such (Fields, H. L. et al., 1983b; and McGaraughty, S., 1998; Meng, I. D. et al., 2005).
Cheng, Z. F. et al., 1986.). Reduced nociceptive thresh- However, direct activation of on-cells has a pronoci-
olds in acute opioid withdrawal and secondary ceptive effect: microinjection of neurotensin or
hyperalgesia during noxious stimulation of a distant cholecystokinin (CCK) at doses that activate on-cells
608 The Brainstem and Nociceptive Modulation

selectively produces thermal hyperalgesia (Heinricher, on- and off-cell firing associated with cutaneous versus
M. M. and Neubert, M. J., 2004; Neubert, M. J. et al., visceral noxious-evoked reflexes emphasizes the basic
2004). Activation of on-cells by CCK likely contributes principle that the functions of RVM neurons must be
to nerve injury pain (Kovelowski, C. J. et al., 2000; inferred from their outputs (i.e., from the behavioral or
Porreca, F. et al., 2001; Burgess, S. E. et al., 2002). physiological effect of selective manipulation of the
Microinjection of a CCK-2 receptor antagonist in the difference cell classes), rather from inputs.
RVM also interferes with the paradoxical hyperalgesia
associated with prolonged opioid administration (Xie, 41.3.3.3 Role of neutral cells
J. Y. et al. 2005), indicating that CCK-mediated activa- All cells that are not classified as on- or off-cells are
tion of on-cells is also involved in this phenomenon. grouped together under the heading of neutral cells.
Viewed collectively, the above data provide strong Whether these neurons have any role in pain mod-
evidence that off-cells suppress nociception, and that ulation has long been an important question. By
on-cells facilitate nociception. The balance of activity definition, the firing of neutral cells is unchanged
between the on- and off-cell populations therefore during behavioral responses to noxious stimulation,
allows graded bidirectional control of nociceptive and neutral cell firing is also unaffected by MOR or
responding (Figure 5). DOR agonists, norepinephrine, neurotensin, or CCK,
Functional studies demonstrate that the RVM all of which alter firing of on- and/or off-cells and
exerts bidirectional control over visceral, as well as nociceptive behavior when applied in the RVM
cutaneous, hyperalgesia (Coutinho, S. V. et al., 1998). (Heinricher, M. M. et al., 1994; 1988; Harasawa, I.
The behavioral response to colorectal distension is et al., 2000; Heinricher, M. M. et al., 2001a; Neubert,
suppressed by microinjection of morphine in the M. J., et al., 2004). KOR agonists depress the firing of
RVM (Friedrich, A. E. and Gebhart, G. F., 2003), some neutral cells, but the functional implications of
which is known to activate off-cells (Heinricher, M. this depression have not been explored (Meng, I. D.
M. et al., 1994). Conversely, microinjection of CCK, at et al., 2005). The differential pharmacology of neutral
a dose that likely activates on-cells (Heinricher, M. M. cells supports their categorization as distinct from on-
and Neubert, M. J., 2004), enhances the behavioral and off-cells. Neutral cells, or subpopulations of neu-
response to colorectal distension (Friedrich, A. E. and tral cells, might be involved in some other functions of
Gebhart, G. F., 2003). Thus, on- and off-cells appear to the RVM, for example thermoregulation (Nakamura,
modulate behavioral responses evoked by colorectal K. et al., 2002; Madden, C. J. and Morrison, S. F., 2003).
distension just as they modulate responses evoked by Despite the lack of direct evidence that neutral
cutaneous heat. The lack of congruence between vis- cells have a role in pain modulation, interest in this
ceral inputs to RVM neurons and heat-evoked reflex- cell class remains high. This is in part because a
related responses of these neurons (Brink, T. S. and subset of neutral cells contains serotonin (Potrebic,
Mason, P., 2004) may be related to the inhibitory effect S. B. et al., 1994; Mason, P., 2001), and serotonin is
of colorectal distension on sensory processing of cuta- implicated in descending facilitation as well as des-
neous stimuli (Bouhassira, D. et al., 1994; 1998; cending inhibition of nociception (Le Bars, D., 1988;
Pertovaara, A. and Kalmari, J., 2002; Kalmari, J. and Calejesan, A. A. et al., 2000; Suzuki, R. et al., 2004).
Pertovaara, A., 2004). More generally, a difference in However, there is significant controversy as to

On-cell

Off-cell

Reflex PW PW
Heat
Normal Analgesia Hyperalgesia
Figure 5 Balance of activity between on- and off-cell populations allows graded bidirectional control of nociceptive
responding. Left column, normal conditions: Application of heat (arrow) to a rats tail or paw evokes a paw withdrawal reflex.
The off-cell pauses and on-cell becomes active just before the reflex occurs. Middle column, analgesia: Manipulations that
cause off-cells to become continuously active also suppress on-cell firing and application of heat does not evoke a response.
Right column, hyperalgesia: Reflexes occur with a shorter latency if heat is applied during a period when on-cells are activated
(for example, as a consequence of a prior noxious stimulus).
 The Brainstem and Nociceptive Modulation 609

the exact role of serotonergic neutral cells in pain (Bowker, R. M. et al., 1983; Mantyh, P. W. and
modulation. Immunohistochemically identified sero- Hunt, S. P., 1984; Menetrey, D. and Basbaum, A. I.,
tonergic neurons express MOR, DOR, and KOR 1987; Millhorn, D. E. et al., 1987a; 1987b; Bowker, R.
(Kalyuzhny, A. E. and Wessendorf, M. W., 1999; M. and Abbott, L. C., 1988; Millhorn, D. E. et al., 1989;
Wang, H. and Wessendorf, M.W., 1999; Marinelli, Palkovits, M. and Horvath, S., 1994; Skinner, K. et al.,
S. et al., 2002). However, neutral cells, including 1997). With the exception of serotonin (see above,
putative serotonergic neutral cells, as a class do not Section 41.3.3.1), the relationship between physiolo-
respond to morphine or MOR agonists given sys- gical/functional class (on-, off-, and neutral cell
temically or microinjected into the RVM (Barbaro, classification) and expression of different neurotrans-
N. M. et al., 1986; Heinricher, M. M. et al., 1994; mitters has yet to be examined.
Gao, K. et al., 1998). Why neurons that express the
MOR fail to respond to MOR agonists remains an 41.3.4.1 Gamma-aminobutyric acid
important puzzle. A similar mismatch between in vivo and glutamate: the off-cell pause and
functional data and immunohistochemical data arises on-cell burst
with the neurotensin receptor. Some immunohisto- One approach to identifying the neurotransmitter (s)
chemically identified serotonergic neurons express mediating defined synaptic inputs is to attempt to
neurotensin receptor-like immunoreactivity (NT1- block those inputs using iontophoretic application of
ir), and behavioral studies show that microinjection neurotransmitter antagonists (Hicks, T. P., 1983).
of high doses of neurotensin or a NT1 agonist pro- Iontophoresis of GABAA receptor antagonists selec-
duce antinociception (Smith, D. J. et al., 1997; tively blocks the off-cell pause, demonstrating that
Neubert, M. J. et al., 2004; Buhler, A. V. et al. 2005). this reflex-related inhibition of off-cell firing is
However, physiologically identified neutral cells do mediated by GABA (Heinricher, M. M. et al., 1991).
not respond to neurotensin when microinjected into Consistent with the idea that the off-cell pause
the RVM at a dose sufficient to produce behavioral removes descending inhibition and permits noxious
antinociception (Neubert, M. J. et al., 2004). Again, information to be processed, microinjection of
there is no obvious explanation for why neurons GABAA receptor antagonists in the RVM blocks the
expressing the NT1 receptor show no change in off-cell pause and produces behavioral antinocicep-
firing following microinjection of neurotensin. tion (Drower, E. J. and Hammond, D. L., 1988;
Resolution of these discrepancies between immuno- Heinricher, M. M. and Kaplan, H. J. 1991;
histochemical and electrophysiological findings will Heinricher, M. M. and Tortorici, V., 1994; Gilbert,
require immunohistochemical labeling of function- A. K. and Franklin, K. B., 2001). Interestingly, Nason
ally identified neurons. M. W., Jr. and Mason P. (2004) recently suggested
A second reason for continued interest in neutral that the effect of RVM GABA receptor antagonism is
cells is the report that the response properties of many selective for stimulation of the tail, with facilitation
neutral cells change to those of on- or off-cells over rather than inhibition of responses evoked by stimu-
the course of hours during prolonged inflammation lation of the foot. However, this is difficult to
(Miki, K. et al., 2002). A subset of neutral cells may reconcile with the antinociception observed by
therefore be involved in long-term facilitation or others using the hot plate and formalin tests, as both
inhibition of nociception, or in other changes in beha- of these tests involve stimulation of the hindpaw
vior or physiology during inflammation. As discussed (Gilbert, A. K. and Franklin, K. B., 2001). The source
above however, the pharmacology of such cells would of GABAergic input to off-cells is unknown, but it is
presumably differ from cells classified as on- or off- presumed to be the target of MOR agonists (see
cells in the absence of inflammation. Section 41.3.4.2).
Although firing of both on-cells and neutral cells
is inhibited by iontophoretically applied GABA,
41.3.4 Pharmacology of Nociceptive
neither cell class displays significant disinhibition
Modulation in the Rostral Ventromedial
during iontophoresis of GABAA receptor antagonists.
Medulla
This observation implies that on-cells and neutral
Among the neuropeptides and neurotransmitters cells express GABA receptors, but that there is little
found in RVM neurons are serotonin, substance P, ongoing GABAergic control of their firing, at least in
enkephalin, thyrotropin-releasing hormone, galanin, the lightly anesthetized rat (Heinricher, M. M. et al.,
somatostatin, CCK, GABA, and acetylcholine 1991).
610 The Brainstem and Nociceptive Modulation

Iontophoretic application of excitatory amino et al., 1993; Pan, Z. Z. and Fields, H. L., 1996;
acids excites all three classes of RVM neurons. Roychowdhury, S. M. and Fields, H. L., 1996) to
However, antagonist studies specifically implicate produce its antinociceptive effect.
excitatory amino acid transmitters in reflex-related The local actions of MOR agonists within the
activation of on-cells and opioid-induced activation RVM are well understood at both the cellular and
of off-cells. Thus, iontophoretic application of the circuit levels. On-cells are directly inhibited by MOR
broad-spectrum excitatory amino acid antagonist agonists. Neither neutral cells nor off-cells are
kynurenate blocks the reflex-related on-cell burst, directly sensitive to MOR agonists, but off-cells are
without affecting the activity of off-cells or neutral activated indirectly, most likely via disinhibition
cells (Heinricher, M. M. and Roychowdhury, R., (Figure 6). Microinjection of MOR agonists in the
1997). Kynurenate microinjected into the RVM at a RVM suppresses on-cell firing and activates off-cells,
dose that blocks on-cell activation does not result in resulting in behavioral antinociception (Heinricher,
increased withdrawal latency to noxious heat, indi- M. M. et al., 1992; 1994). The activation of off-cells is
cating that the on-cell burst does not control required for the antinociceptive effect (Heinricher,
nociceptive threshold, at least under normal condi- M. M. et al., 1997). Suppression of on-cell firing is not
tions (Heinricher, M. M. and McGaraughty, S., 1998). necessary for RVM-mediated analgesia (Heinricher,
However, it is possible that the firing of on-cells M. M. et al., 1994; Neubert, M. J. et al., 2004), although
during and after the withdrawal modulates the mag- it likely contributes. Selective inhibition of on-cell
nitude or force of the reflex, or primes responses to firing reduces the force, although not the latency, of
subsequent stimuli (Ramirez, F. and Vanegas, H., withdrawal to noxious heat (Heinricher, M. M. and
1989; Jinks, S. L. et al., 2004b). McGaraughty, S., 1998; Jinks, S. L. et al., 2004b).
A second important role for excitatory amino acid Studies at the cellular level reveal that some neu-
transmission in the RVM is the recruitment of off- rons are inhibited directly by MOR agonists, whereas
cells by opioids (Heinricher, M. M. et al., 2001b). others are disinhibited via presynaptic inhibition of
Disinhibition of off-cells leads to NMDA-mediated GABAergic transmission (Pan, Z. Z. et al., 1990).
activation of these neurons, amplifying the effect of These findings are thus consistent with the direct
disinhibition (see the next Section 41.3.4.2). inhibition of on-cells and indirect activation of off-
cells observed in vivo. RVM neurons studied in vitro
41.3.4.2 Opioid actions in the rostral
are classified as secondary cells, which are directly
ventromedial medulla
All three opioid receptors, MOR, DOR, and KOR,
are found in the RVM, although their expression is
much less dense than in the PAG (Gutstein H. B.
et al., 1998; Kalyuzhny, A. E. et al., 1996; Wang, H. and
Wessendorf, M. W., 1999). The RVM, and specifi-
cally activation of off-cells, is required for the
analgesic actions of systemically administered mor-
phine (Dickenson, A. H. et al., 1979; Azami, J. et al.,
1982; Mitchell, J. M. et al., 1998; Heinricher, M. M.
et al., 2001a; 2001b; Gilbert, A. and Franklin, K., 2002).
Like the PAG, the RVM supports MOR-mediated
analgesia. Thus, direct local microinjection of mor-
phine or MOR agonists in the RVM produces an
antinociception equivalent to that produced by sys-
temic morphine administration (Heinricher, M. M.
and Morgan, M. M., 1999). Moreover, PAG and
RVM interact in a synergistic fashion (Rossi, G. C. Figure 6 Disinhibition of off-cells, direct inhibition of on-
et al., 1994). This is at least in part because exogenous cells mediates opioid analgesia. On-cells are inhibited,
directly, by morphine and mu-opioid receptor (MOR)
opioid administration at one site recruits endogenous
agonists. Off-cells are not inhibited by MOR agonists, but
opioid mechanisms at the other. Thus, microinjection are activated, via disinhibition. MORs are thus presumed to
of morphine in the PAG apparently evokes release be located on on-cells, and on gamma aminobutyric acid
of endogenous opioids in the RVM (Kiefel, J. M. (GABA)ergic inputs to off-cells.
 The Brainstem and Nociceptive Modulation 611

inhibited by MOR agonists, and primary cells, which 41.3.4.2.(ii) Kappa opioid receptor The role of
are not. Primary cells do however receive opioid- the KOR in the pain-modulating functions of the
sensitive GABAergic inputs, and are disinhibited by RVM remains controversial. Electrophysiological
MOR agonists (Pan, Z. Z. et al., 1990). Secondary cells studies in the RVM slice demonstrate pre- and post-
presumably map to on-cells recorded in vivo, and synaptic effects of KOR agonists in the RVM.
primary cells to off-cells and neutral cells identified However, it is unclear whether neurons in which
in vivo. KOR agonists evoke an outward current are a sepa-
rate population from those inhibited by MOR
agonists (Pan, Z. Z. et al., 1997; Ackley, M. A. et al.,
41.3.4.2.(i) Delta opioid receptor Like MOR ago- 2001; Marinelli, S. et al., 2002; Bie, B. and Pan, Z. Z.,
nists, DOR agonists, particularly delta2 agonists, 2003). Microinjection of a KOR agonist into the
produce behavioral hypoalgesia when microinjected RVM has been reported to either have no behavioral
into the RVM. The effects of delta1 agonists are less effect (Pan, Z. et al., 2000; Meng, I. D. et al., 2005) or to
robust than those of delta2 agonists (see Heinricher, M. produce antinociception, depending on the nociceptive
M. and Fields, H. L., 2003 for review of the role of the test as well as the sex of the animal (Tershner, S. A.
DOR in pain modulation in the RVM). In vitro, the et al., 2000; Ackley, M. A. et al., 2001). KOR agonists also
DOR agonists deltorphin and [D-Pen2, D-Pen5] enke- block the antinociceptive effect of PAG microinjec-
phalin (DPDPE) elicit hyperpolarizing currents in a tion of a MOR agonist or systemic morphine
substantial minority of RVM neurons (Marinelli S. administration (Pan, Z. et al., 2000; Bie, B. and Pan,
et al., 2005). Some, but not all, of these neurons were Z. Z., 2003; Meng, I. D. et al., 2005) as well as MOR-
also hyperpolarized by MOR agonists. It thus appears mediated conditioned hypoalgesia (Foo, H. and
that a subset of DOR-sensitive RVM neurons falls into Helmstetter F. J., 2000). This antianalgesic effect of
the KOR agonist is presumably due to a blockade of
the secondary cell category. In experiments in vivo,
off-cell activation (Bie, B. and Pan, Z. Z., 2003; Meng,
microinjection of deltorphin into the RVM increases
I. D et al., 2005). KOR agonists can also block hyper-
the latency and reduces the amplitude of the reflex-
algesia when microinjected into the RVM,
related on-cell burst, reduces the duration of the
presumably via presynaptic blockade of excitatory
reflex-related off-cell pause, and evokes a small but
inputs to on-cells (Ackley, M. A. et al., 2001; Meng,
statistically significant increase in tail flick latency
I. D. et al., 2005).
(Harasawa, I. et al., 2000).
Molecular, cellular, and behavioral approaches
41.3.4.3 Norepinephrine
point to interactions between MOR- and DOR-
As with the KOR, norepinephrine likely influences
mediated processes, but experiments designed to
both the nociceptive facilitating and inhibiting out-
uncover such interactions within the RVM reveal
puts from the RVM. Behavioral studies have reported
that the effects of MOR and DOR agonists in the
that blockade of 1-adrenergic receptors produces
RVM are independent under basal conditions
antinociception (Sagen, J. and Proudfit, H. K., 1981)
(Hurley, R. W. and Hammond, D. L. 2001; Kalra, A. and reduces hyperalgesia associated with acute
et al., 2001). However, there is a time-dependent opioid withdrawal (Bie, B. et al., 2003). Agonists at
recruitment of supraspinal DOR function in inflam- the 1 receptor produce hyperalgesia and condi-
mation, so that DOR agonists microinjected into the tioned place avoidance, and the increased
RVM are more effective in animals subjected to long- nociception is dissociated from the negative affective
lasting inflammation compared to noninflamed con- state in these animals (Hirakawa, N. et al., 2000).
trols. MOR agonists are also more potent in these Surprisingly, 2 agonists have been reported to
animals (Hurley, R. W. and Hammond, D. L., 2000; produce analgesia (Sagen, J. and Proudfit, H. K., 1985;
2001). This increased opioid action may reflect an Haws, C. W. et al., 1990) as well as to block opioid
increase in the release of endogenous opioids acting analgesia (Bie, B. et al., 2003) when microinjected into
at the DOR (Williams F. G. et al., 1995) and/or the RVM. The neural basis for these conflicting
translocation of DOR from the cytoplasm to the results is unclear, as electrophysiological studies are
plasma membrane (Commons, K. G. et al., 2001; also inconsistent. In vivo, iontophoretically applied
Commons, K. G., 2003). Coexpression of MOR and norepinephrine and 2 agonists suppress the firing
DOR on individual RVM neurons may also play a of on-cells, whilst 1 agonists activate on-cells. Off-
role (Marinelli, S. et al., 2005). cells and neutral cells do not respond to
612 The Brainstem and Nociceptive Modulation

iontophoretically applied norepinephrine or adre- Gebhart, G. F., 2003; Heinricher, M. M. and Neubert,
nergic agents (Heinricher, M. M. et al., 1988). By M. J., 2004). Notably, CCK activates on-cells selec-
contrast, in experiments in the RVM slice, 2 ago- tively when microinjected in a dose sufficient to
nists hyperpolarize primary cells (presumably off- produce behavioral hyperalgesia. This points to on-
cells and/or neutral cells), and 1 agonists activate cells as critical mediators of CCK hyperalgesia in the
all cells (Bie, B. et al., 2003). The findings from the in RVM (Heinricher, M. M. and Neubert, M. J., 2004).
vivo study are thus more consistent with an antinoci- CCK also has antiopioid actions in the RVM,
ceptive effect of 2 receptor activation, whereas the albeit through a different mechanism than that
studies in the slice are more consistent with the through which it produces hyperalgesia. Focal appli-
antiopioid action of these agents. cation of a low dose of CCK in the RVM attenuates
both opioid activation of off-cells and opioid-induced
antinociception without affecting baseline nocicep-
41.3.4.4 Cannabinoids
tive responding or the activity of on-cells
As noted above (Section 41.2.5.4), the PAGRVM
(Heinricher, M. M. et al., 2001a). The signal transduc-
system is now recognized to mediate at least part of
tion mechanisms that underlie this antiopioid action
the antinociceptive effect of cannabinoids. The RVM
of CCK are unknown. In the hippocampus, CCK
is required for the antinociceptive action of systemi-
reduces opioid-induced disinhibition by increasing
cally administered cannabinoid agonists (Meng, I. D.
GABA release (Miller, K. K. et al., 1997), raising the
et al., 1998), and microinjection of CB1 agonists into
possibility that CCK acts presynaptically to block
the RVM produces moderate antinociception
disinhibition of off-cells in the RVM.
(Martin, W. J. et al., 1998; Monhemius, R. et al., 2001;
Meng, I. D. and Johansen, J. P., 2004). Off-cells dis-
41.3.4.6 Neurotensin
play a measurable increase in activity after local
infusion of a CB1 agonist in the RVM, and the Like CCK, neurotensin has multiple effects in the
RVM. Microinjection of this peptide within the
reflex-related activity of on-cells is significantly
RVM gives rise to a dose-related, bidirectional effect
reduced (Meng, I. D. and Johansen, J. P., 2004). Both
on nociceptive behaviors (tail flick, hot plate, and
of these effects are presumed to reflect a
visceromotor responses to colorectal distension) and
presynaptic action, as cannabinoids block excitatory
dorsal horn nociceptive neurons. Extremely low
and inhibitory synaptic currents, but have negligible
doses produce facilitation, whereas high doses pro-
postsynaptic effects in RVM (Vaughan C. W. et al.,
duce antinociception. Intermediate doses are without
1999).
effect (Urban, M. O. and Smith, D. J., 1993; Smith, D.
J. et al., 1997; Urban, M. O. and Gebhart, G. F., 1997;
41.3.4.5 Cholecystokinin Urban, M. O. et al., 1999; Neubert, M. J. et al., 2004.).
Endogenous CCK has long been recognized to coun- Behavioral hyperalgesia is mediated by selective acti-
ter opioid analgesia and to contribute to enhanced vation of on-cells, whereas the hypoalgesia produced
nociception, particularly in neuropathic pain states by higher doses is due to recruitment of off-cells.
(Stanfa, L. et al., 1994; Wiesenfeld-Hallin, Z. et al., Neutral cells do not respond to neurotensin at any
1999). In humans, CCK antagonism increases the dose (Neubert, M. J. et al. 2004).
analgesic actions of morphine and placebo (Price,
D. D. et al., 1985; Benedetti, F. and Amanzio, M., 41.3.4.7 Nociceptin/orphanin FQ
1997; McCleane, G. J., 1998). CCK is found in the The identification of an opioid-like receptor (ORL-
RVM (Skinner, K. et al., 1997), and has both prono- 1) that did not bind classical opioid peptides sug-
ciceptive and antiopioid actions in this region. CCK gested that additional peptides might exist that
in the RVM is implicated in the expression of allo- could modulate nociception. However, despite its
dynia and thermal hyperalgesia in the spinal nerve structural and functional similarities to the opioid
ligation model of nerve injury pain as well as in receptors, activation of ORL-1 by its endogenous
hyperalgesia associated with chronic morphine ligand, referred to as nociceptin or orphanin FQ
administration (Kovelowski, C. J. et al., 2000; Xie, J. (OFQ), does not produce antinociception when
Y. et al., 2005). Furthermore, focal application of applied within the RVM. Rather nociceptin/OFQ
relatively high doses of exogenous CCK in the potently inhibits all neurons in the RVM. As a con-
RVM produces cutaneous and visceral hyperalgesia sequence, focal application of nociception/OFQ in
(Kovelowski, C. J. et al., 2000; Friedrich, A. E. and the RVM is effectively a functional lesion of the
 The Brainstem and Nociceptive Modulation 613

region, and interferes with both antinociceptive and apparent lack of pain observed in some patients sub-
pronociceptive outputs (see Heinricher, M. M., in jected to intense trauma (Wall, P. D. 1979; Melzack, R.
press for a review). et al., 1982) and the from the ability of footshock or other
noxious stimuli to suppress animals responses to other
painful events (Maier, S. F., 1986). Several factors must
41.3.5 Physiological Activation of
be considered in addressing this issue. First, other brain-
Nociceptive Modulatory Neurons in the
stem pathways, such as that through the medullary
Rostral Ventromedial Medulla
subnucleus reticularis dorsalis (Le Bars, D., 2002), act
Given a pronociceptive role for on-cells and an anti- in parallel with the output from the RVM. The net
nociceptive role for off-cells, the obvious question effect of a given noxious stimulus on spinal nociceptive
that arises is how and when these two classes of processing and pain sensation will reflect the actions of
neurons with opposing functions are recruited to these pathways as well as that of the RVM. In addition,
modulate nociception. This question remains a sig- higher-order processes, such as fear or stress, are likely
nificant challenge, in part because a number of factors to be significant when an animal is subjected to an
likely to influence nociception via the RVM involve experimental noxious stimulus, particularly one over
higher-order neural structures and psychological which it has no control. Such higher influences will be
processes that are difficult to study in rodents, parti- most prominent in awake behaving animals, and may
cularly under anesthesia. Nevertheless, we do have shift the balance in the RVM towards activation of off-
some information as to how and when RVM cells rather than on-cells. Such a shift is presumably the
nociceptive modulatory neurons are recruited to basis for stress-induced analgesia or antinociception
modulate nociceptive processing. associated with fear conditioning (Bodnar, R. J. et al.,
1980; Fanselow, M. S. 1986; Helmstetter, F. J. and
41.3.5.1 Response of RVM neurons to Tershner, S. A., 1994). Conditioned fear, for example,
noxious stimulation: does pain inhibit produces antinociception mediated by the amygdala
pain? and the RVM, and direct opioid activation of
Noxious-evoked withdrawals are associated with a the amygdala activates RVM off-cells via the PAG
period of on-cell activation and off-cell inhibition (Helmstetter, F. J. and Landeira-Fernandez, J., 1990;
that can last from less than a second to many minutes. Helmstetter, F. J., 1992; Helmstetter, F. J. and
This period, during which the balance between the Bellgowan, P. S., 1993; Helmstetter, F. J. and Tershner,
on- and off-cell populations is shifted toward on-cell S. A., 1994; Helmstetter, F. J. et al., 1998; McGaraughty,
activation, is associated with heightened nociceptive S. and Heinricher, M. M., 2002; 2004).
responsiveness that can be blocked by lesion of the Thus, because of higher-order inputs to the RVM
RVM (Heinricher, M. M. et al. 1989; Ramirez, F. and and the actions of other descending pathways, the
Vanegas, H., 1989; Morgan, J. L. and Fields, H. L., behavioral response to a given noxious stimulus may
1994; Foo, H. and Mason, P., 2003). Thus noxious be enhanced or suppressed in the presence of pain.
stimulation per se recruits the RVM to facilitate noci- Which behavioral outcome predominates will
ception. This conclusion is consistent with the early depend on the stimulus history, the environment in
suggestion of Cervero F. and Wolstencroft J. H. which the stimulus is applied and the behavioral state
(1984) that the RVM is part of a short-term positive of the animal.
feedback loop activated by noxious stimulation. Such
a positive feedback process presumably prepares the 41.3.5.2 Behavioral state control:
organism to respond more briskly or at a lower anesthesia and sleep/waking cycle
threshold to subsequent damaging inputs. This view The influence of behavioral state variables on noci-
of on-cells as part of a positive feedback loop is ceptive processing at the first central relay in the
further supported by evidence that on-cells are acti- dorsal horn (Hayes, R. L. et al., 1981; Bushnell, M.
vated by acute cutaneous inflammation, and that the C. et al., 1984; Soja, P. J. et al., 1999) suggests that
RVM is necessary for secondary hyperalgesia in this pathways descending from the brain to the dorsal
paradigm (Urban, M. O. et al., 1996; Kincaid, W. et al., horn contribute to the influence of behavioral state
in press). on nociception. One possibility that has been consid-
The evidence for positive feedback mediated by the ered is that RVM neurons might mediate effects of
RVM is plainly at odds with the long-standing idea that general anesthesia or sleep/waking on nociceptive
pain inhibits pain, which arose from observations of an processing.
614 The Brainstem and Nociceptive Modulation

Correlative data suggest that on- and/or off-cell rats does not produce loss of motor tone, even though
firing could contribute to isoflurane-induced motor locomotor activity in an open field is reduced
depression at lighter planes of anesthesia (Jinks, S. L. (Morgan, M. M. and Whitney, P. K., 2000). Finally,
et al., 2004a). However, firing of both on- and off-cells the firing of on- and off-cells does not vary between
is depressed when isoflurane concentration is slow wave sleep and paradoxical sleep (Foo, H. and
increased to levels above that required to suppress Mason, P., 2003). Taken together, these data indicate
nocifensor reflexes (Leung, C. G. and Mason, P., that on- and off-cells are unlikely to mediate atonia
1995). These neurons are thus unlikely to mediate during paradoxical sleep.
the effects of a anesthesia at a surgical plane on
nociceptive withdrawals. 41.3.5.3 Micturition
Analysis of the role of on- and off-cells in mediat- Baez M. A. et al. (2005) recently reported that paw
ing changes in nociception across the sleepwaking withdrawal to noxious heat is attenuated during
cycle is impeded by lack of a clear understanding micturition, presumably preventing potential inter-
of how nociception changes with sleep/waking. In ruption of bladder emptying by noxious stimulus-
cats, Kshatri A. M. et al. (1998) demonstrated that evoked movements. Consistent with the idea that
the latency of the tail flick reflex was slightly off-cells inhibit and on-cells facilitate nociception
increased during slow-wave sleep compared to wak- during micturition, many presumptive off-cells
ing, but substantially longer during paradoxical sleep. were active during urine expulsion, whereas pre-
Consistent with this, noxious-evoked activity of tri- sumptive on-cells were inhibited. However, these
geminal neurons is depressed during paradoxical authors propose a somewhat broader role for RVM
sleep, but not slow-wave sleep, compared to waking neurons, as electrical stimulation in the RVM and
(Cairns, B. E. et al., 1995; 1996). By contrast, Mason P. morphine microinjection in the PAG are known to
et al. (2001) reported that paw withdrawals evoked by suppress bladder contractions (Sugaya, K. et al., 1998;
heat were enhanced during slow-wave sleep com- Matsumoto, S. et al., 2004). Baez M. A. et al. (2005)
pared to waking, suggesting a disinhibition of therefore suggested that off-cell activation suppresses
nociception during slow-wave sleep. Parallel record- voiding, possibly by depressing transmission of blad-
ing studies from this group demonstrated that off- der afferent information.
cells were more active during slow-wave sleep and
on-cells and neutral cells were less active compared 41.3.5.4 Environmental analgesia
to waking (Leung, C. G. and Mason, P., 1999). It is Decreases in pain responsiveness can be induced by a
therefore unlikely that on- or off-cells mediated the wide range of experimental procedures, including
enhanced nociception seen in their behavioral analy- electrical shock, forced swim, and centrifugal rotation
sis. However, these authors noted that rats return to (Watkins, L. R and Mayer, D. J., 1982; Bodnar, R. J.,
sleep more quickly following noxious stimulation if 1986; Maier, S. F., 1986). Biologically relevant threat
the stimulus is delivered when the rats are sleeping stimuli such as odors from stressed animals of the
rather than awake. They suggested that this could same species or exposure to a predator also induce
point to a role for off-cells in controlling arousal. antinociception (Fanselow, M. S., 1985; Lester, L. S.
However, it is not clear that the duration of waking and Fanselow, M. S., 1985; Kavaliers, M. 1988;
triggered by the noxious stimulus could be differen- Lichtman, A. H. and Fanselow, M. S., 1990).
tiated from the ongoing waking pattern in their Activation of endogenous pain-modulating systems
analysis. It remains to be determined whether off- can also enhance the analgesic effects of exogenous
cells control arousal, or mediate the influence of opioids. Thus, presentation of a stressful shock
arousal on nociception. potentiates morphine-induced antinociception, an
A further question is whether on- and off-cells effect that increases as the intensity of stress is
contribute to the atonia of paradoxical sleep. This increased (Grau, J. W. et al., 1981; Rosellini, R. A.
has not been tested directly. However, although sti- et al., 1994). The PAGRVM system is implicated in
mulation throughout the length of the medial at least some of these environmentally induced
pontomedullary reticular formation can produce ato- changes in nociceptive responsiveness. Most impor-
nia in decerebrate rats (Hajnik, T. et al., 2000), the tant, some forms of environmental analgesia can be
most effective sites are generally dorsal to the area attenuated by lesioning the RVM (Prieto, G. J. et al.,
defined as the RVM. Moreover, microinjection of 1983; Watkins, L. R. et al., 1983b). Moreover, admin-
kainate or morphine in the RVM in intact behaving istration of a stressful foot shock blocks both the
 The Brainstem and Nociceptive Modulation 615

pause in firing that characterizes off-cells and the Fernandez, J., 1990; Helmstetter, F. J. and Tershner,
typical on-cell burst of activity (Friederich, M. W. S. A., 1994). The amygdala sends a sparse projection
and Walker, J. M., 1990). This indicates that foot- into the RVM region, raising the possibility that the
shock and possibly other stressors affect the activity amygdala could directly influence the RVM (Price, J.
of on- and off-cell classes in a manner consistent L. and Amaral, D. G., 1981; Hermann, D. M. et al.,
with their respective pronociceptive and antinoci- 1997). However, the projection from the amygdala to
ceptive roles. the PAG is much more robust, and the PAG is
Antinociception is also elicited as a conditioned thought to be a necessary relay in the antinociception
response to previously neutral cues paired with nox- associated with conditioned fear. Amygdala projec-
ious or aversive events (Fanselow, M. S., 1986). tions to the PAG arise from the central nucleus, and
This latter phenomenon is termed conditioned anti- to a lesser extent the medial nucleus (Krettek, J. E.
nociception, and has been widely adopted for and Price, J. L., 1978; Rizvi, T. A. et al., 1991;
experimental analysis of endogenous mechanisms Canteras, N. S. et al., 1995). Terminations from the
involved in nociceptive modulation. In condi- central nucleus are concentrated in PAG regions that
tioned antinociception paradigms, antinociception is in turn send projections to the RVM (Rizvi, T. A.
recruited in concert with other behaviors and auto- et al., 1991). Activation of the central or basolateral
nomic adjustments as part of a defensive reac- nucleus of the amygdala alters the firing of PAG
tion (Williams, F. G. et al., 1990; Fanselow, M. S., neurons, with approximately equal proportions of
1991; Harris, J. A. and Westbrook, R. F., 1995). PAG neurons showing excitation and inhibition
Benzodiazepine receptor inverse agonists have (Sandrew, B. B. and Poletti, C. E., 1984; da Costa
anxiogenic or fear-promoting effects, and yield anti- Gomez, T. M.and Behbehani, M. M., 1995). Effects
nociception after either intracerebral or systemic of basolateral stimulation are mediated primarily
administration (Rodgers, R. J. and Randall, J. I., through the central nucleus (da Costa Gomez, T.
1988; Helmstetter, H. J. et al., 1990; Fanselow, M. S. M. et al., 1996). Finally, microinjection of morphine
and Kim, J. J., 1992). Conversely, hypoalgesia is in the basolateral nucleus of the amygdala activates
reduced by manipulations that would be expected off-cells and suppresses on-cell firing, and these
to reduce fear, such as administration of anxiolytic changes in on- and off-cell activity are associated
agents (Fanselow, M. S. and Helmstetter, H. J., 1988). with behavioral antinociception (McGaraughty, S.
The amygdala, a forebrain structure with a well- and Heinricher, M. M., 2002). Taken together, the
documented role in fear, stress and anxiety, is critical above findings indicate that one way in which the
in organization of the fear-related processes modulatory circuitry of the PAGRVM circuitry is
described above (Davis, M., 1992a; 1992b). Notably, engaged physiologically is via activation of the amyg-
amygdala lesions attenuate freezing and analgesia in dala by stimuli that induce fear.
rats exposed to a cat, which is an innate fear stimulus
for this species (Blanchard, D. C. and Blanchard, R. J.,
1972; Fox, R. J. and Sorenson, C. A., 1994), and to 41.4 Conclusion
intense, nonnoxious noise (Helmstetter, F. J. and
Bellgowan, P. S., 1994; Bellgowan, P. S. and The effort to understand the neural basis of nocicep-
Helmstetter, F. J., 1996). Such lesions have no effect tive modulation by the PAGRVM system highlights
on baseline nociceptive responsiveness (Helmstetter, the importance of studying functionally identified
F. J. 1992; Fox, R. J. and Sorenson, C. A., 1994; neurons. The RVM can both facilitate and inhibit
Manning, B. H. and Mayer, D. J., 1995; Watkins, nociception. Furthermore, this region is also impli-
L. R. et al., 1993). cated in a number of functions other than nociceptive
Antinociception produced by fear-related pro- modulation, including reproductive behaviors, cardi-
cesses organized in the amygdala is mediated by the ovascular and respiratory control, sleepwaking and
PAGRVM system. Thus, conditioned antinocicep- arousal, thermoregulation, and behavioral suppres-
tion involves suppression of nociceptive processing at sion. A meaningful analysis of how the RVM
the level of the spinal cord (Harris, J. A. et al., 1995), contributes to enhanced pain states therefore requires
and is disrupted by lesions of the PAG or the RVM, functional identification of the neurons under study,
or by infusion of an opiate antagonist into the PAG so that mechanisms contributing to nociceptive
(Watkins, L. R. et al., 1983a; 1983b; Kinscheck, I. B. facilitation can be distinguished from those involved
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Acknowledgments Brain Res. 366, 203210.
Barbaro, N. M., Heinricher, M. M., and Fields, H. L. 1989.
Putative nociceptive modulatory neurons in the rostral
MMH was supported by grants from NIDA (DA ventromedial medulla of the rat display highly correlated
05608) and NINDS (NS 40365), and S. L. I. by a firing patterns. Somatosens. Mot. Res. 6, 413425.
Bederson, J. B., Fields, H. L., and Barbaro, N. M. 1990.
NARSAD Young Investigator Award and a grant Hyperalgesia during naloxone-precipitated withdrawal from
from NIDA (DA 015498). morphine is associated with increased on-cell activity in the
rostral ventromedial medulla. Somatosens. Mot. Res.
7, 185203.
Behbehani, M. M. and Da Costa Gomez, T. M. 1996. Properties
of a projection pathway from the medial preoptic nucleus to
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 42 Emotional and Behavioral Significance of the Pain Signal
and the Role of the Midbrain Periaqueductal Gray (PAG)
K Keay and R Bandler, University of Sydney, Sydney, NSW, Australia
2009 Elsevier Inc. All rights reserved.

42.1 More than a Sensation? 627

42.2 Coping with Pain 627
42.2.1 Active versus Passive Emotional Coping 627
42.2.2 Neural Representations of Active Emotional Coping 628
42.2.2.1 Dorsolateral periaqueductal gray 628
42.2.2.2 Lateral periaqueductal gray 630
42.2.3 Neural Representation of Passive Emotional Coping 630
42.2.3.1 Ventrolateral periaqueductal gray 630
42.2.4 Outputs Mediating Active and Passive Emotional Coping 630
42.2.4.1 Lateral periaqueductal gray and ventrolateral periaqueductal gray 630
42.2.4.2 Dorsolateral periaqueductal gray 630
42.3 Conclusion 633
References 633

42.1 More than a Sensation? It is striking that these eminent researchers

although starting from different perspectives arrive
In a provocative article entitled On the Relation of Injury at a shared view, namely, that in addition to knowl-
to Pain, Wall P. D. (1979) suggested that . . . pain is edge of the anatomy and physiology of its ascending
better classified as an awareness of a need state than as a sensory pathways, understanding how pain is loca-
sensation; that it has more in common with the phe- lized in the brain also requires knowledge of the
nomena of hunger and thirst than it has with seeing and neurobiology of the emotional coping strategies trig-
hearing . . . . He also observed, somewhat anecdotally, gered by different classes of pain.
that as pain changed from acute to chronic the emo-
tional coping reaction also changed. Livingston W. K.
(1998) in his posthumously published book Pain and
Suffering stated a similar belief that . . . pain is not 42.2 Coping with Pain
strictly a physical sensation that can be defined simply
by its anatomical and physiological substrates. Writing 42.2.1 Active versus Passive Emotional
even earlier, Lewis T. (1942) observed that different Coping
emotional coping reactions were evoked by pain of Animals (including humans) employ different strate-
different tissue origins, . . . while painful sensations gies to cope with different classes of pain. Active
derived from the human skin are associated with brisk emotional coping, which is characterized by engage-
movements, with a rise of pulse rate and with a sense of ment with the environment (i.e., fight or flight), is the
invigoration . . . painful sensations derived from deeper usual response to escapable pain. Components of
structures are associated often with quiescence, with active coping include increased vigilance, hyperreac-
slowing of the pulse, falling of the blood pressure, tivity, increased somatomotor activity, and increased
sweating and nausea. He went on to suggest that sympathetic activity. Passive emotional coping, is the
. . . the difference in the quality of skin pain and of antithesis of active coping. Instead of engagement
deep pain is so clear . . . that it would perhaps seem with the environment, there is disengagement (e.g.,
unsafe to class both together under the one unqualified conservation-withdrawal (Henry, J. P. and Stephens,
term pain and concluded that . . . we should bear in P. M., 1977)). Components of passive coping include:
mind the possibly serious fallacy of regarding both decreased vigilance, hyporeactivity, quiescence,
types (of pain) as represented in a common centre. and falls in arterial pressure and heart rate. Passive

627
628 Emotional and Behavioral Significance of PAG

emotional coping is the usual response to inescapable 42.2.2.1 Dorsolateral periaqueductal gray
pain, i.e., pain of deep origin (muscle, joint, viscera) The dlPAG is distinguished from the lPAG by the
or any persistent/chronic pain (Lewis, T., 1942; presence (e.g., nitric oxide synthase (NOS), cholecys-
Wall, P. D., 1979; Bandler, R. et al., 2000; Keay, K. A. tokinin (CCK), acetylcholine (ACh), Met-enkephalin)
et al., 2001). or absence (e.g., cytochrome oxidase and Gly-2 trans-
porter) of specific neurochemicals (for a review, see
Keay, K. A. and Bandler, R., 2004). Anatomically,
42.2.2 Neural Representations of Active major descending inputs to dlPAG originate from
Emotional Coping medial prefrontal cortical (PFC) fields and select
medial hypothalamic nuclei, but there is a striking
Research carried out in rats, cats, and primates has absence of ascending afferents arising from either
identified longitudinally oriented, neuronal columns brainstem or spinal cord (Figure 2). The predomi-
within the midbrain periaqueductal gray (PAG) as nance of forebrain, primarily cortical inputs, led to
critical substrates integrating active or passive emo- the suggestion that when triggered by psychological
tional coping. As seen in Figure 1 active emotional (cortical) stimuli, active coping is mediated by the
coping responses (i.e., a coordinated reaction of freez- dlPAG (Bandler, R. et al., 2000). Consistent with
ing, fight or flight, hypertension, tachycardia, and a this suggestion, the potential threat signaled by the
nonopioid-mediated analgesia) are evoked when presence of a cat (sight and/or odor, but without
excitatory amino acids are microinjected into either physical contact) triggers active coping and a selec-
the dorsolateral (dlPAG) or lateral (lPAG) column. tive increase in immediate early gene (c-fos)
(Bandler, R. and Shipley, M. T., 1994). Anatomical expression in the dlPAG of the rat (Figure 2)
and functional studies suggest, however, different (Canteras, N. S. and Goto, M., 1999).
neural circuits and triggers are involved.

Confrontational defense/threat
hypertension and tachycardia
Dorsolateral and lateral PAG:
nonopioid-mediated analgesia
active emotional coping
dm
dl

dm Escape/flight
dl hypertension and tachycardia
nonopioid-mediated analgesia
lat
dm
dl

lat

dm
vl dl
Ventrolateral PAG:
passive emotional coping lat

Quiescence and hyporeactivity

hypotension and bradycardia vl
opioid-mediated analgesia

Figure 1 Schematic illustration of the dorsomedial (dm), dorsolateral (dl), lateral (lat), and ventrolateral (vl) neuronal columns
within the periaqueductal gray (PAG). Injections of excitatory amino acids (EAA) within the dorsolateral and lateral PAG evoke
active emotional coping strategies, whereas passive emotional coping strategies are evoked from the ventrolateral PAG.
Specifically, EAA injections made within the dlPAG and lPAG columns evoke defensive reactions (confrontation, threat,
escape, flight), hypertension, tachycardia, and a non-opioid-mediated analgesia. In contrast, EAA injections made within
the vlPAG evoke cessation of spontaneous activity (quiescence), decreased responsiveness to the environment
(hyporeactivity), hypotension, bradycardia, and an opioid-mediated analgesia. Adapted from Bandler, R. and Shipley, M. T.
1994. Columnar organization in the midbrain periaqueductal gray: modules for emotional expression? Trends Neurosci. 17,
379389.
 Emotional and Behavioral Significance of PAG 629

Prefrontal cortex
FPm ACd ACv
PL
MO IL

Hypothalamus
Dorsolateral PAG
PVN
f dl
LHAa
lat
AHA 3V

opt III

dl
DHA lat
3V
DMH
LHAm vl
f DR

dl

mt 3V
LHAp
PHA
vl
f 3V cnf

Rostral medulla

RVLM

Exposure to a cat

5.8 6.8 7.8 8.8

Figure 2 Schematic illustration of the major afferent and efferent projections of the dorsolateral periaqueductal gray (dlPAG).
Major forebrain inputs to the dlPAG arise from dorsal, medial prefrontal cortical fields, and medial hypothalamic regions. The
dlPAG does not project directly to the medulla, but can influence the rostral ventrolateral medulla via the cuneiform nucleus
(cnf). The lower panel illustrates the pattern of neural activation (c-fos expression) evoked by a psychological stressor
(exposure, without physical contact, of a rat to a cat), which evokes active emotional coping. The shaded area indicates the
dorsolateral PAG column. RVLM, rostral ventrolateral medulla ACd, dorsal anterior cingulate cortex; ACv, ventral anterior
cingulate cortex; AId, dorsal agranular insular cortex; AIp, posterior agranular insular cortex; DLO, dorsolateral orbital cortex;
FPm, medial frontal pole; IL, infralimbic cortex; MO, medial orbital cortex; PL, prelimbic cortex; PRh, perirhinal cortex; VLOm,
ventrolateral orbital cortex, medial part; VO, ventral orbital cortex AHA, anterior hypothalamic area; DHA, dorsal hypothalamic
area; DMH, dorsomedial hypothalamic area; LHAa, lateral hypothalamic area, anterior; LHAm, lateral hypothalamic area,
medial; LHAp, lateral hypothalamic area, posterior; PHA, posterior hypothalamic area; PVN, periventricular hypothalamic
nucleus; f, fornix; mt, mamillothalmic tract; opt, optic tract; 3V, third ventricle; dl, dorsolateral PAG; lat, lateral PAG; vl,
ventrolateral PAG, DR, dorsal raphe. Exposure to a cat: Adapted from Canteras, N. S. and Goto, M. 1999. Fos-like
immunoreactivity in the periaqueductal gray of rats exposed to a natural predator. Neuroreport 10, 413418.
630 Emotional and Behavioral Significance of PAG

42.2.2.2 Lateral periaqueductal gray afferents suggests that passive coping, whether
Although the boundary between the lPAG and evoked by physical or psychological stimuli, is
dlPAG can be established on neurochemical grounds, mediated by the vlPAG. In support of this view,
the boundary between the lPAG and ventrolateral Fos expression is strongly evoked in the vlPAG
periaqueductal gray (vlPAG) rests on functional when passive coping is triggered either (1) as a pri-
criteria (see Figure 1 and Section 42.2.3). mary response to a deep or persistent noxious
Anatomically, the lPAG receives modest descending stimulus (see Figure 4) (Clement, C. I. et al., 1996;
afferents of cortical and dorsal hypothalamic origins. Keay, K. A. et al., 1997; 2001; Keay, K. A. and Bandler,
However, in contrast to the dlPAG, the lPAG R., 2002) or (2) in order to promote recovery and
receives substantial and somatotopically organized healing, as a delayed response to acute injury (for a
ascending inputs from laminae I, II, IV, and V of discussion, see Wall, P. D., 1979; Keay, K. A. and
the spinal cord, as well as the caudal, spinal trigem- Bandler, R., 2004).
inal complex (Sp5) (Figure 3). The predominance of
inputs arising from noci-responsive spinal and SpV
regions suggests that active coping in response to
physical stimuli (e.g., acute cutaneous pain) is 42.2.4 Outputs Mediating Active and
mediated by the lPAG. In support of this hypothesis, Passive Emotional Coping
brief applications of a noxious, cutaneous (thermal)
stimulus evoked increased Fos expression within the 42.2.4.1 Lateral periaqueductal gray and
lPAG, hypertension and active coping (Figure 3) ventrolateral periaqueductal gray
(Keay, K. A. and Bandler, R. 1993; 2002). Although they mediate distinct emotional coping stra-
tegies, the same ventromedial and ventrolateral
medullary regions are projected upon by lPAG and
vlPAG (see Figures 3 and 4) (Keay, K. A. and Bandler,
42.2.3 Neural Representation of Passive R., 2001). Consistent with the opposing functional
Emotional Coping roles of lPAG and vlPAG, electrophysiological studies
Passive coping (i.e., a coordinated, conservation- reveal opposite effects on medullary target neurons.
withdrawal reaction of quiescence, hyporeactivity, For example, presympathetic rostral ventrolateral
hypotension, bradycardia, and an opioid-mediated medulla (RVLM) vasopressor neurons are excited
analgesia) is evoked by microinjection of excitatory by lPAG stimulation, but inhibited by vlPAG stimu-
amino acids into the vlPAG column (Figure 1) lation (Verberne, A. J. and Boudier, H. R. S., 1991;
(Bandler, R. and Shipley, M. T., 1994; Keay, K. A. Lovick, T. A., 1992a; 1992b; Verberne, A. and
and Bandler, R., 2001). Guyenet, P., 1992). In contrast to shared descending
targets, there exist distinct ascending projections
42.2.3.1 Ventrolateral periaqueductal to specific hypothalamic and midline and intralami-
gray nar thalamic fields (Floyd, N. S. et al., 1996; 2000;
In common with the lPAG, substantial ascending Krout, K. E. and Loewy, A. D., 2000; Floyd, N. S. et al.,
inputs to the vlPAG originate from superficial and 2001).
deep dorsal horn of spinal cord (and the transition
zone between caudal and interpolar parts of Sp5,
personal observations) (Keay, K. A. et al., 1997). 42.2.4.2 Dorsolateral periaqueductal
Double-label anatomical tracing studies indicate, gray
however, that vlPAG and lPAG projections arise Although it integrates an identical emotional coping
from distinct and separate populations of spinal neu- strategy to the lPAG, the dlPAG has few direct
rons (Clement, C. I. et al., 2000). Further, spino- medullary outputs. Its influence on the ventrolateral
vlPAG projections are not somatotopically organized medulla is likely mediated indirectly, in part via a
(Keay, K. A. et al., 1997). The vlPAG also receives substantial projection to the cuneiform region (see
substantial ascending inputs from nuclei of the soli- Figure 2) (Redgrave, P. et al., 1988; Mitchell, I. J. et al.,
tary tract (NTS), as well as descending inputs from 1988a; 1988b). The dlPAG also projects to distinct
select orbital and insular cortical fields and the lateral thalamic and hypothalamic fields (Floyd, N. S. et al.,
hypothalamus (Figure 4). The convergence within 1996; 2000; Krout, K. E. and Loewy, A. D., 2000;
the vlPAG of spinal, medullary, and forebrain Floyd, N. S. et al., 2001).
 Emotional and Behavioral Significance of PAG 631

Prefrontal cortex
DLO
AId
FPm ACd

PL
VO/VLO m
MO IL
MO

Lateral PAG
dl
Medullary dorsal horn
Hypothalamus lat

PVN
f III
LHAa
AHA 3V Sp5
opt dl

lat

DHA
3V vl
DMH
LHAm DR
f Spinal cord

dl

mt 3V
LHAp PHA

f 3V

vl

RM
RVLM
CMM
CVLM
Medulla

Intermittent (acute) cutaneous pain

(Keay, K. A. and Bandler, R. 1993)

5.8 6.8 7.8 8.8

Figure 3 Schematic illustration of the major afferent and efferent projections of the lateral periaqueductal gray (lPAG). Major
inputs to lPAG arise from spinal cord and the medullary dorsal horn (spinal trigeminal complex (SpV)). In addition, more
modest inputs arise from a restricted part of medial prefrontal cortex as well as dorsal hypothalamic regions. The lPAG
projects directly to both the rostral and caudal ventrolateral medulla (RVLM, CVLM), as well as the rostral and caudal
ventromedial medulla (RM, CMM). The panel in the lower part of the figure shows the pattern of neural activation (c-fos
expression) evoked by an acute cutaneous noxious stimulus (intermittent radiant heat), which evokes active emotional
coping. The shaded area indicates the lPAG column. For abbreviations see caption to Figure 2.
632 Emotional and Behavioral Significance of PAG

Prefrontal cortex

DLO
AId
FPm ACd ACv
PL
VO/VLO m
MO MO IL

Ventrolateral PAG
Hypothalamus dl Nucleus of the solitary tract
PVN
lat
f
LHAa Sol
AHA 3V vl
opt
DR

DHA
3V dl
DMH
LHAm
f Spinal cord

vl
mt 3V
LHAp
PHA

f 3V

RM
RVLM
CMM
CVLM
Medulla

Muscle pain
(Keay, K. A. et al., 2001)

Persistent cutaneous pain 5.8 6.8 7.8 8.8

(Keay, K. A. et al., 2001)

Figure 4 Schematic illustration of the major afferent and efferent projections of the ventrolateral periaqueductal gray
(vlPAG). Major inputs to vlPAG arise from spinal cord and the nucleus of the solitary tract. In addition, significant inputs arise
also from ventromedial prefrontal cortex and orbital/insular cortices. As well, substantial inputs arise from lateral
hypothalamic fields. The vlPAG projects directly to both rostral and caudal ventrolateral medulla (RVLM, CVLM), as well as the
rostral and caudal ventromedial medulla (RM, CMM). The panel in the lower part of the figure shows the patterns of neural
activation (c-fos expression) evoked by an acute deep noxious stimulus (muscle pain: i.m. formalin) and a persistent
cutaneous noxious stimulus (clip applied to dorsum of neck), each of which evoke passive emotional coping. The shaded area
indicates the vlPAG column. For abbreviations see caption to Figure 2.
 Emotional and Behavioral Significance of PAG 633

42.3 Conclusion Floyd, N. S., Price, J. L., Ferry, A. T., Keay, K. A., and Bandler, R.
2000. Orbitomedial prefrontal cortical projections to distinct
longitudinal columns of the periaqueductal gray in the rat.
Animals (including humans) share the capacity to J. Comp. Neurol. 422, 556578.
respond to escapable or inescapable pain with differ- Floyd, N. S., Price, J. L., Ferry, A. T., Keay, K. A., and Bandler, R.
2001. Orbitomedial prefrontal cortical projections
ent emotional coping strategies. It has been to hypothalamus in the rat. J. Comp. Neurol. 432, 307328.
established that the PAG is divisible into distinct Henry, J. P. and Stephens, P. M. 1977. Stress Health and the
longitudinal neuronal columns, which mediate dis- Social Environment: A Sociobiological Approach to
Medicine. Springer.
tinct coping strategies (dlPAG/lPAG: active Keay, K. A. and Bandler, R. 1993. Deep and superficial noxious
emotional coping; vlPAG: passive emotional coping). stimulation increases fos-like immunoreactivity in different
Further, each PAG column lies embedded within regions of the midbrain periaqueductal grey of the rat.
Neurosci. Lett. 154, 2326.
parallel, but distinct circuits, which extend rostrally Keay, K. A. and Bandler, R. 2001. Parallel circuits mediating
to include specific PFC and hypothalamic regions distinct emotional coping reactions to different types of
(Bernard, J. F. and Bandler, J. F., 1998; Keay, K. A. stress. Neurosci. Biobehav. Rev. 25, 669678.
Keay, K. A. and Bandler, R. 2002. Distinct central
and Bandler, R., 2001). On the output side, each PAG representations of inescapable and escapable
column projects either directly (lPAG/vlPAG) or pain: observations and speculation. Exp. Physiol.
indirectly (dlPAG) onto ventrolateral and ventrome- 87, 275279.
Keay, K. A. and Bandler, R. 2004. Periaqueductal Gray. In: The
dial medullary regions containing (1) somatic and Rat Nervous System, 3rd edn. (ed. G. Paxinos), pp. 243257.
autonomic premotor neurons and (2) neurons med- Elsevier.
iating antinociception. A substantial body of evidence Keay, K. A., Clement, C. I., Depaulis, A., and Bandler, R.
2001. Different representations of inescapable noxious
supports the view that the different PAG columns, stimuli in the periaqueductal gray and upper cervical
and, their associated circuits, play critical roles in spinal cord of freely moving rats. Neurosci. Lett.
integrating the somatic, autonomic and antinocicep- 313, 1720.
Keay, K. A., Clement, C. I., Owler, B., Depaulis, A., and
tive components which characterize the distinct Bandler, R. 1994. Convergence of deep somatic and visceral
emotional coping reactions evoked by pain of differ- nociceptive information onto a discrete ventrolateral
ent tissue origins and/or durations. midbrain periaqueductal gray region. Neuroscience
61, 727732.
Keay, K. A., Feil, K., Gordon, B. D., Herbert, H., and Bandler, R.
1997. Spinal afferents to functionally distinct periaqueductal
gray columns in the rat: an anterograde and retrograde
tracing study. J. Comp. Neurol. 385, 207229.
References Krout, K. E. and Loewy, A. D. 2000. Periaqueductal gray matter
projections to midline and intralaminar thalamic nuclei of the
Bandler, R. and Shipley, M. T. 1994. Columnar organization in rat. J. Comp. Neurol. 424, 111141.
the midbrain periaqueductal gray: modules for emotional Lewis, T. 1942. Pain. MacMillan.
expression? Trends Neurosci. 17, 379389. Livingston, W. K. 1998. Pain and Suffering. IASP Press.
Bandler, R., Keay, K. A., Floyd, N., and Price, J. 2000. Central Lovick, T. A. 1992a. Inhibitory modulation of the cardiovascular
circuits mediating patterned autonomic activity during defence response by the ventrolateral periaqueductal grey
active vs. passive emotional coping. Brain Res. Bull. matter in rats. Exp. Brain Res. 89, 133139.
53, 95104. Lovick, T. A. 1992b. Midbrain influences on ventrolateral
Bandler, R., Price, J. L., and Keay, K. A. 2000. Brain mediation medullo-spinal neurones in the rat. Exp. Brain Res.
of active and passive emotional coping. Progr. Brain Res. 90, 147152.
122, 333349. Mitchell, I. J., Dean, P., and Redgrave, P. 1988a. The projection
Bernard, J. F. and Bandler, R. 1998. Parallel circuits for from superior colliculus to cuneiform area in the rat. Ii.
emotional coping behaviour: new pieces in the puzzle. Defence-like responses to stimulation with glutamate in
J. Comp. Neurol. 401, 429436. cuneiform nucleus and surrounding structures. Exp. Brain
Canteras, N. S. and Goto, M. 1999. Fos-like immunoreactivity in Res. 72, 626639.
the periaqueductal gray of rats exposed to a natural Mitchell, I. J., Redgrave, P., and Dean, P. 1988b. Plasticity of
predator. Neuroreport 10, 413418. behavioural response to repeated injection of glutamate in
Clement, C. I., Keay, K. A., Owler, B. K., and Bandler, R. 1996. cuneiform area of rat. Brain Res. 460, 394397.
Common patterns of increased and decreased fos Redgrave, P., Dean, P., Mitchell, I. J., Odekunle, A., and
expression in midbrain and pons evoked by noxious deep Clark, A. 1988. The projection from superior colliculus to
somatic and noxious visceral manipulations in the rat. cuneiform area in the rat. I. Anatomical studies. Exp. Brain
J. Comp. Neurol. 366, 495515. Res. 72, 611625.
Clement, C. I., Keay, K. A., Podzebenko, K., Gordon, B. D., and Verberne, A. and Guyenet, P. 1992. Midbrain central gray:
Bandler, R. 2000. Spinal sources of noxious visceral and influence on medullary sympathoexcitatory neurons and the
noxious deep somatic afferent drive onto the ventrolateral baroreflex in rats. Am. J. Physiol. 263, R24R33.
periaqueductal gray of the rat. J. Comp. Neurol. Verberne, A. J. and Boudier, H. R. S. 1991. Midbrain central
425, 323344. grey: regional heamodynamic control and excitatory amino
Floyd, N. S., Keay, K. A., and Bandler, R. 1996. A calbindin acidergic mechanisms. Brain Res. 550, 8694.
immunoreactive deep pain recipient thalamic nucleus in Wall, P. D. 1979. On the relation of injury to pain. Pain
the rat. Neuroreport 7, 622626. 6, 253264.
634 Emotional and Behavioral Significance of PAG

Further Reading of spinal or vagal afferents evoked distinct patterns of

fos-like immunoreactivity in the ventrolateral periaqueductal
grey of unanaesthetised rats. Brain Res. 948, 122130.
Bandler, R. and Keay, K. A. 1996. Columnar organization in the Lovick, T. and Bandler, R. 2005. The organization of the
midbrain periaqueductal gray and the integration of Midbrain Periaqueductal Grey and the Integration of Pain
emotional expression. Progr. Brain Res. 107, 285300. Behaviour. In: The Neurobiology of Pain (eds. S. Hunt and
Keay, K. A., Clement, C. I., Matar, W. M., Heslop, D. J., M. Koltzenburg), pp. 267287. Oxford University Press.
Henderson, L. A., and Bandler, R. 1997. Noxious activation
 43 The Thalamus and Nociceptive Processing
J O Dostrovsky, University of Toronto, Toronto, ON, Canada
A D Craig, Barrow Neurological Institute, Phoenix, AZ, USA
2009 Elsevier Inc. All rights reserved.

43.1 Introduction 635

43.2 Comparative Anatomical Findings 635
43.2.1 Thalamic Nuclei Receiving Direct Spinothalamic Tract Inputs 636
43.2.1.1 Ventroposterior nuclei 636
43.2.1.2 Posterior part of the ventromedial nucleus 636
43.2.1.3 Ventral lateral nucleus 637
43.2.1.4 Parafascicular nucleus 638
43.2.1.5 Medial dorsal nucleus 638
43.2.1.6 Central lateral nucleus 638
43.2.1.7 Nucleus submedius 638
43.2.2 Indirect Nociceptive Pathways to the Thalamus 638
43.3 Comparative Physiological Findings 639
43.3.1 Ventroposterior Nuclei 639
43.3.2 Ventroposterior Inferior Nucleus 640
43.3.3 Posterior Thalamus and Posterior Part of the Ventromedial Nucleus 640
43.3.4 Intralaminar Nuclei 642
43.3.5 Medial Dorsal Nucleus 643
43.4 Direct Physiological Evidence in Humans 643
43.4.1 Nociceptive Neurons 644
43.4.2 Innocuous Cooling-Responsive Neurons 644
43.4.3 Stimulation-Induced Pain and Temperature Sensations 645
43.5 The Thalamus and Central Neuropathic Pain 647
43.5.1 Physiological Observations in Central Pain Patients 647
43.5.2 Thalamic Bursting Activity 647
43.5.3 Effects of Thalamic Lesions on Pain 647
43.5.4 The Thalamic Pain Syndrome 648
43.6 Pharmacology 649
References 650

43.1 Introduction originated with the comprehensive analysis of 23 tha-

lamic pain patients by Head and Holmes, and it was
The thalamus has been recognized to play a very emphasized particularly by Melzack R. and Casey K. L.
important role in the higher-level processing of noci- (1968). For many years, the available anatomical and
ceptive inputs ever since the clinical observations physiological evidence suggested that these functions
by Dejerine J. and Roussy G. (1906) and Head H. be ascribed to indirect spinoreticulothalamic (paleos-
and Holmes G. (1911) of pain resulting from strokes pinothalamic) input to medial thalamus and direct
affecting the lateral thalamus. Modern-day anatomical (neospinothalamic) spinothalamic tract (STT)-
and electrophysiological techniques have provided a mediated input to somatosensory lateral thalamus,
wealth of information regarding the processing of which was most prominent in humans. However, the
nociceptive information at the thalamic level. recent evidence summarized below indicates that
The concept that medial thalamus is involved in the lamina I STT input to distinct portions of medial and
affective/motivational aspect of pain and the lateral lateral thalamus can be directly associated with these
thalamus in the sensory/discriminative aspect of pain functions.

635
636 The Thalamus and Nociceptive Processing

This chapter will review the major findings ventrocaudal portion of the medial dorsal nucleus
relating to the anatomy, physiology, and pharma- (MDvc) (Mehler, W. R., 1969; Boivie, J., 1979;
cology of thalamus with respect to understanding its Berkley, K. J., 1980; Craig, A. D., 2003; 2004). Each
role in the mediation of acute and chronic pain. The of these is described in more detail below.
chapter will start with an overview of thalamic
anatomy and the termination sites of ascending
somatosensory pathways focusing on the STT in 43.2.1.1 Ventroposterior nuclei
the primate. Then the physiological findings in The VP that comprises the ventroposterior lateral
animals will be summarized, followed by discussions (VPL) and ventroposterior medial (VPM) nuclei
of findings in humans and a summary of thalamic receives its major ascending afferent input from the
pharmacology. medial lemniscus and is the main relay nucleus for
tactile and kinesthetic information. This nucleus is
also frequently termed the ventrobasal complex and
43.2 Comparative Anatomical in the human, the ventrocaudal nucleus (Vc)
Findings (Hassler, R., 1959; Mehler, W. R., 1966; Jones, E. G.,
1990). However, it is well established that the STT
In contrast to spinal cord, there are significant species and TTT also have terminations in this region. The
differences in the anatomy of the pathways and the STT terminations in VP form clusters (also referred
nuclei mediating nociception at the thalamic level. to as archipelago), which are especially dense along
This chapter will focus on findings from primates its caudal border with the posterior group and pulvi-
including humans and will commence with a descrip- nar and along its rostral border with the VL and near
tion of the thalamic termination sites of ascending the laminae that occur within VP (Stepniewska, I.
pathways involved in the transmission of nociceptive et al., 2003). The terminations are roughly in register
information. The major pathway involved in the with the detailed somatotopic organization of the
relay of nociceptive and thermoreceptive informa- low-threshold mechanoreceptive (LTM) lemniscal
tion is the STT and the functionally equivalent terminations and thalamocortical neurons, with cra-
component of the trigeminothalamic tract (TTT) niofacial inputs to VPM, arm/forelimb inputs to
that originates in the medullary dorsal horn (subnu- medial VPL, and leg/hindlimb inputs to lateral
cleus caudalis of the trigeminal spinal tract nucleus, VPL. Some of the STT axons terminating in VP
also termed the medullary dorsal horn). Although the also send a collateral into the CL (Applebaum, A. E.
STT is frequently described as a single tract, this et al., 1979; Giesler, G. J. Jr. et al., 1981).
chapter will describe separately the terminations of The main source of STT and TTT terminations
its two components, the lateral and the ventral. The in VP is from the neurons in the deep dorsal horn
lateral STT originates largely from lamina I of the (laminae IV and V) as shown in Figure 1(b).
superficial dorsal horn and contains many neurons Interestingly, these terminations appear directed to
responding specifically to noxious stimuli and innoc- VP neurons that are immunopositive for calbindin in
uous thermoreceptive neurons. In contrast, the contrast to the lemniscal afferents from the dorsal
ventral STT originates largely from neurons in dee- column nuclei (DCN) and principal trigeminal
per layers, most of which respond to innocuous nucleus, which terminate in the vicinity of neurons
tactile and proprioceptive inputs in addition to noci- that are immunopositive for parvalbumin (Rausell, E.
ceptive inputs (Craig, A. D. and Dostrovsky, J. O., and Jones, E. G., 1991). Furthermore, the calbindin-
1999; Craig, A. D., 2003). immunopositive neurons project to the superficial
layers of cortex, whereas the parvalbumin neurons
project to the middle layers (Gingold, S. I. et al., 1991;
43.2.1 Thalamic Nuclei Receiving Direct
Rausell, E. and Jones, E. G., 1991). Another difference
Spinothalamic Tract Inputs
between spinothalamic and lemniscal terminations is
There are six major regions of termination of the that the latter but not the former form triadic
STT and TTT within the primate thalamus: the synapses with GABAergic presynaptic dendrites
ventroposterior nucleus (VP), the posterior portion (Ralston, H. J., III and Ralston, D. D., 1994). These
of the ventromedial nucleus (VMpo), the ventro- differences suggest that the spinothalamic nocicep-
lateral nucleus (VL), the central lateral nucleus tive inputs are processed differently than the
(CL), the parafascicular nucleus (Pf), and the innocuous mechanoreceptive inputs in VP.
 The Thalamus and Nociceptive Processing 637

(a) (b)

S1
Area 3a
Area 24c

Inter

Inter
nal c Dorsal
posterior

nal c
insula CL
apsu

apsu
MDvc
le

Thalamus Thalamus VPL

le
Lenticular VPI S2 Lenticular
nucleus nucleus S2
VMpo VPI

Midbrain Midbrain
PAG

Pons Pons
PB ??
A6/A7
Medulla Medulla
A1/C1/A5 SRD

Lamina 1 Lamina V
COLD WDR
Lateral STT
HPC LT
NS Anterior STT

Figure 1 A schematic summary diagram of the ascending projections from the spinal cord to the thalamus and on to the
cortex. (a) The lateral spinothalamic tract arising from nociceptive (HPC and NS) and innocuous thermoreceptive (COLD)
neurons in lamina I. The major thalamic termination sites are in the posterior part of the ventromedial nucleus (VMpo),
ventroposterior inferior nucleus (VPI), and the ventrocaudal part of the medial dorsal nucleus (MDvc). Also shown are the
brainstem terminations in the ventrolateral medulla (A1/C1/A5), the dorsolateral pons (A6/A7), the parabrachial nucleus (PB), the
periaqueductal gray (PAG), and the cortical projections of the thalamic sites receiving the lamina I inputs. (b) The ascending
projections of lamina IVV cells, primarily wide dynamic range (WDR) neurons and low-threshold mechanoreceptive (LTM)
neurons, via the ventral spinothalamic tract (STT) to the VPI, ventroposterior lateral nucleus (VPL), ventrolateral nucleus (VL), and
central lateral nucleus (CL). Terminations also occur in the brainstem in the subnucleus reticularis dorsalis (SRD) and other sites,
probably including the reticular core. Adapted with permission from Craig, A. D. and Dostrovsky, J.O. 1999. Medulla to
Thalamus. In: Textbook of Pain (eds. P. D. Wall and R. Melzack), pp. 183214. Churchill-Livingstone.

Immediately ventral to VPL and VPM lies the almost no terminals within VP (Berkley, K. J., 1980;
ventroposterior inferior nucleus (VPI; roughly equiva- Craig, A. D. and Burton, H., 1985).
lent to the parvicellular part of the ventrocaudal
nucleus (Vcpc) in humans). Curiously, this nucleus 43.2.1.2 Posterior part of
receives STT and TTT inputs from neurons located the ventromedial nucleus
not only in laminae IV and V but also in lamina I, as Recent studies in the monkey have revealed that
well as from vestibular afferents (Ralston, H. J., III and there is a very prominent and dense projection from
Ralston, D. D., 1992). The VPI projects to secondary STT and TTT neurons in lamina I to a region that
somatosensory cortex (SII) and retroinsular (vestibu- has been termed the posterior ventromedial nucleus
lar) cortex (Friedman, D. P. and Murray, E. A., 1986; (VMpo), which lies immediately posterior and infer-
Stevens, R. T. et al., 1993). ior to VP and is contiguous rostrally with the VMb
Species differences . In the rat, there are STT and (Craig, A. D., 2004). Of particular interest and impor-
TTT terminations throughout VP (Mehler, W. R., tance are the findings that the STT and TTT
1969; Peschanski, M., 1984), which originate from terminations are topographically organized (Craig,
both superficial and deep layers of the dorsal horn. A. D., 2003; 2004) and that the VMpo neurons project
In contrast, in the cat, the STT and TTT terminate in a topographic manner to the dorsal posterior
along the ventral aspect of VPL (and VPI and the basal insula, a region that is consistently activated by
part of the ventromedial nucleus (VMb)) and there are thermal and nociceptive stimuli. The VMpo is the
638 The Thalamus and Nociceptive Processing

major projection target of the lamina I neurons in 43.2.1.4 Parafascicular nucleus

the primate, which comprise its almost exclusive The centre median (CM) and Pf regions are fre-
ascending sensory input (Figure 1(a)). The tri- quently cited as playing a major role in nociception.
geminal inputs terminate anteriorly and lumbar However, there is only a weak STT projection to Pf
inputs most posteriorly. This anteroposterior topo- that originates from lamina I and V cells, and more
graphic arrangement contrasts with the mediolateral recent anatomical studies fail to confirm earlier
topography of the VP. reports of STT terminations in CM. The Pf and
The dense lamina I STT and TTT terminations CM nuclei appear to be involved in motor-related
in VMpo are clearly delineated with the use of processing as their main connections are with the
immunohistochemical labeling for calbindin. A basal ganglia, substantia nigra, and motor cortex
region of calbindin-positive terminal labeling has (Jones, E. G., 1985; Royce, G. J. et al., 1989; Sadikot,
been observed in a comparable location in the A. F. et al., 1992).
human thalamus (Blomqvist, A. et al., 2000) and cor-
responds with the area of dense STT terminations 43.2.1.5 Medial dorsal nucleus
observed in thalamus following cordotomies per- Recent studies in the monkey reveal moderately
formed to alleviate pain (Mehler, W. R., 1966). This dense STT and TTT projections to the MDvc.
region can also be delineated in monkeys and humans The terminals are topographically organized along
based on its different cytoarchitecture. Electron an anteroposterior axis, with trigeminal input located
microscopy has revealed that the glutamatergic most posteriorly. The cells of origin of this projection
STT and TTT terminations form triadic synapses are in lamina I of the spinal and medullary dorsal
with the VMpo relay cell dendrites and GABAergic horn (Ganchrow, D., 1978; Craig, A. D., 2004). Cells
presynaptic dendrites (Beggs, J. et al., 2003). This is in MDvc project to area 24c in the cortex at the
similar to the termination of lemniscal afferents in fundus of the anterior cingulate sulcus (limbic
VP but contrasts with the STT terminations in VP. motor cortex), rather than to the orbitofrontal and
The triadic contacts are believed to provide high prefrontal cortex where the remainder of medial dor-
synaptic security and temporal fidelity. VMpo was sal nucleus (MD) projects (Ray, J. P. and Price, J. L.,
not specifically identified in earlier anatomical stu- 1993; Craig, A. D., 2003). It is likely that this region of
dies, but the region was included in the caudal VP MD plays an important role in mediating the
(Mehler, W. R., 1966), the Vc portae (Hassler, R., affective/motivational aspect of pain. There appears
1970), and the posterior complex (Po) ( Jones, E. G., to be no homologous region in the cat or rat (see
1990). Section 43.2.1.7).
Species differences. In the cat, there is a sparse
lamina I terminal field in the ventral VMb, and this 43.2.1.6 Central lateral nucleus
may constitute a primordial homologue of the pri- Dense STT terminations are observed in the caudal
mate VMpo. In support of this notion is the fact that part of CL and in some other portions of the nucleus.
lesions to this part of VMb in the cat disrupt discri- The cells of origin are located in laminae V and VII
minative thermal sensation (Norrsell, U. and Craig, (Applebaum, A. E. et al., 1979; Giesler, G. J., Jr. et al.,
A. D., 1999) and that it projects to insular cortex 1981; Craig, A. D., Jr. et al., 1989). Projections from
(Clasca, F. et al., 1997). A homologous region does different regions of the spinal cord terminate in
not appear to exist in the rat. different cell clusters but do not appear to be topo-
graphically organized (Craig, A. D. and Burton, H.,
43.2.1.3 Ventral lateral nucleus 1985). This nucleus also receives projections from the
Moderately dense STT terminations are observed cerebellum, tectum, substantia nigra, and globus pal-
extending rostrally from VP into caudal VL in cats lidus. Most of the neurons in this region project to the
and monkeys that overlap with inputs from the cer- basal ganglia, but there are also projections to the
ebellum (Berkley, K. J., 1980; Stepniewska, I. et al., superficial and deep layers of posterior parietal and
2003). These inputs probably originate from neurons motor cortices (Jones, E. G., 1985; Royce, G. J. et al.,
in the deep dorsal horn and ventral horn (laminae V 1989). The function of the STT input to this nucleus
and VII). VL projects to the motor cortex (Jones, E. is unknown, but one can speculate that it may be
G., 1985), and this STT component is most likely involved in motor set, attention, and orientation.
related with sensorimotor integration rather than Similar STT terminations have been observed in
nociception. cats, rats, and other vertebrates.
 The Thalamus and Nociceptive Processing 639

43.2.1.7 Nucleus submedius dorsolateral funiculus and terminate in the lateral

In the cat, there are dense and topographically orga- cervical nucleus that is located at the level of C1-2
nized terminations of the TTT and STT arising from just lateral to the superficial dorsal horn (Boivie, J.,
spinal and medullary dorsal horn lamina I neurons in 1983). The neurons of the lateral cervical nucleus
the medial thalamic nucleus submedius (Sm). project to the contralateral VP via the medial lem-
Although Sm originates developmentally from the niscus. This pathway is prominent in the cat and
pronucleus of MD, it projects to the ventrolateral raccoon but very small in the primate. Some of the
orbital cortex rather than to the cingulate cortex and neurons in this pathway respond to nociceptive sti-
thus constitutes a major phylogenetic difference from muli although most respond only to innocuous
the lamina I projection to MDvc in the primate stimuli (Kajander, K. C. and Giesler, G. J., Jr., 1987;
(Craig, A. D., Jr. et al., 1982; Coffield, J. A. et al., Downie, J. W. et al., 1988). Third, evidence obtained
1992). In the cat, the spinal input to the anterior in the rat indicates that spinal input to a dorsal
cingulate relays in the ventral VP (Musil, S. Y. and reticular region in the caudal medulla is relayed by
Olson, C. R., 1988; Yasui, Y. et al., 1988) and also way of a portion of the ventromedial nucleus to
indirectly by way of the parabrachial nucleus (PB) layer 1 of widespread regions of the frontal cortex
(Devinsky, O. et al., 1995). In the rat, input to Sm (Desbois, C. et al., 1999; Desbois, C. and Villanueva,
originates primarily from trigeminal cells at the junc- L., 2001). This region also has a descending projec-
tion of the caudalis and interpolaris subnuclei of the tion to the deep dorsal horn of the spinal cord.
trigeminal spinal tract nucleus and from trigeminal Modulation of activity, such as those associated
and cervical lamina I cells, as well as other cells in the with alerting responses, is suggested as a potential
spinal cord (Dado, R. J. and Giesler, G. J., Jr., 1990; function, but whether this pathway exists in
Yoshida, A. et al., 1992). primates and humans is unknown. Fourth, a spinor-
eticulothalamic pathway (paleospinothalamic tract)
was hypothesized long ago to accommodate clinical
43.2.2 Indirect Nociceptive Pathways to
observations made in patients with thalamic pain
the Thalamus
syndrome (see below). Modern evidence indicates
In addition to the STT, there are several polysy- that spinal terminations in the brainstem reticular
naptic pathways that may be involved in relaying formation do not overlap with the locations of cells
nociceptive signals to the thalamus. First, the post- that project to thalamus, except within the PB
synaptic dorsal column pathway comprises dorsal (Blomqvist, A. and Berkley, K. J., 1992). The PB
horn neurons located primarily in laminae IVVI receives contralateral spinal input and ipsilateral
and X whose axons ascend ipsilaterally in the super- trigeminal input that originates mainly from lamina
ficial dorsolateral funiculus and deep dorsal columns I but also from lamina V cells. There is only a crude
and terminate in the ventral and rostral portions topography, and although there is considerable ana-
of the DCN (gracile and cuneate nuclei). These tomical overlap with ascending homeostatic afferent
portions of the DCN have projections to motor- input from the nucleus of the solitary tract, evidence
related regions of the brainstem rather than to VP in the rat indicates that nociceptive neurons are
(Brodal, A., 1982; Willis, W. D., 1985; Berkley, K. J. mainly modality selective (Bernard, J. F. and
et al., 1986). Although most of the neurons in this Besson, J. M., 1990). In primates, PB projects mainly
pathway respond only to nonnoxious cutaneous to VMb in the thalamus, as well as to hypothalamus
mechanical stimuli, some respond also to noxious and amygdala, but in rats, it projects more broadly
stimuli. Some of the nociceptive neurons in this within medial thalamus and to various cortical
pathway convey visceral activity in the rat and regions associated with autonomic control.
possibly also in the primate (Al-Chaer, E. D. et al.,
1998; see also Villanueva, L. and Nathan, P., 2000).
However, functional imaging studies of visceral pain 43.3 Comparative Physiological
in humans reveal a pattern of activation that does Findings
not appear consistent with the hypothesis that
this pathway is important for visceral pain in The previous section described the pathways and
humans (e.g., Strigo, I. A. et al., 2003). Second, the thalamic termination sites of spinal and trigeminal
spinocervicothalamic tract comprises dorsal horn neurons that include nociceptive neurons. On this
neurons whose axons ascend ipsilaterally in the basis, one would expect to find neurons in the
640 The Thalamus and Nociceptive Processing

thalamic termination sites that would also respond to LTM neurons (Willis, W. D. and Westlund, K. N.,
noxious inputs, and this is particularly the case for 1997). In the monkey, these neurons are concentrated
the regions receiving inputs from lamina I. There in the posterior part of VP and near the major fiber
have indeed been many reports of nociceptive neu- laminae, consistent with the predominant location of
rons in lateral, posterior, and medial areas of the the terminations of the STT and TTT axons of
thalamus. However, it should be kept in mind that lamina V neurons (Applebaum, A. E. et al., 1979;
the existence of neurons responding to noxious sti- Boivie, J., 1979; Kenshalo, D. R., Jr. et al., 1980; Casey,
muli in a given region of thalamus does not K. L. and Morrow, T. J., 1983; Gingold, S. I. et al., 1991;
necessarily signify that the region is involved in Bushnell, M. C. et al., 1993). Anatomical studies have
mediating the sensations of pain that would be provided evidence suggesting that the nociceptive
expected to be elicited by noxious stimuli giving neurons project to areas 3b and 1 of SI cortex.
rise to the noxious responses. For example, the Interestingly, these studies suggest that the cortical
responses could be related to arousal, attentional, terminations of these nociceptive neurons are in the
and motor consequences of the noxious stimulus. most superficial layers of cortex in contrast to the VP
The following section will summarize the physiolo- LTM neurons whose axons terminate in the middle
gical findings reported for each of the main thalamic layer of cortex (Rausell, E. et al., 1992; Shi, T. et al.,
regions receiving nociceptive inputs. 1993), indicating a possible modulatory role for these
nociceptive inputs. Electrophysiological studies have
confirmed that some of these WDR neurons within
43.3.1 Ventroposterior Nuclei
VP can be antidromically activated from areas 3b and
As mentioned earlier, the VPL and VPM are the main 1 of SI (Kenshalo, D. R., Jr. et al., 1980). Curiously,
thalamic targets for innocuous tactile information visceral noxious stimuli activate not only VP WDR
ascending via the medial lemniscus from the DCN neurons but also LTM neurons and do not appear to
and principal trigeminal nucleus. It is thus not sur- be somatotopically organized (Chandler, M. J. et al.,
prising that most of the neurons in this region respond 1992; Al-Chaer, E. D. et al., 1998).
exclusively to innocuous low-threshold mechanical Species differences: In the rat, nociceptive-specific
cutaneous inputs. These neurons are organized in a (NS) and WDR nociceptive cells have been reported
high-resolution somatotopic fashion and project pri- throughout VP intermixed in a roughly topographic
marily to areas 3b and 1 of primary somatosensory manner with the LTM neurons (Guilbaud, G. et al.,
(SI) cortex. There is an anterior and dorsally located 1980). The receptive fields of these nociceptive neu-
shell that contains neurons responding to deep, mus- rons are generally quite large and often bilateral. Some
cle and joint afferent inputs, and this proprioceptive receive convergent visceral input. In rats with experi-
information is relayed to areas 3a and 2 of SI cortex mentally induced arthritis or neuropathies, increased
(Kaas, J. H. et al., 1984; Jones, E. G., 1990). numbers of WDR VP cells that project to the sensori-
It is generally considered that VP is the main relay motor cortex have been reported (Guilbaud, G. et al.,
nucleus for nociceptive inputs involved in mediating 1990). In contrast to the primate and rodent, in the cat,
the sensory aspects of pain (localization and intensity there are virtually no nociceptive cells within VP
discrimination) (Melzack, R. and Casey, K. L., 1968; proper. However, NS and WDR nociceptive neurons
Willis, W. D., 1985), since it receives STT inputs in this species can be found in the dorsal and ventral
(primates and rodents, but not carnivores) and projects aspects of VP, the ventral aspect of VMb and VPI.
to the somatotopically organized SI cortex. Indeed, These are the regions where STT and TTT termina-
there have been many reports of nociceptive neurons tions occur (see above). The nociceptive neurons
within VP in the primate and rodent. In the monkey, within this shell-like region are organized in a crude
about 10% of the neurons are nociceptive and most of mediolateral topographic pattern in parallel (but not in
these are of the wide dynamic range (WDR respond- register) with the somatotopy of the adjacent VP. A
ing to both LTM inputs and nociceptive inputs) type portion of these neurons receive convergent input from
(Figure 2) (Willis, W. D., 1985; Apkarian, A. V. and skin, muscle, viscera, tooth pulp, or cranial vasculature
Shi, T., 1994; Willis, W. D. and Westlund, K. N., 1997). (Davis, K. D. and Dostrovsky, J. O., 1988; Bruggemann,
They usually have moderately large receptive fields J. et al., 1994). Some of these neurons have been shown
(e.g., more than half of the face or arm), and these are to project to area 3a of SI, SII, and/or anterior cingulate
organized in a crude somatotopic manner roughly cortex (Yasui, Y. et al., 1988; Craig, A. D. and
corresponding to the somatotopy of the intermingled Dostrovsky, J. O., 2001).
 The Thalamus and Nociceptive Processing 641

160
(a) (c) Squeeze

140
120
Pinch

100
Impulses s1
(b)

80
LD

60
LP Press.
MD
CL Brush

40
VPLC

20
CMPf
P

0
0 20 40 60 80 100 120
Time (s)
1 mm

(d) 35 C
(f) 35 50 C

(e) 50 C 1 ms
5s

Figure 2 Example of responses of a wide-dynamic-range (WDR) neuron in ventroposterior nuclei (VP). The receptive field
on the leg is shown in (a). The location of the recording site in VPLc (VP) is indicated in (b). The histogram (c) shows the
responses to graded intensities of mechanical stimulation. This neuron also responded to noxious thermal stimulation (d, e,
and f). Modified with kind permission from Kenshalo, D. R., Jr., Giesler, G. J., Jr., Leonard, R. B., and Willis, W. D. 1980.
Responses of neurons in primate ventral posterior lateral nucleus to noxious stimuli. J. Neurophysiol. 43, 15941614.

43.3.2 Ventroposterior Inferior Nucleus various species to contain nociceptive neurons

(Albe-Fessard, D. et al., 1985; Willis, W. D., 1985;
The neurons in the region located immediately infer-
Willis, W. D., Jr., 1997). Although it was formerly
ior (ventral) to VPL and VPM (VPI) project to SII
regarded as undifferentiated, more detailed and
(Krubitzer, L. et al., 1995) and to retroinsular (vestib-
recent observations in monkeys and humans have
ular) cortex. Some of the neurons in this region are
described a distinct nucleus that has been termed
excited by nociceptive inputs (Figure 3). These
VMpo. The VMpo has been shown to be the recipi-
WDR and NS neurons are arranged in a topographic
ent of a major projection from lamina I STT and
representation within this region in the monkey
TTT neurons. In accordance with its lamina I inputs,
(Apkarian, A. V. and Shi, T., 1994).
it has been shown to contain NS and thermoreceptive
neurons (Craig, A. D. et al., 1994). Their responses are
similar to those of lamina I STT and TTT neurons,
43.3.3 Posterior Thalamus and Posterior
and they have small receptive fields that are topo-
Part of the Ventromedial Nucleus
graphically organized in a rostrocaudal axis, in
The posterior thalamus has frequently been impli- correspondence with the topographically organized
cated in pain processing and has been shown in terminations of the afferent inputs (Figure 4).
642 The Thalamus and Nociceptive Processing

(a)
80 Brush press. pinch
VPL
Spikes s1

40
VPM

VPl
0
0 60 120
Time (s)

(b)
180 Brush press. pinch
Spikes s1

VPL

90

VPM

VPl
0
0 60 120
Time (s)

(c)
120 Brush press. pinch
Spikes s1

Pulo VPL

60

Po

MG
0
0 60 120
Time (s)
Figure 3 Example of responses of nociceptive neurons in ventroposterior inferior nucleus (VPI) and posterior complex (Po).
Reproduced with kind permission from Apkarian, A. V. and Shi, T. 1994. Squirrel monkey lateral thalamus. I. Somatic
nociresponsive neurons and their relation to spinothalamic terminals. J. Neurosci. 14, 67796795.

Neuronal recordings in awake monkeys in the pre- studies have consistently found this region of insular
sumed VMpo (originally assumed to be in VPM but cortex to be activated by noxious and thermal stimuli.
subsequently re-interpreted from the original histol- This rostrocaudally organized topography is distinct
ogy to be in VMpo (Craig, unpublished observations) from the mediolateral topography observed in SII
revealed strong correlation between the neuronal and in the adjacent parietal operculum (Disbrow, E.
activity and the behavioral detection of noxious and et al., 2000). There is also a minor projection from
innocuous thermal stimuli applied to the face. VMpo to area 3a of SI cortex.
Furthermore, lidocaine-induced block of activity in Species differences. In the cat and rat where there is
this region reduced the monkeys behavioral no clearly differentiated nociceptive and thermore-
responses to these stimuli (Bushnell, M. C. et al., ceptive relay site in the Po, there are nevertheless
1993; Duncan, G. H. et al., 1993). nociceptive neurons, but they tend to have very large
Further support for the critical role of VMpo in receptive fields and are not arranged in a clear soma-
relaying nociceptive and thermoreceptive informa- totopic fashion (Poggio, G. F. and Mountcastle, V. B.,
tion for perception of pain and temperature is the 1960; Carstens, E. and Yokota, T., 1980). This region
finding that it has a strong and topographic projection does receive some lamina I inputs in the cat. In the
to the dorsal margin of the posterior insula (Craig, A. rat, there is convergent STT input from laminae I
D., 2003), where preliminary fMRI observations in and V to the posterior triangular nucleus, which
monkeys show strong activation with noxious stimuli projects to the region of SII and to the amygdala. It
(Craig, A. D., 2002). Furthermore, human imaging is possible that in the rat, nociceptive inputs also arise
 The Thalamus and Nociceptive Processing 643

125 Noceptive single unit

100

Spikes s1
75

50

25

0
0 100 200 300 400
Time (s)

34 39 43 46 50 54
Temperature (C)
Figure 4 Example of the responses of a posterior part of the ventromedial (VMpo) nociceptive neuron. The histogram
shows the graded responses of a single nociceptive-specific neuron to noxious heat pulses applied with a thermode applied
to the receptive field on the contralateral ulnar hand. Adapted with kind permission from Craig, A. D., Bushnell, M. C., Zhang,
E. -T., and Blomqvist, A. 1994. A thalamic nucleus specific for pain and temperature sensation. Nature 372, 770773.

via a descending corticothalamic route (Diamond, M. nociceptive neurons in this region have large recep-
E. et al., 1992; Berkley, K. J. et al., 1993). Furthermore, tive fields that can be bilateral. Studies employing
there is no direct projection to the portion of insular graded natural stimuli have revealed that some of the
cortex homologous to that targeted by VMpo in the cells can code for stimulus intensity. Although nat-
primate. The part of Po dorsal to VP in the cat, where ural stimulation was used in only some studies, cells
some nociceptive neurons have been reported with graded responses to noxious heat have been
(Hutchison, W. D. et al., 1992; Bruggemann, J. et al., observed. In particular, a study in the awake monkey
1994), projects to area 5a (Jones, E. G., 1985). by Bushnell M. C. and Duncan G. H. (1989) reported
Nevertheless, in both the cat and the rat, there exists the existence of nociceptive neurons in the CMPf
a narrow region along the ventral aspect of VMb that and CL region that responded in a graded fashion to
contains nociceptive and visceroceptive neurons and noxious thermal stimuli applied to the face
projects to the insular cortex adjacent to the gusta- (Figure 5). It is possible that the responses of at
tory cortex (Yasui, Y. et al., 1991; Clasca, F. et al., 1997; least some of these neurons may be related to atten-
Norrsell, U. and Craig, A. D., 1999). tion and arousal rather than to perception of pain. It
is interesting that many intralaminar neurons have
been shown to respond to eye movements (Schlag, J.
43.3.4 Intralaminar Nuclei and Schlag-Rey, M., 1986), which is consistent with
the termination of ascending inputs from the cere-
As described earlier, several regions in medial thala- bellum and the superior colliculus. Indeed, it has
mus receive direct inputs from the spinal cord and been proposed that this region plays a role in gaze
the trigeminal nucleus, but there are also indirect orientation (Jones, E. G., 1985).
inputs to these regions from the brainstem, and in
particular from the PB.
Neurons responding to noxious electrical,
43.3.5 Medial Dorsal Nucleus
mechanical, or heat stimuli have been recorded
throughout the intralaminar thalamus, in particular As mentioned earlier, Craig and colleagues have
in CL and CMPf. Most of these recordings have recently described a direct lamina I input to the
been obtained in anesthetized rats and cats although MDvc in the monkey. A particular interesting feature
some recordings have been performed in monkeys of this projection is that the MDvc contains
(Albe-Fessard, D. et al., 1985; Bushnell, M. C. et al., neurons that project to the anterior cingulate cortex
1993; Bruggemann, J. et al., 1994). Most of the (Craig, A. D. and Zhang, E.-T., 1996), a region
644 The Thalamus and Nociceptive Processing

(a) (b)
40 80 47 C 47.2 C + 47.4 C

35 60

Spikes s1
Mean spikes s1( SE) 40
30

20
25
0
20

0
46 47 48 49 0 2 4
C T2 s RE

Figure 5 Responses of a neuron in posteromedial thalamus of an awake monkey to noxious thermal stimuli to the face. (a)
The mean firing rate to increasing temperatures. (b) A peristimulus histogram with associated dot raster display showing
increased firing when the temperature increased from 47  C to 47.2  C or 47.4  C. In this particular example, the monkey did
not respond until the end of the trial (RE button release). Adapted with kind permission from Bushnell, M. C. and Duncan, G. H.
1989. Sensory and affective aspects of pain perception: is medial thalamus restricted to emotional issues. Exp. Brain Res. 78,
415418.

frequently implicated in pain. There is only very (Dostrovsky, J. O. and Guilbaud, G., 1988). Various
limited information regarding the properties of the studies in the rat that have examined the effects of
neurons in MDvc; however, preliminary studies in lesioning or electrically and chemically induced excita-
anesthetized monkeys indicate that it contains a dis- tion of Sm have provided evidence for a role of Sm and
crete group of NS neurons with large, sometimes its cortical target VLO in the activation of descending
bilateral receptive fields (Craig, A. D., 2003). antinociceptive pathways by way of the periaqueductal
Furthermore, this study reports that their sponta- gray (PAG) (Roberts, V. J. and Dong, W. K., 1994;
neous firing can be inhibited by innocuous thermal Zhang, S. et al., 1998). Although it has been shown in
(cool, warm) stimuli. This phenomenon could pro- the cat that there is also a projection from innocuous
vide an explanation for the well-known cold-induced thermoreceptive cool lamina I STT neurons in addi-
inhibition of pain and the thermal grill illusion of tion to the NS neurons (Craig, A. D. and Dostrovsky, J.
pain (see below). It is possible that some of the O., 2001), neurons excited by cooling stimuli have not
recordings of nociceptive neurons in medial thalamus been found. It is possible, however, that inputs from
in the study of Bushnell M. C. and Duncan G. H. these neurons may provide a basis for the cold-induced
(1989) mentioned above may also have included inhibition of nociceptive processing, similar to MDvc of
neurons located in MDvc. the primate (Ericson, A. C. et al., 1996).
Species differences. Whereas in the monkey (and
presumably the human), the STT and the TTT
project to MDvc, in the cat and the rat, there is a
projection that originates mainly in lamina I to the 43.4 Direct Physiological Evidence in
developmentally related Sm. Several studies have Humans
reported the existence of NS neurons in Sm in the
rat (Dostrovsky, J. O. and Guilbaud, G., 1988; Functional stereotactic surgery for the treatment of
Coffield, J. A. and Miletic, V., 1993; Kawakita, K. chronic pain or tremor provides a unique opportu-
et al., 1993). Receptive fields are generally quite nity to record and stimulate in the thalamus of awake
large and some have inputs from deep tissues. In the patients. There have been several studies that have
Freunds adjuvant-induced arthritic rat, many Sm cells provided information of interest in terms of further-
were found to respond to joint movements, which are ing our understanding of the role of thalamus in pain
normally not effective in activating the Sm neurons and these are described below.
 The Thalamus and Nociceptive Processing 645

43.4.1 Nociceptive Neurons thalamus. It is difficult to evaluate these old reports

since few details were provided and more recent stu-
Lenz and colleagues have searched for neurons in VP
dies have not been able to confirm the findings (Lenz,
(often termed Vc in humans) and the immediately
F. A. et al., 1997). However, there have been very few
adjacent posterior and inferior region of thalamus.
opportunities to record from neurons in medial thala-
Although most of the neurons in Vc respond only to
mus, and the number of neurons that can be tested with
nonnoxious mechanical stimulation of the skin (LTM
noxious stimuli in the awake patient is limited, and this
neurons), they reported that some neurons also had an may account for the lack of definitive evidence.
increased response to noxious mechanical stimuli and
some responded weakly to noxious or innocuous ther-
mal stimuli in addition to innocuous tactile stimuli (i.e.,
WDR neurons) (for review, see Lenz, F. A. and 43.4.2 Innocuous Cooling-Responsive
Dougherty, P. M., 1997; Lee J. et al., 1999). These Neurons
WDR neurons were primarily located in the poster- Neurons responding selectively to innocuous cooling
oinferior portion of VP. In the adjoining posteroinferior of the skin have been found in the human thalamic
area, which includes VMpo (Blomqvist, A. et al., 2000), region located medial and posteroinferior to VP, which
they found NS neurons that responded to noxious heat likely corresponds to VMpo (Davis, K. D. et al., 1999).
but no LTM neurons (Lenz, F. A. et al., 1993a). These neurons had discrete receptive fields on the
Although nociceptive neurons have frequently been contralateral body and responses similar to those of
reported in medial thalamus of anesthetized animals, lamina I spinal and trigeminal neurons (Figure 6(a)).
there have only been a few old reports of neurons This finding is consistent with the findings in animals,
responding to noxious stimuli in awake human medial where cooling-specific neurons have only been found

(a1) (b)
Spikes 100 ms1 Temperature (C)

30
10
20

10 8

8
Cool rating

6 6
4
2
4
0
10 s
(a2)
2
35
10
0

0 2 4 6 8 10 12 14 20 30 40 50
Stimulus intensity (A)
1s
Figure 6 (a1) Recording from a single neuron in the presumed posterior ventromedial nucleus (VMpo) region in an awake
patient showing responses to cooling stimuli applied to receptive field on the fifth digit. The top trace shows the temperature
of the thermode with increasing cooling steps. The bottom trace is a histogram of the neuronal firing showing graded
responses to the increasing cooling steps. (a2) Segment of raw trace of neuronal recording from (a1) showing response to first
part of a cooling step. (b) Thalamic stimulation evoked cool sensations. Verbal ratings (010 scale) of the innocuous cool
sensations evoked by threshold and suprathreshold intensities of thalamic microstimulation obtained in eight patients.
Figurines adjacent to each line depict the location of the thalamic stimulation-evoked sensation at threshold. Reprinted with
kind permission from Davis, K. D., Lozano, R. M., Manduch, M., Tasker, R. R., Kiss, Z. H., and Dostrovsky, J. O. 1999.
Thalamic relay site for cold perception in humans. J. Neurophysiol. 81, 19701973.
646 The Thalamus and Nociceptive Processing

in the cat medial VPM and in the monkey VMpo. opportunity to determine the sensations evoked
Animal studies have also shown that cooling-specific by electrical stimulation within the brain, since
neurons are found only in lamina I of the trigeminal these are performed with the patient awake. As
medullary dorsal horn (nucleus caudalis) and spinal might be expected, electrical stimulation (1 s trains
dorsal horn. Thus, the existence of cooling-specific of 100300 Hz) within VP and adjacent regions of
neurons in the human VMpo is consistent with the the thalamus usually evokes innocuous paresthesia.
evidence cited above that this region receives a major These sensations, which can be elicited by low
input from lamina I neurons. The close association of stimulus intensities (e.g., 2 mA), are perceived as
pain and temperature further supports an important originating from a small region of the contralateral
role of VMpo in the relay of pain signals in the human. side of the body (Figure 7). The projected fields
are usually in register with the receptive fields of
the neurons recorded at the stimulation site.
43.4.3 Stimulation-Induced Pain and
Increasing the intensity of stimulation results in
Temperature Sensations
an increase in the perceived intensity of sensation
The electrophysiological studies in human patients and usually also with an increase in the projected
during stereotactic surgery provide a unique field size, but although the sensation can be very

Depth lntensity
RF PF
(mm) (A)

2.0 P 0 P P P, N
5 5 10 20

1.5 P P P P Sharp
6 12 24 48 jabbing
0.6 P N
12 24
1.3

Sharp
1.0 P jabbing
1.0 N
10 14
Sharp Sharp
1.2 N N
3 10
Sharp
0.6 1.4 N jabbing, deep
5
Quick
1.6 P N and
Deep 9 12 sharp
0.3 P P
8 16
1.8 P
7

N = pain
P = paresthesia

Figure 7 Stimulation-evoked pain in a chronic pain patient (causalgia left arm). The diagram shows a reconstruction of an
electrode track through the ventrocaudal nucleus and on into the posteroinferior region, showing the locations of the receptive fields
(RF) of the low-threshold mechanoreceptive neurons recorded at sites along the electrode track on the left of the vertical line and the
projected fields (PF) induced by stimulation on the right side. The electrode track has been broken down into two contiguous
segments. Depths of recordings are indicated in millimeters to the left of the line at each recording/stimulation site. While the
electrode was in ventrocaudal nucleus (Vc), as evidenced by the low-threshold mechanoreceptive fields, stimulation induced
nonpainful paresthesia (P). However, at the bottom of the Vc (0) and at deeper sites, stimulation induced painful sensations (N).
 The Thalamus and Nociceptive Processing 647

intense, it is not usually reported as painful. 43.5 The Thalamus and Central
Nevertheless, stimulation at some sites in some Neuropathic Pain
patients is reported as eliciting a distinct painful
43.5.1 Physiological Observations in
and/or temperature sensation even at the threshold
Central Pain Patients
for sensation (Hassler, R. and Riechert, T., 1959;
Halliday, A. M. and Logue, V., 1972; Tasker, R. R., There have been many studies that have demonstrated
1984; Lenz, F. A. et al., 1993b; Davis, K. D. et al., the marked plasticity of the brain and in particular the
1996; Dostrovsky, J. O. et al., 2000). These sites are alterations in neuronal properties and somatotopic
generally located near the posteriorinferior border organization at all levels of the somatosensory system
of VP and extend several millimeters posterior, subsequent to peripheral or central damage to the
inferior, and medial to it (Figure 7). The incidence ascending pathways (Wall, J. T. et al., 2002). Although
of sites where pain and/or thermal sensations can much less is known about the plasticity of the pain
be evoked is much higher in the posteroinferior system, especially at the thalamic and cortical levels, it
region than within VP (except in poststroke pain is reasonable to expect that marked changes will occur
patients see below). The sensations are always on following damage to ascending nociceptive pathways.
the contralateral side of the body and can arise Indeed, there have been several animal studies that
from a small region. The sensations are reported examined the effects of STT damage on thalamic
as being quite natural in contrast to the paresthesia physiology (Weng, H. R. et al., 2000; 2003). Studies in
(tingling or shock-like) sensations evoked within patients who have sustained deafferentation due, for
VP. There have been reports of cases where tha- example, to amputation or spinal cord injury provide
lamic stimulation evoked pain arising from deep or evidence for plasticity (Lenz, F. A. et al., 1994; Davis, K.
visceral sites although these are rare (Lenz, F. A. D. et al., 1998). For example, there is evidence suggest-
et al., 1994; Davis, K. D. et al., 1995). Interestingly, ing expansion of the representation of intact regions
Lenz and colleagues found that microstimulation into deafferented regions of VP thalamus. Micro-sti-
within VP at sites where WDR neurons responding mulation in such regions was found to evoke sensations
to noxious mechanical stimuli were found rarely arising from the phantom limb in amputees or to the
elicited pain or temperature sensations, whereas at deafferented body region in spinal injury patients
the sites in the region posteroinferior to VP where (Lenz, F. A. et al., 1994; Davis, K. D. et al., 1998).
stimulation evoked pain, there was a high likeli- Although these stimuli usually resulted in nonpainful
hood of finding nociceptive neurons (Lenz, F. A. paresthesia rather than pain, these types of changes
and Dougherty, P. M., 1997). It seems likely that may be involved in the development of chronic pain
the sites in the region posteroinferior to VP where in these types of patients. Of particular interest have
stimulation evokes pain and temperature are close been the findings of two studies that found changes in
to or in VMpo. It is also notable that at the sites the stimulation-evoked sensations in poststroke pain
where neurons responding selectively to cooling and some other types of chronic central pain patients.
stimuli were found, microstimulation evoked cool- The incidence of stimulation sites in VP where stimu-
ing sensations arising from the same area of skin lation evoked sensations in such patients was markedly
where the receptive fields of the neurons were elevated (Lenz, F. A. et al., 1998; Dostrovsky, J. O. et al.,
located (Figure 6(b)). Increasing the stimulus inten- 2000). In addition, there was an increase in the number
sity at such sites increased the intensity of the cold of sites in the posteroinferior region where pain sensa-
sensation (i.e., it felt colder). tions were evoked and a decrease in sites where
There have been few studies of the effects of innocuous thermal stimuli were evoked (Figure 8).
stimulation in medial thalamus. A few studies These findings provide further evidence suggesting
reported that stimulation in the posterior aspect of that there have been alterations in the thalamic and
medial thalamus can evoke pain (Sano, K., 1979; cortical processing of somatosensory inputs leading to
Jeanmonod, D. et al., 1994). However, in most cases increased perception of pain. It is interesting, however,
the stimuli were delivered from large-tipped electro- that deep brain stimulation within thalamus (Siegfried,
des and the sensations were only elicited at high J., 1987; Kupers, R. C. and Gybels, J. M., 1993) can
current intensities, so current spread (e.g., to the alleviate the pain in some of these chronic pain
STT) is an issue. Several more recent studies have patients, possibly by disrupting the pathological pat-
failed to replicate these findings. terns and balance of activity in these regions.
648 The Thalamus and Nociceptive Processing

(a) (b)
30 25
Motor (n = 345) Motor (n = 1642)
NSP (n = 790)

% of total stimulation sites in

NSP (n = 156)
% of total stimulation sites in
25 20 PSP (n = 388)
PSP (n = 73)

postero inferior region

P < 0.0001

P < 0.0001
20
Vc nucleus

15 P < 0.0001

15 P < 0.001
P < 0.001
10
P < 0.001
10

5
5

0 0
Pain Temperature Pain Temperature
Figure 8 Bar graphs showing the incidence of sites in the ventrocaudal nucleus (a) and the posteroinferior region (b) where
stimulation evoked sensations of pain or innocuous temperature in movement disorder patients (Motor), chronic
nonpoststroke pain patients (NSP), and poststroke pain patients (PSP). Reprinted with kind permission from Dostrovsky, J.
O., Manduch, M., Davis, K. D., Tasker, R. R., Lozano, A. M. 2000. Thalamic Stimulation-Evoked Pain and Temperature Sites in
Pain and Non-pain Patients. In: Proceedings of the 9th World Congress on Pain (eds. M. Devor, M. C. Rowbotham, and Z.
Wiesenfeld-Hallin), pp. 419425. IASP Press.

43.5.2 Thalamic Bursting Activity 1994). Studies in the primate have found that follow-
ing chronic cervical rhizotomy there is a
Thalamic neurons switch to a bursting mode when
downregulation of thalamic GABAA receptors
they are hyperpolarized. This bursting activity is gen-
(Rausell, E. et al., 1992), providing some support for
erated by activation of low-threshold T-type calcium
this suggestion. On the other hand, several positron
channels that give rise to a calcium spike and a burst of
emission tomography (PET) studies have obtained
sodium channel-generated action potentials (so-called
evidence suggesting that chronic neuropathic pain is
LTS bursting). This bursting activity is generally only
associated with decreased blood flow, suggesting
observed during sleep. However, recordings in medial
decreased activity (Apkarian, A. V. et al., 2005).
and lateral thalamus of chronic pain patients during
the awake state revealed the existence of neurons
firing in typical calcium spike-mediated bursts (Lenz,
F. A. et al., 1989; Jeanmonod, D. et al., 1993; Lenz, F. A. 43.5.3 Effects of Thalamic
et al., 1994). It was proposed that this activity may be Lesions on Pain
related to the patients chronic pain and in fact may There have been many studies since the original
mediate the spontaneous pain in these patients (Lenz, pioneering studies of Dejerine and Roussy and
F. A. and Dougherty, P. M., 1997; Llinas, R. R. et al., Head and Holmes which have documented that
1999). However, similar LTS firing in similar regions strokes affecting the lateral thalamus can lead to
of thalamus can also be observed in patients without central pain (poststroke pain and thalamic pain syn-
pain and thus is not specific to pain ( Jeanmonod, D. drome) (Pagni, C. A., 1998). In addition, infarcts and
et al., 1996; Radhakrishnan, V. et al., 1999). More lesions that involve the posteroinferior region can
recently, Llinas R. R. et al. (1999) have proposed that also result in analgesia and thermanesthesia (Head,
LTS bursting in thalamus may be the cause of many H. and Holmes, G., 1911; Hassler, R. and Riechert,
different neurological diseases including chronic pain T., 1959; Hassler, R., 1970; Tasker, R. R., 1984).
and have termed this condition thalamic dysrhythmia. Lesions have been purposefully made in medial tha-
Nevertheless, the role of LTS bursting in chronic lamus of chronic pain patients for relief of their pain
central pain remains unclear. (Gybels, J. M. and Sweet, W. H., 1989). However,
It has been proposed that central neuropathic pain such procedures are rarely undertaken at the present
may be mediated by reduced GABAergic inhibition time as most neurosurgeons do not believe that they
in the thalamus (Roberts, V. J. and Dong, W. K., are effective. It is possible, however, that in some of
 The Thalamus and Nociceptive Processing 649

the reported successes the lesions may have included 43.6 Pharmacology
MDvc and/or VMpo (Jeanmonod, D. et al., 1994;
Lenz, F. A. and Dougherty, P. M., 1997). The pharmacology of thalamic nociceptive neurons
Interestingly, lesions in medial thalamus that spare has not been extensively studied. However, it is rea-
the lateral thalamus and STT do not appear to cause sonable to assume that they share many common
central pain (Bogousslavsky, J. et al., 1988). features with those of other thalamic neurons
about which much more is known. The two most
43.5.4 The Thalamic Pain Syndrome important transmitters are glutamate and GABA
(McCormick, D. A., 1992; Steriade, M. et al., 1997;
The poststroke central (thalamic) pain (CPSP) syn- Sherman, S. M. and Guillery, R. W., 2001). Gluta-
drome was first recognized by Dejerine and Roussy, mate is released by the ascending axons of medial
and it has been comprehensively analyzed clinically lemniscal and STT and TTT pathways to excite
by Head H. and Holmes G. (1911), Riddoch G. and thalamocortical neurons. The released glutamate
Critchley Mc. D. (1937), Schott B. et al. (1986), Boivie acts at both NMDA and non-NMDA (AMPA and/
J. (1994), Pagni C. A. (1998), and others. The first or kainate) ionotropic glutamate receptors. In the
patients had lesions confined to the thalamus, and so cat and primate, there are also many interneurons
the syndrome was first called thalamic pain. within the thalamus and these also receive glutama-
However, patients with central pain have lesions tergic inputs from these afferents that activate in
that interrupt the ascending lateral STT lamina I addition to the ionotropic receptors and metabotro-
spinothalamocortical pathway by way of VMpo to the pic glutamate receptors (Jones, E. G., 1985;
dorsal posterior insula at any level (Schmahmann, J. D.
Magnusson, K. R. et al., 1987; Ericson, A. C. et al.,
and Leifer, D., 1992; Pagni, C. A., 1998; Craig, A. D.,
1995; Blomqvist, A. et al., 1996; Sherman, S. M. and
2003). Such lesions produce loss of pain and tempera-
Guillery, R. W., 2001). The thalamocortical neurons
ture sensation, but in about half of such cases, this
also receive a massive glutamatergic projection from
disruption results, immediately or after a variable
layer 6 of the cortex, which activates both ionotropic
delay, in the paradoxical appearance of ongoing
and metabotropic receptors (Rustioni, A. et al., 1983;
pain in the deafferented region. Head and Holmes
Jones, E. G., 1987; Deschenes, M. and Hu, B., 1990;
inferred that this meant that pain sensation occurred
Eaton, S. A. and Salt, T. E., 1990; 1996; Salt, T. E. and
in the thalamus, but we now know that the thalamus
serves mainly as a relay for activity ascending to the Eaton, S. A., 1996). The thalamocortical neurons
cortex. This syndrome likely results from disruption release glutamate at their cortical terminals
of the interactions at cortical and subcortical levels (Kharazia, V. N. and Weinberg, R. J., 1994; Pirot, S.
between components of the pain processing network. et al., 1994).
One such interaction is the inhibition caused by cool- GABA is the major inhibitory neurotransmitter in
ing on pain, and it has been suggested that the thalamus, and the major source of these inputs
interruption of this interaction could cause central is from the thalamic reticular nucleus (TRN).
pain by disinhibition (Craig, A. D., 1998). The sug- The TRN GABAergic neurons are topographically
gestion is based on similarities with the thermal grill organized and receive excitatory inputs from collat-
illusion of pain, in which reduced activity in cooling- erals of thalamocortical and corticothalamic neurons.
specific lamina I STT neurons can unmask the cold- In cats and primates, but not rats, there are also
activated burning pain elicited by polymodal noci- GABAergic interneurons within the thalamus
ceptive lamina I STT neurons (HPC) in the anterior (Houser, C. R. et al., 1980; Rustioni, A. et al., 1983;
cingulate cortex. A study of thermal sensation in a Jones, E. G., 1985; Steriade, M. et al., 1997). The TRN
central pain patient directly supports that hypothesis produces both short- and long-latency inhibitory
(Morin, C. et al., 2002). A recent imaging study of a responses; the short-latency IPSPs are mediated
central pain patient suggested that the posterolateral by GABAA receptors, whereas the long-latency
thalamic lesion did not involve VMpo but that con- responses are mediated by the GABAB metabotropic
clusion was incompatible with the patients loss of receptors (Lee, S. M. et al., 1994). Interestingly, acti-
thermal sensation (cf. Craig, A. D. et al., 2000; Montes, vation of presynaptic glutamate metabotropic
C. et al., 2005). The indirect ascending nociceptive receptors reduces the release of GABA from the
input to the medial thalamus by way of the PB may TRN receptors (Salt, T. E. and Eaton, S. A., 1995).
also play a role in central pain. Most of the studies of thalamic GABAergic
650 The Thalamus and Nociceptive Processing

mechanisms have involved nonnociceptive neurons; can depress the responses of thalamic nociceptive
however, studies in Pf have shown that thalamic neurons (He, L. F. et al., 1991; Coffield, J. A. and
nociceptive neurons receive inhibitory inputs from Miletic, V., 1993) and produce antinociception
the TRN and are inhibited by GABA (Reyes- when microinjected into Sm (Yang, Z. J. et al., 2002).
Vazquez, C. and Dafny, N., 1983; Jia, H. et al., 2004), Furthermore, the use of radiolabeled -opioid ago-
and microinjection of the GABAA antagonist bicucul- nists with PET has shown high opiate receptor levels
line or agonist muscimol into Sm enhanced or in the human thalamus (see review in Apkarian, A. V.
depressed, respectively, the antinociception induced et al., 2005), and evidence for their involvement in
by the prior microinjection of morphine into the rat endogenous pain modulation.
Sm (Jia, H. et al., 2004). There is also recent evidence
for modulation of GABAB receptors in VP and Po by
noxious inputs (inflammation) (Ferreira-Gomes, J.
et al., 2004). References
The thalamic relay neurons as well as the TRN
Al-Chaer, E. D., Feng, Y., and Willis, W. D. 1998. A role for the
and inhibitory interneurons also receive projections dorsal column in nociceptive visceral input into the thalamus
from serotoninergic neurons in the dorsal raphe of primates. J. Neurophysiol. 79, 31433150.
nucleus, and norardrenergic and cholinergic neurons Albe-Fessard, D., Berkley, K. J., Kruger, L., Ralston, H. J., III,
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in the peribrachial region (see reviews by sensation. Brain Res. 356, 217296.
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identified in thalamic afferents and neurons. For innervation of the rat and cat thalamus. I. Distribution and
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fibers ending in the rat and human thalamus Craig, A. D. 2003. Synaptology of trigemino- and
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Low dose of morphine strongly depresses responses of
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Nishiyama, K. et al., 1995). the rat. Pain 15, 333344.
It is usually assumed that the analgesia produced Berkley, K. J. 1980. Spatial relationships between the
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brainstem levels. However, there is increasing evi- sensory inputs to lateral diencephalon. J. Comp. Neurol.
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Berkley, K. J., Budell, R. J., Blomqvist, A., and Bull, M. 1986.
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opioids on thalamic and cortical nociceptive neurons Res. Bull. 11, 199225.
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1993. Responses of neurons in and near the thalamic
Opioid receptors and enkephalinergic terminals ventrobasal complex of the rat to stimulation of uterus,
have been identified in the thalamus (Mansour, A. cervix, vagina, colon, and skin. J. Neurophysiol.
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Bernard, J. F. and Besson, J. M. 1990. The
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does not prove that effect is due to an action at the processes. J. Neurophysiol. 63, 473490.
Blomqvist, A. and Berkley, K. J. 1992. A re-examination of the
thalamic level. However, there have been studies that spino-reticulo-diencephalic pathway in the cat. Brain Res.
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 44 Psychophysics of Sensations Evoked by Stimulation
of the Human Central Nervous System
S Ohara, C A Bagley, H C Lawson, and F A Lenz, Johns Hopkins Hospital, Baltimore, MD, USA
2009 Elsevier Inc. All rights reserved.

44.1 Introduction 656

44.2 The Spinothalamic Tract 656
44.3 The Dorsal Column Pathway 657
44.4 Thalamic Nuclei 657
44.4.1 Lateral Thalamic Nuclei 657
44.4.2 Medial and Intralaminar Thalamic Nuclei 663
44.5 Cortex 663
44.5.1 Parasylvian Cortex and Pain Memory 663
44.6 Conclusions 665
References 665

Glossary
dorsal columns A pathway carrying axons of somatic sensory nuclear group of the
fibers in the peripheral nerve through the posterior thalamus Nuclei receiving input from the dorsal
aspect of the spinal cord to the dorsal column column nuclei, the spino- or the trigeminothalamic
nucleus at the posterior aspect of the medulla tract by either a direct or a transsynaptic route.
where they synapse and ascend through the medial Summary term for the specific somatosensory
lemnisus to the principal nucleus of thalamus. nuclei of the thalamus. Most commonly used tax-
Classically, this pathway transmits low-threshold onomy is for monkey thalamus: VPL
muscle and cutaneous afferents but recent studies (ventroposterolateral), VPM (ventroposteromedial),
demonstrate that afferents transmitting VPI (ventroposteroinferior) corresponding to
nociceptive, thermal, or visceral inputs are also humans: Vc (ventrocaudal, both medial and lateral).
transmitted. Adjacent, both posterior medial, to VPM ventral
nociceptive specific (NS) neurons Neurons medial posterior (VMpo), a putative pain and tem-
responding only to stimuli which are noxious or perature signaling nucleus may be located (Craig,
painful. A. D. et al., 1994, cf. Willis, W. D., Jr. et al., 2001,
wide dynamic range (WDR) neurons Neurons Graziano, A. and Jones, E. G., 2004; Lenz, F. A.
responding to stimuli across the intensive conti- et al., 2004). VMpo may be located medial to ventral
nuum into the painful or noxious range. caudal portae (Vcpor), while intralaminar nuclei
projected field in the somatosensory system A mostly a thin layer of neurons located medial to
part of the body at which a sensation is evoked in Vim, Vc, and Vcpor (see Figure 1(a)).
response to stimulation of the central nervous spinothalamic tract (STT) The pain and tem-
system. perature signaling pathway from neurons in the
receptive field In the somatosensory system: skin spinal dorsal horn to the lateral and anterolateral
region, stimulation of which can influence the dis- funiculi of the spinal cord and the brainstem to the
charge of the neuron under study lateral, posterior, medial, and intralaminar nuclei of
thalamus.

655
656 Psychophysics of Sensations Evoked by Stimulation of the Human Central Nervous System

spinothalamocortical pathways Cortical areas parietal, and temporal opercula are folds of cortex
which receive input from thalamic nuclei that that extend over the insula and meet each other
themselves receive STT input. These thalamocor- over the insula. The sulcus between the frontal
tical connections include those from Vc to primary parietal opercula and the temporal operculum is the
somatosensory and secondary somatosensory sylvian fissure.
cortex, from Vcpor, Vcpc, and VMpo to parietal Talairach coordinates Initially developed for a
opercular, insular, and retroinsular cortex, and specific stereotactic frame; based on one single
finally from the medial dorsal nucleus to anterior brain; frequently used as common coordinate sys-
cingulate cortex. tem; x: leftright, y: anteriorposterior, z: superior
sylvian fissure Also called lateral sulcus, the syl- inferior; the reference point (0, 0, 0) is the
vian fissure separates the temporal lobe from the anterior commissure (Talairach, J. and Tournoux,
frontal and parietal lobes (see Figure 6, left middle P., 1988).
and lower levels). The insula is a cortical surface, thalamus A subcortical gray matter structure
which is located deep to the lateral fissure and characterized by reciprocal connections with the
parallel to the cortical surface on coronal and axial cortical mantle and by inputs from the periphery
sections (Figure 6, middle and right). The frontal, and nuclei of the brain, such as the pallidum.

44.1 Introduction continuum into the noxious range (WDR wide

dynamic range). These cells arise from both superficial
The dorsal column pathway and the spinothalamic (lamina I) and deep laminae of the dorsal horn (lami-
tract (STT) are the two main somatosensory spinal nae IVV) (Kumazawa, T. and Perl, E. R., 1978; Willis,
tracts afferent to the thalamus. The dorsal column W. D., 1985; Ferrington, D. G. et al., 1987). Some
pathway is formed by the axons of low-threshold neurons in the superficial dorsal horn respond only
mechanoreceptors with cell bodies in the dorsal root to different stimuli including noxious stimuli (noci-
ganglion. These axons terminate to the principal ceptive specific (NS) cells), cold (Kumazawa, T. and
somatic sensory nucleus of the thalamus (human ven- Perl, E. R., 1978; Willis, W. D., 1985; Ferrington, D. G.
tral caudal or Vc, monkey ventral posterior or VP) et al., 1987; Craig, A. D. et al., 1994), injection of
(Jones, E. G. et al., 1982; Kaas, J. H. et al., 1984; Lenz, histamine (itch Andrew, D. and Craig, A. D., 2001;
F. A. et al., 1988; Willis, W. D. and Coggeshall, R. E., Craig, A. D., 2003b), or visceral stimuli (Craig, A. D.,
1991). The dorsal columns do contain a postsynaptic 2003a). The deep and superficial laminae of the dorsal
pain pathway signaling noxious visceral stimuli, which horn project to the brain, respectively, through the
terminates in the Vc (Uddenberg, N., 1968; Rustioni, A. ventral lateral (ventral STT) and the dorsal lateral
et al., 1979; Willis, W. D., et al., 1991; Al Chaer, E. D. spinal funiculi (dorsal STT) (Apkarian, A. V. and
et al., 1996b). The STT is a pain pathway that originates Hodge, C. J., 1989; Cusick, C. G. et al., 1989; Ralston,
from the spinal dorsal horn, and ascends in the ante- H. J. and Ralston, D. D., 1992; Craig, A. D., 1998,
rolateral quadrant of the spinal cord before terminating 2003b; Price, D. D. et al., 2003).
in the thalamus (Willis, W. D., Jr. et al., 2001). The population of neurons in the superficial dorsal
Stimulation of these thalamic nuclei in man reveals horn responding only to noxious stimuli has led to the
the psychophysical dimensions of these pathways description of the STT as a structure signaling only
which is the subject of this chapter.
pain a labeled line (Perl, E. R., 1984; Willis, W. D.,
1985). A recent version of this hypothesis suggests that
the STT is a series of labeled lines for cool and itch, as
44.2 The Spinothalamic Tract well as pain that jointly reflect the internal state of the
body (interoception) (Craig, A. D., 2003a; 2003b). In
Many neurons in the STT are characterized by their this view, pain is the emotion produced by disequili-
response to noxious or painful stimuli. Some of these brium of the internal state. An alternate view is that
respond to the somatic stimuli across the intensive pain is signaled by WDR neurons which transmit a
 Psychophysics of Sensations Evoked by Stimulation of the Human Central Nervous System 657

graded signal, the strength of which might be decoded antidromic invasion from nucleus gracilis (Al Chaer,
in the brain to identify the presence of a painful stimu- E. D. et al., 1996a), and by retrograde and anterograde
lus (Price, D. D. and Dubner, R., 1977; Willis, W. D., tracer studies (Christensen, M. D. et al., 1996).
1985; Price, D. D. et al., 2003). This chapter examines Retrograde tracer studies depositing marker in the
recent evidence from stimulation of the human central nucleus gracilis (Christensen, M. D. et al., 1996) label
nervous system as they impact these hypotheses. cell bodies of origin of the pathway in the medial base
One approach to identifying the pain pathway has of the dorsal horn just above the central canal.
been to stimulate the cord during cordotomy using Injection of tracer into this area resulted in fiber label-
paired-pulse stimulus parameters that selectively ing of the dorsal column midline and in the medial
activate the axons arising in the superficial or deep aspect of the nucleus gracilis. Fibers originating in the
laminae (Mayer, D. J. et al., 1975). This interpulse thoracic cord terminate in the lateral part of nucleus
threshold was more consistent with that of cells in gracilis and adjacent to the medial parts of nucleus
the deep than the superficial dorsal horn (Price, D. D. cuneatus (Wang, C. C. et al., 1999; Willis, W. D. et al.,
and Mayer, D. J., 1975). The results suggest that the 1999). Along with the STT, a major terminus of the
sensory aspect of pain is signaled through the axons postsynaptic dorsal horn pathway is the lateral
in ventral STT which originate from the WDR neu- thalamus.
rons in the deep dorsal horn (Price, D. D. and
Dubner, R., 1977; Dubner, R. et al., 1989).
44.4 Thalamic Nuclei
44.4.1 Lateral Thalamic Nuclei
44.3 The Dorsal Column Pathway
The human principal sensory nucleus (Vc) (Hassler,
The dorsal column pathway is formed by the axons of R., 1959) is divided into a core area (equivalent to
low-threshold mechanoreceptors that project through monkey VP, see Olszewski, J., 1952; Hirai, T. and
the dorsal column nuclei and medial lemniscus to the Jones, E. G., 1989), posterior, and inferior regions.
region of the principle somatic sensory nucleus (ven- These two regions are defined relative to the most
tral caudal, Vc) (Jones, E. G. et al., 1982; Kaas, J. H. et al., posterior and inferior cell with a response to non-
1984; Lenz, F. A. et al., 1988; Willis, W. D. et al., 1991). painful, cutaneous stimuli (cell 57 in Figure 1(b)). In
As shown in Figure 1 receptive fields are quite con- the core, the majority of cells respond to innocuous,
stant within a particular parasagittal plane in Vc. From mechanical, and cutaneous stimulation. This latter
medial to lateral planes, the sequence of neuronal area corresponds to the posterior and inferior sub-
cutaneous receptive fields progresses from intraoral nuclei of Vc, which are ventral caudal portae
through face, thumb, fingers (radial to ulnar), and (Vcpor), ventral caudal parvocellular nucleus
arm to leg. Proximal parts of the limbs are represented (Vcpc) (Mehler, W. R., 1966), the posterior nucleus,
dorsal to the corresponding digits (Lenz, F. A. et al., and the magnocellular medial geniculate (Mehler, W.
1988). R., 1962; Mehler, W. R., 1966; Lenz, F. A. et al., 1993b)
Substantial evidence demonstrates that the dorsal (see Vc and Vcpor in sagittal section in Figure 1(a)).
columns do contain a postsynaptic pain pathway Studies of patients at autopsy following lesions of the
signaling noxious visceral stimuli, rather than nox- STT show terminations in all this nuclei (Bowsher,
ious somatic stimuli, which terminates in the VP D., 1957; Mehler, W. R., 1966; Mehler, W. R., 1969).
(Uddenberg, N., 1968; Rustioni, A. et al., 1979; This area includes the ventral medial nucleus pos-
Willis, W. D. et al., 1991; Al Chaer, E. D. et al., terior part (VMpo), which may receive STT inputs
1996b). This postsynaptic pathway has been demon- and may signal pain and temperature (Craig, A. D.
strated by infusion of neurotransmitter agonists/ et al., 1994; Blomqvist, A. et al., 2000; cf. Willis, W. D.,
antagonists into the dorsal horn, which alters the Jr. et al., 2001; Graziano, A. et al., 2004). The physiol-
response of neurons in the dorsal column nuclei to ogy recapitulates this anatomy.
visceral stimuli. Furthermore, lesions of rat nucleus Cells in Vc responding to painful and thermal
gracilis diminish the response of neurons in VP to stimuli are of several types, including WDR and
visceral as well as cutaneous stimuli (Al-Chaer, E. D. NS cells responding to painful thermal and mechan-
et al., 1997). ical stimuli (see figure 2 in Lee, J.-I. et al., 1999).
The cells of origin of this pathway are located just Figure 2(a) demonstrates the response of a single
dorsal to the central canal, as demonstrated by neuron with WDR properties to a painful 45  C
658 Psychophysics of Sensations Evoked by Stimulation of the Human Central Nervous System

(a) (b)

P2 P1
Z (+)
P1 Core
P2 Vim Posterio
5 r
Vc
10
35
Vop Cool 15
40
20
Vcpor ACPC line
45 50 1 mm
Y (+) 25
AC-PC line PC PC
55
(0, 0)
60
Inferio
r
5 mm
P2 P1
(c)

PF RF PF RF PF RF PF RF
3032 50 1 1516
NR
33 51 2 17
NR NR
34 52 3 18
Tremor 5
related
3536 4 19
53
NR NR
5 5 20
37 54
5
NR

3841 6 5 21
Cool 55
NR NR
42 5 Cool 56 7 22
Tremor
NR related
10
43 8 23
Tremor 5 57
related NR
44 30 9 24
Tremor 58
related
10
45 10 Cool 25
Cool 59
NR
5 4647 5 10
11 Cool 26
NR 60
Tremor
NR related
48 15 12 5 Cool 27
61

49 5
13 28
15 NR
5
14 5 29
NR NR
5
 Psychophysics of Sensations Evoked by Stimulation of the Human Central Nervous System 659

45 C (VAS 3) stimulus. Responses to painful stimuli were charac-

(a)
terized by the mean firing rate during painful
stimulation, recorded through the microelectrode
50/s
(Figure 2(b), x-axis), and by visual analog scale of
1s intensity (VAS) ratings of microstimulation-evoked
pain (y-axis). Some low-threshold cells respond to
(b) Wide dynamic range (WDR)
nonpainful mechanical and cold stimuli (Figure 2)
5 (Lenz, F. A. et al., 1993a; Lenz, F. A. and Dougherty,
P. M., 1998; Lee, J.-I. et al., 1999). Cells in the core and
4
posterior region respond only to noxious heat stimuli
VAS

3 (Lenz, F. A. et al., 1993a; 2004) and to noxious cold

2
stimuli (Davis, K. D. et al., 1999).
Nociceptive cells in Vc appear to signal pain
1 based on temporary lesioning and stimulation stu-
0 10 30 50 70
dies. Blockade of the activity in this region by
Nociceptive specific (NS) injection of local anesthetic into monkey VP, corre-
5 sponding to human Vc (Hirai, T. et al., 1989),
4 * significantly interferes with the monkeys ability to
p = 0.03
discriminate temperature in both the innocuous and
VAS

3 * noxious range (Duncan, G. H. et al., 1993).

VAS (mean SEM)
2 WDR Stimulation within Vc and the regions posterior and
NS inferior to it can evoke the sensations of pain
1 (Hassler, R. and Reichert, T., 1959; Willis, W. D.,
0 10 30 50 70
Response-baseline (Hz)
1985; Dostrovsky, J. O. et al., 1991; Lenz, F. A. et al.,
1993b) and temperature (Lenz, F. A. et al., 1993b;
Figure 2 Activity of neurons in the region of Vc (ventrocaudal)
Davis, et al., 1999).
responding to painful thermal stimuli. The response to
nonpainful and painful heat/mechanical stimuli applied within The largest study of stimulation-evoked pain and
the receptive field (RF) (Lenz, F. A. et al., 1994b; Lee, J.-I. et al., temperature responses examined results of threshold
1999) is compared with the visual analog scale of intensity (VAS) microstimulation of the region of Vc in 124 thalami
evoked by the same stimulus. (a) The response of a cell wide (116 patients), as summarized in Figure 3. The loca-
dynamic range (WDR) to painful heat. (b) VAS and firing rates for
the response to painful stimuli are plotted for nociceptive (NS)
tion of pain and temperature responses is defined
cells which respond only to painful stimuli and WDR neurons, relative to the posterior and inferior borders of the
which respond in a graded fashion to nonpainful and painful principal somatic sensory nucleus (Vc). Warm sensa-
stimuli. Average and one standard deviation scores by decade tions were evoked more frequently in the posterior
to 20 Hz and by 30 Hz steps from 20 to 80 Hz were compared by
region (5.7%) than in the core (2.3%). Otherwise the
MannWhitney U test. Reprinted from Lenz F. A., Ohara, S.,
Gracely, R. H., Dougherty, P. M., and Patel, S. H. 2004. Pain proportions were not significantly different for cool
encoding in the human forebrain: binary and analog or pain sensations between the core or the posterior
exteroceptive channels. J. Neurosci. 24, 6540-6544 with region or both (cool 2.5%, 2.2%; pain 2.8%, 4.1%).
permission 2004 by the Society for Neuroscience).

Figure 1 Map of receptive and projected fields for trajectories in the regions of the Vc (ventrocaudal) in a single patient
(number 193.97). (a) Positions of the trajectories relative to nuclear boundaries as predicted radiologically from the position of
the anterior commissureposterior commissure (ACPC) line. The ACPC line is indicated by the horizontal line in the panel;
the trajectories are shown by the two oblique lines. The positions of nuclei are inferred from the ACPC line and therefore are
only an approximate indicator of nuclear location. Scale as indicated. Abbreviations defined in the text. (b) Location of the
cells, stimulation sites, and trajectories (P1 and P2) relative to the ACPC line (a solid line) and the ventral border of the core of
Vc (a dotted line). The locations of stimulation sites are indicated by ticks to the left of the trajectory; the locations of the cells
are indicated by ticks to the right of the trajectory. Cells with receptive fields (RFs) are indicated by long ticks; those without
are indicated by short ticks. The cold sensation evoked is indicated by filled circles at the end of the tick to the left of the
trajectory. Scale is as indicated. Each site where a cell was recorded or stimulation was carried out or both is indicated by the
same number in (b) and (c). The core of Vc and the regions posterior and inferior are as labelled. (c) P1 and P2 show the site
number, PF, and RF for that site. The threshold (in microamperes) is indicated below the PF diagram. Reprinted from Ohara S.
and Lenz F. A. (2003) Medial lateral extent of thermal and pain sensations evoked by microstimulation in somatic sensory
nuclei of human thalamus. J. Neurophysiol. 90, 2367-2377, used with permission from the American physiological society.
660 Psychophysics of Sensations Evoked by Stimulation of the Human Central Nervous System

(a) Dorsal These latter studies took the anterior commissure

posterior commissure line (ACPC) as the floor of Vc,
z
Anterior contrary to atlas and physiologic maps ( Schaltenbrand,
10
G. and Bailey, P., 1959; Lenz, F. A. et al., 1988). This
recent study suggests that sites where thermal or pain
sensations are evoked are located both within and
y
10 10
posterior, inferior, and medial to Vc. If the proportion
of such sites is larger in posterior and inferior regions,
then those sites must be very close to the borders of the
core.
10 1 mm Patterned stimulation at sites in the region of Vc,
Paresthesia No response an STT terminal region, evokes sensations consistent
(b) with one of two pathways one binary (pain) and
Cool the other analog (pain/) (Figure 3). Specifically,
Warm current was applied at five frequencies (10, 20, 38,
Painful 100, and 200 Hz) in bursts of 4, 7, 20, 50, and 100
pulses in an ascending staircase protocol, the type of
protocol commonly used in studies of pain (Gracely,
R. H. et al., 1988; Yarnitsky, D. and Sprecher, E.,
1994), including our studies (Greenspan, J. D. et al.,
2004). Stimulation at pain sites evoked a constant
high level of pain over large, often cutaneous, pro-
jected fields (PFs). These sites were characterized
both by descriptors, which did not change along the
staircase, or by more intense stimulation-evoked pain
than that evoked at the pain/ sites (Figure 4).
These results suggest that pain sites participate in
a binary, exteroceptive, labeled line which signals the
1 mm
presence of a painful external stimulus.
The thalamic stimulation thresholds for nonpainful
Figure 3 Locations of sites where microstimulation and painful sensations are not significantly different
evoked paresthesias (a, left), no response (NR) (a, right), and (Lenz, F. A. et al., 1993b; Ohara, S. et al., 2003) suggest-
thermal and pain sensations (b). Site location is shown ing that pain/ sites did not result from activation
relative to the posterior and inferior borders of the core of
Vc. Note that thermal and pain sensations were evoked both
of the system transmitting nonpainful sensations (lar-
in the core and posterior regions of Vc. Paresthesic sites are gely medial lemniscal) before that transmitting painful
most dense where NR sites are least dense over the core sensations (largely STT) (Willis, W. D., 1985). In
and the posterior regions. Scale as indicated. Reprinted addition, the equivalence of current, pulse, and fre-
from Ohara S. and Lenz F. A. 2003. Medial lateral extent of quency thresholds for pain at both types of sites
thermal and pain sensations evoked by microstimulation in
somatic sensory nuclei of human thalamus. J. Neurophysiol.
predicts that the neural elements, that is, possibly
90, 2367-2377, used with permission from the American WDR and NS cells, should be activated together if
Physiological Society. they were found at the same site. At such sites, analog
pain responses would be predicted to occur because
the combination of binary plus analog neural elements
Warm sensations were evoked more frequently in the at a site will have analog properties, given the assump-
lateral plane (10.8%) than in the medial planes of the tion of linearity. However, analog pain sites were not
posterior region (3.9%) but no other significant med- observed. For all these reasons, it is plausible that our
ial lateral differences for any sensation were found in observations may be the result of selective activation
the core or posterior region or overall. of two functionally distinct pathways.
These results are in contrast to previous studies Figure 2(a) demonstrates the mean firing rate
reporting a larger proportion of thermal/pain sites minus baseline (Figure 2(b), x-axis) for the response
were evoked in the posterior and inferior regions of multiple single WDR and NS neurons to painful
(Lenz, F. A. et al., 1993b; Davis, K. D. et al., 1996). stimuli versus VAS score (Figure 2(b), y-axis). There
 Psychophysics of Sensations Evoked by Stimulation of the Human Central Nervous System 661

Pain+ Pain/+

(a) RF PF RF PF
12.9 mm 13.0 mm 12.4 mm 13.0 mm
300 Hz, 5 A 300 Hz, 5 A
Unnatural Natural
Surface & deep Deep
Vibration Movement from eye
Painful (7/10) brow to corner of eye
Flicker Painful (9/10)
Electric Hot
200 Hz Same Same Same 200 Hz Warm Warm Warm Hot 8/10
100 Hz Same 100 Hz Tingle Warm Hot 8/10
38 Hz Same Same Same 38 Hz Tingle Warm Warm Hot 8/10
20 Hz 20 Hz Tingle Warm Warm
10 Hz 10 Hz Tightness Warm Warm
5 A 4 Pulses 7 20 50 100 20 A 4 Pulses 7 20 50 100

(b)
VAS VAS
5 5
Pain

0 0
Nonpain

5 5
100 100
50 50
10 20 10 20
se

se
10 7 10 7
20
l

l
Pu

Pu
38 4 20 38 4
100 200 100
Frequen Frequen 200
c y (Hz) cy (Hz)

Figure 4 Pain and pain/ stimulation sites. Sensations evoked by threshold microstimulation were characterized by the
PF and by descriptors from a validated questionnaire, and by a visual analog scale of intensity (VAS) (Lenz, F. A. et al., 1993b;
1998a; Lenz, F. A. and Byl, N. N. 1999). (a, left) Site where stimulation at 300 Hz and 5 mA produced pain in the PF shown in the
figurine and of the quality as described. Pain identical to that evoked by 300 Hz was evoked at most sites with trains 20
pulses and frequencies 200 Hz (yellow rectangle, see text). (a, right) Site where tightness was evoked in the first column at
10 Hz and then tingle at 20, 38, and 100 Hz. At 200 Hz and thereafter, warm was evoked at each step in the staircase,
excepting 7 pulses 10, 20, 100 Hz, until 50 pulses 38 Hz. At this step and further up, the staircase painful heat was evoked.
(b) Average VAS ratings across all pain and pain/ sites. Ratings were taken in response to pulse and frequency pairs
ascending the staircase. The yellow lines along the outside surfaces of the 3D displays indicate the average VAS ratings
across all sites by frequency and number of pulses. Reprinted from Lenz F. A., Ohara, S., Gracely, R. H., Dougherty, P. M.,
and Patel, S. H. 2004. Pain encoding in the human forebrain: binary and analog exteroceptive channels. J. Neurosci. 24,
6540-6544, with permission (2004; by the Society for Neuroscience).

was a significantly steeper initial rise in VAS scores for Zaslansky, R. et al., 1995). The second pathway may
the neurons that only responded to painful stimuli be an analog route in which activity is graded with
(NS neurons), than for WDR neurons. The steep intensity of the painful stimulus, consistent with STT
initial rise of VAS with the firing rate of NS versus neurons, which encode the properties of external sti-
WDR neurons (Figure 2(b)) is consistent with the muli (Willis, W. D., 1985; Price, D. D. et al., 2003). Itch
shorter dynamic range of thalamic NS cells was rarely evoked and never in isolation. Emotion
(Apkarian, A. V. and Shi, T., 1994), and with the binary descriptors (e.g., nauseating, cruel, suffocating) were
response to stimulation at pain sites (Figure 2(b)). uncommonly endorsed at either pain and pain/
We suggest that the first pathway is characterized sites (cf. Lenz, F. A. et al., 1995). Therefore, both
as a binary pain response signaling the presence/ painful responses to stimulation were described in
absence of painful stimuli, consistent with an alert- terms usually applied to external stimuli (exterocep-
ing/alarm function (Becker, D. E. et al., 1993; tion) rather than to internal or emotional phenomena
662 Psychophysics of Sensations Evoked by Stimulation of the Human Central Nervous System

(interoception). Exteroreceptive sensations can be (a) (b) S2

associated with a strong affective dimension. 40
Pain with a strong unpleasant or affective compo- Vop

Vc
45
nent can be evoked by stimulation of the lateral Vim 50
thalamus in the region of Vc. These sensations have PC

the character of memories of a previously experi-

Vcpc 55
enced pain, unlike the pain sensations evoked by
MG
thalamic sensation which are not related to previous
experience (see above) or a diffuse unpleasant sensa-
5 mm 1 mm
tion of the type evoked by stimulation of the medial
thalamus (see below). In the first case, stimulation in (c) S2
Vcpc (Figure 5) evoked chest pain with an affective PF RF PF RF PF RF
dimension in the case of a patient with coronary 40 46 52
artery disease which had been effectively treated by
balloon angioplasty (Lenz, F. A. et al., 1994a).
Microstimulation at site 49 (Figure 5) evoked an
unnatural, painful (visual analog scale 4.6/10),
mechanical sensation in the flank and an unnatural 41 47 53

nonpainful electrical sensation involving the left arm

PC
and leg. At sites 51 and 53 (Figure 5) microstimula-
tion evoked a sensation described by the patient as
heart pain, which was like what I took nitrogly- 40 25
42 48 54
cerin for except that it starts and stops suddenly. It
was not accompanied by dyspnea, diaphoresis, or NR NR
after effects. The PF involved the precordium and
left side of the chest from the sternum in the midline
40 10
to the anterior axillary line. Microstimulation at site 43 49 55
51 also evoked a sensation of nonpainful surface,
tingling in the left leg, which coincided with the NR NR
20
stimulation-associated angina.
Characteristics of the patients stimulation-asso- 5
44 50 56
ciated angina and usual angina were measured by
using a questionnaire. The same descriptors for stimu-
lation-associated angina were chosen intraoperatively NR NR
during stimulation at both sites 51 and 53 (Figure 5)
30 20
including: natural, deep, painful (visual analog scale 45 51 57
10/10), squeezing, frightful, fatiguing, and identical to
her angina. The questionnaire was administered three
times over several months postoperatively to describe
the patients usual angina. The following descriptors 10
30 40
were chosen: natural (3/3 administrations), deep (3/3),
painful (3/3), squeezing (3/3), frightful (2/3), suffocat- Figure 5 (a) Thalamic map of a patient with a history of
angina pectoris successfully treated with coronary artery
ing (2/3), and fatiguing (2/3). Her usual angina
balloon angioplasty. (b) Dark circular balloons on ticks to the
involved the left side of the chest, arm, and neck and right of the line indicate sites (sites 49, 51, 53) where thalamic
was associated with a surface (3/3), nonpainful (3/3), microstimulation evoked painful sensations in PFs indicated
and tingling (3/3) in the left arm and hand. This by stippling in the figurines (c). Open square balloons in (b)
coincidence of descriptors is unlikely to occur at ran- indicate sites where nonpainful, tingling sensations were
dom (p < 106, combinatorial analysis). evoked. All abbreviations and other conventions are as in the
legend to Figure 1. Reproduced from Lenz F. A., Gracely, R.
Similar emotional responses, including crying in H., Hope, E. J., Baker, F. H., Rowland, L. H., Dougherty, P.
response to thalamic stimulation in the same region, M., and Richardson, R. T. 1994a. The sensation of angina can
have been reported in the case of atypical chest pain, be evoked by stimulation of the human thalamus. Pain 59,
dysparunia, and the pain of childbirth (Davis, K. D. 119-125, with permission from Elsevier.
 Psychophysics of Sensations Evoked by Stimulation of the Human Central Nervous System 663

et al., 1995; Lenz, F. A. et al., 1995). Clinical criteria sometimes to exacerbate the patients ongoing pain
including a battery of cardiac tests (enzymes, EKGs, (Sano, K., 1979). This pain was evoked in projected
stress test) ruled out angina of cardiac origin in both fields as large as the whole body or hemibody.
these patients. Explorations in 50 patients without a Stimulation at sites, possibly the medial dorsal and
history of angina found that stimulation-associated periventricular nuclei evoked a generalized unplea-
angina was not evoked at any of the 19 stimulation sant sensation, not localized to a particular body part
sites with PFs on the chest wall. PFs were located on (Sano, K., 1979). Both responses may be consistent
the left chest wall at three sites and the right chest with the STT pathway or a multisynaptic pathway
wall at 16. At one of these 19 sites an unnatural, sharp, traversing the reticular formation (Willis, W. D.,
mechanical, painful, and vibration was described in 1985).
response to stimulation but emotional descriptors
were not endorsed.
Preoperative pain was clearly of cardiac origin in 44.5 Cortex
the patient with angina (Lenz, F. A. et al., 1994a), but
clearly not of cardiac origin in the patient with panic Functional imaging studies of the response to the appli-
disorder. The association of stimulation-associated cation of painful stimuli (Jones, A. K. et al., 1991; Talbot,
angina and the affective dimension was not unex- J. D. et al., 1991; Casey, K. L. et al., 1994; Craig, A. D. et al.,
pected (Lenz, F. A. et al., 1994a) as angina is often 1996; Andrew, D. et al., 2001) have identified three
associated with a strong affective dimension, unlike cortical areas with metabolic activation: primary soma-
other chest pains (Matthews, M. B., 1985; Braunwald, tosensory, parasylvian, and cingulate cortex. These
E., 1988; Procacci, P. and Zoppi, M., 1989; Pasternak, cortical areas all receive input arising from nociceptors
R. C. et al., 1992). Stimulation-evoked sharp chest as demonstrated by cortical potentials evoked by cuta-
pain occurred without an affective dimension in a neous application of a laser (laser-evoked potentials
retrospective analysis of patients without prior LEP) (Ohara, S. et al., 2004b), which selectively activates
experience of spontaneous chest pain with a strong nociceptors (Bromm, B. et al., 1984). Pain-related
affective dimension. Therefore, it is possible that in responses to stimulation have been identified in relation
the case, stimulation-associated chest pain included to the parasylvian cortex.
an affective dimension as a result of conditioning
by the prior experience of spontaneous chest pain
44.5.1 Parasylvian Cortex and Pain
with a strong affective dimension. In retrospect, the
Memory
affective dimension of stimulation-associated angina
might arise by similar conditioning. Parasylvian cortex receives input from the nuclei
around, and subnuclei within Vc (Van Buren, J. M.
and Borke, R. C., 1972) such as Vcpc, Vcpor, and
44.4.2 Medial and Intralaminar Thalamic
putative VMpo. Parasylvian receives nociceptive
Nuclei
input as evidenced by the presence of neurons
The medial and intralaminar nuclei also play a role responding to noxious stimuli (Robinson, C. J. and
in signaling pain sensations. Medial to Vc, the most Burton, H., 1980; Dong, W. K. et al., 1989; 1994) and of
dense STT terminal pattern is found in the intralami- LEP generators (Lenz, F. A. et al., 1998c; Vogel, H.
nar nucleus centralis lateralis (Mehler, W. R., 1962; et al., 2003; Ohara, S. et al., 2004b). The LEP generator
1969), while a much less dense termination is found in is anterior to primary auditory cortex, adjacent to the
other interlaminar nuclei central medial, parafascicu- S2 (secondary somatosensory cortex) generator for
laris (Mehler, W. R., 1962), and the medial dorsal vibratory SEPs (somatosensory-evoked potentials)
nucleus (Mehler, W. R., 1969). Nociceptive neurons (Hamalainen, H. et al., 1990; Ohara, S. et al., 2004b).
have been identified in an area that apparently corre- Source modeling suggests that the LEP generator is
sponds to the human central median nucleus (Ishijima, in the dorsal insulaparietal operculum (Figure 6)
B. et al., 1975; Tsubokawa, T. and Moriyasu, N., 1975; (Lenz, F. A. et al., 1998b; Vogel, H. et al., 2003),
Rinaldi, P. C. et al., 1991). different from the location of the local generator for
Milliampere-current-level stimulation at sites the P3 event-related potentials in the temporal base
probably located in the parafascicular, limitans, and (Lenz, F. A. et al., 1998b). Differences between the
central medial (parvocellular part) nuclei was locations of the late component of the LEP, the P2
reported to evoke a diffuse, burning pain, or wave, and the P3 suggest that the LEP P2 is not a
664 Psychophysics of Sensations Evoked by Stimulation of the Human Central Nervous System

(a) (b) P3-like wave, signaling the alertness evoked by pain-

ful stimuli (Zaslansky, R. et al., 1995; Lenz, F. A. et al.,
CS
1998b). Rather, it is likely that the LEP P2 is primar-
14 23
ily related to the sensation of pain (Ohara, S. et al.,
2004a).
R SF
The pain-related function of this area is consistent
with decreased pain discrimination and tolerance
10 mm
with lesions of the parietal operculum and insula,
(c)
respectively (Greenspan, J. D. et al., 1999). Studies
in which LEPs were recorded through depth electro-
Patient H des implanted in S2 and insula did not show a phase
reversal in the parietal operculum, that is, S2 (Frot,
R L R L M. et al., 1999). Stimulation through electrodes placed
in the posterior, superior insula produced pains
described as burning, stinging, electrical, and dis-
Patient P abling sensations (Ostrowsky, K. et al., 2002). In a
earlier series, stimulation of the exposed insula dur-
ing awake craniotomies (n 5) produced pain
R L R L
uncommonly but did produce nausea, tastes, somatic
sensations in the epigastric area, and rising sensations
LEP in the epigastric and umbilical areas (Penfield, W.
Patient C AEP and Jasper, H. 1954). Thus there is evidence from
human stimulation studies that insula, possibly the
5 cm
R L posteriorsuperior portion, is involved in pain-
related processes.
Figure 6 Verification of subdural grid electrode position
in head model for dipole source analysis. Positions of all Pain with a strong, vivid, affective dimension
subdural grid electrodes were determined in a evoked by stimulation of the region of Vc may
postoperative CT scan. These positions were entered be related activation of its parasylvian cortical pro-
into a spherical head model for dipole source analysis, jection zone (see Section 44.4.1) (Locke, S. et al.,
from which they were projected back into a preoperative
1961; Mehler, W. R., 1962; Van Buren, J. M. and
magnetic resonance imaging (MRI) of this patients brain
(patient H). (a) Intraoperative sketch shows the sylvian Borke, R. C., 1972). These vivid memories are similar
fissure with frontotemporal grid electrode 23 near the to those evoked by stimulation around the lateral
end of the fissure, and electrode 14 just posterior of the sulcus in patients with epilepsy (Halgren, E. et al.,
central sulcus. (b) Location of electrode 23 was 1978; Gloor, P. et al., 1982b; Gloor, P., 1990). These
projected from brain electromagnetic source analysis
memories may be related to cortical activation rather
software (BESA) head coordinates into the preoperative
MRI, showing excellent coincidence of the different than medial temporal structures as they are not fully
coordinate systems used in this study. A similar formed memories like those described here (Halgren,
correlation was found for electrode 14 and the central E. et al., 1978; Gloor, P. et al., 1982a). Furthermore,
sulcus. (c) Projection of dipole source locations for LEP fully formed memories can be evoked after removal
(*) and auditory evoked potentials (AEP) (.). The
of mesial temporal structures (Moriarity, J. L. et al.,
projection was performed in Talairach space of a
standard brain MRI. Left to right, panels show sagittal, 2001). Therefore, pain with a strong affective dimen-
coronal, and axial sections in patients H, P, and C, top sion in response to thalamic stimulation results might
to bottom. The LEP source was located above the be related to the activation of limbic and associated
sylvian fissure, and the AEP was located further posterior cortical structures (Lenz, F. A. et al., 1995). The role
and below the fissure. The axial slice was aligned to
of the medial temporal lobe might be transiently
pass through Heschls gyrus (AEP generator) to illustrate
the relative location of the source of the AEP (available involved in the formation of these memories, before
only in patients H and P). Reprinted from Vogel H., being located in cortex, independent of medial tem-
Port, J. D., Lenz, F. A., Solaiyappan, M., Krauss, G., poral lobe structures (Mishkin, M., 1979; Friedman,
and Treede, R. D. 2003. Dipole source analysis of D. P. et al., 1986; Zola-Morgan, S. and Squire, L. R.,
laser-evoked subdural potentials recorded from
1990).
parasylvian cortex in humans. J. Neurophysiol. 89, 3051
3060, used with permission the American Physiological Painful stimuli sometimes lead to long-term
Society. changes in pain processing, as well as to signaling
 Psychophysics of Sensations Evoked by Stimulation of the Human Central Nervous System 665

the presence of the stimulus. This appears to be the Acknowledgments

situation in the case of stimulation sites in and poster-
ior to Vc, where stimulation can evoke complex, fully This work is supported by the National Institutes of
formed, pain in patients with angina or atypical chest Health National Institute of Neurological Disorders
pain. These memories seem to be due to long term and Stroke (NS38493 and NS40059 to F. A. L.).
changes in forebrain function, that is, conditioning,
since memories of this type have not been reported in
response to STT stimulation.
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 45 Nociceptive Processing in the Cerebral Cortex
R D Treede, Ruprecht-Karls-University Heidelberg, Heidelberg, Germany
A V Apkarian, Northwestern University, Chicago, IL, USA
2009 Elsevier Inc. All rights reserved.

45.1 Introduction 670

45.2 Methods to Study Nociceptive Processing in the Human Cerebral Cortex 671
45.3 Cortical Regions that are Part of the Nociceptive System 673
45.3.1 The Primary Somatosensory Cortex 673
45.3.2 Parasylvian Cortex: the Operculo-Insular Region 674
45.3.2.1 The secondary somatosensory cortex (SII) 675
45.3.2.2 The frontal operculum 675
45.3.2.3 The insula 676
45.3.3 The Posterior Parietal Cortex 676
45.3.4 The Cingulate Cortex 677
45.3.5 The Prefrontal Cortex 678
45.4 Functional Roles of Cortical Nociceptive Signal Processing 678
45.4.1 Location and Quality of Phasic Pain 678
45.4.2 The Time Domain 680
45.4.3 Attention and Distraction Effects on Pain-Evoked Cortical Activity 680
45.4.4 Anticipation and Expectation 681
45.4.5 Empathy 681
45.5 Pain Modulation 681
45.5.1 Psychological Modulation of Pain 681
45.5.1.1 Hypnosis and pain-evoked cortical activity 681
45.5.1.2 Mood and emotional states and pain-evoked cortical activity 682
45.5.1.3 Placebo and pain-evoked cortical activity 682
45.5.2 Pharmacological Modulation of Pain 682
45.5.2.1 Opiates 682
45.5.2.2 Dopamine 683
45.5.2.3 Estrogen 683
45.6 Overview Regarding the Role of the Cortex in Acute Pain Perception 683
45.7 Clinical Applications 684
45.8 Chronic Pain 684
45.8.1 Studying Brain Activity in Chronic Pain with Nonspecific Painful Stimuli 684
45.8.2 Clinical Pain Conditions Studied by Stimulation and the Role of the Cortex 685
45.8.2.1 Migraine 686
45.8.2.2 Cluster headache 686
45.8.2.3 Cardiac pain 686
45.8.2.4 Irritable bowel syndrome 686
45.8.3 Spontaneous Pain as a Confound in Assessing Brain Activity 687
45.8.4 Functional Magnetic Resonance Imaging of Spontaneous Pain 689
45.8.5 Neuropathic Pain 689
45.8.6 Low Back Pain and Fibromyalgia 690
45.8.7 Overview Regarding the Role of the Cortex in Chronic Pain Perception 690
45.9 Conclusions and Outlook 691
References 691

669
670 Nociceptive Processing in the Cerebral Cortex

Glossary
ACC anterior cingulate cortex (n.b.: some authors OIC Operculo-insular cortex, consisting of insular
use this as a summary term for ACC and MCC) cortex plus the frontal, parietal and temporal
CBP Chronic back pain operculum.
CRPS Complex regional pain syndrome PAG Periaqueductal grey
EEG Electro-encephalography PET Positron emission tomography
fMRI Functional magnetic resonance imaging PFC prefrontal cortex
IBS Irritable bowel syndrome PHN Postherpetic neuralgia
IC Insular cortex SI Primary somatosensory cortex
MCC mid-cingulate cortex (n.b.: some authors call SII Secondary soamtosensory cortex
this region the posterior part of ACC) SCI Spinal cord injury
MEG Magneto-encephalography SPECT Single photon emission computed
MRS Magnetic resonance spectroscopy tomography
Th Thalamus

45.1 Introduction electrophysiological studies in humans, using magne-

toencephalography (MEG), electroencephalography
Conscious perception of external stimuli requires (EEG), subdural recordings directly from the surface
encoding by sensory organs, processing within the of the brain, and depth recordings during stereotactic
respective sensory system, and activation of the procedures (for a systematic review see Apkarian, A.
appropriate sensory cortical areas. Based on a small V. et al., 2005).
case series of infra- and supratentorial brain lesions, Meanwhile it has been recognized that painful sti-
Head H. and Holmes G. (1911) postulated that muli activate a vast network of cortical areas, including
the sensation of pain is an exception to this rule and the primary and secondary somatosensory cortex (SI,
that its conscious perception occurs in the essential SII), the insula, posterior parietal cortex, anterior and
organ of the thalamus. In spite of evidence to the mid-cingulate cortex, and parts of the prefrontal cortex
contrary from clinical reports (Marshall, J., 1951; (PFC; Figure 1). These areas are involved in the gen-
Biemond, A., 1956), evoked potentials in humans eration of painful percepts as well as in the descending
(Spreng, M. and Ichioka, M., 1964; Duclaux, R. et al., control of pain (for review see Kenshalo, D. R. and
1974; Carmon, A. et al., 1976; Chen, A. C. N. et al., Willis, W. D., 1991; Treede, R. D. et al., 1999; Price, D.
1979; Bromm, B. and Treede, R. D., 1984), single unit D., 2000; Apkarian, A. V. et al., 2005). Most of these
recordings in animals (Lamour, Y. et al., 1982; areas are also involved in other sensory, emotional,
Kenshalo, D. R. and Isensee, O., 1983), neuroanato- cognitive, motor or autonomic functions. Hence, the
mical tracing (Gingold, S. I. et al., 1991), and some nociceptive system converges with other systems for
early PET studies (Buchsbaum, M. S. et al., 1984), it the generation of the conscious percept of pain. In
was maintained for a long time that the cortical that sense, the nociceptive system is not different
representation of pain is a quantite negligable. from the visual system, for example. But it is still an
This situation changed, when the modern neuroi- open question, to what extent any cortical regions
maging techniques of positron emission tomography can be considered as nociceptive specific.
(PET) and later functional magnetic resonance In this chapter we will briefly review the methods
imaging (fMRI) demonstrated systematic metabolic used to assess nociceptive processing in the human
and perfusion changes in a large number of cortical brain, present connectivity and functional properties
areas following painful stimuli (Talbot, J. D. et al., of each of the principal cortical regions of the noci-
1991; Jones, A. K. P. et al., 1991a; Apkarian, A. V. et al., ceptive system, and summarize the roles of the
1992; Davis, K. D. et al., 1995). These findings cerebral cortex in various aspects of pain perception
were supported by invasive and noninvasive and pain modulation.
 Nociceptive Processing in the Cerebral Cortex 671

(a) M1 S1 (b)
SMA 1 2 3
PPC
2.
ACC PCC

Insula S2
PFC
BG Thalamus AMYG
Amyg HT
PAG
PB

3.
1.
PCC

HT PAG

Figure 1 Cortical regions involved in pain perception, their interconnectivity and ascending pathways. Locations of brain
regions involved in pain perception are color coded in a schematic drawing and in an example magnetic resonance image (MRI).
(a) Schematic diagram shows the regions, their interconnectivity, and afferent pathways. (b) The areas corresponding to those
shown in the schematic are shown in an anatomical MRI, on a coronal slice and three sagittal slices as indicated on the coronal
slice: primary and secondary somatosensory cortices (S1, S2, red and orange), anterior and mid-cingulate cortex (ACC, green),
insula (blue), thalamus (yellow), prefrontal cortex (PFC, purple), primary and supplementary motor cortex (M1 and SMA),
posterior parietal cortex (PPC), posterior cingulate cortex (PCC), basal ganglia (BG, pink), hypothalamus (HT), amygdala (AMYG),
parabrachial nuclei (PB), and periaqueductal gray (PAG). Reproduced from Apkarian, A. V., Bushnell, C., Treede, R. D., and
Zubieta, J. K. 2005. Human brain mechanisms of pain perception and regulation in health and disease. Eur. J. Pain 9, 463484.

45.2 Methods to Study Nociceptive indication for the procedure (e.g., epilepsy, tumors)
Processing in the Human Cerebral may have altered nociceptive signal processing.
Cortex EEG and MEG are noninvasive techniques for the
direct assessment of electrical activity in the brain.
Table 1 summarizes properties of the different brain Mathematical algorithms are available to estimate the
imaging techniques that have been used to define the location of the generators within the brain from the
nociceptive network in the human brain. The most signals recorded at the surface of the head with an
direct approach to learn about the functions of cortical accuracy of about 10 mm (Scherg, M., 1992; Pascual-
neurons is direct intracellular or extracellular record- Marqui, R. D. et al., 1994; Hari, R. and Forss, N., 1999).
ing of their electrical activity during sensory EEG and MEG techniques provide accurate timing
stimulation and in different contexts. This technique information. As a result, both methods have been used
is mostly restricted to animal studies (Kenshalo, D. R. mainly to identify the arrival of information to various
and Isensee, O., 1983; Dong, W. K. et al., 1994) and has cortical regions (stimulus-evoked potentials).
rarely been possible in humans (Hutchison, W. D. et al., Spontaneous fluctuations in EEG and MEG would
1999). Field potentials within the brain invert their provide a view of the interactions between cortical
polarity, when an electrode track passes through or areas. However, the application of the latter to painful
close to their generator source. This technique has states has remained minimal (Chen, A. C. N., 1993;
been used in the course of presurgical epilepsy diag- Ohara, S. et al., 2006). MEG detects brain magnetic
nostics (Frot, M. and Mauguiere, F., 1999), but since activity, a signal that is proportional and orthogonal
the electrode tracks are related to the clinical indica- to the local electrical activity. Depending on the orien-
tions, only few parts of the brain have been sampled tation of a local generator source and the gyral
that way. Presurgical epilepsy diagnostics using sub- geometry of the brain region, evoked potentials in
dural electrode grids samples a much larger part of the different brain areas may be better detected by MEG
brain surface, and dipole source analysis can be used to or EEG. The main weakness of EEG and MEG meth-
estimate the depths of the generators below the grids ods is their limited spatial resolution (on the order of
(Vogel, H. et al., 2003). All of these invasive recordings 1 cm for both methods).
in the human brain need to be interpreted with caution, PET, single photon emission-computed tomogra-
because the cortical pathology that provided the phy (SPECT), and fMRI measure brain activity
Table 1 Brain mapping techniques, their properties, and application in pain studies

Application in pain
Method Energy source Spatial resolution (mm) Temporal resolution (s) Constraints Output measured studies

EEG/MEG Intrinsic 10 0.001 Lack of unique Electrophysiology Increasing in use, mainly

electricity localization of brain events for detecting temporal
sequences
fMRI Radio waves, 45 410 Immobilization, loud, Relative cerebral Most used, mainly for
magnetic cooperation blood flow localizing brain activity
fields
MRS Radio waves, 10 10100 Immobilization, loud Relative chemical Recently used, for
magnetic concentrations detecting long term
fields changes in brain
chemistry
Nuclear (PET/ Radiation 510 601000 Radiation limits, Physiology, Decreasing in use,
SPECT) immobilization neurochemistry, becoming limited to
absolute values neurochemistry
Brain imaging techniques available but rarely or not yet used in pain studies
Single or multiunit Intrinsic 0.011 0.001 Invasive, direct Electrophysiology
electrophysiology electricity access to brain
Near infrared Infrared light 0.05 0.05 Immobilization, Relative cerebral
spectroscopy and surface > depth, blood flow
imaging limited field of view
Transcranial magnetic/ Magnetic/ 10 0.01 Risk of seizures, Electrophysiology,
electric electric immobilization, conduction times
stimulation fields loud
Structural MRI Radio waves, 1 N/A Immobilization, loud Structure, vasculature,
magnetic white matter
fields
Postmortem N/A 0.001 N/A Postmortem Microarchitecture,
chemoarchitecture

EEG, electroencephalography; MEG, magnetoencephalography; fMRI, functional magnetic resonance imaging; MRS, magnetic resonance spectroscopy; PET, positron emission tomography;
SPECT, single photon emission-computed tomography; N/A, not applicable.
 Nociceptive Processing in the Cerebral Cortex 673

indirectly by imaging changes in blood flow, blood developing rapidly and has the potential to become a
oxygenation, or local metabolic changes (Peyron, R. major method in the near future for studying brain
et al., 2000; Davis, K. D., 2003). All three methods can chemistry. In addition, voxel based morphometry
provide similar spatial resolution, although PET and allows to image structural changes related to disease
fMRI methodologies are now far more advanced than states (May, A. et al., 1999).
SPECT. The statistical models and experimental
designs available for PET and fMRI are robust and
very rich. Therefore, these two techniques are cur-
rently most extensively used for detecting brain 45.3 Cortical Regions that are Part of
circuitry underlying many cognitive states, including the Nociceptive System
pain. The temporal resolution of PET and SPECT is in
45.3.1 The Primary Somatosensory Cortex
the order of tens of seconds, while for fMRI it is shorter.
PET and SPECT provide the additional opportunity The primary somatosensory cortex (SI) is located in
for examining in vivo biochemistry and pharmacology the anterior part of the parietal lobe, where it con-
by imaging the distributions of specific neurotransmit- stitutes the postcentral gyrus. It consists of Brodmann
ters or receptors. Recent MRI methods, like magnetic areas 1, 2, 3a, and 3b (Figure 2(a)). Areas 3b and 1
resonance spectroscopy (MRS), have also provided the receive cutaneous tactile input, areas 3a and 2 pro-
ability to examine brain biochemistry. This approach is prioceptive input.

(a) Central sulcus (b)

1

3b

3a

(c) (d)
35
50
30
Peak frequency (impulses s1)

40
25
Impulses s1

20 30

15
20
10
10
5

0
0 20 40 60 80 100 43 45 47 50
Brush Pressure Pinch Stimulus intensity (C)

Figure 2 Nociceptive specific neuron in the primary somatosensory cortex (SI). (a) Left: SI consists of Brodmann areas 1, 2,
3a, and 3b in the postcentral gyrus. Black dot: location of the recorded neuron. (b) The small receptive field is consistent with a
role in spatial discrimination. (c) Stimulus response function to painful mechanical stimuli. (d) Stimulus response function to
painful heat stimuli. Modified from Kenshalo, D. R., Iwata, K., Sholas, M., and Thomas, D. A. 2000. Response properties and
organization of nociceptive neurons in area 1 of monkey primary somatosensory cortex. J. Neurophysiol. 84, 719729.
674 Nociceptive Processing in the Cerebral Cortex

Nociceptive input to monkey SI was demon- and hence are ideally suited to code for the location
strated anatomically. SI receives direct spino- of nociceptive stimuli (Kenshalo, D. R. and
thalamocortical input from the ventrobasal nuclei, Isensee, O., 1983). Somatotopy of nociceptive proces-
in particular the ventro-posterolateral (VPL) nucleus sing in the human SI has been confirmed by EEG and
(Gingold, S. I. et al., 1991). Nociceptive neurons in SI PET studies (Tarkka, I. M. and Treede, R. D., 1993;
are found in clusters, raising the possibility that SI Andersson, J. L. R. et al., 1997). Action potential dis-
may contain nociceptive specific columns (Lamour, Y. charges of nociceptive SI neurons in monkey are
et al., 1983). Since evidence for nociceptive neurons modulated by the intensity of both mechanical and
in the most superficial cortical layers is lacking, this heat stimuli (Figures 2(c) and 2(d)) and their dis-
hypothesis has not yet been confirmed. Nociceptive charges correlate with detection speed (Kenshalo,
neurons are rare in monkey SI and have mainly been D. R. et al., 1988). These findings suggest that noci-
found in area 1 (Kenshalo, D. R. et al., 2000), whereas ceptive SI neurons are involved in the coding of pain
optical imaging techniques have also suggested noci- intensity. This conclusion has been confirmed by a
ceptive input to area 3a (Tommerdahl, M. et al., PET study of hypnotic modulation of perceived pain
1996). Thus, nociceptive signal processing within SI intensity that also modulated perfusion of SI
may be spatially distinct from tactile signal proces- (Hofbauer, R. K. et al., 2001) and by correlation
sing that is primarily directed to area 3b. There is also analysis (Timmermann, L. et al., 2001).
some EEG and MEG evidence in humans that noci-
ceptive areas may be situated more medially within
SI than tactile areas with the same receptive fields, 45.3.2 Parasylvian Cortex: the Operculo-
suggesting that nociceptive and tactile signal proces- Insular Region
sing may occur in different subareas of SI (Ploner, M. The parasylvian cortex has a complicated macro-
et al. 2000; Schlereth, T. et al., 2003). Nociceptive scopic structure and only some of its
input to human SI has been confirmed by subdural cytoarchitectonic areas have been charted in detail
recordings (Kanda, M. et al., 2000; Ohara, S. et al., (Eickhoff, S. B. et al., 2006). In lateral views of the
2004). About 75% of the PET and fMRI studies brain, the Sylvian fissure runs above the temporal
reported activation of SI (Bushnell, M. C. et al., lobe and separates it from the parietal and frontal
1999; Apkarian, A. V. et al., 2005). lobes above the fissure. Hidden deep inside the
Nociceptive neurons in SI have small receptive Sylvian fissure lies a further lobe of the brain: the
fields (Figure 2(b)) that are somatotopically arranged, insula (Figure 3(a)). The insula is covered by the

(a) (b) (c)

Figure 3 Nociceptive regions in parasylvian cortex. (a) Sagittal section shows the insula as a triangular region deep inside
the Sylvian fissure. (b) Transaxial section illustrates the secondary somatosensory cortex (SII) as identified by tactile stimuli
(blue) and regions responsive to nociceptive stimulation (orange) that extend further rostrally and medially. (c) Coronal section
shows that the temporal operculum covers the insula below the Sylvian fissure, and frontal and parietal opercula cover the
insula above the fissure. Frontal and parietal opercula consist of an outer part on the convexity of the brain, a horizontal part
above the Sylvian Fissure, and an inner vertical part facing the insula across its circular sulcus. Arrowheads: Sylvian fissure
(lateral sulcus). Arrows: circular sulcus of the insula. Modified from Treede, R. D., Baumgartner, U., and Lenz, F. A. 2007.
Nociceptive Processing in the Secondary Somatosensory Cortex. In: Encyclopedia of Pain (eds. R. F. Schmidt and W. D.
Willis), pp. 13761379. Springer.
 Nociceptive Processing in the Cerebral Cortex 675

temporal, parietal and frontal opercula. Coronal sec- electrophysiological studies in humans have shown a
tions reveal that the parasylvian cortex consists of the bilateral response to unilateral stimulation, with a
insula itself, the inner vertical surface of the opercula, contralateral preponderance. Functionally, SII is con-
the horizontal banks of the Sylvian fissure, and most sidered to play a role in tactile object recognition and
laterally the outer surface of the convexity of the memory (Seitz, R. J. et al., 1991).
brain (Figure 3(c)). Evoked potential recordings in humans following
The majority of human imaging studies showed brief laser heat stimuli showed that SII was activated
consistent activation of the parasylvian cortex during simultaneously with or even earlier than SI (Ploner,
painful stimulation, and this activation overlapped M. et al., 1999; Schlereth, T. et al., 2003). Combined
only partly with that by tactile stimuli (Treede, R. anterograde and retrograde tracer studies in monkey
D. et al., 2007). Lesions in parasylvian cortex cause (Apkarian, A. V. and Shi, T., 1994) support the con-
deficits in pain perception (Greenspan, J. D. et al., cept that nociceptive input reaches SII more directly
1999), and intracortical electrical stimulation of this than tactile input. Hence, SII has been supposed to
region is painful (Ostrowsky, K. et al., 2002). Thus, be important for the recognition of painful stimuli as
this region is a good candidate to contain some noci- such.
ceptive specific cortical areas, if they exist. In contrast to the abundance of evidence for noci-
About 75% of the PET and fMRI studies reported ceptive activation of the SII region from human
activation of the SII region, and 94% found activa- studies, there are few single neuron recordings in
tion of the insula (Treede, R. D. et al., 2000; Apkarian, this area showing specific nociceptive responses
A. V. et al., 2005). But due to the curvature and (Treede, R. D. et al., 2000). In monkey, some conver-
oblique course of the Sylvian fissure (Ozcan, M. gence with visual input encoding the approach of a
et al., 2005), activated areas are often misallocated, sharp object to the face has raised the possibility of a
even across major sulci. Whereas the operculo-insu- representation of threat. These neurons, however,
lar cortex in the parasylvian region has been were not in SII proper but in area 7b which is adja-
recognized as one of the most important nociceptive cent to SII in monkey but not in humans (Dong, W.
cortical areas, its precise anatomical and functional K. et al., 1994). These neurons are now considered to
organization has yet to be determined. In particular it be part of the posterior parietal cortex (see below).
is not yet known, whether insula and operculum Since neurons in all studies on SII were searched
subserve distinct functions or form one uniform area. using mechanical skin stimulation, it is possible that
these studies missed nociceptive specific neurons,
45.3.2.1 The secondary somatosensory because many primary nociceptive afferents are
cortex (SII) mechanically insensitive (Treede, R. D. et al., 1998).
The secondary somatosensory cortex is located in Thus, an intriguing possibility is that tactile and
the superior bank of the Sylvian fissure, where it nociceptive inputs are represented in different areas
makes up a major part of the parietal operculum within the SII region.
(Figures 1(b), 3(b), and 3(c)). Nociceptive input to
monkey SII was demonstrated anatomically. SII 45.3.2.2 The frontal operculum
receives direct spino-thalamo-cortical input from Dipole source analysis of laser-evoked potentials
the ventrobasal nuclei, in particular the ventro-pos- (LEPs) in healthy volunteers, and subdural and
tero-inferior nucleus VPI (Stevens, R. T. et al., 1993). depth recordings in patients undergoing epilepsy
Nociceptive input to human SII has been confirmed surgery have identified an area in the inner vertical
by subdural recordings (Lenz, F. A. et al., 1998a). surface of the frontal operculum (Figure 4(a)) that
Single neuron recordings in SII have largely was activated by painful heat stimuli with a shorter
focused on the tactile representation (Robinson, C. J. latency (about 150 ms) than any other cortical area
and Burton, H. 1980; Fitzgerald, P. J. et al., 2006). The (Tarkka, I. M. and Treede, R. D., 1993; Valeriani, M.
SII region contains multiple somatotopic representa- et al., 1996; Ploner, M. et al., 1999; Frot, M. and
tions of the body, suggesting the existence of several Mauguiere, F., 2003; Schlereth, T. et al., 2003;
subregions (Disbrow, E. et al., 2000, Fitzgerald, P. J. Vogel, H. et al., 2003). This area anterior of the tactile
et al., 2004). Although most neurons in SII have con- SII area has a different somatotopic orientation (face:
tralateral receptive fields, this is the first part of the anterior, foot: posterior) than SII itself (face: lateral,
somatosensory system with a sizable proportion of foot: medial; Vogel, H. et al., 2003). The thalamic
bilateral receptive fields. Hence, most imaging and source of nociceptive input to this region is not yet
676 Nociceptive Processing in the Cerebral Cortex

(a) (b) 70 Left hemisphere (c) 70 Right hand

Right hemisphere Left hand
60 60

Source activity (nAm)

50 50

Pain rating (%)

*
40 * 40

30 30
SF SF
20 20

10 10

0 0
Dis Easy Diff Dis Easy Diff
Figure 4 Nociceptive input to the frontal operculum. (a) Projection of dipole source locations for the first component of
laser-evoked potentials (LEPs) onto a coronal magnetic resonance image slice at Talairach y 6 mm. The distribution
around the roof of the circular insular sulcus, ranging from the inner vertical face of the frontal operculum to the adjacent
dorsal insula matches the projection area of the nociceptive thalamic nucleus VMpo. (b) Task effects and interhemispheric
differences. (c) The hemispheric asymmetry of opercular activation (left hemisphere > right hemisphere) was not reflected or
caused by different visual analog score pain ratings between both hands. Dis, distraction task; Easy, easy spatial and
intensity discrimination tasks; diff, difficult discrimination tasks. ANOVA: P < 0.05. Modified from Schlereth, T., Baumgartner,
U., Magerl, W., Stoeter, P., and Treede, R. D. 2003. Left-hemisphere dominance in early nociceptive processing in the human
parasylvian cortex. Neuroimage 20, 441454.

clear: it may be VPI like for the posteriorly adjacent ventrobasal nucleus (Lenz, F. A. et al., 1993), a region
SII, or it may be VMpo like for the medially adjacent designated as VMpo by some authors (Craig, A. D.
dorsal insula. Somatotopy in the frontal operculum et al., 1994). VMpo projects to the dorsal insula and
would be consistent with that of a VMpo projection the adjacent frontal operculum. Nociceptive input to
target (Craig, A. D., 1995). Nociceptive input to the the insula in humans has been confirmed by depth
frontal operculum in humans has been confirmed by recordings (Frot, M. et al., 2003). The somatotopic
subdural and depth recordings (Lenz, F. A. et al., representation of pain in the dorsal insula in monkey
1998a; Frot, M. et al., 1999). Responses in this area (face: anterior, foot: posterior) is orthogonal to that in
are modulated during spatial and intensity discrimi- SII (face: lateral, foot: medial; Baumgartner, U. et al.,
nation tasks and show a left-hemisphere dominance 2006b). Direct electrical stimulation of the insula
(Figures 4(b) and 4(c)). is painful with a strong affective component
(Ostrowsky, K. et al., 2002).

45.3.2.3 The insula

The insula is located deep inside the Sylvian fissure,
45.3.3 The Posterior Parietal Cortex
where it can be visualized as a triangular shape in
sagittal sections (Figure 3(a)). It often contains two The posterior parietal cortex is located adjacent and
long sulci in its posterior part and three short sulci posterior to SI. It comprises Brodmann areas 5 and 7
rostrally. Several functional subdivisions of the insula (Figure 5(c)). Nociceptive input to this region is
have been suggested (Dieterich, M. et al., 2003; suggested by studies in monkey that reported short-
Schweinhardt, P. et al., 2006), but there is no consen- latency responses to nociceptive stimuli in area 7b;
sus yet. Parts of the insula subserve varied functions the same neurons also responded to visual stimuli of
in the somatosensory, vestibular, gustatory and auto- sharp objects directed at their receptive field
nomic nervous system, which led to the suggestion (Dong, W. K. et al., 1994). In the tactile system, this
that this region serves for a central representation of region is part of a dorsally directed stream involved
the internal state of the body (Craig, A. D., 2002). in stimulus location, convergence with visual infor-
This concept is consistent with the interoceptive mation and the generation of spatial information for
aspects of nociception. motor control. Nociceptive input to this region in
Another source of nociceptive input into the para- humans has not yet been explored with subdural
sylvian cortex is the posterior inferior part of the recordings, but there is some evidence from EEG
 Nociceptive Processing in the Cerebral Cortex 677

(a) Visual stimulation e (b)

trod
Elec

Impulses / 200 ms bin

Withdraw
A B A B C D E
Syringe
target Hold
15 Approach
10
t.
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D
C

(c)
cs
(d) Thermal stimulus - response function
1
4 20 38 c
3b 45 51 c
2
3a
IPS
15

Spikes s1
5
10
7b
6
5
LS
7b
S2
T3 Ri 0
Pa

5
44 45 46 47 48 49 50 51 52
Temperature (c)
Figure 5 Nociceptive neuron in posterior parietal cortex (Area 7b threat neuron). (a) Bilateral receptive field in the
orofacial region; seeing a syringe approach the receptive field was also an adequate stimulus. (b) Responses to the
stimuli shown in (a). (c) Location of the recorded neuron on a coronal section that passes through both the central
sulcus (CS) and the Sylvian fissure (LS: lateral sulcus). (d) Stimulus response function to painful heat stimuli. IPS,
Intraparietal sulcus. Modified from Dong, W. K., Chudler, E. H., Sugiyama, K., Roberts, V. J., and Hayashi, T. 1994.
Somatosensory, multisensory, and task-related neurons in cortical area 7b (PF) of unanesthetized monkeys. J.
Neurophysiol. 72, 542564.

and MEG studies in humans that nociceptive stimuli input to human cingulate cortex has been confirmed
activate parietal lobe posterior of SI (Schlereth, T. by subdural recordings and by intracortical record-
et al., 2003; Forss, N. et al., 2005). Few fMRI studies ings (Lenz, F. A. et al., 1998b; Hutchison, W. D. et al.,
have assessed this region (Kulkarni, B. et al., 2005; 1999). About 87% of the PET and fMRI studies
Schmahl, C. et al., 2006). reported activation of the cingulate cortex
(Apkarian, A. V. et al., 2005), but no region of the
cingulate cortex is considered to be nociceptive spe-
cific (Vogt, B. A., 2005).
45.3.4 The Cingulate Cortex
Nociceptive neurons in ACC have large or even
The cingulate cortex is located above the corpus whole-body receptive fields (Figure 7, Sikes, R. W.
callosum and around its anterior knee (Figure 6). and Vogt, B. A., 1992; Yamamura, H. et al., 1996). For
The anterior cingulate cortex (ACC) comprises this reason it is unlikely that they contribute to the
Brodmann areas 24 and 32, whereas the posterior sensory dimension of pain. Monkey ACC neurons
cingulate cortex (PCC) contains areas 23 and 31 activate during pain avoidance behavior, reflecting
(Vogt, B. A. et al., 1995). ACC receives nociceptive anticipation, and response selection (Koyama, T.
thalamocortical input from the mediodorsal (MD) et al., 1998; 2001). The cingulate cortex is supposed
and parafascicular (Pf) nuclei (Vogt, B. A. et al., to participate in the affective-motivational dimension
1979). ACC has been further subdivided into mid- of pain. This conclusion has been confirmed by a
cingulate cortex, which is associated with response PET study of hypnotic modulation of perceived
selection and motor efferent functions, and ACC pain affect that also modulated perfusion of ACC
proper that is related to emotion and autonomic (Hofbauer, R. K. et al., 2001) and by correlation ana-
efferent functions (Vogt, B. A., 2005). Nociceptive lysis (Tolle, T. R. et al., 1999).
678 Nociceptive Processing in the Cerebral Cortex

dimension includes intensity discrimination, pain

qualities, stimulus localization and timing discrimi-
nation; this dimension is traditionally thought to
involve lateral thalamic nuclei and the somatosen-
sory cortices SI and SII. The affective-motivational
dimension includes perception of the negative
hedonic quality of pain, autonomic nervous
system manifestations of emotions, and motivated
behavioral responses; this dimension is traditionally
thought to involve medial thalamic nuclei and the
limbic cortices ACC and MCC. The insula has an
intermediate position in that concept, receiving input
Figure 6 Distribution of cingulate cortex regions and from lateral thalamus but projecting into the limbic
subregions. Region borders are marked with arrows.
Cross-hair shows vertical plane at the anterior
system. The cognitive-evaluative dimension includes
commissure (VCA) and the anteriorposterior interaction with previous experience, cognitive
commissural line. A functional overview, derived from the influence on perceived pain intensity and an
analysis of a large volume of literature illustrates general overall evaluation of its salience; this dimension is
regional function. aMCC, anterior mid-cingulate cortex; traditionally thought to involve the PRC. Numerous
cas, callosal sulcus; cgs, cingulate sulcus; dPCC, dorsal
posterior cingulate cortex; irs, inferior rostral sulcus; mr,
neuroimaging studies have assessed various
marginal ramus of cgs; pACC, pregenual anterior experimental paradigms derived from several
cingulate cortex; pcgs, paracingulate sulcus; pMCC, psychological concepts that do not easily fit into the
posterior mid-cingulate cortex; RSC, retrosplenial cortex; traditional three dimensions of pain. Therefore, we
sACC, subgenual anterior cingulate cortex; spls, splenial here report imaging evidence for involvement of
sulci; vPCC, ventral posterior cingulate cortex.
Reproduced from Vogt, B. A. 2005. Pain and emotion
cortical areas in specific functions instead of the
interactions in subregions of the cingulate gyrus. Nat. dimensions of pain.
Rev. Neurosci. 6, 533544.

45.4.1 Location and Quality of Phasic

45.3.5 The Prefrontal Cortex Pain
The PFC (including Brodmann areas 9, 10, 46) Neuroimaging studies have examined brain regions
comprises the major part of the frontal lobe and is activated by many types of painful stimulation, includ-
located anterior of the motor cortical areas. There is ing noxious heat and cold, muscle stimulation using
no evidence that it would receive a direct nociceptive electric shock or hypertonic saline, topical and intra-
thalamo-cortical input, but the PFC receives dermal capsaicin, colonic distention, rectal distension,
cortico-cortical input from the cingulate gyrus gastric distension, esophageal distension, ischemia,
that may convey nociceptive information. About cutaneous electric shock, ascorbic acid, laser heat, as
55% of the PET and fMRI studies reported activa- well as an illusion of pain evoked by combinations of
tion of the PRC in healthy subjects, and 81% of innocuous temperatures (Apkarian, A. V. et al., 2005;
the studies in chronic pain patients (Apkarian, A. V. Bushnell, M. C. and Apkarian, A. V., 2005). Despite the
et al., 2005). The PRC is assumed to participate in differences in sensation, emotion and behavioral
the cognitive-evaluative dimension of pain and in responses provoked by these different types of pain,
endogenous pain control (Lorenz, J. et al., 2003; individuals can easily identify each as being painful.
Schmahl, C. et al., 2006). Thus, there appears to be a common construct of pain
with an underlying network of brain activity in the
areas described above. Nevertheless, despite the simi-
45.4 Functional Roles of Cortical larities in pain experiences and similarities in neural
Nociceptive Signal Processing activation patterns, each pain experience is unique.
Subjects can usually differentiate noxious heat from
Pain perception has been conceived to consist of noxious cold from noxious pressure. This ability to
sensory-discriminative, affective-motivational and differentiate pains is particularly puzzling, since there
cognitive-evaluative dimensions (Melzack, R. and is ubiquitous convergence of information from cuta-
Casey, K. L. 1968). The sensory-discriminative neous, visceral and muscle tissue throughout the
 (b)
(a)
I
(c) IIIII
V
squeezing VI

1
squeezing
squeezing 8 2 D

7 3 1 mm

squeezing squeezing L

6 4
squeezing
squeezing 5

25 mV
100 m
1 sec
squeezing

#6T1.2443
type: N6

Figure 7 Nociceptive neuron in the anterior cingulate gyrus. (a) Responses to painful mechanical stimuli show a whole-body receptive field for this neuron. (b) Intracellular dye
injection reveals a lamina V pyramidal neuron. (c) Location of the recorded neuron in the rat cingulate cortex. Reproduced from Yamamura, H., Iwata, K., Tsuboi, Y., Toda, K.,
Kitajima, K., Shimizu, N., Nomura, H., Hibiya, J., Fujita, S. and Sumino, R. 1996. Morphological and electrophysiological properties of ACCx nociceptive neurons in rats. Brain
Res. 735, 8392.
680 Nociceptive Processing in the Cerebral Cortex

afferent nociceptive system (Willis, W. D. and MEG studies. The dual pain sensation elicited by a
Coggeshall, R. E., 2004). The convergence and the single brief painful stimulus that is due to the differ-
similarities in brain regions activated by different ent conduction times in nociceptive A- and C-fibers
types of pain are consistent with phenomena such as (about 1 s difference) is reflected in two sequential
referred pain, but cannot explain either the ability to brain activations in EEG and MEG recordings from
identify the origin of pain or with contrasting beha- SI, SII, and MCC (Bromm, B. et al., 1983; Bragard, D.
vioral reactions to cutaneous and visceral pain et al., 1996; Magerl, W. et al., 1999; Opsommer, E. et al.,
(withdrawal versus quiescence). 2001; Iannetti, G. D. et al., 2003). EEG mapping
There is evidence from single neuron recordings, studies (Kunde, V. and Treede, R. D., 1993;
MEG, PET, and fMRI that neural activity in SI cortex Miyazaki, M. et al., 1994), source analysis (Tarkka, I.
could underlie the identification of the locus of cuta- M. and Treede, R. D., 1993; Valeriani, M. et al., 1996;
neous pain. Kenshalo and colleagues (Kenshalo, D. R. Ploner, M. et al., 1999), and intracranial recordings
et al., 1988; Kenshalo, D. R. and Isensee, O., 1983) (Lenz, F. A. et al., 1998a; Frot, M. et al., 1999) show
showed that SI nociceptive neurons have discrete that the earliest pain-induced brain activity origi-
receptive fields, so that different neurons respond to nates in the vicinity of SII. In contrast, tactile
painful stimulation in different skin areas. stimuli activate this region only after processing in
Correspondingly, EEG, PET, and fMRI studies have the primary somatosensory cortex (Ploner, M. et al.,
shown a topographic organization of nociceptive 2000). The adjacent dorsal insula is activated slightly
responses in SI cortex similar to the organization of but significantly later than the operculum (Frot, M.
tactile responses, i.e., a medio-lateral organization of and Mauguiere, F., 2003). These observations sup-
foot, hand, face, and intra-abdominal areas (Tarkka, I. port the suggestion derived from anatomical studies
M. and Treede, R. D., 1993; Andersson, J. L. R. et al., that the SII region and adjacent insula are primary
1997; DaSilva, A. F. M. et al., 2002; Strigo, I. A. et al., receiving areas for nociceptive input to the brain
2003; Vogel, H. et al., 2003). Most imaging studies find (Apkarian, A. V. and Shi, T., 1994; Craig, A. D., 2002).
little somatotopic organization of pain in other cortical
areas (Tarkka, I. M. and Treede, R. D., 1993; Xu, X. P.
45.4.3 Attention and Distraction Effects on
et al., 1997), thus suggesting that responses in SI cortex
Pain-Evoked Cortical Activity
may be most important for pain localization. More
recently, a somatotopic organization has also been Early human brain imaging studies examining the
documented for operculo-insular cortex (Vogel, H. effects of attention and distraction show modulation
et al., 2003; Baumgartner, U. et al., 2006a). A left hemi- of pain-evoked activity in a number of cortical
sphere dominance has been reported for the sensory regions, including sensory and limbic structures, as
dimension of pain (Schlereth, T. et al., 2003), whereas well as prefrontal areas (Bushnell, M. C. et al., 1999;
right hemisphere dominance was observed for the Longe, S. E. et al., 2001; Bantick, S. J. et al., 2002;
affective dimension (Pauli, P. et al., 1999; Brooks, J. C. Schlereth, T. et al., 2003). These results generally
W. et al., 2002). show reduced activations in sensory regions of the
Strigo I. A. and colleagues (2003) directly compared cortex and some increased activity in more frontal
brain activations produced by esophageal distension regions, suggesting that attentional modulation is
and cutaneous heat on the chest that were matched mediated through the latter structures resulting in
for pain intensity. They found that the two qualita- reduced sensory processing, where the attentional
tively different pains produced different primary loci distraction is usually reported resulting in reduced
of activation with insula, SI, motor and prefrontal perceived magnitude of pain. A more recent study
cortices. Such local differences in responses within extends these notions by showing that during distrac-
the nociceptive network might subserve our ability to tion there is a functional interaction between
distinguish visceral and cutaneous pain as well as the pregenual ACC and frontal cortex exerting a top-
differential emotional, autonomic, and motor responses down modulation on periaqueductal gray (PAG) and
associated with these different sensations. thalamus to in turn reduce activity in cortical sensory
regions and correspondingly decrease perception of
pain (Petrovic, P. et al., 2000; Tracey, I. et al., 2002;
45.4.2 The Time Domain
Valet, M. et al., 2004). Given that ACC is implicated
Most information about the temporal sequence of in attentional modulation as well as pain perception,
pain-evoked brain activation comes from EEG or a distraction study indicates that some portions of the
 Nociceptive Processing in the Cerebral Cortex 681

pregenual ACC region are decreased with distraction activity reflects the pain experienced by others and
while others are increased, consistent with these two that multiple cortical areas involved in sensory pro-
different functions (Frankenstein, U. N. et al., 2001). cessing of pain are also activated.
The overall notion that empathy involves assess-
ment of the pain experienced by others pain
45.4.4 Anticipation and Expectation
mirroring was tested directly in subjects with alex-
Anticipation or expectation of pain can activate many ithymia, a cognitive and emotional deficit leading to
of the cortical areas related to perception of pain in difficulty in identifying ones own emotional state
the absence of a physical pain stimulus (Ploghaus, A. and also other peoples emotional state. The study
et al., 1999; Hsieh, J. C. et al., 1999b; Sawamoto, N. showed in fact reduced activity in PRC and MCC
et al., 2000; Porro, C. A. et al., 2002). Two studies have during a pain empathy condition in this patient
attempted to identify the circuitry for modulation of population (Moriguchi, Y. et al., 2006). Even though
pain by expectation. In one study MCC, caudate these results are internally consistent, their interpre-
nucleus, cerebellum, and nucleus cuneiformis were tation remains problematic. Simple introspection
modulated by systematic manipulation of pain inten- casts doubt on the notion that empathy means actu-
sity expectation by two different cues (Keltner, J. R. ally experiencing another persons pain. Instead, what
et al., 2006), whereas pain intensity itself modulated is called empathy may be the assessment of the mag-
somatosensory cortex, insula, and rostral ACC. In the nitude of negative emotion that the other person may
second study expectancy was modulated by a placebo be experiencing, i.e., a cognitive function of interper-
procedure, resulting mainly in modulation including sonal communication. According to that concept,
MCC, PRC, cerebellum, pons, and parahippocampal empathy may be defined as a complex form of psy-
gyrus (Kong, J. et al., 2006). The latter study is com- chological inference that enables us to understand the
plicated by the fact that the procedure is a personal experience of another person through cog-
combination of manipulation of expectancy and pla- nitive/evaluative and affective processes. A study in
cebo acupuncture treatment. Generally, there patients with congenital insensitivity to pain
remains a strong need for systematic studies to iden- (Danziger, N. et al., 2006) reported a deficit in rating
tify brain elements that modulate pain responses due pain-inducing events, but normal inference of pain
to expectation. from facial expressions (empathy), indicating that
empathy for pain does not require an intact pain
percept.
45.4.5 Empathy
A provocative study opened the field regarding the
interaction between pain and empathy, where the 45.5 Pain Modulation
authors defined empathy as the ability to have an
45.5.1 Psychological Modulation of Pain
experience of anothers pain. Using this definition
and comparing brain activity for experiencing pain The psychological modulation of pain has been
or knowing that their loved one, present in the same observed very early on and studied in the clinical
room, was experiencing the same pain, the authors and laboratory settings (Beydoun, A. et al., 1993;
showed many cortical regions similarly activated for Villemure, C. and Bushnell, M. C., 2002). Modern
both conditions, especially bilateral operculo-insular brain imaging techniques now provide powerful tools
cortex and MCC (Singer, T. et al., 2004). These with which mechanisms of these modulations can be
results were interpreted as evidence for the affective documented and dissected. Given that these are cog-
component of pain being active in both empathy and nitive/attentional modulations their effects should be
pain, and thus concluded that empathy for pain observed at the cortical level.
involves the affective component, but not the sensory
component, of pain. The study induced a flurry of 45.5.1.1 Hypnosis and pain-evoked
activity in attempting to understand the relationship cortical activity
between empathy and pain. Multiple groups have Hypnosis can alter pain perception. It has been used
replicated the main finding and proposed different to differentially modulate sensory and affective
underlying mechanisms (Morrison, I. et al., 2004; dimensions of pain and thus distinguish the cortical
Botvinick, M. et al., 2005; Jackson, P. L. et al., 2005), regions involved in these dimensions. Such studies
with multiple studies showing that at least MCC indicate that SI activity is preferentially modulated
682 Nociceptive Processing in the Cerebral Cortex

when the hypnotic instructions are directed to the is observed in the above studies as well, which links
intensity of pain, while MCC activity is preferen- opiate descending modulation with prefrontal corti-
tially modulated when hypnosis is directed to the cal control of placebo analgesia. The correspondence
unpleasantness of pain (Rainville, P. et al., 1997, between placebo analgesia and reward was directly
Hofbauer, R. K. et al., 2001). Brain activity for hypno- studied and the results show a strong correspondence
tically induced pain perception seem to be different between brain regions involved in each (Petrovic, P.
from activity for imagined pain in sensory, limbic, et al., 2005).
and prefrontal activation patterns (Derbyshire, S. W.
et al., 2004). The sensory and limbic cortical activa-
45.5.2 Pharmacological Modulation
tions for hypnotically induced and stimulation-
of Pain
induced pain seem relatively similar, the only region
that may be differentiating them seems to be the A league table of analgesic efficacy has been gener-
medial PRC (Raij, T. T. et al., 2005). ated based on pain-related evoked potentials
(Scharein, E. and Bromm, B., 1998). Since these stu-
45.5.1.2 Mood and emotional states and dies used electrical stimuli that circumvent
pain-evoked cortical activity peripheral nociceptive transduction mechanisms,
Studies show that experimental procedures that this table reflects central rather than peripheral
improve mood generally reduce pain, while those analgesic actions, as evidenced, e.g., by the higher
that have a negative effect on mood increase pain efficacy of the antidepressant imipramine than the
(Zelman, D. C. et al., 1991; Marchand, S. and nonsteroidal anti-inflammatory drug (NSAID) acet-
Arsenault, P., 2002). One study showed that looking ylsalicylic acid. Since dipole source analysis has not
at fearful faces increased their level of anxiety and been applied in these EEG studies, possible cortical
discomfort, which also resulted in enhanced esopha- sites of actions were not differentiated. Combining
geal stimulation-evoked activity in limbic regions fMRI and pharmacology promises to provide that
like ACC and insula (Phillips, M. L. et al., 2003). type of information (Tracey, I., 2001; Borsook, D.
et al., 2006). In addition, PET techniques can be
45.5.1.3 Placebo and pain-evoked cortical used for direct tracing of cortical distribution of a
activity given drug, when it has been labeled with the posi-
Placebo is a potent modulator of pain; it afflicts all tron emitting 11C isotope.
clinical studies of pain pharmacology. Placebo effects
have also been seen in depression and in Parkinsons 45.5.2.1 Opiates
disease and recent brain imaging studies show a There is a vast literature regarding opiate-mediated
robust brain and subcortical reward circuitrys invol- descending modulation through the PAG and a simi-
vement in these (Lidstone, S. C. and Stoessl, A. J., larly large literature on its effects on inhibitory
2007). The first neurochemical evidence for opiate interneurons in the spinal cord. At the cortical level,
involvement of placebo was demonstrated about 30 it has been noted that opiate receptors are present in
years ago by showing that placebo analgesia can be many parts of the nociceptive system, with high spe-
blocked by naloxone (Levine, J. D. et al., 1978). cific binding in ACC, insula, and frontal operculum,
Consistent with this notion, changes in endogenous and with moderate specific binding in MCC, SII, and
opiate release are shown to be involved in placebo- SI (Jones, A. K. P. et al., 1991b; Baumgartner, U. et al.,
induced analgesia, where PRC (medial and lateral) as 2006a).
well as insula and ventral striatum seem to be Recent studies of opiate-mediated responses in the
involved, where high placebo responders increased brain have used two approaches, examination of meta-
opiate release in ventral striatum was positively cor- bolic function in response to pharmacological agents
related with pain ratings (Zubieta, J. K. et al., 2005). and direct measurement of opiate receptor binding
Results generally consistent with this brain response potential. Exogenous administration of -opioid recep-
pattern have been demonstrated by a number of tor agonist drugs show dose-dependent increased
other groups (Wager, T. D. et al., 2004; Benedetti, F. metabolic activity in regions rich with -opioid recep-
et al., 2005; Kong, J. et al., 2006); the medial prefron- tors, which in the cortex are mainly localized to PRC
tal/rostral ACC responses for placebo seem to recruit and ACC (Firestone, L. L. et al., 1996; Schlaepfer, T. E.
PAG and amygdala (Bingel, U. et al., 2006); and et al., 1998; Wagner, K. J. et al., 2001). Also, -opioid
involvement of PAG in placebo-induced analgesia agonist fentanyl on brain responses to painful stimuli
 Nociceptive Processing in the Cerebral Cortex 683

have been explored, showing that most cortical follicular menstrual cycle phase compared to late
responses to pain are reduced or eliminated, confirming follicular/mid-cycle; with greater activity found
analgesic effects of the opiate (Casey, K. L. et al., 2000; during early follicular phase in amygdala, hypotha-
Petrovic, P. et al., 2002). Changes in endogenous opioid lamus, hippocampus, orbital frontal cortex, and
system is studied using a selective -opioid radiotracer, ACC, suggesting that estrogen may attenuate
showing activation of opiate neurotransmission in ros- arousal in women through cortical-subcortical
tral ACC, PRC, and insula during a tonic muscle pain control of hypothalamicpituitaryadrenal circuitry
(Zubieta, J. K. et al., 2001). (Goldstein, J. M. et al., 2005). There is also growing
evidence of gender differences in the anatomy of the
45.5.2.2 Dopamine brain, its connectivity, and in cognitive abilities
Dopamine is best known for its role in motor, moti- (Hampson, E., 2002; Becker, J. B. et al., 2005).
vation, and pleasure control. There is accumulating Multiple studies have documented that threshold
evidence to suggest that dopamine acting at the level and tolerance for pain is lower for women
of the basal ganglia may also be involved in pain (Wiesenfeld-Hallin, Z., 2005; Rolke, R. et al., 2006;
modulation. Human brain imaging studies document Wilson, J. F., 2006).
increased pain sensitivity to be associated with lower Gender differences in cortical activity for acute
levels of endogenous dopamine (Pertovaara, A. et al., pain has been observed in early studies (Paulson, P. E.
2004; Martikainen, I. K. et al., 2005; Scott, D. J. et al., et al., 1998). The association of sex hormones with
2006); and sustained experimental pain results in pain perception and pain memory was studied by
release of dopamine in the basal ganglia (Scott, D. J. Zubieta J. K. and colleagues (Zubieta, J. K. et al.,
et al., 2006), and indicate an interaction between 2002; Smith, Y. R. et al., 2006). They scanned healthy
opiate activity and dopamine where alfentanil women during their early follicular phase when
administration results in decreased mechanical pain estrogen levels are low and then repeated the scan
and decreased release of dopamine in the basal gang- during that same phase in another month after they
lia (Hagelberg, N. et al., 2002). Moreover, abnormal had worn for a week an estrogen-releasing skin patch
levels of dopamine in the basal ganglia have been which increased their estrogen to levels normally
associated with chronic pain in burning mouth syn- seen in the menstrual cycle. These studies showed
drome and atypical facial pain (Jaaskelainen, S. K. that more -opioid receptors were available in the
et al., 2001; Hagelberg, N. et al., 2003a; 2003b), and presence of high estrogen levels, and women
perhaps in fibromyalgia (Wood, P. B. et al., 2007). reported less pain in response to acute painful stimuli
Patients with restless legs syndrome display a pro- than when their estrogen levels were low. Moreover,
nounced mechanical hyperalgesia to pinprick stimuli estrogen-associated variations in the activity of
that is slowly reversed by dopaminergic agonists -opioid neurotransmission correlated with indivi-
(Stiasny-Kolster, K. et al., 2004), but this action is dual ratings of the sensory and affective perceptions
probably mediated by extrastriatal dopamine of pain and the subsequent recall of that experience.
receptors. These data demonstrate a significant role of estrogen
in modulating endogenous opioid neurotransmission
45.5.2.3 Estrogen and associated psychophysical responses to an acute
Gender is one of the most important determinants of pain stressor in humans. Approximately similar
human health. Women far outnumber men in sus- results have been reported by another group (De
ceptibility to many autoimmune disorders, Leeuw, R. et al., 2006).
fibromyalgia, and chronic pain, differences in phy-
siological responses to stress may potentially be an
important risk factor for these disorders as physiolo- 45.6 Overview Regarding the Role of
gic responses to stress seem to differ according to the Cortex in Acute Pain Perception
gender, with phase of menstrual cycle, menopausal
status and with pregnancy (Kajantie, E. and The above sections describe the contribution of
Phillips, D. I., 2006). Consistent with this idea recent modern imaging studies to our understanding of
fMRI study shows that brain activity in premenopau- the involvement of the cortex in pain perception.
sal women as studied for negative valence/high Cortical activity is demonstrated to possess proper-
arousal in contrast to neutral visual stimuli show ties necessary for involvement in pain perception,
differences when the task is performed during early like somatotopic representation of painful stimuli,
684 Nociceptive Processing in the Cerebral Cortex

correlation with stimulus intensity, modulation (Treede, R. D., 2001), there is currently no measure
with attention, modulation with expectation and of brain activity that would objectively show whether
other psychological variables, and distinct brain or not a person is in pain. Therefore, neither EEG/
regions showing differential activity for sensory MEG nor imaging with fMRI or PET can be used to
and affective dimensions of pain, as well as verify the presence of ongoing spontaneous pain.
attenuation of responses with analgesic drugs EEG and MEG recordings of evoked potentials,
(Apkarian, A. V. et al., 2005). Thus, human brain however, are sensitive enough to verify whether the
imaging studies have asserted the role of the cortex ascending nociceptive pathways are intact in a given
in acute pain. individual patient (Bromm, B. and Lorenz, J., 1998;
However, because imaging studies identify brain Treede, R. D. et al., 2003; Cruccu, G. et al., 2004). A
responses in a correlative manner, they may all reflect prerequisite for this use of EEG and MEG technol-
secondary processes. Perception of pain automatically ogy is a phasic adequate stimulus for nociceptor
directs attention to the source of pain, results in auto- activation. Radiant heat pulses of a few milliseconds
nomic responses, motor reflexes to escape from the duration, as generated by infrared lasers, have been
pain, and other emotional and cognitive responses that validated for this purpose (Plaghki, L. and Mouraux,
undoubtedly are at least partially mediated through A., 2003), and LEPs can thus be used to verify the
cortical processes. Therefore, the role of the cortex in presence of negative sensory signs of nociception
pain perception in contrast to its activity as a conse- (hypoalgesia).
quence of these secondary responses remains unclear Neither fMRI nor PET are sensitive enough to
and needs to be properly addressed in future studies allow clinical assessment of nociceptive pathways in
(Apkarian, A. V., 2004). individual cases, since so far no activation paradigm
In fact, unpublished data from Apkarians labora- has been developed that would reliably induce a
tory suggest that a large proportion of the brain particular cortical activation pattern in each and
network activated with acute pain may be responses every healthy subject. Thus, negative findings with
that are commonly involved in general magnitude these techniques are inconclusive.
estimation for any sensory modality, and as a result For the study of pathological nociceptive proces-
are not specific for nociception (abstract Society for sing at the group level, however, fMRI and PET
Neuroscience 2006), suggesting that the majority of techniques are extremely powerful. These techni-
cortical activity for acute pain are instead sensory, ques have broadened our understanding of the
cognitive, emotional, and attentional responses to pathophysiology of conditions with decreased pain
nociceptive inputs. Careful clinical and neuropsy- perception such as afferent pathway lesions or bor-
chological examination of patients with small brain derline personality disorder, and conditions with
lesions, combined with high-resolution structural and increased pain perception such as neuropathic pain
neuropharmacological neuroimaging in the same or fibromyalgia (Gracely, R. H. et al., 2004;
patients, will be needed to address the question Maihofner, C. et al., 2005; Garcia-Larrea, L. et al.,
what brain structures are necessary for acute pain 2006; Schmahl, C. et al., 2006; Schweinhardt, P. et al.,
perception. Anatomical tracing studies and single 2006). In addition, PET allows direct estimation of
unit recordings should address the question, to what pharmacological and biochemical processes in the
extent nociceptive specific neurons exist in these brain, such as alterations in dopamine or opioid
brain structures. For most parts of the nociceptive receptor availability (Hagelberg, N. et al., 2003a;
cortical network, as illustrated above, it is likely that Willoch, F. et al., 2004).
they participate only partly in pain perception, by
providing certain feature extraction functions, but
they also participate in other functions in different 45.8 Chronic Pain
contexts.
45.8.1 Studying Brain Activity in Chronic
Pain with Nonspecific Painful Stimuli
45.7 Clinical Applications Chronic pain might result from cortical processing of
chronic nociceptive spinothalamic input according to
It should be emphasized that although the subjective the same mechanisms as in acute pain, or there might
phenomenon of being in pain can be considered be specific changes in cortical processing of nocicep-
an emergent phenomenon of cortical activity tive input in patients with chronic pain. Such changes
 Nociceptive Processing in the Cerebral Cortex 685

could then either be a causal factor for or a conse- The design of the study was to examine brain
quence of the chronicity of the pain condition. activity for thermal stimuli applied to the body part
A recent meta-analysis in fact shows that across where CRPS pain was present, and compare brain
some 100 studies one can establish statistically signifi- responses to this stimulus between CRPS and healthy
cant differences in incidence of different brain areas subjects. Moreover, as the pain in CRPS patients with
activated by experimental painful stimuli between sympathetically maintained pain (SMP) may be
acute and chronic pain conditions: PRC shows a stron- modulated by a sympathetic block, it was reasoned
ger activation in chronic pain patients, whereas other that one could decrease the patients ongoing pain
nociceptive cortical areas and the thalamus show a and then re-examine brain activity responses to the
weaker response (Table 2). A simple interpretation same stimulus. The study was done in a small group
of these findings would be that nociceptive signal of patients and this by itself is an important weakness.
processing for experimental painful stimuli in chronic The main observation was that thermal stimuli in
pain patients involves a reduced sensory discrimina- CRPS evoked more prefrontal cortical activity than
tive component and an increased affective- usually seen in healthy subjects, and this was reversed
motivational or cognitive-evaluational component. (became more similar in pattern to normal subjects
That interpretation would also be consistent with the brain activity to thermal stimuli) following sympa-
stronger affective component of clinical pain as com- thetic blocks. The introduction of sympathetic blocks
pared to experimental pain (Chen, A. C. N. and necessitated the use of the same procedure in healthy
Treede, R. D., 1985). But there are further implica- subjects as well, where its effects were minimal. The
tions: Is the result a consequence of some trivial study also observed that when a placebo block
confounds or does it signify changes in the physiology resulted in decreased pain perception then the cor-
of pain? One could construct a long list of confounds tical response pattern changed similarly to that of
effective blocks. These results show that brain activ-
that may underlie the observation, from attentional
ity may be distinct between CRPS and healthy
shifts, to coping mechanisms, to effects of drug use,
subjects for thermal stimuli.
and heightened anxiety and depression.
The standard approach for studying brain activity
for acute pain is to induce pain by a mechanical or
thermal stimulus and determine brain regions modu- 45.8.2 Clinical Pain Conditions Studied by
lated with the stimulus period and even with the Stimulation and the Role of the Cortex
various intensities used. Therefore, it is natural to A direct approach to studying clinical pain states is to
carry the same technology to the clinical arena and provoke it and examine brain activity. This is doable
apply it to chronic pain patients. As an example, we by drugs in headaches and in cardiac pain. As a result
discuss one study which attempted to identify brain there is growing literature in both fields. There is also
activity in complex regional pain syndrome (CRPS) now good evidence that migraine with aura is accom-
patients using fMRI (Apkarian, A. V. et al., 2001a; panied with decreased blood flow and decreased
2001b). activity in the occipital cortex, and migraine with or

Table 2 Frequency of brain areas active during pain in normal subjects as compared to patients with clinical pain
conditions

ACC SI SII IC Th PFC

Pain in normal subjects in 68 47/54 (87%) 39/52 (75%) 38/51 (75%) 45/48 (94%) 28/35 (80%) 23/42 (55%)
studies
Clinical pain conditions in 30 13/29 (45%) 7/25 (28%) 5/25 (20%) 15/26 (58%) 16/27 (59%) 21/26 (81%)
studies
Comparison between pain in P < 0.001 P < 0.001 P < 0.001 P < 0.001 P 0.095 P 0.038
normal subjects and in
clinical conditions

Incidence values are based on positron emission tomography, single photon emission-computed tomography, and functional magnetic
resonance imaging studies. For details see Apkarian et al., 2005. P values are based on Fishers exact statistics contrasting incidence for
each area. ACC, anterior cingulate cortex; IC, insular cortex; PFC, prefrontal cortex; SI, primary somatosensory cortex; SII, secondary
somatosensory cortex; Th, thalamus.
686 Nociceptive Processing in the Cerebral Cortex

without aura is associated with increased cortical the headache-free state only hypothalamic activity
thickness in visual cortical regions involved in was distinct (May, A. et al., 2000). This highly sig-
motion detection (Granziera, C. et al., 2006). nificant activation was not seen in cluster headache
patients out of the bout when compared to the
patients experiencing an acute cluster headache
45.8.2.1 Migraine
attack. In contrast to migraine, no brainstem activa-
Migraine attacks are characterized by unilateral tion was found during the acute attack compared to
severe headache often accompanied by nausea, pho- the resting state. Newer MRS results further sub-
nophobia and photophobia. Activation of the
stantiate this idea by showing reduced metabolites
trigeminovascular system is thought to be responsible
within the hypothalamus of cluster headache patients
for the pain itself, and cortical spreading depression
in contrast to healthy or migraine headache controls
(CSD) seems to underlie the aura symptoms. This
(Wang, S. J. et al., 2006). These data suggest that while
view has been greatly advanced and substantiated by primary headaches such as migraine and cluster
brain imaging studies. fMRI studies show CSD-typi- headache may share a common pain pathway, the
cal cerebrovascular changes in the cortex of trigeminovascular innervation, and activate similar
migraineurs while experiencing a visual aura cortical regions, the underlying pathogenesis may
(Hadjikhani, N. et al., 2001). The subsequent decrease
be quite different.
in fMRI signal is temporally correlated with the
scotoma that follows the scintillations. These fMRI
signal changes develop first in the extrastriate cortex, 45.8.2.3 Cardiac pain
contralateral to the visual changes. It then slowly Cardiac pain and its variants have been studied by
migrated towards more anterior regions of the visual brain imaging using various drugs that bring about
cortex, representing peripheral visual fields, in agree- these symptoms (Rosen, S. D. et al., 1996; 2002). In
ment with the progressive movement of the patients with myocardial ischemia the perception of
scintillations and scotoma from the centre of vision angina is associated with activity in the hypothala-
towards the periphery. A recent study that analyzed mus, PAG, thalami, rostral ACC, and bilateral PRC.
visually triggered attacks showed hyperemia in the In patients with silent myocardial ischemia it seems
occipital cortex, independently of whether the head- that the silence is not due to impaired afferent signal-
ache was preceded by visual symptoms (Cao, Y. et al., ing, but rather it is associated with a failure of
1999). An alternative view considers migraine aura transmission of signals from the thalamus to the
and headache as parallel rather than sequential pro- frontal cortex. In contrast, in patients with syndrome
cesses, and proposes that the primary cause of X, a condition of chest pain with ischemiclike stress
migraine headache is an episodic dysfunction in brain- electrocardiography but entirely normal coronary
stem nuclei that are involved in the central control of angiogram, activity in the right anterior insula dis-
nociception (Goadsby, P. J. et al., 2002). tinguished these patients from patients with known
coronary disease. These patients appear to have a
deficit in central pain habituation (Valeriani, M.
45.8.2.2 Cluster headache
et al., 2005). Overall, these studies imply that differ-
The pathophysiology of cluster headache is thought
ence between different cardiac pain conditions are
to involve multiple brain regions. Brain imaging stu-
due to central processing, e.g., syndrome X is inter-
dies imply that the associated excruciatingly severe
preted as a cortical pain syndrome, a top-down
unilateral pain is likely mediated by activation of the
process, in contrast with the bottom-up generation
first (ophthalmic) division of the trigeminal nerve,
of a pain percept caused by myocardial ischemia in
while the autonomic symptoms are due to activation
coronary artery disease.
of the cranial parasympathetic out-flow from the
VIIth cranial nerve. The circadian rhythmicity of
cluster headache has led to the concept of a central 45.8.2.4 Irritable bowel syndrome
origin for its initiation (Strittmatter, M. et al., 1996). Irritable bowel syndrome (IBS) is a disorder of
Using PET in cluster headache patients, signifi- abdominal pain or discomfort associated with bowel
cant activations ascribable to the acute cluster dysfunction. Hypersensitivity to visceral, but not
headache were observed in the ipsilateral hypotha- somatic, stimuli has been demonstrated in IBS. A
lamic gray matter and in multiple cortical areas number of groups have examined brain activity in
including cingulate and PRC. When compared to this condition mainly by monitoring responses to
 Nociceptive Processing in the Cerebral Cortex 687

painful and nonpainful rectal distensions, as well as complicated by the fact that the authors do not take
responses to the anticipation of painful distensions. into consideration the influence of spontaneous pain.
Two studies are interesting since both show in nor- Still, this is perhaps one of the best-controlled IBS
mal subjects a significant positive correlation studies, and indicates distinct cortical areas involved
between cingulate cortex activity and subjective rat- in the urge and pain perceptions in each group.
ing of rectal distension pain, and in both studies this There is now evidence that serotonin (5-HT) may
relationship completely disappears in IBS patients be involved in IBS. One study (Nakai, A. et al., 2003)
(Silverman, D. H. et al., 1997; Mertz, H. et al., 2000). examined serotonin synthesis in the brain and indi-
IBS is more prevalent in women than in men. Brain cated greater brain regional serotonin synthesis
imaging studies have now shown gender differences in female IBS. There is also evidence that alosetron,
in brain activity in IBS (Berman, S. M. et al., 2000; a 5-HT3 receptor antagonist, is clinically effective
Naliboff et al., 2001; Berman, S. M. et al., 2002b; Nakai, in treating some subtypes of IBS. Berman S. M. et al.
A. et al., 2003). There are large differences between (2002a) examined brain activity in a large population
the studies, making their synthesis difficult (see of IBS, before and after a randomized, placebo-con-
Ringel, Y., 2006). trolled, 3-week use of alosetron. Treatment
More recent studies show a hint for sensitization improved IBS symptoms and regional cerebral
in IBS patients because subliminal and supraliminal blood flow in brain regions rich with 5-HT3 receptor
distensions of rectal distension seem to indicate small and involved in emotional and aversive functions:
differences between IBS and healthy controls in the amygdala, ventral striatum, and dorsal pons, imply-
total cortical volume activated or in regional activity ing that the therapeutic effects are due to central
as a function of distention volume (Andresen, V. et al., actions and not peripheral. Thus, generally the IBS
2005; Lawal, A. et al., 2006). A study of IBS in contrast studies show that brain responses are different to
to healthy subjects examined thermal and visceral rectal stimuli in patients, and that these central
hyperalgesia and related brain activity (Verne, G. N. events may be critical to IBS.
et al., 2003). This seems the only study where besides
pain intensity and unpleasantness measures, the
45.8.3 Spontaneous Pain as a Confound in
authors also document fear and anxiety and show
Assessing Brain Activity
that all are rated higher by IBS for both heat and rectal
distention, and not surprisingly these increased sensa- A person who has lived for years in the presence of
tions and emotions give rise to larger cortical pain, must have developed some coping mechanisms
activations in IBS. The latter is most likely a reflection that aid in pursuing other everyday life interests in
of a perceptual magnitude mismatch between the spite of the presence of the pain. How does this
groups and says little as to the IBS cortical activity impact the brain? Can one consider the patient in
abnormalities. Such mismatches at least for fear and chronic pain as composed of a brain-signaling pain
anxiety most likely are common in the majority of together with a brain undertaking other tasks as in
studies of IBS. One assumes that the simple introduc- healthy subjects? Or, does the presence of ongoing
tion of a rectal balloon in IBS would result in increased pain interact and impact other processes as well?
anxiety, which undoubtedly effects cortical activity to Certainly our cognitive and anatomic studies suggest
visceral and somatic pain, yet its specific contribution that the latter is more likely. We have now direct
has remained unexplored. evidence of the modulation that ongoing pain
In a more elegant study the authors use percep- imposes on brain activity in general.
tion-related ratings during rectal distention to evoke A recent study reported brain activity for sponta-
either urge to defecate or pain, and compared brain neous pain in PHN patients before and after topical
activity related to the ratings between IBS patients lidocaine treatment (Geha, P. Y. et al., 2007). The
and healthy subjects. (Kwan, C. L. et al., 2005). The PHN patients were imaged before, after 6-hours
approach is similar to the technique used in mapping and 2-weeks treatment with lidocaine. Behaviorally
brain activity for spontaneous pain in chronic back and based on questionnaires most participants
pain (CBP) and in postherpetic neuralgia (PHN; showed a modest but significant decrease in their
Baliki, M. N. et al., 2006; Geha, P. Y. et al., 2007). ongoing pain. The patients were scanned while they
The results show large differences between the two were either rating their ongoing pain or rating a
groups contrasted, with far more extensive brain visual bar that varied in time in a pattern that
activations in the healthy subjects. The results are mimicked their ratings of pain (Figure 8). Thus, the
688 Nociceptive Processing in the Cerebral Cortex

Pain 2

Z-score
2

4
0 20 40 60 80 100
R PP Pain intensity
MPFC

2
Visual

Z-score
2

4
0 20 40 60 80 100
Pain intensity

6.0 2.3 2.3 6.0

Figure 8 Intensity of ongoing pain changes brain activity and thus cognitive processing. Eleven postherpetic neuralgia
patients were studied by functional magnetic resonance imaging (fMRI) once before and twice after lidocaine application on
the painful skin. In all sessions, patients performed two different tasks: in the Pain task they continuously rated the fluctuations
of their spontaneous pain, and brain activity related to this was identified using methods. In the Visual task they rated
fluctuations of a bar varying in time, brain activity was determined with the same approach as for the pain task. The relationship
between brain activity and intensity of ongoing pain was determined by using a covariate analysis, where for each fMRI scan its
related pain intensity was used to determine the effect of this parameter on brain responses. Across-subject and across all
scans average variation of brain activity is displayed for both tasks in the left. Red are brain regions that are positively
correlated and blue regions that are negatively correlated with intensity of ongoing pain (normalized to z-values). The right
scatter-grams show this effect for two brain regions (right posterior parietal cortex, R PP, x 33 y 45 z 50; and medial
prefrontal cortex, MPFC, x 9 y 50 z 40, as respectively circled). Each dot represents a single patients activity at a single
time. Top scatter-gram is for Pain task; bottom for Visual task, red symbols and regression line are for MPFC; blue for R PP.
MPFC exhibited significant positive correlations with pain intensity for pain ( r 0.49, P < 0.05) and visual (r 58, P < 0.01) task,
while R PP showed negative correlation for pain (r 0.48, P < 0.05) and visual (r 0.64, P < 0.01) task. Brain areas that show
increased correlation with ongoing pain are interpreted as a functional compensation for the decreased attentional resources.
Reproduced from Geha, P. Y., Baliki, M. N., Chialvo, D. R., Harden, R. N., Paice, J. A., and Apkarian, A. V. 2007. Brain activity
for spontaneous pain of postherpetic neuralgia and its modulation by lidocaine patch therapy. Pain 128, 88100.

latter is a control task that captures motor and the pain condition, hinting that the effects of
cognitive parts of the task but, of course, it does decreased pain was modulating more than just pain-
not reflect the pain. Brain activity for both tasks related circuitry.
was increasing from first to third session. This To identify the role of spontaneous pain on
observation is similar to earlier reports that decrease brain activity in general, a correlation analysis was
in clinical pain in many cases results in increased done for both tasks with mean spontaneous pain.
brain activity. In this case, however, the internal Figure 8 shows the influence of pain intensity on
control was also changing in a manner parallel to across-sessions averaged brain activity for both tasks.
 Nociceptive Processing in the Cerebral Cortex 689

The resultant map is generally similar for both tasks: therapeutically imperative. Cortical responses to
activity in medial and lateral prefrontal regions was standard mechanical or thermal stimulation are of
positively correlated, while posterior parietal atten- limited value for understanding these clinical pain
tional areas were negatively correlated with mean conditions. Spontaneous pain fluctuates unpredicta-
pain intensity. This result shows that brain activity bly in the time scale of seconds to minutes, and these
for both tasks is influenced by the level of spontaneous fluctuations have characteristic properties that differ-
pain, implying that pain intensity influences task per- entiate between different chronic pain conditions
formance in general. This is in line with previous such as PHN and CBP (Foss, J. M. et al., 2006). This
studies showing that ongoing pain may interfere with variability (specific fractal dimension) can also be
cognitive functions (Lorenz, J. et al., 1997). observed in fMRI signals when such patients rate
This result reinforces the need for correcting their spontaneous pain. Therefore, this technique
brain activity by a control condition performed at was applied to study brain activity in CBP (Baliki,
the same pain level that is the necessity of subtracting M. N. et al., 2006) and PHN patients (Geha, P. Y. et al.,
the visual task from spontaneous pain rating task, at 2007) in relation to their subjective report of fluctua-
each treatment session. For both tasks, the fact that tions of spontaneous pain.
posterior parietal cortical activity was negatively The combination of relating brain activity to spon-
correlated with mean ongoing pain suggests that the taneous pain and correcting for confounds by
attentional abilities of patients are directly related to subtracting brain activity for visual bar lengths, pro-
the intensity of their pain, which would in turn vides a robust approach with which clinical pain may
impact their abilities in performing anything that be studied directly. Note that in this case the brain
would demand concentration. activity is related to exactly the event that the patient
Moreover, multiple prefrontal regions were posi- complains about. With this approach, in CBP patients
tively correlated to the mean pain, suggesting that (Baliki, M. N. et al., 2006) it was shown that the brain
the patients brain regions underlying higher cogni- regions activated when the pain was increasing corre-
tive functions become more active as the pain spond to brain regions seen for acute pain in normal
intensity increases. The exact cognitive implications subjects. In contrast, for time periods when the pain
for these brain activity patterns remain unclear. In was high and sustained, the brain activity was mainly
contrast, the finding indicates that the intensity of limited to medial PRC, a region usually not activated
spontaneous pain impacts brain activity for any task for acute pain. The resultant brain activity was
that the subject attempts to perform, enhancing some strongly correlated to the patients reported pain
aspects and inhibiting others. Therefore, the intensity at the time of the scan, specifically with
decreased brain activity reported for pain tasks in medial prefrontal activity. Also, the duration or
many clinical pain conditions (Peyron, R. et al., chronicity of the pain was captured in the insular
2000; Derbyshire, S. W., 2003; Apkarian, A. V. et al., activity, a region activated only during increases in
2005; Kupers, R. and Kehlet, H., 2006) is most likely a spontaneous pain. Thus, two fundamental properties
reflection of the presence of the spontaneous pain, of CBP its intensity and duration were directly
and is not specific to the task being investigated. reflected in the brain activity identified in these
The fact that pain intensity seems to modulate patients. By applying a thermal painful stimulus in
brain activity in general has another powerful conse- the same patients (as well as in healthy subjects) the
quence. It suggests that simply studying brain activity, same study showed that brain regions reflecting the
in tasks unrelated to pain, one should be able to stimulus intensity were not related to that reflecting
identify the presence of pain and study its effects on the intensity of spontaneous pain. In turn, the brain
sensory/cognitive/motor/attentional processing, an region that reflected spontaneous pain intensity was
exciting prospect that remains to be pursued. only activated for the latter and did not reflect thermal
painful stimulus intensity. Therefore, at least in the
patient group studied spontaneous pain involved a
45.8.4 Functional Magnetic Resonance
different brain activity pattern than acute pain.
Imaging of Spontaneous Pain
Spontaneous pain is highly prevalent in clinical pain
45.8.5 Neuropathic Pain
conditions, and is usually the primary drive for
patients seeking medical care. Thus, understanding Patients with neuropathic pain show decreased
its related brain circuitry is both scientifically and responses in the thalamus to experimental painful
690 Nociceptive Processing in the Cerebral Cortex

stimuli (Peyron, R. et al., 2000). A MRS study showed after pain relief like the medial PRC (Hsieh, J. C.
a decrease in the level of N-acetyl-aspartate, a neu- et al., 1999a). This heterogeneity is not surprising
ronal marker, in the thalami of patients with chronic because pattern of brain activity may be specific to
neuropathic pain after spinal cord injury (SCI), when each neuropathic pain condition.
compared to patients with SCI but without pain
(Pattany, P. M. et al., 2002). Thus, neurochemical
45.8.6 Low Back Pain and Fibromyalgia
brain imaging provides evidence for the occurrence
of long-term changes in the brain chemistry and As mentioned above, brain activity of healthy sub-
morphology of chronic neuropathic pain patients. jects and patients with increased pain sensitivity
Thalamic activity in neuropathic patients was also should be compared in such a way that perceived
reported to increase after pain relief (Hsieh, J. C. et al., intensity has been matched across the two groups. A
1995), and to be significantly negatively correlated recent study used such a design and showed gener-
with the duration of the condition in CRPS patients ally heightened brain activity for painful stimuli of
(Fukumoto, M. et al., 1999). Thus, the reduced equivalent perceptual intensity both in fibromyalgia
activation of the thalamus may also be an altered and CBP patients as compared to healthy subjects
functional state rather than an irreversible degenera- (Gracely, R. H. et al., 2002; Giesecke, T. et al., 2004).
tion. Neuropathic pain patients in addition show a Morphometric and neurochemical brain imaging
reduced availability of opioid receptor binding sites studies provide evidence for the occurrence of
(Maarrawi, J. et al., 2007). This reduction was sym- long-term changes in the brain chemistry and
metric in peripheral neuropathic pain, suggesting a morphology of chronic pain patients. The level of
possible release of endogenous opioids, but latera- N-acetyl-aspartate, a neuronal marker, was decreased
lized to the hemisphere contralateral to the pain in in the medial and lateral PRC of CBP patients com-
central pain patients, consistent with a loss of recep- pared to an age- and gender-matched control group
tors (Jones, A. K. P. et al., 2004, Willoch, F. et al., 2004). (Grachev, I. D. et al., 2000). A morphometric study in
Brain activity differences between healthy sub- chronic pain showed also a decrease in gray matter
jects and patients in activation paradigms are density in the dorsolateral PRC and the thalamus of
difficult to interpret since they do not distinguish CBP patients when compared to matched controls
between brain activity specifically related to the clin- (Apkarian, A. V. et al., 2004). Furthermore, these long-
ical condition and abnormalities in sensory term chemical and morphological changes are signif-
processing secondarily associated with the clinical icantly correlated with different characteristics of
state. Particularly in neuropathic pain, the accompa- pain such as pain duration (Apkarian, A. V. et al.,
nying sensory deficit may be reflected in the imaging 2004), pain intensity (Pattany, P. M. et al., 2002;
results and not the pain. Reduced relevance of the Grachev, I. D. et al., 2002; Apkarian, A. V. et al.,
acute stimulus to subjects who are already in pain 2004), and sensory-affective components (Grachev,
may also account for much of the decreased regional I. D. et al., 2002). The morphometric and neurochem-
brain activity in neuropathic pain. To overcome such ical studies imply an active role of the central
nonspecific brain activity differences one needs to nervous system in chronic pain, suggesting that
compare brain activity for stimuli where perceptual supraspinal reorganization may be critical for chronic
evaluation has been equated between patients and pain.
normal healthy subjects.
Three studies (Hsieh, J. C. et al., 1995; 1999a;
45.8.7 Overview Regarding the Role of the
Apkarian, A. V. et al., 2001b) have looked at the
Cortex in Chronic Pain Perception
regions of the brain modulated by relief of chronic
neuropathic pain: CRPS, peripheral neuropathy, and In spite of a plethora of data there remains a host of
trigeminal neuropathy. Two of these studies show uncertainties about their significance. Overall, the
that the PRC activity is decreased, and all three clinical brain imaging studies indicate reduced infor-
studies report decreased rostral ACC activity, after mation transmission through the thalamus to the
successful pain relief. It is to be noted that in addition cortex, and increased activity in PFC, mostly in
to those regions some areas were also less activated medial PFC coupled with atrophy in dorsolateral
with pain relief such as the insula (Hsieh, J. C. et al., PFC. The number of studies remain very small and
1995) and the anterior limbic thalamus (Hsieh, J. C. hence our confidence as to the reproducibility of
et al., 1999a), whereas others were more activated these changes remain minimal. Still, the observations
 Nociceptive Processing in the Cerebral Cortex 691

regarding cortical and thalamic activity changes in function lies in the parasylvian cortex, in the vicinity
chronic pain are in general consistent with the notion of SII and the dorsal insula. In chronic pain, nocicep-
that chronic pain conditions preferentially engage tive processing in the cerebral cortex is partly
brain areas involved in cognition/emotion and preserved and partly altered, in particular with
decreases activity in regions involved in sensory respect to PRC functions. This reorganization may
evaluation of nociceptive inputs. be a neuroplastic response to the chronicity of pain, it
Evidence has been presented that brain activity, may reflect activation of antinociceptive processes, or
chemistry, and morphology may be reorganized in it may even represent a predisposing factor for the
chronic pain conditions. Does this evidence imply development of chronic pain. The methods available
that there is supraspinal reorganization, above and for the study of nociceptive processing in the brain
beyond what is established in the periphery and allow to address many of these open questions in the
spinal cord? That is, even if we establish a brain near future, and this part of pain research is bound to
pattern of activity for some chronic pain condition, remain a very productive one.
does this reflect some unique contribution of the
brain to this state or is it simply a reflection of
lower level reorganization? The answer is not
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 46 Phantom Limb Pain
H Flor, Central Institute of Mental Health, Mannheim, Germany
2009 Elsevier Inc. All rights reserved.

46.1 Definition 699

46.2 Peripheral Mechanisms of Phantom Limb Pain 700
46.3 Central Factors 701
46.3.1 The Spinal Cord 701
46.3.2 Supraspinal Changes 702
46.4 Implications for the Treatment and Prevention of Phantom Limb Pain 703
46.5 Future Developments 704
References 704

Glossary
cortical reorganization Changes in the maps of tingling or cramping sensations that can be distin-
the primary sensory or motor areas of the cortex guished from phantom sensation as well as from
related to injury or stimulation. Cortical represen- painful sensation.
tations can increase or decrease in size or they can phantom limb pain or phantom pain Pain in a
shift to other locations on the sensory or motor body part that is no longer present. It may be
map. related to a certain position or movement of the
central sensitization Short- or long-term changes phantom and may be elicited or exacerbated by a
in the excitability of central neurons and their range of physical (e.g., changes in weather or
synaptic strength that lead to an enhanced or pressure on the residual limb) and psychological
altered processing of peripheral sensory input factors (e.g., emotional stress). It seems to be more
resulting in hypersensibility. intense in the distal portions of the phantom and
neuroma When a limb is severed, a terminal may have a number of different qualities such as
swelling or endbulb is formed with axonal sprouting stabbing, throbbing, burning, or cramping.
occurring. In the case of an amputation this Phantom limb pain is often confused with pain in
sprouting and endbulb formation lead to neuroma, the area adjacent to the amputated body part.
a tangled mass that is formed when the axons postamputation pain Pain at the site of the
cannot reconnect or can only partially reconnect as wound that must be distinguished from pain in the
is the case in partial lesions. These neuroma gen- residual limb and phantom limb pain that may all
erate abnormal activity that is called ectopic co-occur in the early phase after amputation.
because it does not originate from the nerve preamputation pain Pain that occurred in the
endings. amputated body part before the amputation. It may
nonpainful phantom limbs and nonpainful be related to the incidence, type, and severity of
phantom sensations These phenomena refer to phantom limb pain in the phase following the
the continued presence of the limb (corporeal amputation.
awareness of the limb) and nonpainful sensations residual limb pain or stump pain Pain in the body
such as tingling or pressure sensations. part adjacent to the amputation. It is usually posi-
nonpainful residual limb sensations The residual tively correlated with phantom limb pain.
limb can also have nonpainful sensations such as

46.1 Definition of the continued presence of the limb but also non-
painful and painful phantom sensations such as the
The amputation of a body part is often followed by the feeling of a specific position, shape, or movement of
sensation that the deafferented body part is still pre- the missing limb, feelings of warmth or cold, itching,
sent. These sensations may include not only the feeling tingling or electric sensations, and other paresthesias.

699
700 Phantom Limb Pain

Phantom limb pain (or phantom pain) is pain in the increased excitability and often show spontaneous
body part that is no longer present. This occurs to some impulse activity (ectopic discharge). Mechanical,
degree in 5080% of all amputees (Nikolajsen, L. and chemical, and thermal stimulation may further
Jensen, T. S., 2006). Phantom limb pain may be related exacerbate this ectopic discharge. The increased
to a certain position or movement of the phantom and excitability of injured nerves that results in ectopic
may be elicited or exacerbated by a range of physical discharge seems to result from upregulation or novel
(e.g., changes in weather or pressure on the residual expression, and altered trafficking, of molecules that
limb) and psychological factors (e.g., emotional stress). are responsible for neuronal excitability, such as vol-
It seems to be more intense in the distal portions of the tage-sensitive sodium channels (Devor, M., 2006). In
phantom and may have a number of different qualities addition, abnormal connections between injured
such as stabbing, throbbing, burning, or cramping (Hill, axons, such as ephapses, may contribute to the spon-
A., 1999). Phantom limb pain belongs to the neuro- taneous ectopic activity. Phantom limb pain is often
pathic pain syndromes and is assumed to be related to present very soon after amputation before a palpable,
damage to the axons of peripheral neurons with sec- swollen neuroma could have formed. However, ecto-
ondary changes induced in central neurons. Phantom pic discharge also appears rapidly, apparently
body pain may also occur following spinal cord injury. originating at first in swollen endbulbs at the cut
Phantom limb pain is infrequent if the amputation axon end rather than in outgrowing sprouts.
occurred at a very young age. However, older children Local anesthesia of the stump does not eliminate
exhibit as high an incidence of phantom limb pain as phantom limb pain in all amputees (Birbaumer, N.
adults. Although phantom sensations seem to occur et al., 1997). This fact motivated a search for other
occasionally in congenital amputees (i.e., those born potential sources of ectopic input from the periphery.
without a limb), phantom limb pain seems to be very An additional site of ectopic discharge is the dorsal
rare under these circumstances (Flor, H., 2002). The root ganglion (DRG). Ectopia originating in the DRG
long-term course of phantom limb pain is unclear. can summate with ectopia originating from neuromas
While some authors report a slight decline in pain in the stump. Indeed, processes such as cross-excita-
prevalence over the course of several years, others tion can lead to the depolarization and activation of
have described high prevalence rates also in long- neighboring neurons, significantly amplifying the
term amputees. Both peripheral and central factors overall ectopic barrage (Devor, M., 2006). In experi-
have been discussed as determinants of phantom limb mental preparations and in humans it was found that
pain. Psychological factors do not seem to be a primary anesthetic block of neuromas eliminates spontaneous
cause, but they may well affect the course and the and stimulation-induced nerve activity related to the
severity of the pain (Sherman, R. A., 1997). The general stump, but not ectopic activity potentially originating
view today is that multiple changes along the neuraxis in the DRG (Nystrom, B. and Hagbarth, K. E., 1981).
contribute to the experience of phantom limb pain. Interestingly, there is evidence for genetic factors in
the predisposition to develop ectopic neuroma, DRG
discharge, and neuropathic pain. For example, Devor
46.2 Peripheral Mechanisms of M. et al. (2005) presented evidence for the presence of
Phantom Limb Pain several genes that predisposes to the pain behavior
that follows peripheral neurectomy in rodents. This
Peripheral changes that give rise to nociceptive input neuroma model of neuropathic pain has been consid-
from the residual limb have been viewed as an impor- ered to be a valid animal surrogate of phantom limb
tant determinant of phantom limb pain. This is pain in humans.
supported by the moderately high correlation Elevated sympathetic discharge, as well as
between pain in the residual limb (stump) and phan- increased levels of circulating epinephrine, can trig-
tom limb pain. Ectopic discharge from stump ger and exacerbate ectopic neuronal activity from
neuromas has been postulated as a potential source neuromas (Devor, M., 2006). In addition to such
of such nociceptive input (Devor, M., 2006). When sympatheticsensory coupling at the level of the
peripheral nerves are cut or injured, terminal swel- neuroma, sympatheticsensory coupling also occurs
ling and regenerative sprouting of the injured axon at the level of the DRG. This may account for the
end occur. In this process, neuromas form in the frequent exacerbation of phantom pain at times of
residual limb. The disorganized endings of C fibers emotional distress. Additional factors such as tem-
and demyelinated A fibers in neuromas have perature, oxygenation level, and local inflammation
 Phantom Limb Pain 701

in neuromas and associated DRGs may also play a glycinergic interneurons in the spinal cord may actu-
role. The sympathetic maintenance of phantom limb ally be destroyed by rapid ectopic discharge or other
pain in some patients is supported by evidence that effects of axotomy, contributing to a hyperexcitable
systemic adrenergic blocking agents, and targeted spinal cord (Scholz, J. and Woolf, C., 2006). Changes
chemical or surgical blockade of sympathetic nerves have also been noted in ascending projection neu-
and ganglia, sometimes reduce phantom limb pain. rons. The cascade of biological events that take place
Likewise, injections of epinephrine into stump neu- in the spinal cord after peripheral nerve damage may
romas have been shown to increase phantom limb trigger abnormal firing of spinal origin. Part of this
pain and paresthesias in some amputees. Although sensitization is due to facilitation of the response of
sympathetically maintained pain does not necessarily N-methyl-D-aspartic acid (NMDA) receptors to
covary with regional sympathetic abnormalities, in the primary afferent neurotransmitter glutamate
some patients, sympathetic dysregulation in the resi- (Sandkuhler, J., 2000). A remarkable effect of the
dual limb is apparent. Reduced near-surface blood spinal changes evoked by nerve injury is that low-
flow to a limb has been implicated as a predictive threshold afferents may become functionally con-
physiological correlate of burning phantom limb nected to ascending spinal projection neurons that
pain. Correspondingly, onset and intensity of cramp- carry nociceptive information. When this happens,
ing and squeezing descriptions of phantom pain have normally innocuous A-fiber input from the periph-
been related to muscle tension in the residual limb. ery, ectopic input as well as input from residual intact
This relationship seems not to hold for any other low-threshold afferents, may contribute to phantom
descriptors of phantom pain. In addition, lower skin pain sensation.
temperature in the residual limb in amputees with A number of additional central nervous system
phantom limb pain and phantom sensation and a (CNS) processes are thought to contribute to the
close relationship between phantom limb sensation hyperexcitability of spinal cord circuitry following
and skin conductance responses in the residual limb major nerve damage. For example, there may be
can be viewed as evidence of a sympathetic-efferent downregulation of opioid receptors, both on primary
somatic-afferent mechanism (for review see afferent endings and on intrinsic spinal neurons. This
Sherman, R. A., 1997). is expected to add to disinhibition due to the reduc-
tion of GABA and glycine activity. In addition,
cholycystokinin, an endogenous inhibitor of the opi-
46.3 Central Factors ate receptor, is upregulated in injured tissue (for
review see Woolf, C. J. and Salter, M., 2006).
46.3.1 The Spinal Cord
Another interesting example of changed gene expres-
Anecdotal evidence in human amputees first sug- sion after axotomy is the appearance of the
gested that spinal mechanisms may play a role in neuropeptide substance P in low-threshold A neu-
phantom limb pain. For example, during spinal rons. Substance P is normally expressed only by A
anesthesia, phantom pains have been reported to and C afferents, most of which are nociceptors. The
reoccur in patients who were pain-free at the time injury-triggered expression of substance P by A
of the procedure. Experimental data in human ampu- fibers may render them more like nociceptors. For
tees are lacking, but increasing amounts of evidence example, it may permit ectopic or normal activity in
based on animal models of nerve injury are becoming A fibers to trigger and maintain central sensitiza-
available. tion. Changes such as these in gene expression in
Increased activity in peripheral nociceptors leads injured afferents (and in some postsynaptic spinal
to an enduring change in the synaptic responsiveness neurons), which result in a change in their function-
of neurons in the dorsal horn of the spinal cord, a ing (i.e., their phenotype), are referred to as
process called central sensitization (Woolf, C. J. and phenotypic switch. Recent work based on gene chip
Salter, M., 2006). Central sensitization is also trig- technology indicates that hundreds of genes are up-
gered by nerve injury such as occurs during or downregulated in DRG and spinal neurons fol-
amputation. For example, spinal changes associated lowing peripheral nerve injury.
with nerve injury include increased firing of the Following peripheral nerve injury, degeneration
dorsal horn neurons, structural changes at the central of central projection axon occurs. Massive deafferen-
endings of the primary sensory neurons, and reduced tation is observed when dorsal roots are injured, or
inhibitory processes. Inhibitory GABAergic and avulsed from the spinal cord. Deafferentation may act
702 Phantom Limb Pain

hand-in-hand with the central effects of peripheral

denervation to bring about the changes that contri-
bute to spinal hyperexcitability. A mechanism that
may be of special relevance to phantom phenomena
is the invasion of regions of the spinal cord function-
ally vacated from injured afferents. For example, in
the neuroma model in rats and cats, there is an
expansion of receptive fields on skin adjacent to the
denervated part of the limb, and a shift of activity
from these adjacent areas into regions of the spinal
cord that previously served the part of the limb Figure 1 The representation of lip movements in
that was functionally deafferented by the nerve amputees with phantom limb pain (left) and without
lesion (Devor, M. and Wall, P. D., 1978). Such reor- phantom limb pain (right) in primary somatosensory cortex
ganization of the spinal map of the limb, which is based on functional magnetic resonance imaging. Note that
the amputees with phantom limb pain activate both the
probably due to unmasking of previously silent con- cortical hand and the mouth representation whereas the
nections, is reflected in brainstem and cortical amputees without phantom limb pain activate only the
remapping also. cortical mouth representation.

peripheral input whereas in others central, poten-

46.3.2 Supraspinal Changes tially intracortical changes might be more important.
Supraspinal changes related to phantom limb pain It is so far not known to what extent spinal changes
involve the brain stem, the thalamus, and the cerebral contribute to these supraspinal alterations. It was
cortex (for review see Flor, H. et al., 2006). New shown that axonal sprouting in the cortex underlies
insights into phantom limb pain have come from the reorganizational changes observed in amputated
studies demonstrating changes in the functional and monkeys (Florence, S. L. et al., 1998). Thalamic stimu-
structural architecture of primary somatosensory lation and recordings in human amputees have
cortex subsequent to amputation and deafferentation revealed that reorganizational changes may also
in adult monkeys. In these studies, the amputation of occur at the thalamic level and are closely related to
digits in an adult owl monkey led to an invasion of the perception of phantom limbs and phantom limb
adjacent areas into the representation zone of the pain (Davis, K. D. et al., 1998). Studies in animals have
deafferented fingers. Several imaging studies (for shown that these changes may be relayed from the
review see Flor, H., 2002) have reported that upper spinal and brain stem level; however, changes on the
extremity amputees actually show a shift of the subcortical levels may also originate in the cortex,
mouth into the hand representation in primary soma- which has strong efferent connections to the thalamus
tosensory and motor cortex (Figure 1). Flor H. et al. and lower structures.
(1995) provided evidence that these cortical changes Sometimes pain in the phantom is similar to the
are less related to referred sensations but rather have pain that existed in the limb prior to amputation. The
a close association with phantom limb pain. The likelihood of this ranges from 10% to 79% in different
larger the shift of the mouth representation into the reports and depends on the type and time of assess-
zone that formerly represented the amputated hand ment (see Katz, J. and Melzack, R., 1990). The type
and arm the larger the phantom limb pain. These and time of assessment and potential errors in retro-
cortical changes could be reversed by the elimination spective reports are important determinants of the
of peripheral input from the amputation stump using incidence of these pain memories. It has been pro-
brachial plexus anesthesia. Peripheral anesthesia posed that pain memories established prior to the
completely eliminated cortical reorganization and amputation are powerful elicitors of phantom limb
phantom limb pain in half of the amputees that pain (Katz, J. and Melzack, R., 1990; Flor, H., 2002).
were studied. In the remaining half, both cortical Pain memories may be implicit and not readily acces-
reorganization and phantom limb pain remained sible to conscious recollection. The term implicit pain
unchanged (Birbaumer, N. et al., 1997). This result memory refers to central changes related to nocicep-
suggests that in some amputees, cortical reorganiza- tive input that lead to subsequent altered processing of
tion and phantom limb pain may be maintained by the somatosensory system and do not require changes
 Phantom Limb Pain 703

in conscious processing of the pain experience (Flor, H., mechanisms underlying the production of the pain
2002). In patients with chronic back pain it was (Sindrup, S. H. and Jensen, T. S., 1999). Most studies
shown that increasing chronicity is correlated with are uncontrolled short-term assessments of small
an increase in the representation zone of the back in samples of phantom limb pain patients. The maxi-
primary somatosensory cortex, and it was also mum benefit reported from a host of treatments such
reported that the experience of acute pain alters the as local anesthesia, sympathectomy, dorsal root entry
map in primary somatosensory cortex. These data zone lesions, cordotomy and rhizotomy, neurostimu-
suggest that long-lasting noxious input may lead to lation methods, or pharmacological interventions
long-term changes at the central and especially at the such as anticonvulsants, barbiturates, antidepressants,
cortical level. It has long been known that the pri- neuroleptics, and muscle relaxants seems to be
mary somatosensory cortex is involved in the around 30%. This does not exceed the placebo effect
processing of pain and that it may be important for reported in other studies. Controlled studies have
the sensory-discriminative aspects of the pain experi- been performed for opioids, calcitonin, ketamine,
ence. There have also been reports that phantom dextromethorphan, and gabapentin (see Nikolajsen, L.
limb pain was abolished after the surgical removal and Jensen, T. S., 2006), all of which were found to
of portions of the primary somatosensory cortex and effectively reduce phantom limb pain. Mechanism-
that stimulation of somatosensory cortex evoked based treatments are rare but have been shown to be
phantom limb pain. If a somatosensory pain memory effective in a few small but mostly uncontrolled
has been established with an important neural corre- studies. Lidocaine was found to reduce phantom
late in spinal and supraspinal structures, such as in limb pain of patients with neuromas in two small-
primary somatosensory cortex, subsequent deaffer- sample controlled studies. Biofeedback treatments
entation and an invasion of the amputation zone by resulting in vasodilatation of the residual limb or
neighboring input may preferentially activate corti- decreased muscle tension in the residual limb help
cal neurons coding for pain. Since the cortical area to reduce phantom limb pain and seem promising in
coding input from the periphery seems to stay patients where peripheral factors contribute to the
assigned to the original zone of input, the activation pain (Flor, H., 2002).
in the cortical zone representing the amputated limb Based on the findings from neuroelectric and
is referred to this limb and the activation is inter- neuromagnetic source imaging, changes in cortical
preted as phantom sensation and phantom limb pain. reorganization might influence phantom limb pain.
It is likely that not only the areas involved in sensory- Animal work on stimulation-induced plasticity
discriminative aspects of pain reorganize but also that would suggest that extensive behaviorally relevant
those areas that mediate affectivemotivational stimulation of a body part leads to the expansion of
aspects of pain such as the insula and the anterior its representation zone. It was shown that intensive
cingulate cortex undergo plastic changes that use of a myoelectric prosthesis was positively cor-
contribute to the experience of phantom pain. related with both reduced phantom limb pain and
A prospective study (Larbig et al., unpublished data) reduced cortical reorganization. When cortical reor-
showed that the best predictor of phantom limb pain ganization was partialled out, the relationship
12 months after an amputation is chronic pain before between prosthesis use and reduced phantom limb
the amputation thus supporting the pain memory pain was no longer significant suggesting that cor-
hypothesis. However, these authors tested a sample tical reorganization mediates this relationship. An
that did not include traumatic amputees but mainly alternative approach in patients where prosthesis
amputees with long-standing prior pain problems. use is not viable is the application of behaviorally
Further research is needed to better clarify these relevant stimulation. A 2-week training that con-
relationships. sisted of a discrimination training of electric
stimuli to the stump for 2 h per day led to signifi-
cant improvements in phantom limb pain and a
46.4 Implications for the Treatment significant reversal of cortical reorganization (Flor,
and Prevention of Phantom Limb Pain H. et al., 2001). A control group of patients who
received standard medical treatment and general
Several studies, including large surveys of amputees, psychological counseling in this time period did
have shown that most treatments for phantom limb not show similar changes in cortical reorganization
pain are ineffective and fail to consider the and phantom limb pain. The basic idea of the
704 Phantom Limb Pain

treatment was to provide input into the amputation Acknowledgment

zone and thus undo the reorganizational changes
that occurred subsequent to the amputation. The preparation of this article was supported by a
Mirror treatment (where a mirror is used to trick grant from the Deutsche Forschungsgemeinschaft
the brain into perceiving movement of the phantom (FL 156/29).
when the intact limb is moved) might be effective
but has so far only been tested in an anecdotal
manner.
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 47 Human Insular Recording and Stimulation
F Mauguiere, Lyon I University and INSERM U879, Bron, France
M Frot, INSERM U879, Bron France
J Isnard, Lyon I University and INSERM U879, Bron, France
2009 Elsevier Inc. All rights reserved.

47.1 Anatomy, Connections, and Physiology of the Insula in Primates 708

47.1.1 The Insula as the Fifth Lobe of the Brain 708
47.1.2 The Insula as a Node in a Distributed Cortical Network 709
47.1.3 The Insula as a Polymodal Area 710
47.1.4 The Insula as Part of the Mirror-Neuron System 710
47.2 Rational, Ethical Limitations and Procedure of Insular Recording
and Stimulations in Humans 710
47.2.1 Recording and Stimulation of the Human Insula Are Justified only in the Context of
Presurgical Evaluation of Epilepsy 710
47.2.2 Depth Intracortical Electrodes Are Needed to Explore the Insula 711
47.2.3 The Number of Electrodes Is Limited by Ethical Issues 711
47.2.4 The Site of Electrode Implantation Is Guided by Diagnostic Purposes 711
47.2.5 Only Data Recorded in Nonepileptogenic Cortex are Physiologically Relevant 711
47.3 Insular Recordings of Pain Evoked Responses in Humans 711
47.3.1 The Laser Stimulus 711
47.3.2 Cortical Surface Recordings (Electrocorticography) 712
47.3.3 Intracortical S2 Recordings 712
47.3.4 Intracortical Insular Recordings 712
47.3.5 Topographic Specificity of Insular Laser-Evoked Potentials 714
47.3.6 Interhemispheric Transmission of S2 and Insular Laser-Evoked Potentials 714
47.4 Insular Stimulation 714
47.4.1 The Challenge of Insular Stimulation 714
47.4.2 Pain Evoked by Insular Stimulation 715
47.4.3 Anatomical Location of the Insular Pain Area 715
References 715

Glossary
cytoarchitectony (cytoarchitectonic) primary sensory (visual, auditory, and somatosen-
Anatomical method based on microscopic sory) cortical areas (granular cortex), while large
observation of brain slices used to delineate pyramidal cells are characteristic of the primary
cortical areas according to the organization motor cortex (agranular cortex).
of the different types of gray matter cells (neurons). deep-brain stimulation Technique involving sti-
This technique permits the drawing of mulating the brain with electric current pulses of
cytoarchitectonic maps of the cortex, of which the low intensity using depth electrodes implanted
most widely used is that of Korbinian Brodmann, through the skull. In humans, this technique can be
first described in 1909, which counts 52 distinct used for therapeutic or diagnostic purposes. High-
areas in the human brain. The thickness of two frequency continuous (or chronic) deep-brain sti-
different types of cortical cells layers (granule and mulation is used as a treatment in various
pyramidal cells) and the number of layers are the neurological diseases, e.g., Parkinsons disease.
two main parameters used for cytoarchitectonic For diagnosis purposes, single pulses or trains of
mapping. Granule cells are over-represented in pulses are used mostly in presurgical assessment

707
708 Human Insular Recording and Stimulation

of patients with drug-resistant focal epileptic sei- somatotopy (somatotopic) Representation maps
zures in order to map the seizure focus (or of peripheral skin areas and joints in the somato-
epileptogenic zone) and the functional areas sur- sensory cortex that were first described in the
rounding the focus. Chronically implanted human brain by Wilder Penfield using electric sti-
electrodes are painless; they also permit the mulation of parietal cortex. The even existence of
recording of ongoing brain activity and evoked such maps demonstrates that granule cells receiv-
potentials. ing inputs from the same peripheral areas are
discharge (epileptic discharge, after- grouped together in the somatosensory cortex.
discharge) Abnormal electroencephalographic Microelectrode recordings in mammalian brain
(EEG) activity with abrupt onset and termination including monkeys have shown that separate
lasting at least several seconds seen during an somatotopic maps of the peripheral skin and joint
epileptic seizure; discharges can be recorded receptors can be drawn in the somatosensory cor-
either by scalp or by depth electrodes. They can tical areas.
occur spontaneously or be triggered by deep-brain somatosensory areas (S1, S2) Cortical areas
stimulation and then named after-discharges. containing a full somatotopic map of the contralat-
electroencephalography (EEG) The record of eral body half that have been identified in mammals
electrical activity of the brain taken by means of and human brain by means of electric cortical sti-
electrodes placed on the surface of the scalp. First mulation, evoked potentials, brain imaging
applied to study the activity of the human brain by (functional magnetic resonance imaging), and unit
Hans Berger in 1924, this noninvasive technique is or multiple cell responses. The human primary
still used routinely for the diagnosis of epilepsy. sensory area (S1) is located in the postcentral par-
evoked potentials (EPs) Brain electric response ietal gyrus and the second somatosensory area
(wave or complex of waves) elicited by and time- (S2) is located in the upper bank of the sylvian
locked to a physiological or nonphysiological sti- fissure (parietal operculum).
mulus, the timing of which can be reliably stereotaxy (stereotactic) The stereotactic space
assessed, e.g., a short duration. Laser beam first described by Jean Talairach is a three-dimen-
applied to the skin surface. Because of their low sional representation of the human brain in which
voltage most of EPs are not evident from the each point in the brain can be located according to
ongoing electroencephalographic (EEG) activity its coordinates (x, y, and z) in a Cartesian space.
after a single stimulation. Repeated stimulations The space is defined by the intersection of the mid-
and computer summation techniques are thus sagittal plane and the plane perpendicular to mid-
needed for their detection. The number of single sagittal plane passing through the anterior and
responses to be summated is a function of the posterior commissure of the brain (ACPC). Using
signal-to-noise ratio (ratio between the EP and depth electrodes implanted perpendicular to the
ongoing EEG activity voltages); this number is mid-sagittal plane, any given recording or stimula-
higher for scalp than for depth intracerebral tion site can be localized by its stereotactic
recordings. coordinates.

47.1 Anatomy, Connections, and Due to its anatomical situation in depth of brain lateral
Physiology of the Insula in Primates fissure (see Figure 1 and Tanriover, N. et al., 2004), and
its cytoarchitectonic organization the insula has long be
47.1.1 The Insula as the Fifth Lobe of
considered by anatomists as a rather isolated lobe of the
the Brain
brain mostly devoted to the processing of body and
In an article published in 1896 that was devoted to the visceral sensation including gustation, pain, and other
comparative anatomy of the insula Clark T. E. (1896) emotions, as well as to visceromotor and autonomic
quoted 39 synonyms used in anatomical literature to control. In primates including humans, the insula
name the fifth lobe of the brain buried in the lateral shows a caudorostral sequence of three distinct
sulcus and covered by the opercular parts of the frontal, cytoarchitectonic areas namely; (1) a large area of gran-
parietal, and temporal lobes, among which the term ular cortex, at its upper and posterior part, whose
insula, first proposed by Reil in 1804, has prevailed. structure is very similar to that of the second
 Human Insular Recording and Stimulation 709

(a) (b)

CS
A
ec cn

ic
p 30 mm
SF gp
SF
c ac
T1 ot
t1 B
a
T2 CS
ph
t2
C
T3 phig
fg

Figure 1 Anatomy of the human insular lobe (frontal slices). A) Anatomical fontal brain section showing the deep location of
the insula (in yellow) in the sylvian fissure (SF, white arrow), which separates the frontal and parietal lobes (above) from the
temporal lobe (below). The second somatosensory area (S2 in green), which occupies the superior bank of the SF, has
common borders with the lower part of the primary somatosensory area (S1 in purple), and the insula. The circular sulcus (CS)
separates the insula from the parietal operculum S2 and the first temporal gyrus (T1). The dashed red lines indicates the
trajectory of depth electrodes used for insular, parietal, and temporal lobes recordings in patients with epilepsy of the
temporal lobe and perisylvian cortex (see Isnard, J. et al., 2004). See also Figure 4 for sagittal view of the insula. B) Magnetic
resonance imaging (MRI) slice showing the projection of the recording electrodes implanted horizontally perpendicular to the
midline sagittal plane (dashed vertical white line). Recording contacts appears as small white segments along the electrodes
trajectory. The deepest contacts of electrodes A and B are located in the insular cortex. The depths coordinates (x) of each
contact is calculated from the midline sagittal plane in the human brain stereotactic space of Talairach. The x coordinates of
insular recording sites vary from 27 mm to 36 mm from midline. Electrode A (red circle) contacts explore from depth to
surface: the insula (yellow arrow), the S2 area (green arrow; 38 < x < 46 mm) and the lower part of the S1 area (purple arrow;
48 < x < 63 mm). Electrode B contacts explore the insula and the first temporal gyrus (T1 in Figure 1A). Electrode C contacts
explore the second temporal gyrus (T2 in Figure 1B) and hippocampus. a, amygdala; ac, anterior commissure; c, claustrum;
CS, circular sulcus; cn, caudate nucleus; ec, external capsule; fg, fusiform grus; gp, globus pallidus; ic, internal capsule; ot,
optic tract; phig, parahippocamplal gyrus; SF, sylvian fissure; T1, first temporal gyrus; t1, first temporal sulcus; T2, second
temporal gyrus; t2, second temporal sulcus; T3, third temporal gyrus.

somatosensory (S2) cortical area, which is involved fissure) but also by fuzzy cytoarchitectonic borders
mostly in the processing of somatosensory and pain with neighboring cortical areas and by a dense net-
sensation; (2) a transitional dysgranular field localized work of corticocortical connections with adjacent or
in its anterosuperior part involved in gustation and more distant cortical areas. Therefore its function
visceral sensation; and (3) an anteroventral agranular cannot be sketched as an isolated functional center,
field, which is in continuity with the temporal pole and as suggested by the term insula. A complete descrip-
olfactory proisocortex, and related to olfactory and tion of insular connections is given in the review by
autonomic functions (see Tanriover, N. et al., 2004 for Augustine J. R. (1996) showing that the insula is con-
a review). nected to the limbic areas, amygdalar nucleus, basal
ganglia, and all of the cortical lobes, except the
occipital lobe. Several attempts have been made to
47.1.2 The Insula as a Node in a Distributed
identify functional networks in which the insula
Cortical Network
could play the role of a functional node. Among
The insula is characterized by anatomical borders that these networks the perisylvianinsular, the temporo
are defined by a limiting sulcus (the circuminsular limbicinsular and the mesialorbitofrontalinsular
710 Human Insular Recording and Stimulation

networks (Mesulam, M. M. and Mufson, E. J., 1982) (see Rizzolatti, G. and Craighero, L., 2004 for a
deserve the attention of pain physiologists because review). The concept also refers to regions of the
they are involved in pain localization and intensity brain that are able to encode for a sensation (or emo-
encoding, as well as in emotional and behavioral reac- tion) perceived by the subject and to respond to the
tion to pain. observation of others experiencing that sensation (or
emotion). In the human insula, the regions involved in
visceral sensation or visceromotor responses are also
47.1.3 The Insula as a Polymodal Area responding to faces expressing disgust (Krolak-
The question of insular physiology has been addressed Salmon, P. et al., 2003). Similarly the human insula
by studies using neuroimaging, evoked potentials respond to both pain perception and empathy for
(EPs), and direct stimulation in humans as well micro- others pain ( Jackson, P. L. et al., 2005).
electrode studies in monkeys (Augustine, J. R., 1996).
These studies have confirmed that the insula is
involved in somatosensory and pain sensation as
assessed by numerous functional imaging studies in 47.2 Rational, Ethical Limitations and
humans (see Peyron, R. et al., 2002 for a review), as Procedure of Insular Recording
well as by the somatosensory and pain-evoked and Stimulations in Humans
response recordings and direct electric stimulations 47.2.1 Recording and Stimulation of
of the insular cortex that are detailed below in this the Human Insula Are Justified only in
chapter. However, the insula also proved to be a the Context of Presurgical Evaluation of
highly organized lobe with multiple specific functions Epilepsy
other than pain including in particular:
In humans, electrical stimulation of the cortex, as well
Visceral sensation, visceromotor control. as invasive intracranial recordings of the cerebral
Cardiovascular function as demonstrated by insular activity, including surface recordings using electrodes
stimulation in epileptic patients that produces changes placed on the cortex (electrocorticography) and depth
in heart rate or blood pressure in 50% of cases recordings using electrodes implanted directly in
(Oppenheimer, S. M. et al., 1992), thus leading to sus- the cortex (stereotactic electroencephalography or
pect a role of insular discharges in cardiac arrhythmias SEEG), aims at localizing the area that produces
causing sudden death during epileptic seizures. focal epileptic seizures. This area, referred to as the
Gustation as assessed first by the stimulation epileptogenic zone (EZ; Rosenow, F. and Luders, H.
studies of Penfield W. and Faulk M. E. (1955) and O., 2001), is that which is surgically removed to
further confirmed by neuroimaging studies and obtain seizure freedom in patients whose epilepsy
microelectrode recordings of insular neurons in mon- cannot be controlled by conventional drug treatment.
keys. These physiological findings are consistent Ethically such recordings are justified only when the
with the altered taste perception observed in patients EZ cannot be localized noninvasively either by scalp
with insular lesions. recordings or neuroimaging techniques. In this con-
Audition and language, in particular allocation of text, invasive explorations aim mostly at recording
auditory attention, tuning in to novel auditory sti- spontaneous seizures. They are also used to perform
muli, temporal, and phonological processing of a functional mapping of the cortex located in, or in
auditory stimuli. Furthermore several studies suggest the vicinity of, the EZ, in order to predict the func-
that both right and left insulas are involved in the tional consequences of the surgical treatment, which
control of speech production. consists of removing the EZ cortex. This procedure,
named cortectomy, is possible provided that the EZ is
not located in an eloquent area involved in essential
47.1.4 The Insula as Part of the
cortical functions such as language, memory, motor
Mirror-Neuron System
control, and vision. Both the recording of responses
Some recent studies suggest that the insular lobe could evoked by sensory stimulations and direct electrical
belong to the mirror-neuron system that characterizes stimulation of the cortex are commonly used to
regions of the brain that are able to respond when the achieve this functional mapping in epilepsy surgery
subject performs an action and when the subject centers around the world. This context obviously
observes another individual doing a similar action entails some limitations in the use of depth cortical
 Human Insular Recording and Stimulation 711

recordings and stimulation in studies of human brain 47.2.5 Only Data Recorded in
physiology. Nonepileptogenic Cortex are Physiologically
Relevant

47.2.2 Depth Intracortical Electrodes Are Data from cortical recordings or stimulations can be
Needed to Explore the Insula considered as reflecting the normal physiology of the
brain on the condition that they have been obtained
Due to its deep location in the lateral fissure the in cortical areas that are not involved in the epilepto-
insula can be explored either by electrocorticography genic process and do not show increased excitability
during surgery in patients whose frontoparietal or to peripheral inputs or local electric fields produced
temporal operculum has been removed by a previous by stimulation. Two criteria are commonly used to
cortectomy, as was the case in the few patients first check this, which are first the absence of insular
explored in Montreal by Penfield W. and Faulk M. E. epileptic discharge in the insula during spontaneous
(1955), or by depth SEEG electrodes implanted per- seizures and, second, the absence of sustained after-
pendicular to the mid-sagittal plane through the discharge after electrical stimulation of the insular
opercular cortex (Figure 1). These electrodes usually cortex.
have five to ten contacts, each of 2 mm in length
separated by 1.5 mm and can be left in place chroni-
cally up to 15 days (Isnard, J. et al., 2000; 2004). Only
the deepest contacts of such electrodes reach the 47.3 Insular Recordings of Pain
insular cortex while the superficial ones explore Evoked Responses in Humans
the frontoparietal or temporal opercular cortex.
Therefore, concerning somatosensory responsive In what follows we will describe responses to pain
areas located close to the lateral fissure, it must be and nonpainful cutaneous stimuli because the same
checked on individual brain magnetic resonance ima- contact in the insula most often detects both types of
ging (MRI) whether contacts are located in the responses. Moreover pain intensity rating is subjec-
suprasylvian external parietal cortex (area S1), in tive and corresponds to various levels of stimulus
the parietal operculum cortex (area S2), or in the intensity between subjects. In complement to insular
insula. Oblique electrodes trajectories can also been responses we will also describe responses recorded in
used to explore the insula, thus increasing the num- the second somatosensory (S2) area (Figure 1), which
ber of contacts located in the insular cortex. is located in the superior bank of the lateral fissure,
because it has a blurred anatomical border with the
granular insular cortex and is also involved in build-
47.2.3 The Number of Electrodes Is Limited ing up pain sensation.
by Ethical Issues
Only the minimal number of electrodes that are use- 47.3.1 The Laser Stimulus
ful to diagnosis can be implanted. Consequently the Most of the studies on pain-EPs in normal subjects
spatial resolution of the mapping in each individual is and patients use a laser beam (mostly CO2) applied
low, and pooling interindividual data is necessary to on the skin surface as a stimulus. The laser beam is
draw topographic functional maps of the human known to stimulate the endings of small diameters
insula based on depth electrode data. fibers and mostly those of A delta fibers. When the
power output is fixed, the amount of thermal energy
delivered depends on the duration of the pulse,
47.2.4 The Site of Electrode Implantation Is
which is in the order of a few milliseconds and,
Guided by Diagnostic Purposes
thus, permits an accurate timing for the analysis of
Implantation sites are guided by the hypothesis the electrophysiologic response. The energy density
issued from noninvasive investigations concerning of the laser beam is expressed in milli-Joules per mm2
the most probable EZ location. Furthermore the tra- of skin surface (mJ mm2). Threshold values for pain
jectory of electrode tracts is restricted by the show large interindividual variations between 30 and
anatomy of blood vessels that are particularly dense 40 mJ mm2 using a CO2 laser beam. In most studies,
in the lateral fissure. Therefore, some parts of the the sensation perceived by the subject is that of sharp
insula are rarely or never explored. pinprick, without poststimulus pain, considered as
712 Human Insular Recording and Stimulation

characteristic of the sensations produced by the sti-

Depth Coordinates Talairach (mm)

P230
mulation of A delta fibers. Although they are able to 31 Insula
rate it on a visual analog scale of pain (usually at 47 34
on a 10 level scale), the subjects do not identify this N180
38
sensation as a pain comparable to what they might P170 SII
have experienced in the past. Indeed no natural pain 41
is provoked by selective activation of the A delta 45
fibers, and this must be kept in mind when using
48
laser stimulation in pain studies. In particular, the N140
50 v
+
laser stimulus can be considered as adequate to assess 100 ms
the intensity coding of a pain stimulus, but is sub-
optimal for studying of the emotional reaction to
pain.
E
AC-PC AC-PC

47.3.2 Cortical Surface Recordings

(Electrocorticography)
Figure 2 Pain laser-evoked potentials (LEPs) to skin laser
Lenz F. A. et al. (1998) were the first to record cortical stimulation of the dorsum of the hand. The lower part of the
surface recordings by means of a subdural grid of figure shows frontal magnetic resonance imaging slices of
the right insula with projection of the contacts of a recording
electrodes placed on the surface of the parietal oper-
electrode exploring the insula (red) and S2 (blue). The
culum with CO2-laser-evoked potentials (LEPs) vertical dashed blue line represent the midline sagittal
peaking between 160 and 340 ms after the stimulus. plane, and the horizontal dashed blue line the anterior
The spatial distribution of this response over the commissureposterior commissure (ACPC) horizontal
cortical surface of the perisylvian cortex was consid- plane used for stereotactic implantation of depth
electrodes. Evoked potentials illustrated in the upper part of
ered as compatible with generators located in the
the figure are obtained by averaging responses to repeated
parietal operculum and/or in the insular cortices. stimulations of the left hand by a CO2 laser beam. Figures
However subdural electrodes placed over the sylvian along the vertical axis are the depth coordinates (x) in mm
area do not allow direct recording neither in the deep from midline of each recording contact. Note that distinct
aspect of the frontoparietal opercular cortex nor in responses are obtained in both areas, which are separated
by a delay of 50 ms.
the insula.

47.3.3 Intracortical S2 Recordings 47.3.4 Intracortical Insular Recordings

In humans, LEPs at stimulus intensities below and LEPs contralateral to stimulation recorded in the
above pain threshold are recorded in the suprasylvian insular cortex itself (Frot, M. and Mauguiere, F.,
opercular cortex corresponding to the human S2 area 2003) consist in a N180 negative response (mean
(Frot, M. et al., 2001). These responses are not picked latency: 180 16.5 ms) followed by a P230 positivity
up in the other areas most often explored in presur- (mean latency: 226 16 ms; Figure 2). As for S2,
gical assessment of epilepsy including the amygdala, insular LEPs ipsilateral to stimulation peak 1017 ms
hippocampus, and orbitofrontal cortex. They show a later than contralateral ones. LEP amplitudes increase
biphasic negativepositive waveform peaking at between sensory and pain threshold intensities but,
137 13 ms (N140) and 172 11 ms (P170), respec- contrary to what is observed in S2, continue to
tively, after stimulation of opposite hand (Figure 2). increase significantly at intensities over the pain
Similar responses are equally recorded in the homo- threshold (Figure 3).
logous cortex, ipsilateral to the painful stimulus with The reason for the delay of  50 ms (50 16 ms
a delay of 1017 ms. The LEP voltage in S2 shows a between S2 N140 and insular N180 peaking laten-
significant increase as soon as the stimulus intensity cies) observed between S2 and insular pain responses
reaches the sensory threshold as well as between remains questionable. It is too long for a monosynap-
sensory and pain thresholds while it rapidly reaches tic transmission from S2 to the insula, the two areas
a plateau for intensities above pain threshold (Frot, being interconnected through direct projections.
M. et al., 2007; Figure 3). Alternatively, knowing that both S2 and the insula
 Human Insular Recording and Stimulation 713

(a) SII Insula

50 13
25
12
40 11 20
30 10 15
20 10
10 5
V V
0 0
10 5
20 10
30 15
40 20
50 25
100 0 100 200 300 400 500 600 700 800 900 100 0 100 200 300 400 500 600 700 800 900
ms ms
(b) 90
SII
80 Insula
N_P peak amplitudes (V)

SE
70

60

50

40

30

20

10
I0 I1 I2 I3
Figure 3 Effects of stimulus intensity on laser-evoked potential (LEP) amplitude in S2 and insula. A) LEP traces in S2 and
insula to four different stimulus intensities are superimposed. CO2 Laser pulses were applied at four different intensities in
each subject. The power output being fixed, the amount of thermal energy delivered depends on the duration of the pulse.
Pulse duration is set up according to subjects subjective reports, rated on a visual analog scale (VAS) with an anchor point
corresponding to the pain threshold. The printed scales consist of 10-cm horizontal lines where the left extreme is labeled no
sensation and the right extreme maximal pain, and an anchored level 4 was at pain threshold (Lickert-type scale). The
different stimuli and related subjective sensation are as follows: I0: below sensory threshold (pulse duration: 515 ms, mean
energy density: 7 mJ mm2, no sensation); I1: above sensory threshold (pulse duration: 1545 ms, mean energy density:
19 mJ mm2, producing a detectable nonpainful sensation reported for more than 90% of stimulations; For one-third of
patients this sensation is a warmth sensation and for the others two-thirds a slight nonpainful pinprick sensation; VAS:
1.6 1.09); I2: pain threshold (pulse duration: 2580 ms, mean energy density: 33 mJ mm2, producing a pricking sensation,
like a hair pulling or a drop of hot boiling water on the skin; VAS: 3.9 1.46); I3: 20% above pain threshold (pulse duration: 35
110 ms, mean energy density: 46 mJ mm2, producing a pricking sensation described as clearly painful; VAS: 5.4 1.6). In
S2, a significant increase of the LEP is observed as soon as the stimulus intensity reaches the sensory threshold (between I0
and I1, P < 0.001), as well as between sensory and pain thresholds (I1 to I2, P < 0.001) while amplitudes rapidly reach a plateau
for intensities above pain threshold (no significant amplitude difference between I2 and I3). In the insula, no significant
amplitude increase is observed for low-stimulation intensities between I0 and I1, LEP amplitudes also increase between
sensory and pain threshold intensities (P < 0.05 between I1 and I2) and continue to increase significantly at higher intensities
over pain threshold (P < 0.001 between I2 and I3). B) Insula and S2 LEPs peaking amplitudes as a function of stimulus
intensity. In these polynomial regression curves of the stimulusresponse functions S2 LEPs amplitudes show a S-shaped
profile with increasing stimulus intensities that reaches a plateau above pain threshold, while insular LEP amplitudes show an
exponential profile. Laser stimuli above pain threshold trigger a maximal response in S2 and a pain level-related response in
the insula (Modified from Frot, M., Magnin, M., Mauguiere, F., and Garcia-Larrea, L. 2007. Human SII and posterior insula
differently encode thermal laser stimuli. Cereb. Cortex 17, 610620).
714 Human Insular Recording and Stimulation

receive direct projections from the thalamus (see for studies (e.g., 15 ms between primary visual areas); it
a review Augustine, J. R., 1996) the explanation could is in the same range as that measured between ipsi-
be that the latter are triggered via thalamocortical and contralateral S2 magnetic fields evoked by elec-
fibers with a slower conduction than that of thalamic trical stimulation of the median nerve. The
projections to the S2 area. To our knowledge, how- possibility remains, however, that responses ipsilat-
ever, no electrophysiological demonstration of this eral to the stimulus could be triggered via ipsilateral
hypothesis is hitherto available. A third hypothesis thalamic fibers with slower conduction velocities.
could be that the suprasylvian cortex and the insula Only intracortical recordings of S2 or insular EPs to
are activated by inputs conveyed by peripheral fibers ipsilateral stimuli in patients with a lesion of the
with different conduction velocities. Some studies homologous areas in the opposite hemisphere could
have estimated the A delta conduction velocity in a address this question directly.
large range of 720 m s1 suggesting the existence of
different A delta fiber subpopulations, with different
conduction velocities. One can hypothesize that 47.4 Insular Stimulation
these different subpopulations of peripheral fibers
47.4.1 The Challenge of Insular
could project in distinct cortical regions. However,
Stimulation
to our knowledge, no electrophysiological study has
been devoted to the identification of separate subpo- In humans, it has long been a challenge to stimulate the
pulations of fibers with different conduction insular cortex. Thus, only a few studies have reported
velocities in the spinothalamic tract or thalamocorti- nonnociceptive somesthetic symptoms, cardiovascular
cal projections. effects as well as visceromotor and viscerosensitive
sensations consecutive to direct electrical stimulation
of the insular cortex. Truly painful responses have
47.3.5 Topographic Specificity of Insular
not been reported during stimulation of any area of
Laser-Evoked Potentials
the cerebral cortex in humans by Penfield W. and
Due to the anatomical proximity between the S2 area Faulk M. E. (1955), who extensively stimulated the
and the granular part of the insula the question arises surface of all cortical areas of the human brain, includ-
whether insular responses might reflect the diffusion ing the insula, using surface electrodes. However, they
of S2 LEPs with a polarity reversal across the sylvian did not explore the posterior part of the insula where
fissure, which is almost virtual in that region. The electric stimulation evokes painful sensations.
fact that when two contacts, or more, explore the Electric stimulations are produced by a current-
insular cortex there is an amplitude increase from regulated neurostimulator designed for a safe diagnostic
surface to depth of the N180P230 response suggests stimulation of the human brain. Square pulses of current
that this is not so (see Frot, M. and Mauguiere, F. are applied between two adjacent contacts (bipolar sti-
2003 for a complete discussion). However, the abso- mulation) at a frequency of 50 Hz, pulse duration of
lute proof that the N180P230 is generated in the 0.5 ms, train duration of 5 s, and intensity of 0.8
insular cortex would be its polarity reversal between 6.0 mA. These parameters are used to avoid any tissue
the surface and the depth of the insular gray matter; injury (charge density per square pulse <55 mC cm2;
unfortunately usual stereotactic recordings in Gordon, B. et al., 1990). This stimulation paradigm
patients do not have enough spatial resolution ensures a good spatial specificity with respect to the
(2 mm interval between two neighboring contacts of desired structures to be stimulated. The study of current
1.5 mm) to assess the distribution of potentials per- densities in the cortex for bipolar stimulation with a 10-
pendicular to the cortical surface. mA stimulating current shows that the peak current
density occurs in the region immediately beneath the
bipolar electrodes (0.05 A cm2) and declines rapidly to
47.3.6 Interhemispheric Transmission of
0.02 A cm2 0.5 cm away, and that the current density
S2 and Insular Laser-Evoked Potentials
decreases in relation to the square of the distance into
Insular and S2 responses to pain are bilateral; ipsilat- the cortex. In S1, S2, and the insula, the minimal stimu-
eral potentials peaking with a delay of 1020 ms after lation intensity necessary to elicit a clinical response is
contralateral ones. This delay is compatible with the 1.61.9 mA (Mazzola, L. et al., 2006). With these stimuli
interhemispheric transmission time through fibers of there is virtually no current spread out of the stimula-
the corpus callosum as estimated by numerous tion current dipole as defined by the distance of 5.5 mm
 Human Insular Recording and Stimulation 715

between the outer and inner limits of the superficial and

pg
deep-stimulating contacts, respectively (Ostrowsky, K. pcg
ps
et al., 2002). Moreover it has been demonstrated that no F2 pcs cs
significant cerebral blood flow change occurs at the site
f2 sps
of cortical stimulation, or in its close vicinity, in the
absence of epileptic after discharge. aps

ips T2
47.4.2 Pain Evoked by Insular Stimulation
Painful sensations in response to direct electric sti-
mulation of the insula have been reported by Figure 4 Pain insular area. The locations of stimulation
Ostrowsky K. et al. (2000; 2002) and more recently sites in a population of patients are plotted on a sagittal view
by Mazzola L. et al. (2006). Qualities of the evoked of the insula exposed by removal of the supra and
pain are described as burning, stinging, disabling infrasylvian cortex according to the type of somatosensory
sensation reported by the subjects. Red dots: pain, Orange
sensation, or electrical shock. Pain intensity varies dots: nonpainful warm sensation, yellow dots: nonpainful
from mild to intolerable but is not related to stimula- paresthesias. Note that most of pain sensations are evoked
tion intensity. Pain disappears as soon as the by stimulation the posterior part of the insula situated
stimulation is interrupted in most cases except a behind the tip of the central sulcus (cs), while nonpainful
few where an intense pain is followed by a sore responses are obtained over a larger and more anterior
surface of the insular lobe. Aps, anterior peri-insular sulcus;
feeling that can last up to 1 min after the end of the
cs, central sulcus; F2, second frontal gyrus; f2, second
stimulation. It is located contralateral to the stimula- frontal sulcus; ips, inferior peri-insular sulcus; pcg,
tion site or bilaterally when midline parts of the body precentral gyrus; pg, postcentral gyrus; ps, parietal sulcus;
are involved, and affects large areas of the body (e.g., sps, superior peri-insular sulcus; T2, second temporal
face, upper limb, whole half of the body), suggesting gyrus.
that receptive fields for pain in the insular cortex are
more extended that somatosensory fields in the SI
and SII cortex (Mazzola, L. et al., 2006).
References

47.4.3 Anatomical Location of the Insular Augustine, J. R. 1996. Circuitry and functional aspects of the
insular lobe in primates including humans. Brain Res. Rev.
Pain Area 22, 229244.
Clark, T. E. 1896. The comparative anatomy of the insula.
The area where pain can be elicited by direct electric J. Comp. Neurol. 6, 59100.
stimulation occupies the posterosuperior part of the Frot, M. and Mauguiere, F. 2003. Dual representation of pain in
insular lobe (Figure 4). Surface electrodes cannot the operculo-insular cortex in humans. Brain 126, 438450.
Frot, M., Garcia-Larrea, L., Guenot, M., and Mauguiere, F. 2001.
reach this area unless the lower precentral motor Responses of the supra-sylvian (SII) cortex in humans to
and postcentral somatosensory cortex be removed. painful and innocuous stimuli. A study using intra-cerebral
This is the most probable reason why early electro- recordings. Pain 94, 6573.
Frot, M., Magnin, M., Mauguiere, F., and Garcia-Larrea, L. 2007.
corticographic studies using that type of electrodes Human SII and posterior insula differently encode thermal
failed to produce pain by cortical stimulation in laser stimuli. Cereb. Cortex 17, 610620.
humans. Because only a few contacts can be placed Gordon, B., Lesser, R. P., Rance, N. E., Hart, J., Jr., Webber, R.,
Uematsu, S., and Fisher, R. S. 1990. Parameters for direct
in the insula of a given individual during stereotactic cortical electrical stimulation in the human: histopathologic
exploration of the brain with depth electrodes it is confirmation. Electroencephalogr. Clin. Neurophysiol.
uncertain whether a pain somatopic representation of 75, 371377.
Isnard, J., Guenot, M., Ostrowsky, K., Sindou, M., and
the body, comparable to that demonstrated in the Mauguiere, F. 2000. The role of the insular cortex in temporal
primary somatosensory cortex (S1), does exist in the lobe epilepsy. Ann. Neurol. 48, 614623.
pain insular area. Moreover due to the large skin Isnard, J., Guenot, M., Sindou, M., and Mauguiere, F. 2004.
Clinical manifestations of insular lobe epileptic discharges: a
extent of pains evoked by insular stimulation that stereo-electroencephalographic study. Epilepsia
the somatotopic pain representation is likely to be 45, 10791790.
much more fuzzy, with overlapping receptive fields, Jackson, P. L., Meltzoff, A. N., and Decety, J. 2005. How do
we perceive the pain of the others? A window into the
in the insular pain area than in S1 or S2 somatosen- neural processes involved in empathy. Neuroimage
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Krolak-Salmon, P., Henaff, M. A., Isnard, J., Tallon-Baudry, C., sensation in the human insula: a study of responses to direct
Guenot, M., Vighetto, A., Bertrand, O., and Mauguiere, F. electrical cortical stimulation. Cereb. Cortex 12, 376385.
2003. An attention modulated response to disgust in human Penfield, W. and Faulk, M. E. 1955. The insula. Further
ventral anterior insula. Ann. Neurol. 53, 446453. observations on its function. Brain 78, 445470.
Lenz, F. A., Rios, M., Chau, D., Krauss, G. L., Zirh, T. A., and Peyron, R., Frot, M., Schneider, F., Garcia-Larrea, L.,
Lesser, R. P. 1998. Painful stimuli evoke potentials recorded Mertens, P., Barral, F. G., Sindou, M., Laurent, B., and
from the parasylvian cortex in humans. J. Neurophysiol. Mauguiere, F. 2002. Role of operculo-insular cortices in
80, 20772088. human pain processing. Converging evidence from PET,
Mazzola, L., Isnard, J., and Mauguiere, F. 2006. Somatosensory fMRI, dipole modeling and intra-cerebral recordings of
and pain responses to stimulation of the second evoked potentials. Neuroimage 17, 13361346.
somatosensory area (SII) in humans. A comparison with SI Rizzolatti, G. and Craighero, L. 2004. The mirror-neuron system.
and insular responses. Cereb. Cortex 16, 960968. Ann. Rev. Neurosci. 27, 169192.
Mesulam, M. M. and Mufson, E. J. 1982. Insula of the old world Rosenow, F. and Luders, H. O. 2001. Presurgical evaluation of
monkey. I. Architectonics in the insulo orbito-temporal epilepsy. Brain 124, 16831700.
component of the paralimbic brain. J. Comp. Neurol. Tanriover, N., Rhoton, A. L., Kawashima, M., Ulm, A. J., and
212, 122. Yasuda, A. 2004. Microsurgical anatomy of the insula and
Oppenheimer, S. M., Gelb, A., Girvin, J. P., and Hachinski, V. C. the sylvian fissure. J. Neurosurg. 100, 891922.
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Ostrowsky, K., Isnard, J., Ryvlin, P., Guenot, M., Fischer, C.,
and Mauguiere, F. 2000. Functional mapping of the insular Relevant Websites
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Ostrowsky, K., Magnin, M., Ryvlin, P., Isnard, J., Guenot, M., http://imaging.mrc-cbu.cam.ac.uk CBU Imaging
and Mauguiere, F. 2002. Representation of pain and somatic http://en.wikipedia.org
 48 The Rostral Agranular Insular Cortex
L Jasmin, Neurosurgery and Gene Therapeutics Research Institute, Los Angeles, CA, USA
P T Ohara, University of California, San Francisco, CA, USA
2009 Elsevier Inc. All rights reserved.

48.1 What Is the Rostral Agranular Insular Cortex? 717

48.2 Function of the Rostral Agranular Insular Cortex 717
48.3 Anatomical Location 717
48.4 Neurochemical Signature 718
48.5 Connections of the Rostral Agranular Insular Cortex 718
48.6 Behavioral Studies 719
48.7 Relationship with Adjacent Cortical Regions 719
48.8 Overview in the Rat 720
48.9 From Rats to Humans 720
48.10 Perspective 721
References 721

Glossary
amygdala A group of neurons that is part of the have been shown to be involved in different
limbic system. The amygdala is involved in many aspects of processing painful information.
aspects of behavior such as memory, emotion, and nociception The perception of a stimulus that is
motivation. potentially injurious to ones body.
immunocytochemical staining A method that nucleus accumbens A part of the limbic system
uses antibodies to identify and locate specific cel- involved in reward and addiction.
lular components, such as neurotransmitters. thalamus A part of the brain located at the center
medial frontal, infralimbic, and cingulate that processes sensory and motor information and
cortices Different parts of the cerebral cortex that sends the resulting information to the cerebral cortex.

48.1 What Is the Rostral Agranular 48.2 Function of the Rostral

Insular Cortex? Agranular Insular Cortex

The rostral agranular insular cortex (RAIC) is a small Many parts of the cerebral cortex do not have pre-
area of the cerebral cortex located on the lateral cisely defined functions and usually do not have
aspect of the rats cerebral hemisphere. The RAIC precisely defined borders. Until recently, the RAIC
can be distinguished in terms of its location, cellular was one such region, but a picture indicating that the
architecture, neurochemistry, and connectivity. RAIC is involved in the behavioral responses to
Functionally, the RAIC is involved in processing nociceptive (painful) stimuli is beginning to emerge
pain information and, depending on which cells are from animal and human studies.
activated, can act through connections with other
brain areas to reduce or increase pain responses. 48.3 Anatomical Location
The RAIC therefore is one of a limited number of
cortical regions involved in the perception and In the rat, the RAIC occupies a small area of the
response to pain. lateral aspect of cerebral cortex just above a groove,

717
718 The Rostral Agranular Insular Cortex

concentration of neurons in layer 5 of the RAIC that

SI express the GABAB receptor (Jasmin, L. et al., 2003),
which is one of several types of receptors activated by
SII the inhibitory neurotransmitter, gamma-aminobutyric
OB acid (GABA). Similar concentrations of GABAB
receptor-bearing neurons are also found in the insular
cortex posterior to the RAIC and in a few other brain
RF RAIC AI regions but are less concentrated in the remainder of
the cortex. The neurons in layer 5 of the RAIC that
Primary olfactory cortex
express the GABAB receptor are large pyramidal pro-
Figure 1 Lateral view of the rat brain. The rhinal fissure jection neurons that have axons that leave the cortex
(RF) is seen as a prominent groove starting near the ventral
and innervate other brain regions. This finding sug-
anterior cortex and extending toward the posterior cortex.
The agranular insular cortex is a thin strip of cortex lying just gests that the neurons in the RAIC that have GABAB
above (dorsal to) the RF and the rostral part (RAIC) is receptors play a specific role in how the RAIC com-
indicated in red. The approximate location of the primary municates with other brain areas. A second notable
somatosensory cortex (SI) and the secondary feature of the RAIC is the concentration of nerve
somatosensory cortex (SII) are indicated for reference. OB,
fibers containing the neurotransmitter dopamine. In
olfactory bulb.
the rat, there is only sparse dopamine innervation
throughout most of the cortex dopamine fibers that
which is one of the few surface features of the rat are found in high concentration in then infralimbic
cortex, called the rhinal fissure (Figure 1) (Paxinos, cortex and in the RAIC (Figure 2). These dopamine
G., 1995). Agranular cortex is defined by the absence fibers originate in a region of the brainstem called the
of a layer 4 that gives the cortex a less granular ventral tegmental area (VTA) and probably are
appearance than the adjacent areas. The region was involved in determining the overall level of activity
described by Krettek J. E. and Price J. L. (1977) as a of the RAIC rather than conveying specific sensory
strip of agranular cortex located immediately above information.
the rhinal fissure and bounded dorsally by granular
cortex. Rostrally, the agranular cortex was further
subdivided into a ventral and a dorsal agranular 48.5 Connections of the Rostral
region. This description was later modified Agranular Insular Cortex
(Cechetto, D. F. and Saper, C. B., 1987) to include a
dysgranular field between the agranular and granular The specific connections of a cortical region also
cortices. The dysgranular field had a layer 4 that was help define the boundaries and give insight into the
intermediate in prominence between the fully devel- functions of the region. The RAIC shares the general
oped layer 4 of the granular cortex and the absent pattern of connection common to most regions of
layer 4 of the agranular cortex. Later studies all cortex receiving its major subcortical afferents from
distinguish agranular, dysgranular, and granular the thalamus, having reciprocal connection with
fields although the precise boundaries, particularly other cortical regions, and projecting to a number of
between the dorsal agranular and dysgranular fields, subcortical sites ( Jasmin, L. et al., 2004). The picture
were delineated differently. that emerges from an analysis of the projections is
that the RAIC plays a role in pain mechanisms. The
inputs from the thalamus are from the nuclei that
48.4 Neurochemical Signature convey sensory information from the body. However,
this input is not from thalamic nuclei that subserve
Although classically cortical regions are defined by the fine-detailed discriminative aspect of sensation
cytoarchitecture and location, modern neuroanato- (i.e., those that project to the primary somatosensory
mical techniques have refined our understanding of cortex), rather the input is from nuclei located in the
cortical organization based on connections and neuro- midline of the thalamus that are generally associated
chemical profile. The RAIC has been shown to be with emotional or affective aspects of pain. The
distinguished from surrounding regions by a distinct principal cortical connection of the RAIC is with
pattern of immunocytochemical staining (Figure 2) areas such as the medial frontal, infralimbic, and
(Ohara, P. T. et al., 2003). The first feature is a cingulate cortices, which have long been known to
 The Rostral Agranular Insular Cortex 719

(a) (b) (c) (d)

IL IL

VLO BG

Figure 2 Coronal sections through the left anterior rat brain showing some of the characteristic features of the RAIC. (a) A
section stained with a Nissl stain to show the general appearance and location of the RAIC. The rhinal fissure is indicated by
the arrow and the asterisk indicates the location of the RAIC. (b, c) Sections stained immunocytochemically to show the
location of fibers containing the neurotransmitter, dopamine. The contrast has been digitally enhanced to show the dopamine
more clearly. The arrow indicates the region of dopamine staining that corresponds to the location of the RAIC. Intense
staining is also present in the infralimbic cortex (IL), which is also a pain-related area, and in the basal ganglia (BG), which is a
motor-related site. The ventrolateral orbital cortex (VLO), a site also associated with pain behavior, shows no dopamine
staining. (d) A higher magnification of the boxed region indicated in (c). This section is stained to show the location of neurons
that express the GABAB receptor (green-colored structures) in cortical layer 5. The arrowhead indicated the location of the
rhinal fissure.

be associated with pain responses. The RAIC projects possess GABAA receptors occur through projections
to a number of subcortical sites including the lateral to the brainstem and activation of descending pain
hypothalamus, dorsal raphe, periaqueductal gray mat- inhibitory systems. However, as noted above, the
ter, pericerulear region, rostroventral medulla, and RAIC is characterized by an abundance of cells that
parabrachial nuclei all of which play a role in pain have GABAB receptors. The changes in pain beha-
inhibition mechanisms. Significant to the probable vior mediated by the GABAB receptor-bearing
function of the RAIC are projections to the amygdala neurons point to projections to the amygdala playing
and the nucleus accumbens. These projections include a key role. The proposal that the activity in the RAIC
a large component originating in the cells in layer 5 leads to increased pain was tested by inhibiting
that express the GABAB receptor described above. GABAA receptor-bearing neurons and at the same
The amygdala is known to be involved in several time activating GABAB receptor-bearing neurons.
aspects of behavior including fear, anxiety, and atten- This experiment resulted in an increase in pain beha-
tion (Williams, M. A. et al., 2005). vior ( Jasmin, L. et al., 2003). These studies highlight
two important issues. First, changes in activity of the
RAIC can result in both increases and decreases in
48.6 Behavioral Studies pain response and the overall effect of the RAIC
results from a balance of pro- and antinociceptive
In addition to the connectional studies, there are a activity. Second, the RAIC probably changes pain
number of behavioral studies that give insight into behavior both by activating descending inhibitory
the function of the RAIC. Direct injections of com- circuits and by changing activity in the amygdala
pounds into the RAIC have shown that morphine and other cortical area to change the emotional/
injection, increasing dopamine levels, and increasing affective response to painful stimuli.
GABA levels all result in behavioral antinociception.
Because increasing GABA in the RAIC results in
antinociception and the action of GABA is to inhibit 48.7 Relationship with Adjacent
neuronal activity, it suggests that activity in the Cortical Regions
RAIC is pronociceptive. Additional pharmacological
experiments suggest that the neurons that express the The location and small size of the RAIC makes it
GABAA and those that express the GABAB receptor difficult to unequivocally isolate it from surrounding
change pain thresholds through different pathways. regions. The agranular insular cortex has been most
These antinociceptive effects mediated by cells that associated with visceral and gustatory responses, and
720 The Rostral Agranular Insular Cortex

a number of studies on cells in the region have The advent of high-resolution functional imaging has
indicated that they respond to visceral input. These resulted in the availability of data relating to cortical
functions, however, seem more related to the more function that has previously been difficult to obtain from
posterior regions of the agranular cortex or the dys- animal studies (Figure 3). Specifically, it is possible to
granular cortex immediately above the RAIC. relate verbal descriptions and emotional responses to
Immediately anterior to the RAIC is the ventrolat- painful stimuli to specific cortical sites. In humans, ima-
eral orbital cortex (VLO). This region of cortex in the ging techniques and direct recording of field potentials
rat is also associated with pain responses (Xie, Y. F. show that painful stimuli activate the insular cortex
et al., 2004). As with all parts of the cerebral cortex, it (Coghill, R. C. et al., 1994; Brooks, J. C. et al., 2005). It
is often difficult to define a clear boundary between has been possible to distinguish between activation of the
any two regions. For this, and other technical reasons anterior and of the posterior region of the agranular
such as diffusion of drugs when injected into a spe- cortex located in the insular region (Strigo, I. A. et al.,
cific site, it is possible that some studies previously 2003). The anterior insula, which might be considered
carried out on the VLO may have encroached upon most equivalent to the RAIC, is activated when a strong
the RAIC and vice versa. There are both similarities emotional response is associated with the nociceptive
and differences between the VLO and the RAIC. stimulus. The posterior insula and the adjacent parietal
Morphine injected into the VLO and RAIC both cortex, in turn, are activated by all nociceptive stimuli,
produce antinociception, but the application of independently of their emotional content. These differ-
drugs that inhibit GABA has no effect in the RAIC ences in function between the anterior and the posterior
but do have antinociceptive effects in the VLO. insula are explained by their neural connectivity.
There is less dopamine innervation of VLO, and Notably, ascending nociceptive afferents from the spinal
although the VLO contains GABAB receptor-bearing
cord and brainstem directly activate the posterior insula
cells, they are not as numerous as in the RAIC
(Ohara, P. T. et al., 2003). Whether the VLO and
RAIC are really separate entities with specific, sepa-
rate functions related to pain or whether both regions
are parts of a larger forebrain nociceptive system is
not clear.

48.8 Overview in the Rat

The major connections of the RAIC are with areas

that have established roles in nociceptive information
processing and behavior responses to nociceptive
stimuli. RAIC projections to other cortical areas and
subcortical sites such as the amygdala are likely to
participate in the sensorimotor integration of noci-
ceptive processing, while the hypothalamus and
brainstem projections are most likely to contribute
to descending pain inhibitory control.
Figure 3 A functional magnetic resonance imaging blood
oxygen level-dependent (fMRI BOLD) image averaged from
48.9 From Rats to Humans nine normal healthy subjects who received a painful
stimulus to the back. Note that the insular cortex is activated
(arrows, red and yellow areas) on both sides of the cortex
The agranular insular cortex is found in other mam- although more prominently on the left side. This bilateral
mals including cats, monkeys, and primates including activation of the insular region can be contrasted with the
humans (Augustine, J. R., 1996; Mesulam, M.-M., 2000). unilateral activation of other cortical regions such as the
cingulate cortex (arrowhead) and dorsolateral prefrontal
In primates, the divisions of the insular cortex are the
cortex (double arrowhead). Why the activation is lateralized
same as in rats and the RAIC occupies an area imme- in these latter areas in not known but is a common
diately noticeable, that is, dorsal to the primary occurrence. Photo courtesy of Dr. Vania Apkarian,
olfactory cortex. Northwestern University.
 The Rostral Agranular Insular Cortex 721

only. Accordingly, direct electrical stimulation of the Brooks, J. C., Zambreanu, L., Godinez, A., Craig, A. D., and
Tracey, I. 2005. Somatotopic organisation of the human
posterior insula results in the generation of thermal insula to painful heat studied with high resolution functional
sensations (Ostrowsky, K. et al., 2002), and lesions of the imaging. Neuroimage 27, 201209.
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viscerotopic sensory representation in the cortex and
nition of nociceptive heat (Greenspan, J. D. et al., 1999). thalamus in the rat. J. Comp. Neurol. 262, 2745.
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terms of cytoarchitecture and function, it is not clear Williams, M. A., McGlone, F., Abbott, D. F., and Mattingley, J. B.
2005. Differential amygdala responses to happy and fearful
that they are strictly equivalent areas and our under- facial expressions depend on selective attention.
standing of pain mechanisms in general will have to Neuroimage 24, 417425.
take into account such species-specific similarities Xie, Y. F., Wang, J., Huo, F. Q., Jia, H., and Tang, J. S. 2004. Mu
but not delta and kappa opioid receptor involvement in
and/or differences. ventrolateral orbital cortex opioid-evoked antinociception in
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References
Further Reading
Augustine, J. R. 1996. Circuitry and functional aspects of the
insular lobe in primates including humans. Brain Res. Brain Allen, G. V., Saper, C. B., Hurley, K. M., and Cechetto, D. F.
Res. Rev. 22, 229244. 1991. Organization of visceral and limbic connections in the
Banzett, R. B., Mulnier, H. E., Murphy, K., Rosen, S. D., insular cortex of the rat. J. Comp. Neurol. 311, 116.
Wise, R. J., and Adams, L. 2000. Breathlessness in humans Altier, N. and Stewart, J. 1998. Dopamine receptor antagonists
activates insular cortex. Neuroreport 11, 21172120. in the nucleus accumbens attenuate analgesia induced by
722 The Rostral Agranular Insular Cortex

ventral tegmental area substance P or morphine and by Li, Y., Li, J. J., and Yu, L. C. 2002. Anti-nociceptive effect of
nucleus accumbens amphetamine. J. Pharmacol. Exp. Ther. neuropeptide Y in the nucleus accumbens of rats: an
285, 208215. involvement of opioid receptors in the effect. Brain Res.
Altier, N. and Stewart, J. 1999. The role of dopamine in the 940, 6978.
nucleus accumbens in analgesia. Life Sci. Li, N., Lundeberg, T., and Yu, L. C. 2001. Involvement of CGRP
65, 22692287. and CGRP1 receptor in nociception in the nucleus
Clasca, F., Llamas, A., and Reinoso-Suarez, F. 1997. Insular accumbens of rats. Brain Res. 901, 161166.
cortex and neighboring fields in the cat: a redefinition based Margeta-Mitrovic, M., Mitrovic, I., Riley, R. C., Jan, L. Y., and
on cortical microarchitecture and connections with the Basbaum, A. I. 1999. Immunohistochemical localization of
thalamus. J. Comp. Neurol. 384, 456482. GABA(B) receptors in the rat central nervous system. J.
Clasca, F., Llamas, A., and Reinoso-Suarez, F. 2000. Cortical Comp. Neurol. 405, 299321.
connections of the insular and adjacent parieto-temporal Meyer, S., Strittmatter, M., Fischer, C., Georg, T., and
fields in the cat. Cereb.. Cortex 10, 371399. Schmitz, B. 2004. Lateralization in autonomic dysfunction in
Dupont, S., Bouilleret, V., Hasboun, D., Semah, F., and ischemic stroke involving the insular cortex. Neuroreport
Baulac, M. 2003. Functional anatomy of the insula: new 15, 357361.
insights from imaging. Surg. Radiol. Anat. 25, 113119. Ray, J. P. and Price, J. L. 1992. The organization of the
Gear, R. W., Aley, K. O., and Levine, J. D. 1999. Pain-induced thalamocortical connections of the mediodorsal thalamic
analgesia mediated by mesolimbic reward circuits. J. nucleus in the rat, related to the ventral forebrain-prefrontal
Neurosci. 19, 71757181. cortex topography. J. Comp. Neurol. 323, 167197.
van Groen, T., Kadish, I., and Wyss, J. M. 1999. Efferent Schmidt, B. L., Tambeli, C. H., Barletta, J., Luo, L., Green, P.,
connections of the anteromedial nucleus of the thalamus of Levine, J. D., and Gear, R. W. 2002. Altered nucleus
the rat. Brain Res. Brain Res. Rev. 30, 126. accumbens circuitry mediates pain-induced antinociception
Hof, P. R., Mufson, E. J., and Morrison, J. H. 1995. Human in morphine-tolerant rats. J. Neurosci. 22, 67736780.
orbitofrontal cortex: cytoarchitecture and quantitative Schmidt, B. L., Tambeli, C. H., Gear, R. W., and Levine, J. D.
immunohistochemical parcellation. J. Comp. Neurol. 2001. Nicotine withdrawal hyperalgesia and opioid-
359, 4868. mediated analgesia depend on nicotine receptors in nucleus
Kawaguchi, Y. and Kondo, S. 2002. Parvalbumin, somatostatin accumbens. Neuroscience 106, 129136.
and cholecystokinin as chemical markers for specific Sengupta, J. N., Snider, A., Su, X., and Gebhart, G. F. 1999.
GABAergic interneuron types in the rat frontal cortex. J. Effects of kappa opioids in the inflamed rat colon. Pain
Neurocytol. 31, 277287. 79, 175185.
Kawaguchi, Y. and Kubota, Y. 1998. Neurochemical features Suhara, T., Yasuno, F., Sudo, Y., Yamamoto, M., Inoue, M.,
and synaptic connections of large physiologically-identified Okubo, Y., and Suzuki, K. 2001. Dopamine D2 receptors in
GABAergic cells in the rat frontal cortex. Neuroscience the insular cortex and the personality trait of novelty seeking.
85, 677701. Neuroimage 13, 891895.
Kawaguchi, Y. and Shindou, T. 1998. Noradrenergic excitation Tambeli, C. H., Parada, C. A., Levine, J. D., and Gear, R. W.
and inhibition of GABAergic cell types in rat frontal cortex. J. 2002. Inhibition of tonic spinal glutamatergic activity induces
Neurosci. 18, 69636976. antinociception in the rat. Eur. J. Neurosci. 16, 15471553.
 49 Descending Control Mechanisms
K Ren and R Dubner, University of Maryland, Baltimore, MD, USA
2009 Elsevier Inc. All rights reserved.

49.1 Introduction 724

49.2 Organization of Descending Pathways 725
49.2.1 Key Structures and Pathways 725
49.2.1.1 Midbrain PAG matter 725
49.2.1.2 PAGRVMspinal dorsal horn circuitry 726
49.2.1.3 Dorsolateral pontomesencephalic tegmentum 727
49.2.1.4 Caudal medulla 728
49.2.1.5 Forebrain sites 729
49.2.1.6 Summary 730
49.2.2 Neurotransmitters 730
49.2.2.1 Endogenous opioids and related peptides 730
49.2.2.2 Monoamines 732
49.2.2.3 Amino acids 734
49.2.2.4 Others 735
49.2.2.5 Summary 737
49.2.3 Neuronal Activity in Descending Pathways 738
49.2.3.1 Pain modulatory neurons 738
5.49.2.3.2 Descending influence on dorsal horn neurons 739
49.2.4 Multiplicity of Descending Control 740
49.3 Plasticity of Descending Pathways 740
49.3.1 Descending Modulation in Persistent Pain 741
49.3.1.1 Responses of descending pathways to injury 741
49.3.1.2 Descending inhibition in persistent pain 741
49.3.1.3 Descending facilitation in persistent pain 742
49.3.1.4 Descending facilitation versus inhibition 743
49.3.2 Dynamics of Descending Pain Modulation 743
49.3.2.1 Time-dependent shift in descending control 743
49.3.2.2 Phenotypic changes in pain modulatory neurons 745
49.3.2.3 Involvement of excitatory amino acids and receptors 746
49.3.2.4 Molecular mechanisms of activity-dependent plasticity in descending pathways 746
49.3.2.5 Changes in sensitivity to opioids 747
49.3.2.6 Aging-related alteration in descending pain modulation 747
49.3.3 Significance of Altered Descending Modulation in Persistent Pain 747
49.4 Therapeutic Implications: Activating Descending Pathways 748
49.4.1 Electrical Stimulation 748
49.4.1.1 Brain stimulation 748
49.4.1.2 Spinal cord stimulation 748
49.4.1.3 Vagal stimulation 748
49.4.1.4 Transcutaneous electrical nerve stimulation 748
49.4.2 Pharmacological Approaches 748
49.4.2.1 Opioids 748
49.4.2.2 Alpha 2 adrenoceptor agonists 749
49.4.3 Peripheral Stimulation 749
49.4.3.1 Diffuse noxious inhibitory controls 749
49.4.3.2 Acupuncture 749
49.4.4 Psychological Factors 749

723
724 Descending Control Mechanisms

49.4.4.1 Stress 749

49.4.4.2 Placebo 749
49.5 Concluding Remarks 749
References 749

49.1 Introduction Our knowledge of the descending pain modula-

tory systems spans at least four decades. In 1965,
Pain is a sensory process that reflects the level and Wall and Melzack in their gate control theory of
intensity of noxious stimulus on one hand but is also pain questioned the concept of an invariant rela-
closely controlled or modulated by the central ner- tionship between stimulus and sensation and a
vous system (CNS). In fact, the intensity of the fixed, direct-line connection from the skin to the
perceived pain is not necessarily proportional to the brain. They proposed that central signals arising
amount of stimulus under different conditions from the brain could influence afferent input at the
(Figure 1). Although psychological factors contribute earliest synaptic levels of the somesthetic system,
to the variability of pain, the most important under- the spinal dorsal horn (Melzack, R. and Wall, P. D.,
lying mechanisms responsible for the variant pains 1965). Evidence supporting the concept was avail-
are the existence of the endogenous pain modulatory able even earlier when Hagbarth and colleagues
systems, in which brainstem descending pathways (Hagbarth K. E. and Kerr, D. I., 1954; Hernandez-
play a fundamental role (Fields, H. L. and Basbaum, Peon, R. and Hagbarth, K. E., 1955) showed that
A. I., 1999; Millan, M. J., 2002). These pathways sensory transmission through spinal cord and
provide the neural mechanisms by which attentional, trigeminal sensory nuclei was influenced by des-
motivational, and cognitive variables filter ascending cending fibers from the cerebral cortex. The gate
sensory information. control hypothesis placed this evidence within the
framework of a theory of how pain modulation
Normal subjects
could account, in part, for the variant relationship
between noxious stimuli and the sensations they
produced. The first evidence to support endogenous
Civilian patients
Medical-phobic
pain control came from the study of Reynolds
(Reynolds, D. V., 1969) who demonstrated that
Perceived pain

focal brain stimulation of the midbrain periaque-

Wounded
ductal gray (PAG) produced sufficient analgesia to
soldiers
Injured allow surgery in rats without the use of chemical
athletes anesthetics. Liebeskind and colleagues quickly con-
firmed this finding and concluded that stimulation
of the PAG activated a normal function of the brain:
pain inhibition (Mayer, D. J. et al., 1971; Mayer, D. J.
Stimulus intensity and Liebeskind, J. C., 1974). Later evidence indi-
Figure 1 The variant pains. In this illustrative diagram, the
cates that pain modulation is not limited to
intensity of the perceived pain is plotted against the stimulus inhibition. Descending pathways also facilitate
intensity. The stimulus-response function curve of the pain transmission at the spinal level (Ren, K. et al.,
normal subjects is located in the middle. For wounded 2000; Millan, M. J., 2002; Gebhart, G. F., 2004). Most
soldiers and injured athletes, the curve may be shifted to the
recently, studies have been moved to address the
right and for civilian patients and medical-phobic, the curve
may be shifted to the left. The shift of the curve is translated role of pain modulatory circuitry in persistent, or
as changes in the sensation of pain. At certain stimulus chronic pain conditions. New lines of evidence sug-
intensity (arrow-dashed line), the civilian population may feel gest that the descending pathways exhibit dramatic
more pain and the wounded soldiers show less pain, when plasticity and multiplicity and are actively involved
compared to the normal subjects. The endogenous pain
in the development of persistent pain after tissue or
modulatory system contributes to the variability of pain
under different conditions. Adapted from Cervero, F. and nerve injury (Porreca, F. et al., 2002; Ren, K. and
Laird, J. M. 1996. Mechanisms of touch-evoked pain Dubner, R., 2002; Gebhart, G. F., 2004; Vanegas, H.
(allodynia): a new model. Pain 68, 1323. and Schaible, H.-G., 2004).
 Descending Control Mechanisms 725

49.2 Organization of Descending Cerebral cortex

Pathways Hypothalamus
Amygdala

There are multiple brain sites and pathways that are

involved in descending pain modulation, ranging from
the cerebral cortex to caudal medulla (Fields, H. L.
PAG
and Basbaum, A. I., 1978; 1999; Gebhart, G. F., 1988; Midbrain
Oliveras, J. L. and Besson, J. M., 1988; Sandkuhler, J.,
1996; Willis, W. D. and Westlund, K. N., 1997;
Millan, M. J., 2002) (Figure 2). To date, the most
well characterized endogenous pain modulatory path-
way involves a circuitry linking the midbrain PAG,
LC
rostral ventromedial medulla (RVM), and the spinal
cord. The second important source of descending pain Pons
control is from the dorsolateral pontomesencephalic
tegmentum (DLPT), which includes the cuneiform
nucleus in the midbrain ventrolateral to PAG and
locus coeruleus/subcoeruleus (LC/SC) together with
other neighboring noradrenaline cell groups (Fields,
H. L. and Basbaum, A. I., 1999). In the caudal medulla, Medulla
RVM
ventrolateral medulla (CVLM) including lateral reti-
cular nucleus (LRN), the subnucleus reticularis
dorsalis (SRD), and the nucleus tractus solitarius
DLF VLF
(NTS) have been identified to play a role in descend-
ing pain control. Additionally, hypothalamus and
other forebrain sites also participate in descending Noxious input
pain modulation. DH

Spinal cord

49.2.1 Key Structures and Pathways

Figure 2 Two major brainstem descending pathways
49.2.1.1 Midbrain PAG matter involved in pain modulation. The PAG in the midbrain has
PAG neurons are organized in longitudinal columns efferent projection to the rostral ventromedial medulla
(RVM) and locus coeruleus (LC). The RVM and LC directly
that function to integrate behavioral responses to project to the spinal dorsal horn (DH) where nociceptive
noxious and stressful stimuli (Bandler, R. and input is initially processed. The descending input from the
Shipley, M. T., 1994; Cameron, A. A. et al., 1995a; RVM and LC travels in the dorsolateral (DLF) and
1995b). The dorsal raphe nucleus ventromedial to the ventrolateral funiculus (VLF) and modulates spinal pain
cerebral aqueduct and the cuneiform nucleus lateral transmission (only postsynaptic modulation of dorsal horn
projection neuron is shown). Supraspinal projecting neurons
to the ventrolateral PAG are also considered a func- also issue collaterals to reach pain modulatory structures
tional entity with PAG for descending pain control and the PAG also receives input from forebrain structures
( Jensen, T. S. and Gabhart, G. F., 1988; Fields, H. L. including cerebral cortex, hypothalamus, and amygdala.
and Basbaum, A. I., 1999). The anatomical studies See text for details of other key structures and pathways
implicate the importance of the midbrain PAG in related to descending pain control. The drawings of brain
tissue sections are adapted from Paxinos, G. and Watson,
descending pain modulation. Ascending fibers from C. 2005. The Rat Brain in Stereotaxic Coordinates, 5th edn.
the spinal cord including axons of lamina I neurons Academic Press.
terminate in the PAG (Menetrey, D. et al., 1982;
Hylden, J. L. K. et al., 1986; Azkue, J. J. et al., 1998).
Afferent input from the cerebral cortex, hypothala-
mus, the cuneiform nucleus, caudal brainstem nuclei pain control, including RVM, LC/SC, A5, pontine
such as nucleus raphe magnus (NRM), also enters parabrachial nuclei (PB), NTS, hypothalamus and
PAG (Beitz, A. J., 1982b). The PAG provides major amygdala (Jensen, T. S. and Gabhart, G. F., 1988;
input to most areas that are involved in endogenous Fields, H. L. and Basbaum, A. I., 1999). The PAG
726 Descending Control Mechanisms

also is a site for regulation of other homeostatic critical involvement of the PAGRVMspinal dorsal
functions (Gauriau, C. and Bernard, J. F., 2002). For horn pathway in descending pain control.
instance, PAG neurons are activated by cardiovascu- In the rat, suppression of NRM neurotransmission
lar input (Murphy, A. Z. et al., 1995). Somatovisceral by either lesions, local anesthesia, or microinjection
interaction may occur in distinct organized columns of the N-methyl-D-aspartate (NMDA) receptor
of neurons in the PAG. antagonist reverses the spinal inhibition of nocifen-
Animal studies have demonstrated that electrical sive behavior produced by activation of PAG
stimulation of PAG produces profound antinocicep- neurons (Behbehani, M. M. and Fields, H. L., 1979;
tion, or analgesia (Reynolds, D. V., 1969; Mayer, D. J. Aimone, L. D. and Gebhart, G. F., 1986). Lidocaine
et al., 1971, Mayer, D. J. and Liebeskind, J. C., 1974; block of the NRM blocks PAG stimulation-produced
Fardin, V. et al., 1984a). The stimulation-produced inhibition of spinal nociceptive neurons in cat
analgesia was not due to a generalized sensory and (Gebhart, G. F. et al., 1983).
motor deficit. The animals could move and were RVM does not receive direct input from the
responsive to audiovisual stimuli during stimulation, spinal cord, although indirect spinal input can reach
while the responses to noxious stimuli were absent. RVM through the adjacent Gi (Basbaum, A. I. and
Stimulation of PAG also selectively inhibits Fields, H. L., 1984; Fields, H. L. and Basbaum, A. I.,
responses of dorsal horn and trigeminal sensory neu- 1999). The major source of input to the RVM is from
rons to noxious stimulus (Sessle, B. J. et al., 1981; the PAG and cuneiformis nucleus (Fardin, V. et al.,
Dostrovsky, J. O. et al., 1983; Gebhart, G. F., 1988). 1984b; Fields, H. L. and Basbaum, A. I., 1999). The
The stimulus-produced analgesia has been success- ventrolateral cell column of PAG projects to NRM
fully observed in humans with chronic intractable and the dorsolateral PAG region projects to the GiA
pain. Electrical stimulation of the midbrain periaque- (Cameron, A. A., et al., 1995b). However, Van
ductal and hypothalamus periventricular areas Bockstaele E. J. et al. (1991) show that injections of
produces long-standing analgesia from very brief tracers into the dorsal PAG resulted in anterograde
periods of stimulation. With implanted electrodes
labeling in the NRM.
in the brain, some patients can self-stimulate for
Electrical stimulation and microinjection of an opioid
1015 min, two to three times per day for adequate
receptor agonist into the PAG activate NRM neurons
pain control (Kumar, K. et al., 1997). It has been
that project to the spinal cord (Behbehani, M. M. and
observed in patients who have had failure of the
Pomeroy, S. L.,1978; Fields, H. L. and Anderson, S. D.,
implanted stimulation system, that the actual return
1978). The excitation of NRM neurons by injection of
of pain to its original pre-stimulation level takes 10
glutamate into the PAG is associated with an increase in
days to 2 weeks (Kumar, K. et al., 1997). This would
nociceptive response threshold (Behbehani, M. M. and
indicate that activation of the endogenous pain con-
Fields, H. L., 1979).
trol system has a long-lasting effect.
The RVM has major projections to the spinal
cord. Neurons of all subregions of RVM, NRM,
49.2.1.2 PAGRVMspinal dorsal horn GiA, and LPGi send axons to the spinal cord via
circuitry the dorsolateral funiculus (DLF) (Basbaum, A. I.
Convergent lines of evidence indicate that PAG and Fields, H. L., 1979; 1984). The densest terminal
stimulation-produced analgesia is relayed through fields of RVM descending fibers are in superficial and
other brainstem nuclei. The RVM has been deep dorsal horn where nociceptive information is
identified as the premier relay station between the processed.
PAG and spinal dorsal horn. The RVM is termed for Activation of RVM neurons produces antinoci-
collective structures that are functionally involved in ception. In the cat, electrical stimulation of NRM
descending pain modulation, mainly consisting of the completely suppresses the behavioral responses to
midline nucleus raphe magnus (NRM) and the noxious pinch of the skin and modifies the jaw-open-
adjacent gigantocellular reticular nucleus, alpha part ing reflex threshold to tooth pulp stimulation
(GiA). Anatomically, the raphe pallidus nucleus (Oliveras, J. L. et al., 1975). Electrical stimulation of
(Rpa) and a portion of the medial lateral paragigan- NRM or opioid microinjection inhibits activity of
tocellular nucleus (LPGi) also are located in dorsal horn nociceptive neurons (Fields, H. L. and
this region (Zagon, A., 1995; Paxinos, G. and Anderson, S. D., 1978; Dickenson, A. H. et al., 1979;
Watson, C., 2005). Extensive evidence supports the Jones, S. L. and Light, A. R., 1990).
 Descending Control Mechanisms 727

The DLF mediates descending modulation of dor- combined dopaminebeta-hydroxylase immunohisto-

sal horn substantia gelatinosa neurons (Cervero, F. chemistry and retrograde tracing, Westlund K. N.
et al., 1979). Spinal DLF lesions block antinociception et al. (1983) demonstrated that brainstem descending
produced by brainstem stimulation (Basbaum, A. I. noradrenergic fibers are exclusively from the pontine
et al., 1977). noradrenergic cell groups. In the rat, the LC and SC
Thus, although studies suggest that some PAG provide major descending noradrenergic axons to the
neurons project directly to the spinal cord and spinal spinal cord. These findings form the anatomical basis
trigeminal nucleus (Mantyh, P. W. and Peschanski, M., for a role of the LC/SC in descending control. One
1982; Mantyh, P. W., 1983; Skirboll, L. et al., 1983; concern on the spinal projection of LC/SC neurons is
Li, Y. Q. et al., 1993), the RVM relay is necessary for that they apparently terminate most heavily in lami-
the analgesic effect of PAG. Whether PAG can mod- nae VII, VIII, and IX of the ventral horn. The
ulate dorsal horn sensory transmission through a termination of LC fibers is moderate in the deep
direct pathway to the spinal cord still needs to be dorsal horn and relatively sparse in the superficial
determined. dorsal horn (Clark, F. M. and Proudfit, H. K., 1991).
However, there appears to be a substrain difference in
49.2.1.3 Dorsolateral LC/SC descending projection related to the commer-
pontomesencephalic tegmentum cial sources. More lamina I terminations are from the
In addition to the PAGRVMdorsal horn circuitry, LC in rats raised in Harlan Sprague-Dawley while the
the components of the DLPT constitute separate par- SC of Sasco source provides more lamina I projection
allel descending pathways responsible for pain (Fritschy, J. M. and Grzanna, R., 1990; Sluka, K. A. and
modulation (Dubner, R. and Bennett, G. J., 1983; Westlund, K. N., 1992; Willis, W. D. et al., 1995;
Gebhart, G. F., 1988; Fields, H. L. and Basbaum, A. I., Proudfit, H. K., 2002).
1999). One member of the DLPT, the cuneiform LC/SC stimulation produces inhibition of dorsal
nucleus, is located in the midbrain lateral reticular horn nociceptive neurons in rats and cats (Mokha, S.
formation neighboring ventrolateral PAG and shares S. et al., 1986; Jones, S. L. and Gebhart, G. F., 1986;
anatomical and physiological similarities with PAG Jones, S. L., 1991). Selective spinal lesions indicate
related to descending pain modulation. Electrical sti- that the effect of LC/SC is mediated by the ventro-
mulation of the midbrain lateral reticular formation lateral funiculus (VLF) ( Jones, S. L., 1991), whereas
produces powerful inhibition of dorsal horn nocicep- the NRM stimulation-produced inhibition is con-
tive neurons in cats (Carstens, E. et al., 1980). The other veyed via the DLF (Basbaum, A. I. et al., 1977).
members of the DLPT consist of LC/SC, A7 cell Unlike the PAG, the effect of LC on spinal noci-
group, PB and Kolliker-Fuse nuclei. The critical role ceptive transmission is not mediated through RVM
of these pontine noradrenaline cell groups in descend- since local anesthesia of NRM does not block LC
ing control has been established. stimulation-produced inhibition of dorsal horn neu-
rons ( Jones, S. L., 1991).
49.2.1.3.(i) The PAGLC/SCspinal pathway The Intrathecal yohimbine, an alpha 2 adrenoceptor
LC in the dorsal pontine is the major noradrenergic antagonist, blocks the LC stimulation-produced inhi-
nucleus originally designated the A6 cell group by bition of nociceptive reflex in rats (Gebhart, G. F.,
Dahlstrom A. and Fuxe K. (1964). The SC is located 1988).
immediately ventral to the LC and assigned to the A7 The LC/SC may also relay the effect of PAG
noradrenaline cell group by Westlund K. N. et al. since they receive PAG input. In monkey, dense
(1983). Since the two structures are relatively small projection to the LC is identified from the PAG
and the spinal projecting fibers from the LC are (Mantyh, P. W., 1983). In rat, the dorsolateral PAG
mainly from the ventral part, electrical stimulation cell column innervates LC/SC (Cameron, A. A. et al.,
will affect both nuclei (Jones, S. L., 1991). The fol- 1995b), although another study suggests that PAG
lowing studies provide evidence that the LC/SC does not have major projections to the LC proper
spinal pathway represents an additional source of (Ennis, M. et al., 1991).
descending pain modulation.
The LC/SC neurons project to the spinal cord. 49.2.1.3.(ii) Other noradrenaline cell groups in the
Initially, bulbospinal noradrenergic fibers are thought DLPT participating in descending modulation
to originate from the A1 cell group in the caudal 49.2.1.3.(ii).(a) A7 cell column The A7 cell group
medulla (Dahlstrom, A. and Fuxe, K., 1964). Using spans rostrocaudally the caudal midbrain and rostral
728 Descending Control Mechanisms

pons in the rat (Paxinos, G. and Watson, C., 2005). similarities with other noradrenergic cell groups in
The rostral tip of the A7 group can be seen at the the DLPT such as spinal projection (Westlund, K. N.
same transverse plane as the cuneiform nucleus and et al., 1983; Clark, F. M. and Proudfit, H. K., 1993) and
PAG. In the pons, the A7 is located medial to the extensive connections with other pain modulatory
dorsal part of the ventrolateral principal sensory tri- nuclei including PAG and CVLM (Cameron, A. A.
geminal nucleus and ventral to the supratrigeminal et al., 1995b; Tavares, I. and Lima, D., 2002).
and Kolliker-Fuse nuclei. The caudal A7 cells end Stimulation of the A5 produces inhibition of the
before the appearance of the LC. The A7 cells also nociceptive tail flick reflex in rats and the effect is
provide dense projection to the spinal cord mediated through spinal opioid and alpha adreno-
(Westlund, K. N. et al., 1983). This cell group may ceptors (Burnett, A. and Gebhart, G. F., 1991). Since
play a unique role in the descending circuitry. The A5 has been primarily considered as an important site
A7 group has reciprocal connections with RVM and for cardiovascular regulation, this cell group may
may mediate some effect of RVM on spinal nocicep- play a role in somatoautonomic integration.
tive transmission (Fields, H. L. and Basbaum, A. I.,
1999; Buhler, A. V. et al., 2004). 49.2.1.4 Caudal medulla
Several structures in the caudal medulla have
49.2.1.3.(ii).(b) Parabrachial nucleus The PB emerged as important players in endogenous pain
consists of several distinct subnuclei located in the control. As these sites overlap with those critical in
dorsolateral pons and belongs to the pontine cell groups visceral and cardiovascular functions, the caudal
that give rise to noradrenergic terminals in the spinal medulla provides a point of convergence to facilitate
cord (Westlund, K. N. et al., 1983). The PB has been interactions between somatosensory and autonomic
demonstrated as an important relay station in pain regulation.
pathways. Spinal and subnucleus caudalis lamina I
nociceptive neurons project to PB (Hylden, J. L. K. 49.2.1.4.(i) Caudal ventrolateral medulla
et al., 1989; Bester, H. et al., 2000). PAG neurons project Anatomically, the CVLM is not a clearly defined
to the PB in the rat (Krout, K. E. et al., 1998). The PB has structure, the center of which is about 2 mm ventral
further connections with the amygdala (Bernard, J. F. to the dorsal surface of the medulla and medial to
et al., 1993) and the hypothalamus (Bester, H. et al., the ventral pole of the spinal trigeminal nucleus in
1997), and sends descending projections to the spinal the rat. Rostrocaudally, the CVLM is within the
and medullary dorsal horn (Yoshida, A. et al., 1997). The most caudal segment of the medulla starting from
activation of the PB descending pathway modulates approximately 0.5 mm rostral to the calamus scrip-
spinal and trigeminal nociceptive transmission torius (Aicher, S. A. et al., 1996). This region is also
(Girardot, M. N. et al., 1987; Chiang, C. Y. et al., 1994; referred as the lateral reticular formation and
Willis, W. D. and Westlund, K. N., 1997). includes the A1 catecholamine cell group. The
poorly defined ventromedial border of lamina V of
49.2.1.3.(ii).(c)The Kolliker-Fuse nucleus The the trigeminal subnucleus caudalis may fall into the
Kolliker-Fuse nucleus is immediately dorsal to the territory of the CVLM, or vice versa. It has been
A7 cell group (Paxinos, G. and Watson, C., 2005) and consistently observed that a variety of noxious sti-
is the major source of spinopetal noradrenergic muli, including noxious heating of the hindpaw,
fibers in the cat (Stevens, R. T. et al., 1982). The peritoneal inflammation, urinary bladder inflamma-
Kolliker-Fuse nucleus is more developed in humans tion, deep muscle irritation or inflammation, all
than in other animal species (Lavezzi, A. M. et al., induce Fos protein expression in the CVLM (see
2004). Young R. F. et al. (1992) show that chronic Ren, K., 2002).
stimulation of the Kolliker-Fuse nucleus region can
relieve intractable pain in patients and the effect 49.2.1.4.(i).(a) Lateral reticular nucleus The
is comparable to that of PAG/periventricular LRN is a large nucleus located bilaterally in the
stimulation. ventrolateral medullary reticular formation. It
receives input from a variety of structures including
A5 cell column The A5 cell group
49.2.1.3.(ii).(d) PAG and NRM and projects to the spinal cord
spans the ventrolateral pontine tegmentum at the ( Janss, A. J. and Gebhart, G. F., 1987; Lee, H. S. and
mid-pon level to the rostral ventrolateral medulla Mihailoff, G. A., 1999). Electrical or chemical
(RVLM) in the rat. This cell group shares many stimulation of the LRN produces inhibition of
 Descending Control Mechanisms 729

the nociceptive reflex and spinal dorsal horn unit vagal afferent modulation of nociception is mediated
responses to noxious stimuli. The LRN-produced through NTS (Randich, A. and Gebhart, G. F., 1992).
descending inhibition is partially mediated by bilat- The function of NTS supports somatovisceral and
eral DLF and spinal alpha 2 adrenergic and serotonin somatoautonomic interactions.
receptors ( Janss, A. J. and Gebhart, G. F., 1987; 1988).
49.2.1.5 Forebrain sites
49.2.1.4.(i).(b) Lateral CVLM The CVLMlat, a A number of forebrain structures provide descending
small area of the medullary reticular formation input to brainstem nuclei, particularly the PAG
between the ventral pole of the spinal trigeminal (Beitz, A. J., 1982b).
nucleus and LRN, has recently been proposed to
have a major role in descending pain inhibition 49.2.1.5.(i) Hypothalamus The subregions of
(Tavares, I. and Lima, D., 2002). The CVLMlat has the hypothalamus provide the largest descending
reciprocal connections with spinal lamina I neurons input to the PAG (Beitz, A. J., 1982b). The periven-
and output to the A5 cell group and RVM. These tricular gray (PVG) of the hypothalamus is the rostral
indirect spinopetal pathways from the CVLMlat via extension of PAG and rich in opioids. Axons of PVG
A5 and RVM may mediate noradrenergic and sero- neurons can reach the spinal cord (Jensen, T. S.
tonergic antinociception produced by CVLM and Gabhart, G. F., 1988; Van den Pol, A. N. and
activation. Neurons in this region are reciprocally Collins, W .F., 1994). Stimulation of PVG can produce
connected to the RVM and activated after orofacial effective pain relief in patients (Kumar, K. et al., 1997).
tissue injury (Sugiyo, S. et al., 2005). A group of The lateral hyphthalamus (LH) has reciprocal
reticulohypothalamic tract neurons in the CVLM connections with the PAG and NRM (Aimone, L. D.
may overlap with the CVLMlat (Malick, A. et al., et al., 1988; Cameron, A. A. et al., 1995a; 1995b).
2000). Most of these neurons respond to both innoc- Inhibition of spinal nociceptive transmission from the
uous and noxious stimulation of the head but only to LH requires a bulbar relay in the NRM (Aimone, L. D.
noxious stimulation of extracephalic regions. Their et al., 1988).
response properties suggest that they participate in
pain modulation. 49.2.1.5.(ii) Amygdala The ascending nocicep-
tive input reaches the amygdala through a relay in
49.2.1.4.(ii) Subnucleus reticularis dorsalis The the PB (Gauriau, C. and Bernard, J. F., 2002). The
SRD includes a group of neurons distributed in the amygdala has reciprocal connections with PAG
dorsal medullary reticular formation medial to (Rizvi, T. A. et al., 1991). Activation of amygdala
the trigeminal subnucleus caudalis (also called neurons produces analgesia that is mediated through
medullary reticular nucleus, dorsal part, MdD) PAG (see Fields, H. L. and Basbaum, A. I., 1999). The
(Paxinos, G. and Watson, C., 2005). The SRD con- dorsal PAG interacts with the amygdala to affect fear
tributes to the spinoreticularthalamic pathway by and anxiety and the amygdala is an important site for
receiving nociceptive information from the spinal interrelating sensory and affective dimensions of pain
dorsal horn and relays it to the thalamus (Behbehani, M. M., 1995; Price, D. D., 2002).
(Villaneuva, L., et al. 1998). The diffuse noxious inhi-
bitory control (DNIC), proposed by Le Bars and 49.2.1.5.(iii) Cerebral cortex Recent brain ima-
colleagues, is mediated via the SRD and independent ging studies have generated a wealth of information
of the RVM-spinal pathway (Bouhassira, D. et al., on activation of the cortical areas during pain
1992; 1993). The SRD is also proposed as a pronoci- processing. It also shows that the cerebral cortex is
ceptive center (Lima, D. and Almeida, A., 2002). not only the end point of the pain pathway, but also a
source of descending input that contributes to the
49.2.1.4.(iii) Nucleus tractus solitarius The NTS fine-tuning of pain threshold and modulation of the
is a key structure in somatovisceral processing, since emotional aspect of pain ( Jasmin, L. et al., 2003b;
this nucleus receives somatic and visceral input and Petrovic, P. et al., 2004). One interesting pathway
has extensive efferent connections with other nuclei involves the anterior cingulate cortex (ACC) and
(Randich, A. and Maixner, W., 1984; Ren, K. et al., amygdala in pain modulation. The ACCamygdala
1990). Notably, PAG neurons project to NTS pathway may modulate pain-related stress and
(Cameron, A. A. et al., 1995b). Activation of NTS aversive responses, as well as pain-related fear
neurons produces descending pain inhibition and the memory (Gao, Y. J. et al., 2004; Petrovic, P. et al.,
730 Descending Control Mechanisms

2004; Tang, J. et al., 2005). Transcranial magnetic of pain transmission through their respective recep-
stimulation of the dorsolateral prefrontal cortex tor subtypes. The major neurotransmitters that are
increased human pain tolerance level (Graff- implicated in descending pain control are briefly
Guerrero, A. et al., 2005). discussed below.

49.2.1.6 Summary
It is clear from the above overview that the pivotal 49.2.2.1 Endogenous opioids and related
anatomical substrates for descending pain control are peptides
located within the brainstem. In addition to the mid- 49.2.2.1.(i) Opioid peptides Tsou K. and Jang C.
brain PAG, a number of important sites are in S. (1964) discovered that the most sensitive sites for the
the medulla. The medulla oblongata is classically analgesic action of morphine were located in the PAG
termed the vital center for life with its critical and the adjacent hypothalamic periventricular area.
involvement in cardiovascular and respiratory con- These early findings suggested that opiate drugs like
trols. Since pain is arguably one of the most morphine acted by binding to a receptor in the brain
important body functions for survival, it is not sur- and that there likely were endogenous ligands or
prising that the structures that are vital for pain chemical mediators whose actions were mimicked
modulation are also located in the medulla to facil- by opiates. Opiate binding sites in the brain were
itate interactions between somatosensory and subsequently demonstrated in early 1970s by radioli-
autonomic regulation. The interplay of the descend- gand binding assays (Pert, C. B. and Snyder, S. H.,
ing system with visceral (NTS-PB), cardiovascular 1973; Simon, E. J. et al., 1973; Terenius, L., 1973) and
(CVLM, A5, PAG), respiratory (Kolliker-Fuse the first endogenous opioid peptides enkephalins
nucleus), micturation (RVM, pons), arousal (A6), were identified soon after (Hughes, J. et al., 1975).
and emotional (amygdala, ACC) regulation is exten- To date, four classes of opioid peptides have been
sive and serves to enhance survival (Randich, A. found in the brain: endorphins, enkephalins, dynor-
and Maixner, W., 1984; Randich, A. and Gebhart, phins, and endomorphins (Hughes, J. et al., 1975; Li,
G. F., 1992; Murphy, A. Z. et al., 1995; Mason, P., C. H. and Chung, D., 1976; Goldstein, A. et al., 1979;
2001). Zadina, J. E. et al., 1997). Martin W. R. et al. (1976) first
proposed the existence of three subtypes of opioid
receptors based on three different syndromes pro-
duced by congeners of morphine. All three opioid
49.2.2 Neurotransmitters
receptors,  (MOP),  (DOP), and (KOP), have
A plethora of transmitters (also see Chapter now been cloned (Evans, C. J. et al., 1992; Kieffer, B. L.
Pharmacological Modulation of Pain) have been et al., 1992; Chen, Y. et al., 1993; Meng, F. et al., 1993).
shown to be involved in descending pain control The endogenous opioid peptides and their respective
(Duggan, A. W., 1985; Fields, H. L. et al., 1991; precursors, amino acid sequence, and preferred
Millan M. J., 2002). The most studied and established receptor subtypes are summarized in Table 1. Recent
candidates include endogenous opioid peptides, nor- studies also indicate the existence of alternative spli-
adrenaline, and serotonin (5-HT). All these cing (Pan, L. et al., 2005), subtypes of the mu, delta, and
transmitters participate in inhibition and facilitation kappa receptors (Fowler, C. J. and Fraser, G. L., 1994;

Table 1 Endogenous opioid peptides involved in pain modulation

Precursor Name Amino acid sequence Receptor

POMC -Endorphin Tyr-Gly-Gly-Phe-Met-Thr-Ser-Glu-(human)

(pro-opiomelanocortin) Lys-Ser-Gln-Thr-Pro-Leu-Val-Thr-Leu-Phe-Lys-Asn-Ala-Ile- 
Ile-Lys-Asn-Ala-Tyr-Lys-Lys-Gly-Glu-OH
Proenkephalin Leu-enkephalin Tyr-Gly-Gly-Phe-Leu-OH 
Proenkephalin Met-enkephalin Tyr-Gly-Gly-Phe-Met-OH 
Prodynorphin Dynorphin A Tyr-Gly-Gly-Phe-Leu-Arg-Arg-Ile-Arg-Pro-Lys-Leu-Lys-Trp-
Asp-Asn-Gln-OH
Unidenitified Endomorphin-1 Tyr-Pro-Trp-Phe-NH2 
Unidenitified Endomorphin-2 Tyr-Pro-Phe-Phe-NH2 
 Descending Control Mechanisms 731

Zaki, P. A. et al., 1996; Pasternak, G. W., 2001), and Although there is no question that opioids pro-
opioid receptor gene polymorphism (Ikeda, K. et al., duce potent pain inhibition through their respective
2005). The functional significance of the further mole- receptors at all levels of the descending circuitry, the
cular diversity of opioid receptor subtypes in role of spinopetal opioidergic fibers in mediating
descending pain control is poorly understood. descending modulation from supraspinal sites has
The endogenous opioids and their receptors are yet to be convincingly demonstrated. Intrathecal
distributed in structures involved in descending pain administration of the opioid receptor antagonists in
control at all levels including the amygdala, hypotha- rats has been shown to reverse or attenuate analgesia
lamus, PAG, DLPT, RVM, and spinal cord (Basbaum, produced by intracerebroventricular morphine
A. I. and Fields, H. L., 1984; Ruda, M. A., 1988; (Gogas, K. R. et al., 1996), intra-PAG injection of
Mansour, A. et al., 1995; Fields, H. L. and Basbaum, beta-endorphin (Tseng, L. F. and Tang, R., 1990),
A. I., 1999; Wang, H. and Wessendorf, M. W., 2002). electrical stimulation of the RVM (Zorman, G. et al.,
Notably, beta-endorphin terminals from hypothalamus 1982; Lu, Y. et al., 2004), or the hypothalamic arcuate
descend along the borders of the third ventricle and nucleus (Wang, Q. et al., 1990). However, the con-
reach PAG, DLPT, and NRM (Finley, J. C. et al., 1981; tribution of intrinsic opioidergic interneurons in
Jensen, T. S. and Gabhart, G. F., 1988). Rostral ven- the spinal dorsal horn to supraspinal-produced
tromedial medulla neurons express multiple opioid analgesia cannot be ruled out. In the cat, opioid
receptor phenotypes (Marinelli, S. et al., 2002). In the peptides intrinsic to the spinal cord play a major
RVM, spinally projecting serotonergic and nonseroto- role in inhibition of spinal nociceptive transmission
nergic neurons express mu- and delta-opioid receptor (Duggan, A. W., 1985).
mRNAs (Wang, H. and Wessendorf, M. W., 1999).
49.2.2.1.(ii) Orphanin FQ, nocistatin, and
Enkephalinergic axons in the spinal cord synapse
neuropeptideFF Orphanin FQ, or nociceptin, is a
with thalamic projection neurons (Ruda, M. A., 1982),
heptadecapeptide that was identified as the endogen-
providing postsynaptic modulation of ascending noci-
ous ligand for the orphan-opioid-receptor-like 1
ceptive information.
(ORL-1, now nociceptin/orphanin FQ peptide
Microinjection of morphine into the PAG produces
receptor, NOP) receptor (Meunier, J. C. et al., 1995;
behavioral analgesia (Camarata, P. J. and Yaksh, T. L.,
Reinscheid, R. K. et al., 1995). Genetically, Orphanin
1985). Brain stimulation-produced analgesia can be
FQ and NOP belong to the same family with the
reversed by naloxone, an opioid receptor antagonist
opioids and their receptors and they share homology
(Akil, H. et al., 1976). Behavioral analgesia produced by
with dynorphin A and the kappa opioid receptor,
an opioid receptor agonist is related to the inhibition
respectively. However, orphanin FQ does not
of spinal nociceptive neurons (Yaksh, T. L., 1978).
interact with three classical opioid receptors, largely
However, the effect of opioid agonists on dorsal horn
due to the replacement of the first tyrosine
nociceptive neurons is more variable and can be both residue by phenylalanine. Most studies show that
inhibitory and facilitatory (Gebhart, G. F., 1982; orphanin FQ produces analgesia at the spinal level
Stamford, J. A., 1995). This paradox is likely related and both pro- and anti-nociception supraspinally
to the sampling limitation during single unit recording (Meunier, J. C., 1997; Henderson, G. and McKnight,
and cautions on predicting behavioral outcome from A. T., 1997). However, intrathecal low dose of
activity of single neurons. Using Fos immunoreactiv- orphanin FQ also produces allodynia (Yamamoto, T.
ity to study a population of activated neurons et al., 1999). Orphanin FQ can suppress both
simultaneously, opioids produce behavioral analgesia pain facilitatory and inhibitory neurons in the
that is correlated to an inhibition of dorsal horn neu- RVM, which may explain its dual effects on pain
ronal activation as indicated by a reduction of noxious processing under different conditions (Pan, Z. Z.
stimulus-induced Fos immunoreactivity (Gogas, K. R. et al., 2000; Vaughan, C. W. et al., 2001; Heinricher,
et al., 1991). Is the endogenoous opioid system toni- M. M., 2005).
cally active? The findings from opioid receptor Nocistatin is liberated from the same orphanin FQ
knockout mice suggest a low endogenous opioid tone precursor protein, proorphanin, after posttransla-
with each receptor subtype contributing differentially tional cleavage but binds to a receptor distinct from
to the modulation of transient pain (Matthes, H. W. NOP (Okuda-Ashitaka, E. et al., 1998). Nocistatin
et al., 1996; Sora, I. et al., 1997; Gaveriaux-Ruff, C. and opposes action of orphanin FQ, and can be
Kieffer, B. L., 2002). considered a functional antagonist of orphanin FQ
732 Descending Control Mechanisms

(Yamamoto, T. et al., 1999; Millan, M. J., 2002). intrathecal NA elevates the nociceptive threshold
Proorphanin also yields another heptadecapeptide, (Reddy, S. V. et al., 1980). Opioid analgesia is enhanced
orphanin FQ2, which may participate in pain inhibi- in norepinephrine transporter knockout mice and the
tion (Rossi, G. C. et al., 1998). effect is blocked by yohimbine (Bohn, L. M. et al.,
The octapeptide neuropeptideFF (NPFF, Phe- 2000). Intrathecal administration of the adrenoceptor
Leu-Phe-Gln-Pro-Gln-Arg-Phe-NH2) is originally antagonists produces a decrease in nociceptive thresh-
isolated from the bovine brain and found to possess old in the rat, suggesting that the descending
anti-morphine property (Yang, H. Y. et al., 1985). noradrenergic system is tonically active (Sagen, J.
NPFF fibers project to the PAG from hypothalamus and Proudfit, H. K., 1984). The antinociceptive effect
and to the RVM from the NTS (Aarnisalo, A. A. and of NA in the spinal cord does not appear to require
Panula, P., 1995). In the spinal cord, NPFF-immu- GABAergic, glycinergic, opioidergic, and serotoner-
noreactive cell bodies and terminals are localized to gic activity (Reddy, S. V. et al., 1980), and may
laminae IIV and X and DLF, but a supraspinal involve both pre- and postsynaptic mechanisms
origin of NPFF was not found (Kivipelto, L. and (Sonohata, M. et al., 2004). In vivo whole-cell patch-
Panula, P., 1991). Available evidence indicates that clamp recordings from rat substantia gelatinosa neu-
NPFF produces antinociception at the spinal level rons show that NA reduces EPSC and suppresses
and pronociception (anti-opioid) at the supraspinal action potentials evoked by a noxious stimulus,
level (Millan, M. J., 2002; Mollereau, C. et al., 2005). an effect that is blocked by antagonism of alpha 2,
but not alpha 1, adrenoceptors (Sonohata, M. et al.,
49.2.2.2 Monoamines 2004).
It is now well established that descending monoami- Studies using transgenic mice, such as D79N mice
nergic fibers mediate descending modulation from the with a point mutation of the alpha 2A adrenoceptors,
supraspinal sites (Yaksh, T. L., 1979; Basbaum, A. I. and have examined the subtypes of spinal alpha 2 adre-
Fields, H. L., 1984; Gebhart, G. F., 1988; Tjolsen, A. noceptors involved in descending inhibition. Most
et al., 1991; Hung, K. C. et al., 2003). The major path- studies support a role for the alpha 2A subtype in
ways involve noradrenergic fibers from the DLPT spinal analgesia (Hunter, J. C. et al., 1997; Lakhlani,
and serotonergic fibers from the NRM, the major P. P. et al., 1997; Stone, L. S. et al., 1997; Kingery, W. S.
nucleus of RVM. The two pathways can be co- et al., 2000; Malmberg, A. B. et al., 2001; Mansikka, H.
activated and may act synergistically to participate et al., 2004; Ozdogan, U. K. et al., 2004). However,
in descending pain modulation (Gebhart, G. F., alpha 2C adrenoceptor-bearing terminals synapse
1988; Sawynok, J., 1989; Danzebrink, R. M. and with NK-1 projection cells in the rat dorsal horn
Gebhart, G. F., 1991). (Olave, M. J. and Maxwell, D. J., 2003), suggesting
a role in postsynaptic modulation. The alpha
49.2.2.2.(i) Noradrenaline Intrathecal administra- 2C adrenoceptors have been shown to be involved
tion of the alpha 2-adrenoceptor antagonist in spinal analgesia and adrenergicopioid synergy
yohimbine, but not the nonselective opioid receptor (Fairbanks, C. A. et al., 2002; Lahdesmaki, J. et al.,
antagonist naloxone and alpha 1 adrenoceptor 2003). The alpha 2B adrenoceptors may play an
antagonist prazosin, attenuates descending inhibition important role in the pontine noradrenergic nuclei
of nocifensive tail flick reflex produced by focal involved in analgesic action of nitrous oxide
stimulation of the LC/SC region (Jones, S. L., (Sawamura, S. et al., 2000).
1991). The PAG-produced inhibition of dorsal horn After peripheral nerve injury, there is a decrease
neurons is reduced by microdialysis of alpha 2 adre- in alpha 2A mRNA and protein immunoreactivity in
noceptor antagonists idazoxan and yohimbine into the spinal cord of the rat (Stone, L. S. et al., 1999;
the dorsal horn (Peng, Y. B. et al., 1996). Selective Leiphart, J. W. et al., 2003); these findings suggesting a
depletion of spinal noradrenaline by intrathecal reduced noradrenergic transmission in these rats.
DSP4 attenuates morphine analgesia (Zhong, F. X. However, there is also an increased descending nor-
et al., 1985). These results clearly indicate a role of adrenergic innervation to the spinal dorsal horn after
spinopetal noradrenergic fibers in descending mod- nerve injury, possibly serving to enhance inhibition
ulation since there are no noradrenaline-producing (Ma, W. and Eisenach, J. C., 2003). Interestingly, pain
cell bodies in the spinal cord (Swanson, L. W. and hypersensitivity after nerve injury develops similarly
Hartman, B. K., 1975). Consistently, depletion of in the wild-type mice and those with mutation of
spinal NA by 6-hydroxydopamine decreases and alpha 2A, 2B, or 2C adrenoceptors, suggesting that
 Descending Control Mechanisms 733

the net inhibitory tone is not affected by loss of alpha It has become clear that descending serotonergic
2 adrenoceptors (Malmberg, A. B. et al., 2001). It is fibers also participate in facilitation of nociception
worth noting that, different from previous observa- (Zhuo, M. and Gebhart, G. F., 1991; Millan, M. J.,
tions in normal rats (Sagen, J. and Proudfit, H. K., 2002; Suzuki, R. et al., 2002b; 2004b) The nociceptive
1984), the baseline nociceptive threshold is not facilitation triggered by low intensity vagal afferent
affected in mice with mutation of either of the three stimulation involves descending serotonergic path-
alpha 2 subtypes (Malmberg, A. B. et al., 2001), sug- ways (Ren, K. et al., 1991). Although spinal
gesting that compensatory mechanisms may have application of 5-HT generally produces an analgesic
developed in the mutant mice to balance the loss of effect, excitation of dorsal horn neurons by seroto-
tonic noradrenergic inhibition. nergic agonists is also reported (Todd, A. J. and
In contrast to a predominant inhibitory role of the Millar, J., 1984; Zhang, Y. et al., 2001). In the spinal
alpha 2 adrenoceptors, the alpha 1 adrenoceptor may be slice preparation, serotonin can recruit silent gluta-
involved in descending pain facilitation (Holden, J. E. matergic synapses in the superficial dorsal horn and
et al., 1999; Nuseir, K. and Proudfit, H. K., 2000; enhance synaptic transmission (Li, P. and Zhuo, M.,
Hedo, G. and Lopez-Garcia, J. A., 2001; Holden, J. E. 1998). This result supports a facilitatory role of des-
and Naleway, E., 2001) and supraspinal pronociceptive cending serotonergic fibers in spinal plasticity in
action (Kingery, W. S. et al., 2002). Intracellular record- response to injury. Indeed, intrathecal administration
ings from adult rat spinal slice support the opposing of methysergide has been shown to attenuate neuro-
actions of NA on substantia gelatinosa neurons. pathic pain in the rat (Pertovaara, A. et al., 2001). In
Noradrenaline induces hyperpolarization or increased carrageenan-inflamed rats, the facilitation of dorsal
spontaneous excitatory postsynaptic potentials in sub- horn neuronal responses to noxious stimuli by ago-
populations of dorsal horn neurons, which is selectively nists of 5-HT1A and 5-HT1B receptors is further
blocked by alpha 2 (yohimbine) and alpha 1 (prazosin) enhanced (Zhang, Y. et al., 2001). Blockade of spinal
adrenoceptor antagonists, respectively (North, R. A. 5-HT3 receptors reduces tissue injury-induced per-
and Yoshimura, M., 1984). sistent pain (Zeitz, K. P. et al., 2002).
At least 12 different subtypes of 5-HT receptors
may be involved in pain modulation (Hoyer, D. et al.,
49.2.2.2.(ii) Serotonin There is dense serotonergic 1994; Millan, M. J., 2002). However, their roles in
innervation in the spinal dorsal horn (Ruda, M. A. et al., descending pain control have only been evaluated in
1982; Ruda, M. A., 1988), which is almost exclusively a few subtypes including 5-HT1A, 5-HT1B, 5-HT2A,
derived from supraspinal sources (LaMotte, C. C., 1988; 5-HT3, and 5-HT4 receptors (Millan, M. J., 2002).
Fields, H. L. et al., 1991; Mason, P., 2001; Millan, M. J., Recent studies have identified a pronociceptive role of
2002). The NRM is the major structure that issues the 5-HT3 subtype (Suzuki, R. et al., 2002b; Zeitz, K. P.
descending serotonergic fibers to the spinal cord et al., 2002). A number of studies have indicated a
(Bowker, R. M. et al., 1988), although not all facilitatory role of the 5-HT1A receptor while others
descending axons from the NRM are serotonergic pointed to an opposite effect (Millan, M. J., 2002). The
(Jones, S. L. and Light, A. R., 1992; Wang, H. and equivocal literature on the role of 5-HT receptors in
Wessendorf, M. W., 1999). Descending inhibition pro- pain modulation suggests sophisticated differential
duced by focal electrical stimulation in the PAG or involvement of the 5-HT receptor subtypes in pain
NRM is mediated in part by descending serotonergic modulation and transmission. Moreover, different
pathways acting on spinal serotonergic receptors. experimental conditions including the chosen end-
Microinjection of morphine into the PAG and stimula- points, stimulation modality, anesthetic state, and
tion of NRM evokes the release of serotonin from spinal in vivo versus in vitro preparations are also confound-
cord (Yaksh, T. L. and Tyce, G. M., 1979; Rivot, J. P. ing factors.
et al., 1982). Intrathecal methysergide, a nonselective Despite a well-documented contribution of the
5-HT receptor antagonist, increases the intensity of descending serotonergic pathway to pain modula-
stimulation in the PAG and the NRM for inhibition of tion, the specific role of RVM serotonergic neurons
the nocifensive tail flick reflex while naloxone has no in the operation of descending circuitry is not under-
effect (Gebhart, G. F., 1988). The role of the descending stood. Stimulation of the PAG that produces
serotonergic pathway in descending pain inhibition is analgesia does not excite RVM serotonergic neurons
well established (Fields, H. L. and Basbaum, A. I., 1978; (Gao, K. et al., 1997). Thus, these cells may not
Basbaum, A. I. and Fields, H. L., 1984). mediate the signal from PAG, at least not directly.
734 Descending Control Mechanisms

Stimulation of NRM that induces antinociception including some sites in the PAG can also induce
does not release 5-HT in the spinal cord, although pain-like behavior via activation of NMDA receptors
further increasing the stimulation intensity results in ( Jensen, T. S. and Yaksh, T. L., 1992). Thus, the
5-HT release (Sorkin, L. S. et al., 1993). These results glutamatergic transmission within the pain modula-
are consistent with earlier findings that both seroto- tory circuitry is also capable of bidirectional pain
nergic and nonserotonergic pathways contribute to modulation.
descending inhibition (Rivot, J. P. et al., 1980). Careful Glutamate receptors consist of ionotropic
analysis of the characteristics of RVM serotonergic and metabotropic (mGluR) subfamilies. The former
neurons revealed that their levels of activity depend includes NMDA, alpha-amino-3-hydroxy-5-methyl-
on the conscious state (Mason, P., 2001). Serotonergic 4-isoxazole propionic acid (AMPA), and kainate (KA)
cells exhibit the highest level of discharge during receptor subtypes, and the eight subtypes of metabo-
waking when nociceptive threshold is relatively tropic glutamate receptors (mGluRs) are subdivided
high compared to the sleep state. One hypothesis is into three groups. Studies on the role of specific
that the descending serotonergic modulation of pain glutamate subtypes in descending pain control are
transmission is state dependent and closely related to still in the preliminary stage. In the PAG, group
alterations in serotonergic tone. I mGluR may be involved in pain inhibition
involving an interaction with NMDA receptors
49.2.2.2.(iii) Dopamine Similar to noradrenaline (Maione, S. et al., 1998; Berrino, L. et al., 2001). In
and serotonin, spinal dopamine is virtually exclu- the RVM, activation of group II mGluR produces
sively of supraspinal origin, mainly from the descending inhibition of the nociceptive tail flick
hypothalamus (see Millan, M. J., 2002). The dopa- reflex (Kim, S. J. et al., 2002). Activation of AMPA
mine receptor consists of D1 and D2 subtypes and NMDA receptors contributes to plasticity in
and they both have been localized in the spinal cord the RVM in response to tissue injury (Urban, M. O.
including the dorsal horn. The D1 receptor may and Gebhart, G. F., 1999; Ren, K. and Dubner, R.,
support opioid analgesia from PAG activation 2002).
(Flores, J. A. et al., 2004) and the activity of the D2 Unlike noradrenaline and serotonin, primary
subtype may oppose opioid analgesia (King, M. A. afferent fibers are rich in glutamate (Battaglia, G.
et al., 2001). The contribution of the descending and Rustioni, A., 1988). At the spinal level, glutamate
dopaminergic pathway to modulation of spinal receptors play a major role in the development
nociceptive transmission remains to be determined. of persistent pain. It is difficult to isolate the effect
of descending glutamatergic input from that of
49.2.2.3 Amino acids primary afferent origin. Limited information is
Excitatory and inhibitory amino acid neurotransmit- available on the anatomy of descending glutamater-
ters are widely distributed in the pain modulatory gic pathways. One study shows that descending
circuitry including the PAG, DLPT, RVM, and pathways from the DLPT nuclei contain glutamater-
spinal cord. They clearly participate in the process gic fibers (Liu, R. H. et al., 1995). However, whether
of descending pain control through actions on their they contribute to descending pain modulation is
respective receptors. unclear.

49.2.2.3.(i) Glutamate Activation of supraspinal

glutamate synapses is associated with antinocicep- 49.2.2.3.(ii) Gamma-aminobutyric acid and
tion. Glutamate-containing neurons are observed in glycine In general, gamma-aminobutyric acid
the PAG of the cat (Barbaresi, P. et al., 1997). (GABA) and glycine are used by inhibitory inter-
Stimulation-produced analgesia from the PAG is neurons. In the PAG, GABAergic terminals are
mediated by excitatory amino acid receptors in the abundant (Reichling, D. B. and Basbaum, A. I.,
RVM (Aimone, L. D. and Gebhart, G. F., 1986; van 1990). Blocking GABA receptors in the ventral
Praag, H. and Frenk, H., 1990; Spinella, M. et al., PAG enhances morphine-produced analgesia in this
1996). Stimulation of the cuneiform nucleus excites region, consistent with the view that GABAergic
about 50% of NRM neurons, an effect blocked by an neurons inhibit PAG output neurons involved in
excitatory amino acid antagonist (Richter, R. C. and descending inhibition (Depaulis, A. et al., 1987).
Behbehani, M. M., 1991). However, microinjection GABAergic terminals from the PAG descend to the
of glutamate into the discrete brainstem sites RVM (Reichling, D. B. and Basbaum, A. I., 1990).
 Descending Control Mechanisms 735

RVM GABA-containing neurons and GABAergic the synaptic cleft and access and facilitate nearby
terminals of PAG neurons both synapse with spinally NMDA receptors through spillover (Ahmadi, S.
projecting RVM neurons (Cho, H. J. and Basbaum, A. I., et al., 2003).
1991). Inhibition of GABAergic transmission in the
RVM leads to disinhibition of pain inhibitory activity. 49.2.2.4 Others
Apparently, the GABA system may set up the tone of A number of additional neurotransmitters also parti-
descending inhibition and help to maintain an ade- cipate in descending pain control, although their
quate level of pain sensitivity, which is essential to roles are studied less extensively and a large portion
normal body function. Direct GABAergic innerva- of the study has focused on the different segmental
tion of the spinal dorsal horn by fibers descending levels. Some key observations are discussed below.
from the RVM has been described (Antal, M. et al.,
1996). However, confounded by the presence of 49.2.2.4.(i) Cannabinoids Endocannabinoids
abundant intrinsic GABAergic neurons in the spinal (endogenous marijuana-like compounds) include
cord, the role of descending GABAergic fibers in anandamide and 2-arachidonylglycerol, which act
descending control is unclear. on two subclasses of cannabinoid receptors, CB1
Alterations of GABAergic activity may result in and CB2 (Cravatt, B. F. and Lichtman, A. H., 2004;
either an increase or decrease in pain sensitivity. In Hohmann, A. G. et al., 2005). The CB1 receptors are
the insular cortex of the rat, increasing and decreas- localized in the CNS including important structures
ing GABA transmission produces analgesia and of descending pain control (Tsou, K. et al., 1998).
hyperalgesia, respectively ( Jasmin, L. et al., 2003b). Endogenous cannabinoids produce analgesia via
Thus, GABAergic activity in this region appears to mechanisms apparently different from those of endo-
facilitate cortical output related to descending genous opioids, although they may act in similar
inhibition, which is in contrast to PAG and RVM. neural circuitry involving the PAG and RVM
In the RVM, the GABAB agonist baclofen produces (Walker, J. M. and Huang, S. M., 2002). Stress induces
antinociception at low dose (0.11.0 ng) and hyp- endocannabinoids in the PAG associated with
eralgesia at high dose (30150 ng) (Thomas, D. A. analgesia that can be reversed by CB1 receptor
et al., 1995; Hammond, D. L. et al., 1998). Intrathecal blockade in the region (Hohmann, A. G. et al., 2005).
methysergide antagonizes the baclofen-produced In CB1-mutant mice, the stress-induced analgesia is
antinociception, suggesting that analgesia is related selectively affected, leaving the opioid analgesia
to disinhibition of bulbospinal serotonergic neurons unaffected (Valverde, O. et al., 2000). The analgesic
(Hammond, D. L. et al., 1998). GABAergic neurons in effect of cannabinoids requires the RVM circuitry
the DLPT also exert bidirectional pain modulation and cannabinoids modulate activity of RVM pain
by providing tonic inhibition of two populations of modulating neurons (Meng, I. D. et al., 1998).
noradrenergic neurons in the A7 group (Nuseir, K. Although generally considered to be localized per-
and Proudfit, H. K., 2000). ipherally, increasing evidence indicates that the CB2
Stimulation of the PAG and NRM results in receptor in the spinal cord plays a role in the persis-
release of glycine in the spinal dorsal horn and inhibi- tent pain process (Zhang, J. et al., 2003).
tion of nociceptive neuronal activity (Sorkin, L. S.
et al., 1993; Cui, M. et al., 1999). Iontophoresis of 49.2.2.4.(ii) Brain-derived neurotrophic factor In
glycine and GABA receptor agonists suppresses the adult mammalian brain, brain-derived neuro-
responses of primate spinothalamic tract neurons to trophic factor (BDNF) facilitates excitatory synaptic
noxious stimuli (Lin, Q. et al., 1999). Clearly, disinhibi- transmission and long-term synaptic plasticity.
tion of glycinergic and GABAergic neurotransmission Studies have shown that exogenously applied
constitutes a mechanism of central sensitization and BDNF into the PAG produces analgesia in rats
persistent pain (Cronin, J. N. et al., 2004; Zeilhofer, H. (Siuciak, J. A. et al., 1995; Frank, L. et al., 1997).
U., 2005), although to what extent the descending Overexpression of BDNF in the spinal cord alleviates
control intervenes in this process needs to be speci- neuropathic pain (Cejas, P. J. et al., 2000; Eaton, M. J.
fically addressed. Glycine may contribute to the et al., 2002). The analgesic effect of exogenous BDNF
development of hyperalgesia via an intriguing may be attributed to a downregulation of TrkB after
way. As a required co-agonist for NMDA receptor a large dose of the BDNF treatment (Frank, L. et al.,
activation, glycine released from inhibitory inter- 1997; Chen, H. and Weber, A. J., 2004). If BDNF is
neurons in the superficial dorsal horn can escape microinjected into the RVM at a dose within the
736 Descending Control Mechanisms

physiological range (10100 fmol), pain facilitation O. et al., 1996; Urban, M. O. and Gebhart, G. F.,
occurs (Guo, W. et al., 2006). BDNF-containing neu- 1997). Activation of the NT1 receptor on spinally
rons in the PAG project to and release BDNF in the projecting serotonergic neurons in the RVM pro-
RVM. Intra-RVM sequestration of BDNF by TrkB duces antinociception through release of serotonin
IgG fusion protein or anti-BDNF antibody and in the spinal dorsal horn (Buhler, A. V. et al., 2005).
knockdown of TrkB by RNAi attenuates pain after
inflammation (Guo, W. et al., 2006). The effect of 49.2.2.4.(v) Cholecystokinin Dense immunoreac-
BDNF in the RVM is dependent on the activation tive cholecystokinin (CCK) terminals are localized to
of NMDA receptors. BDNF induces tyrosine phos- PAG and RVM neurons (Skinner, K. et al., 1997).
phorylation of the NMDA receptor NR2A subunit However, CCK-containing neurons in the PAG do
via a signal transduction cascade that involves IP3, not appear to project to the NRM (Beitz, A. J. et al.,
PKC, and Src (Guo, W. et al., 2006). Thus, suprasp- 1983). Most studies agree that CCK acts as an anti-
inal BDNFTrkB signaling contributes to net opioid peptide through CCK2 (CCKB) receptors
descending pain facilitation through a mechanism (Han, J. S., 1995; Fields, H. L. and Basbaum, A. I.,
involving NMDA receptor activity. 1999; Veraksits, A. et al., 2003). Cholecystokinin in
the RVM mediates opioid-induced hyperalgesia and
49.2.2.4.(iii) Substance P Although substance P is antinociceptive tolerance (Xie, J. Y. et al., 2005). Intra-
a major neurotransmitter released from primary RVM pretreatment with a selective CCK2, but not
afferents at the spinal level, it is also widely distrib- CCK1 (CCKA), receptor antagonist significantly
uted in supraspinal structures involved in descending potentiates the antihyperalgesic effect of morphine in
pain control (Beitz, A. J., 1982a). A group of PAG a model of visceral nociception (Friedrich, A. E. and
neurons containing both substance P and cholecys- Gebhart, G. F., 2003). Supraspinal CCK may drive
tokinin (CCK) projects to the spinal cord (Skirboll, L. tonic descending facilitation mechanisms to maintain
et al., 1983). However, the function of this descending neuropathic pain in the rat (Kovelowski, C. J. et al.,
pathway is unclear. A significant population of PAG 2000). The cellular mechanisms of the pronociceptive
neurons possesses neurokinin 1 (NK1) receptors, the and anti-opioid actions of CCK involve an action on
primary binding site for substance P, and many NK1- pain modulatory neurons in the RVM (Heinricher, M.
positive PAG neurons also contain glutamate and to M. et al., 2001; Heinricher, M. M. and Neubert, M. J.,
a lesser extent, enkephalin (Commons, K. G. and 2004).
Valentino, R. J., 2002). This raises the possibility
that activation of NK1 in the PAG may be upstream 49.2.2.4.(vi) Acetylcholine The DLPT nuclei and
to glutamatergic activation and affect activity of the RVM regions have been found to send descending
descending circuitry. Both pro- and antinociceptive cholinergic fibers to the spinal cord (Bowker, R. M.
properties have been attributed to the NK1 receptor. et al., 1983; Jones, B. E. et al., 1986). The cholinergic
Holden J. E. et al. (2002) show that antinociception pedunculopontine tegmental nucleus (PPTg) in the
from lateral hypothalamic stimulation is mediated by DLPT, immediately ventral to the cuneiform
NK1 receptors in the A7 cell group in rats. Suzuki R. nucleus, has attracted attention. The PPTg is one of
et al. (2002b) demonstrate that excitation of spinal the two most sensitive sites for intracranial nicotine-
NK1-bearing projection neurons results in input to produced antinociception (Iwamoto, E. T., 1991).
the RVM, which triggers descending pain facilitation. Neurons in the PPTg are responsive to noxious tail
pinch (Carlson, J. D. et al., 2004). Microinjection of
49.2.2.4.(iv) Neurotensin Microdialysis of mor- nicotine into the NRM, the other sensitive site for
phine into the ventromedial PAG induces the effect of nicotine, produces antinociception
neurotensin release (Stiller, C. O. et al., 1997). The (Iwamoto, E. T., 1991). Intra-NRM ABT-594, a cho-
neurotensinergic input from the PAG to the RVM linergic channel modulator, produces analgesia,
has been described (Beitz, A. J., 1982c). Neurotensin presumably through an action on alpha 4-containing
is involved in bidirectional descending modulation nicotinic acetylcholine receptors coexpressed by ser-
from the RVM. Microinjection of low dose (30 fmol) otonergic neurons (Bitner, R. S. et al., 1998). The
of neurotensin into the RVM produces hyperalgesia dorsal raphe nucleus is also a potential site for nico-
and facilitation of dorsal horn neuronal responses to tinic analgesia (Cucchiaro, G. et al., 2005). Spinal
noxious stimuli and high doses of neurotensin cholinergic receptors contribute to descending inhi-
(0.33 nmol) produces antinociception (Urban, M. bition from the Gi/GiA region, although it is unclear
 Descending Control Mechanisms 737

whether the source of acetylcholine transmitters is (Sakurai, T. et al., 1998). A number of studies have
supraspinal (Zhuo, M. and Gebhart, G. F., 1990). At suggested a role of orexin in descending pain mod-
the spinal level, acetylcholine tonically modulate ulation. Hypothalamic descending axons containing
serotonergic transmission (Cordero-Erausquin, M. orexin innervate all levels of the spinal cord includ-
and Changeux, J. P., 2001) and both inhibitory and ing superficial dorsal horn (van den Pol, A. N., 1999).
excitatory neurons are responsive to nicotinic stimu- Orexin A-like immunoreactive fibers in the spinal
lation (Cordero-Erausquin, M. et al., 2004). cord are mostly confined to axon terminals contain-
ing dense-cored vesicles and mainly forming
49.2.2.4.(vii) Histamine Central histamine is asymmetric synapses (Guan, J. L. et al., 2003). Initial
mainly derived from the tuberomammilllary nucleus analysis indicates that orexin-B modulates superficial
of the hypothalamus and histaminergic terminals are dorsal horn neurons in the rat spinal cord. Orexin-B
found in the PAG, LC, RVM, and spinal cord increases spontaneous inhibitory postsynaptic cur-
(Millan, M. J., 2002). Among four classes of histamine rents in the majority of dorsal horn neurons and
receptors (H14), H13 receptors have been localized to also produces excitatory effects on a smaller popula-
the CNS but their roles in descending pain control are tion of dorsal horn neurons (Grudt, T. J. et al., 2002).
unclear. Activation of central H1 receptor induces Orexins produces analgesia through central but not
behavioral hyperalgesia in mice involving the phospho- peripheral mechanisms and do not appear to involve
lipase Cprotein kinase C pathway (Galeotti, N. et al., the endogenous opioids (Mobarakeh, J. I. et al., 2005).
2004). Intracerebroventricular injection of H3 agonist is Activation of spinal orexin-A receptor produces anti-
hyperalgesic in rodents (Malmberg-Aiello, P. et al., allodynic effect in the rat with carrageenan-induced
1994). However, activation of spinal histamine H3 inflammation (Yamamoto, T. et al., 2003). In the pre-
receptors inhibits mechanical nociception (Cannon, pro-orexin (precursor of orexins) knockout mice, the
K. E. et al., 2003). It is clear that the effects of baseline pain threshold is not altered but the knock-
histamine-related agents may be mediated through out mice exhibit enhanced hyperalgesia after
nonhistamine receptors. A histamine-derived com- inflammation and reduced stress-induced analgesia
pound, improgan, produces analgesia exclusively (Watanabe, S. et al., 2005). These results suggest that
through the supraspinal circuitry involving PAG hypothalamic orexins and related descending path-
and RVM through unknown receptor mechanisms ways are activated by persistent pain and stress to
(Hough, L. B. et al., 1997; Nalwalk, J. W. et al., 2004). produce pain inhibition.

49.2.2.4.(viii)Prostaglandin Recent observa- 49.2.2.5 Summary

tions suggest a role of centrally produced The bulbospinal noradrenergic and serotonergic
prostaglandins in descending pain modulation. pathways are the most established systems with
Microinjection of prostaglandin E2 into the medial regard to descending pain modulation. The roles of
preoptic region and PAG regulates pain modulatory other neurotransmitters and receptors in descending
neurons in the RVM, the effects including activation pain control are mainly documented at the segmental
of ON cells and suppression of OFF cells, and pro- levels. The interactions between these transmitters
duces thermal hyperalgesia (Heinricher, M. M. et al., and receptors, including receptor subtypes, are extre-
2004a; 2004b). These findings support the emerging mely extensive and sophisticated (see Chapter
view that the CNS glia-inflammatory cytokine Pharmacological Modulation of Pain) (Millan, M. J.,
network, in association with the cyclooxygenase 2002), although their contributions are discussed
2-prostaglandin cascade, is involved in the develop- individually here. A striking feature of the involve-
ment of hyperalgesia after tissue and nerve injury ment of neurochemicals in descending pain control is
(Samad, T. A. et al., 2001; Watkins, L. R. et al., 2003). that almost every substance has been shown to be
Such pathways may increase descending facilitation involved in both inhibitory and facilitatory pain con-
from the RVM circuitry and produce pain and trol, either through actions at the different receptor
hyperalgesia. subtypes, different levels of the neuraxis, or different
concentrations; and that the same transmitter may
49.2.2.4.(ix) Orexins (hypocretin) The novel neu- produce opposite modulatory effects at different, or
ropeptides orexin A and B are selectively synthesized even the same, segmental levels. This factor contri-
in the lateral and posterior hypothalamus and origin- butes to some conflicting results in this area and
ally identified as a regulator of feeding behavior seriously complicates the interpretation of the
738 Descending Control Mechanisms

findings derived from genetic approaches that exhibit large GABA-mediated synaptic potentials
involve manipulation of specific receptor(s) at the and are insensitive to opioids, while secondary cells
systemic level. have little or no GABAergic input and are hyperpo-
larized by opioids. Primary and secondary are likely
corresponding to OFF and ON cells, respectively,
49.2.3 Neuronal Activity in Descending
based on their pharmacological characteristics, such
Pathways
that local iontophoretic of morphine directly inhibits
49.2.3.1 Pain modulatory neurons ON cells but does not affect OFF cells (Fields, H. L.
By comparing neuronal activity to the occurrence of and Basbaum, A. I., 1999). Excitatory amino acid
nociceptive reflexes in lightly anesthetized rats, transmission is involved in modulating both ON
Fields H. L. et al. (1983a) identified three classes of and OFF cell activity (Heinricher, M. M. et al.,
neurons in the RVM. One type of cell typically shows 1999), which is in line with the evidence that both
a burst of activity immediately prior to the onset of primary and secondary cell types receive excitatory
the tail flick from a noxious thermal stimulus, thus amino acid input. Models have been proposed to
named an ON cell. Another type of cell exhibits a understand the functional significance of neuronal
pause in activity just before the tail flick and is called activity within the RVM circuitry. In the disinhibition
an OFF cell. The activity of the third class of cell, model, opioids directly inhibit ON cells (correspond-
NEUTRAL cell, exhibits no clear relationship to ing to secondary cells) and release the GABAergic
nociceptive reflexes in response to transient stimuli. inhibition of OFF cells (corresponding to primary
The firing profiles of ON and OFF cells suggest that cells), resulting in pain inhibition (Fields, H. L. and
they are pain modulatory neurons since their activity Basbaum, A. I., 1999). RVM neurons are also subject to
is correlated with facilitation and inhibition of noci- modulation by noradrenergic input (Hammond, D. L.
fensive behaviors, respectively. In fact, both RVM ON et al., 1980), which mainly originated from the DLPT
and OFF cells project to the spinal cord, particularly nuclei (Tanaka, M. et al., 1996; Meng, X. W. et al.,
laminae I, II, and V of the dorsal horn (Fields, H. L. 1997). Opposite to the spinal cord, activation of alpha
et al., 1995), and are modulated by PAG stimulation 2 adrenoceptors hyperpolarizes primary cells and
and opioids (Vanegas, H. et al., 1984; Fields, H. L. et al., attenuates opioid analgesia, and activation of alpha 1
1983b; Fields, H. L. et al., 1991). Although less studied, adrenoceptors depolarizes primary cells and enhances
ON and OFF cells are also found elsewhere involved analgesia (Bie, B. et al., 2003).
in pain modulation, including the PAG (Heinricher, The finding of ON and OFF cells provides a
M. M. et al., 1987), cuneiformis nucleus, PB (Haws, cellular mechanism for bidirectional descending pain
C. M. et al., 1989) and nucleus submedius of the thalamus control and favors our understanding of variability of
(Fu, J. J. et al., 2002). It should be clarified that pain (Fields, H. L., 1988). It should be emphasized that
OFF-cell activity may not be solely responsible for the descending facilitation parallels inhibition and is
suppressing nocifensive withdrawals. In the una- an active process. The enhanced nociceptive respon-
nesthetized rat during slow wave sleep, animals are siveness in morphine-dependent rats after treatment
able to execute withdrawal from noxious stimulation with opioid receptor antagonists is accompanied by an
when OFF cell continues to fire (Leung, C. G. and increased RVM ON-cell firing (Bederson, J. B. et al.,
Mason, P., 1999). Simultaneous recordings from 1990). In spite of a well-documented RVMspinal
spinal and RVM neurons show that ON- and serotonergic projection in descending pain control,
OFF-cell firing reflects well input from the spinal Gao K. and Mason P. (2000) found that serotonergic
dorsal horn, suggesting a role in gating ascending RVM cells that respond to noxious tail heat are not
nociceptive information (Hernandez, N. et al., 1989; ON or OFF cells. The transmitters released by axon
Hernandez, N. and Vanegas, H., 2001). In addition terminals of RVM ON and OFF cells in the spinal
to regulation of nocifensive responses, ON- and cord remain to be verified.
OFF-cell activity may also play a role in integrating Pan Z. Z. et al. (1997) show that activation of the
cardiovascular, visceral, and somatosensory input kappa-receptor hyperpolarizes primary cells that are
(Thurston, C. L. and Randich, A., 1995) and regulat- disinhibited through the inhibition of secondary cells
ing homeostatic functions (Mason, P., 2001). by mu-opioid receptors. Thus, mu- and kappa-opioid
In vitro recordings from medullary slices identified receptors may produce opposing actions on RVM pain
two distinct cell types in the NRM, primary and modulatory neurons. This kappa effect reverses the
secondary cells (Pan, Z. Z. et al., 1990). Primary cells analgesic effect of mu-opioids (Bie, B. and Pan, Z. Z.,
 Descending Control Mechanisms 739

2003). Activation of the kappa-opioid receptor in the dorsal horn activity, consistent with Walls hypoth-
RVM has also been shown to produce antinociception esis (Wall, P. D., 1967).
in a test modality-dependent manner. Microinjection of
a kappa-receptor agonist U69593 into the RVM 49.2.3.2.(ii) Inhibitory effect Descending input
increased withdrawal latency of paw, but not the tail, produces inhibition of dorsal horn neurons through
to noxious thermal stimulus (Ackley, M. A. et al., 2001). multiple synaptic mechanisms (Light, A. R. et al.,
The opioid withdrawal-induced hyperalgesia is attenu- 1986; Zhang, D. X. et al., 1991; Fields, H. L. and
ated by intra-RVM kappa-agonist (Bie, B. and Pan, Z. Z., Basbaum, A. I., 1999). As spinal serotonin and nora-
2003). The cellular mechanisms of these differential drenaline are almost exclusively of supraspinal
actions of the kappa-opioid on RVM neurons and origin, they provide postsynaptic control of spinal
behavioral nociception are unclear and appear to neurons. Westlund K. N. et al. (1990) demonstrated
involve a presynaptic inhibition of glutamatergic that spinothalamic neurons in primate receive direct
input to pain modulatory neurons (Ackley, M. A. catecholaminergic innervation. Stimulation of the
et al., 2001; Bie, B. and Pan, Z. Z., 2003) (also see NRM produces direct postsynaptic inhibition of spi-
Chapter The Brainstem and Nociceptive nothalamic tract neurons (Giesler, G. J. Jr. et al., 1981).
Modulation). Descending input can also modulate secondary order
neurons indirectly through action on interneurons.
49.2.3.2 Descending influence on dorsal NRM and spinal stimulation increases glycine and
horn neurons GABA release into the spinal cord, suggesting activa-
49.2.3.2.(i) Tonic effect Wall P. D. (1967) origin- tion of inhibitory interneurons (Sorkin, L. S. et al.,
ally observed that blocking impulses descending from 1993; Cui, J. G. et al., 1997). Since changes in activity
the brainstem induced an increased activity of lami- of interneurons can lead to changes in presynaptic
nae IV/V dorsal horn neurons in the cat. In contrast, inhibition related to primary afferent depolarization,
the response of lamina VI neurons to movement was primary afferent fibers are also indirectly modulated
mainly reduced. This suggests a tonic brainstem des- by descending input (Martin, R. F. et al., 1979).
cending inhibition on cutaneous sensory processing in
the dorsal horn and a facilitatory control over proprio- 49.2.3.2.(iii) Facilitatory effect Supraspinal struc-
ceptive activity. The tonic inhibition of dorsal horn tures also exert facilitatory influences on spinal
neurons is confirmed by later studies (Besson, J. M. nociceptive transmission. Tattersall J. E. et al. (1986)
et al., 1975; Cervero, F. et al., 1976; Dickhaus, H. et al., report that 44% of neurons recorded from the lower
1985; Ren, K. and Dubner, R., 1996). thoracic spinal cord of the cat show reduced response
It is worth noting that if a portion of the descend- to visceral stimulation after reversible spinalization,
ing pathways is blocked or only NRM is suggesting that they are under descending excitatory
anesthetized, dorsal horn nociceptive neurons do control. Excitation and inhibition of dorsal horn neu-
not show an increased activity (Cervero, F. et al., rons are produced by stimulation of the DLF of the
1979; Gebhart, G. F. et al., 1983). Apparently, tonic spinal cord (Dubuisson, D. and Wall, P. D., 1979;
suppression of dorsal horn activity is implemented by McMahon, S. B. and Wall, P. D., 1988), NRM
multiple descending pathways or supraspinal sites; (Dubuisson, D. and Wall, P. D., 1980), and Gi
and the loss of inhibition from one pathway can be (Haber, L. H. et al., 1980). Even microinjection of
compensated by others. morphine into the NRM produces facilitation of
The findings of Wall also reveal the plastic nature C-fiber responses of some dorsal horn nociceptive
of dorsal horn neurons. In lamina IV, many low neurons (Le Bars, D. et al., 1980). Although in most
threshold neurons became wide dynamic range neu- cases, it is unclear whether facilitation occurs on
rons; and in lamina V, the receptive fields of inhibitory interneurons, excitation of which may
nociceptive neurons were expanded during the spinal result in subsequent inhibition, Haber L. H. et al.
cord block. Collins J. G. and Ren K. (1987) showed (1980) demonstrate the Gi stimulation-produced
that, after a low dose of pentobarbital in the intact excitation of primate spinothalamic tract neurons,
awake cat, dorsal horn neurons (originally only indicating descending facilitation of nociceptive trans-
responsive to noxious stimuli) became responsive to mission. Several studies suggest a spinalbrainstem
low-intensity stimuli, that is, the wide dynamic range loop in the modulation of spinal pain processing.
response profile is unmasked. This effect is likely due The activity of this loop involves descending inhibi-
to a release of inhibitory systems that regulate spinal tion or facilitation of dorsal horn neurons (Cervero, F.
740 Descending Control Mechanisms

and Wolstencroft, J. H., 1984; McMahon, S. B. and receptor agents administered at lower doses and inhi-
Wall, P. D., 1988; Suzuki, R. et al., 2002b). Spinal bition is found at higher intensities of stimulation or
5-HT3 receptors mediate descending facilitation of larger doses of the receptor agents, with few excep-
deep dorsal horn neurons, which involves a circuitry tions (Thomas, D. A. et al., 1995; Hammond, D. L.
consisting of ascending NK1-expressing neurons in the et al., 1998). This may explain why descending facil-
superficial dorsal horn and descending fibers from itation was not recognized in many earlier reports
RVM (Suzuki, R. et al., 2002b). because the facilitation might often be masked by
more intense electrical stimulation or higher drug
doses used to produce net descending inhibition. It
49.2.4 Multiplicity of Descending Control
remains to be explained how and why a manipulation
The bidirectional descending modulation of nocicep- at the same site by varying stimulation intensities
tion has now been appreciated. It appears that some would produce opposing effects on nociceptive trans-
sites in the rostral medulla play an essential role in mission. Pain modulating neurons of different
this balanced descending modulation. Depending on modalities intermingle with each other within the
the intensities used and the specific site, stimulation RVM and the membrane properties of ON and
of the Gi and RVM regions produces facilitation (525 OFF cells are very similar (Zagon, A. et al., 1997).
mA) and inhibition (50200 mA) of nocifensive reflexes One can only speculate that the net effect of
and dorsal horn nociceptive neurons (Zhuo, M. and excitation of brainstem nuclei is related to differen-
Gebhart, G. F., 1992; 1997). RVM neurons may exert tial activation of subsets of neurons with different
bidirectional control of nociception through des- connections and different combinations of neuro-
cending serotonergic pathways (Zhuo, M. and transmitter receptors. In the case of BDNF, brain
Gebhart, G. F., 1991) as well as via the A7 catecho- microinjection at a dose within the normal range, deter-
lamine cell group. Enkephalin-containing neurons in mined by measuring its concentration in vivo,
the RVM project to the A7 cell group and microin- produces nociceptive facilitation (Guo, W. et al.,
jection of morphine in the A7 cell group produces 2006), but higher doses lead to antinociception
facilitation and inhibition of spinal nociception (Siuciak, J. A. et al., 1995; Frank, L. et al., 1997; Guo,
mediated by alpha 1- and alpha 2-adrenoceptors W. et al., 2006). This suggests that activation of
(Holden, J. E. et al., 1999). The other site that is BDNFergic transmission at the physiological level
involved in descending facilitation is the subnucleus results in facilitation and inhibition produced by
reticularid dorsalis (SRD) of the caudal medulla larger doses may result from the recruitment of
(Lima, D. and Almeida, A., 2002). Lesioning the excessive synapses that are not normally active
SRD depresses nociceptive responses to acute and simultaneously. We should also be aware that
inflammatory pain, whereas stimulation produces supraspinal descending influences might exert differ-
pain facilitation. The SRD-mediated facilitation ential effects on motor reflex and dorsal horn
requires a reciprocal disynaptic putative excitatory neuronal activity that are used by most animal stu-
circuit that links the SRD and the spinal dorsal horn. dies as the endpoints (Willer, J. C. et al., 1979). Future
It is interesting that both facilitatory and inhibitory studies should validate pain facilitation and inhibi-
pathways can be triggered by activation of visceral tion by including the affective component of pain and
afferents. Under certain circumstances, vagal afferent not merely measure reflex behaviors (Vierck, C. J. Jr.
stimulation at relative low intensities produces facil- et al., 2004; Neubert, J. K. et al., 2005).
itation of the nociceptive tail flick reflex as well as
dorsal horn nociceptive neuronal activity (Randich, A.
and Gebhart, G. F., 1992). The neural relays involved 49.3 Plasticity of Descending
in vagal afferent-produced facilitation and inhibition Pathways
also include sites within the RVM. Thus, spinal
nociceptive processing is subject to bidirectional con- To be able to feel pain is a normal and essential
trol from supraspinal sites so that the intensity of function for living. Pain warns the body against injury
perceived pain can be fine-tuned by descending and illness so that responses will be undertaken to
pathways. protect the body. Normally, pain goes away when the
In general, when biphasic pain modulation is damage is resolved and tissues are healed, and will
observed at the same site, facilitation is produced by not be long-lasting. However, chronic pain develops
electrical stimulation given at lesser intensities or under a variety of conditions even without the
 Descending Control Mechanisms 741

presence of injury, and the suffering from excessive pain inhibition is shown in rats with partial
persistent pain has been a major health problem. sciatic nerve ligation (Azami, J. et al., 2001). In the
Most earlier studies on the descending pain control CCI model, there is an upregulation of spinal CB1
mechanisms have focused on responses to acute or receptors and an increased antihyperalgesic and anti-
transient noxious stimuli, which has built up the allodynic effect of Win 55,212-2, a CB receptor agonist
fundamentals for our understanding of the system. (Lim, G. et al., 2003), although it is unclear whether
Recent studies have turned attention to chronic pain descending input contributes to this enhanced pain
conditions and examined the effects of pain that last inhibition.
for hours, days, or longer, following tissue damage or Intraplantar injection of formalin has been used
nerve injury. Persistent pain conditions are associated extensively as a model of prolonged chemical noci-
with prolonged functional changes in the nervous ception. In rats receiving formalin injection, there is
system, evidenced by the development of dorsal an increase in noradrenaline and serotonin concen-
horn hyperexcitability, or activity-dependent plasti- trations in the spinal dorsal horn. Intrathecal
city, also commonly referred to as central pretreatment with yohimbine, an alpha 2 adrenocep-
sensitization. Evidence has accumulated to indicate tor antagonist, and methysergide, a 5-HT receptor
that descending pathways also exhibit plasticity that antagonist, significantly enhanced nocifensive beha-
contributes to altered pain processing after injury vior (Omote, K. et al., 1998). These results suggest
(Porreca, F. et al., 2002; Ren, K. and Dubner, R., activation of descending monoaminergic pathways
2002; Vanegas, H. and Schaible, H.-G., 2004). by formalin. In the PAG, formalin injection leads to
a decrease in GABA release (Maione, S. et al., 1999). A
decreased GABAergic tone in the PAG presumably
49.3.1 Descending Modulation in
will disinhibit PAG neurons and result in activation
Persistent Pain
of opioid interneurons in the RVM, leading to
49.3.1.1 Responses of descending increased descending inhibition.
pathways to injury In humans with secondary hyperalgesia after
In the rat inflammation model, intraplantar injection heat/capsaicine sensitization, the PAG and the
of carrageenan induces inflammation and hyperalge- cuneiform nucleus activation is observed using
sia that is associated with an increased Fos protein whole-brain functional magnetic resonance imaging
expression, a marker of neuronal activation, in the PB (Zambreanu, L. et al., 2005). These multiple lines of
and LC/SC (Bellavance, L. L. and Beitz, A. J., 1996; evidence indicate that, not only can stimulation of
Buritova, J. et al., 1998; Tsuruoka, M. et al., 2003). In pain modulatory circuitry attenuate hyperalgesia and
adjuvant-induced inflammation and the sciatic nerve dorsal horn neuronal hyperexcitability after inflam-
ligation model, there is a selective increase in mation and nerve injury (Morgan, M. M. et al., 1991;
proneurotensin mRNA expression in the PAG Sotgui, M. L., 1993), descending pathways are also
(Williams, F. G. and Beitz, A. J., 1993). In contrast, activated under persistent pain conditions.
veratridine-induced GABA release is decreased
in the PAG after hind paw inflammation (Renno, W. 49.3.1.2 Descending inhibition in
M. and Beitz, A. J., 1999). Orofacial inflammation persistent pain
activates the coeruleotrigeminal antinociceptive There is now considerable evidence that inflamma-
pathway (Matsutani, K. et al., 2003) and hind paw tory injury induces an enhanced net descending
inflammation induces an increase in NA levels in inhibition at sites of primary hyperalgesia (Ren, K.
the spinal dorsal horn, suggesting activation of des- and Dubner, R., 2002). In cats with knee joint inflam-
cending noradrenergic pathways (Tsuruoka, M. et al., mation, descending inhibition is greater in neurons
1999). with input from the inflamed knee as revealed by
In rats receiving chronic constriction injury of the reversible spinalization with a cold block and DNIC-
sciatic nerve (the CCI model), in addition to the induced inhibition (Schaible H. G. et al., 1991;
sensory structures of the cortex and thalamus, Danziger, N. et al., 1999; 2001). Similar phenomenon
changes in brain neural activity are observed in a is observed in rats with adjuvant-induced hind paw
variety of regions involved in descending control inflammation. Local anesthetic block of the thoracic
including the hypothalamic arcuate nucleus and spinal cord leads to an increased activity of dorsal
central gray matter, LC, PB, and Gi (Mao, J. et al., horn nociceptive neurons that is greater in inflamed
1993). The activation of GiA-mediated descending than that in noninflamed rats (Ren, K. and Dubner, R.,
742 Descending Control Mechanisms

1996). Studies using Fos protein expression as a mar-

ker of neuronal activation have reached a similar Thalamus and cortex
conclusion. There are more inflammation-induced
Fos-expressing neurons in the spinal dorsal horn in Pain
spinally transected or DLF-lesioned rats, when com- modulatory circuitry
Pain
pared to sham-operated inflamed rats (Ren, K. and hyperexcitability
Dubner, R., 2002). Kauppila P. et al. (1998) show that
thermal but not mechanical nociceptive responses are +
further enhanced in hind paw-inflamed and spinal Injury Dorsal horn
nerve-ligated rats after spinalization at the midthor- input hyperexcitability
acic level. Furthermore, behavioral hyperalgesia is
intensified in rats with lesions of the dorsal lateral Figure 3 Diagram illustrating descending pain facilitation
quadrant of the spinal cord after inflammation or for- and inhibition in persistent pain. An enhanced primary afferent
input after injury may lead to dorsal horn hyperexcitability and
malin injection (Abbott, F. V. et al., 1996; Ren, K. and
produce dynamic changes in excitability within brain stem
Dubner, R., 1996). These studies reveal the net des- pain modulatory circuitry. The net effect of descending
cending inhibitory effects after tissue injury. The modulation depends on a balance between inhibitory () and
enhanced descending net inhibition serves to dampen facilitatory () input to the spinal cord. An increase in
injury-induced dorsal horn hyperexcitability and descending net facilitatory modulation may lead to the
development of persistent pain. Adapted from Ren, K., Zhuo,
hyperalgesia.
M., and Willis, W. D. 2000. Multiplicity and Plasticity of
The source of the enhanced inhibition can be Descending Modulation of Nociception: Implications for
traced back to brainstem structures. Local anesthesia Persistent Pain. In: Proceedings of the 9th World Congress on
of the RVM results in a further increase in dorsal Pain (eds. M. Devor, M. C. Rowbotham, and Z. Wiesenfeld-
horn nociceptive neuronal activity in hind paw- Hallin), pp. 387400. IASP Press.
inflamed rats (Ren, K. and Dubner, R., 1996). Focal
lesions of the RVM and LC are followed by an
increase in spinal Fos expression and hyperalgesia inflammation-induced spinal Fos expression (Wei, F.
after inflammation (Tsuruoka, M. and Willis, W. D., et al., 1999). The descending facilitatory effect may
1996; Wei, F. et al., 1999). When lidocaine was micro- also originate from the medullary SRD (Lima, D.
injected into the NRM in adjuvant-inflamed rats, and Almeida, A., 2002) and other brain sites such as
increases in activity were observed in over 75% of the the anterior cingulate cortex (Calejesan, A. A. et al.,
spinal dorsal horn neurons (Ren, K. and Dubner, R., 2000).
1996). Interestingly, there appears to be laminar selec- Descending facilitation significantly contributes
tivity in the effect of RVM serotoninergic and LC to the deveolpment of certain types of persistent
noradrenergic descending pathways. The descending pain, particularly those associated with secondary
serotoninergic and noradrenergic pathways differen- hyperalgesia or nerve injury. Spinalization blocks
tially suppress the responses of spinal neurons, mustard oil-produced secondary allodynia and hyper-
including spinoparabrachial neurons, in the deep and excitability of dorsal horn neurons (Mansikka, H. and
superficial dorsal horn, respectively (Ren, K. et al., Pertovaara, A., 1997). RVM lesions prevent formalin-
2000). It appears that both RVM and LC descending induced nocifensive behavior (Wiertelak, E. P. et al.,
pathways are major sources of enhanced net inhibition 1997) and inhibit secondary hyperalgesia produced by
in inflamed animals. topical application of mustard oil (Urban, M. O. and
Gebhart, G. F., 1999). Masseter muscle inflammation
49.3.1.3 Descending facilitation in produces mechanical hyperalgesia of the overlying
persistent pain skin that is blocked by RVM lesions, and likely
Descending input also exerts a facilitatory effect fol- includes secondary hyperalgesia of the skin and pri-
lowing injury. Emerging evidence indicates that mary hyperalgesia of the muscle (Sugiyo, S. et al.,
descending facilitation not only parallels inhibition 2005). The same phenomenon occurs in models of
but also can be a driving force for the development of neuropathic pain. Suzuki R. et al. (2004a) show that
persistent pain (Porreca, F. et al., 2002; see Figure 3). descending facilitatory control of mechanically
The selective destruction of the Gi with a soma- evoked responses is enhanced in dorsal horn neu-
selective neurotoxin, ibotenic acid, leads to an rones after nerve injury. Local anesthesia of the
attenuation of hyperalgesia and a reduction of PAG or RVM attenuates allodynia in rats receiving
 Descending Control Mechanisms 743

ligation of the two spinal nerves (Pertovaara, A. et al., dominated by the facilitation. After tissue or nerve
1996; Kovelowski, C. J. et al., 2000). The tactile allo- injury, there is an increase in synaptic strength for
dynia and cold hypersensitivity after nerve injury both descending inhibition and facilitation. Both
depends upon activation of bulbospinal descending facilitatory and inhibitory circuitry may be activated
facilitatory pathways (Ossipov, M. H. et al., 2000; by ascending input after injury (Herrero, J. F. and
Urban, M. O. et al., 2003). Microinjection of the Cervero, F., 1996; Gozariu, M. et al., 1998). There is
CCK2 receptor antagonist L365,260 into the RVM an increase in ON- and OFF-cell activity after
reverses tactile allodynia and thermal hyperalgesia inflammation (Miki, K. et al., 2002). Thus, the balance
after L5/L6 spinal nerve ligation and intra-RVM between synaptic excitation and inhibition under
CCK-8 produces allodynia and hyperalgesia different conditions appears important. The Gi
(Kovelowski, C. J. et al., 2000). These findings indicate plays a role in descending facilitation of nociceptive
that CCK may drive descending facilitation to main- transmission after transient noxious stimuli (Zhuo, M.
tain neuropathic pain in rats, which is consistent with and Gebhart, G. F., 1992). Lesions of the Gi produce
the anti-opioid action of CCK in RVM. In nerve- an attenuation of hyperalgesia and spinal Fos
injured rats, lesions of the DLF, local anesthetic block expression after inflammation (Wei, F. et al., 1999).
of the RVM, and lesions of RVM mu-opioid receptor However, combined Gi and NRM lesions reverse
expressing cells do not prevent the onset, but reverse the opposite Gi or NRM lesion-induced effects.
the later maintenance of tactile and thermal hyper- It is possible that severe persistent pain may be
sensitivity (Porreca, F. et al., 2001; Burgess, S. E. et al., enhanced when the facilitatory network overrides
2002). These observations point to an ascending the inhibition. In cases of neuropathic pain and
descending loop that is activated in response to pro- secondary hyperalgesia, descending facilitation pro-
longed stimulation to facilitate spinal nociceptive vides a driving force for the maintenance of
processing. The ascending dorsal column pathways persistent pain (also see Chapter Neuropathic Pain:
likely contribute to this pain facilitatory circuitry in Basic Mechanisms (Animal)).
neuropathic pain models (Miki, K. et al., 2000). There
is no evidence of activation of the dorsal column
49.3.2 Dynamics of Descending Pain
pathway in models of inflammatory primary and
Modulation
secondary hyperalgesia.
49.3.2.1 Time-dependent shift in
49.3.1.4 Descending facilitation versus descending control
inhibition There is a gradual build-up of descending inhibition
Clearly, descending modulation of persistent pain in the early phase of tissue inflammation (Schaible, H.
involves both inhibition and facilitation. Although G. et al., 1991; Ren, K. and Dubner, R., 1996; Hurley, R.
there may exist selective circuitry that is responsible W. and Hammond, D. L., 2000). The difference in
for multiple phases of modulation, it is rather surpris- inflammation-induced Fos expression between trans-
ing that the facilitatory sites in the rostral medulla ected and sham controls is clear at 3 days but not
generally overlap with the sites that also produce apparent shortly (2 h) after inflammation (Ren, K.
inhibition. The inhibition and facilitation may also and Ruda, M. A., 1996). In early stages of inflamma-
share some synaptic mechanisms with subtle differ- tion, the dynamic response is similar in both the
ence in sensitivity. Although many of the above cited spinally intact and transected rats, suggesting that
studies have concluded that the hyperalgesia is descending inhibitory inputs play less of a role in
dependent completely on facilitatory influences the early response to peripheral inflammation. In
from the brainstem, it should be noted that the contrast, at 3 days after inflammation, descending
same effects could be produced by a reduction in inhibition is enhanced to counteract continuous nox-
descending inhibition leading to a dominance of des- ious input. The results from electrophysiological
cending facilitation. The serotonin concentration in studies also suggest a progressive enhancement of
the NRM is reduced in mice with diabetic neuropa- descending inhibition of dorsal horn neurons during
thy and sciatic nerve ligation, which is associated the development of inflammation (Schaible, H. G.
with a reduced opioid analgesia (Sounvoravong, S. et al., 1991). An effect of Freunds adjuvant on the
et al., 2004). Thus, neuropathic hyperalgesia may be contralateral spinal cord may also be subject to the
dependent on a net descending facilitatory effect inhibitory control of descending inputs as suggested
wherein a simultaneous descending inhibition is by a clear contralateral increase in Fos-positive cells
744 Descending Control Mechanisms

in transected rats after inflammation (Ren, K. and (a)

ES
Ruda, M. A., 1996). Williams F. G. and Beitz A. J. Drugs R
(1993) demonstrate that after peripheral inflamma-
tion or nerve injury, the increase in neurotensin
RVM Medulla
mRNA expression was confined to the ventromedial
PAG and the dorsal raphe nucleus initially and
spreads to the structures neighboring the PAG
including the cuneiform nucleus and the PPTg as
the injury persists. The enhanced descending inhibi-
tion in inflamed animals has been shown to subside in Noxious input
the late stage at 2 weeks (Guan, Y. et al., 2003). In the DH
ankle monoarthritis rat model, tonic descending inhi- Spinal cord
bition of convergent neurons with input from the
inflamed ankle was enhanced during the acute stage Nocifensive
response
(12 d) and then decreased in the chronic stage (34
weeks) of monoarthritis (Danziger, N. et al., 1999;
2001).
(b) 1 h-CFA
The dynamic changes in descending inhibition
after inflammation can be examined over time by
monitoring antinocifensive responses in lightly
24 h-CFA
anesthetized rats during RVM stimulation Response latency (s)
(Figure 4). Terayama R. et al. (2000) directly com-
pared the potency of descending inhibition during
the development of inflammation in the lightly 3 h-CFA
anesthetized rat preparation. Following a unilateral
hind paw inflammation, the stimulus-response func-
tion (S-R) curve for the inflamed paw was initially
shifted to the right of the noninflamed paw at 3 h and
then gradually shifted to the left and reached the
maximal potency at 24 h after inflammation RVM stimulus intensity (A)
(Figure 4(b)). These findings indicate that inflamma- Figure 4 Time- and stimulus intensity-dependent
tion induces dramatic changes in the excitability of changes in RVM electrical stimulation-produced
RVM pain-modulating circuitry. Early (up to 3 h) in suppression of nocifensive responses. (a) Schematic
the development of inflammation there is an representation of the experimental setup. In lightly
anesthetized animals, nocifensive responses such as paw
increased descending facilitation also shown by
withdrawal can be evoked by noxious stimulus. Electrical
Urban M. O. and Gebhart G. F. (1999), which reduces stimulation (ES) or drugs can be delivered through a guide
the net effect of the inhibition. Over time, the level of cannula placed in the RVM. Neuronal activity can be
descending inhibition increases, or descending facil- recorded (R) in this preparation (see Figure 5). (b) The
itation decreases, leading to a net enhancement of nocifensive response latency is plotted against the RVM ES
intensity. The stimulus-response function (S-R) curve
antinocifensive behavior. Direct stimulation of the
illustrates intensity-dependent inhibition of nocifensive
DLF that bypasses brainstem synaptic mechanisms responses, as indicated by an increase in response latency.
does not produce a change in excitability, indicating Animals were inflamed with complete Freunds adjuvant
that the changes are due to supraspinal mechanisms (CFA), an inflammatory agent. The S-R curve at 1 h post-
at the level of the RVM or higher. CFA is located in the middle. At 3 h post-CFA, the curve
shifted to the right, suggesting a decrease in
Thus the activity of descending pathways exhibits
descending inhibition at this time. At 24 h post-CFA, the S-R
considerable plasticity. Brainstem descending path- curve shifted to the left, suggesting an enhanced
ways become progressively more involved in descending inhibition. These results demonstrate dynamic
suppressing incoming nociceptive signals in primary changes in descending pain control during the
hyperalgesic zones following inflammation. Injury- development of inflammatory hyperalgesia. Adapted from
Terayama, R., Guan, Y., Dubner, R., and Ren, K. 2000.
related, persistent primary afferent input is most
Activity-induced plasticity in brain stem pain
likely responsible for triggering this ascendingdes- modulatory circuitry after inflammation. Neuroreport 11,
cending feedback circuit. 19151919.
 Descending Control Mechanisms 745

49.3.2.2 Phenotypic changes in pain NEUTRAL cell responds in an ON or OFF fashion

modulatory neurons to stimuli applied to the craniofacial region, although
The time-dependent plasticity in descending pain it is nonresponsive to noxious stimulation of the tail.
modulatory circuitry involves changes in the The switch in the response profile of RVM neu-
response profiles of RVM neurons. Montagne- rons correlates with the temporal changes in
Clavel J. and Oliveras J. L. (1994) have shown excitability in the RVM after inflammation. This
changes in RVM neuronal properties after inflamma- phenotypic change of RVM neurons is verified in a
tion in the awake, freely moving rat, which suggests population study that shows a significant increase in
an increase in the population of neurons involved in the percentage of ON- and OFF-like cells, and a
pain modulatory activity. Using paw withdrawal as a decrease in the NEUTRAL-like cell population
behavioral correlate to assess the relationship 24 h after inflammation as compared to control ani-
between nocifensive behavior and RVM neural mals (Miki, K. et al., 2002). The studies of RVM
activity, ON-, OFF-, and NEUTRAL-like cells can neurons support the view that enhanced descending
be identified (Miki, K. et al., 2002). Interestingly, modulation after inflammation involves both facilita-
some NEUTRAL-like cells change their response tion and inhibition since there are changes in the
profile and can be reclassified as ON- or OFF-like responses of both ON and OFF cells (Miki, K. et al.,
cells through continuous recordings during the 2002; de Novellis, V. et al., 2005). After inflammation,
development of inflammation (Figure 5). These find- there is a greater increase in ON-like cell responses
ings suggest that NEUTRAL cells have potential to before the onset of paw withdrawal as compared to
become involved in pain modulation after injury. that in nave controls. ON-cell activity is associated
Another study by Schnell C. et al. (2002) shows that with facilitation of nocifensive behavior (Fields, H. L.
rat RVM neurons may display different properties and Basbaum, A. I., 1999). In contrast, OFF-like cell
depending on the site of peripheral stimulation. A responses are reduced after inflammation indicated

(a) (b) (c) (d)

10 Hz
5s

48 C 48 C 48 C 48 C

PW PW PW PW
Figure 5 Examples of phenotypic changes of RVM neurons following inflammation. (a) Neutral-like cell, 9 h post-CFA. (b)
Panels illustrate that at 14.5 h post-CFA, the same neuron shown in A exhibited a reduction of activity immediately before the
paw withdrawal, characteristic of an OFF-like cell. (c) Neutral-like cell, 2 h post-CFA. (d) The same cell shown in (c) became an
ON-like cell at 5.5 h post-CFA when the cell showed a burst of activity immediately before the paw withdrawal. All histograms
were averaged from three repeated trials (200 ms bin width). The histograms are time-locked to the thermal stimuli (upper row)
or the onset of paw withdrawal (lower row), respectively. Small arrowheads indicate the onset of paw withdrawal s (upper row)
or onset of thermal stimuli (48  C, lower row) for individual trials, respectively. Note that the changes in neuronal activity are
best correlated with the paw withdrawal response. Adapted from Miki, K., Zhou, Q. Q., Guo, W., Guan, Y., Terayama, R.,
Dubner, R., and Ren, K. 2002. Changes in gene expression and neuronal phenotype in brain stem pain modulatory circuitry
after inflammation. J. Neurophysiol. 87, 750760.
746 Descending Control Mechanisms

by a less reduction in neuronal activity after the circuitry after injury. Williams F. G. and Beitz A.
noxious stimulus, and a lack of a complete pause. J. (1993) show that adjuvant-induced hindpaw
The pause in OFF-cell activity is associated with inflammation increased neurotensin mRNA expres-
disinhibition; the lack of the pause and the less reduc- sion in the PAG and the lateral tegmental nuclei,
tion in neuronal activity suggest an increased from which neurotensinergic neurons project to the
inhibition of nocifensive behavior. RVM. The increased sensitivity of EAA receptors
in the descending circuitry during the development
49.3.2.3 Involvement of excitatory amino of inflammatory hyperalgesia is related to transcrip-
acids and receptors tional and translational modulation of the receptors
What are the cellular mechanisms that underlie these (Ren, K. and Dubner, R., 2002). Examination of the
changes? Excitatory amino acids (EAAs) have been mRNA expression of the NR1, NR2A, and NR2B
shown to mediate descending modulation in response subunits of the NMDA receptor in the RVM
to transient noxious stimulation and early inflamma- reveals an upregulation that parallels the time
tion (Urban, M. O. and Gebhart, G. F., 1999; course of the RVM excitability changes. This is
Heinricher, M. M. et al., 1999), and they are involved accompanied by an increase in NMDA receptor
in the development of RVM excitability associated protein. Protein phosphorylation is a major
with inflammatory hyperalgesia (Terayama, R. et al., mechanism for regulation of receptor function.
2000; Guan, Y. et al., 2002; Miki, K. et al., 2002). The native NMDA receptor is likely a tetramer
Microinjection of NMDA into the RVM produces that consists of two NR1 and two NR2 subunits.
effects that are dependent upon the postinflamma- Phosphorylation of multiple sites in the cytoplas-
tory time period. At 3 h post inflammation, low doses mic C-termini of the NR1 and NR2 subunits,
of NMDA produce facilitation of the response to
including tyrosine, serine, and threonine residues,
noxious heat of the inflamed hind paw and the non-
is known to modulate NMDA receptor activity and
inflamed hind paw and tail, supporting findings that
affect synaptic transmission. There was an increase
descending facilitatory effects are NMDA receptor
in NR2A, but not NR2B, tyrosine phosphorylation
dependent and occur early after inflammation
in the RVM after inflammation (Guo, W. et al.,
(Urban, M. O. and Gebhart, G. F., 1999). Higher
2006). In contrast, inflammation induces an increase
doses of NMDA at 3 h post inflammation only pro-
in NR2B, but not NR2A subunit tyrosine phos-
duce inhibition. At 24 h post inflammation, NMDA
phorylation in the spinal cord. A time-dependent
produces only inhibition. All of these effects are
blocked by administration of NMDA receptor increase in NR1 serine phosphorylation has also
antagonists. AMPA, a selective AMPA receptor ago- been identified in the RVM after inflammation.
nist, produces dose- and time-dependent inhibition There is also an increase in the AMPA receptor
that is significantly greater at 24 h compared to that at GluR1 subunit levels in the RVM post inflammation
3 h post inflammation. There is a leftward shift of the (Guan, Y. et al., 2003). AMPA receptor subunits
dose-response curves of NMDA- and AMPA-pro- (GluR14) exist in the two flip and flop isoforms
duced inhibition at 24 h post inflammation as that differentially affect the desensitization proper-
compared to 3 h. The leftward shift of the dose- ties of the receptor. Reverse transcription polymerase
response curves of EAA receptor agonists parallels chain reaction analysis indicates that inflammation
the time-dependent enhancement of net descending induces a significant upregulation of mRNAs encod-
inhibition produced by RVM electrical stimulation, ing the GluR1-flip (524 h post inflammation),
which is also attenuated by NMDA receptor antago- GluR2-flip (24 h post inflammation) and GluR2-
nists (Terayama, R. et al., 2000). These results suggest flop (24 h post inflammation) isoforms in the RVM,
that the enhanced EAA neurotransmission contri- whereas the levels of GluR1-1 flop mRNAs do not
butes to the time-dependent functional changes in exhibit significant changes (Guan, Y. et al., 2003).
descending modulation. GluR1 serine 831 phosphoprotein levels are also
increased as early as 30 min after inflammation. The
49.3.2.4 Molecular mechanisms of increase in GluR1 phosphorylation depends on pri-
activity-dependent plasticity in mary afferent input and interestingly, is blocked by
descending pathways NMDA receptor antagonists, suggesting an NMDA
Correlate changes in gene transcription and protein receptor-mediated modification of AMPA receptor
translation have been found in the descending function in the RVM circuitry.
 Descending Control Mechanisms 747

These findings indicate that EAA receptors in the pathways from the RVM and an increase in CCK
RVM undergo selective transcriptional, translational, activity may contribute to hyperalgesia after pro-
and posttranslational modulation following inflamma- longed morphine exposure (Ossipov, M. H. et al.,
tion. The activity-induced plasticity in descending 2005).
circuitry complements the hyperexcitability in ascend-
ing pain transmission pathways (Dubner, R. and Ruda, 49.3.2.6 Aging-related alteration in
M. A., 1992). The increased neuronal barrage at the descending pain modulation
spinal level activates spinal projection neurons that Iwata K. et al. (2002) have reported age-related plas-
then activate glutamatergic, opioidergic, and presum- ticity in descending pathways in rats. It is found that,
ably GABAergic neurons at the brainstem level leading in aged rats, there is an increased dorsal horn noci-
to a similar but not necessarily identical form of neu- ceptive neuronal activity and nocifensive behavior
ronal plasticity. associated with a significant loss of serotonergic and
noradrenergic fibers in the spinal dorsal horn. These
49.3.2.5 Changes in sensitivity to results suggest a reduced descending inhibition with
opioids advancing age. Human studies concur that the
49.3.2.5.(i) Increased analgesic potency Rats DNIC, a form of endogenous pain inhibition, is
with inflammatory hyperalgesia exhibit an increased reduced in the older adults (Edwards, R. R. et al.,
sensitivity to opioid analgesics (Neil, A. et al., 1986). 2003).
Typically, there is a leftward shift of the dose-
response curve for opioids from the inflamed hyper-
49.3.3 Significance of Altered Descending
algesic paw when compared to the noninflamed paw
Modulation in Persistent Pain
(Hylden, J. L. K. et al., 1991). Kayser V. et al. (1991)
suggest that this increased opioid sensitivity in Descending pathways modulate the intensity of per-
inflamed animals is related to a peripheral mechan- ceived pain under normal conditions and persistent
ism as it is significantly attenuated after local pain as a result of injury. The activity-dependent
injections of very low doses (0.51 mg) of naloxone. plasticity within the pain modulatory circuitry is a
Ossipov M. H. et al. (1995) demonstrate that the normal function of the brain and presumably is acti-
increased morphine analgesic potency is mediated vated to protect the organism from further
through delta opioid receptors. This delta-mediated environmental injury. The dynamic changes in des-
effect may require mu-opioid receptor-induced traf- cending modulation after inflammation and primary
ficking of delta-receptors to neuronal membranes hyperalgesia and allodynia are protective. The early
(Morinville, A. et al., 2003). The increased opioid facilitation may function to enhance nocifensive
sensitivity after inflammation may also be explained escape behavior whereas the dominant late inhibition
by changes in central pain modulating pathways. may provide a mechanism by which movement of the
Hurley R. W. and Hammond D. L. (2000; 2001) injured site is suppressed or reduced to aid in healing
demonstrate enhancement and plasticity of the des- and recuperation. Gebhart G. F. (2004) proposes that
cending inhibitory effects of mu and delta 2 opioid the need to escape from a predator requires enhanced
receptor agonists microinjected into the RVM during control of pain and thus more descending inhibition,
the development and maintenance of inflammatory which is supported by opioid and nonopioid mechan-
hyperalgesia. isms of stress-induced analgesia seen in animals
(Hayes, R. L. et al., 1978). However, the significance
49.3.2.5.(ii) Opioid tolerance and hyperalgesia of the enhanced descending facilitation found at sites
Hyperalgesia may occur after long exposure to of secondary hyperalgesia and after nerve injury is
opioids as opioid tolerance develops (Mao, J., 2002). unclear. Gebhart G. F. (2004) hypothesizes that des-
The underlying mechanisms may involve activity of cending facilitation is necessary to maintain
the pain modulatory circuitry. Morphine-tolerant hyperalgesia as the tissue heals and to protect the
rats exhibit an increased glutamate synaptic transmis- injured tissue from further insult. The enhanced des-
sion in RVM neurons (Bie, B. and Pan, Z. Z., 2005). cending pain modulation clearly includes shifts in the
Chronic morphine treatment induces delta receptor- balance between inhibitory and facilitatory compo-
mediated synaptic inhibition in PAG neurons that nents. Present evidence suggests that there is a
is not seen in the untreated mice (Hack, S. P. et al., different balance in neural networks receiving input
2005). The activation of descending pain facilitatory from zones of secondary hyperalgesia where there is
748 Descending Control Mechanisms

no primary injury. The balance toward facilitatory considered on patients suffering from intractable
influences appears to be maintained for longer peri- pain refractory to other treatments.
ods. Activation of these sites would lead to an
enhancement of movement behavior that could also 49.4.1.2 Spinal cord stimulation
be protective. Effective pain control can be achieved by direct
Injury to neural tissues will also upset the balance spinal cord stimulation (SCS) through an implanted
between facilitation and inhibition. The dynamic stimulator in the epidural space (Shealy, C. N. et al.,
plasticity of descending pathways after peripheral 1967; Janfaza, D. R. et al., 1998). In theory, spinal
nerve injury leading to neuropathic pain may render stimulation can bypass brainstem circuitry to directly
the system vulnerable and lead to pathological activate descending fibers or antidromically excite
consequences. In this situation, the initiation of collaterals of ascending fibers that indirectly inhibit
hyperalgesia is not dependent upon descending facil- projection neurons, leading to inhibition of pain
itation, whereas the maintenance of the hyperalgesia transmission at the spinal level. However, the
for long periods of time is dependent on such des- mechanisms of SCS-produced pain relief are more
cending facilitation. Nerve injury may activate a complex (Meyerson, B. A. and Linderoth, B., 2000).
descending nociceptive system that normally is pro- One interesting aspect is that SCS is selectively
tective but can become a source of persistent pain effective against neuropathic pain and hyperalgesia,
after pathology in the nervous system. leaving normal nociceptive threshold unaffected
(Meyerson, B. A. and Linderoth, B., 2000).

49.4.1.3 Vagal stimulation

49.4 Therapeutic Implications: Animal studies have provided evidence that stimula-
Activating Descending Pathways tion of vagal afferent fibers produces antinociception,
although low intensity may facilitate pain (Randich, A.
The mechanisms underlying various clinical and Gebhart, G. F., 1992). Taking advantage of the
approaches for analgesia and pain control involve approved vagal stimulation approach for epilepsy, the
activation of descending pathways. One important effect of vagal stimulation on pain has been con-
area of research is to find efficient and selective firmed in humans (Kirchner, A. et al., 2000; Ness, T.
ways to activate endogenous descending pathways J. et al., 2000). Vagal stimulation may offer another
for pain control. alternative method for activating the bodys pain
control system.
49.4.1 Electrical Stimulation
49.4.1.4 Transcutaneous electrical nerve
Electrical stimulation has been applied to the all stimulation
levels of the nervous system to produce analgesia or Peripheral stimulation can be applied through trans-
pain attenuation. Presumably, this approach activates cutaneous electrical nerve stimulation (TENS), an
endogenous pain control including the descending alternative modality for pain relief (Shealy, C. N.,
pathways to the spinal cord. However, the underlying 2003). Although the results have been controversial,
mechanisms for the stimulation-produced analgesia, the clinical efficacy of this approach has been demon-
particularly the selectivity of the stimulation against strated by many studies. The effect of TENS is likely
certain types of chronic pain in humans, remain to be produced through modulation of the intrinsic spinal
elucidated. pain processing and release of endogenous opioids
(Sluka, K. A. and Walsh, D., 2003).
49.4.1.1 Brain stimulation
Stimulation-produced analgesia has been confirmed
49.4.2 Pharmacological Approaches
in humans (Kumar, K. et al., 1997; Wallace, B. A. et al.,
2004). For pain control, the most commonly used 49.4.2.1 Opioids
sites for deep brain stimulation have been the peri- Opioid agents have been used extensively for analge-
ventricular gray and PAG regions, which is sia. Even applied peripherally, a central action
consistent with findings in animal studies that these involving activation of the endogenous pain control
areas are involved in opioid analgesia and descending system is required for the analgesic effect of the
pain inhibition. Practically, this approach is only opioids. The opioids are potent and efficacious
 Descending Control Mechanisms 749

analgesics. However, their side effects have limited induces antinociception involving the amygdala, PAG,
their use in chronic pain conditions. and RVM (Helmstetter, F. J. and Tershner, S. A., 1994).
The relative painless state in the injured athlete or the
49.4.2.2 Alpha 2 adrenoceptor agonists wounded soldiers may be explained by stress-induced
Extensive documentation of the role of descending pain control related to a particular setting.
noradrenergic pathways in pain inhibition provides
the bases for the use of 2 adrenoceptor agonists for 49.4.4.2 Placebo
analgesic purpose. Epidural clonidine for cancer pain Placebo analgesia may be mediated by endogenous
is the approved application (Kamibayashi, T. and opioids (Levine, J. D. et al., 1978; Benedetti, F. and
Maze, M., 2000). The use of the 2 agonist can Amanzio, M., 1997). The imaging studies show that
reduce the dose of fentanyl because of their synergis- the neural substrate related to placebo effect on pain
tic action. Intrathecal clonidine has been shown to lies in the prefrontal and dorsal anterior cingulate cortex
improve the effect of SCS (Schechtmann, G. et al., (Lieberman, M. D. et al., 2004). It appears that expecta-
2004). tion is a factor for triggering the placebo response (see
Chapter The Placebo Effect) (Benedetti, F. et al., 2003).
49.4.3 Peripheral Stimulation
49.4.3.1 Diffuse noxious inhibitory 49.5 Concluding Remarks
controls
Heterotopic noxious stimulation produces analgesia Tremendous progress has been made in the past four
in humans (Reinert, A. et al., 2000), which can be decades on descending pain control mechanisms. In
understood by the DNIC hypothesis proposed by addition to well-established descending pain inhibi-
Le Bars and colleagues (Le Bars, D. et al., 1979a; tion, the demonstration of descending pain
1979b). DNIC provides a rationale for understanding facilitation provides a more complete picture of the
counterirritation-produced hypoalgesia and help in bodys endogenous pain modulatory system. The
the design of nonpharmacological approaches for simultaneous descending excitatory and inhibitory
pain relief (see Chapter Diffuse Noxious Inhibitory inputs contribute to a balance between perception
Controls (DNIC)). Additionally, non-noxious and suppression of noxious events. We now appreci-
manipulations also provide pain relief (Simkin, P. P. ate that the pain modulatory system undergoes
and O9Hara, M., 2002), presumably through modula- dramatic changes after injury. There is a concurrent
tion of pain processing by interfering stimuli. activation of brainstem descending inhibitory and
facilitatory pathways and the interaction between
49.4.3.2 Acupuncture these pathways will dictate, or affect, the develop-
Acupuncture and moxibustion have been used for ment of neuronal hyperexcitability and behavioral
pain relief for thousands of years. The analgesic hyperalgesia. The dynamic plasticity of descending
effect of acupuncture is related to the recruitment pathways may sometimes render the system vulner-
of endogenous opioids (Han, J. S., 2004; Zhang, R. X. able and lead to pathological consequences. The
et al., 2004) and pain modulatory circuitry (Ma, F. imbalance between these modulatory pathways may
et al., 2004). Since a typical needling during the acu- be one mechanism underlying variability in acute
puncture treatment does not induce pain in humans, and chronic pain conditions. For patients suffering
the mechanisms of acupuncture analgesia are likely from deep pains such as temporomandibular disor-
different from that of DNIC. ders, fibromyalgia, and low back pain, the diffuse
nature and amplification of persistent pain, in part,
may be the result of a net increase in endogenous
49.4.4 Psychological Factors
facilitation.
49.4.4.1 Stress
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 50 Diffuse Noxious Inhibitory Controls (DNIC)
D Le Bars, INSERM U-713, Paris, France
J C Willer, INSERM U-731, Paris, France
2009 Elsevier Inc. All rights reserved.

50.1 Introduction 764

50.2 A Spinally Mediated Process 764
50.3 A Spinally Mediated Process Involving Supraspinal Structures 768
50.4 A Model in the Rat 769
50.5 The Role of Wide-Dynamic-Range Neurons 769
50.6 Summary and Conclusions 772
References 772

Glossary
allodynia Pain caused by stimuli that would not Hoffmann reflex (H reflex) Monosynaptic reflex
normally cause pain (i.e., non-noxious stimuli). from the calf muscles elicited by electrical stimula-
basic somesthetic activity Description of the tion of the sciatic nerve.
ongoing activity in somatosensory pathways in the inhibitory receptive field Area of the body (sur-
absence of any deliberate stimuli but including sti- face or interior) which when stimulated will produce
muli provided by the environment. inhibitory effects on the activity of a given neuron.
blink reflex Exteroceptive reflex of the orbicularis jaw-opening reflex Exteroceptive reflex pro-
oculi muscles usually evoked by stimuli such as a duced by stimulation of facial or intraoral afferents
mechanical tap on the glabella or by electrical cur- and involving activation of jaw opening muscles
rents delivered to the skin in the periorbital region. (e.g., the digastric) and/or inhibition of activity in jaw
Brown-Sequard syndrome Patient with a hemi- closing muscles (e.g., the masseter).
section of the spinal cord of traumatic origin. jerk reflex Myotatic monosynaptic reflex usually
bulbospinal control A neural control mechanism elicited by a percussion of a muscles tendon using
originating in the brainstem and projecting to the a reflex testing hammer.
spinal cord. Lasegues maneuver Elevation of a lower limb in
dermatome Area of skin innervated by nerves an extended position on a patient lying on a bed.
from a given segment of the spinal cord. This technique explores the Lasegues sign which
descending inhibition Inhibition of spinal cord is the painful limitation of the angle of elevation in
function produced by pathways originating in the brain. cases of radicular compression of the sciatic
diffuse noxious inhibitory controls (DNICs) Neural nerve.
controls triggered by noxious stimulation of microelectrophoretic application Application of
widespread areas of the body which exert an inhibi- a substance in its ionic form from a microelectrode
tory influence on wide-dynamic-range (WDR) by the application of electrical current.
neurons. monoarthritis Inflammation of a single joint.
electromyography (EMG) All electrophysiological mononeuropathy A disturbance of function or
methods for exploring the physiology and patho- pathological changes in a single nerve.
physiology of peripheral nerves and muscles. nociceptive stimulus Stimulus that activates
excitatory receptive field Area of the body (sur- nociceptive afferents (which could be a noxious
face or interior) which when stimulated will produce stimulus or an electrical stimulus).
excitation of a given neuron. noxious stimulus Stimulus that causes or threa-
hemianalgesia Loss of pain sensation from one tens to cause tissue damage.
half of the body. polyarthritis Inflammation of several joints.
heterotopic Part of the body remote from the area of propriospinal mechanism Neural mechanism
interest (e.g., the excitatory receptive field of a neuron). mediated entirely within the spinal cord.

763
764 Diffuse Noxious Inhibitory Controls (DNIC)

RIII reflex Electromyographic response elicited by hypoalgesia by electrically stimulating peripheral

electrical painful stimulation of the (purely cuta- nerves through the skin (i.e., with electrodes placed
neous) sural or ulnar nerve by recording from the on the skin).
biceps femoris (lower limb) or biceps brachialis trigeminal nucleus caudalis Most caudal part of
(upper limb), respectively. So-named because it the trigeminal sensory nuclear complex (or of the
involves activation of group III (A) afferents. trigeminal spinal nucleus); also sometimes called
subnucleus reticularis dorsalis (SRD) Brainstem the medullary dorsal horn.
nucleus located ventral to the cuneate nucleus, ventrolateral quadrant Part of the spinal cord
between trigeminal nucleus caudalis and the where spinoreticular and spinothalamic fibers
nucleus of the solitary tract. travel.
supraspinal mechanism Neural mechanism that Wallenbergs syndrome Patient with a unilateral
involves structures above the spinal cord. lesion of the retro-olivary part of the brainstem
tetraplegic Patient with a complete transection of resulting from the ischemia of a posterior cerebellar
traumatic origin at a high level of the spinal cord artery.
thus affecting all four limbs. wide-dynamic-range (WDR) neurons Neurons of
transcutaneous electrical nerve stimulation the dorsal horn activated by both noxious and non-
(TENS) Technique for inducing analgesia/ noxious stimuli.

50.1 Introduction are involved in mediating both pain and nociceptive

reflexes, DNIC can be studied at three related end-
Painful stimuli can diminish or even mask pain eli- points: spinal neuronal activities, reflexes, and
cited by stimulation of a remote (extrasegmental) sensations. The first two are accessible in animals
part of the body (see references in Melzack, R., while the last two can be studied in human beings
1989; Le Bars, D. and Willer, J. C., 2002). This phe- (Figure 1).
nomenon has been known since ancient times as
illustrated by the Hippocrates aphorism: If a patient
be subject to two pains arising in different parts of the
50.2 A Spinally Mediated Process
body simultaneously, the stronger blunts the other.
Numerous popular therapeutic methods for the alle-
Combined psychophysical measurements and
viation of pain some used spontaneously by
recordings of nociceptive reflexes in man have
patients take advantage of this common clinical shown that the spinal cord is involved in the phe-
observation. It is often referred to as counter-irrita- nomenon of pain inhibiting pain (Willer, J. C. et al.,
tion or counter-stimulation. In Kabylia, for example, 1984). Electrical stimulation of the sural nerve at the
healers treat rheumatic pain by placing a red-hot ankle simultaneously induces a nociceptive reflex in
scythe close to the abdomen of the patient and then a knee flexor muscle (the RIII reflex) and a pinprick
vibrating it at a frequency of about 3 Hz, in order to type of painful sensation in the territory of the nerve
create a series of acute burning-type painful (Willer, J. C., 1977). Both the reflex and the sensation
sensations. have strong relationships with the stimulus intensi-
A working hypothesis was developed that some of ties with the thresholds for both parameters being
the neurons involved in the transmission of nocicep- practically identical.
tive signals might be inhibited by nociceptive One line of study involved recording these para-
stimulation of peripheral territories outside their meters (i.e., the reflex and pain) before, during, and
own excitatory receptive fields. Such a hypothesis after the application of heterotopic conditioning sti-
was found to be correct at as early a stage in the muli such as the immersion of a hand in a
processing of nociceptive signals as the spinal cord. thermoregulated water bath at various temperatures
This phenomenon was termed diffuse noxious inhi- (Figure 2, upper panels). When the temperature of
bitory control (DNIC). Since these spinal neurons the bath was lower than 45  C, the immersion of the
 Diffuse Noxious Inhibitory Controls (DNIC) 765

Pain

(c)
Pain

V
Bulbar
reticular Nociceptive
formation conditioning
stimulus

DNIC

DLF DLF

A and C
fibers
Conditioned
stimulus
Ventrolateral
quadrant
(a)
Neuronal activity

Dorsal root

+
Motoneurones
(b)
Ventrolateral
Electromyographic quadrant
reflex activity

Figure 1 Nociceptive signals activate dorsal horn neurons, which in turn transfer the information to the brain to elicit pain
and to spinal interneurons (broken blue line) to elicit somatic and vegetative reflexes. The effects on motoneurons through
polysynaptic pathways trigger a movement that moves the stimulated area away from the stimulus. This movement results
from both excitatory (e.g., contraction of flexor muscles) and inhibitory mechanisms the latter affecting antagonist muscles.
In animals, one can record from neurons (a) and from muscles (b) but the sensation (c) cannot be assessed. In man, on the
other hand, one cannot record from neurons but it is possible to monitor both the sensation and the reflex activities. In an
experimental paradigm designed for the study of pain inhibiting pain phenomena, one requires a conditioned stimulus (here in
blue) with the corresponding endpoints (a, b, or c) and a conditioning stimulus (here in red). A painful focus activates dorsal
horn neurons that send an excitatory signal through the ventrolateral quadrant toward higher centers, including the lower
brainstem. This signal activates DNICs, which will inhibit WDR neurons through the dorsolateral funiculi (DLF). With two
painful foci, the resulting sensation will depend on the balance of the nociceptive information elicited from the two sources.
The power of such nociceptive information depends on the three dimensions of any nociceptive stimulus, namely strength,
duration, and area. In an experimental situation designed to observe inhibitions of the response to the conditioned stimulus,
the beam of the balance should tilt in favor of the conditioning stimulus. This is illustrated in Figure 2 where the response to a
20 ms duration series of electrical pulses is inhibited by the stimulation of the hand for 2 min.
766 Diffuse Noxious Inhibitory Controls (DNIC)

10 mA Rlll reflex Pain sensation

15 mA 0.2 mV % T mr a.u. T up
20 mA 50 ms 100 10 10
9

25 mA 8
7
6
50 5 5
Stimulation Recording
4
3
2
Biceps 1 Tp
0 0
0 20 40 mA 0 20 40 mA
Sural nerve
Tr

150 150 150 150

(a) T r increase (b) T mr increase (c) T p increase (d) T up increase
% % % %

100 100 100 100

50 50 50 50

0 0 0 0
40 42 44 46 48 C 40 42 44 46 48 C 40 42 44 46 48 C 40 42 44 46 48 C

(A) 42 C 44 C 45 C 46 C 47 C

100

50

(B) 200 ml 400 ml 600 ml 800 ml 1000 ml

100

50

0
(C) (a) Healthy (b) Tetraplegic (c) Wallenberg (d) Thalamic lesion

150

100

50

0
 Diffuse Noxious Inhibitory Controls (DNIC) 767

hand did not elicit any change in the stimulus (Figure 2(A)). Very similar effects were produced
response relationships. By contrast, temperatures by visceral stimulation (gastric or rectal distension)
between 45 and 47.5  C caused the stimulus in healthy volunteers (Bouhassira, D. et al., 1994;
response curves to shift to the right as a direct func- 1998). During gastric distension for example
tion of the temperature. Similar effects were observed (Figure 2(B)), the inhibitory effects increased with
when other painful procedures were applied as con- the volume of distension and the resultant sensation.
ditioning stimuli (e.g., cold, pressure, exercise under As with the inhibitions elicited by stimulation of
ischemia, high-intensity transcutaneous electrical somatic structures, these visceral-evoked inhibitions
nerve stimulation) (Willer, J. C. et al., 1984; 1989; were correlated to the intensity of the conditioning
Danziger, N. et al., 1998; France, C. R. and stimuli. However, they could be triggered by stimuli
Suchowiecki, S., 1999; Sandrini, G. et al., 2000). which were unpleasant but not quite painful
The temporal evolution of these phenomena can (Figure 2(C)). When they become clearly painful,
be studied by applying a heterotopic conditioning the duration of the inhibitory effects outlasted the
stimulus while the sural nerve is stimulated at a duration of the distension by several minutes.
constant intensity (Figure 2, lower panels). This pro- Clinical pains can also activate this type of inhibi-
cedure revealed aftereffects of long duration. For tion. Thus, in patients suffering from sciatic pain due
example, a 2 min immersion of the hand in a 47  C to a herniated disk, a Lasegues maneuver of the
water bath completely abolished the RIII reflex and it affected leg, which elicits a typical radicular pain,
took more than 9 min to achieve a full recovery also blocks the RIII reflex, both in the affected and

Figure 2 (Upper panels) Experimental setup for recording the nociceptive RIII flexion reflex from the biceps femoris muscle
in the lower limb using surface electrodes. The reflex is evoked by electrical stimulation of the ipsilateral sural nerve. The insert
shows typical examples of electromyographic (EMG) responses elicited by increasing stimulus intensities (from top to
bottom). The subjective sensation was estimated by the subject on a 10-level scale (box with switches shown in insert), with
the pain threshold being defined as level 3. Examples of stimulusresponse curves for the nociceptive reflex and subjective
reports of sensations produced by a wide range of stimulation intensities applied randomly are shown on the left and right
graphs, respectively. Note the close correlations between (1) the reflex threshold (Tr) and the pain threshold (Tp) around
10 mA; and (2) the stimulus intensities producing maximal responses, that is, the threshold of the maximal reflex recruitment
(Tmr) versus the threshold of unbearable pain (Tup). Such recruitment curves were built before and during a 2 min period of
immersion of the right hand in a thermoregulated water bath. No effects were seen at lower temperatures but a shift of the
curves to the right was elicited when the temperature reached painful levels (45  C and above). The shift applied to the reflex
(a, b) and sensation (c, d) thresholds. (Lower panels) The temporal evolution of the reflex can be monitored by employing
repeated, constant, stimulation of the sural nerve (1.2 times threshold every 6 s). The individual RIII reflex is expressed in terms
of the percentages of the mean value recorded during a control period. Each vertical bar represents the mean of ten
responses recorded during 1 min. Graphs show pooled data from several subjects. (A) Effects of immersion of the hand. The
nonpainful temperature (42 and 44  C, left) did not modify the reflex. By contrast, the painful temperatures (45, 46, and 47  C)
depressed the reflex during and after the period of conditioning. The extent of these depressions was temperature
dependent. Note the long duration (around 10 min) of the inhibitory posteffects following the application of the highest
temperature (47  C, right). (B) Effects of visceral stimulation. Gastric distension by means of a balloon introduced into the
proximal part of the stomach was applied for a 3 min period. The inflation of the balloon in volumes of 200 or 400 ml did not
modify the RIII reflex, whereas volumes of 600, 800, and 1000 ml produced inhibitions correlated to the volume of distension.
The 600 and 800 ml distensions were unpleasant while the 1000 ml volume was clearly painful. (C) Effects of nociceptive
electrical stimuli (2025 mA) applied to the upper limb of healthy volunteers or patients with lesions of the spinal cord or the
brain. (a) In healthy volunteers; (b) in tetraplegic patients, the conditioning stimulus did not inhibit the RIII reflex; (c) in patients
with a Wallenberg syndrome (unilateral lesion of the retro-olivary part of the brainstem) the conditioning stimuli did or did not
inhibit the RIII reflex depending on whether it was applied to the normal hand (ipsilateral to the brain lesion) or the analgesic
hand (contralateral to the brain lesion); and (d) in patients with thalamic lesions, the conditioning stimulation applied to the
analgesic hand (contralateral to the brain lesion) did produce inhibition of the RIII reflex. (Upper panels) Adapted from Willer, J.
C., Roby, A., and Le Bars, D. 1984. Psychophysical and electrophysiological approaches to the pain relieving effect of
heterotopic nociceptive stimuli. Brain 107, 10951112. (A) Adapted from Willer, J. C., De Broucker, T., and Le Bars, D. 1989.
Encoding of nociceptive thermal stimuli by diffuse noxious inhibitory controls in humans. J. Neurophysiol. 62, 10281038.
(B) Adapted from Bouhassira, D., Chollet, R., Coffin, B., et al. 1994. Inhibition of a somatic nociceptive reflex by gastric
distention in humans. Gastroenterology 107, 985992. (C) Adapted from Roby-Brami, A., Bussel, B., Willer, J. C., and Le Bars,
D. 1987. An electrophysiological investigation into the pain-relieving effects of heterotopic nociceptive stimuli: probable
involvement of a supraspinal loop. Brain 110, 14971508; and De Broucker, T., Cesaro, P., Willer, J. C., Le Bars, D. 1990.
Diffuse noxious inhibitory controls (DNIC) in man: involvement of the spino-reticular tract. Brain 113, 12231234.
768 Diffuse Noxious Inhibitory Controls (DNIC)

in the nonaffected leg (Willer, J. C. et al., 1987). When nociceptive conditioning stimuli applied to the
the Lasegues maneuver is performed on the healthy fourth and fifth fingers of the left hand were com-
limb, it is painless and has no effect on the reflex. In pared in normal subjects and tetraplegic patients with
patients suffering from neuropathic pain, the RIII lesions of traumatic origin at the cervical-57 level
reflex is inhibited when pain is triggered by pressure (Roby-Brami, A. et al., 1987). In the normal subjects
(static allodynia), but not when it is triggered by (Figure 2(Ca)), the painful conditioning stimuli
light brushing (dynamic allodynia) (Bouhassira, D. strongly depressed both the RIII reflex and the asso-
et al., 2003). Interestingly, it is generally agreed that ciated pain. By contrast, in the tetraplegic patients,
the former is due to the activation of nociceptive nociceptive stimulation of the same territories,
processes via A- and C-fibers, while the latter is which, being in the cervical-8 and thoracic-1 derma-
due to the activation of A-fibers. tomes, were clinically unaffected by the spinal lesion,
In summary, a painful conditioning stimulus can and did not produce any depression of the RIII reflex
depress both a preexisting pain and its associated (Figure 2(Cb)). One can therefore conclude that the
nociceptive reflex at the first spinal relays for inhibitory effects triggered by heterotopic nocicep-
the transmission of nociceptive information. tive stimuli are sustained by a loop that includes
Interestingly, nociceptive brainstem reflexes such as supraspinal structures.
the blink or jaw-opening reflexes are also inhibited In order to identify, or at least localize, these
by remote painful stimulation (Pantaleo, T. et al., supraspinal structures, observations were made on
1988; Maillou, P. and Cadden, S. W., 1997; Ellrich, patients with cerebral lesions causing contralateral
J. and Treede, R. D., 1998). By contrast, nonnocicep- hemianalgesia (De Broucker, T. et al., 1990). These
tive reflexes such as jerk or Hoffmann reflexes were patients with either a lesion of the retro-olivary
remain unaffected during such procedures. One part of the medulla (Wallenbergs syndrome,
must stress here that there has to be an obvious Figure 2(Cc)) or a unilateral thalamic lesion
imbalance between the conditioned and the condi- (Figure 2(Cd)). In the former group, no inhibitions
tioning stimuli for a clear depressive effect to be seen. were observed when the (not felt) nociceptive con-
For instance, in the examples shown in Figure 2, the ditioning stimulus was applied to the affected side of
conditioned stimuli were short (20 ms) trains of elec- the body, contralateral to the brain lesion. If this
trical shocks, whereas the conditioning stimuli were stimulus was applied to the normal side, ipsilateral
applied over time periods of several seconds to large to the brain lesion, it was felt as painful and triggered
areas of the body. Clearly, spatiotemporal summation inhibitory effects very similar to those seen in normal
is required to elicit visible inhibitory effects because subjects. By contrast, in the patients with a thalamic
of the reciprocal nature of the phenomenon triggered lesion, the RIII reflex was strongly depressed, as in
by two remote painful foci. A balance, the beam of normal subjects, by the nociceptive conditioning sti-
which tips (or does not tip) in favor of one or other mulus applied to the affected side (which was not
pain, could symbolize the net result of such a reci- felt as painful). Thalamic structures and conse-
procal process (Figure 1, upper left). Of course, quently spinothalamic pathways, are therefore not
clinical situations are always much more complicated involved in DNIC, whereas brainstem probably
than these experimental situations which are delib- reticular structures seem to play a key role in
erately designed to simplify the problem under these phenomena. These observations also demon-
analysis. More particularly, it seems likely that the strate unambiguously that such phenomena can be
existence of multiple painful foci would produce elicited in the absence of a painful sensation, pro-
very complicated interactions between excitatory vided the nociceptive nature of the information
and inhibitory processes. reaches some brain structures. This suggests that
the observed phenomenon does not result from a
competition between the two attentional foci that
50.3 A Spinally Mediated Process the conditioned and the conditioning pain represent
Involving Supraspinal Structures in the healthy volunteers. This is an important obser-
vation as it is known that the perception of pain is
Are the inhibitory mechanisms described above due sensitive to attentional processes (Bushnell, M. C.
to propriospinal mechanisms or do they involve et al., 1985; Willer, J. C. et al., 1979).
supraspinal structures? To answer this question, the An exceptional case was also reported in a patient
effects on the RIII reflex in the right leg, of with a Brown-Sequard syndrome due to a hemisection
 Diffuse Noxious Inhibitory Controls (DNIC) 769

of the spinal cord (left side, T6 level) produced by a injections of an irritant agent or noxious heat all
knife-stab in the back (Bouhassira, D. et al., 1993). produce an increase in nociceptive thresholds in dis-
The RIII reflexes elicited by stimulation of cutaneous tant somatic structures, notably the tail (e.g., tail flick
afferents in the ulnar and sural nerves were studied in test) and paws (e.g., paw withdrawal test). (See refer-
the upper and lower limbs by recording from the ences in Le Bars, D. et al., 2001.)
biceps brachialis and biceps femoris muscles, respec-
tively. For each limb, the RIII reflex was elicited
regularly before, during, and after periods of nocicep- 50.5 The Role of Wide-Dynamic-
tive electrical conditioning stimulation applied Range Neurons
successively to the other three limbs. Inhibitions of
around 90% followed by strong poststimulus effects In several species (rat, mouse, cat, and probably
were observed in all situations except that (1) monkey), most WDR and some nociceptive-specific
no inhibition could be obtained when the conditioning neurons are strongly inhibited by a noxious stimulus
stimuli were applied to the lower right limb applied outside their excitatory receptive fields. Such
(contralateral to the spinal lesion), and (2) the RIII effects do not appear to be somatotopically organized
reflex in the lower left limb was completely insensitive but apply to the whole body. Figure 3 shows exam-
to any of the conditioning stimuli. These results ples of recordings from lumbar WDR neurons in the
strongly suggest that in human beings (1) the ascending rat. The activities of these neurons whether elicited
part of the loop subserving DNIC is completely by pinching their receptive field or by applying an
crossed at the spinal level, and (2) the descending excitatory amino acid to their membranes were
part is confined to the white matter ipsilateral to the strongly inhibited by noxious mechanical (pinch of
limb being tested. the muzzle, the contralateral hindpaw or the tail) or
thermal stimuli (immersion of the tail in a 52  C
water bath) (Figure 3(A)). These examples illustrate
50.4 A Model in the Rat the whole body inhibitory receptive fields of WDR
neurons as recorded in intact rats. Interestingly, such
Another nociceptive reflex this time recorded in inhibitory controls are exacerbated during clinical
the anaesthetized rat is the C-fiber reflex which can pains, for example, when an animal suffers from
be elicited by electrical stimulation of the sural nerve monoarthritis, polyarthritis or a peripheral mono-
and recorded from the biceps femoris muscle. This neuropathy (Danziger, N. et al., 2000; 2001).
reflex can be strongly inhibited by both mechanical These properties of dorsal horn neurons were
and thermal noxious heterotopic stimuli applied to observed at the level of various segments of the spinal
the muzzle, a paw or the tail, and by colorectal dis- cord and in both nucleus caudalis and nucleus oralis
tension. These inhibitory effects on the C-fiber reflex of the trigeminal system thus suggesting a general
did not occur in spinal animals, or ipsilateral to a organization. In keeping with the human studies
rostral unilateral lesion of the dorsolateral funiculus shown in Figure 2, there is a clear relationship
(DLF). Such observations are consistent with several between the intensity of the conditioning stimulus
reports showing the inhibition of reflexes or the and the strength of the resultant DNIC. With strong
increase of nociceptive thresholds elicited by hetero- stimuli, the inhibitory effects are powerful and fol-
topic noxious conditioning. For example, the reflex lowed by long-lasting poststimulus effects, which can
discharge in the common peroneal nerve following persist for several minutes. In some cases, the inhibi-
electrical stimulation of the sural nerve in the rat was tory effects can involve a complete abolition of
inhibited by pinching the muzzle or tail; the gastro- activity for a long period of time following removal
cnemius medialis reflex evoked by sural nerve of the conditioning stimuli (switch off) and the
stimulation in the decerebrate rabbit was inhibited activity can be restored to preconditioning levels by
by electrical stimulation of the contralateral common further manipulations of the excitatory receptive
peroneal or of the ipsi- or contralateral median field (switch on) (Cadden, S. W., 1993). (See refer-
nerves; the digastric reflex evoked by tooth pulp ences in Le Bars, D., 2002.)
stimulation in the cat was inhibited by toe pinch, With regard to the viscera, some differences should
percutaneous electrical stimulation of a limb or elec- be noted: visceral stimuli, for example, distension of
trical stimulation of the saphenous nerve. In the colon or urinary bladder, generally produce inhi-
behavioral experiments, intraperitoneal or cutaneous bitions with slower rates of onset and recovery but
770 Diffuse Noxious Inhibitory Controls (DNIC)

(A) Pinch-evoked GLU-evoked

Muzzle (pinch)

Paw (pinch)
Electrophoresis
of DLH
Recording

Tail (pinch)

Peripheral 10 Hz
excitatory 1 min
field

Tail (52 C)

(B) 20

(a) 10

0
200 0 200 400 600 800 ms

Electrophoresis
N
of DLH
Recording 10

0
200 0 200 400 600 800 ms
Peripheral
excitatory
field 7 ms 210 ms
(b)
25 ms 330 ms
20

100 mm N

10
(c)

0
200 0 200 400 600 800 ms
 Diffuse Noxious Inhibitory Controls (DNIC) 771

starting at intensities below a painful level 7.3 and 0.7 m s1, which fall into the A- and C-fiber
(Cadden, S. W. and Morrison, J. F. B., 1991). It was ranges, respectively. Such biphasic inhibitions could
proposed that these differences might have reflected be elicited from any part of the body and recorded
different amounts and patterns of activity in the rele- from any WDR neurons. Figure 3(B) shows a record-
vant primary afferent fibers rather than being due to ing from a lumbar WDR neuron with an excitatory
different central neural mechanisms. Again, this is receptive field located on the extremity of the ipsi-
consistent with human studies (e.g., Figure 2(B)). lateral hind paw: two components of inhibition were
In any case, these data suggest that DNICs are induced by the activation of A and C-fibers, respec-
triggered specifically by the activation of peripheral tively, when a single 2 ms duration shock of 10 mA was
nociceptors whose signals are carried by A- and applied to the muzzle, the base or the tip of the tail.
C-fibers. In order to investigate further the types of DNICs are not observed in anesthetized or decere-
peripheral fiber involved in DNIC, use was made of brate animals in which the spinal cord has been
the facts that (1) trigeminal and spinal dorsal sectioned. It is therefore obvious that the mechanisms
horn neurons respond with relatively steady dis- underlying DNICs are not confined to the spinal cord
charges to the microelectrophoretic application of and that supraspinal structures must be involved. Such
excitatory amino acids; and (2) DNICs act by a final a system is therefore completely different from seg-
postsynaptic inhibitory mechanism involving hyper- mental inhibitory systems, which work both in intact
polarization of the neuronal membrane. It was found and in spinal animals, and can be triggered by the
that when trigeminal WDR neurons were directly activation of low-threshold afferents. DNICs are also
excited by the microelectrophoretic application of very different from the propriospinal inhibitory pro-
glutamate, the percutaneous application of single cesses that can be triggered by noxious inputs. It should
square-wave, electrical stimuli to the tail always also be noted that DNIC are blocked by increasing the
induced a biphasic depression of the resultant activ- anesthetic regimen (Jinks, S. L. et al., 2003).
ity. Both the early and late components of this The ascending and descending limbs of this loop
inhibition occurred with shorter latencies when the travel through the ventrolateral and dorsolateral
base rather than the tip of the tail was stimulated. funiculi, respectively. Since thalamic lesions do not
Such differences in latency were used to estimate the affect DNICs, it has been proposed that they result
mean conduction velocities of the peripheral fibers from a physiological activation of some of the brain-
triggering the inhibitions: the means were found to be stem structures that produce descending inhibition

Figure 3 (A) Example of inhibitions of a spinal WDR neuron elicited by noxious heterotopic stimuli. Recordings were made
in the lumbar dorsal horn from a WDR neuron with a receptive field located on the ipsilateral hind paw. The neuron was
activated either by a sustained pinch of its receptive field or by regular (once a minute) microelectrophoretic applications of
the excitatory amino acid, glutamate (GLU, 20 nA, horizontal bars). Conditioning stimuli were pinch of the muzzle, the
contralateral hind paw or the tail, and immersion of the tail in a 52  C water bath (from top to bottom, respectively). Note that all
these conditioning stimuli virtually blocked both the pinch-evoked and the DLH-induced firing. In the latter case, the
inhibitions remained for several minutes in most cases, suggesting that WDR neurons were hyperpolarized for a long time
following the conditioning stimulation. (B) Example of heterotopic activation of A- and C-fibers triggering inhibitions in a
spinal WDR neuron. Left: Schematic representation of the experimental design. Recordings were made in the lumbar dorsal
horn from a WDR neuron with a receptive field located on the ipsilateral hind paw. The continuous microelectrophoretic
application of the excitatory amino acid, DL-homocysteic acid (DLH) induced a steady discharge from the neuron under
study. The repetitive application of individual percutaneous electrical stimuli of adequate intensities to the contralateral
muzzle (a), the base (b), or the tip (c) of the tail induced biphasic depressions of the neuronal activity. Right: Peristimulus
histograms (bin width 5 ms) prepared during the continuous microelectrophoretic application (15 nA) of DLH onto the
membrane of the neuron. The broken white lines show the timing of percutaneous electrical stimulation (10 mA; 2 ms duration;
0.66 Hz; 200 ms delay; 100 sweeps). The broken black line represents the mean firing calculated during the prestimulation
control period (200 to 0 ms). Two waves of inhibition can be seen. They occurred earlier when the base of the tail (b) was
stimulated instead of the tip (c). The time gaps are shown as yellow areas between the histograms, for both inhibitory
components. The gap was 7 and 25 ms for the beginning and the end of the first component; it was 150 and 290 ms for the
beginning and the end of the second component. Knowing that the distance between b and c was 100 mm, one can easily
calculate the conduction velocities of fibers that elicited the first and second components: 414 m s1 and 0.30.7 m s1
respectively. These fibers therefore belong to the A- and C- groups, respectively. (A) Adapted from Villanueva, L.,
Cadden, S. W., and Le Bars, D. 1984. Evidence that diffuse noxious inhibitory controls (DNIC) are mediated by a final
post-synaptic inhibitory mechanism. Brain Res. 298, 6774.
772 Diffuse Noxious Inhibitory Controls (DNIC)

(see Chapter 5.49). Surprisingly, DNICs were not Many sources of descending inhibition from the
modified by lesions of the following structures: the brain that modulate the spinal transmission of noci-
periaqueductal gray (PAG), cuneiform nucleus, para- ceptive information have been described in animals.
brachial area, locus coeruleus/subcoeruleus, rostral To date, the only descending inhibitory mechanisms
ventromedial medulla (RVM). By contrast, lesions of that have been described in man are DNICs.
subnucleus reticularis dorsalis (SRD) in the caudal
medulla strongly reduced DNICs. The SRD is
located ventral to the cuneate nucleus, between tri-
geminal nucleus caudalis and the nucleus of the
solitary tract and contains neurons with characteris-
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 Diffuse Noxious Inhibitory Controls (DNIC) 773

0.8 and 1.2 minimum alveolar anesthetic concentration. Sandrini, G., Milanov, I., Malaguti, S., Nigrelli, M. P., Moglia, A.,
Anesth. Analg. 97, 111116. and Nappi, G. 2000. Effects of hypnosis on diffuse noxious
Le Bars, D. 2002. The whole body receptive field of dorsal horn inhibitory controls. Physiol. Behav. 69, 295300.
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Le Bars, D. and Willer, J. C. 2002. Pain Modulation Triggered by medullary subnucleus reticularis dorsalis (SRD) as a key link
High-Intensity Stimulation: Implication for Acupuncture in both the transmission and modulation of pain signals. Pain
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Maillou, P. and Cadden, S. W. 1997. Effects of remote deep Supraspinal influences on nociceptive flexion reflex and pain
somatic noxious stimuli on a jaw reflex in man. Arch. Oral. sensation in man. Brain Res. 179, 6168.
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Melzack, R. 1989. Folk Medicine and the Sensory Modulation of nociceptive thermal stimuli by diffuse noxious inhibitory
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 51 Fibromyalgia
R Staud, University of Florida, Gainesville, FL, USA
2009 Elsevier Inc. All rights reserved.

51.1 Introduction 775

51.2 Pathogenesis of Fibromyalgia 776
51.3 Muscle Nociception 776
51.4 Triggering Events for Fibromyalgia 776
51.5 Abnormal Response to Stressors 776
51.6 Posttraumatic Stress Disorder and Fibromyalgia 777
51.7 Fibromyalgia and Affective Spectrum Disorders 777
51.8 Central Sensitization in Fibromyalgia 777
51.9 Temporal Summation of Second Pain (Wind-Up) 777
51.10 Abnormal Wind-Up of Fibromyalgia Patients 778
51.11 Wind-Up Measures as Predictors of Clinical Pain Intensity in Fibromyalgia Patients 778
51.12 Fibromyalgia and Other Chronic Pain Conditions 778
51.13 Myofascial Pain Syndrome 778
51.14 Trigger Points 778
51.15 Relationship between Myofascial Pain and Fibromyalgia 779
51.16 Inflammatory Connective Tissue Diseases 779
51.17 Treatment of Fibromyalgia 779
51.18 Prognosis of Fibromyalgia 780
51.19 Conclusions 780
References 780

Glossary
chronic pain Pain for more than 3 months. TPs represent areas of mechanical allodynia (pain
central sensitization Increased sensitivity to threshold 4 kg cm2). Eleven or more TPs as well
painful stimuli due to functional changes of the as chronic widespread pain are required to fulfill
central nervous system (neuroplasticity). fibromyalgia criteria.
peripheral sensitization Increased sensitivity to trigger point (TrP) Painful muscle areas that show
painful stimuli due to decreased thresholds of pri- increased muscle tension (taut bands), a twitch
mary nociceptive afferents. response to palpation, and referred pain.
tender point (TP) Nine paired body areas at ten- wind-up Central pain mechanisms that depend
don insertion points of the neck, chest, buttocks, as on N-methyl-D-aspartate and substance P receptor
well as all upper and lower extremities as defined mechanisms.
by the 1990 American College of Rheumatology.

51.1 Introduction Like many other syndromes, FM has no single specific

feature but is a symptom complex of self-reported or
Chronic nonmalignant pain is very frequent in the elicited findings. In 1990, the American College of
general population (Croft, P. et al., 1993). Fibromyalgia Rheumatology (ACR) published diagnostic criteria
(FM) represents the combination of chronic wide- for FM which include chronic widespread pain
spread pain with mechanical allodynia and dispropor- (>3 months) and mechanical tenderness in at least 11
tionably affects women (ratio women:men 9:1). out of 18 tender points (TP; Wolfe, F. et al., 1990). Most

775
776 Fibromyalgia

TP sites are located at tendon insertion areas and have mostly substance P and calcitonin gene-related pep-
shown few detectable tissue abnormalities. Besides tide (CGRP). There is also a retrograde migration of
musculoskeletal pain and mechanical tenderness most substance P from cell bodies which is then released in
FM patients also complain of insomnia, fatigue, and the region of the free nerve endings. This retrograde
distress. In addition FM coaggregates with major flow can sensitize tissues and increase their respon-
mood disorders (Raphael, K. G. et al., 2004). siveness to less intense stimuli. Muscle nociceptors
are activated by mechanical stimuli (stretching or
pressure), bradykinin, serotonin, and potassium ions,
but are not activated by normal muscle movements
51.2 Pathogenesis of Fibromyalgia or even increased muscle tension. Sensory input from
muscle, as opposed to skin, is a much more potent
FM can be characterized as a pain amplification
effector of central sensitization and this may be par-
syndrome of patients who often display widespread ticularly relevant for FM (Wall, P. D. and Woolf, C.
hyperalgesia and allodynia to mechanical, thermal, J., 1984).
and chemical stimuli. However, the hypersensitivity
of FM patients is not limited to pain, but also
includes light, sound, and smell. The cause for this
51.4 Triggering Events for
heightened sensitivity is unknown, but central ner-
Fibromyalgia
vous system (CNS) sensory processing abnormalities
have been reported in several studies (Desmeules, J.
The onset of FM has frequently been associated with
A. et al., 2003; Staud, R., 2004). Most of these studies
certain triggers or stressors include physical trauma,
found evidence of central sensitization of dorsal horn
infections, emotional distress, endocrine disorders,
neurons of the spinal cord and the brain including
and immune activation (Greenfield, S. et al., 1992;
those that are related to pain. The pathogenesis of Middleton, G. D. et al., 1994; Waylonis, G. W. and
central sensitization in FM is unclear but may be
Perkins, R. H., 1994). These stressors have been asso-
related to prior stressors, including infections and
ciated with the degree of pain, disability, life
traumas. Because most FM patients relate their pain
interference, and affective distress of FM patients
to deep tissues, particularly muscles, these structures
(Turk, D. C. et al., 1996). Strong evidence for trauma
may play an important role in the initiation and
as a trigger of FM symptoms comes from studies of
maintenance of central sensitization and pain. No
patients with acute injuries who developed chronic
consistent tissue abnormalities have been reported
widespread pain at much higher rates than controls
in FM patients that would explain persistent pain. (Al Allaf, A. W. et al., 2002; Buskila, D. and Neumann,
However, focal muscle areas, like trigger points
L., 2002). Additional evidence for such an association
(TrPs), may play an important role for FM pain
include postinjury reported sleep abnormalities
because little nociceptive input from peripheral tis-
(Berglund, A. et al., 2001), local injury sites as a source
sues is required for the maintenance of central
of chronic distant regional pain (Arendt-Nielsen, L.
sensitization (Bengtsson, M. et al., 1989).
and Svensson, P., 2001), and recent evidence of
injury-related CNS neuroplasticity in FM (Carli,
G., 2000). Further prospective studies, however, are
51.3 Muscle Nociception needed to confirm these associations and to identify
the mechanisms that are relevant for posttraumatic
Nociceptive input from muscle may be particularly FM pain (White, K. P. et al., 2000).
relevant for FM pain. Muscle pain travels in myeli-
nated A-delta fibers (group III) and in unmyelinated
type C fibers (group IV). Pain receptors are mostly 51.5 Abnormal Response to
located at free nerve endings which are concentrated Stressors
around small arterioles and capillaries between the
muscle fibers. The cell bodies for these nerves are The biological response to stressors appears predict-
found in the dorsal root ganglion and synapse in able in animals and humans. Particularly, events
lamina I and IVV of the dorsal horn of the spinal that are perceived as inescapable or unavoidable,
cord. Activation of peripheral nociceptors leads to a or which appear unpredictable, evoke the strongest
release of neurotransmitters in the spinal cord, adverse biological responses (Gold, P. W. et al., 1988a;
 Fibromyalgia 777

1988b; Chrousos, G. P. and Gold, P. W., 1992; Viau, 51.7 Fibromyalgia and Affective
V. et al., 1993; Sapolsky, R. M. et al., 2000). This may Spectrum Disorders
explain why victims of trauma appear to develop
much higher rates of FM than persons with Several studies have reported that FM is comorbid
self-inflicted injuries (Greenfield, S. et al., 1992). In with major depressive disorder (MDD; Arnold, L. M.
addition, early life stressors can have a permanent et al., 2000; White, K. P. et al., 2002). In addition,
and profound impact on subsequent biological anxiety is very prevalent in patients with FM
responses to stress in animals and humans. Studies (Thieme, K. et al., 2004). The outcome of a recent
in rodents have demonstrated that exposure to multi- large family study of FM probands was consistent
ple stressors including trauma or separation in the with the hypothesis that FM and MDD are charac-
neonatal period can lead to permanent changes in terized by shared, genetically mediated risk factors
the biological response to stress (Sapolsky, R. M., (Raphael, K. G. et al., 2004). Although the findings of
1996; McNamara, R. K. et al., 2002). This permanent this study should not be interpreted that MDD and
effect of early stressors could explain the higher FM represent different forms of the same syndrome,
than expected incidence of traumatic childhood they strongly suggest that FM and MDD share
events in individuals who later develop chronic important CNS mechanisms.
pain (Anderberg, U. M. et al., 2000; Bailey, B. E.
et al., 2003).
51.8 Central Sensitization in
Fibromyalgia

51.6 Posttraumatic Stress Disorder Whereas peripheral sensitization is related to

and Fibromyalgia changes of primary nociceptive afferent properties,
central sensitization requires functional changes in
Posttraumatic stress disorder (PTSD) often occurs the CNS (neuroplasticity). Behaviorally, centrally
after a significant traumatic event and is character- sensitized patients, like FM sufferers report abnormal
ized by behavioral, emotional, functional, and or heightened pain sensitivity with spreading of
physiological symptoms. Relevant traumatic events hypersensitivity to uninjured sites and the generation
related to PTSD are usually perceived as threaten- of pain by low threshold mechanoreceptors that are
ing ones life or physical integrity and can lead to normally silent in pain processing (Torebjork, H. E.
emotional responses including horror, helplessness, et al., 1992). Thus, tissue injury may not only cause
or intense fear. The psychological symptoms of pain but also an expansion of dorsal horn receptive
PTSD include re-experience of the traumatic fields and central sensitization.
event, avoidance, and increased arousal. It has been
shown that the experience of trauma is associated
with increased somatic and physical complaints, 51.9 Temporal Summation of Second
including pain (Beckham, J. C. et al., 1997; 1998). Pain (Wind-Up)
More than 50% of patients with FM suffer from
PTSD in the USA and Israel (Sherman, J. J. et al., In 1965, Mendell and Wall described for the first
2000; Cohen, H. et al., 2002). Compared to the pre- time, that repetitive C-fiber stimulation can result
valence of PTSD in the general population (6%), in a progressive increase of electrical discharges
FM patients show greatly increased rates similar to from second-order neurons in the spinal cord
Vietnam veterans and victims of natural disasters (Mendell, L. M. and Wall, P. D., 1965). This impor-
(Shore, J. H. et al., 1986; Green, M. M. et al., 1993). tant mechanism of pain amplification in the dorsal
Not surprisingly, the incidence of FM is also horn neurons of the spinal cord is related to temporal
increased in patients with PTSD (21%) and often summation of second pain or wind-up (WU) and has
associated with increased pain ratings, more distress, been used in FM patients for the evaluation of central
and higher functional impairment (Amir, M. et al., sensitization (Staud, R. et al., 2001). This technique
1997). As with several other disorders, however, it is reveals sensitivity to input from unmyelinated (C)
unclear whether PTSD is the cause or consequence afferents and the status of the N-methyl-D-aspartate
for FM. (NMDA) receptor systems that are implicated in a
778 Fibromyalgia

variety of chronic pain conditions. Temporal sum- Sjogrens syndrome, inflammatory myopathies
mation depends upon activation of NMDA (Sultan, S. M. et al., 2002), systemic lupus erythema-
transmitter systems by C nociceptors, and chronic tosus (Tench, C. et al., 2002), multiple sclerosis, and
central pain states like FM can result from excessive joint hypermobility syndrome (Nef, W. and Gerber,
temporal summation of pain (Staud, R. and N. J., 1998). Furthermore, several infectious diseases
Smitherman, M. L., 2002). including hepatitis C, Lyme disease, coxsackie B
infection, HIV, and parvovirus infection, frequently
result in chronic pain states (Barkhuizen, A., 2002).
51.10 Abnormal Wind-Up of Although the majority of FM patients report the
Fibromyalgia Patients insidious onset of pain and fatigue, approximately
half of all patients describe the start of chronic pain
Several recent studies have demonstrated psychophy- after a traumatic or infectious event.
sical evidence that input to central nociceptive
pathways is abnormally processed in patients with
FM (Staud, R. and Domingo, M., 2001a; 2001b; 51.13 Myofascial Pain Syndrome
Staud, R. et al., 2001; Vierck, C. J. et al., 2001; Price,
D. D. et al., 2002; Staud, R., 2002). When WU pain is Myofascial pain or regional musculoskeletal pain is
evoked both in normal controls and FM patients, the one of the most common pain syndromes encoun-
perceived magnitude of the experimental stimuli tered in clinical practice. Myofascial pain represents
(heat, cold, pressure, electricity) is greater for FM the most common cause of chronic pain, including
patients compared to controls, as is the amount of neck and shoulder pain, tension headaches, and lower
temporal summation within a series of stimuli. back pain (Granges, G. and Littlejohn, G., 1993;
Following a series of stimuli, after-sensations are Fricton, J. R., 1994; Macfarlane, G. J. et al., 1996;
greater in magnitude, last longer, and are more fre- Borg-Stein, J. and Simons, D. G., 2002). The term
quently painful in FM subjects. These results indicate myofascial pain was introduced in the early 1950s by
both augmentation and prolonged decay of nocicep- Janet Travell. She also defined the term myofascial
tive input in FM patients and provide convincing TrP and demonstrated with David Simons that indi-
evidence for the presence of central sensitization. vidual muscles have specific nondermatomal patterns
of TrP pain referral (Simons, D. G. et al., 1999). In
1983, both authors first described the clinical picture
51.11 Wind-Up Measures as and pathophysiology of a new syndrome which they
Predictors of Clinical Pain Intensity in named myofascial pain syndrome (MPS; Long, S. P.
Fibromyalgia Patients and Kephart, W., 1998; Simons, D. G. et al., 1999).
MPS has been defined as a chronic pain syndrome
The important role of central pain mechanisms, like accompanied by TrPs in one or more muscles or
WU, for FM patients is also supported by their ability groups of muscles. Similar to FM, it is more fre-
to predict clinical pain intensity. Thermal WU ratings quently found in women compared to men. Besides
correlate with clinical pain intensity (Pearsons the presence of TrPs and referred pain, MPS is fre-
r 0.529), thus emphasizing the important role of quently associated with limitation of movement,
these pain mechanisms for FM. In addition, statistical weakness, and autonomic dysfunction (Long, S. P.
prediction models that includes TP count, pain- and Kephart, W., 1998) similar to FM.
related negative affect, and WU ratings have been
shown to account for nearly 50% of the variance in
FM clinical pain intensity (Staud, R. et al., 2003; 2004). 51.14 Trigger Points

TrPs represent areas of local mechanical hyperalge-

51.12 Fibromyalgia and Other sia that can be found in MPS and several chronic pain
Chronic Pain Conditions conditions, including FM, osteoarthritis, and rheu-
matoid arthritis. They are defined as specific areas of
Many systemic illnesses can also present with diffuse hyperirritability in muscle, but can also be detected
pain similar to FM, including polymyalgia rheuma- in ligaments, tendons, periosteum, scar tissue, or skin
tica (Gowin, K. M., 2000), rheumatoid arthritis, (Han, S. C. and Harrison, P., 1997; Simons, D. G. et al.,
 Fibromyalgia 779

1999). TrPs are located in palpable taut bands and elevated erythrocyte sedimentation rate (ESR),
produce local and referred pain, which is specific for C-reactive protein (CRP), anemia, autoantibodies
the particular muscle. When TrPs are mechanically (rheumatoid factor, cyclic citrullinated peptide
stimulated, so-called taut bands within a muscle, antibodies, anti-nuclear antibodies (ANA)), etc.
rather than the entire muscle, will contract (Chu, J.,
1998). They are often associated with a local muscle
twitch response, which can easily be elicited by need- 51.17 Treatment of Fibromyalgia
ling or palpation of the TrP (Chu, J., 1998; 1999).
Latent TrPs are similar to active TrPs, but they are Treatment of patients with chronic widespread pain
not associated with spontaneous pain and no referral like FM needs to be individually tailored to each
of pain occurs. However, latent TrPs are painful patients needs. This includes the assessment of biop-
when palpated. sychosocial abnormalities which are readily
detectable in most FM patients. Importantly the
identification of pain generators is essential for an
51.15 Relationship between effective treatment plan. Thus patients with arthritis,
Myofascial Pain and Fibromyalgia particularly osteoarthritis of the spine will benefit
from muscle relaxants, physical therapy, and mas-
Approximately 70% of patients with FM have TrPs sage. In addition, these patients may respond well to
(Granges, G. and Littlejohn, G., 1993). A TP is con- therapy with cyclo-oxygenase (COX) inhibitors.
sidered to be different from a TrP because of the Identification and treatment of mood abnormalities
absence of referred pain, local twitch response, and a is crucial because affective spectrum disorders seem
taut band in the muscle. The distinction between to share important mechanisms with FM.
TPs and TrPs requires careful physical examination. Pharmacotherapy for FM has been most success-
TrPs are frequently located in areas of muscular TPs ful with antidepressant, muscle relaxant, or
of patients with FM (Wolfe, F. et al., 1992; Borg-Stein, anticonvulsant drugs. These drugs affect the release
J. and Stein, J., 1996) suggesting that some muscular of various neurochemicals (e.g., serotonin, norepi-
TPs in patients with FM may actually be TrPs nephrine, substance P) that have a broad range of
(Inanici, F. et al., 1999). The presence of TrPs in activities in the brain and spinal cord, including
most if not all FM patients represents evidence for modulation of pain sensation and tolerance. None
local muscle abnormalities in this chronic musculos- of these drugs, however, is currently approved by
keletal pain syndrome. Although it is unclear if TrPs the US Food and Drug Administration for the treat-
are the cause or effect of muscle injury, they repre- ment of FM.
sent abnormally contracted muscle fibers. This Most FM patients will respond to low-dose tri-
muscle contraction can lead to accumulation of his- cyclic medications, such as amitriptyline and
tamine, serotonin, tachykinins, and prostaglandins, cyclobenzaprine, as well as cardiovascular exercise,
which may result in the activation of local nocicep- cognitive behavioral therapy (CBT), patient educa-
tors. Prolonged muscle contractions may also result tion, or a combination of these for the management of
in local hypoxemia and energy depletion (Simons, D. FM. Also tramadol, selective serotonin reuptake inhi-
G. et al., 1999). bitors (SSRI), selective norepinephrine re-uptake
inhibitors (SNRI), and anticonvulsants have been
found to be moderately effective. There is some
51.16 Inflammatory Connective evidence for the efficacy of strength training exercise,
Tissue Diseases acupuncture, hypnotherapy, biofeedback, massage,
and warm water baths. However, many commonly
Many patients (up to 25%) with chronic arthritis used FM therapies like guaifenesine have been found
have also wide-spread pain similar to FM. These to be ineffective.
patients may also present with symptoms of chronic The efficacy of FM patients to manage their pain
fatigue, impaired memory and concentration, and seems to correlate with their functional status. Brain
mood abnormalities. Most of these patients, how- imaging and psychological profiles have identified at
ever, will have findings suggestive of inflammation, least three FM subgroups, that is, patients who are
including joint pain/swelling, rashes, muscle weak- highly dysfunctional, interpersonally distressed, or
ness, as well as laboratory abnormalities, like are effective copers (Giesecke, T. et al., 2003). Such
780 Fibromyalgia

studies provide an explanation why some treatments References

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 52 Pain Perception Nociception during Sleep
G J Lavigne, Universite de Montreal, Montreal, QC, Canada
K Okura, Tokushima Graduate School, Tokushima, Japan
M T Smith, John Hopkins Medical School, Baltimore, MD, USA
2009 Elsevier Inc. All rights reserved.

52.1 Pain in Relation to Sleep 784

52.1.1 Definition, Epidemiology, and Relevance 784
52.1.2 Cognitive Disturbances 785
52.1.3 Sleep Fragmentation and Deprivation 785
52.1.4 Circadian Variation in Pain Perception 786
52.2 Perception of Pain during Sleep: Nociception or Sleep Arousal 786
52.2.1 Nociception Pain Perception during Sleep 786
52.2.2 Sleep Arousal: Brain and Autonomic Activation 789
52.3 Circular Relation between Pain and Poor Sleep and Putative Consequence on
Health Cost 790
References 791

Glossary
alpha EEG wave intrusion The intrusion of fast- physiological activity; it tends to be repeated 815
frequency electroencephalographic (EEG) alpha times per hour of sleep.
(7.511 Hz) activity into slow wave sleep (SWS) or nociception The receiving and transmitting of
into deep sleep (stages 3 and 4). SWS is dominated noxious sensory signals from the periphery to the
by large and slow EEG waves of delta type (0.54.0 central nervous system.
or 0.754.5 Hz); it also characterizes sleep stages 3 pain An unpleasant and sensory, cognitive, and
and 4. SWS is not a specific biological marker of emotional experience normally associated or not
pain during sleep. with tissue damage, which usually requires a mini-
sleep awakening (major arousal) A sudden mal level of consciousness for expression.
increase in EEG and heart rate frequency with a rise pain threshold The lowest experimental stimula-
in muscle tone that lasts more than 10 or 15 s. tion perceived as being painful by a conscious
Subject is not usually aware of external influences. subject in 50% of the trials.
An excessive number of major arousals during sleep A natural physiological and behavioral state
sleep is frequently followed by a complaint of usually characterized by a partial isolation from the
unrefreshing sleep and cognitive impairment the environment except when an unpleasant or poten-
following day. tially harmful or life-threatening event is present.
cyclic alternating pattern Repetition of micro- sleep stage A division of the specific sleep period
arousals every 2060 s as a sentinel that allows for into sleep stages (St): light sleep (St 1 or 2), deep
a reset of physiological functions (e.g., heart rate, sleep (St 3 and 4) are both described as non-REM
respiration, muscle tone) or prepares the body for sleep, and the REM or paradoxical sleep. Most
an appropriate response in relation to potential body movements occur in light non-REM sleep
disrupting events. (periodic limb movement, sleep bruxism). Sleep
micro-arousal A sudden increase, during sleep, in apnea (cessation of breathing for more than 10 s) is
EEG activity and heart rate frequency under a car- dominant in deep sleep and in REM sleep. REM
diac sympathetic dominance with a possible rise in behavior disorder (RBD) is characterized by sudden
muscle tone. It should last more than 3 s but less movement in REM sleep that is normally associated
than 10 or 15 s depending on scoring method. A with muscle hypotonia (behaviorally described as
sleeping subject is normally unaware of this motor paralysis).

783
784 Pain Perception Nociception during Sleep

sleep fragmentation Interruption of any sleep deep St 3 and 4 sleep and possible alpha EEG
stage by isolated or repetitive events such as sleep wave intrusion. As a result, sleep continuity may be
stage shifts (deeper to lighter), micro-arousals, impaired and poor sleep quality (e.g., unrefreshing)
frank awakenings, with or without low duration of is frequently reported upon awakening.

52.1 Pain in Relation to Sleep 2001; Moldofsky, H., 2001). A cross-sectional study
done in Hungary with over 12 643 adults showed that
52.1.1 Definition, Epidemiology, and
insomnia-related complaints were 3 times higher in
Relevance
chronic pain patients than in nonpain subjects
Pain is a sensation that usually requires a certain (Novak, M. et al., 2004). Finally, the concomitance of
level of consciousness before it is interpreted as an sleep disorders and pain may modify the subjective
unpleasant sensory experience. Most persistent pain sensation related to sleep quality (e.g., refreshing)
states can trigger complaints of poor sleep quality. since 50% of insomnia patients with pain reported
Poor sleep quality is reported in 5090% of patients that pain interfered with daily activities (Novak, M.
with the following chronic pain conditions: arthritis, et al., 2004). Moreover, in the presence of bodily pain
cervical and orofacial pain, low back pain, cancer there is an increase (OR of 1.73) in self-reports of
pain, fibromyalgia. (Atkinson, J. H. et al., 1988; Dao, daytime sleepiness (Foley, D. et al., 2004) a topic
T. T. T. et al., 1994; Morin, C. M. et al., 1998; Smith, further described in the next section.
M. T. et al., 2000; Dauvilliers, Y. and Touchon, J., In explorations of the interaction between high
2001; Riley, J. L. III et al. 2001; Roizenblatt, S. et al., pain intensity and poor sleep, it is important to con-
2001; McCracken, L. M. and Iverson, G. L., 2002). sider that chronic pain is frequently concomitant
Sleep quality is also impaired in the presence of other with anxiety, fatigue, mood disturbance, depression,
medical conditions of which pain is a common symp- and poor physical fitness (Menefee, L. A. et al., 2000b;
tom, including: headaches, irritable bowel syndrome, Riley, III. J. L. et al., 2001; Smith, M. T. et al., 2000;
spinal cord injury, and metastatic breast cancer McCraken, L. M. and Iverson, G. L. 2002b; Nicassio,
(Cohen, M. et al., 2000; Menefee, L. A. et al., 2000; P. M. et al., 2002; Sayar, K. et al., 2002; Yatani, H. et al.,
Moldofsky, H., 2001; Widerstrom-Noga, E. G. et al., 2002). For example, fatigue is frequently reported in
2001; Rains, J. C. and Penzien, D. B., 2002; Koopman, chronic pain patients and can explain 6.5% of the
C. et al., 2002; Okifuji, A. and Turk, D. C., 2002; variability between pain intensity and complaints of
Lavigne, G. J. et al., 2005). According to a recent poor sleep. Depression may further explain up to
survey from the National Sleep Foundation (USA), 40% of this variability. The presence of frank sleep
up to 20% of the adult population reports that their apnea, a major risk for fatigue and daytime sleepiness,
sleep is disrupted by somatic discomfort and pain. seems to be relatively low in chronic pain patients, as
The interaction between pain and poor sleep is reported in a recent review (Dauvilliers, Y. and
supported by cohort and case control studies. Patients Touchon, J. 2001; Donald, F. et al., 1996). However,
with orofacial pain are 3.7 more times at risk of the cumulative effect of poor sleep over a given night
reporting poor sleep (Macfarlane, T. V. and and possibly over several nights also needs to be
Worthington, H. V., 2004). In a survey done in the considered. While it is recognized that daytime slee-
USA with 1506 community women and men aged piness may result from long interruptions of
between 55 and 84, it was found that bodily pain respiration (more than 10 s is defined as sleep
was associated with a risk for complaints of insomnia apnea), less information is available on the cumula-
(e.g., difficulty falling asleep, middle of the night/ tive consequence of several briefer hypoxic events,
early morning awakenings, or nonrestorative sleep). which may alter cognitive function and brain meta-
Odds ratios for insomnia among those reporting bod- bolism (Bonnet, M. H. 1985; Wesenten, N. J. et al.,
ily pain were estimated to be between 1.88 and 2.68 1999; Stepanski, E. J., 2002; Martin, S. E. et al 1996;
(Foley, D. et al., 2004). It has also been reported that Van Dogen, H. P. et al., 2003; Gozal, D. and OBrien,
the prevalence of insomnia is 2 times higher in L. M., 2004). Recent work in fibromyalgia, for
Canadian chronic pain patients in comparison to the instance, suggests the possibility of a high rate of
general population without pain (Sutton, D. A. et al., sleep-related upper airway resistance syndrome that
 Pain Perception Nociception during Sleep 785

when reversed by continuous positive airway pres- The increase in sleep fragmentation in normal
sure therapy, may improve daytime function (Gold, subjects or in patients with sleep apnea seems to be
A. R. et al., 2004). The impact of the above-mentioned clearly associated with a reduction in next-day per-
variables on pain and sleep interaction and their formance, sleepiness, and impaired alertness (Bonnet,
reversal by continuous positive airway pressure M. H., 1985; Wesenten, N. J. et al., 1999; Stepanski, E. J.,
(CPAP) needs to be further assessed in controlled 2002; Martin, S. E. et al., 1996; Van Dogen, H. P. et al.,
and blind studies. 2003). A minimum of 56 h of sleep seems to be the
threshold below which daytime performance is altered
and, interestingly, several days are required before a
52.1.2 Cognitive Disturbances
return to baseline levels (Van Dogen, H. P. et al., 2003).
Very few studies have investigated the effect of pain There is still no definitive evidence indicating that
on cognitive functions such as memory and attention. lack of deep sleep (St 3 and 4) or of slow wave EEG
Three studies using chronic musculoskeletal pain and activity (SWA) increases pain sensitivity, although
fibromyalgia patients suggest a causative association some animal and human experimental studies suggest
(Kewman, D. G. et al., 1991; Landro, N. I. et al., 1997; such a link (see review from Kundermann, B. et al.,
Cote, K. A. and Moldofsky, H., 1997). The impact of 2004a; Kundermann, B. and Lautenbacher, S., 2007).
cognitive deficits and their potential aggravation by To our knowledge only one study has compared
disturbed sleep needs to be further studied. The deep-sleep deprivation with other sleep stages or
potential impact of several classes of pain medication total sleep deprivation (Onen, A. H. et al., 2001).
(e.g., opioids, anticonvulsants, muscle relaxants, anti- Moreover, the absence of deep sleep is a feature of
depressants) on sleep and daytime alertness is also a all chronic-pain conditions (Okura, K. et al., 2004).
particularly important line of research with real-life Before summarizing the studies on sleep depriva-
consequences ranging from sleep-related industrial tion and changes in pain perception, it is important to
accidents to impaired driving ability (George, C .F. reiterate that study design and data analysis need to
P., 2003; Chapman, S., 2001). A recent experimental control for: (1) the differences between groups for
study done in normal volunteers showed that a one age, since after 40 years of age, slow wave density, a
night infusion of morphine during sleep time can alter dominant feature of deep sleep, tends to decline; (2)
sleep quality by reducing deep-sleep duration and presence of respiratory disturbances during sleep,
increasing sleep stage 2 duration (Shaw, I. R., et al., since patients with fibromyalgia who complain of
2005). Evidence-based data on long-term use of mor- daytime hypersomnolence tend to present with
phine on sleep homeostasis are missing. more micro-arousals, a higher number of short
respiratory disturbances and longer oxygen desatura-
tion periods than matched normal controls, an effect
52.1.3 Sleep Fragmentation and
reversed by CPAP (Sergi, M. et al., 1999; Gold, A. R.
Deprivation
et al., 2004); (3) the concomitant effect of fatigue,
Sleep fragmentation is usually characterized by a mood depression, anxiety, poor lifestyle habits, etc.,
rapid shift from a deeper sleep stage to a lighter one as described above. In addition to the above issues, it
or by an increase in the number of brief arousals up to is also important to note that the literature of sleep
longer awakenings during the night (see Glossary for deprivation and pain is generally limited by experi-
definitions). The number of body movements and mental studies done with small sample sizes and a
respiratory disturbance events, which are often asso- lack of adequate control groups.
ciated with arousals, seems to have an additive effect Several studies have been conducted in normal
on daytime functioning over the following day or subjects to assess the effect of deep-sleep deprivation
days. Sleep deprivation is a more general term that on next-day pain perception. The disruption of sleep
either characterizes the absence or very short dura- stage 4 or SWS by sound was associated with an
tion of a given sleep stage and/or the near absence or increase in muscle tenderness and a reduction in pain
reduction of sleep as tested by experimental manip- threshold (mechanical pressure) the next morning, but
ulations. As recently demonstrated, measures of the this effect was less dominant when studies controlled
impact of chronic sleep loss fall into a clear hierarchy: for concomitant influences of fatigue (Moldofsky, H.
mood measures being more sensitive than cognitive and Scarisbrick, P., 1976; Lentz, M. J. et al., 1999; Older,
tasks, which are in turn more sensitive than perfor- S. A. et al., 1998). Negative findings have been reported
mance of motor tasks (Bonnet, M. H., 2005). by others (Drewes, A. M. et al., 1997; Kundermann, B.
786 Pain Perception Nociception during Sleep

et al., 2004a; Older, S. A. et al., 1998; Arima, T. et al., involve pain that directly interferes with sleep; in
2001; Smith, M. T., et al., 2007). Surprisingly, when patients with torticolis, pain is rapidly reduced by
normal subjects were deprived of SWA sleep, REM adopting the supine posture (Lobbezoo, F. et al.,
sleep or total sleep duration, no change in thermal pain 1996; Bentley, A. J., 2007).
threshold was noted during any of the sleep interrup-
tions (Onen, A. H. et al., 2001). In this study, however,
total sleep deprivation slightly reduced mechanical 52.2 Perception of Pain during Sleep:
pain tolerance scores (8%) and in the period after Nociception or Sleep Arousal
sleep recovery, a significant rebound effect was noted,
52.2.1 Nociception Pain Perception
such that mechanical pain tolerance was 15% higher. A
during Sleep
recent study done in normal volunteers showed that
two nights of total sleep deprivation reduced both heat While awake, tactile and thermal sensory input of
and cold pain thresholds, but had no effect on warmth sufficient intensity can activate A delta, A beta, and C
and cold detection threshold; results suggested that sensory fibres projecting to the thalamus and several
change in pain threshold was not specific to alterations cortical areas (Bromm, B. and Lorenz, J. 1998;
in somatosensory function (Kundermann, B. et al., Coghill, R.C. et al., 1994; Le Pera, D. et al., 2000;
2004b). Kundermann and colleagues also suggested, Peyron, R. et al., 2000). The long duration of painful
in a comprehensive review, that the role of stress and hypertonic saline infusions seems to activate A delta
changes in endogenous serotonin and opioid activity and C afferent fibers that in the wake state reach the
can also play an important role in arousal and changes thalamus, amygdale, and other cortical areas asso-
in pain perception (Kundermann, B. et al., 2004a). ciated with pain processing (Le Pera, D. et al., 2000;
It is important to clarify that experimental sleep Zubieta, J. K. et al., 2001). During sleep, however, a
deprivation, which has been induced in the above putative gating process of somatosensory inputs is
studies, is an extreme manipulation that may be dif- thought to prevent awakening resulting from irrele-
ferent from the natural ongoing sleep disruption vant input or non-life-threatening events. Several
experienced by chronic pain patients. As described studies suggest that sleep-preserving sensory gating
above, fragmentation or deprivation, not necessarily mechanisms are likely to be located at the upper
limited to St 3 and 4, seems to alter cognitive function brain stem area (see Figure 1) in the so-called reti-
and memory consolidation; all sleep stages seem to be cular activating system (Pompeiano, O. and Swett, J.
equally important for the preservation and mainte- E. 1963; Hernandez-Peon, R. et al., 1965; Soja, P. J.
nance of daytime functions (Stickgold, R. et al., 2000; et al., 2001; Soja, P. J., 2007; Nofzinger, E. and
Mednick, S. et al., 2003; Gais, S. et al., 2000). Derbyshire, S. W. G., 2007). However, the balance
between wakefulness system(s) and sleep-promoting
neuronal network(s) in relation to sensory discrimi-
52.1.4 Circadian Variation in Pain
nation and pain during sleep is poorly understood
Perception
and requires further investigation.
Surprisingly, most studies using normal nonpain sub- Regarding sensory perception during sleep, it is
jects without mood alteration have failed to crucial to discriminate between studies made using
demonstrate a clear circadian variation in experimen- sound and those using tactile stimulations. The
tal pain threshold or pain intensity reports over a 24 h results of sound studies have made it apparent that
schedule or when comparing evening and morning acoustic stimulation triggers more sleep arousals and
data (Rogers, E. J. and Vilkin, B. 1978; Strian, F. et al., awakenings in non-REM light sleep stages (St 1 and 2)
1989; Koltyn, K. F. et al., 1999; Lavigne, G. J. et al., than in deep sleep (St 3 and 4) or in REM sleep
2000; Bentley, A. J. et al., 2003; Lavigne, G. J. et al., (Rechstaffen, A. and Kales, A., 1968; Carley, D. W.
2004). In patients with chronic pain, however, symp- et al., 1997; Kato, T. et al., 2004). Sound is a different
tom presentation often has a typical circadian pattern. sensory modality from pain: its main function with
Arthritic pain is often worse in the morning, while respect to sleep is to alert the individual to wake up
pain levels related to fibromyalgia, myofascial orofa- (alarm clock) or to parental duties (a babys cry).
cial pain, and some headache conditions are typically Tactile stimulation, however, is more likely to be
higher in the afternoon (Bellamy, N. et al., 1991; interpreted as a touch to the body, which may or
Reilly, P. A. and Littlejohn, G. O., 1993; Dao, T. T. may not enforce the awakening process and is likely
T. et al., 1994). Furthermore, not all pain conditions to be interpreted differently than sound by a sleeping
 Pain Perception Nociception during Sleep 787

Wake Sleep
Cortex Cortex

Thalamus Thalamus

Hypothalamus Hypothalamus
Brainstem Brainstem
reticular formation reticular formation

Stimulus Stimulus

Figure 1 During wake state, there is a free flow in neuronal activity from spinal cord or/and brainstem to and from thalamus
and cortical networks. During sleep, a gating mechanism seems to isolate the upper brain from most incoming inputs
originating at somatosensory level. However, some circuits, for example, auditory processing, remain fully active. The exact
anatomical site of the separation (see dashed line) between spinal and brainstem networks and thalamo-cortical pathways
remain to be demonstrated.

brain (Velluti, R. A. et al., 2000; Kato, T. et al., 2004; suppressed (Cairns, B. E. et al., 1996; Soja, P. J. et al.,
Soja, P. J. et al., 2001) Moreover, during sleep the term 2001). It was further suggested that the serotoninergic
nociception is probably more accurate to describe the and nonserotoninergic brainstem raphe magnus cells
process of potentially harmful inputs, interpreted in are involved to an important extent in the sensory
conscious and awake subjects as pain (Bromm, B. and modulation of pain inputs (Foo, H. and Mason, P.,
Lorenz, J., 1998; Lavigne, G. J. et al., 2000). 2003). Interestingly, recording of thalamic cells in
Studies done in the 1960s, using tactile-cutaneous one human has confirmed previous animal findings
sensory stimulations during wake/sleep states, are a suggesting that thalamic neurons discharge differently
rich source of information (Pompeiano, O. and Swett, during states of waking and sleeping. It was observed
J. E., 1963; Hernandez-Peon, R. et al., 1965) It was that the discharge of thalamic neurons was in a tonic
suggested that a filtering mechanism is active during mode while the patient was awake and in a bursting
synchronized or non-REM sleep at the first relay of mode when the patient was in light sleep; nevertheless
neurons in the brainstem mesencephalic-reticular for- none of these cells had a clear sensory receptive field
mation and in the trigeminal sensory nucleus. More in the periphery (Tsoukatos, J. et al., 1997). These
recent work lends further support to the concept that findings are similar to those from animal studies show-
sleep involves a selective isolation from the external ing that thalamic neurons are under a global inhibition
milieu. Recordings of ascending spinoreticular and during sleep while cortical neurons, by contrast, are
trigemino-thalamic tract neurons show that a gating highly active (Steriade, M. and Timofeev, I., 2003).
mechanism seems to be present during sleep when The exact location of the mechanism(s) supporting the
brief sensory or pain stimulations are applied to the somatosensory dissociation during sleep remains
skin of the animal. Furthermore, the neurons behaved under investigation (see Figure 1). The balance
differently across sleep stages, that is, during non REM between mechanisms that maintain vigilance or pro-
or REM sleep: air puff activated trigemino-thalamic mote sleep continuity, however, seems to be state-
tract neurons during REM sleep whereas neuronal dependent and related to the modulation of neuronal
activity following tooth pulp stimulation was networks at the level of the reticular activating system.
788 Pain Perception Nociception during Sleep

The initiation of an awakening and conscious reaction the first 2 days, and a reduction in duration of deep
during sleep may be secondary to a decrease in the sleep (St 3 and 4 in humans) and REM sleep stages
sensory gating in sleep resulting in the recovery of the (Carli, G. et al., 1987) Interestingly, after 2 days, the
free flow between ascending and descending brain- animals recovered, exhibiting total sleep time and
stem, thalamus and cortical activities (Siegel, J., 2004; deep sleep of longer duration; a typical pattern seen
Skinner, R. D. et al., 2004). In other words, it is as if the after sleep deprivation (Onen, A. H. et al., 2001;
reptilian brain maintains basic survival function dur- Kundermann, B. et al., 2004a). Another sleep and
ing sleep, and interaction with thalamo-cortical pain study in rats, using the experimental adjuvant
networks depends on the nature (sound, touch, and arthritis model in both hind paws, also showed frag-
harmful stimulus), magnitude, and duration of the mentation of the typical sleep and wake pattern
stimulation. Obviously, this hypothesis needs to be (Landis, C. A. et al., 1988). The number of brief
supported by further evidence. sleep periods was increased over the circadian
A recent paper summarizing the evidence on brain rhythm; total sleep time and deep sleep duration
activation during sleep provides relevant information were also shorter in rats with arthritis. The above-
(Peigneux, P. et al., 2003). During light sleep, pontine mentioned animal studies suggest that a sleeping
tegmentum and thalamus are deactivated while brain is not completely isolated from the external
mesencephalic regions remain active. In deep sleep, milieu and that pain-nociception can activate net-
all three structures are relatively deactivated while works related to vigilance and body protection.
activity is largely maintained at the cortical temporal This activation appears to be related to the type
and insular lobe levels. Paradoxically, all the above (phasic versus tonic, intensity, etc.) and duration of
structures are activated during REM sleep, although the painful stimuli (Velluti R. A. et al., 2000; Soja, P. J.
a strong dissociation is present: there is no peripheral et al., 2001; Kato, T. et al., 2004; Lavigne, G. J. et al.,
efferent activity (e.g., the motor cortex does not trig- 2004).
ger a movement except in the presence of sleep In humans, the perception of pain during sleep
pathology: the REM behaviour disorder). There is also depends on the duration and type of stimulus
little evidence strongly supporting a clear-cut gating used; brief sensory nerve-related or mild duration
of sensory information at the lower brain level, to thermal or long duration chemical or mechanical
prevent awakening from irrelevant sensory inputs. stimulations initiate different types of responses that
However, the role of the amygdala and the hypotha- need to be interpreted accordingly. In a study using
lamus is discussed in the interpretation of sensory the spinal motor responses, following brief stimula-
gating outside the higher thalamocortical levels tion of the leg sural nerve to evoke a flexion reflex, it
(Morrison, A. R. et al., 2000; Lee, R. S. et al., 2001). A was noted that the reflex latency was prolonged dur-
review of behavioral animal and human studies is also ing sleep in comparison to wake state (Sandrini, G.
useful for a better understanding of the modulation of et al., 2001). In parallel, the threshold was 60% higher
sensory inputs during sleep. in non-REM and 200% higher in REM sleep than
The interaction between the pain system and vig- during wake state. This study supports the above-
ilance versus sleep is further supported by a cat study mentioned animal findings, suggesting that during
using the classical tail flick sensory motor reflex sleep lower brain neurons modulate pain inputs in
(Kshatri, A. M. et al., 1998). It was suggested that the such a way that a hypoalgesic response seems to be
gating of sensory inputs during sleep depends on the present. In contrast, when sensory processing from
integrity of the cholinergic system at the medial periphery to cortex was estimated with extremely
pontinereticular level. This work also clearly brief (<60 ms) sensory-laser or electrical evoked
showed, in comparison to the wake state and in the responses to pain, the somatosensory cortically
absence of any medication, that the reflex latency was evoked potential completely disappeared during
3 times longer in non-REM sleep (quiet sleep in light sleep (Bedyoun, A. et al., 1993; Wang, X. et al.,
animal) and 5 times longer in REM sleep. This 2003). This absence of somatosensory evoked poten-
finding again supports the concept of higher respon- tials during sleep stands in contrast to similarly
siveness to brief sensory inputs during non-REM in present auditory stimuli in which evoked potentials
comparison to REM sleep. However, another cat are largely preserved during sleep. Sound appears to
study revealed that under tonic sensory nociceptive play a more sensitive role in preserving bodily integ-
influences (formalin injection in the foot) the animals rity during sleep than tactile-somatosensory input
tended to remain awake with delayed sleep onset for (Kato, T. et al., 2004). Alternatively, in collaboration
 Pain Perception Nociception during Sleep 789

with the Italian Milano sleep group, we found that Lavigne, G. J. et al., 2001; Bentley, A. J. et al., 2003;
longer (612 s) moderate heat pain stimulation (46  C) Lavigne, G. J. et al., 2004). It is interesting to note that
applied over the upper arm can evoke a clear sleep the so-called fast alpha EEG intrusion, which for a
behavioral response: nearly twice the number of long time was believed to be a marker of pain in
micro-arousals (see Glossary for definition) in light relation to sleep, is not considered to provide the
non-REM sleep compared to deep or REM sleep sole explanation for poor sleep in pain patients
(Lavigne,G. J. et al., 2000). These results were con- (Moldofsky, H., 2001; Mahowald, M. L. and
firmed by another group that found that higher Mahowald M. W., 2000; Roizenblat, S. et al., 2001;
thermal pain temperatures were required to elicit Rains, J. C. and Penzien, D. B., 2003). More recent
arousal during deep sleep and REM in comparison thinking conceptualizes pain as a disruptor of sleep
to light sleep. These temperatures were closer to homeostasis that may increase sleep arousal instabil-
waking pain tolerance levels instead of pain threshold ity, described as an augmentation in the normal
level (Bentley, A. J. et al., 2003). More recently, with cyclical alternating pattern in which brain and car-
the objective of challenging sensory pain processing diac activation tends to recur in clusters (Staedt, J.
across sleep stages, we used a hypertonic saline infu- et al., 1993; Moldofsky, H., 2001; Roizenblatt, S. et al.,
sion in the arm muscle to mimic longer clinical pain 2001; Parrino, L., Zucconi, M. and Terzano, M. G.,
episodes (>100 s). A similar procedure has been used 2007).
by others during sleep (Drewes, A. M. et al., 1997). Cardiac activation during sleep is used as an indir-
Interestingly, we found that micro-arousal responses ect estimate of sympathetic (e.g., accelerator) and
were also more dominant in light sleep in comparison parasympathetic (decelerator or brake) balance dur-
to deep and REM sleep, and a clear and dominant ing wake and sleep states. During wake state, a
sleep awakening response was present with an equi- sympathetic dominance predominates, maintaining a
potent response rate in all sleep stages (Lavigne, G. J. high level of vigilance. Sympathetic-cardiac domi-
et al., 2004). nance is also present during REM sleep, a state with
This evidence suggests that when a painful stimu- high cortical and cardiac activity but with a beha-
lus or a clinical pain episode lasts long enough, the vioral muscle paralysis. Conversely, during non-
protective mechanism that maintains sleep continu- REM sleep, parasympathetic-cardiac activity is domi-
ity (the gating barrier to irrelevant input) is released nant and this may provide a recuperative function
and a clear behavioral response may occur with a since there is an inverse coupling: a low sympathetic
potential return to consciousness. Recent clinical dominance is inversely coupled with a high degree of
evidence for impaired somatosensory gating during SWA in each consecutive non-REM to REM cycle
sleep in chronic pain conditions has been suggested (Brandenberger, G. et al., 2001). Interestingly, in
by findings of decreased sleep spindle activity in insomnia and fibromyalgia patients, a sympathetic
chronic low back pain and fibromyalgia patients dominance persists across NREM sleep cycles at
(Harman, K. et al., 2002; Landis, C. A. et al., 2004). levels similar to wake and REM sleep states
Thus it can be hypothesized that brief sensory sti- (Bonnet, M. H. and Arand D. L., 1998; Martnez-
muli, such as electrical or thermal pain, are too brief Lavn, M. et al., 1998). It has been hypothesized that
to be processed by a sleeping brain as potentially enhanced sympathetic activity during sleep may
harmful, whereas longer-lasting or tonic painful sti- over-facilitate arousal mechanisms and cyclical pat-
muli akin to clinical pain states are more likely to terns of brain activation. However, this hypothesis
elicit a full-blown arousal response. was not supported in a study of medication-free
women suffering from fibromyalgia in comparison to
women with efficient sleep and no pain (McMillan, D.
52.2.2 Sleep Arousal: Brain and Autonomic
et al., 2004). By contrast, when patients with chronic
Activation
fibromyalgia (widespread pain) and pain (low back,
The above arousal and awakening responses etc.) presenting with poor sleep efficiency were
observed with thermal and deep muscle infusion are matched for age and gender to normal sleepers, we
not only restricted to a cortical activation as seen on found (see Figure 2) that the normal parasympathetic
electroencephalogram. They are also associated with and sympathetic balance was lost in both pain groups
a clear increase in heart rate and a change in respira- during non-REM sleep with an inverse coupling of
tion that is frequently interrupted with the onset of SWA, that is, a lower density of SWA was observed
the pain stimulation (Lavigne, G. J. et al., 2000; during consecutive sleep stages compared to control
790 Pain Perception Nociception during Sleep

500 500
400 p = 0.03 FM 400 p = 0.09 Pain

(amplitude V)
SWA power 300 300

200 200

100 100

0 0
NREM REM NREM REM NREM REM NREM REM NREM REM NREM REM

1st sleep cycle 2nd sleep cycle 3rd sleep cycle 1st sleep cycle 2nd sleep cycle 3rd sleep cycle

0.9 0.9

0.8 FM 0.8 Pain

LF/(HF + LF)

0.7 0.7

0.6 0.6

0.5 0.5

0.4 0.4
n = 10 n = 10
Normal
FM
Pain

Figure 2 Dynamics of SWA and HRV over three consecutive sleep cycles. In normal subjects, SWA shows an inverse
coupling with sympathetic activity (low-frequency divided by high-frequency plus low-frequency. One of the heart rate
variability: HRV). During non-REM sleep, SWA was high when the sympathetic cardiac activity was low, and, conversely, SWA
was low when HRV was high in REM sleep. In comparison to normal subjects, fibromyalgia (FM) chronic widespread pain
patients showed a significant reduction in SWA for the first sleep cycle only (t-test: p 0.03). Chronic pain patients (PAIN) also
presented a similar trend in SWA, but not statistically significant (t-test: p 0.09). PAIN patients did not show a rise in
sympathetic activity during REM sleep (p NS: repeated ANOVA); FM patients showed a trend toward no rise in REM sleep.

subjects (Okura, K. et al., 2005). Furthermore, like 1998; Smith, M. T. et al., 2000; Riley, J. L., III et al.,
previous studies of men with fibromyalgia in a wake 2001). In other words, in retrospective accounts, sleep
state, our preliminary analysis suggests that men disturbance is generally not considered a major pro-
maintain a high sympathetic activity across all sleep blem for many patients until an injury is experienced;
stages (Cohen, H. et al., 2001). Women, however, do then pain and sleep become salient interacting issues.
not show the usual rise in sympathetic activity during Intensive, time-series diary studies using multilevel
REM sleep. Our data differ from those of (Martinez- modeling statistical techniques consistently show that
Lavin, M., et al., 1998) since we separated data into over time, a day with intense pain is frequently fol-
consecutive non-REM and REM cycles. Caution is lowed by a poor sleep quality and subsequent ratings of
needed when interpreting the above-mentioned poor sleep are in turn linked to next-day increases in
observations since parasympathetic and sympathetic clinical pain. This circular relationship, also described
balance tends to be altered with age and acute stress in as a vicious circle, has been observed in patients with
relation to sleep (Brandenberger, G. et al., 2003; Hall, fibromyalgia, severe skin burns, and rheumatoid arthri-
M. et al., 2004). tis (Affleck, G. et al., 1996; Raymond, I. et al., 2001;
Stone, A. A. et al., 1997; Nicassio, P. M. et al., 2002;
Lavigne, G. J. et al., 2005). As has been mentioned
52.3 Circular Relation between Pain above, the influences of fatigue, mood alteration,
and Poor Sleep and Putative other sleep disorders (such as sleep apnea or periodic
Consequence on Health Cost limb movement), and medication use have not been
well controlled for in these studies.
The initiation of pain (e.g., acute post-op or trauma) No direct data, to our knowledge, reports on the
usually precedes or coincides with the onset of poor cost of pain and its interaction with poor sleep.
sleep in close to 5389% of patients (Morin, C. M. et al., However, an extensive study done in a Canadian
 Pain Perception Nociception during Sleep 791

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 53 Pharmacological Modulation of Pain
A Dray, AstraZeneca Research and Development, Montreal, PQ, Canada
2009 Elsevier Inc. All rights reserved.

53.1 Introduction 795

53.2 G-Protein-Coupled Receptors 796
53.3 Opioids and Their Receptors 796
53.4 Kinins and Their Receptors 798
53.5 Protease-Activated Receptors 799
53.6 Cannabinoids and Their Receptors 799
53.7 Mrg-Related GPCR and Their Ligands 801
53.8 Prostanoids and Receptors 801
53.9 Cytokines, Chemokines, and Their Receptors 802
53.10 Adrenoceptors 803
53.11 Glutamate Regulation and Glutamate Receptors 804
53.12 Gamma-Aminobutyric Acid Receptors 805
53.13 Ion Channels 805
53.13.1 Ligand-Gated Channels 805
53.14 Purinergic Receptors 806
53.14.1 P2X Receptors 806
53.14.2 Acid-Sensing Channels 807
53.15 Sodium Channels 807
53.16 Calcium Channels 808
53.17 Neurotrophins and Their Receptors 809
53.18 Kinases 810
53.19 Botulinum Toxin 810
53.20 Nitric Oxide 811
53.21 Summary and Conclusions 811
References 811

53.1 Introduction processes. Furthermore there is the evolving view

that the pharmacology of pain may alter with time-
Chronic pain is a complex integration of sensory, dependent expression of targets, in the initiation ver-
affective, and cognitive processes encompassing a vari- sus the maintenance phases of pain, and that target
ety of clinical conditions that have been generalized as expression is state-dependent (related to relative
nociceptive (rheumatoid and osteoarthritis (OA)), activity of the pain process or pathway). Thus poor
neuropathic (postdiabetic neuropathy, posttraumatic understanding of pain pharmacology has seriously
neuralgia), or etiologically mixed (low back pain, can- hampered the introduction of new drugs as there
cer pain; for references see Merskey, H. et al., 2005; have been few drugs introduced with a new mechan-
Campbell, J. N. et al., 2006). Pain conditions are etio- ism of action within living memory.
logically diverse and there is limited understanding of A recent step to building rational pain therapy
clinical pain mechanisms and of environmental and has embraced a mechanism-based approach guided
social factors that aggravate pain such as stress and by advances in pain pharmacology. Thus major
anxiety. In addition, it has been difficult to provide a symptoms such as hyperalgesia, allodynia, and
unifying linkage of pain symptoms with the cellular spontaneous pain have been linked with some of the
mechanistic processes underlying these symptoms that identifiable, and often overlapping, neural cellular
would rationalize chronic pain therapy. The complex- processes. The key processes are considered to be
ity of chronic pain is further reflected in the diversity sensitization (reduced threshold for stimulation of
of molecular targets considered to drive pain pain pathways), hyperexcitability (amplification or

795
796 Pharmacological Modulation of Pain

prolongation of nerve discharges in pain pathways), In this chapter I have selected from the emerging
and spontaneous nerve activity (ectopic and/or spon- pharmacology of pain molecular approaches (sum-
taneous discharges in pain pathways). These marized in Table 1) that will provide direction for
processes or mechanisms are driven by molecular future pain treatments.
changes that provide the targets for pharmacological
characterization and analgesia intervention (sum-
marized in Figure 1). It is important to remember 53.2 G-Protein-Coupled Receptors
that for the most part these neural mechanisms and
their molecular drivers have been explored in animal GPCRs have been a relatively tractable family of drug
models and rarely consolidated to the same degree in targets in a number of therapeutic arenas. A variety of
human studies. GPCRs are involved in regulating neural excitability
There have been a number of excellent reviews of in pain pathways, often through the action of a variety
pain mechanisms and molecules (Hunt, S. P. and of inflammatory mediators. These mediators act per-
Mantyh, P., 2001; Scholz, J. and Woolf, C. J., 2002; ipherally or centrally, via an equally large variety of
Watkins, L. R. and Maier, S., 2002) in the peripheral specific membrane receptors (Scholz, J. and Woolf, C.
nervous system and central nervous system (CNS). J., 2002). In general, GPCRs signal via the production
Causative factors for pain are inflammatory media- of second messengers (cyclic adenosine monopho-
tors, factors released and made in response to nerve sphate (cAMP), cyclic guanosine monophosphate
injury, phenotypic changes in neural pathways as (cGMP), diacylglycerol (DAG), phospholipase C
well as supporting glial cells, structural modifications (PLC)) coupled with intracellular protein kinases
including nerve sprouting (sympathetic and sensory and phosphatases (protein kinase A, several isotypes
fibers), nerve rewiring (sensory and CNS fibers), of protein kinase C (PKC)) which in their turn, phos-
and neurodegeneration of specific neurons in the phorylate and dephosphorylate specific cellular
CNS resulting in a hyperexcitable nervous system. proteins including ligand- (e.g., TRPV1) and vol-
Specific molecular interactions that drive the afore- tage-gated membrane ion channels (e.g., NaV1.8)
mentioned excitability changes may differ according which are important in regulating nerve excitability.
to the specific injury and consequent chemical envir-
onments that are created. The molecular interactions
involve all major families of regulatory proteins (G- 53.3 Opioids and Their Receptors
protein-coupled receptors (GPCRs), ion channels,
regulatory enzymes, neurotrophins, kinases), offering Opioids, and morphine in particular, are among the
an abundance of analgesic targets and therapeutic most effective pain medications. They act at peripheral,
opportunities. The pharmacology of these targets spinal, and supraspinal sites through a variety of opioid
is evolving and is dependent on the availability of receptors (-, -, and -opioid receptors). These
reliable chemical and biopharmaceutical tools. receptors are considered to be targets for an endogen-
ous opioid system that has been extensively reviewed
(Yaksh, T. L., 1997). -Opioid receptor activation com-
Central sensitization
Brain
monly causes a variety of well-documented, target-
and hyper-excitability
Opioids, NMDA, mGluR5 related, CNS side effects including sedation, dysphoria,
GABA,
respiratory depression, and constipation. Thus there
has been a vigorous attempt to exploit the peripheral
Spinal cord antinociceptive actions of opioids as a means of avoid-
Skin/Joint/GI DRG ing CNS side effects. In support of this approach,
sensory neurons express and transport opioid receptors
to both the central and peripheral terminals. At central
Spontaneous activity terminals, opioids reduce transmitter release from pri-
NaVs, CaVs, TRPV1, CB1, NGF/TrKA
Peripheral sensitization mary afferent nociceptors, thus blocking synaptic
and hyper-excitability
COX, mPGES, NOS transmission while in the periphery opioid receptor
CB1, B1/2,
TRPV1, NaVs, CaVs, P2X3
activation directly hyperpolarize sensory neurons and
NGF/TrKA attenuate nerve sensitization or hyperexcitability
Figure 1 Mechanistic processes in chronic pain and key induced by inflammation or injury (Hurley, R. W.
molecular drivers. and Hammond, D. L., 2000; Sawynok, J., 2003).
 Pharmacological Modulation of Pain 797

Table 1 Pharmacological modulators of pain and emerging mechanisms of action and clinical indications.

Pharmacological tools
Molecular Analgesia mechanism
Target Agonist Antagonist and targeted tissue Pain indications

-Opioid Morphine, fentanyl Naloxone, naltrexone Agonist: reduced Acute and chronic
receptors peripheral and CNS pain states
excitability
-Opioid Enkephalin, DPDPE, Naltrindole Agonist: Modulation of Nociceptive and
receptor SNC80 CNS pathways inflammatory
B1 receptor Des Arg9 bradykinin des Arg10, HOE-140; Antagonist: Nociceptive and
SSR240612, NVP- inflammatory tissue: inflammatory
SAA 164 pain pathway: PNS,
CNS
B2 receptor Bradykinin Icatibant, bradyzide Antagonist: Nociceptive and
inflammatory tissue: inflammatory
pain pathway: PNS,
CNS
PAR2 Proteases Unknown Antagonist: PNS Nociceptive and
inflammatory
CB1 THC, anandamide, SR141716A Agonist: Pain pathways Nociceptive and
2-arachidonylglycerol, (rimonabant), PNS and CNS neuropathic
palmitoylethanolamide, SR147778,
WIN55, 212-2, ajulemic
acid
CB2 THC, anandamide, SR144428 Agonist: Pain pathways Nociceptive and
2-arachidonylglycerol, PNS and CNS neuropathic
palmitoylethanolamide,
WIN55, 212-2, ajulemic
acid, GW405833, HU-
308, AM 1241
SNSR BAM8-22, 2-MSH Unknown Antagonist: PNS Nociceptive
COX-1 Ibuprofen, naproxene Inhibition of PG Nociceptive and
constitutive synthesis inflammatory
COX-2 induced Ibuprofen, naproxene, Inhibition of PG Nociceptive and
celecoxib, rofecoxib synthesis in PNS and inflammatory
CNS
mPGES Unknown Inhibition of PG Nociceptive and
synthesis in PNS and inflammatory
CNS
IL-1 IL-1 Unknown Antibody versus ligand Nociceptive and
neuropathic
TNF- TNF- Anti-TNF, etanercept, Antibody versus ligand Nociceptive and
adalimumab neuropathic
-2 Clonidine, Inhibition of Nociceptive and
dexmedetomidine neuroexcitability in neuropathic
PNS and CNS
iGluR Glutamate, kainate, AMPA, LY293558, GV196771, Antagonist: CNS Nociceptive and
NMDA ifenprodil, CP-101, pathways neuropathic
606
mGluR (5) Glutamate MPEP, SIB1757 Antagonist: CNS Nociceptive and
pathways neuropathic
GABAA Muscimol Bicuculline Agonist: reduced CNS Nociceptive and
excitability neuropathic
GABAB Baclofen CGP35348 Agonist: reduced CNS Nociceptive and
excitability neuropathic
TRPV1 Capsaicin Capsazepine, Antagonist: PNS Nociceptive and
DD161515, excitability neuropathic
SB705498
TRPA1 Cinnamaldehyde Unknown Antagonist: PNS Neuropathic

(Continued )
798 Pharmacological Modulation of Pain

Table 1 (Continued)

Pharmacological tools
Molecular Analgesia mechanism
Target Agonist Antagonist and targeted tissue Pain indications

TRPM1 Menthol, icilin Unknown Antagonist: PNS Neuropathic

P2X3 ATP, , -methy ATP A-3174919 Antagonist: PNS Nociceptive
P2X4 ATP TNP-ATP Antagonist: CNS Neuropathic
P2X7 ATP Unknown Antagonist: CNS and Nociceptive and
PNS excitability neuropathic
ASIC1-3 Low pH A-317567 Antagonist: PNS Nociceptive
NaV1.3, NaV1.8, Lidocaine, mexiletine, Blocker: PNS Nociceptive and
NaV1.7, lamotrigine excitability neuropathic
NaV1.9
CaV2.2 SNX-111, NMED-160 Blockers: PNS and Nociceptive and
CNS excitability neuropathic
CaV3.2 Ethosuximide Blocker: PNS Neuropathic
NGF NGF ALE0540, PD90780, Antiligand: PNS Nociceptive and
RN624 (mAb) excitability neuropathic
P38-kinase ATP SB203580, CNI-14930 Blocker: PNS and CNS Nociceptive and
excitability neuropathic
iNOS L-NAME, GW27415 Blocker: PNS and CNS Nociceptive and
excitability neuropathic

AMPA, -amino-3-hydroxy-5-methyl-4-isoxazolepropionic acid; ATP, adenosine triphosphate; B, bradykinin; CaV, voltage-gated calcium
channels; CB, cannabinoid; CNS, central nervous system; COX, cyclo-oxygenase; DPDPE, [D-Pen2,D-Pen5]-enkephalin; GABA, gamma-
aminobutyric acid; iNOS, inducible nitric oxide synthase; L-NAME, L-N-nitro-L-arginine methyl ester; mGluR, metabotropic glutamate
receptor; MPEP, 2-methyl-6-(phenylethynyl)-pyridine; mPGES, microsomal-associated prostaglandin E synthase; NaV, voltage-gated
sodium channel; NGF, nerve growth factor; NMDA, N-methyl-D-aspartic acid; PAR, protease-activated receptor; PG, prostaglandin; PNS,
peripheral nervous system; SNSR, sensory neuron specific receptor; THC, tetrahydrocannabinol; mAb, monoclonal antibody; TNP-ATP,
29,39-O-(2,4,6-trinitrophenyl) adenosine 59-triphosphate.

In addition peripherally mediated opioid analgesia 390) and antagonists (e.g., naltrindole; see Dray, A.,
may be enhanced because -opioid receptor expres- 1999 for references).
sion is increased by inflammation and nerve injury  Ligands may provide less analgesic efficacy than
(Stein, C. et al., 2003). This is also accompanied by a morphine but their effectiveness appears to depend on
peripheral increase in the expression of opioids pep- the pain stimulus, the type of injury, and the influence
tides. Novel opioids such as [8-(3,3-diphenyl-propyl)- of their neurochemical environment. Thus, systemi-
4-oxo-1-phenyl-1,3,8-triazaspiro[4.5]dec-3-yl]-acetic cally administered  ligands have low analgesic
acid (DiPOA) and the antidiarrheal drug loperamide, efficacy in acute pain models but show robust analge-
whose distribution is peripherally restricted, have sia efficacy in a variety of chronic pain conditions
shown efficacy in a number of post-operative, inflam- accompanies by inflammation (Cahill, C. M. et al.,
matory, and neuropathic pain models (Whiteside, G. 2001). Enhanced activity has been attributed to stimu-
T. et al., 2005a; Shidoda, K. et al., 2006). lus-induced trafficking of DOR from the cytoplasm to
The pharmacology and analgesic efficacy of nerve membranes in CNS neurons (Cahill, C. M. et al.,
-opioid receptors (DOR) have also been explored 2001). Activity and environment-dependent receptor
since there is a potential for analgesic efficacy with- trafficking also appears as an important regulatory
out the confounding side effects of other opioid ( mechanism in sensory neurons (Bao, L. et al., 2003)
and ) receptor therapies. A variety of studies have but further studies are required to understand the
supported the concept that -selective ligands pro- significance of this in clinical pain and analgesia.
duce analgesia without notable sedation, respiratory
depression, or inhibition of gastrointestinal motility.
These studies have used genetic methods ranging 53.4 Kinins and Their Receptors
from the knockdown of receptors to deletion of the
DOR gene as well as pharmacological tools such as Bradykinin is an important peptidic mediator of inflam-
selective peptides and nonpeptide agonists ([D- matory pain causing nociceptor activation and sensi-
Pen2,D-Pen5]-enkephalin (DPDPE), SNC80, AM- tization via constitutively expressed bradykinin B2
 Pharmacological Modulation of Pain 799

receptors (BK2) on peripheral nociceptors (Dray, A., Kinin antagonists have not been extensively eval-
1997). It also stimulates the production and release of uated in clinical pain although they inhibit other
prostaglandin (PGs) cytokines and nitric oxide (NO). kinin-mediated changes in humans, for example, can-
The abundant metabolite of bradykinin, des Arg9 cer, allergy, and vasodilatation. However, recent
bradykinin (kallidin), activates BK1 receptors which clinical study using a single dose intra-articular
also occur constitutively, but in low abundance, in the joint administration of Icatibant (Flechtenmacher, J.
periphery (Dray, A. and Perkins, M. N., 1993; et al., 2004) produced long-lasting (up to 1 week)
Wotherspoon, G. and Winter, J., 2000) and in the relief of movement-related pain in patients suffering
primate CNS (Shughrue, P. J. et al., 2003). Both recep- from OA. This study highlights a strong proof of
tors are coupled to similar transduction mechanism principle for BK2 involvement in OA and that loca-
including G-i and G-q causing stimulation of lized monotherapy with a selective antagonist can
DAG, PLC, and the release of intracellular calcium. produce significant pain relief.
BK2 receptors are considered to undergo desensi-
tization following prolonged kinin exposure, whereas
BK1 receptors do not desensitize rapidly and thus 53.5 Protease-Activated Receptors
may promote more prolonged pain and inflamma-
tion. In keeping with this, BK1 receptors are Several proteases, from circulating inflammatory
dramatically unregulated in many tissues, including cells and vascular epithelium, cleave a variety of
sensory neurons (Levy, D. and Zochodne, D. W., protease-activated receptors (PARs) that are asso-
2000) and spinal cord (Fox, A. et al., 2003; Ferreira, ciated with sensory neurons (Vergnolle, N. et al.,
J. et al., 2005), following tissue or ultraviolet injury 2003). PAR2 in particular has been highlighted as a
(Eisenbarth, H. et al., 2004) or traumatic nerve injury regulator of sensory nerve excitability and plays an
and by mediators such as interleukin (IL)-1 or the important role in inflammatory hyperalgesia. Thus
neurotrophin glial cell line-derived neurotrophic PAR2 agonists contribute to neurogenic inflamma-
factor (GDNF; Fox, A. et al., 2003; Valleni, V. et al., tion through the stimulation of peripheral substance
2004). These kinins cause a cascade of secondary P and calcitonin gene-related peptide (CGRP)
changes, including prostanoid production, phosphor- release and to spinal hyperexcitability via release of
ylation of signaling proteins such as PKC, and the these neuropeptides in the spinal cord. PAR2 recep-
sensitization of sensory transducers such as the tor activation directly increases dorsal root ganglion
TRPV1 receptor (Marceau, F. et al., 1998). These (DRG) excitability and sensitizes TRPV1 receptors
events are linked with heat and mechanical hyper- via a PKC mechanism (Amadesi, S. et al., 2004);
algesia (Liang, Y. F. et al., 2001; Fox, A. et al., 2003). thus contributing to inflammation-induced hyperal-
There is an abundance of evidence supporting the gesia. In keeping with this, deletion of the TRPVR1
analgesic potential of BK2 receptor antagonists gene or pretreatment with the TRPV1 antagonist
(Stewart, J. M., 2004). For example, BK2 antagonists capsazepine, abolished PAR2-induced thermal
(icatibant [HOE-140], bradyzide) and BK1 antagonist hyperalgesia, suggesting that TRPV1 is required for
(des Arg10 HOE-140; SSR240612) produce robust PAR2-induced hyperalgesia. PAR1 is also coex-
antihyperalgesic effects in a variety of animal models pressed on peptide-containing DRGs, and although
of inflammatory hyperalgesia (Burgess, G. M. et al., PAR1 agonists induce neurogenic inflammation they
2000; Fox, A. et al., 2003, Gougat, J. et al., 2004) as do not sensitize TRPV1 receptors, nor do they
well as in models of nerve injury-induced hyperalgesia induce hyperalgesia. Overall these data suggest that
(Gabra, B. H. and Sirois, P., 2003). In keeping with this PAR2 antagonists would have antihyperalgesic activ-
deletion of the BK receptor genes reduces immune cell ity (Mantyh, P. W. and Yaksh, T. L., 2001)
chemotaxis, spinal sensitization, as well as heat and
mechanical allodynia following traumatic and diabetic
nerve lesions (Pesquero, J. B. et al., 2000; Ferreira, J. 53.6 Cannabinoids and Their
et al., 2005; Gabra, B. H. et al., 2005). To strengthen the Receptors
translation of BK1 receptor therapy to humans, trans-
genic mice expressing human BK1 receptors, show Two major cannabinoid receptor subtypes, CB1 and
robust inflammatory pain that was attenuated by the CB2 are associated with pain modulation and have
oral administration of the human-selective BK1 received much attention in recent years (reviewed
antagonist NVP-SAA 164 (Fox, A. et al., 2005). by Fox, A. and Bevan, S., 2005). These receptors are
800 Pharmacological Modulation of Pain

widely distributed in the nervous system but CB1 analgesia has been proposed to be driven by activa-
receptors are located in greatest abundance in the tion of descending inhibitory mechanisms. In keeping
CNS and on peripheral sensory neurons. CB2 recep- with this, block of the descending noradrenergic sys-
tors have been found mainly in peripheral tissues tems (Gutierrez, T. et al., 2003) has been proposed to
including immune tissues and keratinocytes account, in part, for CB1-induced antinociception.
(Hohmann, A. G. and Herkenham, M., 1999; Rice, A. Several studies have now shown that CB2 agonists
S. C. et al., 2002) but more recently on some sensory modulation acute pain, for example, AM1241 and
and CNS cells. Several fatty acids, for example, GW405833 (incision model) (LaBuda, C. J. et al.,
anandamide, 2-arachidonylglycerol, and palmitoy- 2005) while GW405833 has been reported to exhibit
lethanolamide, have been identified as endogenous efficacy in neuropathic and inflammatory pain mod-
ligands for these receptors and are termed endocanna- els (Valenzano, K. J. et al., 2005). Interestingly another
binoids. They are released from postsynaptic nerve CB2 agonist, JWH-133 shows antihyperalgesia as
terminals to modulate postsynaptic excitability and well as anti-inflammatory activity (Elmes, S. J. R.
transmitter release at presynaptic elements via et al., 2005). The effects of CB2 agonists appear not
Gi/Go-coupled activity. Specific antagonists such as to be due to the release of endogenous opioids as
SR141716A and SR147778 for CB1 and SR144428 for actions were unaffected by naltrexone (Whiteside, G. T.
CB2 have been used to characterize receptor et al., 2005b).
functions. Several clinical studies have supported the use of
For the most part CB1 receptors are located on cannabinoids, such as tetrahydrocannabinol (THC),
neurons where activation causes attenuation of pre- to reduce pain. However this action is commonly
synaptic N-calcium channel activity and inhibition of accompanied by adverse effect such as euphoria,
transmitter release (Kreitzer, A. C. and Regeher, W. dizziness, and sedation (Campbell, F. A. et al., 2001;
G., 2002). In contrast, activation of neuroglia (astro- Wade, D. T. et al., 2003; Svenden, K. B. et al., 2004).
cytes, microglia) is accompanied by de novo The reduction of CNS side effects has been
expression of both CB1 and CB2 and increased approached through targeting peripheral CB recep-
release of endocannabinoids (Stella, N. 2004). tors, since both CB1 and CB2 receptor activation
Specifically CB2 receptors are expressed in spinal produce antinociception. In support of this approach,
microglia following peripheral nerve lesions but not the systemic administration of WIN55,212-2, a non-
by peripheral inflammation (Zhang, J. M. et al., 2003). selective CB agonist, attenuated inflammatory
The significance of this is presently unclear although hyperalgesia and this action was reversed by peripheral
prolonged CB receptor activation has been associated (intraplantar) but not spinal administration of a CB1
with increased expression of neuroprotective modu- antagonist (Fox, A. and Bevan, S., 2005). Moreover a
lators such as brain-derived neurotrophic factor peripherally restricted CB1 agonist (e.g., NVP-001,
(BDNF; Marsicano, G. et al., 2003). personal communication) was shown to reduced
Both CB1 and CB2 receptor types have been inflammatory hyperalgesia by directly reducing noci-
shown to have a key role in the modulation of noci- ceptor excitability (Richardson, J. D. et al., 1998).
ceptors. Although CB1 effects have been best Finally, CT-3 (ajulemic acid) a nonspecific CB1 and
characterized, the mechanism and sites of action are CB2 agonist, with limited CNS availability produced
incompletely understood. It is likely that an interplay analgesia in a number of inflammatory and neuropathic
of several CB1-driven systems occurs during acute or pain models (Burstein, S. H. et al., 2004; Dyson, A. et al.,
chronic pain. Brainstem structures appear to be 2005) as well as in human neuropathic pain, at doses
important. For example, CB1-induced local activa- which cause minimal CNS side effects (Karst, M. et al.,
tion of the periaqueductal gray (PAG; by stress- 2003). The mechanism of action of CT-3 appears to
induced release of endocannabinoids) has been sug- involve mainly CB1 receptors and a number of other
gested to be involved in aversive responses and acute indirect actions including activation of peroxisome
stress-induced analgesia (Finn, D. P. et al., 2004; proliferator-activated receptor (PPAR)- receptors
Hohmann, A. G. et al., 2005). This was blocked by and inhibition of inflammatory cell activity and med-
SR141716A (rimonabant, the CB1 antagonist) but not iator release (Liu, J. et al., 2003).
by naltrexone (-opioid antagonist). Paradoxically, Selective CB2 ligands (HU-308, AM-1241, and
recent data suggest that deletion of the CB1 receptor GW405833) as well as nonselective compounds
in mice is associated with a greater vulnerability to (HU-210 and CP55940) have shown antinociceptive
stress (Fride, E. et al., 2005). Finally CB1-induced effects in a variety of inflammatory and neuropathic
 Pharmacological Modulation of Pain 801

pain models (Malan, T. P. et al., 2003, Valenzano, K. J. 53.7 Mrg-Related GPCR and Their
et al., 2005). These effects are still present in CB1- Ligands
knockout (KO) mice, absent in CB2-KO animals, can
be revered by CB2 antagonist such as SR144528, and The mas-related gene (Mrg) family is a large family
occur without major CNS side effects (sedation, cat- of GPCRs which vary in numbers depending on
alepsy, or motor impairments). It is unclear how species, are rather exclusively expressed in subsets
these effects are produced since CB2 receptors have of small sensory neurons (Dong, X. et al., 2001), and
not been found on sensory neurons (Sokal, D. M. may be co-localized with markers for nociceptive
et al., 2003). Rather an indirect effect via immune neurons such as TRPV1 (Lembo, P. M. C. et al.,
cell modulation and the naloxone-reversible release 2002). Selective manipulations of individual Mrgs
of -endorphin from keratinocytes has been pro- have provided exquisite markers for subsets of small
posed (Ibrahim, M. M. et al., 2005). In contrast, a sensory neurons showing selective epidermal and
number of reports have suggested that CB2 agonists, spinal innervation, suggesting a close anatomical
for example, JWH-133, may reduce C-fiber excit- and functional relationship (Zylka, M. J. et al., 2005).
ability directly (Ross, R. A. et al., 2001; Sagar, D. R. Focus on the human mrgX subfamily (six genes in
et al., 2005). There is also a possibility that CB2 humans), also called sensory neuron-specific recep-
agonists cause antinociception via the CNS as CB2 tors (SNSR), has revealed a complex pharmacology.
receptors are expressed in spinal microglia after The endogenous ligand for SNSR is unconfirmed,
nerve injury (Zhang, J. M. et al., 2003), and CB2 but several mammalian peptide ligands have been
activation may modulate microglial activity (Water, identified with high affinity for this receptor. These
L. et al., 2003). Indeed the direct spinal administration peptides include bovine adrenal medullary peptide
of JWH-133 attenuated mechanical allodynia (Sagar, 22 (BAM22), and its fragment BAM8-22 derived
D. R. et al., 2005). Clearly greater understanding is from the endogenous opioid precursor pro-enkephalin
A, as well as two unrelated peptides 2-melanocy-
still required with respect to the mechanisms of CB2-
tostimulating hormone (MSH) and CT-2-MSH.
mediated analgesia.
Apart from BAM22, these substances do not activate
Other approaches to address the role of endogen-
opioid receptors, rather they increase peripheral and
ous cannabinoid systems in pain and analgesia have
spinal excitability to potentiate thermal and
targeted fatty acid amide hydrolysis (FAAH), the
mechanical nociception (Grazzini, E. et al., 2004).
major degradation pathway for endogenous canna-
This suggests that the same gene products can facil-
binoids (Cravatt, B. F. and Lichtman, A. H., 2004).
itate as well as inhibit sensory neurons and that an
Thus in mice lacking this enzyme (Cravatt, B. J.
SNSR antagonist should be sought to inhibit hyper-
et al., 2001), or treatment of nave mice with a
algesia. Although these data strongly support a role
novel FAAH inhibitor such as OL135 (Lichtman,
for SNSRs in the modulation of pain, the endogen-
A. H. et al., 2004), increases the analgesic efficacy ous ligand for this receptor has not been confirmed
of anandamide and significantly elevated brain ana- and there is little direct evidence linking SNSRs in
ndamide levels were measured. In addition, OL135 the etiology of acute or chronic pain.
increased pain threshold in acute pain models,
decreased the mechanical allodynia of neuropathic
pain, and decreased thermal hyperalgesia in models
of persistent pain (Chang, L. et al., 2006). Surprisingly 53.8 Prostanoids and Receptors
a CB2 rather than CB1 antagonist attenuated the effect
of OL135 in the persistent pain models and its effects A variety of prostanoid cyclo-oxygenase (COX)
appeared to be mediated by a naloxone-reversible enzyme products (PGE2, PGD2, PGF2, thrombox-
mechanism. In this latter respect several reports have ane, PGI2) occur during inflammation but PGE2 is
indicated analgesic synergy, between -opioid and CB considered to be the major contributor to inflamma-
receptors. Thus combinations of these agonist have tory pain. Thus blocking the major synthetic
been shown to provide pain reduction with minimal enzymes, COX-1 (constitutive) and COX-2 (induci-
side effects in acute pain models (Cichewicz, D. L. and ble), or inhibition of prostanoid receptors continue to
McCarthy, E. A., 2003). However, it is still unclear be important approaches for reducing inflammatory
whether such synergy can be exploited in chronic pain (Flower, R. J., 2003). Experience with selective
pain treatment. inhibitors of COX-2 (celecoxib, rofecoxib) shows
802 Pharmacological Modulation of Pain

improved gastrointestinal tract safety but little inflammation, peripheral nerve injury, or by admin-
improvement in analgesic efficacy. A relatively infre- istration of cytokines, leading to increased
quent though increased cardiovascular risk (coronary production of spinal PGE2. In keeping with this,
constriction) has been observed with some COX several NSAIDs have been shown to reduce inflam-
inhibitors. The mechanism is not understood but matory hyperalgesia via inhibition of spinal COX
has led to Vioxx (rofecoxib) being withdrawn from activity (Yaksh, T. L. et al., 2001). Several mechanisms
therapy and considerable clinical debate about the have been proposed for the PGE2-induced increase
safety of future approaches to COX inhibition. in spinal excitability. Prominent are the contribution
An alternative approach for the clinical improve- of EP1 receptors and spinal release of glutamate.
ment of COX inhibitors has been to combine COX Recently however the spinal effects of PGE2 have
inhibition with NO donation (CINOD). Such mole- been linked with a glycine receptor. Thus deletion of
cules (NO-naproxene, NO-ibuprofen) have been the GlyR3 subunit gene, reduced pain sensitivity
claimed to show improved efficacy and safety over caused by PGE2 administration or inflammation
the parent nonsteroidal anti-inflammatory drugs (Harvey, R. J. et al., 2004).
(NSAIDs) due to actions (improved side effects and Finally, PGE2 exerts its effects via a variety of EP
efficacy) of cleaved NO (Fiorucci, S. et al., 2001). receptors (EP1, 2, 3 4) present both in peripheral
Recently a splice variants of COX-1 has been sensory neurons and in the spinal cord. Activation
identified, COX-3 (Chandrasekharan, N. V. et al., of these receptors produces a complexity of effects,
2002). However it has low enzymic capability and ranging from Ca2 influx to cAMP activation or
its distribution and low abundance in the CNS and in inhibition. In the periphery, sensitization by PGE2
periphery does not make this a compelling target for as been shown to be cAMP mediated causing the
analgesia. However several NSAIDs (acetamino- enhancement of tetrodotoxin-resistant (TTX-r)
phen, diclofenac, phenacetin) show low efficacy sodium currents in nociceptors via channel phos-
but some degree of selectivity for COX-3. Overall phorylation (England, S. et al., 1996; Gold, M. S.
however the analgesic efficacy of acetaminophen is et al., 1998). However in the spinal cord, excitability
poorly correlated with COX inhibition. Interestingly was enhanced by EP1 receptors but reduced by an
endogenous acetaminophen metabolism produces EP3 agonist (ONO-AE-248) suggesting further
a conjugated arachidonic acid derivative (AM404) complexity in the prostanoid regulation of pain
that shows cannabinoidlike analgesia properties mechanisms (Bar, K. J. et al., 2004).
(Hogestatt, E. D. et al., 2005; Ottani, A. et al., 2006).
Furthermore gene deletion studies suggested that
COX-3 inhibition may be linked with the antipyretic 53.9 Cytokines, Chemokines, and
and analgesia properties of NSAIDs. Clearly further Their Receptors
studies are needed to link this enzyme to inflamma-
tory pain pathophysiology. Cytokines are produced by a variety of immune cells
Another route of inhibiting PGE2 synthesis is via as well as brain neuroglial cells in response to injury
the blockade of PGE synthase (PGES), a major route and inflammation. They are powerful mediators of
of conversion of PGH2 to PGE2. Two isoforms of the hyperalgesia (see Abbadie, C., 2005).
enzyme have been identified, membrane or micro- Probably the most characterized are IL-1 and
somal-associated PGES (mPGES-1) and a cytosolic tumor necrosis factor (TNF)- that act via specific
enzyme (cPGES/p23) which are linked with COX- receptors on sensory neurons. Their effects can be
2- and COX-1-dependent PGE2 production, respec- attenuated by receptor antagonists (IL-1-r) that
tively (Jakobsson, P. J. et al., 1999; Claveau, D. et al., sequester the ligand as well as by neutralizing anti-
2003). Both isoforms are upregulated by inflamma- bodies. Indeed the TNF antibody, etanercept, has
tory mediators and gene deletion studies in mice been developed for the treatment of chronic inflam-
indicate an important role for mPGES in acute and mation and the presence of TNF- has been
chronic inflammation and inflammatory pain correlated with a number of painful inflammatory
(Trebino, C. E. et al., 2003). clinical conditions (Lindenlaub, T. and Sommer, C.,
Apart from prominent roles in regulating periph- 2003). Cytokines induce hyperalgesia by a number of
eral excitability during inflammatory pain, COX-1 direct and indirect actions. Thus IL-1 activates
(glia) and COX-2 (ventral horn cells) are constitu- nociceptors directly and produces heat sensitization
tively present in spinal cord and are increased by via intracellular kinase activation, but it may also
 Pharmacological Modulation of Pain 803

cause indirect nociceptor sensitization via the pro- behavioral signs of hyperexcitability (Watkins, L. R.
duction of kinins and prostanoids (Sommer, C. and and Maier, S., 2002; Raghavendra, V. et al., 2003).
Kress, M., 2004). TNF- also activates sensory neu- Chemokines are important peripheral and central
rons directly via TNFR1 and TNFR2 receptors, mediators of inflammation. The major chemokines
(Pollock, J. et al., 2002; Ohtori, S. et al., 2004) and and their respective receptors are macrophage-
initiates a cascade of inflammatory reactions through derived chemokine (MDC)/CCR4, regulated upon
the production of IL-1, IL-6, and IL-8. It is significant activation, normal T-cell expressed, and secreted
that direct TNF- application in the periphery (RANTES)/CCR5, fractalkine/CX3CR1, and SDF-
induces neuropathic pain behavior that is blocked 1/CXCR4. Receptors are located on leukocytes, on
by ibuprofen and celecoxib (Schafers, M. et al., central neurons, sensory neurons, and neuroglial cells
2004), while nerve ligation causes increased TNF- (Watkins, L. R. and Maier, S., 2002). Apart from their
in damaged as well as adjacent undamaged axons major chemoattractant effects, chemokines contri-
(Schafers, M. et al., 2003a). Interestingly anti-TNF- bute directly to inflammatory hyperalgesia through
 treatment with TNF antibody, adalimumab, pro- G-protein-coupled activation and sensitization of
duced a prolonged reduction of pain symptoms in sensory neurons (Oh, S. B. et al., 2001). Block of
OA (Grunke, M. and Schulze-Koops, H., 2006). CX3CR1 by a fractalkine receptor-neutralizing anti-
There appears to be a complex interplay between body induces antiallodynic effects in models of
cytokines, and it should be noted that like other peripheral nerve inflammation (Milligan, E. D. et al.,
mediators of pain not all should be considered as 2005b). However some chemokines appear beneficial.
detrimental. Thus IL-6 has been associated with Thus gene therapy (AV-333) has been used to
potential beneficial effects after nerve injury includ- increase IL-10 production through the delivery of
ing protection against cell death and the promotion viral and nonviral vectors by acute spinal intrathecal
of growth. However there is also compelling evi- delivery. This experimental treatment has been
shown to reverse mechanical allodynia in the chronic
dence of a strong association with this cytokine in
constriction injury (CCI) model of neuropathic pain
several human pain conditions including herniated
(Milligan, E. D. et al., 2005a).
lumbar disc pain and pain caused by failed back
surgery (De Jongh, R. F. et al., 2003).
A variety of studies have demonstrated an impor-
tant role for spinal inflammatory and neuroimmune
53.10 Adrenoceptors
processes triggered by peripheral inflammation and
nerve injuries. These processes involve the regula-
A number of chronic pain disorders termed sym-
tion of a variety of receptors, channels, and enzymes
pathetically maintained pain have highlighted the
with patterns that are likely to differentiate one pain importance of the release of sympathetic transmitters
state from another (Honore, P. et al., 2000). In addi- (epinephrine or norepinephrine) from sympathetic
tion, activation of spinal neuroglial cells (microglia, varicosities and the involvement of adrenergic recep-
astrocytes, satellite cells) stimulates a cascade of sec- tors in pain etiology. For example following
ondary excitability changes (Watkins, L. R. and peripheral nerve injury sprouting of sympathetic
Maier, S., 2002). Neuroglia make close-junctional nerve endings occurs at several sites. Thus sympa-
connections with other cells, providing a means of thetic/sensory coupling at the level of the DRG
spreading excitability changes beyond the bound- (Zhang, J. M. et al., 2004), at the site of injury (neu-
aries of spinal segmental input. Neuroglia also roma), and in the periphery (Shinder, V. et al., 1999)
secrete a number of mediators such NO, neurotro- have been demonstrated. In keeping with this some
phins, IL-1, TNF-, free radicals, and glutamate. In neuropathic pain symptoms have been attenuated by
addition, neuroglial mediators may contribute to sympathetic blocks or surgical sympathectomy.
spinal excitability by causing dysfunction or degen- Other studies have shown the expression of 1-
eration of inhibitory spinal neurons (Moore, A. K. and 2-adrenoceptors on sensory neurons or post-
et al., 2002). In keeping with an important role for ganglionic sympathetic terminals (Sato, J. and Perl, E.
neuroglial mediators, treatments with anti-inflamma- R., 1991; Tracey, D. J. et al., 1995; Lee, D. H. et al.,
tory agents or modulators of neuroglial activity such 1999) after nerve injuries. Under these conditions
as propentofylline and minocycline inhibit glial acti- sensory neurons can be directly activated by the
vation and the release of glial products with reduced endogenous release of sympathetic transmitters (via
804 Pharmacological Modulation of Pain

1 receptors) or in the clinic by intradermal injection excitability during chronic pain and repetitive stimu-
of norepinephrine (Ali, Z. et al., 2000). lation of pain pathways. Indeed NMDA antagonists
On the other hand, transdermal applications of the show robust attenuation of pain behaviors but pro-
2-agonist, clonidine via transdermal patches or voke a number of side effects (sedation, confusion,
creams, has proven efficacious in a variety of neuro- and motor incoordination) and thus have an insuffi-
pathic pain conditions (Byas-Smith, M. G. et al.,1995). cient therapeutic window. There has been a refocus
Since clonidine has been shown to decrease the excit- on more specific NMDA-receptor subtype blockers
ability of small sensory neurons from nerve injured (NR1 and NR2) directed toward the glycine modu-
animals (Ma, W. et al., 2005) the antinociceptive latory site to avoid side effects. This site actively
efficacy of clonidine is considered to be due to 2 modulates the NMDA channel only during the sus-
receptor-coupled inhibition. Clonidine and other tained stimulation of the receptor, which is
2-agonists such as dexmedetomidine have also been considered to occur during chronic pain. Selective
used systemically to inhibit of sensory transmission in NR1-Gly antagonists have been claimed to reduce
the spinal cord by block of pre- and postsynaptic pain with reduced side effects (Danysz, W. and
membrane excitability. Unfortunately sedation and Parsons, C. G., 1998; Quartaroli, M. et al., 2001).
hypotension are major target-related side effects of However clinical experience has not yet confirmed
these compounds. Great efforts have been made to this. GV196771 did not show efficacy against clinical
identify ligands with improved 2-receptor subtype pain, possible due to inadequate penetration into the
selectivity, to avoid side effects, but thus far this has CNS (Wallace, M. S. et al., 2002).
not been particularly successful. Alternative initiatives have targeted other NMDA
receptor subtypes such as the NR2B receptor, which
has a specific distribution in sensory pathways.
53.11 Glutamate Regulation and Blockade of this receptor has also been claimed to
Glutamate Receptors produce antinociception (ifenprodil, CP-101,606)
with reduced side effects (Taniguchi, K. et al., 1997).
Glutamate is the major excitatory neurotransmitter This concept has yet to be evaluated in the clinic.
in the CNS with important regulatory roles for pain Metabotropic glutamate receptors, particularly
transmission. However there is also considerable evi- mGluR1 and mGluR5, have been reported to play a
dence that it plays a role in pain processing in the key role in sustaining heightened central excitability
periphery (Carlton, S. M. et al., 1995; Bahave, G., et al., in chronic pain with minimal involvement in acute
2001). Glutamate acts through receptor-coupled nociception. Thus spinal administration of selective
ligand-gated ion channels (-amino-3-hydroxy-5- agonists such as dihydroxy phenyl glycine produced
methyl-4-isoxazolepropionic acid (AMPA)/kinate allodynia, while mGluR5 was shown to be signifi-
receptors: iGluRs) and GPCR-coupled receptors cantly overexpressed in some, but not all, chronic
(metabotropic glutamate receptors (mGluRs)). pain models (Hudson, L. J. et al., 2002). Furthermore
Injections of glutamate or metabolically stable antisense-oligonucleotide and antibody treatments,
receptor-selective agonists such as N-methyl-D- directed at these receptors, reduce chronic inflamma-
aspartic acid (NMDA), AMPA, and kainate, cause tory and neuropathic pain behaviors (Fundytus, M. E.
a reduction in thermal and mechanical thresholds et al., 2002). In keeping with these observations, the
for pain, while application of iGluR and mGluR mGluR5-selective antagonists 2-methyl-6-(phenyl-
antagonists attenuate pain in acute models (Zhou ethynyl)-pyridine (MPEP) did not reduce acute
et al.,1996; Bhave, G., et al., 2001). physiological pain responses but reduced heat hyper-
Building upon this has been significant progress in algesia and allodynia in models of inflammatory and
development of a glutamate antagonist optimized for neuropathic pain (Walker, K. et al., 2001; Hudson, L.
clinic testing. For example, the AMPA-kinate receptor J. et al., 2002; Urban, M. O. et al., 2003).
antagonist, LY293558 has been shown to have efficacy Peripheral mGluR5 receptors have also been
in models of neuropathic pain (Blackburn-Munro, G. claimed to modulate pain (Jang, J. H. et al., 2004).
et al., 2004) and to reduce capsaicin-induced pain in Thus local administration of mGluR5 (MPEP,
human volunteers as well as clinical dental pain with SIB1757) have been effective in reducing pain beha-
minimal side effects (Sang, C. N. et al., 1998). vior (Dogrul, A. et al., 2000; Zhu, C. Z. et al., 2004)
There has been a great deal of information linking suggesting a potential for using peripheral mGlu5
NMDA receptors in central sensitization and CNS antagonists in pain therapy.
 Pharmacological Modulation of Pain 805

Metabotropic group II receptors (mGluR2 and ion gradient in afferent terminals. This allows the out-
mGluR3) also modulate pain transmission. The flow of chloride ions associated with the primary
mGluR2 is located in sensory neurons and presynap- afferent depolarization (PAD) and presynaptic inhibi-
tic nerve terminals whereas mGluR3 is found all tion, following GABAA receptor activation of afferent
over the brain. MGluR3 can be selectively increased terminals. Deletion of the NKCC1 gene induced
in the spinal dorsal horn neurons after peripheral touch-evoked pain caused by light stroking. This was
ultraviolet injury (Boxall, S. J. et al., 1998). mGluR2/3 shown to be due to increased nerve terminal excitabil-
receptor activation appears necessary to reduce ity caused by a reduced capability to generate
nerve terminal excitability and modulate pain trans- presynaptic inhibition (Laird, J. M. A. et al., 2004).
mission since treatment with L-acetyl-carnitine On the other hand increasing the expression of
reduced inflammatory hyperalgesia and mechanical this cotransporter or increasing its efficacy through
allodynia and increased the expression of mGlur2/3. AMPA mediated Ca/CaM-kinase II phosphorylation
The effect of L-acetyl-carnitine were attenuated has been postulated (Cervero, F. et al., 2003) to
by LY379268, a mGlu2/3 antagonist (Chiechio, S. increase PAD sufficient to reach the firing threshold
et al., 2002). of the nerve terminal. This mechanism may also
contribute to spinal sensitization following nerve
injury and may indeed generate antidromic action
53.12 Gamma-Aminobutyric Acid potentials triggering a dorsal root reflex or the ecto-
Receptors pic discharges in sensory nerves which cause, or
contribute to spontaneous pain.
Gamma-aminobutyric acid (GABA) receptors are Spinal sensitization that occurs following periph-
emerging as important regulators of pain, particularly eral nerve injury may also arise by other mechanisms
in the spinal cord as they are abundantly expressed of spinal disinhibition. This has been hypothesized to
both on spinal afferent nerve terminals and spinal involve the reduced expression of the potassium-
neurons. Activation of both subtypes of GABA recep- chloride exporter KCC2, seen only in superficial
tor by muscimol (GABAA agonist) or baclofen spinal lamina 1 neurons after injury. In addition,
(GABAB agonist) reduced pain behavior. In contrast, knockdown of KCC2 with antisense or block of this
GABAA (bicuculline) and GABAB (CGP35348) transporter with ((dihydronindenyl)oxy) alkanoic
antagonists cause pain when injected into the spinal acid (DIOA) causes a similar shift in the transmem-
intrathecal space (Malan, T. P. et al., 2002). Loss of brane ionic gradient in spinal lamina 1 neurons and a
GABA (Ibuki, T. et al., 1997) and impaired GABA- consequent behavioral hyperalgesia resembling that
mediated inhibition through loss of spinal interneur- seen after nerve injury (Coull, J. A. M. et al., 2003). In
ons has been demonstrated in models of neuropathic addition, normal spinal inhibitory currents, mediated
pain (Moor, A. K., et al., 2002). by GABA and glycine interneurons, are reversed in
Improvement of GABA synthesis through promo- the absence of KCC2, so that the effects of GABA
tion of its synthetic enzyme, glutamic acid become predominant excitatory, due to the outward,
decarboxylase (GAD), has been achieved by gene rather than inward, flow of chloride ions. The emer-
transfer of a GAD promoter (in herpes simplex ging view suggests that a variety of changes in ionic
virus vector) via peripheral injection. This was regulation occur during chronic pain. This adds com-
taken up into sensory neurons and transported to plexity to the puzzle of pain but opportunity for
DRGs to reduce spinal excitability, as well as intervention. It will be important to learn how these
mechanical and thermal allodynia caused by nerve disinhibitory mechanisms contribute in different
injury (Hao, S. et al., 2005). chronic pain conditions and whether they are critical
Other important changes in ionic regulation have for pain initiation as well as its maintenance.
been highlighted to occur in chronic pain which indir-
ectly affects GABA- and glycine-mediated inhibition
and excitability in the spinal cord. In particular changes 53.13 Ion Channels
in transmembrane ion transporters have been associated
53.13.1 Ligand-Gated Channels
with modulation of primary afferent excitability and
related to neuropathic pain disorders. Thus the chloride Transient receptor potential (TRP) channels form a
transporter NKCC1, localized in primary sensory neu- large family of sensory transducers involved in cel-
rons, is responsible for maintaining the high chloride lular calcium regulation. Many are localized to
806 Pharmacological Modulation of Pain

mammalian sensory nerves and are involved in the emergence of a novel therapy. Indeed recent clinical
transduction of temperature and chemical signals studies in human volunteers have shown that oral
that result in pain. The TRPV family has received SB705498 attenuated capsaicin and ultraviolet C
considerable attention and is considered a good tar- (UVC)-induced pain and hyperalgesia (Chizh, B.
geted for developing analgesics (Krause, J. E. et al., et al., 2006).
2005). TRPV1, originally called the vanilloid recep- Other TRP channels (TRPV3, TRPV4, TRPA1)
tor because of its selective activation by capsaicin and have been suggested to be involved in transduction.
pungent vanilloids, has the properties of a noxious Thus TRPA1 (ANKTM1) is colocalized with
heat transducer. The effects of capsaicin on sensory TRPV1, is activated by capsaicin and mustard oil
neurons are complex, involving activation through but can also be sensitized by inflammatory mediators
ligand-operated cation channels, and the extracellu- including bradykinin to produce cold-induced burn-
lar influx as well as the intracellular increase in ing pain (Bandell, M. et al., 2004). In addition TRPV1
calcium ions. Calcium-induced stimulation of phos- can oligomerize with other TRP family members
phatase such as cacineurin then reduces excitability including TRPV3. TRPV3 is also a heat transducer
via inactivation voltage-gated calcium (CaV) chan- that it sensitive in the physiological temperature
nels as well as rapid CaV2.2 internalization (Wu, Z. range and responds to noxious heat, but is insensitive
Z. et al., 2005). This mechanism explains the transient to capsaicin. This is found in keratinocytes and
and reversible analgesia effects of capsaicin either on appears to be upregulated in inflammatory pain con-
peripheral or spinal afferent nerve terminals. TRP ditions. It is unclear how activity of TRPV3 leads to
channel regulation of heat transduction is complex as nociceptor activation. So far there are few chemical
animals with deleted TRPV1 and TRPV2 genes tools to help characterize the functions of these TRP
appear to have normal heat sensitivity (Woodbury, receptors and support their value as analgesia targets.
J. et al., 2004) whereas gene deletion of TRPV1 pre-
vented inflammatory heat hyperalgesia (Caterina, M.
J. et al., 2000). 53.14 Purinergic Receptors
The TRPV1 receptor has assumed an increasingly
53.14.1 P2X Receptors
prominent role in regulating sensory neuronal excit-
ability in inflammatory pain. A variety of The unique localization of the P2X3 receptor to
inflammatory agents including protons, bradykinin, small sensory fibers has highlighted its importance
adenosine triphosphate (ATP), PGE2, 12-lipoxygen- in pain. Large amounts of the endogenous ligand,
ase products, PAR2, anandamide, and nerve growth ATP are released after tissue injury and during
factor (NGF) indirectly sensitize TRPV1, or regulate inflammatory injuries while both ATP and a stable
its expression, to cause thermal hyperalgesia. A com- analogue ,-Me ATP, induce pain and are prono-
mon mechanism appears to be GPCR-coupled ciceptive when administered intradermally in human
stimulation of PLC, the formation of intermediates volunteers.
inositol triphosphate (IP3) and DAG and the activa- In chronic inflammatory pain, P2X3-mediated
tion and mobilization of PKC (Vellani, V. et al., 2001) excitability is enhanced while reduction of P2X3
which can phosphorylate the TRPV1 receptor. receptors, by antisense oligonucleotide administra-
Earlier analgesia strategies, targeting TRPV1, tions, reduced inflammatory hyperalgesia as well
were focused on capsaicin-like agonists that induced as that evoked by ,-Me ATP (Honore, P. et al.,
functional inactivation of sensory fibers by causing 2002). In keeping with this a number of antagonists
reversible subepidermal degeneration. This has been including 29,39-O-(2,4,6-trinitrophenyl) adenosine
successfully translated into the clinic with the intro- 59-triphosphate (TNP-ATP), pyridoxal-phosphate-
duction of a number of topical capsaicin therapies for 6-azophenyl-29,49-disulfonate (PPADS), and suramin,
inflammatory pain. Currently there is a focus on reduce pain behavior. More selective, and druglike,
TRPV1 channel blockers or selective antagonists antagonists such as A-3174919 reduced pain in a num-
against the TRPV1 receptor (Garcia-Martiez, C. ber of acute and chronic pain models supporting the
et al., 2002). Supporting these approaches, competi- possibility for future analgesia therapy (Jarvis, M. F.
tive (AMG-9810, personal communication) and et al., 2002).
noncompetitive vanilloid receptor-1 (VR1) antago- It should be noted that a number of other puri-
nists (DD161515; Sachez-Baez, F. et al., 2002) block nergic receptor subtypes, for example, P2X4 and
chemical and thermal pain sensitivity, heralding the P2X7, have also been suggested to modulate pain
 Pharmacological Modulation of Pain 807

through altered central excitability and the release of stinging pain produced by intradermal injections of
neuroglial-cell products. Thus activated microglial, acidic solutions and low extracellular pH enhancing
astrocytes, and satellite cells release a variety of the effects of other inflammatory mediators (Krishtal,
inflammatory mediators including IL-1, TNF-, O., 2003; Mamet, J. et al., 2003).
prostanoids, and NO upon ATP stimulation. Indeed Exogenously administered acidic solutions pro-
increased expression of P2X4 has been shown to duce a rapid but transient increase in membrane
occur in spinal microglial after peripheral nerve cation permeability as well as a more prolonged
lesions and this was related to painful mechanical permeability increase in sensory neurons. This can
allodynia. This behavior was blocked by spinal give rise to sustained nerve activation as well as an
administrations of the selective P2X4 antagonist enhanced mechanosensitivity. The mechanism of
TNP-ATP (Tsuda, M. et al., 2003). Remarkably proton-induced activation of sensory neurons under-
spinal administration of activated microglia repro- lying pain has not been fully elucidated, but appears
duced TNP-ATP sensitive mechanical allodynia in to be driven by a number of acid-sensing ion channel
nave animals. (ASICs) particularly ASIC1 and ASIC3 (also called
Increased P2X7 expression has been found in per- DRASIC). ASIC3 has been shown to have a strong
ipheral macrophages following inflammation but this antinociceptive phenotype following deletion of this
receptor is also expression in spinal neurons and gene in KO mice (Sluka, K. A. et al., 2003).
microglia following peripheral nerve injury A novel blocker (A-317567) of peripheral ASIC1,
(Deuchars, S. A. et al., 2001). In keeping with an ASIC2, and ASIC3 channels has been described
important role in chronic pain both microglia and (Dube, G. R. et al., 2005). This produces antihyper-
P2X7 receptors are upregulated in human chronic algesia in models of inflammatory and postoperative
pain patients while deletion of the P2X7 receptor pain but there have been no reports of therapeutic
gene produced a complete absence of mechanical advances with more selective inhibitors.
and thermal pain in mice (Chessell, I. P. et al., 2005).
It is worth noting that other nucleotide-gated ion
channels have also been shown to be important for 53.15 Sodium Channels
regulating peripheral excitability. Thus the sodium/
potassium repolarizing pacemaker current, Ih, which Voltage-gated sodium channels are characterized by
is activated during membrane hyperpolarization is their primary structure and sensitivity to tetrodotoxin
important for generation of rhythmic and spontaneous (TTX). A variety of TTX sensitive (NaV1.1, Nav1.2,
action potentials in sensory neurons following nerve NaV1.6, and Nav1.7) and TTX insensitive (Nav1.8,
injury. Ih currents are controlled by cyclic nucleotides NaV1.9) channels are involved in regulating sensory
(cAMP and cGMP) via a family (HCN14 channels) neural excitability (Matzner, O. and Devor, M., 1994;
of ligand-gated ion channels that are constitutively Eglen, R. M. et al., 1999). Changes in the expression,
expressed in sensory nerves and differentially distrib- trafficking, and redistribution of NaVs, following
uted after crush or inflammatory nerve injuries inflammation or nerve injury is considered to account
(Chaplan, S. R. et al., 2003; Yao, H. et al., 2003). for the abnormal firing (ectopic generators) of afferent
Nerve injury has been shown to enhance the Ih and nerves (Devor, M., 2005). However channel redistri-
this can be blocked with ZD7288 which also prevents bution appears complex as most channels appear to be
repetitive firing in damaged sensory neurons and downregulated in DRGs after nerve injury whereas,
reverses touch hypersensitivity in neuropathic pain for example, NaV1.8 is redistributed along small
models (Chaplan, S. R. et al., 2003). This approach axons. It has also been important to recognize that
clearly has great potential for addressing peripheral channel expression and nerve excitability are changed
excitability in neuropathic pain disorders. dramatically in uninjured axons that are closely
apposed to injury (Gold, M. S. et al., 2003).
The TTX-R sodium channel, NaV1.8, is uniquely
53.14.2 Acid-Sensing Channels
expressed in all sensory neurons (IB4 positive and
Proton production is increased in inflammation and NGF sensitive) and appears to be an important con-
is likely to be involved in inflammatory hyperalgesia tributor in the generation of abnormal excitability in
and in the sensation of muscle aching and discomfort sensory axons. Thus knockdown of NaV1.8 produces
due to the hypoxia/anoxia of muscle exercise. Indeed a marked reduction in abnormal pain responsiveness
direct activation of nociceptors accounts for the sharp in pain models (Lai, J. et al., 2003).
808 Pharmacological Modulation of Pain

Inflammation also causes the overexpression of is unclear whether they will impact on nerve excit-
NaV1.7 in several types of sensory neurons in models ability associated with specific pain etiology.
of inflammatory pain (Gould, H. J. et al., 2004) and in NaV1.3 is another ion channel that is dramatically
inflamed human tooth pulp. Interestingly NaV1.7 regulated after injury. This channel has been found
overexpression can be prevented by pretreatment abundantly in fetal but not in adult tissue. However
with COX-1 and COX-2 inhibitors (ibuprofen, NS- NaV1.3 is dramatically upregulated in adult sensory
398). neurons and spinal cord following peripheral and
The clinical utility of nonselective sodium chan- CNS injury (Hains, B. C. et al., 2004). The expression
nel blockade in pain treatment has been well of NaV1.3 has been suggested to make an important
established with the use of local anesthetics such as contribution to the sustained and high-frequency fir-
lidocaine. Clinical experience has confirmed that ing found in injured afferents related to neuropathic
local block of abnormally active afferents reduces pain (Cummins, T. R. and Waxman, S. G., 1997; Kim,
secondary pain in established neuropathic pain con- C. H. et al., 2002). Further it has been proposed that
ditions (Gracely, R. et al., 1992). Interestingly this can NaV1.3 expression may not be directly related to
be achieved at lower concentrations than are neural injury as it was not correlated with activating
required for block of conduction in quiescent fibers transcription factor (ATF)-3 expression, a neuronal
highlighting a potential therapeutic advantage for cell death marker (Lindia, J. A. et al., 2005), but caused
selective fiber block (Devor, M. et al., 1992). The by a deficiency of growth factor. In this respect,
basis of this is due to the state-dependent increase GDNF administration reduced NaV1.3 expression
in the affinity of channel blockers for ion channel and attenuated neuropathic pain behavior (Boucher,
proteins (Devor, M., 2005). An additional utility of T. J. et al., 2000). In contrast, antisense treatments that
lidocaine is that intravenous administration has been significantly reduced sensory neuronal NaV1.3
reported to produce long-lasting pain relief both in expression in the spared nerve injury (SNI) model of
animal models (Araujo, M. C. et al., 2003) and in neuropathic pain did not attenuate nerve injury-
induced allodynia (mechanical or cold; Lindia, J. A.
intractable neuropathic pain (Kastrup, J. et al., 1987).
et al., 2005) whereas this was reduced by the nonselec-
The major disadvantages of nonselective sodium
tive channel blockers mexiletine and lamotrigine.
channel blockers are cardiotoxicity and CNS seda-
tion and confusion produced by NaV1.5 and NaV1.2
channel block, respectively. Thus great activity is
53.16 Calcium Channels
currently focused on discovering novel but selective
sodium channel blockers.
A variety of calcium channels (CaV) have been iden-
In this regard novel activity-dependent sodium
tified and characterized. Several have been shown to
channel blockers (e.g., NW-1029) have been shown
be prominently involved in pain regulation (Yaksh,
to reduce excitability of peripheral nerves and cause
T. L., 2006). The N-type calcium channel CaV2.2 is
antihyperalgesia in models of inflammatory and neu- an important regulator of nerve terminal excitability
ropathic pain (Veneroni, O. et al., 2003). Overall and neurotransmitter release. It has been well estab-
however, channel blockers still appear insufficiently lished that there is complex chemical regulation of
selective to avoid cardiac and CNS side effects. N-channels, particularly through GPCR signaling.
An alternative approach to selectively regulate ion For example, N-channel activity is attenuated by
channels is to block the trafficking of channels to the analgesic drugs such as opioids, with a resultant
nerve membrane. For example the functioning of modulation of sensory transmitter release, e.g., sub-
NaV1.8 may be reduced by preventing its interaction stance P, CGRP, and glutamate, at both spinal and at
with p-11, an annexin II-related protein that tethers peripheral sensory nerve terminals. N-channel traf-
the channel to the nerve membrane (Okuse, K. et al., ficking may also be affected by GPCRs. For example,
2002). In addition, channel-associated cell surface gly- activation of the orphanin FQ receptor (ORL) by
coproteins such as contactin may be involved in nociceptin causes channel internalization and down-
concentrating specific channel subtypes, e.g., NaV1.8 regulation of calcium entry (Altier, C. et al., 2005).
and NaV1.9 (IB4 positive) but not NaV1.6 and NaV1.7 Of additional importance is the fact that deletion of
(IB4 negative) in DRG nerve membranes, with an the N-channel gene reduces inflammatory and neuro-
associated increased in current density (Rush, A. M. pathic pain (Kim, C. et al., 2001; Saegusa, H. et al.,
et al., 2005). Although these approaches are attractive it 2001). Moreover selective blockers such as ziconotide,
 Pharmacological Modulation of Pain 809

(a naturally occurring conopeptide) and verapamil phenotype including excitability (Sah, D. W. H.

have been used to characterize channel activity while et al., 2003; Zweifel, L. S. et al., 2005). Several neuro-
ziconitide (SNX-111, Prialt) has been used experi- trophins have been identified including NGF, brain-
mentally and clinically by intrathecal administration, derived growth factor (BDNF) and neurotrophins
to show utility for pain relief (Xiao, W. H. and (NT) 3, neurotrophin 4/5. Each neurotrophin binds
Bennett, G. J., 1995; Snutch, T. P., 2005). Building on with high affinity to receptor tyrosine kinases (Trk):
this concept, small molecule CaV2.2 channel blockers NGF to TrkA, BDNF and NT4/5 to TrkB, and NT3
with oral availability are now reported to have been to TrkC. NT3 also binds with TrkA and TrkB. Mature
developed, e.g., NMED-160, showing efficacy in neurotrophins also bind to a structurally distinct
chronic pain models (Snutch, T. P., 2005). receptor p75 which affects neuronal development
While it is believed that blockers of CaV2.2 exert through downstream signaling. Neurotrophins arise
their major analgesic effects via the spinal cord, per- from proneurotrophin precursors following extracel-
ipheral actions are also possible. Thus N-channels lular cleavage by metalloproteinases and plasmin. It
expression also occurs in peripheral terminals and should be observed that proneurotrophins also signal
has been shown to increase following axotomy through the p75 receptor and may produce opposite
(Baccei, M. L. and Kocsis, J. D., 2000). This suggests effects from neurotrophins, e.g., apoptosis rather than
that peripheral N-channels can be targeted for neu- cell survival (Lu, B. et al., 2005).
ropathic pain treatments; thus avoiding potential NGF has been most studies with respect to
CNS side effects such as sedation. inflammatory hyperalgesia as its production is unre-
Low-voltage-activated T channels (CaV3.1, gulated by inflammation in macrophages, fibroblasts,
CaV3.2, and CaV3.3) also appear important for pain and Schwann cells. NGF has emerged as a key reg-
transmission and as targets for pain therapy. Thus ulator of sensory neuron excitability and as an
they are expressed in superficial laminas of the spinal important mediator of injury-induced nociceptive
cord and in DRG neurons (for references see Altier, and neuropathic pain (Ro, L. S. et al., 1999;
C. and Zamponi, G. W., 2004; Yaksh, T. L., 2006). Theodosiou, M. et al., 1999; Hefti, F. F. et al., 2005).
T-channels appear unaffected in DRGs after axotomy NGF acts via TrkA and p75 to activate a number of
(Baccei, M. L. and Kocsis, J. D., 2000) but may play a kinase pathways (e.g., p38 kinase; Ji, R. R. et al., 2002)
more prominent role in regulating spinal excitability leading to altered gene transcription and the
and spinal sensitization following repetitive C-fiber increased synthesis of sensory neuropeptides (sub-
stimulation (Ikeda, H. et al., 2003). Moreover nerve stance P, CGRP), ion channels (TRPV1, NaV1.8,
injury-induced hyper-responsiveness was blocked by ASIC3; Fjell, J. et al., 1999; Ji, R. R. et al., 2002;
the T-channel blocker ethosuximide (Matthews, E. A. Mamet, J. et al., 2003), membrane receptors such as
and Dickenson, A. H., 2001) which also attenuated bradykinin and P2X3 (Petersen, M. et al., 1998;
mechanical allodynia in animal models of vincristine- Ramer, M. S. et al., 2001), and structural molecules
and paclitaxel-induced neuropathic pain (Flatters, S. J. including neurofilament and channel anchoring pro-
and Bennett, G. J., 2004). teins such as the annexin light chain p11 (Okuse, K.
Finally a strongly validated approach for neuro- et al., 2002).
pathic pain has targeted the 2,1 calcium channel Increased expression and release of NGF have
subunit, the substrate for the antiallodynic drugs, been demonstration in several painful conditions in
gabapentin and pregabalin. This subunit is important animal models (e.g., ultraviolet injury, surgical injury;
for channel assembly, is expressed in small DRGs and Oddiah, D. et al., 1998; Miller, L. J. et al., 2002) and in
in spinal neurons, and its overexpression has been human conditions including arthritis, cystitis, prosti-
associated with allodynia in a number of specific pain tis, and headache (Aloe, L. et al., 1992; Halliday, D. A.
models (Luo, Z. D. et al., 2002). et al., 1998; Sarchielli, P. et al., 2001). Administration
of exogenous NGF induces thermal and mechanical
hyperalgesia in animals and humans (Andreev, N.
53.17 Neurotrophins and Their et al., 1995; Apfel, S. C., 2002), which is considered
Receptors to be due in part to mast cell degranulation and by
directly increasing sensory neuronal excitability
The neurotrophins represent an important family of (Sah, D. W. H. et al., 2003).
regulatory proteins essential for sensory nerve devel- Few small molecule NGF antagonists are avail-
opment, survival, and determination of chemical able but ALE0540, which inhibits the binding of
810 Pharmacological Modulation of Pain

NGF to TrkA and p75, and PD90780 which inhibits Although there appears not to be a specific role in
NGF binding to p75, have been proposed to have inflammation, GDNF has been shown to have neu-
efficacy in chronic pain models (Owolabi, J. B. et al., roprotective and restorative properties in a number
1999; Colquhoun, A. et al., 2004). In contrast, the use of neurodegenerative and neuropathic pain states
of TrkA-immunoglobulin G (IgG) and NGF mono- (Sah, D. W. H. et al., 2003). Specifically GDNF treat-
clonal antisera have confirmed the importance of ment has been shown to restore peripheral sensory
NGF in pain. Thus anti-NGF treatment have pro- neuron function, including peptide and ion channel
vided positive behavioral and biochemical readouts expression patterns, following painful peripheral
in several models of chronic nociceptive (including nerve injury accompanied by an attenuation of pain
visceral) and neuropathic pain (Obata, K. et al., 2004; behaviors. Unfortunately clinical observations using
Hefti, F. F. et al., 2005; Sevcik, M. A. et al., 2005; GDNF have shown unacceptable side effects such as
Shelton, D. L. et al., 2005). These studies have also weight loss and allodynia, which has discouraged
indicted a lack of effect on acute nociceptive proces- therapeutic developments (Nutt, J. G. et al., 2003).
sing or on sympathetic nerves whose phenotype is
also regulated by NGF.
The importance of NGF has also received clinical 53.18 Kinases
confirmations since RN624, a humanized anti-NGF
monoclonal antibody (mAb), has been reported to be As mentioned earlier inflammatory mediators also
efficacious in reducing pain and improved mobility activate a number of protein kinases in sensory neu-
in OA (Lane, N. et al., 2005). Anti-NGF mAb therapy rons and in the spinal cord. These include PKA, PKC,
appears as an attractive therapeutic approach with and mitogen-activated protein kinases (MAPK) con-
the potential for long-lasting pain treatment, similar sidered to be important downstream regulators of
in efficacy to morphine, without necessarily compro- excitability through altering gene transcription and
posttranslational modification of target proteins
mising physiological nociception.
(Woolf, C. J. and Salter, M. W., 2000). There are
NGF also induces the synthesis of another neuro-
several types of MAPKs including extracellular sig-
trophin, BDNF, from peptide-containing sensory
nal-regulated kinases (ERK), cJUN, N-terminal kinase
neurons. BDNF also accumulates in sensory neurons
(JNK), and p38 kinase that are considered as targets for
following painful nerve injury (see Sah, D. W. H.
inflammatory pain. For example, several inhibitors of
et al., 2003). Release of BDNF in the spinal dorsal
p38 kinase (e.g., SB203580, CNI-14930) posses anti-
horn increasing spinal excitability and pain sensitiza-
inflammatory as well as antihyperalgesic properties in
tion via TrkB receptors. This initiates a variety of
a variety of animal models (Schafers et al., 2003b).
effects, including direct neural excitation, activation
of a signaling cascade via the phosphorylation of
NMDA receptors, and an altered regulation of the 53.19 Botulinum Toxin
neural chloride transporter that contributes to pain
hypersensitivity (Coull, J. A. M. et al., 2005). Another approach to pain modulation has been the
In keeping with these observations, spinal BDNF use of botulinum toxins (BoTNs). This family of
administration induces thermal and mechanical neurotoxins has been traditionally used as an experi-
allodynia whereas anti-BNDF neutralization or mental tool to study muscle nerve interactions.
TrkB IgG administration reduces inflammation or Recently BoTN-A has been approved for clinical
nerve injury-mediated hypersensitivity pain in a use to induce muscle relaxation. The mechanism of
number of animal models (Kerr, B. J. et al., 1999; action of BoTN is related to inhibition of transmitter
Theodosiou, M. et al., 1999; Deng, Y. S. et al., 2000). release from motor fibers through proteolytic clea-
Finally glial cell-line-derived neurotrophic factor, vage of a number of synaptosomal regulatory
GDNF represents an extensive family of ligands and proteins (SNARE, syntaxin, SNAP-25, synaptobre-
membrane receptor complexes which have an impor- vin). More recent studies also indicated potential for
tant role in regulating peripheral and central neural inhibition of neuropeptide transmitter release from
phenotypes. GDNF-related ligands include neur- small afferent neurons (Welch, M. J. et al., 2000;
turin and artemin, which act via the complex RET Mense, S., 2004). In keeping with this BoTN has
Trk receptor and co-receptors GFR1, GFR2, been shown to provide long-lasting pain relief fol-
GFR3, and GFR4. lowing administration into human osteoarthritic
 Pharmacological Modulation of Pain 811

joints and improve bladder dysfunction in overactive derived for the most part from our studies of animal
bladder patients. This was correlated with loss of models (see Figure 1). The complexity and heteroge-
both P2X3 and VR1 receptors in the bladder neity of chronic pain states will clearly remain an
(Apostolidis, A. et al., 2005). enormously challenging area for future research.
There is still a great need for human data showing
the regulated expression of pain molecules and clinical
53.20 Nitric Oxide data that provides more rapid evaluation of emerging
pharmacological concepts. This will allow the most
NO induces a delayed burning pain upon intrader- valuable targets to be selected for analgesia develop-
mal injection (Holthusen, H. and Arndt, J. O., 1994) ment in the clinic.
and NO donors have been postulated to activate Clearly the improvements in our pharmacological
cerebral sensory fibers directly, causing release of understand of pain has also provided an abundance of
the sensory vasodilator CGRP (Wei, P. et al., 1992). opportunities for developing analgesics. Among the
Indeed NO has been suggested to contribute to most comprehensively studied molecules in chronic
migraine and other types of head pain (Olesen, J. pain are inflammatory mediators and their key recep-
et al., 1994) and there is an abundance of evidence tors. These include PGE2/EP1, bradykinin/BK1 and
linking NO in the etiology of a number of chronic BK2, ATP/P2X3, cytokines/ IL-1, chemokines/
pain conditions (Millan, M. J., 1999). CCr2 and neurotrophins/NGF/TrkA that sensitize
Peripheral nerve injury and inflammation increases and increase excitability in peripheral and central
the expression of NO in sensory neurons and causes pain pathways. Many of these mediators act through
the upregulation of the nitric oxide synthase (NOS) GPCRs and there appears to be some convergence of
isozymes neuronal NOS (nNOS) and inducible NOS mechanisms with changes in expression and regula-
(iNOS) in the spinal cord (Gordh, T. et al., 1998) as a tion of ion channel. Key channels for pain are either
result of axonal neurodegeneration, neuroglial cell
ligand gated, for example, TRPV1, P2X3, P2X4,
activation and inflammation (Levy, D. et al., 2001). In
P2X7, or voltage gated, for example, NaV1.8,
keeping with these observations, treatment with non-
NaV1.7, NaV1.3, CaV2.2.
specific NOS inhibitors such as L-N-nitro-L-arginine
A variety of additional cellular process including
methyl ester (L-NAME) has been shown to prevent or
protein phosphorylation of membrane receptors and
reduce pain hypersensitivity (Meller, S. T. et al., 1992;
channels via key kinases, for example, MAPK, as well
Hao, J. X. and Xu, X. J., 1996; Handy, R. L. and Moore,
as complex phenotype change regulated by neuro-
P. K., 1998). Indeed particular interest has been
trophins require further understanding. In addition,
devoted to producing selective blockers of iNOS, to
the emerging importance of neuroglia and their phar-
avoid side effects associated with nonspecific block of
nNOS and endothelial NOS (eNOS). Recently macology needs exploration and consolidation.
GW27415, a selective iNOS inhibitor has been Finally despite the great advances in pain phar-
shown to partially reduce Freunds complete adjuvant macology, many pain molecules remain poorly
(FCA)-induced inflammatory pain as well the hyper- validated as good targets for analgesia. Stronger clin-
sensitivity associated with the CCI neuropathy model. ical translation is highly likely with the progression
This action was most likely to have been caused via a of pharmacological opportunities into humans.
peripheral mechanism as no iNOS expression was
detectable in sensory ganglia or spinal cord (Alba, J.
D. et al., 2006).
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botulinum neurotoxins. Toxicon 38, 245258.
Whiteside, G. T., Boulet, J. M., and Walker, K. 2005a. The role of
central and peripheral mu opioid receptors in inflammatory Further Reading
pain and edema: a study using morphine and DiPOA ([8-(3,3-
diphenyl-propyl)-4-oxo-1-phenyl-1,3,8-triaza-spiro[4.5]dec- Abdulla, F. A. and Smith, P. A. 2001. Axotomy- and autotomy-
3-yl]-acetic acid). J. Pharmacol. Exp. Ther. 314, 12341240. induced changes in the excitability of rat dorsal root ganglion
Whiteside, G. T., Gottshall, S., Boulet, J. M., Chaffer, S. M., neurons. Am. J. Physiol. 85, 630643.
Harrison, J. E., Pearson, J. E., Turchin, P. I., Mark, L., Cahill, C. M., Morinville, A., Hoffert, C., ODonnell, D., and
Garrison, A. E., and Valenzano, K. J. 2005b. A role for Beaudet, A. 2003. Up-regulation and trafficking of delta
cannabinoid receptors, but not endogenous opioids in the opioid receptor in a model of chronic inflammation:
antinociceptive activity of the CB(2) selective agonist implications for pain control. Pain 101, 199208.
GW405833. Eur. J. Pharmacol. 528, 6572. Davis, K. D., Treede, R. D., Raja, S. N., Meyer, R. A., and
Woodbury, J., Zwick, M., Wang, S., Lawson, J. J., Campbell, J. N. 1991. Topical application of clonidine
Caterina, M. J., Koltzenbury, M., Albers, K. M., relieves hyperalgesia in patients with sympathetically
Koerber, H. R., and Davis, B. M. 2004. Nociceptors Lacking maintained pain. Pain 47, 309317.
TRPV1 and TRPV2 Have Normal Heat Responses. J. Dray, A. 1994. Tasting the inflammatory soup: the role of
Neurosci. 24, 64106415. peripheral neurones. Pain Rev. 1, 153171.
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Ferreira, J., Campos, M. M., Pesquero, J. B., Araujo, R. C., Kim, S. F., Huri, D. A., and Snyder, S. H. 2005. Inducible nitric
Bader, M., and Calixto, J. B. 2001. Evidence for the oxide synthase binds, N-nitrosylates and activates
participation of kinins in Freunds adjuvant induced cyclooxygenase-2. Science 310, 19661970.
inflammatory and nociceptive responses in kinin B1 and B2 Kress, M., Izydorcyk, I., and Kuhn, A. 2001. N- and L-but not P/
knockout mice. Neuropharmacology 41, 10061012. Q-type calcium channels contribute to neuropeptide release
Hao, S., Mta, M., and Goins, W. 2003. Transgene-mediated from rat skin in vitro. Neuroreport 12, 867870.
enkephaline release enhances the effect of morphine and Ossovskaya, V. S. and Bunnett, N. W. 2003. Protease-activated
evades tolerance to produce a sustained antiallodynic effect. receptors: contribution to physiology and disease. J.
Pain 102, 135142. Physiol. 552, 589601.
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 54 Forebrain Opiates
J-K Zubieta, University of Michigan, Ann Arbor, MI, USA
2009 Elsevier Inc. All rights reserved.

54.1 Introduction 821

54.2 Endogenous Opioid Mechanisms in the Regulation of Pain 821
54.2.1 m-Opioid-Receptor-Mediated Pain Processing 821
54.2.2 -Opioid-Receptor-Mediated Processing 824
54.2.3 d-Opioid-Receptor-Mediated Processing 825
54.2.4 Opioid Receptor-like (ORL-1, NOP Receptor)-Mediated Processing 826
References 827

54.1 Introduction () (Chen, Y. et al., 1993; Thompson, R. C. et al., 1993),

and kappa ( ) (Li, S. et al., 1993; Meng, F. et al., 1993).
The initial studies on the neurobiology of pain and These are seven transmembrane domain G-protein-
the discovery and understanding of the endogenous coupled receptors inhibiting adenyl cyclase and mod-
opioid systems have been historically intrinsically ulating calcium and potassium conductance.
linked. In the 1960s, early research by Liebeskind
and coworkers (Mayer, D. J. et al., 1971) demonstrated
that the electrical stimulation of the ventral peria-
queductal and periventricular gray produced 54.2 Endogenous Opioid
profound analgesia, an effect that was independently Mechanisms in the Regulation of Pain
observed by Reynolds D. V. (1969). During the same 54.2.1 m-Opioid-Receptor-Mediated Pain
time, Tsou K. and Jang C. S. (1964) observed similar Processing
analgesic effects when morphine was microinjected
into these locations. As a consequence, and before This chapter will cover the regulation of pain by
opioid receptors or their ligands had been isolated, supraspinal opioid systems. Investigation of these
Mayer D. J. et al. (1971) proposed that electrical mechanisms has been most extensive for those per-
stimulation induced analgesia by activating neural taining to the -opioid receptor, as this is the site of
substrates that are involved in the blockage of pain action of opiate analgesics and the receptor most
and that were related to the analgesic action of mor- consistently associated with pain suppression.
phine. These observations were followed by the Knockout animals devoid of these receptors display
subsequent use of electrical stimulation of the poster- shorter latencies in the tail flick and hot plate tests,
thought to involve spinal and supraspinal mechan-
ior aspect of the paraventricular nucleus of the
isms, respectively, as well as a lack of morphine
thalamus for the treatment of pain (Richardson, D.
effects (Sora, I. et al., 1997). Its endogenous ligands
E. and Akil, H., 1977a; 1977b). In 1973, Snyder and
include -endorphin, derived from the peptide pre-
Pert published the first manuscript demonstrating the
cursor proopiomelanocortin (POMC), the recently
existence of opioid receptors (Pert, C. B. and Snyder,
described endomorphins 1 and 2, and in some cir-
S. H., 1973), a development that was paralleled by the
cuits, the enkephalins (cleaved from the larger
work of Eric Simon (Simon, E. J. et al., 1973) and Lars
preproenkephalin precursor). POMC-containing
Terenius (Terenius, L., 1973). These initial findings
neurons are located in the arcuate nucleus of the
were followed by the identification of met- and hypothalamus and periarcuate regions of the med-
leu-enkephalin (Hughes, J. et al., 1975), -endorphin ial-basal hypothalamus, with a smaller group
(Li, C. H. et al., 1976; Loh, H. H. et al., 1976), and the localized in the nucleus of the tractus solitarius of
dynorphin peptides (Goldstein, A. et al., 1979). The the medulla. Projections are extensive to the periven-
1990s saw the cloning of the opioid receptors delta () tricular thalamus, septum, and amygdala, dorsally
(Evans, C. J. et al., 1992; Kieffer, B. L. et al., 1992), mu descending to the periaqueductal gray (PAG), midline

821
822 Forebrain Opiates

raphe, and reticular formation, and laterally to the Last, the most recently identified family of opioid
anterior and lateral hypothalamus. Lighter innervation peptides, the endomorphins 1 and 2, and in a manner
is observed in the medial portion of the nucleus accum- similar to that for -endorphin, is synthesized in the
bens and olfactory cortex (Khachaturian, H. et al., 1985). brain only in hypothalamic cells (periventricular,
Animals devoid of a functioning POMC gene show a arcuate, and ventromedial and dorsomedial nuclei),
lack of naloxone-reversible analgesia induced by mild and project to pain-processing regions of the brain-
swim stress, but greater nonopioid (naloxone nonrever- stem and pons (trigeminal tract, parabrachial nucleus,
sible) analgesic effects and normal effects of morphine. nucleus of the tractus solitarious, PAG, and locus
These data demonstrated the involvement of -endor- coeruleus) as well as to midline thalamic nuclei and
phin in stress-induced analgesic responses, while also the amygdala. Besides the hypothalamic cell bodies,
highlighting the presence of alternate mechanisms not the only other known areas containing endomorphin-
mediated by -endorphin and -opioid receptors positive cell bodies include the nucleus of the tractus
(Rubinstein, M. et al., 1996). solitarius and the dorsal root ganglia. Consistent with
In contrast to the narrow localization of POMC- its actions at -opioid receptors, intracerebroventri-
containing cells, enkephalinergic cells and projections cular administration of endomorphins induces
are widely distributed, forming both local circuits and profound analgesia that may be dissociated from the
long projections, with similar distributions for met- and rewarding properties associated with -opioid recep-
leu-enkephalin, albeit with a higher level of met-enke- tor agonists (Zadina, J. E., 2002).
phalin in all regions. The highest concentrations of Initial studies on the role of supraspinal -opioid-
terminals with these peptides are encountered in the receptor-mediated antinociception focused on the
globus pallidus and central and lateral amygdala nuclei. role of the PAG and its connections with the rostro-
Globus pallidus and ventral pallidal projections form ventral medulla (RVM). These are regions that,
part of the enkephalinergic striatopallidal pathway, together with the dorsolateral pontine tegmentum,
with cell bodies arising primarily in the nucleus accum- control nociceptive transmission via projections
bens and to a lesser extent in the ventromedial through the spinal cord dorsolateral funiculus to the
hypothalamus. Cell bodies containing preproenkepha- dorsal horn laminae (Fields, H. L. and Basbaum, A. I.,
lin are also encountered in the olfactory bulb, septal 1978). Much of the focus during the following dec-
nuclei, bed nucleus of the stria terminalis, throughout ades was then dedicated to the understanding of the
the basal ganglia and in the cortex. Terminal densities mechanisms underlying this modulation. Both pain
are moderate to dense in the cingulate, piriform, and facilitatory and inhibitory cells have been described
entorhinal cortex, anterior and periventricular thala- at the level of the RVM (Moreau, J. L. and Fields, H.
mus, and preoptic hypothalamus (the latter from both L., 1986; Heinricher, M. M. et al., 1987; Urban, M. O.
local projections and terminals arising from the amyg- and Smith, D. J., 1994; Zhuo, M. and Gebhart, G. E.,
dala and periamygdalar area). Both cell bodies and rich 1997). -Opioid receptors, through the activation of
local projections are encountered in the PAG, raphe, enkephalinergic neurotransmission and GABA inter-
locus coeruleus, nucleus ambiguus, and nucleus of the neurons in the PAG and RVM (al-Rodhan, N. et al.,
solitary tract (Petrusz, P. et al., 1985). Enkephalin-defi- 1990), increase the activity of inhibitory (off) cells
cient mice display large increases in the concentration reducing pain transmission. The administration of -
of -opioid receptors in the globus pallidus, central opioid receptor agonists systemically, or directly
nucleus of the amygdala, bed nucleus of the stria ter- into the PAG or RVM, increases the activity of these
minalis and substantia innominata, preoptic, medial and off cells, with blockade of the activation of these neu-
lateral hypothalamus, medial thalamus, PAG, raphe, rons preventing morphines antinociceptive effects
nucleus ambiguus, and nucleus of the tractus solitarius, (Heinricher, M. M. et al., 1999). Not surprisingly, in
suggesting substantial enkephalinergic activity on view of these results, the possible role of -opioid
-opioid receptors in these regions, with compensatory neurotransmission beyond the PAG, RVM, and des-
upregulation in the knockouts (Brady, L. S. et al., 1999). cending pathways into the spinal cord (e.g.,
Enkephalin-deficient mice display reductions in supraspinal mechanisms) was thought to be less cri-
supraspinal analgesic responses as measured with the tical and mostly regarded as secondary to the
hot-plate test and rapid nocifensive responses in the analgesia mediated primarily by brainstem and spinal
formalin assay, but unaltered spinal analgesic responses cord regions.
(tail-flick test) and stress-induced analgesia (Konig, M. Nevertheless, examination of the distribution of
et al., 1996). -opioid receptors, mRNA in the rodent (Mansour,
 Forebrain Opiates 823

A. et al., 1995) and in vivo -opioid receptor avail- 1977; Watkins, L. and Mayer, D., 1982). It is therefore
ability in the human brain (Frost, J. et al., 1985) possible that the progressive engagement of suprasp-
demonstrate a broad distribution of these sites in inal -opioid mechanisms may be of greater
telencephalic areas. Administration of -opioid importance for the control of noxious stimuli as it
receptor agonists increases the blood flow of brain progresses from a relatively simple sensory experi-
regions rich in -opioid receptors (cingulate, pre- ence to one that requires the assessment of its
frontal, temporal and insular cortices, thalamus, significance and an appropriate response by the
hypothalamus, basal ganglia, amygdala, and brain- organism.
stem) (Adler, L. J. et al., 1997; Schlaepfer, T. et al., Two studies have examined the activity of
1998; Casey, K. et al., 2000; Wagner, K. J. et al., 2001), -receptor-mediated opioid neurotransmission
and reduces pain-induced activation in the thalamus in humans during the experience of pain. Both
(Casey, K. et al., 2000). Some of these areas, such as studies employed the -opioid-receptor-selective
thalamic and hypothalamic nuclei, the central radiotracer [11C]carfentanil and positron emission
nucleus of the amygdala, the agranular insular cortex tomography. Bencherif B. et al. (2002) utilized the
and lateral orbitofrontal cortex, have been associated capsaicin model of experimental pain, while in
with the suppression of pain behavior in animal mod- Zubieta J. K. et al. (2001), jaw muscle pain was
els (Mayer, D. J. and Liebeskind, J. C., 1974; Yaksh, T. induced by the intramuscular infusion of hypertonic
L. and Rudy, T. A., 1978; Bodnar, R. J. et al., 1980; saline. The former, in a smaller sample, showed evi-
Coffield, J. A. et al., 1992; Burkey, A. R. et al., 1996; dence of activation of -opioid-receptor-mediated
Manning, B., 1998; Manning, B. and Franklin, K., neurotransmission in the thalamus. In the latter,
1998; Harte, S. et al., 2000; Manning, B. H. et al., engagement of this neurotransmitter system was
2001). However, the inhibition of pain signaling eli- observed in a number of brain areas, including the
cited by the activation of -opioid receptors in these PAG, dorsal and rostral anterior cingulate, dorsolat-
supraspinal regions is thought to take place through eral prefrontal cortex, anterior and posterior insular
their connections with the PAG, since inactivation of cortex, medial and lateral thalamus, hypothalamus,
the PAG or the administration of opioid antagonists ventral basal ganglia, and amygdala. Invariably, the
in this area abolishes the analgesic response. In turn, relationship between -opioid activation and subjec-
PAG-induced analgesia is blocked by -opioid tive ratings of pain was in the negative direction (the
receptor antagonists microinjected in the RVM, larger the activation the lower the pain ratings), with
then reducing nociceptive transmission from the sensory or pain affect elements being modulated by
spinal cord (Kiefel, J. M. et al., 1993; Roychowdhury, different regions. Some of these regions had been
S. M. and Fields, H. L., 1996). previously implicated in opioid-mediated suppres-
In view of the critical role of the PAG and RVM sion of nociceptive responses in animal models (e.g.,
on -opioid-receptor-mediated antinociception, per- PAG, thalamus, hypothalamus, amygdala, and to a
haps the question to be answered regarding the lesser extent areas of the insular cortex), as noted
involvement of opioid systems in supraspinal nuclei above. Conversely, while pain-induced increases in
is their respective role in the experience of pain as a synaptic activity of the prefrontal cortex (Baron, R.
complex phenomenon. Opioid neurotransmission et al., 1999; Lorenz, J. et al., 2002) and dorsal anterior
is activated in humans during clinical pain, or experi- cingulate cortex (Rainville, P. et al., 1997) had been
mental pain of some duration, as evidenced described and associated with hyperalgesia and pain
by enhancements in pain ratings after the adminis- affect, respectively, the involvement of the endogen-
tration of naloxone, a nonselective opioid receptor ous opioid system on these processes had not been
antagonist (Levine, J. et al., 1978). Conversely, experi- previously explored. Similarly, the ventral basal
mental pain of relatively short duration (e.g., <10 ganglia (nucleus accumbens, ventral pallidum) have
min) does not elicit an apparent activation of opioid been typically associated with either responses to
neurotransmission, evidenced by a lack of increases natural rewards or drugs of abuse, stress, or, as is
in pain ratings after naloxone administration more recently postulated, with salient stimuli irre-
(Grevert, P., and Goldstein, A., 1977). These findings spective of its valence (Horvitz, J., 2000). These data
are similar to those observed in response to nonpain- then appear to indicate that, besides its traditional
ful stressors, whereby prolonged or unpredictable role in the modulation of descending inhibitory path-
stimuli activate opioid neurotransmission, leading to ways into the PAG and RVM, supraspinal -opioid
the term stress-induced analgesia (Madden, J. T. et al., receptors are additionally involved in the local
824 Forebrain Opiates

processing of complex assessments of the pain P., 2001; Fillingim, R. B. and Gear, R. W., 2004). A
experience. In this regard, -opioid receptors appear less prominent activation of -opioid-receptor-
implicated in regulating emotional responses, as mediated neurotransmission, as measured with neu-
examined in knockout rodents devoid of these recep- roimaging techniques, has been shown in women
tors. These animals display an enhancement of studied under conditions of low estradiol, low pro-
anxiety-like responses in the elevated arm maze gesterone during an experimental pain challenge
(Filliol, D. et al., 2000) and deficits in attachment (Zubieta, J. K. et al., 2002). This effect, observed in
behavior (Moles, A. et al., 2004). Proenkephalin the thalamus, amygdala, and nucleus accumbens, was
knockouts also show a lack of conditioned place reversed after selectively increasing estradiol plasma
aversion to naloxone (Skoubis, P. D. et al., 2005), levels, which also increased -opioid receptor con-
increased aggressive behavior to an intruder and centrations in these regions (Smith, Y. R. et al., 2006).
anxiety-like behavior in the open arm maze, as well These data then suggest an active regulation of
as alterations in supraspinal but not spinal pain pro- -opioid-receptor-mediated mechanisms by gonadal
cessing (Konig, M. et al., 1996), additionally involving steroids.
this neurotransmitter system in emotional behavioral
responses.
54.2.2 -Opioid-Receptor-Mediated
Work in humans has also shown that in the case of
Processing
some of the brain regions in which -opioid systems
become active during pain (e.g., rostral anterior cin- The prodynorphin-derived peptides, which present
gulate, dorsolateral prefrontal cortex, nucleus the highest affinity and colocalization with -opioid
accumbens, and insular cortex), cognitive expecta- receptors, consist of extended forms of leu-enkepha-
tions associated with the administration of a placebo lin coded by a different gene and mRNA. These
with expected analgesic properties are also capable of peptides include dynorphin A, B, and neoendorphins
activating this neurotransmitter system to mediate  and  (Goldstein, A. et al., 1979). Their involve-
the placebo analgesic effect (Benedetti, F. et al., ment in pain regulatory mechanisms has been
2005; Zubieta, J. K. et al., 2005). controversial, as both anti- and pronociceptive
A relatively less explored area in the understand- responses have been described in response to -ago-
ing of -opioid receptor modulation of pain is the nists depending on the brain regions studied.
additional influences of age and gonadal steroids (e.g., Similarly to -endorphin and endomorphin systems,
sex effects). In rodents, age-associated declines in there is a dense accumulation of dynorphin-contain-
-opioid receptor concentrations have been reported ing cells in the multiple regions of the hypothalamus,
(Piva, F. et al., 1987). However, humans demonstrate forming both local circuits and longer projections,
opposite effects, with increases in most brain regions particularly to the amygdala and descending to
shown in postmortem material (Gross-Isseroff, R. brainstem, pontine, and medullary/spinal regions
et al., 1990; Gabilondo, A. et al., 1995) and in neuroi- (e.g., PAG, but also substantia nigra). However, and
maging studies (Zubieta, J. K. et al., 1999), albeit the closer to the distribution of the enkephalin peptides,
psychophysical correlates of these changes in recep- cell bodies are also encountered in limbic areas of the
tor concentrations have not been determined. In cortex, such as the anterior cingulate, orbitofrontal,
human subjects, women further demonstrate declines insular, periamygdaloid, and entorhinal cortex, in the
in -opioid receptor levels after menopause (Zubieta, basal ganglia (caudate, putamen, globus pallidus),
J. K. et al., 1999). In this regard, trophic effects of nucleus accumbens, amygdala, particularly in its cen-
estradiol have been shown at the level of -opioid tral nucleus, PAG, raphe nuclei, locus coeruleus,
receptor levels and their mRNA, POMC content, nucleus ambiguus, and nucleus of the solitary tract.
and -endorphin release in animal models Long pathways have been described from the stria-
(Hammer, R. P. and Bridges, R. S., 1987; Dondi, D. tum to the substantia nigra and ventral pallidum, as
et al., 1992; Quinones-Jenab, V. et al., 1997; Eckersell, well as from the amygdala to brainstem and pons
C. et al., 1998). The clinical implications of these regions. Minimal innervation of the thalamus has
effects have not been well developed, albeit sex dif- been observed, which is confined to midline and
ferences have been observed in the effects of intralaminar nuclei (Fallon, J. and Ciofi, P., 1990).
-opioid receptor agonists, including their side Mice devoid of the dynorphin gene show an upregu-
effects and cardiovascular parameters, which appear lation of -receptors in the nucleus accumbens shell
to be more prominently affected in women (Zacny, J. and core, hypothalamus, amygdala, and substantia
 Forebrain Opiates 825

nigra, consistent with their high level of dynorphin patients, albeit the clinical consequences of these
innervation, and suggesting an active role of this alterations are presently unknown (Hagelberg, N.
peptide in the modulation of the -receptors in et al., 2004).
these brain regions (Clarke, S. et al., 2003).
Intrathecal administration of -opioid agonists or
54.2.3 d-Opioid-Receptor-Mediated
their microinjection in the RVM has been shown to
Processing
result in antinociceptive responses in various rat
models (Harada, Y. et al., 1995; Ackley, M. A. et al., The enkephalins are thought to be the endogenous
2001), albeit modest, and typically requiring high ligand for the -opioid receptor, in addition to acting
doses. These effects have also been demonstrated to on -opioid receptors, as noted above. Their distri-
be sex dimorphic, with female rats demonstrating a bution was described in this chapter under Section
more profound analgesic effect than the males in the 54.2.1. Knockout mice devoid of the enkephalin gene
tail-flick test (Tershner, S. A. et al., 2000). In -recep- display increases in the regional concentration of -
tor knockout animals, female but not male rats and -opioid receptors (Brady, L. S. et al., 1999),
demonstrate decreased withdrawal latencies in the suggesting an effect of this peptide in both receptor
tail-immersion test, compared to wild-type animals systems. In the case of the -receptor, these increases
(Martin, M. et al., 2003). Consistent with these find- were observed in the caudateputamen, nucleus
ings, one study has shown a preferential accumbens core and shell, ventral pallidum, globus
antinociceptive effect of opiates with -opioid activ- pallidus, the central nucleus of the amygdala, preop-
ity in women, with respect to men, after surgical tic area, and lateral hypothalamus. Microinjection of
extraction of the third molar (Gear, R. et al., 1996). -receptor agonists in the PAG and medullary reti-
-Opioid agonists, however, have also been shown cular formation has been shown to induce
to antagonize morphine analgesia, by the hyperpo- antinociception (Jensen, T. S. and Yaksh, T. L.,
larization of pain-inhibitory cells activated by - 1986). Their intracerebroventricular administration
opioid receptors in the RVM (Pan, Z. Z. et al., 1997; has also been shown to have antinociceptive proper-
Meng, I. D. et al., 2005). Therefore pronociceptive, ties in tests of spinal and supraspinal nociceptive
antianalgesic effects of -opioids may exist simulta- mechanisms in wild-type and -opioid-receptor-
neously with antinociceptive activity, depending on deficient mice (Porreca, F. et al., 1984; Vaught, J. L.
the brain regions involved or the existing level of et al., 1988; Ossipov, M. H. et al., 1995). Rodent,
-opioid receptor activity. Opposing effects of - primate, and human -opioid receptors are uniformly
and -receptors have also been shown at the level distributed throughout the cortex, diffusely in the
of the medial thalamus, whereby microinjection of striatum, and moderately to densely in the nucleus
-agonists increased, and -agonists reduced, the accumbens, ventral pallidum, hippocampus, amyg-
thresholds for pain-induced vocalization and motor dala, medial thalamus, hypothalamus, PAG, and
responses in male rodents (Carr, K. D. and Bak, T. H., raphe nuclei. Low concentrations are observed in
1988). Dynorphin also appears to be involved in the lateral thalamic nuclei and the cerebellum (Lewis,
maintenance of hyperalgesia and persistent pain, M. E. et al., 1981; 1983; Moskowitz, A. S. and
with only spinal mechanisms having been investi- Goodman, R. R., 1985; Smith, J. S. et al., 1999;
gated at this time (Wang, Z. et al., 2001). Cahill, C. M. et al., 2001b). This distribution, which
Dynorphin and -opioid agonists further antago- emphasizes a limbic and paralimbic organization as
nize dopamine release in the basal ganglia (Zhang, Y. well as its relationship with pain circuitry, has sug-
et al., 2004), whether induced by -opioid receptor gested the involvement of this receptor and
agonists, psychostimulants, or stressors, by the inhibi- neurotransmitter system in emotional and stress reg-
tion of dopamine (DA) cells in the ventral tegmental ulation in addition to antinociceptive mechanisms.
area and substantia nigra or possibly indirectly Regarding the latter, -opioid-receptor-agonist-
through the regulation of DA reuptake at the level of induced analgesia and respiratory depression has
the nucleus accumbens (Thompson, A. C. et al., 2000; been shown either reduced (Matthes, H. W. et al.,
Margolis, E. B. et al., 2003; McLaughlin, J. P. et al., 2003; 1998) or in some cases abolished (Scherrer, G. et al.,
Narita, M. et al., 2005). Dopaminergic mechanisms, 2004), depending on the assays employed, in animals
typically associated with responses to drugs of abuse devoid of -opioid receptors. These results then sug-
and more generally to salient stimuli regardless of its gested the presence of functional interactions between
valence, have been found dysregulated in chronic pain - and -opioid-receptor types. Conversely, -opioid
826 Forebrain Opiates

stress-induced analgesia after forced swim stress was opioid receptor, with high homology with other opioid
found maintained in -opioid receptor knockout mice. receptors, but does not bind to traditional opioid pep-
These data supported a lack of interactions between tides. It is present in high concentrations in the septum,
- and -opioid receptors when endogenous opioid diagonal band of Broca, hypothalamus (preoptic, arcu-
transmission was activated by a stressor, as opposed to ate, paraventricular, ventromedial), hippocampus,
exogenously administered agonists. However, and in medial amygdala, substantia nigra, locus coeruleus,
the same studies, two phases of the stress-induced raphe nuclei, PAG, as well as in the dorsal and ventral
analgesia were observed, one early response mediated horns of the spinal cord and in the trigeminal nucleus.
by -opioid receptors, and a later response that was It is also moderately concentrated in the thalamus,
abolished in the -opioid receptor knockouts and cortex (olfactory, cingulate, claustrum, and frontopar-
unaffected by -opioid receptor antagonists (LaBuda, ietal areas), the shell of the nucleus accumbens, and
C. J. et al., 2000). islets of Calleja. Low levels are observed in the cau-
Utilizing single, double, and triple knockouts for dateputamen, core of the nucleus accumbens, and
-, -, and -opioid receptors Martin M. et al. (2003) cerebellum (Mollereau, C. et al., 1994). The 17-amino
have suggested that -opioid receptors, together with acid peptide orphanin FQ/nociceptin (OFQ /N), the
-opioid receptors, were involved in mechanical endogenous ligand for this receptor, was subsequently
nociception and some, but not other phases of inflam- isolated. It presented high homology with opioid pep-
matory pain. Besides their independent functions, tide transmitters, particularly dynorphin (Meunier, J.
interactions between - and -receptors have also C. et al., 1995; Reinscheid, R. K. et al., 1995). This
been demonstrated at various levels. Treatment peptide is synthesized as part of a larger precursor,
with -opioid agonists has been shown to increase preproOFQ/N, with a distribution largely overlapping
the concentration of membrane-bound -receptors in with that of the NOP receptor in the rat brain
the spinal cord by increasing their trafficking from (Mollereau, C. et al., 1996; Darland, T. et al., 1998).
intracellular sites, an effect that was abolished in Like opioid agonists, OFQ/N inhibits adenylate
-opioid receptor knockout animals (Cahill, C. M. cyclase and calcium channels, and induces an inhibi-
et al., 2001a; Morinville, A. et al., 2003). Furthermore, tory effect on postsynaptic neurons by opening
interacting complexes of - and -receptors (hetero- potassium channels and hyperpolarization (Hawes, B.
dimers) have been demonstrated in spinal cord and E. et al., 2000; Moran, T. D. et al., 2000). This inhibitory
neuroblastoma cell lines, as well as synergistic inter- effect has been invoked in explaining the often-times
actions in analgesic effects between the two receptor contradictory effects of generalized routes of OFQ/N
types (Gomes, I. et al., 2004). peptide administration (e.g., intracerebroventricular)
Regarding other possible effects of -opioid neu- (Darland, T. et al., 1998). For example, initial reports
rotransmission, stressors have been shown to described increases in pain behavior (Meunier, J. C.
decrease the concentration of -opioid receptors in et al., 1995; Reinscheid, R. K. et al., 1995), no effect on
limbic areas of the rat (Fadda, P. et al., 1991) and pain responses (Mogil, J. S. et al., 1996a; 1996b; Tian, J.
increase that of enkephalin mRNA in the ventral H. et al., 1997), or even hyperalgesia (Rossi, G. C. et al.,
medulla (Mansi, J. A. et al., 2000) and hypothalamus 1996; 1997), and a reversal of the analgesic actions of
(Helmreich, D. L. et al., 1999; Dumont, E. C. et al., morphine and endogenous opioid peptides (Mogil, J. S.
2000), confirming their involvement in the modula- et al., 1996a; 1996b). It is thought that these effects
tion of the stress response, albeit not in its depend on the populations of cells being inhibited, as
neuroendocrine components, at least in primates OFQ/N can hyperpolarize neurons that both facilitate
(Williams, K. L. et al., 2003). Antidepressant-like and inhibit pain signal transmission at the level of the
effects have also been demonstrated in animal models RVM (Heinricher, M. M. et al., 1997), as well as most of
of depression, suggesting the participation of this the neuronal populations rich in the NOP receptor
system in complex behavioral functions (Jutkiewicz, (e.g., hypothalamus, medulla, hippocampus, amygdala,
E. M. et al., 2003; Torregrossa, M. M. et al., 2004). ventral tegmental area, locus coeruleus, and PAG) (see
Heinricher, M. M., 2003 for a recent review). Using
data acquired by the microinjection of OFQ /N in the
54.2.4 Opioid Receptor-like (ORL-1, NOP
RVM, Heinricher M. M. et al. (1997) proposed that the
Receptor)-Mediated Processing
behavioral effects of this peptide depend on the state of
The opioid receptor-like (ORL1) receptor (now activity of the output neurons affected by the peptide.
denominated NOP) is the most recently identified Under conditions of stimulation (opioid withdrawal,
 Forebrain Opiates 827

stress, and opioid agonist effects) the effects of OFQ / Cahill, C. M., Morinville, A., Lee, M. C., Vincent, J. P., Collier, B.,
and Beaudet, A. 2001a. Prolonged morphine treatment
N would oppose the state of activation in the relevant targets delta opioid receptors to neuronal plasma
circuits, inducing an opposing effect, with additional membranes and enhances delta-mediated antinociception.
influencing variables, such as sex or genotype, also J. Neurosci. 21, 75987607.
Cahill, C. M., McClellan, K. A., Morinville, A., Hoffert, C.,
suggested to influence its effects (Mogil, J. S. and Hubatsch, D., ODonnell, D., and Beaudet, A. 2001b.
Pasternak, G. W., 2001; Flores, C. A. et al., 2003). On Immunohistochemical distribution of delta opioid receptors
the other hand, and during basal states, OFQ /N in the rat central nervous system: evidence for
somatodendritic labeling and antigen-specific cellular
effects would be minimal (Heinricher, M. M. et al., compartmentalization. J. Comp. Neurol. 440, 6584.
1997). Consistent with these findings, OFQ /N Carr, K. D. and Bak, T. H. 1988. Medial thalamic injection of
enhanced stress-induced ACTH production at the opioid agonists: mu-agonist increases while kappa-agonist
decreases stimulus thresholds for pain and reward. Brain
level of the pituitary and plasma corticosterone, an Res. 441, 173184.
effect that in this case was also observed to a lesser Casey, K., Svensson, P., Morrow, T., Raz, J., Jone, C., and
extent in nonstressed animals (Devine, D. P. et al., Minoshima, S. 2000. Selective opiate modulation of
nociceptive processing in the human brain. J. Neurophysiol.
2001). Knockout rodents devoid of either OFQ /N or 84, 525533.
NOP receptors do not appear to have significant Chen, Y., Mestek, A., Liu, J., Hurley, J. A., and Yu, L. 1993.
changes in nociceptive responses or changes in opiate Molecular cloning and functional expression of a mu-opioid
receptor from rat brain. Mol. Pharmacol. 44, 812.
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et al., 1997; Koster, A. et al., 1999; Kest, B. et al., 2001; 2003. Region selective up-regulation of micro-, delta- and
Mamiya, T. et al., 2001). In a neuropathic pain model, kappa-opioid receptors but not opioid receptor-like 1
receptors in the brains of enkephalin and dynorphin
NOP mRNA expression increased in pain regulatory knockout mice. Neuroscience 122, 479489.
circuitry (raphe magnus, PAG, and RVM), albeit the Coffield, J. A., Bowen, K. K., and Miletic, V. 1992. Retrograde
behavioral consequences of these increases have not tracing of projections between the nucleus submedius, the
ventrolateral orbital cortex, and the midbrain in the rat.
been explored (Ma, F. et al., 2005). J. Comp. Neurol. 321, 488499.
Darland, T., Heinricher, M. M., and Grandy, D. K. 1998.
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Reinscheid, R. K., Nothacker, H. P., Bourson, A., Ardati, A., Terenius, L. 1973. Characteristics of the receptor for narcotic
Henningsen, R. A., Bunzow, J. R., Grandy, D. K., Langen, H., analgesics in synaptic plasma membrane fraction from rat
Monsma, F. J., Jr, and Civelli, O. 1995. Orphanin FQ: a brain. Acta Pharmacol. Toxicol. (Copenh) 33, 377384.
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 55 Neuropathic Pain: Basic Mechanisms (Animal)
M H Ossipov and F Porreca, University of Arizona, Tucson, AZ, USA
Published by Elsevier Inc.

55.1 Introduction 833

55.2 Injury-Induced Sensitization of Peripheral Nerves and Neuropathic Pain 833
55.2.1 Role of Peripheral Nerve Degeneration in Neuropathic Pain 835
55.2.2 Role of Nerve Growth Factor in Immediate Pronociceptive Function 836
55.2.3 Nerve Growth Factor May Mediate Posttranslational Pronociceptive Functions 836
55.2.4 Sodium Channels and Enhanced Peripheral Nerve Activity 837
55.2.5 Role of Calcium Channels 839
55.3 Enhanced Afferent Discharges Lead to Central Sensitization in the Spinal Cord 840
55.4 Excitatory Transmitters Promote Central Sensitization 841
55.5 Descending Facilitation Promotes Enhanced Pain 842
55.5.1 Role of the Rostralventromedial Medulla in Descending Facilitation of Pain 842
55.5.2 The ON Cells of the Rostralventromedial Medulla Promote Pain Through Descending
Facilitation 843
55.5.3 Cholecystokinin in the Rostralventromedial Medulla May Mediate Pain Facilitation 844
55.6 Second-Order Projection Neurons Expressing NK1 Receptors May Result in
Activation of Descending Pain Facilitation 844
55.7 Descending Facilitation Maintains a Sensitized Spinal Cord: Upregulation of Spinal
Dynorphin and Enhanced Release of Primary Afferent Transmitters 845
55.7.1 Descending Facilitation Maintains a Sensitized Spinal Cord: Upregulation of Spinal
Dynorphin and Enhanced Release of Primary Afferent Transmitters 846
55.7.2 Descending Facilitation Maintains a Sensitized Spinal Cord: Serotonergic Contributions 846
55.8 Summary 847
References 848

55.1 Introduction described as shooting, burning, lancinating, prickling,

and electrical. Evoked, or stimulus-dependent, neu-
Neuropathic pain is defined by the International ropathic pain includes allodynia, defined by IASP as
Association for the Study of Pain (IASP) as pain pain due to a stimulus which does not normally
initiated or caused by a primary lesion or dysfunction provoke pain. These stimuli may be nonnoxious
in the nervous system. Although this definition heat, light touch, or even a puff of cool air.
functions well as a guide for the clinical assessment Moreover, mechanical allodynia may be static, as
of neuropathic pain, a definitive categorization of evoked by a light touch, or dynamic, as evoked by a
neuropathic pain is difficult because there are many light brush of the skin. Hyperalgesia is identified
descriptions of the pain syndrome. Neuropathic pain when a stimulus that normally produces a nocicep-
may arise from an actual injury to a peripheral nerve tive responses produces an exaggerated responses.
or it may arise in the absence of any obvious nerve The different forms through which neuropathic
damage (e.g., trigeminal neuralgia and complex pain manifests suggest that differing mechanisms
regional pain syndrome (CRPS), type I). Disease are likely to mediate the different features of the
processes, such as diabetic neuropathy, postherpetic condition. The numerous features of neuropathic
neuralgia resulting from varicella zoster infection pain can complicate an accurate clinical diagnosis.
(shingles), and complications from AIDS also may For this reason, efforts have emphasized the develop-
cause neuropathic pain, as may alterations in periph- ment of highly specific questionnaires emphasizing a
eral nerve function induced by chemotherapeutic unique set of symptoms that can aid clinical diagnosis
regimens. Neuropathic pain may include sponta- with relatively little effort. Features that appear to be
neous, or stimulus-independent, pain that has been most often related to neuropathic pain include tingling

833
834 Neuropathic Pain: Basic Mechanisms (Animal)

or numbness, pain evoked by heat or cold, and espe- spontaneous activity of nerve fibers in adjacent, unin-
cially, a sensation of heat or a burning-like quality is jured nerve roots and in peripheral nerves (Ali, Z.
associated with the neuropathic pain state (Krause, S. J. et al., 1999; Wu, G., et al., 2001). Such studies raised
and Backonja, M. M., 2003; Backonja, M. M. and the question of relative contributions of enhanced
Stacey, B., 2004; Bennett, M. I. et al., 2007). More excitability from injured and uninjured fibers in eli-
recently, a questionnaire identified at DN4 was citing neuropathic pain. Yoon Y. W. et al. (1996) used
designed to assess the presence of pain combined a model of neuropathic pain in which the L5 spinal
with parasthesias or dysesthesias (Bouhassira, D. et al., nerve was injured (the spinal nerve ligation or SNL
2005). model, Kim, S. H. and Chung, J. M., 1992) and
showed that the behavioral signs of pain were abol-
ished by dorsal rhizotomy or bupivacaine applied to
55.2 Injury-Induced Sensitization of the injured spinal nerves emphasizing the importance
Peripheral Nerves and Neuropathic of afferent drive. Moreover, dorsal rhizotomy or
Pain bupivacaine at L4 and L5 also abolished behavioral
signs of mechanical and cold allodynia, but not signs
There are several factors common to the neuropathic of ongoing pain (Yoon, Y. W. et al., 1996). The con-
pain state, regardless of the etiology that may initiate tribution of uninjured nerves, or uninjured adjacent
the condition. It has been repeatedly demonstrated fibers, to neuropathic pain has been debated. In an
that peripheral nerve injury is associated with a insightful study, Belzberg and coworkers (Li, Y. et al.,
heightened sensitivity of nerves to sensory stimuli, a 2000) used the SNL model and found that the beha-
phenomenon that has been termed peripheral sensi- vioral signs of experimental neuropathic pain
tization. Importantly, peripheral sensitization is not produced by ligation of the L5 spinal nerve were
restricted to the injured nerve fibers, but also not abolished by rhizotomy at L5, but rather were
includes adjacent, uninjured (but abnormal) nerves. abolished by rhizotomy at L4. These findings
Injured nerves may form a tangled mass of connec- suggested a prominent role for uninjured but abnor-
tive tissue termed a neuroma with distally sprouting mal adjacent nerve fibers following nerve injury and
regenerating axons. Physical manipulation of the were consistent with the suggested importance of
neuroma elicits sensations ranging from minor dys- uninjured, but abnormal, L4 afferents; these fibers
esthesias to intense pain. Electrophysiologic studies show significant redistribution of sodium channels
showed that both myelinated and unmyelinated such as NaV1.8 following L5 injury (Lai, J. et al.,
fibers in neuromas show a basal spontaneous activity 2002) and this may reflect a mechanism which drives
as well as enhanced discharges provoked by touch or neuropathic pain (see below). It has also been sug-
movement, indicating that the neuroma may serve as gested that sensitization of uninjured peripheral
an ectopic generator of action potentials. In addition, nerves may result, in part, from ephaptic excitation
cell bodies of the dorsal root ganglia (DRG) may also caused by the close proximity of injured and non-
generate ectopic discharges, thus producing sponta- injured nerves within the nerve trunks or in the DRG
neous hyperesthesias and pain as well as enhanced (Devor, M. and Wall, P. D., 1990; Attal, N. and
sensitivity to stimuli. Traumatic injury of the rat Bouhassira, D., 1999). Sensitization of peripheral
sciatic nerve produced by loose ligatures, the chronic nerves may mediate SNL-induced thermal hyperal-
constriction injury or CCI model, resulted in sponta- gesia, but may not mediate tactile allodynia. Unlike
neous discharges generated from the DRG. In this tactile allodynia, thermal hyperalgesia was abolished
study, discharges were recorded from 15% to 35% of by the intravenous infusion of the quaternary lido-
myelinated and 3% of unmyelinated fibers (Kajander, caine analogs QX-314 and QX-222, which do not
K. C. et al., 1992). Electrophysiological recordings per- cross the bloodbrain barrier (Chen, Q. et al., 2004).
formed on sciatic nerve bundles after CCI also In contrast, both tactile and thermal parameters of
demonstrated spontaneous and evoked ectopic dis- enhanced pain were abolished by systemic lidocaine,
charges from the region of injury (Tal, M. and or microinjection of lidocaine or QX-222 into the
Bennett, G. J., 1994). Surprisingly, these spontaneous rostralventromedial medulla (RVM) (Chen, Q. et al.,
discharges of DRG neurons occurred from cells of 2004). These observations emphasize the contribu-
both injured as well as intact nerves (Michaelis, M. tion of a central mechanism (addressed below) which
et al., 1996). Additionally, electrophysiologic studies may act in addition to peripheral mechanisms to
showed spinal nerve transection enhanced the elicit neuropathic pain states.
 Neuropathic Pain: Basic Mechanisms (Animal) 835

55.2.1 Role of Peripheral Nerve sensitivity to noxious stimuli perhaps as a conse-

Degeneration in Neuropathic Pain quence of upregulation of NGF by keratinocytes
(Taherzadeh, O. et al., 2003). Increased NGF produc-
Degeneration of peripheral nerves provokes the
tion promotes sprouting of peripheral nerve
release of proinflammatory cytokines such as inter-
terminals into the region into the boundary between
leukins and tumor necrosis factor (TNF), and
the denervated region and that with normal innerva-
neurotrophic factors in the region of the target tissue.
tion (Rajan, B. et al., 2003). Collateral sprouting of
The initial reaction is followed by macrophage infil-
peripheral axons together with the pronociceptive
tration that further mediate proinflammatory signals.
actions of peripheral NGF can promote nociceptive
This accumulation of proinflammatory substances
hypersensitivity in the region immediately surround-
may sensitize adjacent nerve terminals of uninjured
ing the zone of denervation (Griffin, J. W., 2006).
fibers. Myelinated peripheral nerve fibers under- Other growth factors, such as those in the glial-cell-
going Wallerian degeneration elicit the release of line-derived neutrotropic factor (GDNF) family
several pronociceptive factors in the target tissue including GDNF and artemin, may play a significant
and cytokines along the peripheral nerve, thereby role in expression of neuropathic pain. NGF and
sensitizing the adjacent nerve endings of uninjured GDNF family members may act at peripheral
nerve fibers, and exciting and potentially producing nociceptive primary afferents, leading to enhancement
an ectopic focus in the uninjured adjacent C fibers of sensitivity to noxious inputs (Zwick, M. et al., 2002;
(Li, Y. et al., 2000). Wallerian degeneration of the Elitt, C. M. et al., 2006; Griffin, J. W., 2006; Malin, S. A.
injured, spontaneously active myelinated fibers may et al., 2006). Injury-induced upregulation of growth
excite unmyelinated fibers through the comingling of factors such as NGF results in a situation where intact,
injured and uninjured nerves that occurs within the uninjured nociceptive C fibers expressing the trkA
Remak bundles of the sciatic nerve trunk (Wu, G. receptor are exposed to a surplus of NGF and may
et al., 2001; 2002). Furthermore, Wallerian degenera- become overtrophed in response to nerve injury
tion associated with injured nerves causes alterations (Griffin, J. W., 2006). In these fibers, NGF would
in the Schwann cells that provoke them to release promote nociceptive transmission through phosphor-
proinflammatory mediators, including glutamate as ylation of TRPV1 ion channels, as well as
well as nerve growth factor (NGF) (Shamash, S. et al., posttranslational mechanisms that include upregula-
2002; MacInnis, B. L. and Campenot, R. B., 2005). tion of ion channels, redistribution of NaV1.8 and
Within the nerve trunk, these substances may diffuse increased synthesis of substance P and CGRP
to adjacent uninjured nerve fibers and enhance sen- (Boucher, T. J. et al., 2000; Boucher, T. J. and
sitivity such that an ectopic locus may develop. McMahon, S. B., 2001; Griffin, J. W., 2006). In contrast,
Release of proinflammatory mediators including the axons of the injured peripheral fibers, which
NGF in the peripheral terminal fields that include include C fiber nociceptors as well as some myelinated
uninjured neurons may sensitize these fibers to ensu- sensory fibers, are not able to traffic target-derived
ing stimulation, thus producing peripheral growth factors to the cell body. These peripheral
sensitization. It has been shown that nerve injury nerves may be considered to be undertrophed
causes significant elevations in NGF and its mRNA (Griffin, J. W., 2006) and can demonstrate increased
in Schwann cells, macrophages, and fibroblasts at the expression of tetrodotoxin (TTX)-sensitive (TTX-S)
site of injury (Shamash, S. et al., 2002; MacInnis, B. L. sodium channels and enhanced spontaneous activity,
and Campenot, R. B., 2005). Treatment of cultured adaptations which may underlie such painful
fibroblasts or keratinocytes with inflammatory med- conditions as postamputation pain and neuropathic
iators, including histamine and interleukin-1 also pain associated with mastectomy and thoracotomy
provoke upregulation of NGF (Kanda, N. and scars (Griffin, J. W., 2006). This hypothesis was tested
Watanabe, S., 2003). Moreover, NGF promotes by providing support with exogenous trophic factors
degranulation of mast cells, which release inflamma- including GDNF and artemin. Administration of
tory mediators and thereby promotes peripheral exogenous GDNF normalized expression of sodium
sensitization of nociceptors as well as further release channels and the reversed the behavioral signs of
of NGF (Lewin, G. R. et al., 1994; Stempelj, M. and neuropathic pain (Boucher, T. J. et al., 2000; Boucher,
Ferjan, I., 2005). Denervation of the skin due to T. J. and McMahon, S. B., 2001; Wang, R. et al., 2003).
degeneration of peripheral nerves produce regions In addition, exogenous GDNF delivered by the
that are insensate surrounded by regions of increased intrathecal route also abolished electrophysiologic
836 Neuropathic Pain: Basic Mechanisms (Animal)

signs of enhanced firing, ectopic discharges, and nor- studies also showed that exogenous NGF produced
malized sodium channel currents in injured peripheral long-lasting mechanical allodynia and hyperalgesia
nerves (Boucher, T. J. et al., 2000; Boucher, T. J. and and mild-to-moderate myalgia (Apfel, S. C., 2000;
McMahon, S. B., 2001) as well as multiple biochemical Svensson, P. et al., 2003). Conversely, manipulations
markers in the DRG (Wang, R. et al., 2003). Systemic that block NGF activity alleviate signs of hyperalgesia.
treatment with artemin to rats with SNL also normal- Blockade of NGF activity with the trkAIgG fusion
ized the expression of biochemical markers of protein, that antagonist ALE-540 or with antiserum to
neuropathy, including the expression and redistribu- NGF abolished mechanical and thermal hyperalgesia
tion of NaV1.8 and abolished signs of neuropathic pain in animal models of inflammation, incision, or nerve
(Gardell et al., 2003b). The pronociceptive role of injury (Owolabi J. B. et al., 1999; Banik, R. K. et al.,
neurotrophic factors is discussed below. 2005). More recently, intrathecal injection of antibody
to NGF blocked behavioral signs of cold allodynia
after nerve ligation (Obata, K. et al., 2005). In addition,
55.2.2 Role of Nerve Growth Factor in
anti-NGF given by intrathecal injection or local appli-
Immediate Pronociceptive Function
cation onto the L4 spinal nerve abolished signs of
NGF binds preferentially to the high-affinity tyrosine thermal and mechanical hyperalgesia in rats with per-
kinase receptor subtype trkA and to the low-affinity, ipheral nerve injury (Obata, K. et al., 2004; 2006). The
nonselective neurotrophin receptor p75 (Sah, D. W. prominent role of NGF in the expression of chronic
et al., 2005a; 2005b). The neuronal distribution of the pain states has led a number of pharmaceutical com-
trkA receptor indicates a substantial role of NGF in panies, including Pfizer/Rinat, Amgen, and
nociceptive processes, since trkA is found almost PainCeptor Pharma to initiate drug discovery and
exclusively on small-diameter unmyelinated pepti- development efforts targeting NGF or trkA (Shelton,
dergic DRG neurons (Sah, D. W. et al., 2003). D. L. et al., 2005; Hefti, F. F. et al., 2006). Drug devel-
Double-labeling studies indicate that 92% of trkA- opment approaches include sequestering or binding
expressing DRG neurons also express CGRP and NGF so that it does not interact with the trkA recep-
substance P, whereas it is present on about 20% of tor, trkA receptor antagonists, or disruption of the
myelinated DRG neurons (Bennett, D. L. et al., 2000; signaling pathways linked to trkA receptor activation
Averill, S. et al., 2004). Investigations performed with a (Hefti, F. F. et al., 2006).
skinnerve preparation showed that NGF increased
mechanical hyperalgesia mediated by thin myelinated
55.2.3 Nerve Growth Factor May Mediate
fibers and thermal hyperalgesia mediated by unmye-
Posttranslational Pronociceptive Functions
linated nociceptors (Stucky, C. L. et al., 2002).
Electrophysiologic studies revealed that endogenous In addition to its immediate actions in the periphery,
NGF regulates the sensitivity of unmyelinated NGF can also initiate long-lasting posttranscriptional
nociceptors to chemical and thermal stimuli changes in peripheral nerves to promote and sustain
(Bennett, D. L. et al., 1998). Sensitization to heat sti- an enhanced pain state. When NGF binds to the trkA
mulus or to capsaicin application is induced rapidly by receptor at the peripheral terminal, it is rapidly inter-
NGF (Shu, X. and Mendell, L. M., 2001). Within nalized and the phosphorylated trkANGF complex
10 min of exposure to NGF, there is a rapid potentia- is accumulated in signaling endosomes that are
tion of capsaicin-evoked ion currents in cultured transported to the DRG, where it triggers the activa-
DRG neurons gated by TRPV1 (Shu, X. and tion of several intracellular signaling cascades,
Mendell, L. M., 2001). Activation of the trkA receptor including expression of the MAP kinases, ERK, and
by NGF leads to, among other events, phosphoryla- p38 ( Ji, R. R. et al., 2002; McMahon, S. B. and Cafferty,
tion of TRPV1, in part through the signaling cascades W. B., 2004). Inflammation or exogenous NGF admi-
associated with PKA, PKC, MEK, and the MAP nistered spinally causes phosphorylation of p38 in
kinases (Shu, X. and Mendell, L. M., 1999; 2001). nociceptors in the DRG, but not the dorsal horn,
Moreover, NGF increases nociceptor activity by a along with a substantial upregulation of TRPV1 in
rapid facilitation of sodium currents and suppression the DRG (Ji, R. R. et al., 2002; McMahon, S. B. and
of an outward potassium current. Behavioral hyperal- Cafferty, W. B., 2004; Malik-Hall, M. et al., 2005).
gesia is evoked by NGF administered systemically or Moreover, there is evidence that the TRPV1 receptor
into the endoneurium of a peripheral nerve (Ruiz, G. is preferentially trafficked to the peripheral terminals
et al., 2004; Ruiz, G. and Banos, J. E., 2005). Clinical of the nociceptors, where it can contribute to
 Neuropathic Pain: Basic Mechanisms (Animal) 837

peripheral sensitization and hyperalgesia (Ji, R. R. injured peripheral nerves are related to increased
et al., 2002). Furthermore, hyperalgesia may also be activity of voltage-gated sodium channels (VGSCs)
maintained by the increased expression of peptidic (Lai, J. et al., 2002; 2003). The VGSCs have been linked
neurotransmitters specific to primary afferent noci- to excitability of primary afferent nociceptors and may
ceptors. The exposure of DRG cultures to NGF mediate sensitized pain states (Lai, J. et al., 2003).
produced 15- to 60-fold increases in mRNA for sub- Peripheral nerve injury is also associated with changes
stance P and CGRP in trkA-expressing cells, and in the expression and distribution of isoforms of the
increased capsaicin-evoked release of CGRP or sub- VGSCs (Novakovic, S. D. et al., 2001). Three sodium
stance P from cultured trigeminal neurons and from channels merit special consideration.
central terminals of primary afferent nociceptors in The TTX-S sodium channel NaV1.3 is fast-activat-
spinal cord sections (McMahon, S. B. and Cafferty, W. ing channel normally present in brain tissue but scarce
B., 2004; Puntambekar, P. et al., 2005). Animals with in the DRG. However, peripheral nerve injury results
diabetic or chemotherapy-induced neuropathy trea- in a substantial expression this channel in peripheral
ted with NGF showed abnormally elevated levels of nerves, and it is believed that this novel expression may
substance P and CGRP protein and mRNA be partly responsible for the enhanced excitability of
(Tomlinson, D. R. et al., 1997). These functional con- the primary afferents (Waxman, S. G. et al., 1999; Lai, J.
sequences of NGF expression, including stimulation et al., 2003). However, although its biophysical proper-
of BDNF production, may enable transition from ties favor generation of spontaneous discharges, a causal
peripheral to central sensitization after injury, result- relationship between expression of NaV1.3 in the DRG
ing in chronic pain (Sah, D. W. et al., 2003; McMahon, and neuronal discharges has not been established. It had
S. B., et al., 2005; Sah, D. W. et al., 2005a; 2005b). been hypothesized that upregulation of NaV1.3 in per-
Moreover, retrogradely transported NGF promotes ipheral nerves might mediate neuropathic pain as
the upregulation of ion channels that promote activa- intrathecal treatment with GDNF abolished behavioral
tion of peripheral nerves and enhances nociceptive signs of enhanced pain, injury-induced enhanced per-
transmission. For example, hyperalgesia is also main- ipheral neuronal excitability, and the upregulation of
tained by NGF-mediated upregulation of ASIC3 and NaV1.3 (Boucher, T. J. et al., 2000). Other studies, how-
upregulation and redistribution of NaV1.8 on primary ever, showed that intrathecal GDNF normalized many
afferent nociceptors (McMahon, S. B. and Jones, N. changes observed in the DRG following peripheral
G., 2004). Importantly, it was shown that peripheral nerve injury (Wang, R. et al., 2003) making specific
nerve injury is associated with enhanced retrograde conclusions about NaV1.3 difficult. Observations that
transport of NGF to the cell bodies of spared, or knockdown of NaV1.3 by antisense treatment abolished
uninjured adjacent, peripheral nerves, thus promoting both the enhanced neuronal activity and behavioral
pronociceptive transcriptional changes in these signs of neuropathic pain in nerve-injured rats seemed
nerves as well. For example, ligation of the L5 spinal to support this hypothesis (Hains, B. C. et al., 2004).
nerve increased the expression of p75NTR in the However, more recent evidence has clearly shown that
DRG that was abolished by spinal administration of this channel may not be relevant to enhanced pain after
anti-NGF (Obata, K. et al., 2006). Moreover, the appli- nerve injury. Studies employing two different condi-
cation of anti-NGF onto the L4 spinal nerve after tional mutant mice lines, one with knockout of NaV1.3
ligation of the L5 spinal nerve blocked the injury- in nociceptors and another with global knockout of
induced upregulation of p75NTR (Obata, K. et al., NaV1.3 throughout the nervous system, showed that
2006). The spinal injection of the NGF/TrkA antago- behavioral signs of neuropathic pain developed nor-
nist K252a blocked injury-induced thermal and cold mally in both mutant lines after peripheral nerve
hyperalgesia and the upregulation of phosphorylated injury (Nassar, M. A. et al., 2006). Knockout of NaV1.3
trkA, p38, and of TRPV1 and TRPA1 in L4 DRG also did not alter responses to acute nociceptive stimuli
(Obata, K. et al., 2006). or to inflammation (Nassar, M. A. et al., 2006).
Accordingly, it is believed that the NaV1.3 channel is
neither sufficient nor necessary for the expression of
55.2.4 Sodium Channels and Enhanced
neuropathic pain (Nassar, M. A. et al., 2006).
Peripheral Nerve Activity
The TTX-insensitive sodium channel NaV1.8
Because of their critical role in the generation of action exhibits properties consistent with a role in neuro-
potentials and neuronal excitation, the increased pathic pain (Sangameswaran, L. et al., 1997; Lai, J.
abnormal spontaneous and ectopic discharges of et al., 2002). It is expressed primarily in small-diameter,
838 Neuropathic Pain: Basic Mechanisms (Animal)

unmyelinated peripheral nerve fibers (C fibers) and in nerve has been also observed in the chronic constric-
other cells in the DRG, and is not found in other tion injury model of neuropathic pain, and NaV1.8
peripheral or central neurons or nonneuronal tissue immunoreactivity is evident in peripheral nerve tissues
(Sangameswaran, L. et al., 1997; Akopian, A. N. et al., from patients with chronic neuropathic pain
1999). Studies employing mutant mice lacking the (Novakovic, S. D. et al., 1998; Coward, K. et al., 2000).
NaV1.8 channel or antisense knockdown techniques Immunohistochemical studies of human tooth pulp
have shown that this channel is not essential for resting obtained from painful and nonpainful teeth demon-
membrane potentials, neuronal excitation thresholds, strated that fine unmyelinated nerve fibers expressed
or basal nociceptive thresholds (Akopian, A. N. et al., NaV1.8 (Renton, T. et al., 2005). In addition, there was
1999; Renganathan, M., et al., 2001; Lai, J. et al., 2003). an increased expression of NaV1.8 in nerve fibers
In contrast, knockdown of NaV1.8 in rats with SNL obtained from painful teeth (Renton, T. et al., 2005).
reversed behavioral signs of neuropathic pain NaV1.7, a TTX-S VGSC is found predominantly
(Porreca, F. et al., 2000; Lai, J. et al., 2003), which is in small-diameter sensory DRG neurons and in sym-
consistent with the ability of NaV1.8 to enable DRG pathetic nerves and plays a prominent role in
neurons to fire repetitively after stimulation (Akopian, enhanced pain states (Wood, J. N. et al., 2004b).
A. N. et al., 1999; Renganathan, M., et al., 2001). There Recent investigations confirm that this channel med-
is evidence that suggests that behavioral neuropathic iates the intense burning pain associated with familial
pain may be partly mediated through an abnormal erythermalgia. Primary erythermalgia is a rare, auto-
redistribution of NaV1.8 after nerve injury (Lai, J. somal-dominant condition that manifests itself by
et al., 2003). Recent investigations found that the O- intermittent bouts of swelling of the feet or hands,
conotoxin MrVIB produced a selective block of the intense burning pain, and erythemia (Yang, Y. et al.,
NaV1.8 current (Ekberg, J. et al., 2006). Intrathecal 2004). The syndrome may be provoked by warmth or
injections of MrVIB produced dose-dependent exercise and is refractory to therapeutic interventions
attenuation of tactile allodynia and thermal hyperal- (Yang, Y. et al., 2004). It was found that individuals
gesia in rats with peripheral nerve injury, leading to afflicted with erythromelalgia expressed a T to A
the conclusion that blockade of NaV1.8 may provide transversion mutation in the gene SCN9A, which
one therapeutic avenue for the alleviation of neuro- codes the alpha-subunit of the NaV1.7 sodium chan-
pathic pain (Ekberg, J. et al., 2006). nel (Yang, Y. et al., 2004). These mutations of the
Interestingly, immunological and electrophysiolo- NaV1.7 sodium channel were found to promote
gical studies showed that SNL resulted in a abnormal activity of small-diameter nociceptors,
downregulation of NaV1.8 protein or conductance in which would be consistent with a small-fiber neuro-
the DRG of injured neurons but upregulation in adja- pathy (Drenth, J. P. et al., 2005). Electrophysiologic
cent, uninjured unmyelinated, and myelinated DRG studies demonstrated that the mutated channel pro-
neurons (Porreca, F. et al., 1998; Gold, M. S. et al., 2003). duced a hyperpolarizing shift in activation, slowed
This redistribution of NaV1.8 after SNL was blocked deactivation of the channel and markedly increased
by antisense knockdown techniques (Gold, M. S. et al., the ramp current, with the net effect being a marked
2003; Lai, J. et al., 2003). The injury-induced changes in hyperexcitation of the cells (Dib-Hajj, S. D. et al.,
immunoreactivity of NaV1.8 was correlated with an 2005). The NaV1.7 sodium channel is upregulated
increase in tetrodotoxin-resistant (TTX-R) compound after carrageenan-induced inflammation, and geneti-
action potential at C fiber conduction velocity along cally modified mice with knockdown of the NaV1.7
with a minor contribution at A fiber conduction channel of sensory neurons showed an attenuated
velocity, suggesting a functional reorganization of inflammation-induced hyperalgesia (Wood, J. N.
NaV1.8 along unmyelinated fibers and in some myeli- et al., 2004a). More recently, voltage clamp studies
nated fibers (Gold, M. S. et al., 2003). Antisense- performed on transfected mouse DRG cells showed
mediated knockdown of NaV1.8 immunoreactivity that the human mutations lowered the thresholds for
and TTX-R current in these uninjured axons corre- generation of action potentials and repetitive firing,
lates with the reversal of both mechanical and thermal thus provoking high-frequency firing of nociceptive
hypersensitivity, suggesting that this reorganization of small-diameter sensory neurons in the DRG (Dib-
NaV1.8 activity along the uninjured axons may be Hajj, S. D. et al., 2005). It is now believed that
necessary for expression of neuropathic pain in the enhanced sensitivity of this sodium channel may
injured rat (Gold, M. S. et al., 2003; Lai, J. et al., 2003). exacerbate activation of the TRPV1 channels, thus
A redistribution of NaV1.8 along the injured sciatic driving nociceptor hyperexcitability.
 Neuropathic Pain: Basic Mechanisms (Animal) 839

More recent studies were undertaken with patients 1987). This channel is predominant on nerve terminals
with the autosomal dominant spontaneous pain con- of primary afferent terminals in the superficial lamina
dition, paroxysmal extreme pain disorder (PEPD), of the spinal dorsal horn and participates in nociceptive
which manifests as paroxysmal episodes of burning transmission since it is critical to depolarization-
pain in ocular, mandibular, and rectal regions. coupled release of neurotransmitters, including sub-
Mutational analysis of SCN9A obtained from indivi- stance P, CGRP, and glutamate, from these terminals
duals with PEPD and unafflicted family members (Santicioli, P. et al., 1992). It was suggested that nerve
revealed that the mutation was associated with injury results in either increased frequency of opening
enhanced NaV1.7 current activity (Fertleman, C. R. of the N-type calcium channel, or an increase in the
et al., 2006). Although primary erythermalgia and population (see Snutch, T. P. et al., 2001; Snutch T. P.,
PEPD are due to mutations of SCN9A, these familial 2005, for reviews). Blocking the activity of the N-type
disorders are considered allelic variants that are clini- calcium channel with the omega-conotoxinGVIA
cally and mechanistically distinct, related to the abolished enhanced electrically evoked responses of
specific effects of the mutations on the sodium chan- dorsal horn neurons as well as windup and post-
nel activity (Fertleman, C. R. et al., 2006). On the other discharge phenomena in nerve-injured rats, which is
side of the spectrum, individuals with one of three consistent with inhibition of signs of enhanced pain
distinct nonsense mutations of SCN9A show a loss of (Matthews, E. A. and Dickenson, A. H., 2001b). The
function of the NaV1.7 channel activity show a spinal administration of a novel N-type calcium
marked insensitivity to pain (Cox, J. J. et al., 2006). channel blocker ziconitide produced antinociception
This sodium channel appears to be critical to noci- in uninjured rats and abolished tactile and thermal
ceptive processing, both acutely and in chronic pain hyperesthesias in those with peripheral nerve injuries
states (Cox, J. J. et al., 2006; Fertleman, C. R. et al., (Scott, D. A. et al., 2002; Wang, Y. X. et al., 2000a; 2000b).
2006). In spite of the evidence pointing to a prominent Recent clinical trials with ziconotide (PRIALT)
role of voltage-gated sodium channels in nociception provide strong evidence that blockade of the N-type
and enhanced pain due to nerve injury, therapeutic calcium channel may provide a significant clinical
interventions based on blockade of the sodium chan- benefit in the management of severe, refractory pain,
nels have, to this point, been disappointing. A large such as deafferentation pain or pain due to cancer or
multicenter clinical investigation that was recently neuropathies associated with HIV/AIDS and posther-
concluded showed that lamotrigine, which is a non- petic neuralgia (Wang, Y. X., et al., 2000a; 2000b;
selective sodium channel blocker, failed to produce Miljanich, G. P., 2004; Prommer, E. E., 2005).
changes that were different from placebo in patients The T-type calcium channel is present in DRG
with diabetic neuropathy (Vinik, A. I. et al., 2007). A neurons and on second-order neurons of the dorsal
multicenter clinical investigation with oxcarbazepine horn of the spinal cord (Talley, E. M. et al., 1999; see
for diabetic neuropathy suggested that this nonselec- Hildebrand, M. E. and Snutch, T. P., 2006, for a
tive sodium channel blocker produced a moderate review). Activation of the NK1 receptor and of the
effect that may be clinically meaningful (Dogra, S. T-type voltage-gated calcium channels (VGCCs)
et al., 2005). Though currently unclear, one conclusion acts synergistically with activation of the NMDA/
is that sodium channel blockers may not be suitable calcium channel complex to facilitate firing of the
for routine use against neuropathic pain, but may be NK1-expressing cells of lamina I, thus promoting
appropriate for responsive subgroups of neuropathic central sensitization (Ikeda, H. et al., 2003). This
pain patients (Sindrup, S. H. and Jensen, T. S., 2007). observation is consistent with observations that
One such example may be patients with postherpetic blockade of the T-type VGCC produced dose-
neuralgia mediated by irritable nociceptors (Sindrup, dependent reversal of behavioral signs of neuropathic
S. H. and Jensen, T. S., 2007). pain in rats with peripheral nerve injury (Dogrul, A.
et al., 2003). Moreover, pharmacologic blockade of
T-type VGCCs also diminished electrophysiologic
55.2.5 Role of Calcium Channels
signs of postsynaptic sensitization indicated by
Calcium channels also are critical elements that mod- windup and after-discharges in nerve-injured rats
ulate neuronal excitation (Snutch, T. P. et al., 2001). (Matthews, E. A. and Dickenson, A. H., 2001a). The
The N-type calcium channel, which is blocked by role of this VGCC in pain modulation makes the
omega-conotoxin, has been identified in DRG neurons T-type calcium channel an attractive target for
(Nowycky, M. C. et al., 1985; McCleskey, E. W. et al., drug development. Although several compounds are
840 Neuropathic Pain: Basic Mechanisms (Animal)

under preclinical investigation as T-type channel The state of central sensitization is generally char-
blockers, no clinical data exist with which to gauge acterized by electrophysiologic parameters that include
their clinical utility against neuropathic pain a reduction in sensory thresholds, increased responsive-
(Hildebrand, M. E. and Snutch, T. P., 2006). ness of second-order neurons to afferent inputs along
with after-discharges, and expansion of the receptive
field along with recruitment of afferent fibers that are
55.3 Enhanced Afferent Discharges not normally nociceptive (Cook, A. J. et al., 1987).
Lead to Central Sensitization in the Behaviorally, a reduction in sensory threshold is
Spinal Cord demonstrated by the significantly reduced withdrawal
latencies to noxious thermal stimuli or reduced thresh-
The pronociceptive changes that occur in peripheral olds to noxious mechanical stimuli (Woolf, C. J. and
afferent fibers after nerve injury promote enhanced Thompson, S. W., 1991; Hedo, G. et al., 1999). In
activity of these fibers, and this afferent drive is addition, light tactile stimuli that do not elicit
thought to result in hyperalgesia and allodynia behavioral responses in normal animals produce noci-
(Amir, R. et al., 2002). However, although although fensive behaviors in the sensitized state (Woolf, C. J.
onset of neuropathic pain is consistent with the initia- and Thompson, S. W., 1991; Hedo, G. et al., 1999;
tion of the increased afferent discharges, the enhanced Ossipov, M. H. and Porreca, F., 2005). For example,
activity of peripheral fibers diminishes fairly rapidly, sensitization induced by the intradermal injection of
whereas the behavioral signs of enhanced pain persist capsaicin reduced pain thresholds to noxious heat and
long after the initial insult, indicating that mechanisms elicited allodynia to light brush in human volunteers
which sustain neuropathic pain may differ from those (Simone, D. A. et al., 1989). Similarly, intradermal injec-
which initiate such pain (Liu, C. N. et al., 2001; tion of capsaicin reduced the stimulus threshold
Ossipov, M. H. et al., 2005; Sun, Q. et al., 2005). There required to elicit responses of the spinothalamic tract
is general agreement that these behavioral manifesta- (STT) neurons to light brush and also increased the
tions of enhanced pain are mediated in large part by response intensity of these second-order neurons
an enhanced responsiveness of the spinal cord to (Simone, D. A. et al., 1991). Peripheral nerve injury
sensory inputs, or central sensitization, which may be was shown to increase the spontaneous activity of dor-
both initiated, and in part, sustained by the primary sal horn neurons and to enhance the responses of these
afferent barrage. Sensitization of second-order neurons neurons to noxious heat or light touch applied to the
of the spinal dorsal horn has been illustrated experi- injured hindpaw (Suzuki, R. and Dickenson, A., 2005a;
mentally by the phenomena of windup. Windup is 2005b). Moreover, nerve injury also provoked pro-
characterized by progressively increasing responses nounced after-discharges and significantly enlarged
of spinal dorsal horn neurons in response to repetitive the receptive field of dorsal horn neurons in response
electrical stimulation of C fibers (Mendell, L. M., 1966; to noxious heat or pinch (Suzuki, R. et al., 2004a; Suzuki,
Woolf, C. J., 1996). Windup is relatively short term in R. and Dickenson, A., 2005a; 2005b). Peripheral nerve
duration and occurs only in response to stimulation of injury also caused a shift to the left of the stimulus
C fibers, and, in the uninjured state, not of A-beta response function of wide dynamic range (WDR) neu-
fibers, and is nociceptive specific (Woolf, C. J., 1996; rons in response to tactile, but not thermal, stimuli
Ji, R. R. et al., 2003; Ji, R. R. and Strichartz, G., 2004). (Kauppila, T. et al., 1998; Pertovaara, A. et al., 2001).
Conditioning stimuli in the form of electrically evoked In addition to electrophysiologic means, immuno-
trains of C fiber activity has been found to produce histochemical detection of the proto-oncogene product
long-term responsiveness of dorsal horn units to addi- FOS in the spinal cord has been widely employed as an
tional stimuli, a profile that appears similar to long- indicator for neurons that are activated by noxious
term potentiation (LTP) (Woolf, C. J., 1996; Ji, R. R. inputs, and its enhanced expression in the spinal dorsal
et al., 2003; Ji, R. R. and Strichartz, G., 2004). Although horn is suggestive of sensitization (Hunt, S. P. et al.,
these dorsal horn neurons are unreactive to A-beta 1987; Shortland, P. and Molander, C., 1998). Peripheral
fiber inputs in the resting state, they become sensitized nerve injury produced significant increases in noxious
through repetitive C fiber stimulation which can ren- and touch-evoked FOS expression in the superficial
der them excitable by A-beta fiber inputs, and this may and intermediate laminae of the spinal dorsal horn
represent a mechanism which contributes to touch- (Catheline, G. et al., 1999; Vera-Portocarrero, L. P.
evoked allodynia (Woolf, C. J., 1996; Ji, R. R. et al., et al., 2006; Zhang, E. T. et al., 2007). Electrical stimula-
2003; Ji, R. R. and Strichartz, G., 2004). tion at A-beta fiber intensity or application of light
 Neuropathic Pain: Basic Mechanisms (Animal) 841

touch evoked FOS expression, but only after nerve afferent terminals evoked by spinal NMDA was abol-
injury or inflammation (Ma, Q. P. and Woolf, C. J., ished by capsaicin, indicating that presynaptic NMDA
1996; Shortland, P. and Molander, C., 1998; Nomura, receptors provoke further release of excitatory trans-
H. et al., 2002). Recently, gentle stroking of the hindpaw, mitters (Liu, H. et al., 1997). Glutamate may also act at
which did not evoke FOS expression in normal rats, but presynaptic metabotropic receptors to promote sensi-
caused significant increases in FOS expression after tization (Ohishi, H. et al., 1995). Prostacyclin and PGE2
nerve injury (Catheline, G. et al., 1999; Zhang, E. T. enhanced capsaicin-evoked release of substance P and
et al., 2007). The enhanced expression of FOS was found CGRP through presynaptic excitation (Southall, M. D.
in the superficial and deep laminae of the spinal dorsal et al., 1998; Southall, M. D. and Vasko, M. R., 2001;
horn (Zhang, E. T. et al., 2007). Together, these studies Southall, M. D. et al., 2002). These observations are
indicate that peripheral nerve injury can promote a consistent with the fact that central sensitization eli-
state of sensitization of spinal dorsal horn neurons. cited by C fiber stimulation or inflammation was
blocked by NMDA antagonists (Woolf, C. J. and
Salter, M. W., 2000).
55.4 Excitatory Transmitters A measure indicative of the level of primary affer-
Promote Central Sensitization ent drive into the spinal dorsal horn is the
determination of internalization of the NK1 receptor
Sensitization of the spinal cord elicited by enhanced on second-order neurons that are activated by sub-
peripheral afferent inputs results in part from the stance P released from primary afferent nociceptors
recruitment of subthreshold stimuli to evoke a neuronal (Allen, B. J. et al., 1999; Honore, P. et al., 1999). Under
response that is related to increased release of excitatory nonsensitized conditions, noxious stimulation, such
neurotransmitters form the primary afferent terminals as electrical stimulation at C fiber intensity, subcuta-
(Li, Q. J. and Lu, G. W., 1989; Li, J. et al., 1999). neous capsaicin or noxious heat or pinch, will evoke
Microdialysis studies demonstrated the release of glu- rapid internalization of the NK1 receptor almost
tamate and aspartate from primary afferents in response exclusively in the outer lamina of the spinal cord
to the capsaicin injected intradermally in the plantar (Allen, B. J. et al., 1999; Honore, P. et al., 1999).
aspect of the hindpaw of the rat or electrical stimulation Moreover, nonnoxious stimulation does not evoke
of the sciatic nerve at C fiber intensity (Paleckova, V. internalization of NK1. However, when sensitization
et al., 1992; Sluka, K. A. and Willis, W. D., 1998). occurs due to inflammation or peripheral nerve
Similarly, behavioral hypersensitivity and central sen- injury, there is a marked enhancement of noxious-
sitization due to peripheral nerve injury or induced NK1 internalization, and NK1 internaliza-
inflammation are associated with increased evoked tion is found in the deeper laminae of the spinal cord
release of glutamate and aspartate from primary afferent as well (Allen, B. J. et al., 1999; Honore, P. et al., 1999).
terminals (Paleckova, V. et al., 1992; Kawamata, M. and In addition, nonnoxious stimuli such as light brush
Omote, K., 1996; Sluka, K. A. and Willis, W. D., 1998; also evoke NK1 internalization in the intermediate and
Kawamata, M. and Omote, K., 1999). Spontaneous and deep laminae of the spinal dorsal horn (Allen, B. J. et al.,
stimulus-evoked release of substance P and CGRP 1999; Honore, P. et al., 1999; Khasabov, S. G. et al.,
from primary afferent terminals is increased after 2002). Critically, electrical stimulation at A-beta fiber
peripheral nerve injury (Paleckova, V. et al., 1992; intensity does not elicit NK1 internalization after
Sluka, K. A. and Willis, W. D., 1998; Wallin, J. and nerve injury, indicating that the effect produced by
Schott, E., 2002; Gardell, L. R. et al., 2003a; 2003b). light brush is not a direct result of large-diameter
Increased release of excitatory neurotransmitters, fiber stimulation, but is likely to be secondary to
including glutamate, substance P, and CGRP is other mechanisms driving sensitization (Allen, B. J.
believed to contribute to central sensitization and to et al., 1999; Khasabov, S. G. et al., 2002). NK1 interna-
hyperalgesia. For example, CGRP release after intra- lization observed in the deeper laminae may be due in
dermal capsaicin increased WDR activity in the dorsal part to diffusion of substance P to NK1 receptors on
horn in response to brush or pinch (Sun, R. Q. et al., sensitized deep lamina neurons, or partly due to the
2003; Sun, R. Q. et al., 2004). small number of direct synaptic contacts between C
The existence of excitatory NMDA autoreceptors fibers and deep lamina neurons (Allen, B. J. et al., 1999;
on central terminals of primary afferents has been Khasabov, S. G. et al., 2002).
described (Liu, H. et al., 1997). Nociception and Recent studies demonstrated that the postsynaptic
release of substance P and glutamate from primary second-order neurons of the spinal dorsal horn
842 Neuropathic Pain: Basic Mechanisms (Animal)

express message for the bradykinin B2 receptor Gardell, L. R. et al., 2003a; 2003b). Most recently, it
(Wang, H. B. et al., 2005). Patch-clamp studies was shown that activation of descending facilitation
showed that activation of the B2 receptor produces from the RVM was subsequent to stimulation of
a dose-dependent potentiation of AMPA and NMDA NK1-expressing lamina I postsynaptic cells and
currents in dorsal horn neurons. Excitation of the B2 increased the neuronal responses of WDR neurons
receptor mediates increased frequency and ampli- of the deeper laminae of the dorsal horn after nerve
tude of AMPA-mediated excitatory postsynaptic injury or inflammation (Suzuki, R. et al., 2002; 2004a;
potentials, whereas B2 antagonists abolished electro- Rahman, W. et al., 2004; 2006). Spinal ondansetron
physiologic signs of central sensitization in dorsal blocked the sensitization of these WDR neurons,
horn neurons (Wang, H. B. et al., 2005). These studies which is consistent with a descending serotonergic
suggest that spinal bradykinin B2 receptors represent facilitatory pathway from the RVM (Suzuki, R. et al.,
an important postsynaptic mechanism in promoting 2002; Rahman, W. et al., 2004; Suzuki, R. et al., 2004a;
central sensitization and enhanced pain through the Rahman, W. et al., 2006).
potentiation of glutaminergic synaptic transmission
(Wang, H. B. et al., 2005). While the importance of
55.5.1 Role of the Rostralventromedial
bradykinin receptors in enhancing pain, including
Medulla in Descending Facilitation of Pain
chronic neuropathic pain (see below) seem clear,
the mechanism by which the receptors are activated The RVM and surrounding tissue has long been
is not known. recognized as a region critical to nociceptive proces-
sing and receives inputs from ascending nociceptive
pathways as well as from descending pain modula-
55.5 Descending Facilitation tory sites. The RVM is considered to be the final
Promotes Enhanced Pain common relay for descending inhibition of nocicep-
tive inputs. More recently, this region has also been
The existence of a pronociceptive pain facilitatory demonstrated to mediate a significant descending
system projecting from the RVM has been well estab- pain facilitatory system as well (Fields, H. L. and
lished (see Porreca, F. et al., 2002; Heinricher, M. M. Basbaum, A. I., 1999; Porreca, F. et al., 2002;
et al., 2003; Ossipov, M. H. and Porreca, F., 2005 for Heinricher, M. M. et al., 2003; Ossipov, M. H. and
reviews). It has been demonstrated that the micro- Porreca, F., 2005). Stimulation of this region, either
injection of lidocaine into the RVM abolished through electrical stimulating electrodes or through
behavioral signs of enhanced pain after nerve injury the microinjection of glutamate or neurotensin,
or inflammation, and abolished the sensitization of resulted in biphasic effects on behavioral and
spinal WDR neurons to mustard oil induced inflam- electrophysiologic responses to noxious stimuli
mation (Mansikka, H. and Pertovaara, A., 1997; (Zhuo, M. and Gebhart, G. F., 1997; Fields, H. L.
Burgess, S. E. et al., 2002; Pertovaara, A. and and Basbaum, A. I., 1999). Whereas high levels of
Almeida, A., 2006). Lesions of the descending projec- stimulation inhibited nociceptive responses, lower
tions from the RVM in the dorsolateral funiculus also levels of stimulation facilitated the same responses
abolished behavioral hypersensitivity after inflam- (Zhuo, M. and Gebhart, G. F., 1997; Urban, M. O.
mation or nerve injury (Wei, F. et al., 1998; 1999; et al., 1999). Similarly, microinjection of the neuro-
Ossipov, M. H. et al., 2000; Burgess, S. E. et al., 2002). tensin antagonist SR48692 into the RVM also
Finally, it was found that selective ablation of RVM produced a biphasic effect on nociceptive responses.
neurons that express the mu-opioid receptor, and Taken together, these observations indicate that the
which may label cells which contribute to descending RVM is capable of mediating both pain inhibitory
facilitation, also abolished behavioral signs of sensiti- functions, which are likely to predominate under
zation and the enhanced capsaicin-evoked release of basal conditions, and facilitation of pain, which
CGRP (Burgess, S. E. et al., 2002; Gardell, L. R. et al., would predominate under conditions of enhanced
2003a; 2003b; Porreca, F. et al., 2002). Importantly, pain (Ossipov, M. H. et al., 2001). Pharmacologic
disruption of descending facilitation abolished manipulations that attenuate RVM activity also
behavioral hypersensitization during later, but not block enhanced nociception. For example, thermal
initial, phases after nerve injury, indicating that it is hyperalgesia induced by naloxone-precipitated with-
essential to the maintenance of a sensitized state drawal or by previous prolonged exposure to a
(Burgess, S. E. et al., 2002; Porreca, F. et al., 2002; nociceptive stimulus can be blocked by the
 Neuropathic Pain: Basic Mechanisms (Animal) 843

microinjection of lidocaine into the RVM (Kaplan, H. and behavioral parameters of nociception through
and Fields, H. L., 1991; Morgan, M. M. and descending inhibition from the RVM (Fields, H. L.
Heinricher, M. M., 1992). It is likely that activation and Heinricher, M. M., 1985; Neubert, M. J. et al.,
of these pronociceptive circuits in the RVM in 2004). In contrast, the electrophysiologically deter-
response to persistent or prolonged noxious inputs mined response characteristics of the ON cells are
leads to a facilitation of more pain (Porreca, F. et al., consistent with a pronociceptive function, and it is
2002). Tail withdrawal responses to noxious thermal believed that these neurons serve as the source of
or mechanical stimuli can be facilitated by the sub- descending facilitation from the RVM (Fields, H. L.
cutaneous injection of formalin into a hindpaw, and and Heinricher, M. M., 1985; Heinricher, M. M. et al.,
this effect can be reversed by local infusion of the 2003; Neubert, M. J. et al., 2004). Accordingly, manip-
hydrophilic lidocaine derivative QX-314 injected at ulations that promote nociceptive responsiveness
the site of formalin (Calejesan, A. A. et al., 1998). It is also increase ON cell activity, which is consistent
currently understood that descending pain facilitatory with pain facilitation.
systems arising from the RVM are likely mediated The enhanced behavioral responses to noxious
through spinopetal serotonergic projections that stimuli elicited during naloxone-precipitated with-
enhance nociceptive inputs at the level of the spinal drawal from morphine correlates with increased
cord. The relationship among noxious inputs, the spontaneous activity of the RVM ON cells, and
RVM, and descending serotonergic projections to both behavioral hyperalgesia and enhanced on cell
chronic pain states is described in greater detail below. activity are abolished by lidocaine microinjected into
the RVM (Bederson, J. B. et al., 1990; Kaplan, H. and
Fields, H. L., 1991). Prolonged application of a nox-
55.5.2 The ON Cells of
ious thermal stimulus increased ON cell activity and
the Rostralventromedial Medulla Promote
facilitated behavioral nociceptive responses, and both
Pain Through Descending Facilitation
the electrophysiologic and behavioral responses were
Neuronal populations within the RVM have been abolished by microinjection of lidocaine into the
described as ON cells, OFF cells and neutral cells RVM (Kaplan, H. and Fields, H. L., 1991; Morgan,
based upon electrophysiologic responses to noxious M. M. and Fields, H. L., 1994). Electrophysiologic
heat applied to the tail of lightly anesthetized rats monitoring of RVM neurons demonstrated that the
(Fields, H. L., 2000; Ossipov, M. H. et al., 2001; microinjection of either cholecystokinin (CCK) or
Neubert, M. J. et al., 2004). By definition, the OFF neurotensin at doses that selectively activated on
cells are tonically active and pause in firing immedi- cells produced facilitated behavioral responses to
ately before a withdrawal response from a noxious nociceptive stimuli in lightly anesthetized rats
thermal stimulus, whereas the ON cells accelerate (Heinricher, M. M. and Neubert, M. J., 2004;
firing immediately before the nociceptive reflex Neubert, M. J. et al., 2004). Application of mustard
occurs (Fields, H. L. and Heinricher, M. M., 1985; oil to the skin of the rat produced increased firing of
Neubert, M. J. et al., 2004). As the name implies, the ON cells and thermal hyperalgesia that were both
activity of the neutral cells does not correlate with blocked by NMDA antagonist application into the
nociceptive inputs. Characterization of the neurons RVM (Kincaid, W. et al., 2006; Xu, M. et al., 2007). It
of the RVM were initially determined in lightly was also shown that inflammation induces an upre-
anesthetized rat preparations, and it was found that gulation of NMDA receptors on the ON cells, along
tail flick latency was longer during periods of with increased phosphorylation of AMPA receptors,
increased OFF cell activity and shorter when the thus enhancing ON cell activity and facilitating noci-
ON cells were active. The considerable body of ceptive responses (Guan, Y. et al., 2003; 2004). Finally,
evidence collected subsequent to these studies con- recent studies showed that inflammation results in
firmed the role of these neurons in the modulation of upregulation of BDNF, which facilitates ON cell
nociception and extended the findings to other mod- firing through an NMDA-dependent mechanism
els of acute and enhanced pain. It is now generally (Guo, W. et al., 2006). Most recently, it was discov-
accepted that the activity of OFF cells correlates with ered that capsaicin injected into a hindpaw or
inhibition of nociceptive inputs and of behavioral iontophoretic application of Sar9,Met(O2)11-sub-
nocifensive responses to nociception. In the presence stance P into the RVM enhanced ON cell activity
of opioids, OFF cells continue firing after a noxious and potentiated the responses of ON cells to ionto-
stimulus is applied and inhibit electrophysiologic phoretic NMDA (Budai, D. et al., 2007). Conversely,
844 Neuropathic Pain: Basic Mechanisms (Animal)

iontophoretic administration of an NK1 antagonist intermediate laminae of the dorsal horn and promoted
into the RVM abolished the enhanced responses of novel expression of FOS in response to light brush
ON cells (Budai, D. et al., 2007). Taken together, (Vera-Portocarrero, L. P. et al., 2006; Zhang, E. T. et al.,
these studies indicate that ON cells promote noci- 2007). Microinjection of dermorphinsaporin conju-
ception, and that enhanced ON cell activity underlies gate into the RVM abolished enhanced FOS
hyperalgesia associated with chronic pain states. In expression due to SNL (Vera-Portocarrero, L. P.
addition, it is suggested that substance P in the RVM et al., 2006). These observations indicate that the main-
may modulate enhanced ON cell activity, and con- tenance phase of the neuropathic pain state is
sequently, enhanced pain states. dependent on the certain neuroplastic changes that
In light of these observations, it has been proposed result in activation of a tonic descending pain facilita-
that neuropathic pain due to peripheral nerve injury tion from the RVM, whereas the earlier initiation
may be initially mediated through enhanced afferent phase is independent of descending systems.
activity, but that the enhanced discharges lead to a
sensitized spinal cord and neuroplastic changes cul-
55.5.3 Cholecystokinin in
minating in an enhancement of descending
the Rostralventromedial Medulla May
facilitation mediated through ON cells of the RVM
Mediate Pain Facilitation
(Heinricher, M. M. et al., 2003; Ossipov, M. H. et al.,
2005; Ossipov, M. H. and Porreca, F., 2006). It was recently shown that CCK opposes the anti-
Considerable electrophysiologic evidence exists to nociceptive activity of morphine by attenuating the
indicate that the population of RVM neurons that morphine-induced activation of OFF cells, which
express the mu-opioid receptor, or a subset of these constitute part of the descending inhibition of noci-
cells, are likely to drive descending facilitation ceptive inputs (Heinricher, M. M. et al., 2001;
(Heinricher, M. M. et al., 1994; Neubert, M. J. et al., Heinricher, M. M. and Neubert, M. J., 2004). The
2004). Based on this evidence, presumptive pain microinjection of CCK into the RVM produces
facilitatory neurons of the RVM were selectively enhancement of responses to light tactile or noxious
ablated by using the mu-opioid receptor as a portal thermal stimuli similar to that observed after SNL
of entry into these cells for dermorphin conjugated (Kovelowski, C. J. et al., 2000; Xie, J. Y. et al., 2005).
with the cytotoxin saporin (Porreca, F. et al., 2001; Moreover, the microinjection of CCK2 antagonists
Burgess, S. E. et al., 2002; Vera-Portocarrero, L. P. into the RVM attenuated behavioral signs of neuro-
et al., 2006). With this technique, when the agonist pathic pain in rats with SNL (Kovelowski, C. J. et al.,
binds to the receptor, internalization occurs, bringing 2000; Xie, J. Y. et al., 2005). The behavioral hyper-
the conjugated agonist and saporin complex into the esthesias and hyperalgesia elicited by RVM CCK
cell body, where saporin is released and leads to were abolished by dorsolateral funiculus (DLF)
inhibition of ribosome activity, thus causing cell lesions or microinjection of L365,260 (Kovelowski,
death within 23 weeks (Mantyh, P. W. et al., 1997). C. J. et al., 2000; Xie, J. Y. et al., 2005). In addition to its
Selective ablation of pain facilitatory neurons of the activities in inhibiting morphine-induced activation
RVM by microinjection of the dermorphinsaporin of OFF cells, it was also recently demonstrated that
conjugate either 7 days prior to SNL or once tactile CCK provokes a direct and selective activation of
and thermal hyperesthesias were well established, ON cells of the RVM and that this activation is
respectively, prevented and reversed the behavioral directly related to the production of hyperalgesia
signs of neuropathic pain (Porreca, F. et al., 2001; (Heinricher, M. M. and Neubert, M. J., 2004).
Burgess, S. E. et al., 2002). Importantly, these beha-
vioral signs of neuropathic pain were abolished 1
week after SNL, and not earlier. Similarly, physical 55.6 Second-Order Projection
disruption of descending facilitatory tracts from the Neurons Expressing NK1 Receptors
RVM or microinjection of lidocaine at different time May Result in Activation of Descending
points after SNL produced similar results (Porreca, F. Pain Facilitation
et al., 2001; Burgess, S. E. et al., 2002). Enhanced expres-
sion of FOS in the spinal cord is indicative of Converging evidence strongly indicates that central
sensitization. Recently, it was shown that SNL sensitization and hypersensitivity to sensory stimuli
enhanced both the intensity and laminar distribution is dependent predominantly on the long-term sensi-
of FOS evoked by noxious stimuli to include the tization of the lamina I cells that express the NK1
 Neuropathic Pain: Basic Mechanisms (Animal) 845

receptor (Ikeda, H. et al., 2003). Electrophysiologic 55.7 Descending Facilitation

studies have revealed that approximately 75% of Maintains a Sensitized Spinal Cord:
the neurons of lamina I are nociceptive specific, Upregulation of Spinal Dynorphin and
and the remainder are either WDR neurons or poly- Enhanced Release of Primary Afferent
modal nociceptors that also respond to noxious Transmitters
cold (Suzuki, R. and Dickenson, A. H., 2005a;
2005b). Approximately one-half of the lamina I neu- The enhancement of descending facilitation from the
rons express the NK1 receptor and project to RVM is believed to lead to neuroplastic changes in
supraspinal sites that process nociception, including the spinal cord that favor a sensitized state and
the parabrachial region and the thalamus (Suzuki, R. enhanced nociceptive inputs. A number of studies
and Dickenson, A. H., 2005a; 2005b). Recent studies have demonstrated that enhanced descending facil-
showed that selective ablation of lamina I neurons itation elicited by peripheral nerve injury is
that express the NK1 receptor abolish behavioral associated with an upregulation of spinal dynorphin
signs of neuropathic pain in rats with SNL content and enhanced release of excitatory neuro-
(Nichols, M. L. et al., 1999; Honore, P. et al., 1999; transmitters from primary afferent neurons. Based on
Suzuki, R. et al., 2002). In these studies, ablation was these observations, we formulated the hypothesis that
performed by spinal injection of substance P conju- neuropathic pain is initially driven by enhanced pri-
gated to saporin. Internalization of the NK1 receptor mary afferent the ectopic discharge but is then
stimulated by substance P introduced the conjugate maintained in large part through descending facilita-
into the cell body, where saporin interfered with tion and upregulation of spinal dynorphin (Lai, J.
protein synthesis, causing eventual death of the cell et al., 2001; Porreca, F. et al., 2002; Ossipov, M. H.
(Honore, P. et al., 1999; Nichols, M. L. et al., 1999). and Porreca, F., 2006). Dynorphin upregulation
Electrophysiologic studies indicated that signs of occurs in the same postsynaptic lamina I cells that
spinal sensitization induced by intraplantar capsaicin also express FOS in response to sensitization (Ji, R.
or peripheral inflammation were abolished by selec-
R., 2004; Kawasaki, Y. et al., 2004). Elevated levels of
tive ablation of the NK1-expressing neurons of
spinal dynorphin enhance capsaicin-evoked release
lamina I (Khasabov, S. G. et al., 2002; 2005).
of CGRP and substance P from primary afferent
Additional electrophysiologic studies demonstrated
terminals and of PGE2, presumably from postsynap-
that SNL-induced enhancement of neuronal
tic cells, which further enhances the release of
responses to noxious thermal stimuli or light brush,
excitatory amino acids from afferent terminals
neuronal after-discharge, and enlargement of the
(Gardell, L. R. et al., 2003a; 2003b; Koetzner, L.
receptive field were abolished in rats with SNL and
et al., 2004). Sequestration of dynorphin with anti-
in which NK1-expressing spinal neurons were selec-
tively ablated with spinal injections of substance P serum or surgical and neurochemical manipulations
conjugated to saporin (Suzuki, R. et al., 2004a; 2004b; that blocked the upregulation of spinal dynorphin
Suzuki, R. and Dickenson, A. H., 2005a; 2005b). also abolished enhanced transmitter release and
Moreover, evoked expression of FOS in the RVM behavioral neuropathic pain (Burgess, S. E. et al.,
and in neurons in the deep laminae of the spinal cord 2002; Porreca, F. et al., 2002; Gardell, L. R. et al.,
was also abolished by selective ablation of NK1- 2003a; 2003b).
expressing neurons of lamina I (Suzuki, R. et al., It is well established that elevated levels of spinal
2002; Suzuki, R. and Dickenson, A. H., 2005a; dynorphin and its fragments have a prominent prono-
2005b). Based on these observations, especially the ciceptive role in chronic pain states (Lai, J. et al., 2001).
critical finding that the projection neurons of the The spinal injection of antiserum to dynorphin did not
superficial lamina of the spinal cord appear to pro- abolish behavioral signs of neuropathic pain in mice
mote sensitization of dorsal horn neurons in the deep with SNL when administered 2 days after the injury,
lamina through an indirect mechanism, led to the but was effective 10 days after SNL (Wang, Z. et al.,
hypothesis that injury-induced enhanced noxious 2001; Gardell, L. R. et al., 2004). Moreover, knockout
inputs are transmitted to supraspinal sites, leading mice that do not express dynorphin still develop
to the activation of a pronociceptive descending behavioral signs of neuropathic pain after SNL, but
pain facilitatory system that maintains the state of these signs resolve within 1 week (Wang, Z. et al., 2001;
spinal sensitization and maintains the neuropathic Gardell, L. R. et al., 2004). The dynorphin-dependent
pain state. maintenance phase is dependent on descending
846 Neuropathic Pain: Basic Mechanisms (Animal)

facilitation form the RVM. Surgical and pharmacolo- recently been suggested to be mediated through brady-
gic manipulations that disrupt descending facilitation kinin receptors (Lai, J. et al., 2006). Bradykinin B1 and
abolish both dynorphin upregulation and the mainte- B2 receptors are present on nerve terminals of primary
nance, but not the initiation, of behavioral signs of afferent fibers (Botticelli, L. J. et al., 1981; Cloutier, F.
neuropathic pain. Rats with SNL and either lesions et al., 2002), and are in close proximity to interneurons
of the DLF or selective ablation of facilitatory RVM that express dynorphin (Botticelli, L. J. et al., 1981).
neurons after microinjection of the dermorphin Moreover, the B1 and B2 receptors are upregulated
saporin conjugate did not show an upregulation of after SNL (Lai, J. et al., 2006). A pronociceptive role of
spinal dynorphin levels and the behavioral signs of these receptors is indicated since intrathecal adminis-
neuropathic pain resolved over the time course during tration of bradykinin produced behavioral signs of
which dynorphin upregulation would be expected hyperalgesia (Wang, H. B. et al., 2005). Behavioral
after SNL (Burgess, S. E. et al., 2002; Gardell, L. R. signs of enhanced pain after nerve injury were attenu-
et al., 2003a). Thus, in the absence of elevated spinal ated in B1 or B2 knockout mice (Ferreira, J. et al., 2002;
dynorphin, neuropathic pain is not maintained. Wang, H. B. et al., 2005). Curiously, although the bra-
Elevated levels of spinal dynorphin promote the dykinin receptors are present in the spinal cord, there is
release of transmitters from primary afferent term- little evidence of bradykinin synthesis within the spinal
inals, thereby enhancing nociceptive input. The time cord, suggesting the either bradykinin must diffuse
course and profile of the effect of spinal dynorphin on from the circulatory system to act at the receptors, or
capsaicin-evoked release of CGRP from primary there may be another endogenous ligand (Lai, J. et al.,
afferent terminals correlates closely with the beha- 2006). It was shown that dynorphin induced a transient
vioral studies. Addition of antiserum to dynorphin increase in calcium conductance in embryonic DRG
abolishes the enhanced capsaicin-evoked release of cells that was blocked by B2 antagonists (Lai, J. et al.,
CGRP 10 days after SNL, but not 2 days after SNL 2006). Dynorphin-induced calcium currents were also
(Burgess, S. E. et al., 2002; Gardell, L. R. et al., 2003a). blocked by B1 or B2 antagonists in transfected cells
Disruption of descending facilitation by microinjec- expressing these receptors (Lai, J. et al., 2006). In the
tion of dermorphinsaporin conjugate into the RVM same study, it was shown that dynorphin demonstrated
or by lesioning the DLF also abolishes SNL-induced specific binding to the B1 and B2 receptors (Lai, J. et al.,
enhancement of capsaicin-evoked release of CGRP 2006). Finally, antagonists of the B1 or B2 receptors
(Burgess, S. E. et al., 2002; Gardell, L. R. et al., 2003a). given intrathecally produced dose-dependent attenua-
The significance of this line of investigation requires tion of behavioral parameters indicative of neuropathic
additional examination, however. pain in rats with SNL, but only at the time points when
spinal dynorphin was elevated (Lai, J. et al., 2006). The
antagonists were without effect during the dynorphin-
55.7.1 Descending Facilitation Maintains a
independent early phase of nerve injury. Taken
Sensitized Spinal Cord: Upregulation of
together, these studies indicate that dynorphin may
Spinal Dynorphin and Enhanced Release of
mediate its pronociceptive enhancement of neuro-
Primary Afferent Transmitters
pathic pain through actions at the bradykinin
The mechanism through which dynorphin may exert a receptors. The activity of dynorphin at bradykinin
pronociceptive function in the spinal cord is not fully receptors could underlie the sensitization of dorsal
understood. It was once suggested that dynorphin may horn neurons as described above (Wang, H. B. et al.,
act at the glycine coagonist site to potentiate the effect 2005).
of glutamate at the NMDA receptor (Zhang, L. et al.,
1997). However, other studies provided evidence that
55.7.2 Descending Facilitation Maintains a
dynorphin could inhibit opening of ion channels
Sensitized Spinal Cord: Serotonergic
mediated by the NMDA receptor (Chen, L. and
Contributions
Huang, L. Y., 1998). Bakalkin and coworkers suggested
that dynorphin could promote the formation of pores More recent studies suggest that descending seroto-
through the cell membrane, allowing the leakage of nergic projections from the RVM may promote
sodium and calcium ions, thus sensitizing neurons in spinal sensitization through an action at the 5-HT3
this fashion, but this mechanism was not explicitly receptor. The 5-HT3 receptor facilitates release
demonstrated (Hugonin, L. et al., 2006). The mechanism or transmitter from nerve terminals through a
by which dynorphin may promote nociception has ligand-gated Ca2 channel (Miquel, M. C. et al.,
 Neuropathic Pain: Basic Mechanisms (Animal) 847

2002; Zeitz, K. P. et al., 2002). This receptor is pre- maintenance of the tonic descending facilitatory sys-
dominant in the superficial laminae of the spinal tem, thus perpetuating a chronic pain state. The
dorsal horn (Doucet, E. et al., 2000; Miquel, M. C. existence of a pronociceptive spinalsupraspinal
et al., 2002). Of this population, the major portion, loop that maintains a sensitized spinal cord may
approximately 85%, are found on primary afferent also provide a mechanism through which tactile
nerve terminals and the remainder at postsynaptic hyperesthesia might be mediated.
sites (Doucet, E. et al., 2000; Miquel, M. C. et al.,
2002). Electrophysiologic studies indicated that pri-
mary afferent nerves that express the 5-HT3 55.8 Summary
receptor on their terminals include C fiber nociceptors
and myelinated A-delta nociceptors sensitive to Tremendous progress in our understanding of
mechanical stimuli (Zeitz, K. P. et al., 2002). This mechanisms that initiate and maintain neuropathic
distribution is consistent with the modulation of pain has been made over the past two decades.
mechanical-evoked nociceptive signals, and the 5- Importantly, such increased understanding has led
HT3 receptor has been shown to facilitate nociceptive to programs aimed at the development of novel ther-
transmission at the level of the spinal cord (Zeitz, K. P. apeutics for neuropathic pain. It has been well
et al., 2002). Double-labeling studies in the nucleus appreciated that peripheral nerve injury produces
raphe magnus (NRM) detected a population of sero- increased activity of the injured and adjacent unin-
tonergic neurons that express FOS in response to jured peripheral nerve fibers. This enhanced activity,
noxious stimulation, suggesting the existence of a pro- or peripheral sensitization, is maintained in part by
nociceptive serotonergic projection from the RVM increased or relocation of ion channels that promote
(Suzuki, R. et al., 2002). Moreover, it was found that axonal firing. Programs are in place in the pharma-
serotonergic terminals from the RVM are juxtaposed ceutical industry to develop subtype selective
with cell bodies in the intermediate laminae of the sodium channel blockers that may diminish enhance
dorsal horn, some of which also express the NK1 activities of injured or adjacent nerves. Increase in
receptor (Todd, A. J. et al., 2000; Todd, A. J., 2002). pronociceptive mediators, including cytokines, che-
Antagonists of the 5-HT3 receptor or depletion of mokines, and growth factors have been noted in
serotonin from the spinal cord abolished the enhanced multiple experimental models of neuropathic pain.
responses of dorsal horn neurons to mechanical stimu- The enhanced primary afferent activity leads to a
lation in rats with SNL (Rahman, W. et al., 2006). The state of sensitization of the spinal cord neurons,
5-HT3 antagonist ondansetron abolished behavioral such that they are more responsive to sensory inputs.
signs of neuropathic pain and electrophysiologic para- Approaches aimed at decreasing sensitization of
meters indicative of sensitization of superficial and nociceptors through anti-NGF strategies, or through
deep dorsal horn neurons in rats with SNL (Suzuki, the administration of exogenous growth factors of the
R. et al., 2004a; 2004b). The effects of ondansetron GDNF family, are either in development or under
were blocked by disrupting enhanced RVM activity consideration by the pharmaceutical industry.
through the ablation of NK1-expressing dorsal horn Enhanced activity of sensitized nociceptors can
neurons with substance Psaporin conjugate, indicat- increase ascending sensory inputs to supraspinal cen-
ing that the 5-HT3 receptors were activated by a ters and neuroplastic changes that help maintain a
descending serotonergic pain facilitatory system from sensitized state. One prominent change is increased
the RVM (Suzuki, R.et al., 2004a; 2004b). availability of CCK in the RVM and enhanced des-
Collectively, these observations paint a picture cending facilitation of pain from this region. The
suggesting that neuropathic pain states are main- enhanced pain facilitation leads to further changes
tained, at least in part, by increased sensitivity of in the spinal cord, notably upregulation of spinal
primary afferent neurons to noxious stimuli. In addi- dynorphin to pathological levels. Dynorphin, along
tion, it is hypothesized that persistent nociceptive with pronociceptive transmitters such as PGEs and
inputs elicits supraspinal neuroplastic changes that glutamate, enhance the release of excitatory trans-
results in the activation of a tonic descending facil- mitters from primary afferent terminals. The result is
itation of nociceptive inputs, which is mediated by an a net increase in nociceptive inputs that help perpe-
upregulation of spinal dynorphin content. Thus, tuate this pronociceptive spinal/supraspinal/spinal
increased spinal dynorphin serves to promote exci- pain facilitatory loop. Such knowledge underlies
tatory transmitter release, which in turn promotes the efforts to development of N-type calcium channel
848 Neuropathic Pain: Basic Mechanisms (Animal)

blockers. Understanding the changes that occur at derived neurotrophic factor family receptor components are
differentially regulated within sensory neurons after nerve
each of these nodes in this cycle could lead to injury. J. Neurosci. 20, 427437.
means through which this self-perpetuating chain of Bennett, D. L., Koltzenburg, M., Priestley, J. V., Shelton, D. L.,
events may be broken, thus abolishing the mechan- and McMahon, S. B. 1998. Endogenous nerve growth factor
regulates the sensitivity of nociceptors in the adult rat. Eur. J.
isms that perpetuate chronic pain. The accelerating Neurosci. 10, 12821291.
understanding of pain regulatory systems advanced Botticelli, L. J., Cox, B. M., and Goldstein, A. 1981.
by the discovery of underlying mechanisms driving Immunoreactive dynorphin in mammalian spinal cord and
dorsal root ganglia. Proc. Natl. Acad. Sci. U. S. A.
enhanced abnormal pain will be continuously vali- 78, 77837786.
dated by clinical trials of drugs with novel Boucher, T. J. and McMahon, S. B. 2001. Neurotrophic factors
mechanisms of action. Critically, new mechanisms and neuropathic pain. Curr. Opin. Pharmacol. 1, 6672.
Boucher, T. J., Okuse, K., Bennett, D. L., Munson, J. B.,
of action will be tested clinically with the expectation Wood, J. N., and McMahon, S. B. 2000. Potent analgesic
that improved therapy can be provided to patients. effects of GDNF in neuropathic pain states. Science
290, 124127.
Bouhassira, D., Attal, N., Alchaar, H., Boureau, F., Brochet, B.,
Bruxelle, J., Cunin, G., Fermanian, J., Ginies, P., Grun-
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 56 Animal Models and Neuropathic Pain
I Decosterd and T Berta, University of Lausanne, Lausanne, Switzerland
2009 Elsevier Inc. All rights reserved.

56.1 Introduction 857

56.2 Animal Models of Neuropathic Pain 857
56.2.1 Models of Central Neuropathic Pain: Spinal Cord Injuries 857
56.2.2 Models of Peripheral Neuropathic Pain: Injury to Roots, Dorsal Root Ganglia,
and Spinal Nerves 859
56.2.3 Models of Peripheral Neuropathic Pain: Injury to Peripheral Nerves 859
56.2.4 Models of Peripheral Neuropathic Pain: Distal Nerve Branch Injuries 860
56.2.5 Nontraumatic Models of Neuropathic Pain 861
56.3 Behavioral Assessment of Neuropathic Pain in Animal Models: Spontaneous Pain,
Stimulus-Evoked Pain, and Stimulus-Induced Pain 861
56.4 Conclusions 862
References 862

Glossary
allodynia Pain due to a stimulus which does not neuropathy A disturbance of function or patholo-
normally provoke pain. gical change in a nerve.
autotomy Self amputation by animal that severs spontaneous pain Pain without an evident
its own appendages (usually distal phalanges). stimulus.
hyperalgesia An increased response to a stimulus stimulus-evoked pain pain evoked by a
which is normally painful. mechanical/thermal/chemical stimulus.
neuropathic pain Pain initiated or caused by a Wallerian degeneration The degeneration of a
primary lesion or dysfunction in the nervous nerve fiber induces by injury or disease, character-
system. ized by segmentation of the myelin and resulting in
atrophy and destruction of the axon.

56.1 Introduction the mechanisms involved. Therefore, it is important

to select the best model for each specific research
Animal models in pain research offer great promise interest or a combination of appropriate but distinct
for both the identification of pain mechanisms, and models. In this chapter we will argue for the most
the investigation of possible therapeutic applications. frequently used animal models of NP, compare their
Our understanding of neuropathic pain (NP) has advantages and disadvantages, and summarize the
progressed during last few decades and although behavioral characterization.
animal models have provided important information,
in humans a clear pattern of the mechanisms respon-
sible for its generation and maintenance is still 56.2 Animal Models of Neuropathic
unclear. Therefore, it is essential to put forward Pain
advanced animal models based, on the investigation
of specific mechanisms, as well as refining the way in At present, animal models of NP cover various etiol-
which the data is interpreted. Moreover, it is clear ogies and are related to symptoms leading to an
that the quest of a unique and universal animal model extensive picture of clinical NP manifestations. The
of all symptoms in patients ignores the complexity of majority of animal models of NP involve traumatic

857
858 Animal Models and Neuropathic Pain

injuries to peripheral nerves, nerve roots, or spinal of injury involves functional and structural changes,
cord by transection, ligation, or compression partial loss of normal descending pathway, and hyper-
(Figure 1). Other experimental models are related excitability of dorsal horn neurons (Woolf, C. J., 1983;
to direct or indirect nerve inflammation, ischemia, Woolf, C. J. and Fitzgerald, M., 1983; Hulsebosch, C. E.,
drug toxicity, or systemic metabolic disorders. Each 2002). Spinal cord transection, contusion, compression,
animal model has been characterized by precise and chemical toxicity induce all at-level or below-level
behavioral-based evaluations using different modal- signs of pain hypersensitivity (Vierck, C. J. et al., 1999;
ities of sensory stimulus. Many temporal and spatial Sjolund, B. H., 2002; Rosenzweig, E. S. and McDonald,
profiles of molecular, physiological, and structural J. W., 2004). In the hemisection model (Christensen, M.
changes occurring during the development of NP D. et al., 1996), a lateral spinal hemisection is performed
have been investigated and most of the rodent models at T13 level. After a period of motor dysfunction,
are compatible with actual electrophysiological mea- ipsilateral to the injury, rats display persistent stimu-
surements, functional imaging techniques, as well as lus-evoked hypersensitivity of the four limbs
genomic and proteomic screenings. associated with vocalization. This model was success-
fully used to observe molecular and structural
reorganization of the spinal cord in NP (Christensen,
56.2.1 Models of Central Neuropathic Pain: M. D. and Hulsebosch, C. E., 1997b; Bennett, A. D. et al.,
Spinal Cord Injuries 2000; Hains, B. C. et al., 2003b). The anterolateral
Partial or complete spinal cord injuries result in loss of cordotomy model (Vierck, C. J. and Light, A. R.,
function in varying degrees below the level of injury, 1999) consists in an unilateral injury of the anterolat-
and are often associated with central NP pain syn- eral column at the T9T11 spinal segments. The
dromes (Christensen, M. D. and Hulsebosch, C. E., injury is associated with overgrooming and signs of
1997a; Siddall, P. J. and Loeser, J. D., 2001). This type allodynia/hyperalgesia. The model has been adapted

Spinal cord injury

Transection toxicity
Dorsal root
ganglia Spinal Contusion
cord
L2
Dorsal root injury
L3 Rhizotomy

L4 Ventral root injury

L5 VRT

DRG injury
L6
Femoral CCD
nerve
Spinal nerve injury

SNL
Sciatic
nerve Nerve trunk injury

Saphenous Axotomy PSL

nerve
CCI PST
Common Sural Nerve branch injury
peroneal Tibial nerve
nerve nerve SNI SPL

Figure 1 Anatomy of the rat lumbar plexus and the location of the different injuries used to create the different neuropathic
pain models. VRT, ventral root transection; CCD, chronic compression of dorsal root ganglia; SNL, spinal nerve ligation; CCI,
chronic constriction injury; PSL, partial sciatic ligation; PST, partial sciatic nerve transection; SNI, spared nerve injury (tibial
and c. peroneal nerves are injured, sural nerve is intact); SPL, saphenous nerve partial ligation (3050% of the saphenous is
transected).
 Animal Models and Neuropathic Pain 859

to primates and much information has been acquired sensory and motor fibers and a clear segmental loca-
on thalamus circuitry in NP (Weng, H.-R. et al., 2000). tion of injured and noninjured primary afferents.
The model of spinal cord contusion (Gruner, J. A., Spontaneous and stimulus-evoked pain are sustained
1992; Siddall, P. et al., 1995) is produced by an impact by different mechanisms such as deafferentation,
injury on the spinal cord at the level of T12/L1 using a ectopic activities in dorsal root ganglion (DRG),
weight dropped with a standard impact device and spinal cord neurons, Wallerian degeneration of
(MASCIS, Multi-Center Animal Spinal Cord Injury the peripheral axons and central reorganization
Study). The incidence of pain hypersensitivity is cor- (Eschenfelder, S. et al., 2000; Li, Y. et al., 2000). For
related to the extent of the injury (Hulsebosch, C. E. dorsal and ventral root rhizotomies, typically at L5
et al., 2000; Mills, C. D. et al., 2001; Gwak, Y. S. et al., level, the root is ligated and transected 34 mm prox-
2004) and evidence of hyperexcitability of dorsal horn imal to the DRG (Colburn, R. W. et al., 1999; Li, L.
neurons and regulation of voltage-gated sodium chan- et al., 2002). The ventral root transection model pro-
nels have been shown (Hains, B. C. et al., 2003a). vides an additional feature to investigate NP, in the
Squeezing the spinal cord is the basis of the compres- sense that lesion of a motor nerve induces alteration
sion injury model (Fehlings, M. G. and Nashmi, R., of pain sensitivity (Li, L. et al., 2002; Sheth, R. N. et al.,
1995). A 35- or 50-g clip compression is placed on the 2002). Robust mechanical and cold allodynia-like
dura mater at the juncture of T12 and T13 spinal behaviors have been documented (Sheen, K. and
segments and leads to microvasculature disruption, Chung, J. M., 1993), whereas loose ligation of
ischemia, and progressive axonal injury associated L4L6 dorsal roots produces only mechanical allo-
with the development of tactile allodynia (Bruce, J. C. dynia (Tabo, E. et al., 1999). The chronic compression
et al., 2002). Other central models of NP use toxic of dorsal root ganglia (CCD), proposed as a model of
substances. The photochemically induced lesion of lower back pain/disc herniation, consists of the pla-
the spinal cord (Hao, J. X. et al., 1991) is produced by cement of a fine stainless-steel rod into the L5/L4
the intravenous injection of erythrosine B (a photosen- intervertebral foramen (Hu, S. J. and Xing, J. L.,
sitizing dye) immediately followed by irradiation with 1998). Heat hyperalgesia and a tactile allodynia are
an argon laser beam at T13 level. This procedure the resulting behaviors encountered in this model
results in an ischemic lesion of thoracic spinal cord (Zhang, J. M. et al., 1999; Ma, C. and LaMotte, R.
by occlusion of spinal cord vessels and produces a H., 2005). The widely used spinal nerve ligation
particularly severe mechanical and cold allodynia- (SNL), where L5/L6 spinal nerve are tightly ligated
like behavior in skin territories innervated by the ros- (Kim, S. H. and Chung, J. M., 1992), produces robust
tral border of the injury (Hao, J. X. et al., 1991; 1992). and consistent sympathetic-dependent NP-related
Excitotoxic spinal cord injury (ESCI) is directly behaviors including indirect signs of spontaneous
mediated by the intraspinal injection at levels ranging pain, heat hyperalgesia, mechanical allodynia/hyper-
from T10 to L4 of quisqualic acid (QUIS), an - algesia, and cold allodynia (Choi, Y. et al., 1994). This
amino-3-hydroxy-5-methylisoxazole-4-propionic acid type of damage has also been applied successfully to
(AMPA) metabotropic receptor agonist (Yezierski, R. mice and primates (Choi, Y. et al., 1994; Ali, Z. et al.,
P. et al., 1993; 1998). This model offers the considerable 1999; Mogil, J. S. et al., 1999). The additional and
advantage of avoiding surgical laminectomy, and considerable advantage of SNL is the possibility to
although less representative of a clinical pathology, it distinguish injured (L5/L6) and noninjured (L4)
induces sustained NP-like symptoms related to a pre- DRG and spinal segments.
cise mechanism. The QUIS model leads to a gradual
and specific loss of neurons associated to NP-like
56.2.3 Models of Peripheral Neuropathic
behaviors and the extension of the injury and the
Pain: Injury to Peripheral Nerves
intensity of changes are dependent on the depth and
volume of the QUIS injections. The first models of NP used complete transection of
major peripheral nerve. Wall and co-workers produce
an entire denervation of the distal hindlimb, damaging
56.2.2 Models of Peripheral Neuropathic
the sciatic and saphenous nerves, to model anesthesia
Pain: Injury to Roots, Dorsal Root Ganglia,
dolorosa, a type of NP referred to denervated zone
and Spinal Nerves
without any sensory input from that area. Nerves
Surgical procedures on selective root, dorsal root were transected at midthigh level and the proximal
ganglia, and spinal nerves allows direct access to stump was either tightly ligated or encapsulated into a
860 Animal Models and Neuropathic Pain

polyethylene tube (Wall, P. D. et al., 1979a; 1979b). grooming of the face, mechanical allodynia, and
The total sciatic transection prevents stimulus-evoked thermal hyperalgesia (Imamura, Y. et al., 1997).
assessment of the hindlimb but self-mutilation (autot- Sensory assessment of the animal face is difficult;
omy) was the major feature of the experimental however, the recent introduction of an automated
anesthesia dolorosa mode. Degrees of autotomy have thermal behavioral assay should provide valuable
been defined according to the extension of self-muti- characterizations of orofacial pain models (Neubert,
lation in the hindpaw and are used to evaluate the J. K. et al., 2005).
progression of the neuropathy. The subsequent
chronic constriction injury (CCI) model, four catgut
56.2.4 Models of Peripheral Neuropathic
ligations loosely placed around the sciatic nerve Pain: Distal Nerve Branch Injuries
(Bennett, G. J. and Xie, Y. K., 1988) or the partial
sciatic ligation (PSL) model, ligation of one-third to Injuries to distal nerves branches offer the advantage
one-half of the sciatic nerve in rats (Seltzer, Z. et al., that they can be anatomically defined, helping the
1990) or mice (Malmberg, A. B. and Basbaum, A. I., standardization of surgical procedures. The spared
1998), allow sensory testing in the hindpaw (Figure 2). nerve injury (SNI) in rats and mice is based on the
Moderate autotomy, guarding, and excessive groom- ligation and transection of the tibial and common
ing of the injured limb were noticed as well as peroneal nerves leaving the sural nerve intact
thermal/mechanical hyperalgesia- and allodynia-like (Decosterd, I. and Woolf, C. J., 2000; Bourquin, A. F.
behavior were recorded (Bennett, G. J., 1993). et al., 2006). This model results in early, robust, and
Alternative to the CCI models include application of intense mechanical allodynia/hyperalgesia and cold
fixed-diameter polyethylene cuffs around the sciatic allodynia in the territory of the sural nerve
nerve, favoring a standardization of the loose, but (Figure 3). It offers the advantage of sparing an
constrictive ligatures in CCI (Mosconi, T. and almost sensory nerve in combination with a clear
Kruger, L., 1996). Another valuable alternative is the separation of injured and noninjured axons distal to
partial sciatic nerve transection (PST) model where the injury. SNI has also been adapted to neonatal
animals (Howard, R. F. et al., 2005) and variants of
half of the diameter of the sciatic nerve is only trans-
the model have been described in rats and mice (Lee,
ected, avoiding inflammation caused by suture
B. H. et al., 2000; Shields, S. D. et al., 2003). Recently,
material with PSL. Chronic constriction of the tri-
the saphenous nerve partial ligation (SPL) model
geminal nerve (infraorbital nerve CCI, IONCCI)
(3050% of the saphenous nerve, a branch of the
was designed by Vos B. P. et al. (1994) as a model of
femoral nerve that innervates the medial part of the
trigeminal pain. IONCCI model elicits excessive
paw, is tightly ligated) leads to the development of NP-
related symptoms (Walczak, J. S. et al., 2005). The
relevance in this model is that the injury of an exclu-
sively sensory nerve is easily transferable for
investigations with skinnerve preparations (Reeh, P.
W., 1986). Injury to peripheral branches has also been

Dorsal Plantar

Sural Lateral Medial Lateral

Common peroneal
Saphenous
Tibial
Overlaping territories

Figure 2 In many experimental models of neuropathic

pain (NP), stimulus-evoked pain sensitivity is assessed in Figure 3 Areas of the dorsal and plantar paw innervated
the plantar side of the hindpaw with various stimuli, usually by the sciatic and femoral nerves in the rat. Some overlap
in the territory of spared or regenerated afferents. between territories may occur (pink).
 Animal Models and Neuropathic Pain 861

applied to the trigeminal nerve to study orofacial pain nerve injury), plus the localization, the intensity,
and similar to the SNI model, transection of a branch of and the time course of symptoms. NP in humans is
the trigeminal nerve (infra-alveolar nerve) leads to characterized by modification of basal pain sensitiv-
hypersensitivity in the whisker pad area, which is ity with ongoing/spontaneous pain, and stimulus-
innervated by a spared nerve (ION) (Tsuboi, Y. et al., evoked hyper- or hyposensitivity (Woolf, C. J. and
2004). Decosterd, I., 1999). Spontaneous pain appears inde-
pendently of any external stimulus and cannot be
assessed directly in animals. Only indirect physiolo-
56.2.5 Nontraumatic Models
gical or behavioral surrogates of spontaneous pain are
of Neuropathic Pain
measurable: spontaneous discharges in sensory neu-
Vincristine, taxol, and cisplatin may lead to periph- rons, autotomy, posture (e.g., guarding behavior)
eral neuropathy and NP in cancer patients. Models movements (e.g., brisk paw flinches), or changes in
based on these substances have been created with general behavior (e.g., grooming, sleep patterns,
different features of NP-related symptoms, usually vocalization, exploratory behavior) (Mogil, J. S. and
in a dose-dependent manner (de Koning, P. et al., Crager, S. E., 2004). All raised substantial issues that
1987; Cavaletti, G. et al., 1995; Aley, K. O. et al., have meant laboratories favoring the recording of
1996; Authier, N. et al., 2000; Mimura, Y. et al., 2000; stimulus-evoked pain.
Nozaki-Taguchi, N. et al., 2001). Diabetes is a major Stimulus-evoked pain is elicited by distinct sti-
cause of painful neuropathy in humans. The diabetic muli like touch, pinprick, or temperature (heat or
model was induced by intraperitoneal injections of cold). Change in the threshold of response and/or
streptozotocin (STZ), which leads to pancreatic islet quality of response are generally used as outcomes of
destruction and chronic hyperglycemia (Courteix, C. allodynia/hyperalgesia-like behavior.
et al., 1993; 1994). NP symptoms such as hyperalge- Mechanical allodynia-like behavior is usually
sia- and allodynia-like behavior appear 2 weeks after. measured by using a series of calibrated monofila-
STZ produces functional, biochemical, and struc- ments of logarithmic incremental force (von Frey
tural abnormalities in the sciatic nerve similar to hairs) (Bennett, G. J., 2001). Monofilaments are
those seen in human diabetic neuropathy (Sima, A. A. applied in various territories of the animals body,
and Sugimoto, K., 1999). Prolonged ischemia of the and positive responses are recorded when the appli-
hindpaw was proposed as a surrogate to complex cation induces a withdrawal, movement, or
regional pain syndrome (CRPS) type I (Coderre, T. J. vocalization. The evaluation of nociceptive threshold
et al., 2004). After 8 h and lasting for 4 weeks, mechan- in sensible regions is the result of different proce-
ical hyperalgesia and bilateral mechanical/thermal dures, such as the updown method of Dixon
allodynia was observed along with excessive shaking (Chaplan, S. R. et al., 1994). Cold allodynia-like beha-
and licking of hindpaws (Coderre, T. J. et al., 2004). vior is assessed by placing an acetone drop on the
Axonal damage is combined with inflammation and area of interest, the acetone drop quickly evaporates
nerve inflammatory models, without direct nerve eliciting a cold sensation (Choi, Y. et al., 1994).
injury were developed by wrapping the sciatic Frequency of response to multiple applications as
nerve with cellulose-containing inflammatory well as the type and duration of response have been
reagents such as carrageenan or complete Freunds used to score cold allodynia. Another method for
adjuvant (CFA) (Eliav, E. et al., 1999). In these cases, measuring response to cold is the use of a cooled
the course of pain hypersensitivity is very short (25 metal floor, usually at noxious low temperatures,
days only). thus evaluating cold hyperalgesia (Bennett, G. J.
and Xie, Y. K., 1988; Choi, Y. et al., 1994). Thermal
hyperalgesia-like behavior is typically evaluated by
56.3 Behavioral Assessment of measuring the latency and duration of reaction
Neuropathic Pain in Animal Models: toward a heat noxious stimulus induced by a radial
Spontaneous Pain, Stimulus-Evoked infrared beam (Hargreaves, K. et al., 1988).
Pain, and Stimulus-Induced Pain Mechanical hyperalgesia-like behavior is evoked by
a pinprick test, using a safety pin stimulus that pro-
Tests based on pain sensitivity consider the type of vokes, in a normal animal, a small brisk withdrawal
injury, the remaining capacity of withdrawal reflexes reflex, while abnormal responses are evoked in
(e.g., when neuromuscular defect is caused by the experimental NP rats. A large duration of withdrawal
862 Animal Models and Neuropathic Pain

and paw guarding/shaking/licking follows the stimu- pain mechanisms, but they may reflect them.
lus (Tal, M. and Bennett, G. J., 1994). Similarly, all outcome measures in experimental
Stimulus-induced pain refers to the capacity of models are not equivalent. A sophisticated approach
external stimuli not only to directly evoke pain, but using multiple models (Decosterd, I. et al., 2004) is
also to alter sensory processing such as to generate a then required in order to correlate information
state of pain hypersensitivity that outlast the primary obtained in experimental models with clinical syn-
stimuli. In animals, stimulus-evoked pain after cap- dromes of NP. Although all models of NP have weak
saicin, inflammation, or nerve crush (Bennett, G. J. points in modeling only partially human NP and the
and Xie, Y. K., 1988; Choi, Y. et al., 1994; Ma, Q. P. impossibility to have direct behavioral measurement
and Woolf, C. J., 1996; Kim, H. T. et al., 2001; of spontaneous pain, we believe that current and
Decosterd, I. et al., 2002) has been investigated after future models of NP will strengthen our knowledge
repeated low-threshold mechanical stimuli and of the underlying mechanisms and favor the devel-
further measurement of mechanical sensitivity is car- opment of new treatment strategies for this currently
ried out using von Frey hairs. In this case, stimulus- intractable form of pain.
induced pain has therefore been termed progressive
tactile hypersensitivity.
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Hao, J. X., Xu, X. J., Aldskogius, H., Seiger, A., and Wiesenfeld- Mogil, J. S., Wilson, S. G., Bon, K., Lee, S. E., Chung, K.,
Hallin, Z. 1991. Allodynia-like effects in rat after ischaemic Raber, P., Pieper, J. O., Hain, H. S., Belknap, J. K.,
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irradiation. Pain 45, 175185. Heritability of nociception I: responses of 11 inbred mouse
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Neubert, J. K., Widmer, C. G., Malphurs, W., Rossi, H. L., Vierck, C. J. and Light, A. R. 1999. Effects of combined
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 57 Neuropathic Pain: Clinical
R Baron, Christian-Albrechts-Universitat Kiel, Kiel, Germany
2009 Elsevier Inc. All rights reserved.

57.1 Definition of Neuropathic Pain 866

57.2 Epidemiology of Neuropathic Pain 866
57.3 Classification 866
57.3.1 Disease/Anatomy-Based Classification 866
57.3.1.1 Painful peripheral (focal, multifocal, generalized) neuropathies 867
57.3.1.2 Central pain syndromes 871
57.3.1.3 Complex painful neuropathic disorders 872
57.3.1.4 Mixed pain syndromes 873
57.3.2 Mechanism-Based Classification 873
57.4 Signs and Symptoms in Neuropathic Pain 874
57.5 Pathophysiological Mechanisms in Neuropathic Pain 876
57.5.1 Peripheral Sensitization of Primary Afferent Nociceptors 876
57.5.2 Central Sensitization 878
57.5.3 Central Disinhibition and Fascilitation 879
57.5.4 Inflammation and Neuropathic Pain 880
57.6 Pathophysiological Mechanisms in Patients in Relation to Their Somatosensory Profile 880
57.6.1 Peripheral Sensitization of Primary Afferent Neurons in Patients 880
57.6.2 Sensitization to Catecholamines in Patients 882
57.6.3 Central Sensitization in Patients 884
57.6.4 Central Disinhibition Leading to Cold Hyperalgesia 885
57.6.5 Deafferentation: Hyperactivity of Central Pain Transmission Neurons 885
57.6.6 Inflammation in Patients 886
57.7 Diagnostic Tools for Neuropathic Pain 886
57.7.1 Questionnaires 887
57.7.2 Bedside Assessment of Neuropathic Pain 887
57.7.3 Apparative Assessment of Somatosensory Phenotypes 889
57.7.4 Skin Biopsies 889
57.7.5 Microneurography 891
57.7.6 Indirect Test of Afferent Unmyelinated Fiber Function 891
57.7.7 Imaging Techniques 892
57.7.8 Do We Have Diagnostic Tool to Dissect Individual Mechanisms in Neuropathic Pain? 892
57.8 Therapy 893
57.8.1 Antidepressants 894
57.8.2 Anticonvulsants (Ca Channel Modulators) 894
57.8.3 Anticonvulsants (Na Channel Blockers) 895
57.8.4 Tramadol and Opioid Analgesics 895
57.8.5 Topical Medications 895
57.8.5.1 Topical capsaicin 895
57.8.5.2 Topical lidocaine 895
57.8.6 N-Methyl-D-Aspartic Acid Receptor Antagonists 895
57.8.7 Cannabinoids 895
57.8.8 Treatment Guideline 896
References 897

865
866 Neuropathic Pain: Clinical

Glossary
CRPS Complex regional pain syndrome. PET Positron emission tomography.
DRG Dorsal root ganglion, spinal ganglion. TRP Ion channels of transient receptor potential
MEG Magnetic encephalography. family.
NCF Nucleus cuneiformis. TNF Tumor necrosis factor.
PAG Periaqueductal gray. QST Quantitative sensory testing.

57.1 Definition of Neuropathic Pain nociceptive and neuropathic pain types who were
referred to pain specialists in Germany
The current NeuPSIG (Neuropathic pain special (Freynhagen, R. et al., 2005/2006) revealed that
interest group of the International Association for the already 13% of these patients suffer from the two
Study of Pain) taxonomy of chronic pain, defines classical neuropathic disorders, postherpetic neural-
neuropathic pain as a Pain arising as a direct conse- gia (PHN) and painful diabetic neuropathy and 40%
quence of a lesion or disease affecting the of all patients at least have a neuropathic component
somatosensory system. The term disease refers to to their discomfort (especially patients with chronic
identifiable disease processes such as inflammatory, back pain and radiculopathy). In neuropathic patients
autoimmune conditions, or channelopathies, while pain therapy was already performed for 24 years,
lesions refer to macro- or microscopically identifiable one-third of the patients mentioned three or more
damage. The restriction to the somatosensory system doctor visits for pain therapy in the past month and
is necessary, because diseases and lesions of other parts 820% already applied for pension (Table 1).
of the nervous system may cause nociceptive pain. For Comorbidities such as poor sleep, depressed mood,
example, lesions or diseases of the motor system may and anxiety are common in neuropathic pain and have
lead to spasticity or rigidity, and thus may indirectly a significant impact on the global pain experience. The
cause muscle pain. The latter pain conditions are now German survey revealed that about 60% of neuro-
explicitly excluded from the condition neuropathic pathic patients were moderate or severely depressed
pain. Where possible, neuropathic pain should be qua- compared with 25% in the nociceptive patient group.
lified as being of peripheral or central origin in terms Optimal sleep quality was only scored by 27% of the
of the location of the lesion or disease process. This neuropathic patients and in 49% of the nociceptive
distinction is important, as lesions or diseases of the pain patients, respectively.
central nervous system (CNS) and peripheral nervous
system (PNS) are distinct in terms of clinical manifes-
tations and underlying pathophysiology. 57.3 Classification
57.3.1 Disease/Anatomy-Based
Classification
57.2 Epidemiology of Neuropathic It is common clinical practice to classify neuropathic
Pain pain according to the underlying etiology of the
disorder and the anatomical location of the specific
Chronic neuropathic pain is common in clinical lesion (Jensen, T. S. and Baron, R., 2003). The major-
practice, greatly impairs the quality of life of patients ity of patients fall into four broad classes (Table 2):
and is a major economical health problem. Estimates painful peripheral neuropathies (focal, multifocal, or
of point prevalence for neuropathic pain in the gen- generalized, e.g., traumatic, ischemic, inflammatory,
eral population are as high as 5% (Daousi, C. et al., toxic, metabolic, and hereditary), central pain syn-
2004), a quarter of them suffering from severe inten- dromes (e.g., stroke, multiple sclerosis (MS), and
sity (McDermott, A. M. et al., 2006). Moreover, a spinal cord injury (SCI)), complex painful neuro-
recent prospective cross-sectional survey in more pathic disorders (complex regional pain syndromes,
than 12 000 chronic pain patients with both CRPSs) and mixed pain syndromes (combination of
 Neuropathic Pain: Clinical 867

Table 1 Sociodemographic and clinical characteristics in patients with postherpetic neuralgia (PHN) and diabetic painful
neuropathy (DPN) (%)

PHN DPN

Number of patients 498 1144

Male 199 (40.0) 596 (52.1)
Female 299 (60.0) 548 (47.9)
Age (years)
Mean 60.6 61.7
SD 15.4 13.2
Duration of pain treatment (months)
Mean 24 52
SD 41 63
No. of doctor visits for pain during the past 4 weeks
One visit 188 (37.8) 348 (30.4)
Two visits 135 (27.1) 339 (29.6)
Three or more 132 (26.5) 348 (30.4)
Ongoing psychotherapy 8 (1.6) 34 (3.0)
Intended/submitted pension application 42 (8.2) 221 (19.3)
Pain intensity on VAS during the past 4 weeks
Mild 154 (30.9) 492 (43.0)
Moderate 239 (48.0) 424 (37.1)
Severe 105 (21.1) 228 (19.9)
VAS worst pain (mean) 7.4 6.3
VAS average pain (mean) 5.5 4.9

VAS, visual analogue scale.

nociceptive and neuropathic pain, e.g., chronic low 57.3.1.1 Painful peripheral (focal,
back pain with radiculopathy). multifocal, generalized) neuropathies
The anatomical distribution pattern of the Although most of the different neuropathic pain syn-
affected nerves provides valuable differential diag- dromes are common and well known to the pain
nostic clues as to possible underlying causes. specialists, there are certain entity-specific features
Therefore, painful peripheral neuropathies are that should be mentioned in detail (See also Table 3):
grouped into symmetrical generalized polyneuropa-
thies, disease affecting many nerves simultaneously
and into asymmetrical neuropathies with a focal or 57.3.1.1.(i) Diabetic neuropathy
multifocal distribution or processes affecting the bra-
chial or lumbosacral plexuses. High prevalence

One important subgroup of painful polyneuropa- Several types of polyneuropathies, most preva-
lent symmetrical distally, sensory predominant
thies is characterized by a predominant or in some neuropathy
cases even isolated affection of small afferent fibers,
i.e., unmyelinated C fibers and small myelinated A
Numbness and paresthesias (sometimes burn-
ing quality) are common
fibers. In many cases, autonomic efferent small fiber
systems are also affected. Several different etiologies
Sometimes spontaneous, deep, aching pain, and
lightning pain
may lead to small-fiber polyneuropathies (see
below), up to 20% of cases, however, are of unknown
Severe sensory neuropathy in diabetes may
lead to painless perforating foot ulcers often asso-
cause. It is important to realize that conventional ciated with autonomic neuropathy
electrophysiological techniques like nerve conduc-
tion study (NCS), sensory-evoked potential (SEP),
Patients without pain tend to have areflexia
with distal sensory loss particularly involving joint
etc., only assess the function of myelinated peripheral position sense
axonal systems, the affection of small fibers are
missed. Therefore, especially in small-fiber neuropa-
Patients with severe burning pain and hyper-
esthesia tend to have sensory loss with a relative
thies alternative diagnostic procedures have to be preservation of position sense and intact reflexes
used (see below). and often have accompanying autonomic neuropathy
868 Neuropathic Pain: Clinical

Table 2 Disease-/anatomy-based classification of painful peripheral neuropathies

Painful peripheral neuropathies

Focal, multifocal
Phantom pain, stump pain, nerve transection pain (partial or complete)
Neuroma (posttraumatic or postoperative)
Posttraumatic neuralgia
Entrapment syndromes
Mastectomy
Postthoracotomy
Mortons neuralgia
Painful scars
Herpes zoster and postherpetic neuralgia
Diabetic mononeuropathy, diabetic amyotrophy
Ischemic neuropathy
Borreliosis
Connective tissue disease (vasculitis)
Neuralgic amyotrophy
Peripheral nerve tumors
Radiation plexopathy
Plexus neuritis (idiopathic or hereditary)
Trigeminal or glossopharyngeal neuralgia
Vascular compression syndromes
Generalized (polyneuropathies)
Metabolic or nutritional
Diabetic, often Burning feet syndrome
Alcoholic
Amyloid
Hypothyroidism
Beri beri, pellagra
Drugs
Antiretrovirals, cisplatin, disulfiram, ethambutol, isoniazid, nitrofurantoin, thalidomid, thiouracil, vincristine,
chloramphenicol, metronidazole, taxoids, gold
Toxins
Acrylamide, arsenic, clioquinol, dinitrophenol, ethylene oxide, pentachlorophenol, thallium
Hereditary
Amyloid neuropathy
Fabrys disease
CharcotMarieTooth disease type 5, type 2B
Hereditary sensory and autonomic neuropathy (HSAN) type 1, type 1B
Malignant
Carcinomatous (paraneoplastic), myeloma
Infective or postinfective, immune
Acute or inflammatory polyradiculoneuropathy (GuillainBarre syndrome), borreliosis, HIV
Other polyneuropathies
Erythromelalgia
Idiopathic small-fiber neuropathy
Trench foot (cold injury)
Central pain syndromes
Vascular lesions in the brain (especially brainstem and thalamus) and spinal cord
Infarct
Hemorrhage
Vascular malformation
Multiple sclerosis
Traumatic spinal cord injury including iatrogenic cordotomy
Traumatic brain injury
Syringomyelia and syringobulbia
Tumors
Abscesses
Inflammatory diseases other than multiple sclerosis; myelitis caused by viruses, syphilis
Epilepsy
Parkinsons disease

(Continued )
 Neuropathic Pain: Clinical 869

Table 2 (Continued)

Complex painful neuropathic disorders

Complex regional pain syndromes type I and II (reflex sympathetic dystrophy, causalgia)
Mixed pain syndromes
Chronic low back pain with radiculopathy
Cancer pain with malignant plexus invasion
Complex regional pain syndromes

Table 3 Special clinical features that are relevant for specific diagnoses

Special feature Possible diagnosis

Cold allodynia Traumatic nerve injury

Trench foot syndrome
Complex regional pain syndrome
Oxaliplatin-induced polyneuropathy
Central poststroke pain
Deep somatic allodynia Complex regional pain syndrome
SMP Complex regional pain syndrome, acute herpes
Isolated small-fiber affection Diabetic polyneuropathy
Amyloid polyneuropathy
Fabrys disease
Hereditary polyneuropathy
Idiopathic small-fiber polyneuropathy
Painful polyneuropathy in several family members Amyloid polyneuropathy,
Fabrys disease
CharcotMarieTooth disease type 5, type 2B
Hereditary sensory, autonomic polyneuropathy (HSAN) type 1, 1B

SMP, sympathetically maintained pain.

Small sensory and autonomic fibers are Sensorimotor small- and large-fiber neuropathy
involved early in diabetic neuropathy as well as in
impaired glucose tolerance (IGT)
Sometimes early small-fiber deficit

Abnormal epidermal C fiber density is highly

correlated with warm detection threshold (QST) 57.3.1.1.(iii) Amyloid neuropathy
Regeneration capacity of cutaneous C fibers
(experimental challenge with topical capsaicin) is Predominant painful small fiber neuropathy in
both inherited and sporadic forms
reduced
Hyperglycemia in diabetic patients may itself Predominant distal sensory loss of pain and
thermal sensations, often autonomic involvement
be an important factor for acute exacerbations of pain
Up 40% of patients with a so-called burning Severe pain usually of a deep aching quality,
sometimes shooting pains
feet syndrome have a small-fiber neuropathy and
many already demonstrate an IGT indicative of
early diabetes 57.3.1.1.(iv) Fabrys disease (angiokeratoma
corporis diffusum)
57.3.1.1.(ii) Alcoholic neuropathy
Lipid (glycosphingolipid) storage disorder, sex-
Incidence of neuropathy in chronic alcoholism linked recessive
about 10% Heterozygous carrier females occasionally
Paresthesias, burning pain, and tenderness of develop symptoms later in life
the feet Deficiency of -galactosidase A lead to
Painful symptoms not related to the severity of
somatosensory deficit
accumulation of glycosphingolipid (globotriaosylcer-
amide) in tissues
870 Neuropathic Pain: Clinical

Painful small-fiber neuropathy early and pre-

senting clinical feature
57.3.1.1.(vii) Erythromelalgia (erythermalgia)

Boys or young men present with tenderness of Rare condition characterized by painful, red,
hot extremities
the feet and severe spontaneous burning pain in the
legs Acquired form sometimes associated with poly-
cythemia rubra vera
Stroke in young adults due endothelial vascu-
lopathy, about 5% of young male patients with Rare hereditary form caused by a genetic defect
of the sodium channel Nav1.7
cryptogenic stroke have Fabrys disease
Extraneural manifestations: cutaneous telan- Symptoms are aggravated by heat
giectasia, cardiac, renal, pulmonary failure
limb
Immediate pain relief by cooling the affected

Patients with preserved renal function involves

exclusively small myelinated and unmyelinated fibers Feet affected in more than 80%, hands in 25%

Normal results of nerve conduction studies

cases
Underlying small fiber neuropathy in some

Normal results of large-fiber quantitation by

sural nerve biopsy
57.3.1.1.(viii) Acute herpes zoster and
57.3.1.1.(v) Idiopathic small-fiber neuropathy postherpetic neuralgia

Predominant or isolated loss of small fibers Acute zoster: neurocutaneous disease with uni-
lateral efflorescences within one or few dermatomes
involving somatic and autonomic neurons
Often elderly patients Reactivation of varicella zoster virus, which
resides in latent form in sensory trigeminal and dorsal
40% of patients with a so-called burning feet
syndrome have a small-fiber neuropathy suffering root ganglia following primary infection as varicella
from burning sensation, dysesthesia and paresthesias Preherpetic neuralgia (pain before cutaneous
symptoms of acute zoster) of some days
in the feet, lancinating pains, and minimal or no distal
weakness Every dermatome may be affected, thoracic
54%, especially Th510 (15%), trigeminal 20%,
Abnormalities of epidermal skin innervation
(skin biopsies) especially first branch (13%)
Normal results of nerve conduction studies PHN: chronic pain syndrome for more than 3
months after healing of acute zoster lesions
Normal results of large-fiber quantitation by
sural nerve biopsy Incidence of pain 915% after 1 month, 25%
of all age groups after 1 year
Main risk factor for PHN is age
57.3.1.1.(vi) Human immunodeficiency virus- and
highly active antiretroviral therapyassociated
Spontaneous burning pain, stimulus-evoked
pain, and shooting pains possible
neuropathy

Incidence about 1530% in the human immu-

nodeficiency virus (HIV) population
57.3.1.1.(ix) Phantom limb pain

Painful sensory polyneuropathy Phantom limb: amputation of an extremity or

Distal painful paresthesias, spontaneous pain,
evoked shock-like pain, and mechanical allodynia
other body part or transection of a peripheral nerve
leads almost in all cases to a sensation of the missing
with symmetric stocking-like distribution associated or denervated limb
with autonomic neuropathy Phantom limb pain: pain in the missing body
Neuropathy is due to a combination of the virus
itself and antiretroviral drugs (nucleoside analogue
part, up to 80% of patients, mainly after extremity
amputation, cramp-like severe pain in distal phan-
reverse transcriptase inhibitors, highly active antire- tom, sometime painful phantom movements
troviral therapy (HAART)) Stump pain: the amputation stump itself shows
Clinically, these two neuropathies are
indistinguishable
mechanical allodynia especially close to the skin flap
scar or near tender neuromas
Nerve pathology in the painful sensory poly- Telescoping: seemingly shrinking phantom
neuropathy includes axonal atrophy and endoneurial
capillary thickening
Aggravation of both painful and nonpainful
phantom sensations by external stimuli (noxious or
 Neuropathic Pain: Clinical 871

innocuous mechanical stimuli at the stump, some- subcortical white matter, and in the cerebral cortex
times at face or remote ipsilateral body parts) have all been reported (Leijon, G. et al., 1989).
Aggravation of both painful and nonpainful
phantom sensation by internal stimuli (micturition,
The most common are cerebrovascular lesions,
MS, and traumatic spinal cord injuries (SCIs). In
yawning) or emotional stimuli these diseases the incidences are around 8%, 28%,
Main reasons for amputations in times of peace:
peripheral vascular disease (60%), half of the patients
and 40%, respectively (Siddall, P. J. et al., 2003). In
stroke, there is a particularly high prevalence among
also suffer from additional diabetes mellitus, acci- patients with brainstem and thalamic lesions, whereas
dents (20%), rare causes like tumor, arterial there is no difference in the prevalence after ischemic
thrombosis, and osteomyelitis (20%) and hemorrhagic events. In patients with thalamic
Pathophysiological mechanisms: it seems
obvious that the phantom limb image with its com-
infarction only patients with lesions that includes
the ventroposterior region, which receives a particu-
plex perceptual, emotional and cognitive qualities larly dense spinothalamic projection develop central
involves interaction of some cerebral structures. pain. In MS it has not been possible to determine the
This, however, may not preclude the notion that location of the lesions that cause central pain, but
neuronal mechanisms at a much lower level of the clinical observations indicate that many of them are
CNS or even within the severed peripheral nerves located in the spinal cord.
may furnish the brain with the information necessary In stroke the pain is most frequently a hemipain
for creating the phantom. (75%). In MS it affects one or both sides more often in
the legs (87%) than in the upper extremities (31%).
57.3.1.2 Central pain syndromes Trigeminal neuralgia, caused by a lesion in the brain
Central pain is defined as chronic pain following a stem, occurs in 5% of all MS patients. In all conditions,
lesion or disease of the CNS. The cause of pain is a there is a large variation in the quality of central pain.
primary process within the CNS. All lesions that Often central pain develops with a latency of
cause central pain affect the somatosensory pathways. weeks or month after the inciting event. The most
They may be located at any level of the neuraxis. frequent clinical characteristics are spontaneous
Thus lesions at the first synapse in the dorsal horn of burning pain, shooting pain and evoked pain within
the spinal cord or trigeminal nuclei, along the the affected body part. Allodynia to touch or light
ascending pathways through the spinal cord pressure as well as to heat or cold are common. Many
(Figure 1) and brainstem, in the thalamus, in the internal and external events influence central pain,

Figure 1 Different pains in spinal cord injury patients. (a) Typical pattern of below-level neuropathic pain following spinal
cord injury (SCI) at T7. The dark shading represents the distribution of pain. (b) Typical pattern of at-level neuropathic pain
following SCI at T7. The dark shading represents the distribution of pain. Adapted from Siddall, P. 2006. Pain Following Spinal
Gord Injury. In: Walland Melzacks Textbook of Pain, 5th edn. (eds. McMahan, S. B. and Koltzenburg, M.), pp. 10431056.
Elsevier.
872 Neuropathic Pain: Clinical

such as cutaneous stimuli, body movements, visceral

stimuli, emotions, and changes in mood.
Musculoskeletal nociceptive pain is the most
common (58%)
In order to diagnose central pain it is necessary to
ensure that the patient has a CNS disorder by a detailed
At-level neuropathic pain 42%, sharp, shoot-
ing, electric, burning, and stabbing pain located in a
history and a neurological examination, computed region of sensory disturbance
tomography (CT), magnetic resonance imaging
(MRI), cerebrospinal fluid analysis, or electrophysiolo-
Below-level neuropathic pain 34%, develops
months and even years following initial injury, spon-
gical techniques (in particular QST to assess taneous and/or evoked pain diffusely caudal to the
spinothalamic function). It is necessary to exclude pain- level of SCI, burning, aching, stabbing, or electric
ful peripheral neuropathy as a cause. Polyneuropathy is shocks, often with hyperalgesia
not uncommon in, for instance stroke patients, a group Fluctuation with mood, activity, infections
with a high incidence of diabetes. Triggered by sudden noises, jarring movements
The pathophysiological mechanisms of central Both complete and partial injuries may be asso-
ciated with the diffuse, burning pain
pain are unclear. An affection of the spinothalamo-
cortical pathways seems to be crucial for the Incomplete injuries often show allodynia due to
sparing of lemniscal tracts conveying touch
development of central pain, whereas isolated lesions
of the lemniscal system are never associated with sensations
pain. Thus, the pain is always correlated with sensory
in 90%
Mainly not painful phantom sensation present
disturbances, dominated by abnormalities in the
sensibility to temperature and pain. Decreased sensi-
bility to touch, vibration, and joint movements may 57.3.1.3 Complex painful neuropathic
be present, but is often less pronounced. It is disorders
hypothesized that central pain occurs only in patients In addition to the classical neuropathic syndromes
who have lesions of the spinothalamocortical path- like painful diabetic neuropathy, PHN, or phantom
ways. Lesioning of the spinothalamic tract leads to a limb pain there are certain chronic painful condi-
tions that share many clinical characteristics. These
loss of input to normal pain-modulating systems in
syndromes were formerly called reflex sympathetic
particular in thalamic neurons. This loss may creates
dystrophy, M. Sudeck or causalgia and are now
neuronal signs of hyperexcitability. Furthermore,
classified under the umbrella term CRPSs. CRPS
infarction of the dorsolateral thalamus may lead to
are painful disorders that may develop as a dispro-
neuronal disinhibition in medial thalamic nuclei.
portionate consequence of trauma typically
Incomplete SCI may be associated with sensitization
affecting the limbs (Janig, W. and Baron, R., 2003;
of spinal dorsal horn neurons.
Baron, R. and Janig, W., 2004). CRPS type I usually
develops after minor trauma with a small or no
obvious nerve lesion at an extremity (e.g., bone
57.3.1.2.(i) Spinal cord injury pain fracture, sprains, bruises or skin lesions, surgeries).
Prevalence of pain about 65% (Siddall, P. J.
et al., 2003)
CRPS type II develops after trauma with a mostly
large nerve lesion. The patients often report a burn-
Four types of pain (Figure 1): ing spontaneous pain felt in the distal part of the
affected extremity. Stimulus-evoked pains are a
1. Nociceptive pain (musculoskeletal, spastic, striking clinical feature; they include mechanical
visceral) and thermal allodynia and/or hyperalgesia. These
2. Neuropathic pain above level (secondary to nerve sensory abnormalities are most pronounced distally,
lesions: compressive mononeuropathies, CRPS) and have no consistent spatial relationship to indi-
3. Neuropathic pain at level (segmental, circumfer- vidual nerve territories or to the site of the inciting
ential band-like distribution: nerve root lesion. Typically, pain can be elicited by movement
compression, syringomyelia, spinal cord trauma, of and pressure on the joints (deep somatic allody-
or ischemia) nia), even if these are not directly affected by the
4. Neuropathic pain below level (below the level inciting lesion.
of the spinal cord lesion perceived diffusely in Unlike other classical neuropathic pain syn-
anesthetic regions: spinal cord trauma, or dromes CRPS show additional nonsensory features
ischemia) like signs of autonomic dysfunction (abnormal
 Neuropathic Pain: Clinical 873

regulation of blood flow and sweating), edema of skin based classification is often insufficient. First, despite
and subcutaneous tissues, active and passive move- obvious differences in etiology, many of these condi-
ment disorders, and trophic changes of skin, tions share common clinical phenomena; for example,
appendages of skin and subcutaneous tissues. touch-evoked pain in PHN and painful diabetic neu-
Especially for the acute phase, there is accumulating ropathy. Conversely, different signs and symptoms can
evidence that inflammatory processes might be be present in the same disease; for example, pain
involved. Thus, it is likely that nociceptive as well paroxysms and stimulus-evoked abnormalities in
as neuropathic mechanisms are responsible for pain PHN. Second, classification on the basis of anatomical
generation in CRPS, which would then fit into the location also has its shortcomings, as neuroplastic
classification if a mixed pain syndrome (see below). changes following nervous system lesions often give
rise to sensory and pain distributions that do not
57.3.1.4 Mixed pain syndromes respect nerve, root, segmental or cortical territories.
There is agreement that both nociceptive and neuro- Third, and most importantly, decades of rather dis-
pathic processes contribute to many chronic pain couraging systematic research on chronic pain therapy
syndromes and that these different mechanisms may have revealed that a disease based strategy is of no or
explain the qualitatively different symptoms and little help to these patients and their pain.
signs that patients experience. In particular, patients These observations have raised the question of
with chronic low back pain, cancer pain, and CRPS whether an entirely different strategy, in which pain
seem to fit into this theoretical construct (Baron, R. is analyzed on the basis of underlying mechanisms
and Binder, A., 2004). (Jensen, T. S. and Baron, R., 2003), could provide an
In chronic low back pain, the nociceptive com- alternative approach for examining and classifying
ponent stems from activation of intact nociceptors patients, with the ultimate aim of obtaining a better
that innervate ligaments, small joints, muscle, and treatment outcome (Woolf, C. W. et al., 1998;
tendons. The neuropathic pain component is caused Dworkin, R. H. et al., 2003). Our increasing under-
by mechanical compression of the nerve root or by standing of the mechanisms that underlie chronic
action of inflammatory mediators originating from pain, together with the discovery of new molecular
the degenerative disc on the nerve root. Cytokines therapy targets, has strengthened the demand for
and chemokines have been implicated in the che- alternative concepts.
mical pathomechanism of radicular pain. Increased One theoretical possibility to identify pain
levels of tumor necrosis factor alpha (TNF-), mechanisms in patients is to assess differences in
interleukin (IL)-1, IL-6, and granulocyte-macro- the somatosensory phenotype as precisely as possible.
phage colony stimulating factor (GM-CSF) have These specific patterns of signs and symptoms could
been detected in herniated disc tissue compared be compared with the knowledge derived from ani-
with normal, nondegenerated disc tissue. TNF- mal experiments where the association of signs and
applied to nerve roots reproduces the pathologic symptoms and underlying mechanisms has been elu-
changes observed with nerve root exposure to cidated. This concept has led to the development of a
nucleus pulposus extracts. Finally, in an animal symptom-oriented diagnostic approach to neuro-
model of radiculopathy induced by intervertebral pathic pain conditions that supplements the
disc material, inhibitors of TNF- reversed the etiology-based classification scheme, which recog-
radicular pathology. So far it is unclear whether nizes the fact that neuropathic pains are usually a
these mediators are capable of activating intact composite of several pain symptoms. A symptom-
nerve root fibers even without any mechanical com-
oriented approach does not negate the fact that dis-
pression or if the combination of mechanical nerve
tinct neuropathies present differently clinically, and
damage and cytokine attack is the important
that some neuropathic disease states may predispose
prerequisite.
to certain constellations of pain symptoms (e.g.,
touch-evoked pain in PHN). The rationale of this
57.3.2 Mechanism-Based Classification approach recognizes several principles:
As described above neuropathic pain represents het-
erogeneous conditions, which neither can be
Clinically distinct pain symptoms such as
ongoing stimulus-independent pain may be caused
explained by one single etiology or disease nor by a by similar, if not identical, neural mechanisms, even
specific anatomical lesion, i.e., a disease- and anatomy- if the underlying neuropathies differ. For example,
874 Neuropathic Pain: Clinical

nociceptive afferents that develop ongoing activity Beside these negative somatosensory signs (deficit
after axon injury could mediate ongoing pain regard- in function), which are bothering but not painful also
less of the precipitating or sustaining neuropathic positive signs are characteristic for neuropathic con-
cause. ditions. Paresthesias (ant crawling, tingling) are not
More than one pain mechanism is usually pre-
sent in an individual patient.
painful. Painful positive signs are spontaneous (not
stimulus-induced) ongoing pain and spontaneous
Some symptoms, such as mechanical hypersen-
sitivity, can be explained by several distinct neural
shooting, electric schock-like sensations. Many
patients with peripheral neuropathic pain also have
mechanisms that may even coexist in an individual evoked types of pain (stimulus-induced pain, hyper-
patient. sensitivity), which are characterized by several
As said, a symptom-based approach to painful neu- sensory abnormalities. They may be adjacent to or
ropathies can be useful for dissecting the underlying intermingled with skin areas of sensory deficit. Most
neural mechanisms, and this knowledge may even- often, patients report mechanical hypersensitivity
tually be harnessed for the development of novel followed by hypersensitivity to heat and cold. Two
analgesic drugs that differentially target these types of hypersensitivity can be distinguished. First,
mechanisms. A very promising but still hypothetical allodynia is defined as pain in response to a nonno-
approach is summarized in Table 5. ciceptive stimulus. In case of mechanical allodynia
even gentle mechanical stimuli such that even slight
bending of hairs may evoke severe pain. Second,
hyperalgesia is defined as an increased pain sensitiv-
57.4 Signs and Symptoms in ity to a nociceptive stimulus. Another evoked feature
Neuropathic Pain is summation, which is the progressive worsening of
pain evoked by slow repetitive stimulation with
Pain associated with nerve injury has several clinical mildly noxious stimuli, for example, pinprick. A
characteristics (Baron, R., 2006) (Tables 4 and 6). If a small percentage of patients with peripheral nerve
mixed peripheral nerve with a cutaneous branch or a injury have a nearly pure hypersensitive syndrome in
central somatosensory pathway are involved, there is which no sensory deficit is demonstrable.
almost always an area of abnormal sensation and the The quality of the reported sensation may also be
patients maximum pain is coextensive with or within a clue; neuropathic pain commonly has a burning
an area of sensory deficit. This is a key diagnostic and/or shooting quality with unusual tingling, crawl-
feature for neuropathic pain. The sensory deficit is ing or electrical sensations (dysesthesiae).
usually to noxious and thermal stimuli, indicating Although all these characteristics are neither uni-
damage to small-diameter afferent fibers. versally present in, nor absolutely diagnostic of

Table 4 Symptoms and signs of neuropathic pain

Peripheral Postherpetic Complex regional pain

neuropathic pain neuralgia syndrome

Ongoing pain 100% 100% 100%

Brush-evoked pain (dynamic mechanical 36% 60% 60%
allodynia)
Pinprick hyperalgesia 38% 37% Present
Static allodynia 35% 52% Present
Deep somatic allodynia None None 90%
Heat allodynia 29% 29% 30%
Cold allodynia 46% 21% 30%
Edema Low Low 50%
Trophic changes Present Scars 90%
Sympathetically maintained pain Low Low 90% (early)

Source: Data from Pappagallo M. et al. (2000) and unpublished data from Peters and Nurmikko (for a group of patients with mixed peripheral
neuropathic pain) and Baron (for complex regional pain syndrome). Adapted from Scadding, J. W. and Koltzenburg, M. 2006. Painful peripheral
neuropathies. In: Wall and Melzacks Textbook of Pain. 5th edition. (eds. S. B. McMahon and M. Koltzenburg), pp. 973-1000. Elsevier.
Table 5 Proposed model for the relationship between neuropathic pain machanism and clinical symptoms and signs, and possible targets for the rapeutic interventions

Symptom Neuronal processes, mechanisms Targets Optimal compounds Available

Peripheral nociceptor
Spontaneous pain (shooting)
hyperexcitability
Ectopic impulse generation, oscillations in DRG Na channels Selective Na channel blocker Lidocaine, carbamazepine,
oxcarbazepine, lamotrigine, TCA
Spontaneous pain (ongoing) Peripheral nociceptor sensitization
Inflammation within nerves
Cytokine-release Cytokines Cytokine antagonists TNF- antagonists
Heat hyperalgesia Cyclooxygenase blocker NSAIDS?
Reduced activation threshold to:
Heat TRPV1 receptor TRPV1 receptor antagonists Capsaicin cream
Cold hyperalgesia
Cold TRPM8 receptor TRPM8 receptor antagonists Menthol?

Mechanical stimuli ASCI receptor? ASCI receptor antagonists ?

Static mechanical
hyperalgesia Noradrenaline receptor receptor antagonists Phentolamine, sympathetic block, TCA

Histamine H1 receptor H1 receptor antagonists TCA

SMP
Central dorsal horn hyperexcitability
Central sensitization on spinal level Presynaptic:
Ongoing C Input induces increased receptors receptor agonists Opioids
synaptic transmission
Ca channels(2) Ca channel blocker, 2- ligands Gabapentin, pregabalin

Amplification of C fiber input

Gating of A fiber input Postsynaptic:

(mechanical dynamic hyperalgesia) NMDA receptors NMDA receptor antagonists For example, ketamine, dextromethorphan
NK1 receptors NK1 receptor antagonists ?
Dynamic Gating of A fiber input NA channels Selective Na channel blocker For example, carbamazepine
mechanical (mechanical punctate hyperalgesia) Intracellular cascads Mitogen activated protein kinase ?
hyperalgesia mediators

Central spinal disinhibition, ? ? ?

A cold fiber function

Intraspinal inhibitory interneurons

(Functional, degneration)
Punctate
mechanical GABA-ergic GABA-B receptors GABA-B agonists Baclofen
hyperalgesia Opioidergic receptors receptor agonists Opioids

Changes
of supraspinal descending modulation

Inibitory control (noradrenaline, 5-HT) 2 receptors 2 receptor agonists Clonidine

Serotonine receptors NA/5-HT reuptake blocker TCA, venlafaxine, duloxetine
? ?
Faciliatory control ?

5-HT, 5-hydroxytryptamine (serotonin); ASIC, acid-sensing ion channel; GABA, -aminobutyric acid; MAPK, mitogen-activated protein kinase; NK1, neurokinin 1; NMDA, N-methyl-D-
aspartate; NSAIDS, nonsteroidal anti-inflammatory drugs; SMP, sympathetically maintained pain; TCA, tricyclic antidepressants; TNF-, tumor necrosis factor-.
Adapted from Baron R. 2006. Machanism of disease: neuropathic pain a clinical perspective. Nat. Clin. Pract. Neurol. 2, 95106.
876 Neuropathic Pain: Clinical

neuropathic pain, when they are present the diagno- channel genes (Nav1.8 and Nav1.9) are expressed
sis of neuropathic pain is likely (Table 4). selectively in nociceptive primary afferent neurons
and an embryonic channel (Nav1.3) is also upregulated
in damaged peripheral nerves and associated with
57.5 Pathophysiological increased electrical excitability and spontaneous activ-
Mechanisms in Neuropathic Pain ity in neuropathic pain states. The accumulation of
sodium channels at sites of ectopic impulse generation
Most of the present pathophysiological ideas are (sodium channel clusters) may be responsible for low-
derived from experimental work with animal models ering of action potential threshold (Omana-Zapata, I.
for neuropathic pain (Figure 2). This work has et al., 1997). There is evidence that small primary
delineated a series of partially independent pathophy- afferent fibers may acquire a unique sodium channel
siological mechanisms presumed to be responsible for expression profile after nerve lesion (e.g., a specific
different types of neuropathic pain and different soma- relation of up- and downregulation of channel pro-
tosensory abnormalities (Baron, R., 2006). teins), which makes them an interesting target (Wood,
J. N. et al., 2004).
After peripheral nerve damage the accumulation of
57.5.1 Peripheral Sensitization of Primary sodium channel clusters does not only occur at the site
Afferent Nociceptors of the nerve lesion but also far proximally within the
Pain sensations are normally elicited by activity in intact dorsal root ganglion (DRG). In the DRG a
unmyelinated (C) and thinly myelinated (A) reciprocation between a phasically activating voltage-
primary afferent neurons. After peripheral nerve dependent, tetrodotoxin (TTX)-sensitive sodium
lesion these neurons acquire an abnormal sensitiza- conductance and a passive, voltage-independent potas-
tion, i.e., an increased responsiveness of primary sium leak generates characteristic membrane potential
nociceptors to stimulation of their receptive field. oscillations. Ectopic firing is triggered when the ampli-
The characteristic features of sensitized nociceptors tude of oscillation sinusoids reaches threshold (Amir,
are: pathological spontaneous discharge, a lowered R. et al., 2002). Pathological membrane properties
activation threshold for thermal and mechanical sti- within the DRG that occur after nerve lesion are of
muli, and an enhanced discharge to suprathreshold particular therapeutic interest as the DRG is spared of
stimulation (hyperalgesia). the blood brain barrier and might be easily accessible
A large number of dramatic molecular and cellu- for systemic therapies (Jacobs, J. M. et al., 1976). In
lar changes at the level of the primary afferent addition to sodium currents, impaired potassium con-
nociceptor that are triggered by the nerve lesion ductances in myelinated fibers were demonstrated in
underlie these pathological changes. experimental diabetes and may underlie hyperexcit-
Nerve injury is matched by increased expression of ability in these fibers.
messenger RNA for voltage gated sodium channels in Damage to peripheral primary nociceptors
primary afferent neurons. Two voltage-gated sodium also induces upregulation of a variety of receptor

(a) (b)
-adrenoceptor
A/A TRPV1 receptor
c Na channel

Spinal cord NGF

dorsal horn

NE

Figure 2 (continued)
 Neuropathic Pain: Clinical 877

(c) (d)
Opioid receptor
Opioid receptor
Glutamate receptor Glutamate receptor
NA/5-HT receptor NA/5-HT receptor
GABA receptor GABA receptor
-adrenoceptor -adrenoceptor
TRPV1 receptor
TRPV1 receptor Kainate receptor
Kainate receptor
Na channel
Na channel
Ca channel
Ca channel
(2- subunit)
(2- subunit)

C C

A +
A

Figure 2 Mechanisms of peripheral sensitization and central sensitization in neuropathic pain. (a) Primary afferent pathways
and their connections in the spinal cord dorsal horn. Nociceptive C fibers (red) terminate at spinothalamic projection neurons
in upper laminae (orange neuron) whereas nonnociceptive myelinated A fibers project to deeper laminae. The second-order
projection neuron is of wide dynamic range (WDR) type, that is, it receives direct synaptic input from nociceptive terminals
and also multisynaptic input from myelinated A fibers (nonnoxions information, blue neuron system). -aminobutyric acid
(GABA)-releasing interneurons (green neuron) normally exert inhibitory synaptic input on the WDR neuron. Furthermore,
descending modulatory systems synapse at the WDR neuron (green descending terminal). Spinal cord glia cells (gray cell)
also communicate with WDR neuron. (b) Peripheral changes at primary afferent neurons after partial nerve lesion leading to
peripheral sensitization. Some axons are damaged and degenerate (upper two axons), whereas others (lower two axons) are
still intact and connected with the peripheral end organ (skin). The lesion triggers the expression of sodium channels on
damaged neurons. Furthermore, products such as nerve growth factor, which are associated with Wallerian degeneration,
are released in the vicinity of spared fibers (arrows), triggering channel and receptor expression (sodium channels, TRPV1
receptors, adrenoceptors) on uninjured fibers. (c) Spontaneous activity in C nociceptors induces secondary changes in the
central sensory processing, leading to spinal cord hyperexcitability (central sensitization of second-order WDR neurons
indicated by star in orange neuron). This causes input from mechanoreceptive A fibers (light touch and punctate stimuli; blue
neuron system) to be perceived as pain (dynamic and punctate mechanical allodynia; indicates gating at synapse via
AMPA/KA (-amino-3-hydroxy-5-methyl-4-isoxazole propionic acid and kainate) receptors). Several presynaptic (opioid
receptors, calcium channels) and postsynaptic molecular structures (glutamate receptors, NA (adrenceptors), 5-HT
(serotonin) receptors, GABA receptors, sodium channels) are involved in central sensitization. Inhibitory interneurons and
descending modulatory control systems (green neurons) are dysfunctional after nerve lesions, leading to disinhibition or
facilitation of spinal cord dorsal horn neurons and to further central sensitization. (d) Peripheral nerve injury activates spinal
cord nonneural glia cells (gray cell), which further enhances excitability in WDR neurons by releasing cytokines and glutamate.
Adapted from Baron, R. 2006. Mechanisms of disease: neuropathic pain a clinical perspective. Nat. Clin. Pract. Neurol. 2,
95106.

proteins at the membrane some of them only senses noxious heat (>43  C) (Catarina, M. J.
marginally expressed under physiological condi- et al., 2000). Accordingly, TRPV1-deficient
tions. Vanilloid receptors (TRPV1) are located knockout (KO) mice have obvious deficits in
predominantly on nociceptive afferent fibers and chemical and thermal heat nociception but nor-
can be activated by the ingredient of hot chilli mal reactions to noxious mechanical and to
pepper (capsaicin). Physiologically, this receptor noxious cold stimuli.
878 Neuropathic Pain: Clinical

After partial nerve injury and in streptozotocin- adrenergic sensitivity (Figure 2(b)). I.v. epineph-
induced diabetic rats the lesion triggers a TRPV1 rine or physiological noradrenaline release after
downregulation on many damaged afferents but a stimulation of sympathetic efferents that have
novel expression of TRPV1 on uninjured C and A regenerated into the neuroma can excite afferent
fibers (Figure 2(b)) (Hudson, L. J. et al., 2001; Hong, S. nociceptors. After section and reanastomosis of
and Wiley, J. W., 2005) which is likely involved in the peripheral nerves, electrical stimulation of the
development C nociceptor sensitization and the asso- sympathetic trunk at physiological stimulus fre-
ciated symptom of heat hyperalgesia. Recent studies quencies activates regenerated C nociceptors
also provides evidence for a an upregulation of through an 1 adrenoceptor mechanism (Habler,
TRPV1 in medium and large injured dorsal root H. J. et al., 1987). Furthermore, sympathetic activity
ganglion cells (Ma, W. et al., 2005). However, can sensitize identified intact nociceptors follow-
TRPV1 does not appear to be the only transduction ing damage to the nerve in which they run via an 2
mechanism for heat sensitization after nerve injury. adrenoceptor mechanism (Sato, J. and Perl, E. R.,
After partial sciatic nerve ligation, wild-type and 1991). The concept of a pathological coupling
TRPV1-null mice exhibited comparable persistent between sympathetic postganglionic fibers and
enhancement of mechanical and thermal nociceptive afferent neurons via noradrenaline forms the con-
responses (Catarina, M. J. et al., 2000). In this context, ceptual framework for the therapeutic application
a recent study examined strain differences in the of sympathetic blocks in certain pain syndromes,
normal sensitivity to noxious heat in mice. These e.g., CRPS (Price, D. D. et al., 1998; Baron, R. et al.,
differences reflect differential responsiveness of pri- 1999b).
mary afferent thermal nociceptors to heat stimuli due There is increasing evidence that also uninjured
to a genetic variance in CGRP expression and sensi- fibers running in a partially lesioned nerve may take
tivity (Mogil, J. S. et al., 2005). part in pain signaling (Wasner, G. et al., 2005).
In taxol-induced small-fiber painful polyneuropa- Uninjured fibers comingle with degenerating fibers in
thy TRPV4 which is normally activated by heat of the same nerves. Products associated with Wallerian
more than 30  C seems to play a crucial role in degeneration released in the vicinity of spared fibers
producing taxol-induced mechanical hyperalgesia (e.g., nerve growth factor (NGF)) may be the trigger for
(Alessandri-Haber, N. et al., 2004). channel and receptor expression and may alter the
Investigations into temperature-sensitive excita- properties of uninjured afferents (Hudson, L. J. et al.,
tory ion channels also identified several cold- 2001; Wu, G. et al., 2001). Expression of sodium chan-
sensing ion channels in peripheral neurons. The nels, TRPV1 receptors, adrenoreceptors and an
cold- and menthol-sensitive TRP channel increase of TNF- sensitivity has been shown to play
(TRPM8) is activated within the range of 828  C a role in uninjured fibers adjacent to lesioned axons.
(Patapoutian, A. et al., 2003) and sensitized by
menthol. This receptor is expressed in around 10%
of all afferent ganglion neurons of rats, primarily 57.5.2 Central Sensitization
within small-diameter cells (McKemy, D. D. et al.,
As a consequence of periphereal nociceptor hyper-
2002). TRPA1 is activated at lower temperatures and
activity also dramatic secondary changes in the spinal
its exogenous ligand is cinnamaldehyde, a constitu-
cord dorsal horn occur. Peripheral nerve injury leads
ent of cinnamon oil, mustard oil, and horseradish.
to an increase in the general excitability of nocicep-
Peripheral nerve lesions have been shown to upre-
tive and multireceptive spinal cord neurons (multiple
gulate the expression of the latter cold-sensing ion
synaptic input from C as well as A fibers, wide
channels in DRG cells of rats that developed cold
dynamic range neurons). This phenomenon is called
allodynia (Obata, K. et al., 2005). Therefore, upregu-
central sensitization and is defined as an increased
lation or gating of these channels after injury may
responsiveness of nociceptive neurons in the CNS to
lead to the peripheral sensitization of cold-sensitive
their normal afferent input. Central sensitization is
C nociceptors, resulting in the sensory phenomenon
manifested by at least three different modes:
of cold hyperalgesia.
Besides these temperature-sensitive receptors 1. increase of neuronal activity to noxious stimuli,
experimental nerve injury also triggers the expres- 2. expansion of size of neuronal receptive fields, and
sion of functional 1 or 2 adrenoceptors on 3. spread of spinal hyperexcitability to other
cutaneous afferent fibers, these neurons develop segments.
 Neuropathic Pain: Clinical 879

Central sensitization is initiated and maintained by corresponding part on the opposite side of the body.
activity in pathologically sensitized C fibers, which New data suggest that communication of activated
sensitize second-order spinal cord dorsal horn astrocytes via gap junctions may mediate such spread
neurons by releasing glutamate acting on N-methyl- of pain.
D-aspartic acid (NMDA) receptors and the neuropep-
tide substance P. Furthermore, central neuronal
57.5.3 Central Disinhibition and
voltage-gated N-calcium channels located at the
Fascilitation
presynaptic sites on terminals of primary afferent
nociceptors are involved in central sensitization by Physiologically dorsal horn neurons receive a strong
facilitation of the release of glutamate and substance intraspinal inhibitory control by gamma-aminobutyric
P. This channel is overexpressed after peripheral acid (GABA)-ergic interneurons. Partial peripheral
nerve lesion and in rats with streptozotocin-induced nerve injury may promote a selective apoptotic loss
diabetes (Luo, Z. D. et al., 2001). As a consequence of of these GABA-ergic inhibitory neurons in the super-
peripheral nerve lesion the dorsal horn neurons abnor- ficial dorsal horn of the spinal cord (Moore, K. A. et al.,
mally express HNav1.3 (Hains, B. C. et al., 2004) also 2002), a mechanism which would further increases
enhancing central sensitization. Several intracellular central sensitization. Furthermore, there is a novel
cascades contribute to central sensitization, in particu- mechanism of disinhibition following peripheral
lar the mitogen-activated protein kinase (MAPK) nerve injury. This mechanism involves a trans-synap-
system ( Ji, R. R. and Woolf, C. J., 2001). If central tic reduction in the expression of the potassium
sensitization is established, normally innocuous tactile chloride exporter KCC2 in lamina I nociceptive neu-
stimuli become capable of activating spinal cord pain rons. This induces a consequent disruption of the
signalling neurons via A and A low-threshold anion homeostasis in these dorsal horn neurons. The
mechanoreceptors (Tal, M. and Bennett, G. J., 1994). resulting shift in the transmembrane anion gradient
By this mechanism, light innocuous mechanical sti- caused normally inhibitory anionic synaptic currents
muli to the skin induces pain, i.e., punctuate and to be excitatory. Due to this mechanism the release of
dynamic mechanical allodynia. Besides these dramatic GABA from normally inhibitory interneurons now
changes in the spinal cord, sensitized neurons were exerts an excitatory action on lamina I neurons via
also found in the thalamus and primary somatosensory GABAA receptors (Coull, J. A. et al., 2003). The
cortex after partial peripheral nerve injury (Guilbaud, changes in lamina I neurons is induced by brain-
G. et al., 1992). derived neurotrophic factor (BDNF) released from
There is increasing evidence that neuropathic activated spinal cord glia (Coull, J. A. et al., 2005).
pain is in part mediated by an interaction of non- Dorsal horn neurons receive a powerful descend-
neural spinal cord glia and nociceptive neurons ing modulating control from supraspinal brainstem
(Figure 2(d)). In experimental pain states in animals, centers (inhibitory as well as faciliatory) (Vanegas, H.
astrocytes and microglia are activated by neuronal and Schaible, H. G., 2004) (Figures 2(a) and 2(c)). It
signals including substance P, glutamate, and frac- was hypothesized that a loss of function in descend-
talkine (Wieseler-Frank, J. et al., 2005). Activation of ing inhibitory serotonergic and noradrenergic
glia by these substances in turn leads to the release of pathway contributes to central sensitization and
mediators that then act on other glia and also on pain chronicity. This idea nicely explained the effi-
central nociceptive neurons. These include proin- cacy of serotonin and noradrenaline reuptake
flammatory cytokines and most likely also other blocking antidepressants in neuropathic pain.
neuroexcitatory compounds like glutamate. By this However, in animals, mechanical allodynia after per-
interaction central sensitization is augmented. While ipheral nerve injury was dependent upon tonic
traditional therapies for pathological pain have activation of descending pathways that facilitate
focused on neuronal targets, glia might be new ther- pain transmission indicating that structures in the
apeutic targets. mesencephalic reticular formation possibly the
Some patients with neuropathic pain, in particular nucleus cuneiformis (NCF) and the periaqueductal
patients with CRPS characteristically report extra- gray (PAG) are involved in central sensitization in
territorial and/or mirror- image pain. The pain is neuropathic pain (Ossipov, M. H. et al., 2000).
experienced not only in the area of trauma but also in Interestingly, exactly the same brainstem structures
neighboring healthy tissues. In cases of mirror-image were shown to be active in humans with allodynia
pain, the pain is perceived from the healthy, using advanced functional MRI (fMRI) techniques
880 Neuropathic Pain: Clinical

(Zambreanu, L. et al., 2005). Because in most animal be fostered in the future. The solution of the problem
pain models descending facilitation and inhibition neuropathic pain can only be unraveled in the human
are triggered simultaneously, it will be important to experiment, that is, in patients that really feel all
elucidate why inhibition predominates in some neu- components of this complex sensation of neuropathic
ronal pools and facilitation in others. pain.

57.5.4 Inflammation and Neuropathic

Pain 57.6.1 Peripheral Sensitization of Primary
Afferent Neurons in Patients
The connective tissue sheath of peripheral nerves is
innervated by sensory fibers, the nervi nervorum, Microneurographic single-fiber recordings support
which enter the sheath with the nutrient blood ves- the idea of peripheral sensitzation of primary afferent
sels. Some of these fine diameter primary afferents neurons in patients with painful nerve lesions by
are nociceptors (Zochodne, D. W., 1993). Nervi ner- demonstrating abnormal activity and reduced thresh-
vorum are a potential source of pain in diseases of olds in cutaneous afferents. Abnormal ectopic activity
peripheral nerve especially in those conditions with of myelinated mechanosensitive fibers were found in
an inflammatory component. traumatic nerve lesions, entrapment neuropathies, or
A different mechanism involves the production of radiculopathies (Figure 3). Because the ectopic nerve
inflammatory mediators at the site of nerve injury activity correlated in intensity and time course to the
(local inflammation) that might be a critical factor in perceived paresthesias it is likely that pathological
the cascade of events leading to neuropathic pain. activity in A fibers is the underlying mechanism of
Activated macrophages infiltrating from endoneurial positive nonpainful sensations.
blood vessels have been demonstrated in experimen- Recordings from transected nerves in awake
tally injured nerves (Sommer, C. and Myers, R. R., human amputees with phantom limb pain have
1996) and also the dorsal root ganglia after nerve demonstrated spontaneous ectopic activity as well
transection (Lu, X. and Richardson, P. M., 1993). as barrages of action potential firing in afferent A
Proinflammatory cytokines and in particular TNF- and C fibers projecting into the neuroma (Nystrom,
 released by activated macrophages can induce B. and Hagbarth, K. E., 1981). Ectopic excitation
ectopic activity in injured but also in adjacent unin- occurred at multiple sites in damaged sensory neu-
jured primary afferent nociceptors at the lesion site rons. Ongoing activity and mechanical sensitivity
(Sorkin, L. S. et al., 1997) and thus is a potential cause were recorded proximal to the nerve neuroma.
of pain and hyperalgesia (Sommer, C. et al., 1998; Following local anesthetic blockade of the nerve dis-
Wagner, R. et al., 1998; Sommer, C., 2003; tal to the recording site, impulses evoked by
Marchand, F. et al., 2005). mechanical stimulation of the neuroma were abol-
ished, but ongoing activity at the recording site
continued, suggesting that this residual activity
57.6 Pathophysiological arose from the DRGs. As these patients suffered
Mechanisms in Patients in Relation to from spontaneous burning pain and electric shock-
Their Somatosensory Profile like sensations it is very likely that these symptoms
are associated with ectopic firing in primary afferent
Although we have achieved enormous progress in the C fibers.
understanding of pathophysiological mechanisms In few patients with characteristic burning pain
generating neuropathic pain the most important and heat hyperalgesia microneurographic recordings
question is whether it is possible to translate these have provided evidence for sensitized C nociceptors.
concepts into the clinical situation. In the following In patients with erythromelalgia nociceptors dis-
section the findings of the correlation of mechanisms played ongoing activity, which is normally not
and somatosensory abnormalities in animal experi- observed in nociceptors, and there was a sensitization
ments will be matched by observations in human of mechanically insensitive afferents to nonpainful
experimental and clinical pain conditions. tactile stimuli. These abnormalities were only pre-
In order to get more insight into the puzzle of sent in nociceptive fibers, but not of the sympathetic
neuropathic pain and even solve the therapeutic unmyelinated fibers, as an indicator of a neuropathic
dilemma this type of translational research should process (Figure 4) (Orstavik, K. et al., 2003). In the
 Neuropathic Pain: Clinical 881

(a)

Stimulation

Recording 1s

(b)

Intensity of paraesthesiae
5s

Straining during chinchest manoeuvre

Figure 3 Microneurographic multiunit recording from the sural nerve in a patient with a compression of the S1 spinal root by
a herniated disc. (a) Excitation of mechanosensitive units in the receptive field by tactile stimulation (bars). (b) Straining and
chinchest maneuver-provoked paresthesiae and an ectopic discharge of afferent fibers originating from the compressed
root. Adapted from Nordin, M., Nystrom, B., Wallin, U., and Hagbarth, K. E. 1984. Ectopic sensory discharges and
paresthesiae in patients with disorders of peripheral nerve, dorsal roots and dorsal columns. Pain 20, 231245. Elsevier Ltd.

autosomal dominantly inherited form of erythrome- PHN patients cutaneous iontophoresis of histamine
lalgia a mutation in SCN9A was found, a gene that evoked a burning pain sensation whereas only itch
encodes the Nav1.7 sodium channel, leading to an was elicited in normal skin. Again, this phenomenon
altered firing pattern in afferent neurons (Dib-Hajj, S. indicates that nociceptive neurons in the affected
D. et al., 2005). Hence, erythromelalgia is the first skin are abnormally sensitive to histamine (Baron,
channelopathy associated with chronic pain. R. et al., 2001) probably due to expression of a novel
Several clinical observations also support the con- receptor pattern.
cept of sensitized nociceptors in PHN patients. These observations suggest that spontaneous
About 30% of patients with PHN do not show any burning pain and heat hyperalgesia at least in part
loss of sensory function in the affected extremity are associated with sensitization of primary afferent
indicating that in these particular group of patients C fibers to TRPV1 agonists and histamine or with an
loss of neurons is minimal or absent. Accordingly, altered firing pattern in these fibers due to abnormal
thermal sensory thresholds in their region of greatest sodium channels.
pain are either normal or even decreased (heat Cutaneous hypersensitivity to cold, i.e., cold hyper-
hyperalgesia) by up to 24  C (Rowbotham, M. C. algesia, is particularly prominent in patients with
and Fields H. L., 1996; Pappagallo, M. et al., 2000). posttraumatic neuralgias, some small-fiber polyneuro-
The decrease of heat pain perception thresholds is a pathies and chronic CRPS. Another condition of acute
well-known phenomenon of peripheral nociceptor cold intolerance occurs after systemic injection of the
sensitization. Using skin punch biopsy, it was shown cancer chemotherapeutic agent oxaliplatin, which is
that thermal sensitivity is directly correlated with associated with paresthesiae and painful hypersensi-
density of cutaneous innervation in the area of most tivity aggravated by cold. Psychophysical studies of
severe pain (Rowbotham, M. C. et al., 1996). human volunteers using the topical menthol model
Moreover, in PHN patients with heat hyperalgesia suggest that sensitization of cold-sensitive nociceptors
acute topical application of the vanilloid compound can produce cold hyperalgesia in normal volunteers
capsaicin (TRPV1 agonist), enhances pain, a sign that (Wasner, G. et al., 2004). Peripheral sensitization also
is indicative of an increased capsaicin sensitivity of appears to occur in acute oxaliplatin-induced periph-
nociceptors in the affected skin area and has been eral neuropathy (Lehky, T. J. et al., 2004). Therefore, it
attributed to a sensitization of nociceptors as the is likely that cold hyperalgesia in some patients is
driving element in some patients (Petersen, K. L. induced by sensitization of primary afferent cold-
et al., 2000). Furthermore, in a similar group of sensitive nociceptors.
882 Neuropathic Pain: Clinical

57.6.2 Sensitization to Catecholamines

in Patients
Several clinical observations support the idea that
nociceptors acquire a sensitivity to catecholamines
Control subject Patient with erthermalgia
that permits an abnormal excitation by either nora-
drenaline or by circulating catecholamines. This
chemical sensitization makes the nociceptors suscep-
1/8 0.125 Hz
tible for noradrenaline released from efferent
1/4 0.25 Hz sympathetic fibers in the periphery. The consecutive
1/2 pathological sympatheticafferent coupling forms
Electrical stimulation

0.5 Hz
the conceptual framework for sympathetically main-
0.25 Hz tained pain states.
Noradrenergic sensitivity has been described in
several human neuropathies and suggests that the
1/4
ongoing pain can be caused or maintained by the
sympathetic nervous system in selected patients: In
amputees, perineuromal administration of physiologi-
cal doses of norepinephrine induces intense pain as
compared with saline injections (Raja, S. N. et al.,
530 560 590 500 530 560 1998). Intraoperative stimulation of the sympathetic
Response latency (ms)
chain induces an increase of spontaneous pain in
Figure 4 Microneurographic C fiber recordings from the patients with causalgia (CRPS II) but not in patients
peroneal nerve of a normal volunteer (left) and a patient with hyperhidrosis. In PHN, application of norepi-
suffering from erythromelalgia (right). The first trace
shows the original nerve signal. The subsequent
nephrine into a symptomatic skin area increased
recordings are a falling leaf display in which each action spontaneous pain and dynamic mechanical hyperalge-
potential is symbolized by a series of dots. Each row of sia (Choi, B. and Rowbotham, M. C., 1997). In CRPS II
dots represents the latency, at different electrical and posttraumatic neuralgias, intracutaneous norepi-
stimulation frequencies of the fiber, of its receptive field
nephrine rekindles pain and hyperalgesia that had
in the skin. Simultaneous recording of three afferent C
fibers in a control subject shows two units with activity- been relieved by sympathetic blockade. Also intrader-
dependent latency increases at low-frequency mal norepenephrine, in physiologically relevant doses,
stimulation, and one unit that displays latency increases was demonstrated to evoke greater pain in the affected
when excitation frequency increases to 1 impulse per 2 s. regions of patients with sympathetically maintained
These biophysical features are characteristic for
pain (SMP) than in the contralateral unaffected limb,
mechanically insensitive and mechanically sensitive
nociceptive C fibers, respectively. Stimulation of the and in control subjects (Ali, Z. et al., 2000). Because
receptive field with a 750-mN von Frey hair (arrow) noradrenaline-induced pain occurs during a differen-
excited only the mechanosensitive C nociceptor, as tial blockade of myelinated fibers, unmyelinated fibers
evidenced by a strong latency shift to electrical appear to signal sympathetically maintained pain
stimulation. Mechanically insensitive afferents are not
activated by this stimulus. The right panel shows a C
(Torebjork, E. et al., 1995).
fiber recording from a patient with erythromelalgia. The We performed a study in patients with CRPS I
unit had the biophysical properties of a mechanically using physiological stimuli of the sympathetic
insensitive C fiber but responded reproducibly to nervous system (Baron, R. et al., 2002). Cutaneous
mechanical stimulation (arrows). This is consistent with sympathetic vasoconstrictor outflow to the painful
the hypothesis that mechanically insensitive afferents
become sensitized in erythromelalgia. An alternative extremity was experimentally activated to the high-
explanation would be that mechanically sensitive est possible physiological degree by whole body
nociceptors start to display a different pattern of activity- cooling. During the thermal challenge the affected
dependent latency slowing in this disease. Adapted from extremity was clamped to 35  C in order to avoid
Orstavik, K., Weidner, C., Schmidt, R., Schmelz, M.,
thermal effects at the nociceptor level. The intensity
Hilliges, M., Jorum, E., Handwerker, H. and Torebjork, E.
2003. Pathological C-fibres in patients with a chronic as well as area of spontaneous pain and mechanical
painful condition. Brain 126, 567578. Copyright 2003 allodynia (dynamic and punctate) increased signifi-
Oxford University Press. cantly in patients that had been classified as having
SMP by positive sympathetic blocks but not in SIP
 Neuropathic Pain: Clinical 883

patients (Figure 5). The experimental setup used extremities, i.e., piloarrector, sudomotor, and muscle
in the latter study selectively alters sympathetic vasoconstrictor neurons. Therefore, the interaction
cutaneous vasoconstrictor activity without influen- of sympathetic and afferent neurons measured
cing other sympathetic systems innervating the here is likely to be located within the skin as

(a) Affected limb

300
Skin blood flow (PU)

200

100
Unaffected limb

(b)
Affected limb
Skin temperature (C)

34

29

24 Unaffected limb

0 40 80 0 40 80

(c) High sympathetic activity Low sympathetic activity

Dynamic
mechanical
hyperalgesia

Figure 5 Experimental modulation of cutaneous sympathetic vasoconstrictor neurons by physiological thermoregulatory

reflex stimuli in 13 complex regional pain syndrome (CRPS) patients. With the help of a thermal suit, whole-body cooling and
warming was performed to alter sympathetic skin nerve activity. The subjects were lying in a suit supplied by tubes, in which
running water of 12  C and 50  C, respectively (inflow temperature) was used to cool or warm the whole body. By these means
sympathetic activity can be switched on and off. (a) High sympathetic vasoconstrictor activity during cooling induces
considerable drop in skin blood flow on the affected and unaffected extremity (laser Doppler flowmetry). Measurements were
taken at 5-min intervals (mean SD). (b) On the unaffected side, a secondary decrease of skin temperature was documented.
On the affected side, the forearm temperature was clamped at 35  C by a feedback-controlled heat lamp to exclude
temperature effects on the sensory receptor level. Measurements were taken at 5-min intervals (mean SD). (c) Effect of
cutaneous sympathetic vasoconstrictor activity on dynamic mechanical hyperalgesia in one CRPS patient with
sympathetically maintained pain (SMP). Activation of sympathetic neurons (during cooling) leads to a considerable increase of
the area of dynamic mechanical hyperalgesia. From Baron, R., Schattschneider, J., Binder, A., Siebrecht, D. and Wasner, G.
2002. Relation between sympathetic vasoconstrictor activity and pain and hyperalgesia in complex regional pain syndromes:
a casecontrol study. Lancet 359, 16551660, with permission.
884 Neuropathic Pain: Clinical

predicted by the pain-enhancing effect of intracuta- tactile sensations in healthy skin (Gracely, R. H.
neous norepinephrine injections (Ali, Z. et al., 2000). et al., 1992; Price, D. D. et al., 1992), and
Interestingly, the relief of spontaneous pain after 3. using differential nerve blocks dynamic allodynia
sympathetic blockade was more pronounced than is abolished at time points when tactile sensations
changes in spontaneous pain that could be induced is lost, but other modalities remain unaffected
experimentally by sympathetic activation. One (Campbell, J. N. et al., 1988; Ochoa, J. L. and
explanation for this discrepancy might be that a com- Yarnitsky, D., 1993).
plete sympathetic block affects all sympathetic
Therefore, patients with dynamic mechanical allo-
outflow channels projecting to the affected extremity.
dynia would be expected to have central sensitization
It is very likely that in addition to a coupling in the
as their underlying mechanism.
skin, a sympatheticafferent interaction may also
Hyperalgesia to pinprick stimuli, typically elicited
occur in other tissues, in particular in the deep
by probing of the skin with a stiff von Frey hair is
somatic domain such as bone, muscle, or joints.
distinct from dynamic mechanical allodynia because
Supporting this view, especially these structures are
of its different spatial and temporal profile and the
extremely painful in some cases with CRPS (Baron,
fact that it is signaled by nonsensitized, heat-insensi-
R. and Wasner, G., 2001). Furthermore, there may be
tive, A nociceptors.
patients who are characterized by a selective or pre-
Notably, in many neuropathic pain patients the
dominant sympatheticafferent interaction in deep
mechanically sensitive skin area expands widely into
somatic tissues sparing the skin (Wasner, G. et al.,
1999). In summary, it is likely that sympathetically the secondary zone, i.e., the area not affected by the
maintained pain is a consequence of an adrenergic primary nerve lesion, which is also indicative for
sensitization of C nociceptors. CNS mechanisms involved. Furthermore, the area
of secondary mechanical hypersensitivity is a
dynamic phenomenon. In PHN patients with signs
of peripheral nociceptor sensitization (heat hyperal-
57.6.3 Central Sensitization in Patients gesia, see above) cutaneous capsaicin application into
One hallmark of central sensitization of spinal the primary skin area leads to an increase of the
cord neurons in animals is that activity in A fiber allodynic zone into previously nonallodynic and
mechanoreceptors is allowed to gain access to nonpainful skin that had normal sensory function
the nociceptive system and induces pain. These and cutaneous innervation. These observations sup-
phenomena are called mechanical allodynia or port the hypothesis that allodynia in a subgroup of
hyperalgesia. PHN patients is a form of chronic secondary hyper-
Mechanical hypersensitivity is a common phe- algesia dynamically maintained by input from intact
nomenon in neuropathic pain states in patients. and possibly sensitized (irritable) primary afferent
There are several lines of evidence that also in nociceptors to a sensitized CNS (Fields, H. L. et al.,
patients central mechanisms contribute to these sen- 1998; Petersen, K. L. et al., 2000). Because central
sory phenomena. Two distinct types have been sensitization involves the NMDA receptor, the fact
described in patients, dynamic mechanical and pin- that the NMDA receptor antagonist ketamine
prick mechanical hypersensitivity. relieves some neuropathic pain disorders further sup-
There is consensus that dynamic mechanical allo- ports the concept of central sensitization.
dynia is signaled out of the skin by afferent Besides these dramatic changes in the spinal cord,
mechanoreceptors with large myelinated axons that there is now evidence that higher centers of the
normally encode nonpainful tactile stimuli: neuraxis demonstrate an increased excitability as
well as fundamental changes in the somatosensory
1. Reaction time measurements show dynamic representation. Magnetic encephalography (MEG),
mechanical allodynia in patients to be signaled by positron emission tomography (PET), and fMRI stu-
afferents with conduction velocities appropriate for dies revealed cortical changes in patients with
large myelinated axons (Lindblom, U. and Verrillo, phantom limb pain, CRPS, and central pain syn-
R. T., 1979; Campbell, J. N. et al., 1988), dromes (Flor, H. et al., 1995; Willoch, F. et al., 2000;
2. transcutaneous or intraneural stimulation of Pleger, B. et al., 2004; Willoch, F. et al., 2004;
nerves innervating the allodynic skin can evoke Maihofner, C. et al., 2005a) as well as experimental
pain at stimulus intensities which only produce pain models (Baron, R. et al., 1999a; Baron, R. et al.,
 Neuropathic Pain: Clinical 885

2000). Interestingly, these changes correlated with This is an entirely different mechanism as com-
the intensity of the perceived pain and disappeared pared with the peripheral sensitization process of
after successful treatment of the pain (Maihofner, C. cold-sensitive C nociceptors described above leading
et al., 2004; Pleger, B. et al., 2005). to the same somatosensory phenomenon, cold
hyperalgesia.

57.6.4 Central Disinhibition Leading to

Cold Hyperalgesia 57.6.5 Deafferentation: Hyperactivity of
Central Pain Transmission Neurons
Cutaneous hypersensitivity to cold, i.e., cold hyper-
algesia is particularly prominent in patients with The above data convincingly support a role for sen-
posttraumatic neuralgias, some polyneuropathies, sitization mechanisms in the peripheral as well as the
and chronic CRPS. Some observations in neuro- CNS in the generation of neuropathic pain.
pathic pain patients point to central disinhibition as However, in some patients there is a profound cuta-
the underlying mechanism in a subgroup of patients. neous deafferentation of the painful area. Up to 60%
Cold stimuli are usually transmitted centrally by of PHN patients show considerable signs of neuronal
cold-sensitive A fibers, whereas cold pain is con- degeneration and loss of sensory function within the
veyed via nociceptive cold-sensitive C fibers. In affected tissues. Interestingly, some of these patients
patients with polyneuropathies a disproportionate still suffer from severe dynamic mechanical allodynia
loss of A axons and relative sparing of C fibers although the function of nociceptors is diminished or
have been described (Ochoa, J. L. and Yarnitsky, D., absent in the same skin area (Wasner, G. et al., 2005).
1994). These patients suffer from the so-called triple Punch skin biopsies and the anti-PGP 9.5 anti-
cold syndrome (CCC syndrome): cold hypoesthesia body, a panaxonal marker, were used in PHN
in combination with cold hyperalgesia and a cold patients and zoster patients without pain to quantify
skin. A selective damage of cold-sensitive A fibers sensory nerve endings in the affected skin and com-
leads to a lack of inhibition (disinhibition) on C pared the numbers with the homologous
nociceptors transmission normally exerted by conco- contralateral site (Rowbotham, M. C. et al., 1996;
mitant activation of myelinated cold A fibers. This Oaklander, A.L., 1998; Oaklander, A.L., 2001). A
mechanism of a central interaction between cold severe loss could be demonstated on the affected
specific afferents and nociceptors would nicely side (20% as compared with the controls). Neurite
explain the combination of cold hyperalgesia and loss was more prominent in the epidermis than in the
cold hypoesthesia in the above patients. dermis. Furthermore, the PHN group also had lost
Furthermore, it is suggested to play a crucial role in half of the neurites in the contralateral epidermis
central pain syndromes after infarction of the dorso- whereas distant areas where unaffected. However,
lateral thalamus with cold hyperalgesia and these contralateral changes could not be observed
paradoxical heat sensation. In these patients the with QST or axon reflex measurements (Baron, R.
lesion in the lateral thalamus might disinhibit the and Saguer, M., 1994).
nociceptive system projecting through the medial Functional studies support the concept of degen-
thalamus. eration of cutaneous C nociceptors. By using these C
Recently, direct evidence was provided for the fiber axon reflex reactions it is possible to objectively
central disinhibhition theory of cold hyperalgesia in assess cutaneous C fiber function in the human skin.
an experimental pain model in humans. Selective A In some patients the histamine evoked axon reflex
fiber block induces the symptom combination of cold vasodilatation and flare size were impaired or abol-
hyperalgesia and cold hypoesthesia and therefore ished in skin regions with intense dynamic allodynia
mimics the situation in polyneuropathy patients (Baron, R. and Saguer, M., 1993). Similarly, during
with A fiber degeneration. With fMRI the disinhibi- capsaicin stimulation, a subgroup did not experience
tion of the medial nociceptive system (medial worsening of pain (Petersen, K. L. et al., 2000). Using
thalamus, anterior cingulate cortex (ACC), and fron- quantitative sensory testing (QST) some chronic
tal cortices) could be demonstrated. PHN patients have extremely high thermal thresh-
Thus, a loss of peripheral A fiber function disin- olds in areas with marked dynamic allodynia
hibits the nociceptive system centrally leading to (Nurmikko, T. and Bowsher, D., 1990; Nurmikko,
cold hyperalgesia. T. et al., 1994; Choi, B. and Rowbotham, M. C.,
886 Neuropathic Pain: Clinical

1997). Thus, there is a subset of PHN patients with resolves in several weeks. However, a small subgroup
pain and loss of cutaneous C nociceptor function. of patients with PHN has inflammatory infiltrates
Assuming that the DRG cells and the central throughout the affected peripheral nerve, DRG, and
afferent connections are lost in such patients, their dorsal root (Watson, C. P. et al., 1991). Moreover,
pain must be the result of intrinsic CNS changes. In Gilden D. H. et al. (1991) have described a subpopu-
animal studies, following complete primary afferent lation of PHN patients with evidence of continuing
loss of a spinal segment, many dorsal horn cells begin low-level viral expression whose pain responds to
to fire spontaneously at high frequencies (Lombard, antiviral agents. Thus continuing VZV expression
M. C. and Larabi, Y., 1983; Fields, H. L. et al., 1998). could produce sensitization and activity in primary
There is some evidence that a similar process may afferents secondary to inflammation.
underlie the pain that follows extensive denervating
injuries in human. Recordings of spinal neuron activ-
ity in a pain patient whose dorsal roots were injured 57.7 Diagnostic Tools for
by trauma to the cauda equina revealed high-fre- Neuropathic Pain
quency regular and paroxysmal bursting discharges
(Loeser, J. D. et al., 1967). That patient complained of Modern research into the mechanisms of neuropathic
spontaneous burning pain in a skin region that was pain clearly revealed that the nerve lesion leads to
anesthetic by the lesion (anesthesia dolorosa). Thus, dramatic changes in the PNS and CNS that makes it
extensive degeneration of primary afferents asso- distinct from other chronic pain types in which the
ciated with severe somatosensory deficits points to nociceptive system is intact (chronic nociceptive
an increased excitability of deafferented central neu- pain, e.g., osteoarthritis). Furthermore, neuropathic
rons as underlying mechanism. pain states require different therapeutic approaches,
e.g., anticonvulsants, that are not effective in noci-
ceptive pain. To make the situation even more
57.6.6 Inflammation in Patients
complex, many chronic pain states are characterized
In patients with inflammatory demyelinating neuro- by a combination of both the pain types. Best exam-
pathies such as acute GuillainBarre syndrome or ples for the so-called mixed pain syndromes are
vasculitic neuropathies deep proximal aching pain chronic radicular back pain, tumor pain, or CRPS.
in addition to paroxysmal types of pains is a charac- For the clinician, it is, therefore, of utmost impor-
teristic phenomenon. Accordingly, COX2 was found tance to have valid diagnostic tools that differentiate
to be upregulated in nerve biopsy specimens from neuropathic from nociceptive pain or estimate the
patients with chronic inflammatory demyelinating neuropathic pain component in mixed pain
neuropathy and an increased expression of proin- syndromes.
flammatory cytokines have been demonstrated in The easiest approach to this would be to use
peripheral nerves of most vasculitic neuropathies somatosensory symptoms assessed by questionnaires
(Lindenlaub, T. and Sommer, C., 2003). About 15 or history questions or simple signs testable at the
50% of AIDS patients suffer from distal predomi- bedside that are characteristic for neuropathic pain.
nantly sensory neuropathy, which very often is However, a recent study on this issue raised sev-
painful. In sarcoidosis, painful small-fiber neuropathy eral caveats (Rasmussen, P. V. et al., 2004): This study
may be present in a subgroup of patients. In the fluid prospectively looked at symptoms and signs in 214
of artificially produced skin blisters significantly patients with suspected chronic neuropathic pain that
higher levels of IL-6 and TNF- were observed in were a priori classified by pain experts as having the
CRPS-affected extremities as compared with the so-called definite, possible or unlikely neuropathic
uninvolved extremity. In CRPS patients with hyper- pain. Pain symptoms including pain descriptors were
algesia higher levels of the soluble TNF- receptor recorded and sensory tests including repetitive pin-
type I were found (Maihofner, C., et al., 2005b). prick stimulation, examination for cold-evoked pain
Accordingly, a significant increases in IL-1 and by an acetone drop and brush-evoked pain were
IL-6, but not TNF- was demonstrated in the CSF carried out in the maximal pain area and in a control
of individuals afflicted with CRPS as compared with area in order to determine if symptoms and signs
controls (Alexander, G. M. et al., 2005). cluster differentially in groups of patients with
Acute zoster is accompanied by intense inflamma- increasing evidence of neuropathic pain. Several
tion along the affected peripheral nerve that typically symptoms (touch- or cold-provoked pain) and signs
 Neuropathic Pain: Clinical 887

(brush-evoked allodynia) were more prominent in diagnosed according to the results of two indepen-
patients with definite or possible neuropathic pain; dent pain specialists who determined the
however, there was considerable overlap with the predominant pain type (neuropathic versus nocicep-
clinical presentation of patients with unlikely neuro- tive) by means of clinical experience as well as
pathic pain. Even worse, the used pain descriptors neurological examination, electrophysiologic, or
could not at all distinguish between the three clinical imaging techniques. This questionnaire showed a
categories. correct classification rate of 82.5% with a sensitivity
of 80.8% and a specificity 84.7%.
57.7.1 Questionnaires
57.7.2 Bedside Assessment of Neuropathic
Other approaches to find easy screening tools for
Pain
neuropathic pain gave more promising results: a clin-
ician-administered 10-item questionnaire (DN4) Patients with neuropathic pain demonstrate a variety
consists of sensory descriptors (seven items) as well of distinct sensory symptoms and signs that can coexist
as signs related to bedside sensory examination (three in combinations (see above). Therefore, the sensory
items) (Bouhassira, D. et al., 2004). The interview bedside examination should include the following
questions address the quality of the pain (burning, qualities: touch, pinprick, pressure, cold, heat, vibra-
painful cold, and electric shocks) and associated tion, temporal summation, and after sensations
symptoms (tingling, pins and needles, numbness, (Bouhassira, D. et al., 2004; Cruccu, G. et al., 2004,
and itching). The examination consists of the assess- definition in Table 6). To assess either a loss (negative)
ment of hypoesthesia to touch, prick and allodynia to or a gain of somatosensory function (positive sensory
brush. This questionnaire was validated in a prospec- signs) the responses can be graded as normal,
tive study of 160 patients presenting with pain of decreased or increased. The stimulus-evoked (posi-
nociceptive and neuropathic origin and showed tive) pain types are classified as hyperalgesic or
86.0% of correctly identified patients (sensitivity allodynic, and according to the dynamic or static
82.9%, specificity 89.9%). character of the stimulus (Rasmussen, P. V. et al., 2004).
A different approach to distinguish neuropathic Touch can be assessed by gently applying cotton
from nonneuropathic pain uses a patient-based ques- wool to the skin, pinprick sensation by the response
tionnaire with nine questions without the need of to sharp pinprick stimuli, deep pain by gentle pres-
examinations by the physician (PainDetect) sure on muscle and joints, and cold and heat sensation
(Freynhagen, R. et al., 2006). The questionnaire con- by measuring the response to a thermal stimulus, for
sists of slightly different sensory descriptors (seven example, thermorollers kept at 20 or 45  C. Cold
items, burning pain, tingling or prickling (electricity), sensation can also be assessed by the response to
sensitivity to touch (clothes, blanket), pain caused by acetone spray. Vibration can be assessed by a tuning
light pressure (e.g., with finger), shooting pain or fork placed at strategic points (interphalangeal joints,
electric shock-like pain, occasional painful cold or etc.). Abnormal temporal summation is the clinical
heat (e.g., bath tub), and numbness), the question equivalent to increasing neuronal activity following
whether the pain is spatially radiating and a questions repetitive C fiber stimulation >0.3 Hz. This windup-
addressing the individual pain pattern. In the latter, like pain can be produced by mechanical and thermal
the patients have to choose one of four graphically stimuli. After-sensations the persistence of pain
illustrated pain patters (permanent pain with a light long after termination of a painful stimulus is
variability, stable permanent pain with paroxysmal another characteristic feature of neuropathic pain,
pain attacks, paroxysmal pain attacks with no perma- which is closely related to a coexistent dynamic or
nent pain, and pain attacks with fluctuating static hyperalgesia. When present, allodynia or
permanent pain in between). This questionnaire was hyperalgesia can be quantified by measuring inten-
validated in 392 patients recruited at 10 highly spe- sity and area. At present, it is generally agreed that
cialized pain centers with either pain of pure assessment should be carried out in the area of max-
predominant neuropathic origin (n 167, e.g., imal pain using the contralateral area as control.
PHN, painful polyneuropathies, and nerve trauma) However, contralateral segmental changes following
or pure or predominant nociceptive origin (n 225, a unilateral nerve or root lesion cannot be excluded,
e.g., ostheoathritis, mechanical low back pain, so an examination at mirror sites may not necessarily
and inflammatory arthropathies). Patients were represent a true control site.
888 Neuropathic Pain: Clinical

Table 6 Definition and assessment of negative and positive sensory symptoms or signs in neuropathic pain

Assessment Bedside Expected pathological

Symptom/sign Definition exam response

Negative signs and symptoms

Hypoesthesia Reduced sensation to nonpainful Touch skin with painters Reduced perception,
stimuli brush, cotton swab, or numbness
gauze
Pall hypoesthesia Reduced sensation to vibration Apply tuning fork on bone Reduced perception threshold
or joint
Hypoalgesia Reduced sensation to painful Prick skin with single pin Reduced perception,
stimuli stimulus numbness
Thermhypoesthesia Reduced sensation to cold/warm Contact skin with objects Reduced perception
stimuli of 10  C (metal roller,
glass with water,
coolants like acetone)
Contact skin with objects
of 45  C (metal roller,
glass with water)
Spontaneous sensations/pain
Paraesthesia Nonpainful ongoing sensation (ant Grade intensity (010) Area
crawling) in cm2
Paroxysmal pain Shooting electrical attacks for Number per time Grade
seconds intensity (010)
Threshold for evocation
Superficial pain Painful ongoing sensation often of Grade intensity (010) Area
burning quality in cm2
Evoked pain
Mechanical Normally nonpainful light moving Stroking skin with painters Sharp burning superficial pain
dynamic stimuli on skin evoke pain brush, cotton swab, or Present in the primary affected
allodynia gauze zone but spread beyond into
unaffected skin areas
(secondary zone)
Mechanical static Normally nonpainful gentle static Manual gentle mechanical Dull pain
allodynia pressure stimuli at skin evoke pressure at the skin Present in the area of affected
pain (damaged or sensitized)
primary afferent nerve
endings (primary zone)
Mechanical Normally not painful/slightly Manual pricking the skin Sharp superficial pain
punctuate, stinging stimuli evoke pain with a safety pin, sharp Present in the primary affected
pinprick stick, or stiff von Frey zone but spread beyond into
hyperalgesia hair unaffected skin areas
(secondary zone)
Temporal Repetitive application of identical Pricking skin with safety Sharp superficial pain of
summation single noxious stimuli is pin at interval <3 s for increasing intensity
perceived as increasing pain 30 s
sensation (windup like pain)
Cold allodynia Normally nonpainful/slightly painful Contact skin with objects Painful often burning
hyperalgesia cold stimuli evoke pain of 20  C (metal roller, temperature sensation
glass with water,
coolants like acetone)
Control: contact skin with Present in the area of affected
objects of skin (damaged or sensitized)
temperature primary afferent nerve
endings (primary zone)
Heat allodynia Normally nonpainful/slightly painful Contact skin with objects Painful burning temperature
hyperalgesia heat stimuli evoke pain of 40  C (metal roller, sensation
glass with water)

(Continued )
 Neuropathic Pain: Clinical 889

Table 6 (Continued)

Assessment Bedside Expected pathological

Symptom/sign Definition exam response

Control: contact skin with Present in the area of affected

objects of skin (damaged or sensitized)
temperature primary afferent nerve
endings (primary zone)
Mechanical deep Normally nonpainful pressure on Manual light pressure at Deep pain at joints or muscles
somatic allodynia deep somatic tissues evoke pain joints or muscle

It is important to realized the spatial distribution of analysis of the exact somatosensory phenotype of
abnormal sensations. In neuropathic conditions, the neuropathic pain patients (Figure 6). To evaluate
distinction between primary and secondary area cor- plus or minus signs in patients, an age- and gender-
responds to the tissue supplied by damaged nerves and matched database for absolute and relative QST
the area outside this innervation territory. Mechanical reference data was established for healthy human
hypersensitivity often expands into the secondary area. subjects. Until now this nationwide multicenter trial
In Table 6 several sensory signs that can be found in comprises complete sensory profiles of 180 healthy
peripheral neuropathies are defined and the appropri- human subjects and more than 1500 neuropathic pain
ate tests to assess these signs clinically are summarized. patients of a variety of entities. Thermal detection
and pain thresholds including a test for the presence
of paradoxical heat sensations, mechanical detection
57.7.3 Apparative Assessment of thresholds to von Frey filaments and a 64-Hz tuning
Somatosensory Phenotypes fork, mechanical pain thresholds to pinprick stimuli
and blunt pressure, stimulus/response functions for
It is important to realize that conventional electro- pinprick and dynamic mechanical allodynia (pain to
physiological techniques like NCS, SEP, etc., only light touch), and pain summation (windup ratio)
assess the function of myelinated peripheral axonal using repetitive pinprick stimulation are determined.
systems, the affection of small fibers including noci- Data of healthy human subjects were analyzed for the
ceptors are missed. Similarly, SEP measures the influence of body side and region, age, and gender.
conduction in the lemniscal system (dorsal columns) For most variables pathological values of positive and
and not the spinothalamic tract, which conveys nox- negative signs can be detected on the basis of refer-
ious information. Therefore, alternative diagnostic ence data.
procedures have to be used to analyze nociceptive
fibers and tracts in the peripheral or CNS, especially
to diagnose small fiber neuropathies and central pain
57.7.4 Skin Biopsies
states that are associated with a isolated damage to
central nociceptive systems. The advent of skin biopsy as a diagnostic and inves-
A sophisticated neurophysiologic technique to tigative tool has changed the diagnostic picture in
test both systems in the PNS and the CNS is QST, neuropathic pain; skin biopsies can be done at several
which uses standardized mechanical and thermal sti- sites to establish a spatial profile of nerve fiber invol-
muli (graded von Frey hairs, several pinprick stimuli, vement, and can be repeated over time. They have
pressure algometers, quantitative thermotesting, etc.). generally met good patient acceptance in neuropa-
Another advantage of QST is that it assesses a loss of thies, and even in allodynic states like PHN. They
function (minus signs) as well as a gain of function can be used to assess epidermal nerve fibers, predo-
(positive signs). For example Allodynia or hyperal- minantly C fiber nociceptors, sympathetic fibers
gesia, can be quantified by measuring intensity, innervating the cutaneous sweat glands, and myeli-
threshold for elicitation, duration, and area. nated A and A fibers in the dermis.
A standardized protocol for QST was recently Immunostaining of skin punch biopsies for the
proposed by the nationwide German Network on panaxonal marker protein gene product 9.5
Neuropathic Pain (Rolke R. et al., 2006) including (PGP9.5), a neuronal ubiquitin hydrolase is believed
13 parameters of sensory testing procedures for the to identify axons of all categories of nerve fibers. In a
890 Neuropathic Pain: Clinical

Figure 6 Quantitative sensory testing (QST) a battery of sensory tests: figure of methods. The standardized QST protocol
assesses 13 variables in seven test procedures (ag). All procedures are presented including a time frame for testing over one
area. (a) (thermal testing; CDT, cold detection threshold; WDT, warm detection threshold; TSL, thermal sensory limen for
alternating warm and cold stimuli; PHS, number of paradoxical heat sensations during the TSL procedure; CPT, cold pain
threshold; HPT, heat pain threshold) comprises detection and pain thresholds for cold, warm, or hot stimuli (C and A fiber
mediated). (b) (MDT, mechanical detection threshold) tests for A fiber function using von Frey filaments. (c) (MPT,
mechanical pain threshold) tests for A-fiber-mediated hyper- or hypoalgesia to pinprick stimuli. (d) (stimulus/response
functions: MPS for pinprick stimuli, and ALL for dynamic mechanical allodynia) assesses again A-mediated sensitivity to
sharp stimuli (pinprick), and also A-fiber-mediated sensitivity to stroking light touch (CW, cotton wisp; QT, cotton wool tip;
BR, brush). (e) (WUR, windup ratio) compares the verbal ratings within five trains of a single pinprick stimulus (a) with a series
(b) of 10 repetitive pinprick stimuli to calculate WUR as the ratio: b/a. (f) (VDT, vibration detection threshold) tests again for A
fiber function using a RydelSeiffer 64-Hz tuning fork. (g) (PPT, pressure pain threshold) is the only test for deep pain
sensitivity, most probably mediated by muscle C and A fibers.

variety of neuropathies affecting small caliber nerve densities have been documented with improvement
fibers, denervation has been documented to occur in in neuropathic status spontaneously or after therapies
a length-dependent manner. These include diabetes (Hart, A. M. et al., 2002).
mellitus, HIV-associated sensory neuropathy Cutaneous innervation is usually reduced in PHN
(Polydefkis, M. et al., 2002), Fabrys disease (Scott, skin compared to mirror-image skin when assessed
L. J. et al., 1999), restless legs syndrome, and idio- with PGP (Rowbotham, M. C. et al., 1996; Oaklander,
pathic small-fiber sensory neuropathies (Holland, N. A.L. et al., 1998; Petersen, K. L. et al., 2000) (Figure 7).
R. et al., 1997). However, a significant proportion of PHN subjects
Punch skin biopsies have more recently been used have similar fiber densities in both PHN and mirror-
as a serial measure, to examine changes in epidermal image skin, suggesting that the symptoms of pain and
innervation over time, or after therapeutic interven- sensory dysfunction is not due to a mere loss in over-
tions in various neuropathic conditions. For example, all innervation density (Rowbotham, M. C. et al., 1996;
epidermal reinnervation occured concomitant with Petersen, K. L. et al., 2000). Oaklander demonstrated
symptomatic improvement in sensory neuropathy that the average cutaneous fiber density was lower in
(Nodera, H. et al., 2003), and impressive increases in subjects with established PHN than in those who had
epidermal, dermal, and sweat gland innervation recovered without PHN after herpes zoster.
 Neuropathic Pain: Clinical 891

Normal PHN

Figure 7 Representative skin biopsy samples from a postherpetic neuralgia (PHN) patient. PGP 9.5 immunofluorescence in
normal and mirror-image painful PHN skin. The patient suffered from spontaneous pain and allodynia, but had a marked loss
of thermal sensation (heat pain threshold elevated by 7.3  C). D, dermis; E, epidermis. (Reprinted from Rowbotham, M. C.,
Yosipovitch, G., Connolly, M.K., Finlay, D., Forde, G. and Fields, H. L. 1996. Cutaneous innervation density in the allodynic
form of postherpetic neuralgia. Neurobiol. Dis. 3, 205214. Copyright 1996 Academic Press.)

Furthermore, mirror-image skin had lower fiber den- effector component of C fibers as indirect means
sities than distant control skin in subjects with (Baron, R. and Saguer, M., 1993; Schuller, T. B.
established PHN (Oaklander, A.L. et al., 1998). et al., 2000). Studies of the axon reflex vasodilatation
(neurogenic flare) using visual inspection (Figure 8),
thermography, or laser Doppler flowmetry in
57.7.5 Microneurography
response to a chemical stimulus that activates cuta-
Microneurography has been used to assess functional neous C fibers (histamine, capsaicin) are commonly
abnormalities in single small fibers. This technique used. While independent of patient cooperation, a
makes it possible to subgroup C nociceptors in major drawback of this technique is the dependence
healthy controls into a variety of physiologically on a variety of other factors that affect the effector
different classes. The technique requires huge tech- response.
nical skills from the researcher and is very time
consuming.
Regarding pathological states it has been possible
to identify spontaneously active C fibers in patients
e.g., with erythromelalgia (Orstavik, K. et al., 2003)
(Figure 4). However, the results about abnormal fir-
ing patterns in diabetic neuropathy patients obtained
so far have not been conclusive (Handwerker, perso-
nal communication). One major problem has been
that normal aging changes the functional properties
of C afferents dramatically. As the patient popula-
tions are generally older, normative data from a
cohort of healthy older subjects have to be obtained.
Furthermore, as recordings are performed from sin- Figure 8 Histamine flares in a patient with severe
gle afferent C fibers and not from whole nerves it is postherpetic neuralgia in the right segment Th6. Histamine
not possible to obtain reliable quantitative data about iontophoresis was performed within the allodynic
transitional zone at the cranial border of the affected and
fiber numbers. adjacent segment and on the contralateral corresponding
site. The formerly affected dermatome is characterized by
heavy scar tissue, whereas the allodynic area shows no
57.7.6 Indirect Test of Afferent scars or secondary pigmentation. Note the difference in
Unmyelinated Fiber Function flare size (ipsilateral 260 mm2, contralateral 2060 mm2)
indicating an impairment of nociceptive C fiber function in
The function of afferent unmyelinated fibers in the the allodynic region. Adapted from Baron, R. and Saguer,
skin can be objectively assessed by using the efferent M. 1993. Copyright 1993 Oxford University Press.
892 Neuropathic Pain: Clinical

57.7.7 Imaging Techniques mechanism by assessing a specific sensory symptom.

There are, however, some important caveats with such
A new MRI technique, MR neurography makes it
an approach. Recent clinical experimental studies indi-
possible to identify small patches of inflammation in
cate that it is not appropriate to link one single
peripheral nerves. This might be of value in inflam-
symptom with exactly one mechanism. It was shown
matory neuropathies, acute zoster, or schwannoma
that one specific symptom may be generated by several
and neuroma detection (Haanpaa, M. et al., 1998;
entire different underlying pathophysiological mechan-
Bendszus, M. et al., 2004) (Figure 9). More patients
isms. It became clear that only a specific symptom
with visable lesions associated with acute herpes zos-
constellation, a symptom profile, i.e., a combination of
ter still had pain after 3 months. No data are available
negative and positive sensory phenomena, might be
to demonstrate changes that occur in chronic PHN.
able to predict the mechanisms and not just one single
MRI can also identify changes in peripheral nerves
symptom. In order to translate these ideas into the
and secondary neurogenic alterations in skeletal
clinical framework of neuropathic pain the most impor-
muscle (Koltzenburg, M. and Bendszus, M., 2004).
tant approach is to characterize the somatosensory
Prolongation of the T2 relaxation time and gadoli-
phenotype of the patients as precisely as possible.
nium enhancement of denervated muscle develop in
For example, standardized QST methods described
parallel with the development of pathological spon-
above can nicely distinguish between phenotypic sub-
taneous activity on EMG that is a feature of axonal
types of PHN patients with distinct sensory symptom
damage. Axonal nerve lesions cause a hyperintense
constellations that are likely correlated with different
signal on T2-weighted MRI of the affected nerve at
underlying mechanisms (sensitization type, deafferen-
and distal to the lesion site, which correlates with
tation type, see above). Distinct pathophysiological
Wallerian degeneration and nerve edema. While
changes in the excitability of peripheral and central
the MRI changes do not distinguish between painful
neurons are likely involved in pain generation (for
and painless nerve lesions, they supplement the
details, see Figure 10).
differential diagnosis of peripheral nerve disease.
As attractive a subtype classification based on the
Furthermore, diffusion tensor imaging technique is
nociceptor function and evoked pain types might be it
capable of detecting degeneration of fibers in the
should be emphasized that not all PHN patients fit
dorsal columns in severe peripheral neuropathies.
exactly into one category or the other. It rather seems
to be a continuum. Furthermore, in a large group of
57.7.8 Do We Have Diagnostic Tool PHN patients many heterogeneous patterns of sensory
to Dissect Individual Mechanisms in dysfunction were detected (Pappagallo, M. et al., 2000).
Neuropathic Pain? Accordingly, detailed testing of sensory function, che-
mical stimulation, and cutaneous innervation in one
The modern theoretical concept of a mechanism-based PHN patient clearly showed areas of relative preserva-
therapy uses the assumption that a specific symptom tion in close vicinity to impaired thermal sensation, both
predicts a specific underlying mechanism. It should be within the affected dermatome (Petersen, K. L. et al.,
possible for a clinician to identify a particular 2000). Furthermore, the sensory patterns showed a var-
iation over the time course of PHN. However, by
classification of PHN patients due to sensory perception
thresholds within the most painful skin area it is possible
to detect the predominant individual sensory profile and
the most likely underlying pain-generating mechanism.
Another caveat addressing the predictive value of
QST measurements for therapy was recently revealed
by two independent studies. It was hypothesized that
topically applied lidocaine that is believed to act on
Figure 9 Magnetic resonance image of a patient with ectopic discharges in nociceptive fibers would be in
acute herpes zoster. Spinal cord in the cervical affected particular beneficial for patients with sensitized per-
region. Note the contrast enhancement in the dorsal root,
ipheral nociceptors as compared with patients with a
dorsal horn of the spinal cord, and lesser also in the ventral
root. The patients was severely affected with pain and motor
loss of dermal nociceptors. In contrast to the hypoth-
weakness at the left arm (T1 image with contrast medium). esis, however, skin biopsies, QST, histamine test as
Photo courtesy of Prof. Dr. T. Tolle, Munich. well nerve conduction studies could not identify
 Neuropathic Pain: Clinical 893

PHN I: peripheral and central sensitization

PHN II: deafferentation of small and large fibers

Dynamic
>>6 mechanical
allodynia
Gain of function
5 Static hyperalgesia
to blunt pressure
4 Pinprick
Heat
hyperalgesia
3 hyperalgesia

1
Z-score

2
Static hypoalgesia
Loss of function

3 Pinprick to blunt pressure

Tactile hypoalgesia
4
hypoesthesia
Cold and warm
5 hypoesthesia
<<6
CDT WDT TSL PHS CPT HPT MDT MPT MPS ALL WUR VDT PPT
QST parameter
Figure 10 Z-score sensory profiles of two patients suffering from postherpetic neuralgia (PHN). Patient PHN I (open
circles) presents the Quantitative sensory testing (QST) profile of a 70-year-old woman suffering from PHN for 8 years.
Ongoing pain was 80 on a 0 to 100 numerical rating scale. The profile shows a predominant gain of sensory function in terms
of heat pain hyperalgesia (HPT), pinprick mechanical hyperalgesia (MPS), dynamic mechanical allodynia (ALL), and static
hyperalgesia to blunt pressure (PPT) outside the 95% confidence interval of the distribution of healthy subjects (gray zone).
This profile is consistent with a combination of peripheral and central sensitization. Patient PHN II (filled circles) shows the
QST profile of a 71-year-old woman with pain for 8 months. Ongoing pain was 70 on a 0 to 100 numerical rating scale. The
QST profile shows predominant loss of sensory function. Note the cold (CDT), warm detection thresholds (WDT), thermal
sensory limen (TSL), heat pain thresholds (HPT), tactile detection thresholds (MDT), and mechanical pain thresholds to
pinprick stimuli (MPT) outside the normal range as presented by the gray zone. This profile is consistent with a combined
small- and large-fiber sensory deafferentation. Z-score: Numbers of standard deviations between patient data and group-
specific mean value. Adapted from Rolke, R., Baron, R., Maire, C., Tolle, T. R., Treede, R. D., Binder, A., Birbaumer, N.,
Birklein, F., Botefur, C. I., Braune, S., Flor, H., Huge, V., Klug, R., Landwchrmeyer, G. B., Magerl, W., Maihofer, G., Rolko, C.,
Scherens, A., Sprenger, T., Valet, M., and Wasserka, B. 2006. Quantitative Sensory testing in the German Research Network
on Neuropathic Pain (DFNS): standardized protocol and reference values. Pain 123, 231243.

lidocaine responders in painful neuropathies G., 2006;; McQuay H. et al., 1995; McQuay, H. J. et al.,
(Herrmann, D. N. et al., 2006) and PHN (Wasner, G. 1996; Sindrup, S. H. and Jensen, T. S., 1999; Dworkin,
et al., 2005). Alternatively, it might be possible that R. H. et al., 2003; Finnerup, N. B. et al., 2005; Siddall, P.
surviving A fibers in C-nociceptor-deprived skin may J. and Middleton, J. W., 2005). For central neuropathic
express sodium channels, develop ectopic firing and pain there are limited data. The treatments discussed
might therefore be the target for lidocaine. below have all been demonstrated to provide statisti-
cally significant and clinically meaningful treatment
benefits compared with placebo in multiple rando-
57.8 Therapy mized controlled trials (Table 7).
Nonmedical treatments, i.e., interventional proce-
The number of trials for peripheral neuropathic pain dures, neurostimulation techniques, neurosurgical
has expanded greatly in the past few years. These have destructive techniques, psychological therapy as
been summarized in several recent meta-analyses that well as physiotherapy and occupational therapy are
are referred to in the following (Baron, R. and Wasner, not the focus of this review.
894 Neuropathic Pain: Clinical

Table 7 Pharmacological therapy of neuropathic pain syndromes

Compound Evidence

Antidepressants
Amitriptyline PHN **, PNP **, PTN *, STR *
Venlafaxine PNP **
Duloxetine PNP **
Anticonvulsants (Na channel)
Carbamazepine PNP *, TGN **
Oxcarbazepine PNP *
Lamotrigine HIV *, PNP *, STR *
Anticonvulsants (Ca channel)
Gabapentin PHN **, PNP **, HIV *, CRPS *, PHAN *, SCI *,
MIX *, CANC *
Pregabalin PHN **, PNP **, SCI *
Tramadol PHN *, PNP **
Long-acting strong opioids
Morphine PHN *, PHAN *
Oxycodone PHN *, PNP **
Cannabinoids MS **, PA ,, MIX *
Topical therapy
Capsaicin cream PHN *, PNP *, PTN *
Lidocaine patch PHN **, MIX *

CANC, neuropathic cancer pain; CRPS, complex regional pain syndrome; HIV, HIV neuropathy; MIX, mixed neuropathic pain cohort; MS,
central neuropathic associated with MS (multiple sclerosis); PA, central neuropathic pain after plexus avulsion; PHAN, phantom pain; PHN,
postherpetic neuralgia; PNP, polyneuropathy (mainly diabetic); PTN, posttraumatic neuralgia; SCI, spinal cord injury; STR, poststroke pain;
TGN, trigeminal neuralgia.
Levels of evidence: **, several randomized clinical trials (RCTs) or meta-analyses; *, at least 1 RCT; ,, unclear.

57.8.1 Antidepressants 57.8.2 Anticonvulsants (Ca Channel

Modulators)
The effectiveness of tricyclic antidepressants
(TCAs) in neuropathic pain may account for their There are extensive clinical trials of gabapentin for
broad range of pharmacological actions. These com- chronic neuropathic pain. These studies examined
pounds are inhibitors of the reuptake of patients with PHN, DPN, mixed neuropathic pain
monoaminergic transmitters. They are believed to syndromes, phantom limb pain, GuillainBarre syn-
potentiate the effects of biogenic amines in CNS drome, and acute and chronic pain from SCI.
pain-modulating pathways. In addition, they block Improvements in sleep, mood, and quality of life
voltage dependent sodium channels and  adrener- were also demonstrated. Pregabalin, the successor
gic receptors. drug of gabapentin was shown to be efficacious in
Venlafaxine and duloxetine that blocks both ser- PHN, DPN, and SCI. There is growing evidence
otonin and norepinephrine reuptake (serotonin supporting the mechanism of action on the 2 sub-
noradrenaline reuptake inhibitors (SNRI)) were effi- unit of neuronal calcium channels partly located at
cacious in diabetic painful neuropathy (DPN). In a the presynaptic spinal terminals of primary afferent
comparison of venlafaxine and imipramine in nociceptors. One advantage over gabapentin is its
patients with painful polyneuropathy, both antide- superior bioavailability, which makes it easier to use
pressants demonstrated superior pain relief without the need of long titration periods.
compared with placebo but did not differ from each Gabapentin and pregabalin are generally well toler-
other. ated, safe, have no drug interactions, and no negative
Selective serotonin reuptake inhibitors (SSRI) impact on cardiac function. This advantages makes
have fewer adverse effects and are generally better them a suitable option especially for the elderly, a
tolerated than TCAs. However, they did not show population very often suffering from several comor-
convincing efficacy in neuropathic pain states. bidities that need multiple drug therapies.
 Neuropathic Pain: Clinical 895

57.8.3 Anticonvulsants (Na Channel of primary nociceptive afferents. On initial applica-

Blockers) tion, it has an excitatory action and produces burning
pain and hyperalgesia, but with repeated or pro-
For lamotrigine there is evidence of efficacy for HIV
longed application it inactivates the receptive
sensory neuropathy, DPN, and central poststroke
terminals of nociceptors. Therefore, this approach is
pain. Carbamazepine is very effective in trigeminal
reasonable for those patients whose pain is main-
neuralgia. However, the strength of evidence is much
tained by anatomically intact sensitized nociceptors.
lower for the benefit of these drugs in other types of
Capsaicin has been reported to reduce the pain of
neuropathic pain. Oxcarbazepine which has fewer
PHN and DPN.
side effects and drugdrug interactions than carba-
mazepine was shown to be effective in painful
57