A stable and systematic daily change in light levels at dawn and dusk provides the most reliable ... more A stable and systematic daily change in light levels at dawn and dusk provides the most reliable indicator of the phase of the day. It is likely that organisms have evolved mechanisms to use these twilight transitions as the primary zeitgeber to adjust their circadian phases. In this study, we investigated under natural illumination conditions the effects of daylight exposure restricted to twilights on the timing of testicular regression and locomotor activity of the house sparrow (Passer domesticus), which possesses a strongly self-sustaining circadian system. Two experiments were performed on adult male house sparrows. Beginning in the third week of April, the first experiment examined whether exposure to natural daylight only during twilights influenced the timing of testicular regression and concomitant changes in testosterone-dependent beak color of reproductively mature sparrows. Interestingly, there was a significant delay in testicular regression and depigmentation of the be...
Reproductive biology and endocrinology : RB&E, Jan 26, 2004
Little is known about how hormones interact in the photoperiodic induction of seasonal responses ... more Little is known about how hormones interact in the photoperiodic induction of seasonal responses in birds. In this study, two experiments determined if the treatment with melatonin altered inhibitory effects of prolactin on photoperiodic induction of seasonal responses in the Palearctic-Indian migratory male redheaded bunting Emberiza bruniceps. Each experiment employed three groups (N = 6-7 each) of photosensitive birds that were held under 8 hours light: 16 hours darkness (8L:16D) since early March. In the experiment 1, beginning in mid June 2001, birds were exposed to natural day lengths (NDL) at 27 degree North (day length = ca.13.8 h, sunrise to sunset) for 23 days. In the experiment 2, beginning in early April 2002, birds were exposed to 14L:10D for 22 days. Beginning on day 4 of NDL or day 1 of 14L:10D, they received 10 (experiment 1) or 13 (experiment 2) daily injections of both melatonin and prolactin (group 1) or prolactin alone (group 2) at a dose of 20 microgram per bird...
Avian behavior and physiology are temporally regulated by a complex circadian clock on both a dai... more Avian behavior and physiology are temporally regulated by a complex circadian clock on both a daily and an annual basis. The circadian secretion of the hormone melatonin is a critical component of the regulation of circadian/daily processes in passerine birds, but there is little evidence that the gland regulates annual changes in primary reproductive function. Here it is shown that locomotor rhythms of house sparrows, Passer domesticus, which are made arrhythmic by either pinealectomy or maintenance in constant light, can be synchronized by daily administration of melatonin of different durations to simulate the melatonin profiles indicative of long and short photoperiods. Pinealectomized male sparrows maintained in constant darkness were entrained by both melatonin regimens. In both cases, testes were regressed and the song control nuclei were small. Intact male house sparrows maintained in constant light were also entrained to both melatonin regimens. However, sparrows that recei...
Background Chick pinealocytes exhibit all the characteristics of a complete circadian system, com... more Background Chick pinealocytes exhibit all the characteristics of a complete circadian system, comprising photoreceptive inputs, molecular clockworks and an easily measured rhythmic output, melatonin biosynthesis. These properties make the in vitro pineal a particularly useful model for exploring circadian control of gene transcription in a pacemaker tissue, as well as regulation of the transcriptome by primary inputs to the clock (both photic and noradrenergic). Results We used microarray analysis to investigate the expression of approximately 8000 genes within cultured pinealocytes subjected to both LD and DD. We report that a reduced subset of genes was rhythmically expressed in vitro compared to those previously published in vivo, and that gene expression rhythms were lower in amplitude, although the functional distribution of the rhythmic transcriptome was largely similar. We also investigated the effects of 6-hour pulses of light or of norepinephrine on gene expression in free-...
There are two effects of long day length on reproductive responses in birds, one is the photoindu... more There are two effects of long day length on reproductive responses in birds, one is the photoinduction of gonadal growth and maturation and the other is the induction of gonadal regression and photorefractoriness. Although it is likely that the same photoreceptors are involved in the photoinduction of gonadal growth and the onset and maintenance of photorefractoriness, and so the influence of wavelength should be similar, this has not been investigated. Therefore, we investigated the influence of light wavelength on reproductive photorefractoriness in the migratory male blackheaded bunting held under long photoperiods. In mid May, when photoperiod was approximately 14L:10D (14 hours light:10 hours darkness), eight groups of sexually mature birds were moved indoors on an artificial photoperiod of 14L:10D (L-450 lux, D-0 lux). Then after 3 weeks, for six groups, a 4-h light period in the morning (zt 0-4; zt 0 [zeitgeber time 0] refers to the beginning of lights-on period) or in the evening (zt 10-14) was substituted with green (428 nm), red (654 nm) or white light at 16 ± 2 lux intensity. Of the remaining two groups, one was maintained on 14L:10D and the other transferred to 10L:14D; these served as controls. At the end of 4 weeks, all birds were found to have undergone testicular regression, irrespective of LD cycle they were exposed to. When these gonadally regressed birds were subjected to 16L:8D for another 4 weeks, to test their responsiveness to the stimulatory effects of long day lengths, only those exposed to 10L:14D and 14L:10D with a 4-h green light period showed testicular regrowth. On the other hand, birds exposed to 14L:10D with a 4-h white or red light period remained fully regressed, similar to 14L:10D controls. Except for some individual difference, there was no difference in response between the groups that received a 4-h light period in the morning and that received it in the evening. These results suggest that the wavelengths of light influence induction of buntings from the photosensitive state into the photorefractory state. Whereas the short light wavelengths facilitated recovery from the photorefractoriness, the long light wavelengths were more effective in maintaining the photorefractoriness.
Subcutaneous daily injections of ovine prolactin (PRL) inhibit photoperiodic induction of gonads ... more Subcutaneous daily injections of ovine prolactin (PRL) inhibit photoperiodic induction of gonads and fattening in the female redheaded bunting (Emberiza bruniceps). However, birds respond to the same photoperiod (15L:9D) after withdrawal of the exogenous PRL. Further, PRL injections given in late hours during the subjective days induce ovarian regression in the photostimulated females. The results suggest that the PRL has an inhibitory role on gonadal photoperiodic responses in redheaded buntings, and extend our understanding of the regulatory function of PRL in migratory birds.
SUMMARY The effects of the duration and time of food availability on stimulation of the photoperi... more SUMMARY The effects of the duration and time of food availability on stimulation of the photoperiodic responses (fattening and gain in body mass, and growth and development of testes) were investigated in the migratory blackheaded bunting(Emberiza melanocephala). Two experiments were performed. Experiment 1 examined the effects of a reduction in the duration of food supply in buntings that were subjected to long day lengths (16 h:8 h L:D) and received food ad libitum (group I) or for restricted durations, coinciding with the end of the lights-on period, of 8 h (group II) and 4 h (group III). Buntings of group I gained in body mass, whereas there was a mixed response in group II (half the birds gained and half lost body mass), and all birds of group III lost body mass. There was no effect on testis growth in groups I and II, but testes grew more slowly in group III. Experiment 2 investigated the effects of both the duration and the time of food availability. Of five groups of birds, ...
To investigate if the plumage colour mutation relates to circadian activity behaviour in the zebr... more To investigate if the plumage colour mutation relates to circadian activity behaviour in the zebra finch, Taeniopygia guttata, wild type grey and pied mutant males were sequentially subjected for three weeks each to 12 h light:12 h darkness (12L:12D) and constant dim light (LL(dim)) condition. During the first 3 h of the 12 h day, pied finches were significantly greater active than grey finches. Also, as compared to grey, pied finches had longer activity duration in the day, with early activity onsets and late activity offsets. This was changed under free-running condition (LL(dim)), when the activity later in the subjective day (clock hour 9 and 11) was significantly greater in grey than in pied finches.Two colour morphs differed in daily activity profile, but not in the total daily activity or circadian rhythm period. Results suggest that greyzebra finches represent late chronotype, and could perhaps be better adapted to a seemingly stressful environment, such as low intensity LL(...
Comparative Biochemistry and Physiology Part A: Molecular & Integrative Physiology, 2015
Eukaryotic cells produce chemical energy in the form of ATP by oxidative phosphorylation of metab... more Eukaryotic cells produce chemical energy in the form of ATP by oxidative phosphorylation of metabolic fuels via a series of enzyme mediated biochemical reactions. We propose that the rates of these reactions are altered, as per energy needs of the seasonal metabolic states in avian migrants. To investigate this, blackheaded buntings were photoperiodically induced with non-migratory, premigratory, migratory and post-migratory phenotypes. High plasma levels of free fatty acids, citrate (an intermediate that begins the TCA cycle) and malate dehydrogenase (mdh, an enzyme involved at the end of the TCA cycle) confirmed increased availability of metabolic reserves and substrates to the TCA cycle during the premigratory and migratory states, respectively. Further, daily expression pattern of genes coding for enzymes involved in the oxidative decarboxylation of pyruvate to acetyl-CoA (pdc and pdk) and oxidative phosphorylation in the TCA cycle (cs, odgh, sdhd and mdh) was monitored in the hypothalamus and liver. Reciprocal relationship between pdc and pdk expressions conformed with the altered requirements of acetyl-CoA for the TCA cycle in different metabolic states. Except for pdk, all genes had a daily expression pattern, with high mRNA expression during the day in the premigratory/migratory phenotypes, and at night (cs, odhg, sdhd and mdh) in the nonmigratory phenotype. Differences in mRNA expression patterns of pdc, sdhd and mdh, but not of pdk, cs and odgh, between the hypothalamus and liver indicated a tissue dependent metabolism in buntings. These results suggest the adaptation of oxidative phosphorylation pathway(s) at gene levels to the seasonal alternations in metabolism in migratory songbirds.
The present study was carried out on a Palearctic-Indian migratory species, the blackheaded bunti... more The present study was carried out on a Palearctic-Indian migratory species, the blackheaded bunting (Emberiza melanocephala), to understand the importance of photoperiodism and circannual rhythms in determining seasonality in changes in body mass and testis size in birds. An initial experiment determined the effects of duration and intensity of light on photoperiodic induction. The birds were exposed to different photoperiods (hours of light:hours of darkness; 11.5L:12.5D, 12L:12D, 12.5L:11.5D and 13L:11D) at the same (~ 450 lux) light intensity, and to 13L:11D at different light intensities (50-, 100-, 400-, 800-and 1000-lux). The induction and subsequent regression of photoperiodic responses were dependent upon duration and intensity of the light period until these reached threshold. A second experiment investigated if an endogenous seasonal rhythm underlies photoperiodism in buntings. Birds maintained since February on a 8L:16D photoperiod (a non-inductive short day length invariably used to ensure photosensitivity in photoperiodic species) were subjected periodically to 16L:8D (a long day length), one group every month from mid-March to mid-August. The magnitude of long day response in body mass and testes decreased as the duration of the short days progressed, but testicular response was restored in birds that were exposed to long days in July and August. The birds exposed simultaneously to short, long, and natural day lengths for 32 weeks underwent an induction-regression cycle under long days and natural day lengths, but not under short days in which a decrease in body mass occurred after about 20 weeks. The last experiment examined the importance of latitudinal migration on photoperiodism, by comparing the response to long days of three groups which included birds from populations those were held in the outdoor aviary for 1 or 2 years at 27° N and those immediately arrived from their breeding grounds (~ 40° N). There was no difference in the photoperiodic induction among the three groups, indicating that neither experience to changing photoperiods during a migratory journey, nor to long photoperiods at breeding grounds, were critical for a subsequent response (initiationtermination-reinitiation) cycle. Taken together, these findings suggest that (1) the blackheaded bunting has its own endogenous timing program, which is regulated by the photoperiod, and (2) the photoperiodic programs of bunting are flexible enough to accommodate variations in the amplitude of environmental cycles. Thus, it appears that photoperiodism has evolved independently of the evolution of migration in this species.
Birds seasonally switch from one life history state (LHS) to another to maximize their fitness. A... more Birds seasonally switch from one life history state (LHS) to another to maximize their fitness. Accordingly, they exhibit distinct differences in their physiological and behavioral phenotypes between seasons. Possible molecular mechanisms underlying changes through the seasons have scarcely been examined in migratory birds. The present study measured key genes suggested to be involved in the metabolic regulation of 4 photoperiodically induced seasonal LHSs in a long-distance migratory songbird, the blackheaded bunting (Emberiza melanocephala). Buntings were held under short days (8 h light:16 h darkness, 8L:16D), during which they maintained the winter nonmigratory phenotype. Then they were exposed for several weeks to long days (13L:11D). Differences in the activity-rest pattern, body fattening and weight gain, testis size, organ (heart, intestine) weights, and blood glucose and triglyceride levels confirmed that buntings sequentially exhibited spring migration-linked premigratory,...
This study investigated photoperiodic plasticity in hypothalamic expression of genes implicated i... more This study investigated photoperiodic plasticity in hypothalamic expression of genes implicated in the photoperiodic light perception (rhodopsin, melanopsin, neuropsin and peropsin), transduction (pax6, bmal1, clock, per2 and casr), induction (eya3, tshβ, dio2 and dio3, gnrh and gnih) and metabolism (NPY, sirtuin1, foxO1, hmgcr, citrate synthase and dehydrogenases) in photosensitive and photorefractory redheaded buntings. There was a significant increase in eya3, tsh β, dio2, pax6 and rhodopsin and decrease in dio3 mRNA expression at hour 15 and/or 19 on the day photosensitive buntings were subjected to a 13- or 16 h, but not to 8- and 11 h light exposure. Downstream reproductive and metabolic gene expression was not altered, except for an increase in those genes coding for succinate and malate dehydrogenase enzymes involved in lipogenesis. Photorefractory buntings had high dio3 mRNA expression which significantly declined after 1 short day exposure, suggesting possible involvement ...
Proceedings of the Royal Society B: Biological Sciences, 2011
We investigated the effects of temperature on photoperiodic induction of the phenologies linked w... more We investigated the effects of temperature on photoperiodic induction of the phenologies linked with migration (body fattening and premigratory night-time restlessness, Zugunruhe ) and reproduction (testicular maturation) in the migratory blackheaded bunting. Birds were exposed for four weeks to near-threshold photoperiods required to induce testicular growth (11.5 L:12.5 D and 12 L:12 D) or for 18 weeks to a long photoperiod (13 L:11 D) at 22°C or 27°C (low) and 35°C or 40°C (high) temperatures. A significant body fattening and half-maximal testicular growth occurred in birds under the 12 L, but not under the 11.5 L photoperiod. Further, one of six birds in both temperature groups on 11.5 L, and four and two of six birds, respectively, in low- and high-temperature groups on 12 L showed the Zugunruhe . Buntings on 13 L in both temperature groups showed complete growth-regression cycles in body fattening, Zugunruhe and testis maturation. In birds on 13 L, high temperature attenuated ...
We have investigated phasic response of the photoperiodic clock to wavelength (color) and intensi... more We have investigated phasic response of the photoperiodic clock to wavelength (color) and intensity of light in the male redheaded bunting (Emberiza bruniceps). Two experiments were performed. Experiment I examined whether varying the wavelength and intensity of first (entraining, E) and/or second (inducing, I) light pulse will alter the effects of a skeleton photoperiod. Birds were subjected for a period of 6 weeks to skeleton photoschedules (6L:5D:1L:12D; 6 h E-light pulse, 1 h I-light pulse) containing either an E-pulse at 5 or 20 lx coupled with I-pulse at 100 lx, or an E-pulse at 100 lx, coupled with I-pulse at 5-or 20-lx intensity. Additional two groups that received both E-and I-pulses at 100 lx served as controls. All photoschedules were employed in two colors-white and red (654 nm). There was the wavelength-and intensity-dependent stimulation of the testis growth and development. Long wavelengths of light (red light) induced faster and greater gonadal response, but the effects were also intensity dependent. Experiment II tested whether in the photostimulated birds held on long photoperiods the change in wavelength and intensity of light hours in the morning (entraining period) or the evening (inducing period) will influence the maintenance of the photoperiodic sensitivity. Birds were subjected initially to a long photoperiod of 14L:10D (L ϭ ف 500 lx; D ϭ 0 lx) and then after 3 weeks, a 4-h light period in the morning (zeitgeber time, zt, 0-4) or in the evening (zt 10-14) of 14 L was substituted with white, green (528 nm), or red (654 nm) light at ف 20-lx intensity. One group maintained on 14L:10D and other exposed to 10L:14D served as controls. After another 7 weeks, all birds were subjected to 16L:8D for a further 4 weeks to test for their responsivity to long-day photostimulation as a consequence of exposure to different experimental photoperiods. Testes regrew under 16L:8D only in birds that were exposed to 10L:14D or to 14L:10D with a green light pulse. However, there was no effect of the timing (morning, evening) of the light pulse. Taken together, the results from both the experiments indicate that in the redheaded bunting (1) the photoperiodic clock responds differentially to different wavelengths (colors, spectra) and intensities of light, and (2) the effects of wavelength and intensity of light on the clock are phase dependent, and such phasic effects can be seen in skeleton photoperiods in which light is applied discretely at different circadian phases.
This investigation attempts to identify the mechanism (s) involved in the fat deposition in a pho... more This investigation attempts to identify the mechanism (s) involved in the fat deposition in a photoperiodic migratory species, the Blackheaded bunting (Emberiza melanocephala). Groups of photosensitive male buntings were exposed to resonance, interrupted night, ...
We investigated whether pineal is part of the circadian clock system which regulates circadian rh... more We investigated whether pineal is part of the circadian clock system which regulates circadian rhythms of activity and photosensitivity in the Indian weaver bird (Ploceus philippinus). Two experiments were performed. The first experiment examined the induction of testicular growth, and androgen-dependent beak pigmentation and luteinizing hormone (LH)-specific plumage coloration in pinealectomised (pinx) and sham-operated (sham) birds exposed to short day
Olfactory and visual sensory mechanisms seem to play a critical role in migratory orientation and... more Olfactory and visual sensory mechanisms seem to play a critical role in migratory orientation and navigation. How these two mechanisms are functionally linked with other migratory processes is unknown. We investigated this, in relation to the profound behavioural shift that occurs during migration in the night-migratory blackheaded bunting (Emberiza melanocephala). Photosensitive unstimulated birds singly housed in activity cages were subjected to long days (LD 16/8). The activity of each bird was continuously monitored. Daily activity pattern defined the nonmigratory phase (no nocturnal activity) and migratory phase (intense nocturnal activity, Zugunruhe). Body mass and testis size were measured at the beginning and end of the experiment. Long days induced the migratory phenotype (body fattening and Zugunruhe) and testis maturation. The c-fos (Fos) immunoreactivity, as marker of the neural activity of the olfactory and visual subsystems, was measured at midday (8 h after lights-on) and midnight (4 h after lights-off) after the first seven long days (nonmigratory phase) and after seven nights of the Zugunruhe (migratory phase). In the nonmigratory phase, Fos-like immunoreactive (Fos-lir) cells in the olfactory and visual subsystems were high in the day and low at night. In the migratory phase, this was reversed; Fos-lir cells were high at night and low in the day. The phase inversion of neural activity in the olfactory and visual systems in parallel with the behavioral shift suggests a functional coupling between the systems governing migratory flight (expressed as Zugunruhe) and migratory orientation and navigation.
To investigate whether the photoperiodic clocks of species possessing strongly self-sustaining ci... more To investigate whether the photoperiodic clocks of species possessing strongly self-sustaining circadian clocks share identical features, we compared the full response cycle (initiation and termination of the response) in body mass and testes of the non-migratory house sparrow (Passer domesticus) with that of the migratory redheaded bunting (Emberiza bruniceps) under Nanda-Hamner experiments. Birds were exposed to a 36 h day (L:D=6:30 h), controls exposed to a 24 h day (L:D=6:18 h), for a period of 31 weeks. By week 18 of L:D=6:18 h, there was a small increase in body mass among sparrows, but not among buntings, and the testes of bunting did not grow, while those of sparrow grew slightly. The response to L:D=6:30 h is of particular interest. There was a rapid gain and subsequent loss in the body mass of bunting, but not of sparrows. Further, both species underwent a testicular cycle as if they were exposed to long days, but the response of sparrows was slower and hence delayed the attainment of peak testicular size. Such a differential response to exotic light cycles between these two photosensitive species, despite their similar circadian oscillatory properties (strong self-sustainment), could suggest a species-specific adaptation of the endogenous clock involved in photoperiodic regulation of avian seasonality.
This study investigated the functional linkage between food availability and activity behavior in... more This study investigated the functional linkage between food availability and activity behavior in the Palaearctic Indian night migratory blackheaded bunting (Emberiza melanocephala) subjected to artificial light-dark (LD) cycles. Two experiments were performed on photosensitive birds. In the first one, birds were exposed to short days (LD 10/14; Experiment 1A), long days (LD 13/11; Experiment 1B), or increasing daylengths (8 to 13 h light/d; Experiment 1C) and presented with food either for the whole or a restricted duration of the light period. In Experiments 1A and 1B, illumination of the light and dark periods or of the dark period, alone, was changed to assess the influence of the light environment on direct and circadian responses to food cycles. In the second experiment, birds were exposed to LD 12/12 or LD 8/16 with food availability overlapping with the light (light and food presence in phase) or dark period (light and food presence in antiphase). Also, birds were subjected to constant dim light (LL dim) to examine the phase of the activity rhythms under synchronizing influence of the food cycles. Similarly, the presentation of food ad libitum (free food; FF) during an experiment examined the effects of the food-restriction regimes on activity rhythms. A continuous measurement of the activity-rest pattern was done to examine both the circadian and direct effects of the food and LD cycles. Measurement of activity at night enabled assessment of the migratory phenotype, premigratory restlessness, or Zugunruhe. The results show that (i) light masked the food effects if they were present together; (ii) birds had a higher anticipatory activity and food intake during restricted feeding conditions; and (iii) food at night alone reduced both the duration and amount of Zugunruhe as compared to food during the day alone. This suggests that food affects both the daily activity and seasonal Zugunruhe, and food cycles act as a synchronizer of circadian rhythms in the absence of dominant natural environmental synchronizers, such as the light-dark cycle.
A stable and systematic daily change in light levels at dawn and dusk provides the most reliable ... more A stable and systematic daily change in light levels at dawn and dusk provides the most reliable indicator of the phase of the day. It is likely that organisms have evolved mechanisms to use these twilight transitions as the primary zeitgeber to adjust their circadian phases. In this study, we investigated under natural illumination conditions the effects of daylight exposure restricted to twilights on the timing of testicular regression and locomotor activity of the house sparrow (Passer domesticus), which possesses a strongly self-sustaining circadian system. Two experiments were performed on adult male house sparrows. Beginning in the third week of April, the first experiment examined whether exposure to natural daylight only during twilights influenced the timing of testicular regression and concomitant changes in testosterone-dependent beak color of reproductively mature sparrows. Interestingly, there was a significant delay in testicular regression and depigmentation of the be...
Reproductive biology and endocrinology : RB&E, Jan 26, 2004
Little is known about how hormones interact in the photoperiodic induction of seasonal responses ... more Little is known about how hormones interact in the photoperiodic induction of seasonal responses in birds. In this study, two experiments determined if the treatment with melatonin altered inhibitory effects of prolactin on photoperiodic induction of seasonal responses in the Palearctic-Indian migratory male redheaded bunting Emberiza bruniceps. Each experiment employed three groups (N = 6-7 each) of photosensitive birds that were held under 8 hours light: 16 hours darkness (8L:16D) since early March. In the experiment 1, beginning in mid June 2001, birds were exposed to natural day lengths (NDL) at 27 degree North (day length = ca.13.8 h, sunrise to sunset) for 23 days. In the experiment 2, beginning in early April 2002, birds were exposed to 14L:10D for 22 days. Beginning on day 4 of NDL or day 1 of 14L:10D, they received 10 (experiment 1) or 13 (experiment 2) daily injections of both melatonin and prolactin (group 1) or prolactin alone (group 2) at a dose of 20 microgram per bird...
Avian behavior and physiology are temporally regulated by a complex circadian clock on both a dai... more Avian behavior and physiology are temporally regulated by a complex circadian clock on both a daily and an annual basis. The circadian secretion of the hormone melatonin is a critical component of the regulation of circadian/daily processes in passerine birds, but there is little evidence that the gland regulates annual changes in primary reproductive function. Here it is shown that locomotor rhythms of house sparrows, Passer domesticus, which are made arrhythmic by either pinealectomy or maintenance in constant light, can be synchronized by daily administration of melatonin of different durations to simulate the melatonin profiles indicative of long and short photoperiods. Pinealectomized male sparrows maintained in constant darkness were entrained by both melatonin regimens. In both cases, testes were regressed and the song control nuclei were small. Intact male house sparrows maintained in constant light were also entrained to both melatonin regimens. However, sparrows that recei...
Background Chick pinealocytes exhibit all the characteristics of a complete circadian system, com... more Background Chick pinealocytes exhibit all the characteristics of a complete circadian system, comprising photoreceptive inputs, molecular clockworks and an easily measured rhythmic output, melatonin biosynthesis. These properties make the in vitro pineal a particularly useful model for exploring circadian control of gene transcription in a pacemaker tissue, as well as regulation of the transcriptome by primary inputs to the clock (both photic and noradrenergic). Results We used microarray analysis to investigate the expression of approximately 8000 genes within cultured pinealocytes subjected to both LD and DD. We report that a reduced subset of genes was rhythmically expressed in vitro compared to those previously published in vivo, and that gene expression rhythms were lower in amplitude, although the functional distribution of the rhythmic transcriptome was largely similar. We also investigated the effects of 6-hour pulses of light or of norepinephrine on gene expression in free-...
There are two effects of long day length on reproductive responses in birds, one is the photoindu... more There are two effects of long day length on reproductive responses in birds, one is the photoinduction of gonadal growth and maturation and the other is the induction of gonadal regression and photorefractoriness. Although it is likely that the same photoreceptors are involved in the photoinduction of gonadal growth and the onset and maintenance of photorefractoriness, and so the influence of wavelength should be similar, this has not been investigated. Therefore, we investigated the influence of light wavelength on reproductive photorefractoriness in the migratory male blackheaded bunting held under long photoperiods. In mid May, when photoperiod was approximately 14L:10D (14 hours light:10 hours darkness), eight groups of sexually mature birds were moved indoors on an artificial photoperiod of 14L:10D (L-450 lux, D-0 lux). Then after 3 weeks, for six groups, a 4-h light period in the morning (zt 0-4; zt 0 [zeitgeber time 0] refers to the beginning of lights-on period) or in the evening (zt 10-14) was substituted with green (428 nm), red (654 nm) or white light at 16 ± 2 lux intensity. Of the remaining two groups, one was maintained on 14L:10D and the other transferred to 10L:14D; these served as controls. At the end of 4 weeks, all birds were found to have undergone testicular regression, irrespective of LD cycle they were exposed to. When these gonadally regressed birds were subjected to 16L:8D for another 4 weeks, to test their responsiveness to the stimulatory effects of long day lengths, only those exposed to 10L:14D and 14L:10D with a 4-h green light period showed testicular regrowth. On the other hand, birds exposed to 14L:10D with a 4-h white or red light period remained fully regressed, similar to 14L:10D controls. Except for some individual difference, there was no difference in response between the groups that received a 4-h light period in the morning and that received it in the evening. These results suggest that the wavelengths of light influence induction of buntings from the photosensitive state into the photorefractory state. Whereas the short light wavelengths facilitated recovery from the photorefractoriness, the long light wavelengths were more effective in maintaining the photorefractoriness.
Subcutaneous daily injections of ovine prolactin (PRL) inhibit photoperiodic induction of gonads ... more Subcutaneous daily injections of ovine prolactin (PRL) inhibit photoperiodic induction of gonads and fattening in the female redheaded bunting (Emberiza bruniceps). However, birds respond to the same photoperiod (15L:9D) after withdrawal of the exogenous PRL. Further, PRL injections given in late hours during the subjective days induce ovarian regression in the photostimulated females. The results suggest that the PRL has an inhibitory role on gonadal photoperiodic responses in redheaded buntings, and extend our understanding of the regulatory function of PRL in migratory birds.
SUMMARY The effects of the duration and time of food availability on stimulation of the photoperi... more SUMMARY The effects of the duration and time of food availability on stimulation of the photoperiodic responses (fattening and gain in body mass, and growth and development of testes) were investigated in the migratory blackheaded bunting(Emberiza melanocephala). Two experiments were performed. Experiment 1 examined the effects of a reduction in the duration of food supply in buntings that were subjected to long day lengths (16 h:8 h L:D) and received food ad libitum (group I) or for restricted durations, coinciding with the end of the lights-on period, of 8 h (group II) and 4 h (group III). Buntings of group I gained in body mass, whereas there was a mixed response in group II (half the birds gained and half lost body mass), and all birds of group III lost body mass. There was no effect on testis growth in groups I and II, but testes grew more slowly in group III. Experiment 2 investigated the effects of both the duration and the time of food availability. Of five groups of birds, ...
To investigate if the plumage colour mutation relates to circadian activity behaviour in the zebr... more To investigate if the plumage colour mutation relates to circadian activity behaviour in the zebra finch, Taeniopygia guttata, wild type grey and pied mutant males were sequentially subjected for three weeks each to 12 h light:12 h darkness (12L:12D) and constant dim light (LL(dim)) condition. During the first 3 h of the 12 h day, pied finches were significantly greater active than grey finches. Also, as compared to grey, pied finches had longer activity duration in the day, with early activity onsets and late activity offsets. This was changed under free-running condition (LL(dim)), when the activity later in the subjective day (clock hour 9 and 11) was significantly greater in grey than in pied finches.Two colour morphs differed in daily activity profile, but not in the total daily activity or circadian rhythm period. Results suggest that greyzebra finches represent late chronotype, and could perhaps be better adapted to a seemingly stressful environment, such as low intensity LL(...
Comparative Biochemistry and Physiology Part A: Molecular & Integrative Physiology, 2015
Eukaryotic cells produce chemical energy in the form of ATP by oxidative phosphorylation of metab... more Eukaryotic cells produce chemical energy in the form of ATP by oxidative phosphorylation of metabolic fuels via a series of enzyme mediated biochemical reactions. We propose that the rates of these reactions are altered, as per energy needs of the seasonal metabolic states in avian migrants. To investigate this, blackheaded buntings were photoperiodically induced with non-migratory, premigratory, migratory and post-migratory phenotypes. High plasma levels of free fatty acids, citrate (an intermediate that begins the TCA cycle) and malate dehydrogenase (mdh, an enzyme involved at the end of the TCA cycle) confirmed increased availability of metabolic reserves and substrates to the TCA cycle during the premigratory and migratory states, respectively. Further, daily expression pattern of genes coding for enzymes involved in the oxidative decarboxylation of pyruvate to acetyl-CoA (pdc and pdk) and oxidative phosphorylation in the TCA cycle (cs, odgh, sdhd and mdh) was monitored in the hypothalamus and liver. Reciprocal relationship between pdc and pdk expressions conformed with the altered requirements of acetyl-CoA for the TCA cycle in different metabolic states. Except for pdk, all genes had a daily expression pattern, with high mRNA expression during the day in the premigratory/migratory phenotypes, and at night (cs, odhg, sdhd and mdh) in the nonmigratory phenotype. Differences in mRNA expression patterns of pdc, sdhd and mdh, but not of pdk, cs and odgh, between the hypothalamus and liver indicated a tissue dependent metabolism in buntings. These results suggest the adaptation of oxidative phosphorylation pathway(s) at gene levels to the seasonal alternations in metabolism in migratory songbirds.
The present study was carried out on a Palearctic-Indian migratory species, the blackheaded bunti... more The present study was carried out on a Palearctic-Indian migratory species, the blackheaded bunting (Emberiza melanocephala), to understand the importance of photoperiodism and circannual rhythms in determining seasonality in changes in body mass and testis size in birds. An initial experiment determined the effects of duration and intensity of light on photoperiodic induction. The birds were exposed to different photoperiods (hours of light:hours of darkness; 11.5L:12.5D, 12L:12D, 12.5L:11.5D and 13L:11D) at the same (~ 450 lux) light intensity, and to 13L:11D at different light intensities (50-, 100-, 400-, 800-and 1000-lux). The induction and subsequent regression of photoperiodic responses were dependent upon duration and intensity of the light period until these reached threshold. A second experiment investigated if an endogenous seasonal rhythm underlies photoperiodism in buntings. Birds maintained since February on a 8L:16D photoperiod (a non-inductive short day length invariably used to ensure photosensitivity in photoperiodic species) were subjected periodically to 16L:8D (a long day length), one group every month from mid-March to mid-August. The magnitude of long day response in body mass and testes decreased as the duration of the short days progressed, but testicular response was restored in birds that were exposed to long days in July and August. The birds exposed simultaneously to short, long, and natural day lengths for 32 weeks underwent an induction-regression cycle under long days and natural day lengths, but not under short days in which a decrease in body mass occurred after about 20 weeks. The last experiment examined the importance of latitudinal migration on photoperiodism, by comparing the response to long days of three groups which included birds from populations those were held in the outdoor aviary for 1 or 2 years at 27° N and those immediately arrived from their breeding grounds (~ 40° N). There was no difference in the photoperiodic induction among the three groups, indicating that neither experience to changing photoperiods during a migratory journey, nor to long photoperiods at breeding grounds, were critical for a subsequent response (initiationtermination-reinitiation) cycle. Taken together, these findings suggest that (1) the blackheaded bunting has its own endogenous timing program, which is regulated by the photoperiod, and (2) the photoperiodic programs of bunting are flexible enough to accommodate variations in the amplitude of environmental cycles. Thus, it appears that photoperiodism has evolved independently of the evolution of migration in this species.
Birds seasonally switch from one life history state (LHS) to another to maximize their fitness. A... more Birds seasonally switch from one life history state (LHS) to another to maximize their fitness. Accordingly, they exhibit distinct differences in their physiological and behavioral phenotypes between seasons. Possible molecular mechanisms underlying changes through the seasons have scarcely been examined in migratory birds. The present study measured key genes suggested to be involved in the metabolic regulation of 4 photoperiodically induced seasonal LHSs in a long-distance migratory songbird, the blackheaded bunting (Emberiza melanocephala). Buntings were held under short days (8 h light:16 h darkness, 8L:16D), during which they maintained the winter nonmigratory phenotype. Then they were exposed for several weeks to long days (13L:11D). Differences in the activity-rest pattern, body fattening and weight gain, testis size, organ (heart, intestine) weights, and blood glucose and triglyceride levels confirmed that buntings sequentially exhibited spring migration-linked premigratory,...
This study investigated photoperiodic plasticity in hypothalamic expression of genes implicated i... more This study investigated photoperiodic plasticity in hypothalamic expression of genes implicated in the photoperiodic light perception (rhodopsin, melanopsin, neuropsin and peropsin), transduction (pax6, bmal1, clock, per2 and casr), induction (eya3, tshβ, dio2 and dio3, gnrh and gnih) and metabolism (NPY, sirtuin1, foxO1, hmgcr, citrate synthase and dehydrogenases) in photosensitive and photorefractory redheaded buntings. There was a significant increase in eya3, tsh β, dio2, pax6 and rhodopsin and decrease in dio3 mRNA expression at hour 15 and/or 19 on the day photosensitive buntings were subjected to a 13- or 16 h, but not to 8- and 11 h light exposure. Downstream reproductive and metabolic gene expression was not altered, except for an increase in those genes coding for succinate and malate dehydrogenase enzymes involved in lipogenesis. Photorefractory buntings had high dio3 mRNA expression which significantly declined after 1 short day exposure, suggesting possible involvement ...
Proceedings of the Royal Society B: Biological Sciences, 2011
We investigated the effects of temperature on photoperiodic induction of the phenologies linked w... more We investigated the effects of temperature on photoperiodic induction of the phenologies linked with migration (body fattening and premigratory night-time restlessness, Zugunruhe ) and reproduction (testicular maturation) in the migratory blackheaded bunting. Birds were exposed for four weeks to near-threshold photoperiods required to induce testicular growth (11.5 L:12.5 D and 12 L:12 D) or for 18 weeks to a long photoperiod (13 L:11 D) at 22°C or 27°C (low) and 35°C or 40°C (high) temperatures. A significant body fattening and half-maximal testicular growth occurred in birds under the 12 L, but not under the 11.5 L photoperiod. Further, one of six birds in both temperature groups on 11.5 L, and four and two of six birds, respectively, in low- and high-temperature groups on 12 L showed the Zugunruhe . Buntings on 13 L in both temperature groups showed complete growth-regression cycles in body fattening, Zugunruhe and testis maturation. In birds on 13 L, high temperature attenuated ...
We have investigated phasic response of the photoperiodic clock to wavelength (color) and intensi... more We have investigated phasic response of the photoperiodic clock to wavelength (color) and intensity of light in the male redheaded bunting (Emberiza bruniceps). Two experiments were performed. Experiment I examined whether varying the wavelength and intensity of first (entraining, E) and/or second (inducing, I) light pulse will alter the effects of a skeleton photoperiod. Birds were subjected for a period of 6 weeks to skeleton photoschedules (6L:5D:1L:12D; 6 h E-light pulse, 1 h I-light pulse) containing either an E-pulse at 5 or 20 lx coupled with I-pulse at 100 lx, or an E-pulse at 100 lx, coupled with I-pulse at 5-or 20-lx intensity. Additional two groups that received both E-and I-pulses at 100 lx served as controls. All photoschedules were employed in two colors-white and red (654 nm). There was the wavelength-and intensity-dependent stimulation of the testis growth and development. Long wavelengths of light (red light) induced faster and greater gonadal response, but the effects were also intensity dependent. Experiment II tested whether in the photostimulated birds held on long photoperiods the change in wavelength and intensity of light hours in the morning (entraining period) or the evening (inducing period) will influence the maintenance of the photoperiodic sensitivity. Birds were subjected initially to a long photoperiod of 14L:10D (L ϭ ف 500 lx; D ϭ 0 lx) and then after 3 weeks, a 4-h light period in the morning (zeitgeber time, zt, 0-4) or in the evening (zt 10-14) of 14 L was substituted with white, green (528 nm), or red (654 nm) light at ف 20-lx intensity. One group maintained on 14L:10D and other exposed to 10L:14D served as controls. After another 7 weeks, all birds were subjected to 16L:8D for a further 4 weeks to test for their responsivity to long-day photostimulation as a consequence of exposure to different experimental photoperiods. Testes regrew under 16L:8D only in birds that were exposed to 10L:14D or to 14L:10D with a green light pulse. However, there was no effect of the timing (morning, evening) of the light pulse. Taken together, the results from both the experiments indicate that in the redheaded bunting (1) the photoperiodic clock responds differentially to different wavelengths (colors, spectra) and intensities of light, and (2) the effects of wavelength and intensity of light on the clock are phase dependent, and such phasic effects can be seen in skeleton photoperiods in which light is applied discretely at different circadian phases.
This investigation attempts to identify the mechanism (s) involved in the fat deposition in a pho... more This investigation attempts to identify the mechanism (s) involved in the fat deposition in a photoperiodic migratory species, the Blackheaded bunting (Emberiza melanocephala). Groups of photosensitive male buntings were exposed to resonance, interrupted night, ...
We investigated whether pineal is part of the circadian clock system which regulates circadian rh... more We investigated whether pineal is part of the circadian clock system which regulates circadian rhythms of activity and photosensitivity in the Indian weaver bird (Ploceus philippinus). Two experiments were performed. The first experiment examined the induction of testicular growth, and androgen-dependent beak pigmentation and luteinizing hormone (LH)-specific plumage coloration in pinealectomised (pinx) and sham-operated (sham) birds exposed to short day
Olfactory and visual sensory mechanisms seem to play a critical role in migratory orientation and... more Olfactory and visual sensory mechanisms seem to play a critical role in migratory orientation and navigation. How these two mechanisms are functionally linked with other migratory processes is unknown. We investigated this, in relation to the profound behavioural shift that occurs during migration in the night-migratory blackheaded bunting (Emberiza melanocephala). Photosensitive unstimulated birds singly housed in activity cages were subjected to long days (LD 16/8). The activity of each bird was continuously monitored. Daily activity pattern defined the nonmigratory phase (no nocturnal activity) and migratory phase (intense nocturnal activity, Zugunruhe). Body mass and testis size were measured at the beginning and end of the experiment. Long days induced the migratory phenotype (body fattening and Zugunruhe) and testis maturation. The c-fos (Fos) immunoreactivity, as marker of the neural activity of the olfactory and visual subsystems, was measured at midday (8 h after lights-on) and midnight (4 h after lights-off) after the first seven long days (nonmigratory phase) and after seven nights of the Zugunruhe (migratory phase). In the nonmigratory phase, Fos-like immunoreactive (Fos-lir) cells in the olfactory and visual subsystems were high in the day and low at night. In the migratory phase, this was reversed; Fos-lir cells were high at night and low in the day. The phase inversion of neural activity in the olfactory and visual systems in parallel with the behavioral shift suggests a functional coupling between the systems governing migratory flight (expressed as Zugunruhe) and migratory orientation and navigation.
To investigate whether the photoperiodic clocks of species possessing strongly self-sustaining ci... more To investigate whether the photoperiodic clocks of species possessing strongly self-sustaining circadian clocks share identical features, we compared the full response cycle (initiation and termination of the response) in body mass and testes of the non-migratory house sparrow (Passer domesticus) with that of the migratory redheaded bunting (Emberiza bruniceps) under Nanda-Hamner experiments. Birds were exposed to a 36 h day (L:D=6:30 h), controls exposed to a 24 h day (L:D=6:18 h), for a period of 31 weeks. By week 18 of L:D=6:18 h, there was a small increase in body mass among sparrows, but not among buntings, and the testes of bunting did not grow, while those of sparrow grew slightly. The response to L:D=6:30 h is of particular interest. There was a rapid gain and subsequent loss in the body mass of bunting, but not of sparrows. Further, both species underwent a testicular cycle as if they were exposed to long days, but the response of sparrows was slower and hence delayed the attainment of peak testicular size. Such a differential response to exotic light cycles between these two photosensitive species, despite their similar circadian oscillatory properties (strong self-sustainment), could suggest a species-specific adaptation of the endogenous clock involved in photoperiodic regulation of avian seasonality.
This study investigated the functional linkage between food availability and activity behavior in... more This study investigated the functional linkage between food availability and activity behavior in the Palaearctic Indian night migratory blackheaded bunting (Emberiza melanocephala) subjected to artificial light-dark (LD) cycles. Two experiments were performed on photosensitive birds. In the first one, birds were exposed to short days (LD 10/14; Experiment 1A), long days (LD 13/11; Experiment 1B), or increasing daylengths (8 to 13 h light/d; Experiment 1C) and presented with food either for the whole or a restricted duration of the light period. In Experiments 1A and 1B, illumination of the light and dark periods or of the dark period, alone, was changed to assess the influence of the light environment on direct and circadian responses to food cycles. In the second experiment, birds were exposed to LD 12/12 or LD 8/16 with food availability overlapping with the light (light and food presence in phase) or dark period (light and food presence in antiphase). Also, birds were subjected to constant dim light (LL dim) to examine the phase of the activity rhythms under synchronizing influence of the food cycles. Similarly, the presentation of food ad libitum (free food; FF) during an experiment examined the effects of the food-restriction regimes on activity rhythms. A continuous measurement of the activity-rest pattern was done to examine both the circadian and direct effects of the food and LD cycles. Measurement of activity at night enabled assessment of the migratory phenotype, premigratory restlessness, or Zugunruhe. The results show that (i) light masked the food effects if they were present together; (ii) birds had a higher anticipatory activity and food intake during restricted feeding conditions; and (iii) food at night alone reduced both the duration and amount of Zugunruhe as compared to food during the day alone. This suggests that food affects both the daily activity and seasonal Zugunruhe, and food cycles act as a synchronizer of circadian rhythms in the absence of dominant natural environmental synchronizers, such as the light-dark cycle.
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