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Activation of neuronal P2X7 receptor–pannexin-1 mediates death of enteric neurons during colitis

Nature Medicine volume 18, pages 600–604 (2012)Cite this article

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Abstract

Inflammatory bowel diseases (IBDs) are chronic relapsing and remitting conditions associated with long-term gut dysfunction resulting from alterations to the enteric nervous system and a loss of enteric neurons1,2. The mechanisms underlying inflammation-induced enteric neuron death are unknown. Here using in vivo models of experimental colitis we report that inflammation causes enteric neuron death by activating a neuronal signaling complex composed of P2X7 receptors (P2X7Rs), pannexin-1 (Panx1) channels, the Asc adaptor protein and caspases. Inhibition of P2X7R, Panx1, Asc or caspase activity prevented inflammation-induced neuron cell death. Preservation of enteric neurons by inhibiting Panx1 in vivo prevented the onset of inflammation-induced colonic motor dysfunction. Panx1 expression was reduced in Crohn's disease but not ulcerative colitis. We conclude that activation of neuronal Panx1 underlies neuron death and the subsequent development of abnormal gut motility in IBD. Targeting Panx1 represents a new neuroprotective strategy to ameliorate the progression of IBD-associated dysmotility.

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Figure 1: P2X7R activation is necessary and sufficient for enteric neuron death.
Figure 2: Inflammation-induced enteric neuron death requires Panx1 and Asc but not Nlrp3.
Figure 3: Inhibiting neuronal Panx1 during colitis protects against post-inflammation deficits in inhibitory neuromuscular transmission.
Figure 4: Panx1 inhibition is neuroprotective in a recurrent model of colitis, and human enteric neuron Panx1 expression changes are observed in Crohn's disease.

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Acknowledgements

This work was supported by grants from the Canadian Institutes of Health Research (CIHR, to K.A.S., R.J.T. and D.M.M.), the Crohn's & Colitis Foundation of Canada (CCFC to D.M.M.) and US National Institutes of Health grant DK62267 (to G.M.M.). Some of the equipment used in the study was provided by funds from the Canadian Foundation for Innovation and the Alberta Science and Research Authority. B.D.G. holds fellowships from the Canadian Association of Gastroenterology/CIHR and Alberta Innovates-Health Solutions (AI-HS)/CCFC. K.A.S. is an AI-HS Medical Scientist and holds the CCFC Chair in IBD Research at the University of Calgary. R.J.T. is an AI-HS Scholar. P.L.B. is supported by AI-HS, CIHR and CCFC. J.A.M. is an AI-HS Senior Scholar and Canada Research Chair. D.M.M. is an AI-HS Scientist and holds a Canada Research Chair (Tier 1). S.A.H. holds fellowships from CIHR and AI-HS. We thank C. MacNaughton, W. Ho and A. Wang for technical support, D.M. McCafferty (University of Calgary) for providing Il10−/− mice, V. Dixit (Genentech) and J. Tschopp (University of Lausanne) for providing male Pycard−/− and Nlrp3–/– mice, respectively, N. Hyman (University of Vermont), O. Bathe (University of Calgary) and the IBD Tissue Bank for providing human tissue samples and J. Bains for reviewing and commenting on the manuscript.

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Authors and Affiliations

  1. Hotchkiss Brain Institute, University of Calgary, Calgary, Alberta, Canada

    Brian D Gulbransen, Mohammad Bashashati, Roger J Thompson & Keith A Sharkey

  2. Department of Physiology and Pharmacology, University of Calgary, Calgary, Alberta, Canada

    Brian D Gulbransen, Mohammad Bashashati, Derek M McKay & Keith A Sharkey

  3. Snyder Institute of Infection, Immunity and Inflammation, University of Calgary, Calgary, Alberta, Canada

    Brian D Gulbransen, Mohammad Bashashati, Simon A Hirota, Justin A MacDonald, Daniel A Muruve, Derek M McKay, Paul L Beck & Keith A Sharkey

  4. Department of Medicine, University of Calgary, Calgary, Alberta, Canada

    Simon A Hirota, Daniel A Muruve & Paul L Beck

  5. Libin Cardiovascular Institute, University of Calgary, Calgary, Alberta, Canada

    Simon A Hirota & Justin A MacDonald

  6. Department of Biochemistry and Molecular Biology, University of Calgary, Calgary, Alberta, Canada

    Simon A Hirota & Justin A MacDonald

  7. Department of Pathology and Laboratory Medicine, University of Calgary, Calgary, Alberta, Canada

    Xianyong Gui

  8. Department of Anatomy & Neurobiology, University of Vermont College of Medicine, Burlington, Vermont, USA

    Jane A Roberts & Gary M Mawe

  9. Department of Cell Biology and Anatomy, University of Calgary, Calgary, Alberta, Canada

    Roger J Thompson

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Contributions

Overall project design and hypotheses were developed by B.D.G. under the supervision of R.J.T. and K.A.S. B.D.G. coordinated the project, conducted all experiments unless otherwise noted, analyzed the data, prepared figures and wrote the manuscript. M.B. carried out the colon contractility experiments. P.L.B., X.G., S.A.H., J.A.R., J.A.M., D.A.M., D.M.M. and G.M.M. contributed to experimental design and prepared and provided mice, tissues and reagents. All authors participated in revising the manuscript and agreed to the final version. R.J.T. and K.A.S. supervised the overall project.

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Correspondence to Brian D Gulbransen or Keith A Sharkey.

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The authors declare no competing financial interests.

Supplementary information

Supplementary Text and Figures

Supplementary Figures 1–7, Supplementary Table 1 and Supplementary Methods (PDF 83820 kb)

Supplementary Video 1

A myenteric ganglia containing enteric neurons and glia (loaded with the Ca2+ indicator Fluo4) is challenged with the P2X7R agonist BzATP. Note how the response begins in neurons, followed closely by glia. (AVI 1205 kb)

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Gulbransen, B., Bashashati, M., Hirota, S. et al. Activation of neuronal P2X7 receptor–pannexin-1 mediates death of enteric neurons during colitis. Nat Med 18, 600–604 (2012). https://doi.org/10.1038/nm.2679

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