P009 IL-17 mediates visceral hypersensitivity by sensitising colonic nociceptors

Abstract Background Comparison of cytokine expression in colitic tissue with respective cytokine receptor expression in pain sensing neurons highlights a likely contribution of IL-17 to the production of abdominal pain during flare. To explore this pro-nociceptive potential of IL-17 in colitis we examined the ability of IL-17 to stimulate sensory neurons and the colonic afferent response to noxious distension. Methods IL-17 mediated sensory neuron activation: changes in the intracellular [Ca2+]i within thoracolumbar (T12-L5 spinal segments) dorsal root ganglia sensory neurons were examined in cells cultured overnight and loaded with Fluo-4-AM (30min) prior to imaging. Responses were measured to IL-17 (10-300 ng/mL) alone and following pre-treatment with either TRPV1 or TRPA1 inhibitors. Afterwards cells were exposed to capsaicin (1 µM) followed by 50 mM KCl, to confirm the presence of nociceptors and neurons respectively by positive response. In a separate set of experiments the effect of IL-17 was also examine on a neuron pure cell culture in which non-neuronal cells were removes by magnet assisted cell sorting. IL-17 mediated colonic nociceptor activation: electrophysiological recordings were made of lumbar splanchnic nerve activity and the response to noxious luminal distension of the isolated colorectum (ex-vivo), perfused with carbogenated Krebs solution, examined following pre-treatment with IL-17 (50 ng/mL) alone or in the presence of respective inhibitors. All experiments were performed using tissue from male C57BL/6J mice euthanaised by rising concentration of CO2followed by exsanguination in accordance with Schedule 1 of the UK Animals Scientific Procedures act (1986). Results Consistent with the ability of IL-17 to stimulate sensory neurons including those classified as nociceptors by their co-sensitivity to capsaicin, pre-treatment with IL-17 also produced a significant sensitisation of the colonic afferent response to luminal distension at noxious distending pressures. The increase in [Ca2+]i produced by IL-17 in sensory neurons was blocked by pre-treament with a TRPV1 antagonist but not a TRPA1 antagonist. Furthermore, the proportion of neurons stimulated by IL-17 was comparable between standard DRG cell cultures and neuron pure cell cultures demonstrating a direct effect of IL-17 on sensory neurons. Finally, IL-17 mediated mechanosensitisation of colonic nociceptors was unaffected by TRPV1 inhibition. Conclusion Our findings demonstrate that IL-17 stimulates capsaicin-sensitive nociceptors and mediates visceral hypersensitivity in colonic nociceptors consistent with a causative role for IL-17 is the generation of abdominal pain in people with colitis.