Project Summary/Abstract Our current understanding of mechanisms underlying visceral hypersensitivity, such as that associated with irritable bowel syndrome (IBS), remains rudimentary. Importantly, treating this and other functional gut disorders is limited, with a clear need for alternative treatment options. For the growing population afflicted by IBS, GI hypersensitivity and pain persist long after signs of tissue injury have resolved. Unlike other intestinal disorders, patients with IBS are hypersensitive with a lower pain threshold following colonic rectal distention (CRD) testing. Identifying the molecular and cellular components that mediate both the acute and persistent phases of visceral pain is a critical first step in understanding how environmental and endogenous factors produce long-term changes in the nervous system or associated tissues to engender chronic pain syndromes. In this new multi-PI application, we have taken a team-science approach and a multifaceted strategy designed to maximize the relevance of our pre-clinical basic research discoveries. Enterochromaffin (EC) cells are key sensory cells in the intestinal epithelium that release serotonin onto primary sensory nerve fibers, thereby evoking a sensation of discomfort and pain in response to luminal irritants, such as bacterial metabolites, inflammatory agents, or ingested chemicals. Our group recently established that EC cell-mucosal afferent signaling is a major mediator of visceral pain. We also show that the strength of this signal differs in males versus females. We will leverage our new colitis-free chemogenetic model of visceral hypersensitivity to zero in on the contribution of EC cells to visceral pain and identify molecular mechanisms through which these cells modulate the activity of nearby sensory nerve fibers. We will also ask how estrogen signaling contributes to the strong female bias that is characteristic of human IBS. Our team brings expertise in neurophysiology, pharmacology, visceral tissue anatomy, sex differences, and hormone signaling in female physiology and an unusually wide-ranging set of innovative approaches to tackle a prevalent gut-brain disorder.