Studies are planned for examining the role of intrinsic neurons in regulation of secretory and absorptive function of the infant and adult human colon and in animal model. The overall objective is threefold. The first is to determine the involvement of intramural cholinergic and peptidergic neurons in regulation of colonic ion transport. The second is to identify putative transmitters that are released during neural stimulation. The third is to identify receptors for putative transmitters on epithelial membranes. The studies will be done on flat sheets of surgically resected human colon. When human tissue is not available, segments of guinea pig colon, or human T84 colonic cells grown as monolayers will be used. The approach will be to mount tissues or cell monolayers in Ussing flux chambers that are modified for electrical field stimulation of intramural neurons and to measure and evaluate changes in short-circuit currents and ionic fluxes that are evoked or modified by electrical activation of intramural nerves. Whole thickness preparations of colonic mucosa or sheets of submocosa/mocosa without the myenteric ganglia will be used to assess interactions between myenteric and submucosal ganglia in regulation of epithelial ion transport. The principal method for determining the types of nerves that influences the colonic mucosa will be pharmacological analysis of effects of agonists and antagonists of putative transmitters. Radioisotopic or radioimmunological methodologies will be used to determine whether putative neurotransmitter substances are released into the organ bath by electrical field stimulation. The presence of receptors on neurons, on epithelial cells or on both will be identified by autoradiography and ligand binding techniques. The project is expected to yield new information that will increase understanding of regulation of mucosal transport function by the enteric nervous system. It should improve understanding of abnormal secretory and absorptive processes that underlie constipation and diarrheal states in humans.
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