Based on technique developed by the Principal Investigator that allows the muscularis mucosae (the innermost muscle layer of the intestine) from different anatomical locations to be studied in a uniform manner it has been shown that this muscle is not homogeneous. It exhibits clearly defined differences in its innervation and pharmacologic behavior along the length of the gut and in corresponding intestinal regions in different species. The proposed project utilizes three species, the North American opossum, the rabbit and the rat, to investigate potential mechanisms that may explain these observation.
Specific aim 1 is to characterize the nature of the neurotransmitters released by the nerves of the submucosal plexus innervating the muscularis mucosae in different intestinal regions. This is to test the hypothesis that disparate responses of different muscularis mucosae preparations are a function of their respective intrinsic innervation. This will be studied by the established in vitro technique of electrical field stimulation, in association with the use of selective pharmacologic agents that mimic or block the observed responses via receptor occupation or inhibition of transmitter synthesis or release.
Specific aim 2 is to comprehensively classify the receptor subtypes that mediate muscularis mucosae responses to the endogenous substances identified in specific aim 1. This is designed to test the hypothesis that differences in the pharmacologic behavior of the muscularis mucosae from different anatomical locations or species are a function of the receptor subtypes that the muscle possesses. This will be determined through the use of selective agonists and antagonists and the derivation of values such as the pA2 and the dissociation constant.
Specific aim 3 is to determine which second messengers are utilized by the muscle cells of the muscularis mucosae once endogenous compounds have interacted with the receptors identified in specific aim 2. This is to test the hypothesis that stimulation of the same receptor subtype in different preparations elicits contrasting effects because the receptor is not coupled to the same intracellular second messenger system. This will be achieved by quantitating agonist-evoked responses of the muscularis mucosae in terms of their respective phasic and tonic components and reevaluating these parameters following exposure to agents that selectively block different intracellular signal transduction pathways. These studies will, for the first time, allow region- and species-dependent differences in the characteristics of the muscularis mucosae to be quantitatively explained.
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