Brainstem nAChR Subtypes & Their Effect on GI Function

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DESCRIPTION (provided by applicant): Administration of nicotine and/or smoking has been shown to have important effects on gastric function. Evidence of others and our own data strongly indicate that these effects are due to nicotine acting on neuronal nicotinic acetylcholine receptors (nAChRs) in the dorsal motor nucleus of the vagus (DMV) and medial subnucleus of the tractus solitarius (mNTS). The goals of our proposed studies are to: (i) investigate the influence of DMV and mNTS nAChRs in the effects of systemically administered nicotine on gastric function and (ii) to use DMV and mNTS nAChRs as model systems to learn more about native CNS nicotinic receptors, their nature, their function and their regulation. The Specific Aims of our research are to: (i) determine the subunit composition of nAChRs in the rat DMV and mNTS that mediate the effects of nicotine on gastric tone and motility; (ii) examine regulation of the function and density of nAChRs in rat DMV and mNTS after chronic administration of nAChR-stimulating doses of nicotine, and the agonist-induced responsiveness of nAChRs in the DMV and mNTS after acute exposure to sub-activating doses of nicotine; and (iii) test the hypothesis that nAChRs observed to exist in the DMV and mNTS as revealed by pursuing Specific Aims 1 & 2 play an important role in the gastric effects (DMV and mNTS) and cardiovascular effects (mNTS) of systemically administered nicotine. To achieve these Specific Aims, a focused but multidisciplinary approach will be used consisting of the following techniques: (i) pharmacological tools applied either in vivo by microinjection of drugs into the DMV and mNTS of anesthetized rats while monitoring gastric tone and motility, or in vitro to rat brain slice preparations containing the DMV and mNTS while monitoring electrophysiological events using the whole-cell patch-clamp recording method; (ii) in vitro autoradiography taking advantage of the high affinity epibatidine binding to nAChRs combined with selective denervation of efferent and afferent vagal fibers; (iii) immunofluorescent identification of nAChR subtypes in DMV and mNTS neurons; (v) fluorescent in situ hybridization technique to identify nAChR subunits associated with DMV neurons projecting to the fundus and antrum, (vi) knockout mice, in which one gene of interest is silenced, and the contribution of a particular nAChR subunit (e.g., a7 subunit) in eliciting a specific gastric response can be assessed. Findings made to date suggest that: (I) there are five different nAChRs in the DMV and mNTS that influence gastric function; (ii) there might be a viscerotopic organization of nAChR subunits on DMV motoneurons. That is a7 containing nAChRs might be located primarily on DMV neurons projecting to the fundus but not on DMV neurons projecting to the antrum; and (iii) iv. administered nicotine might act in the mNTS on a ct4B2-nAChR subtype to inhibit tonic contraction of the fundus. These and future data obtained from pursuing our Specific Aims will extend current understanding of the different receptor mediated effects of nicotine on parasympathetic nervous system control of the upper GI tract.