MOLECULAR AND FUNCTIONAL ANALYSIS OF COLONIC IONIC CONDUCTANCES

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Regulation of colonic motility is intimately coordinated with the modulation of ionic conductances expressed in colonic smooth muscle and non-muscle cells. The long-term goal of this project is to gain a more complete understanding of the role of ionic conductances in regulating colonic motility. The molecular identification of the components for these conductances will aid in the analysis of their modulation and receptor mediated regulation. In addition, this information will provide a foundation for the identification or development of therapeutics that might act on these channels. It is much easier to study these conductances and develop blocking drugs in expression systems than in native GI muscle cells. This project will focus on the relationship between ionic currents in native colon cells and their molecular counterparts. We will use the information gained from determining this relationship to study: (1) the role of specific ion channels in colonic electrical activity, (2) how these channels are regulated in response to neuronal and hormonal input, (3) the cellular and subcellular distribution of ion channels in the colon. The experiments are based upon several important advances in the molecular identification of colonic ion channels made during the previous funding period. We will address several hypotheses in this study. One aim focuses on the relationship between cloned Kv channels and native colonic electrical activity. We also propose that members of the trp gene family encode the colonic non-selective cation current, central to excitatory neurotransmission in the colon. We will test the hypotheses that Kir3.1/3.2 encodes I/KACh in the colon, contributing a hyperpolarizing influence on the excitatory response. We will determine whether colonic ion channels can be targets on regulatory pathways in order to make testable predictions for native cells and tissues. Finally, we will determine the cellular and subcellular distribution of several K+ channels in colon muscle and non-muscle cells.