Studies are proposed to further identify ion transport processes and the factors that regulate them in guinea pig gallbladder epithelial cells. They will initially focus ont he characterization and regulation of influx and efflux pathways for potassium (K+) and bicarbonate (HCO3-). We have postulated that two distinct K+ efflux pathways exist in the apical membrane of guinea pig gallbladder cells based upon the differential sensitivity of transepithelial K+ fluxes to the k+ channel blockers, tetraethylammonium (TEA+) and Charybdotoxin (CHTX). In addition, a quinidine-sensitive, TEA-insensitive pathway has been observed at the basolateral membrane. Our studies will combine transepithelial fluxes, conventional electrophysiology and patch clamp techniques to distinguish the individual K+ efflux pathways underlying these conductances and their physiologic roles. The regulation of K_ transport by cyclic nucleotides, intracellular calcium, and intracellular metabolites such as ATP will be investigated from the effects of the exogenous addition of these agents and appropriate inhibitors on membrane voltage and conductance and single channel characteristics. Measurements of changes in the intracellular concentrations in Ca2+, cAMP and ATP will be correlated with changes in K+ transport. Experiments are also proposed to further define he HCO3- entry step at the basolateral membrane essential for PGE2-stimulated HCO3- secretion. This process appears to be mediate by an electrogenic NaHCO3-cotransproter. Conventional electrophysiologic techniques will be used to define the Na+ and HCO3- dependency of the process, its electrogeneity and its contribution to basolateral membrane conductance. We will initiate studies to determine the intracellular factors regulating its activity from measurements of intracellular pH in cells loaded with fluorescent dyes. The results of these studies will lead to an enhanced understanding of ion transport by the mammalian gallbladder and will be applicable to a variety of other leaky epithelia.
|Bayse, Gladys S; Hammonds-Odie, Latanya P; Jackson, Kimberly M et al. (2013) Permeation of roxarsone and its metabolites increases caco-2 cell proliferation. Adv Biol Chem 3:389-396|