Principal Investigator: NANCY WILLS
Abstract: The long term goal of this project is to understand how sodium and potassium are transported across epithelia. Two of the most important sites for the regulation of plasma K+ and Na+ levels are the renal collecting duct system and the colon. The present study proposes to use 2 model epithelial systems to investigate the regulation of Na+ and K+ transport: 1) the A6 cell line from toad kidney, and 2) primary cell cultures of rabbit distal tubule. These epithelia both absorb Na+ by mineralocorticoid-sensitive mechanisms, similar to those in other so-called tight epithelia. In addition rabbit distal tubule cells possess well-studied K+- transport mechanisms. The focus of the project is the regulation of apical and basolateral ionic channels by aldosterone, cyclic nucleotides, and other factors. Electrophysiological methods including conventional and ion-sensitive microelectrode methods, impedance analysis, current fluctuation analysis and patch clamp techniques will be used. From these methods new information will be obtained concerning membrane potentials, conductances and areas as well as single channel properties. Using this information, it will be possible to determine how these channels are regulated, e.g. by changes in channel kinetics, alteration of open-channel conductance or changes in membrane area and the number of conducting channels. The first specific aim is to characterize membrane conductances, areas, and Na+ channel properties in cultured A6 epithelia. The following questions will be asked: 1) Do aldosterone and anti-diuretic hormone activate the same population of Na+ channels? 2) What factors regulate the basolateral membrane potassium conductance? 3)What are the long term effects of aldosterone on membrane conductances and areas? 4) After removal of hormonal stimulation how are epithelial channels "deactivated"? The second objective is to characterize apical membrane K+ channels in renal distal tubule cells. Specifically, the following questions will be asked: 1) Does aldosterone activate a distinct population of K+ channels or does it increase K+ conductance by modifying channel-gating or other properties. 2) What is the time course of hormonal stimulation of this K+ conductance and is it downregulated or upregulated in relation to Na+ absorption? The results will provide important quantitative information at the single-membrane and channel level concerning the effects of chronic hormonal stimulation on epithelial membrane properties.
Funding Period: 1987-01-01 - 1995-12-31
more information: NIH RePORT