The long term objective of this proposal is to characterize the ion transport systems in subpopulations of apical and basolateral membranes and of intravesicular organelles of renal cells. Another long term objective is to define the factors which regulate ion transport at the cell membrane level. The hypothesis that there is a dynamic relationship between ion transporters in the cell membrane and in intracellular vesicle populations will be examined.
The specific aims of the project are: 1. To identify and characterize the transport systems for Na+, H+, K+, C1-, and other anions in renal cell membranes. 2. To evaluate the effects of in vivo conditioning (diet, acidosis, hormones, etc.) on the activity of renal cell membrane transport systems. 3. To investigate the ion transport systems in continuous cell lines of renal origin. 4. To determine the role played by intracellular pools of transpoerrts in the regulation of renal cell membrane transport systems (""""""""membrane recycling hypothesis""""""""). 5. To evaluate the nature and regulatory role of secondary sites (""""""""modifier sites"""""""") on ion transport systems which may regulate transport activity by kinetic mechanisms. Cell membrane populations will be prepared by differential centrifugation, density gradient fractionation techniques, and counter-current distribution in aqueous 2-phase systems. Membrane populations will be identified by marker enzyme determinations and morphologic studies with an electron microscope. Ion transport studies will be done with standard isotopic techniques and fluorescence methods for measuring transmembrane pH and voltage gradients. Understanding the mechanisms of ion transport and factors which regulate these systems will be an important step in furthering the overall understanding of renal function, as well as defining the nature of adaptive changes to physiologic pertubations.
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