ROLE OF ALTERED MEMBRANE FUNCTION IN XENOBIOTIC TOXICITY

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Renal secretory systems for organic anions (OA) and cations (OC) govern the elimination of most small (<500mol.wt.) foreign chemicals. We have previously shown that OA transport is indirectly coupled to metabolic energy through Na/alpha-ketoglutarate (alphaKG) co-transport and OA/alphaKG exchange. We are currently examining a) plasma membrane and intracellular events associated with secretory transport (in collaboration with D.S. Miller), b) molecular biology of these systems, c) energetics and mechanisms of extrarenal OA and OC transport, and d) toxicology of OA and OC in kidney and extrarenal sites. The choroid plexus mediates transport of potentially toxic OA and OC across the blood-CSF barrier into the blood for subsequent elimination by kidney or liver. We have developed a monolayer cultured plexus preparation which actively transports OA and OC in vitro. In addition to flux studies which have characterized the basic transport properties of this epithelium, we have shown mediated uptake of both classes of xenobiotics into intracellular vesicles. OC containing vesicles were shown to move to the basolateral side of the cell (blood side) and release their contents, indicating direct participation of vesicular movement in transepithelial flux - the first such demonstration in a secretory epithelium. Vesicular release of OC at the basolateral face of the cell was reversibly disrupted by depolimerization of microtubules with nocodazole. We have also demonstrated that the rate and effectiveness of secretory transport of OA by intact renal tubular cells was determined by the intracellular concentration and gradient of alphaKG across the basolateral membrane. In turn the cytoplasmic alphaKG concentration was determined by its intracellular compartmentation (primarily into mitochondria) and metabolsim. Finally, expression cloning and PCR probes have been utilized to screen a cDNA library prepared from rat kidney mRNA. These studies are preliminary to identification and cloning of the genes coding for these two important excretory transport systems.