This project's goal is to so define the biologic importance of cytochrome P-450 NADPH-dependent arachidonate metabolites in the mammalian kidney. Arachidonic acid metabolites from this pathway affect peptide hormone secretion, platelet aggregation, smooth muscle relaxation, and epithelial sodium transport. The kidney produces large quantities of P-450 metabolites of arachidonic acid. To achieve our overall goal, we propose to test four hypotheses: (1) An endogenous cytochrome P-450 metabolite of arachidonic acid is a potent inhibitor of volume absorption in rabbit proximal tubule; (2) There is a functional link between organic ion secretion and cytochrome P-450 metabolism of arachidonic acid in renal proximal straight tubule; (3) Cytochrome P-450 metabolite(s) of arachidonic acid produced in medullary thick ascending limb regulate ion transport in this nephron segment; (4) The cortical collecting tubule water permeability response to arginine vasopressin is blunted by arachidonic acid metabolites of cytochrome P-450 via a mechanism different than that observed with PGE2. We will conduct studies using in vitro microperfusion of rabbit nephron segments coupled with studies on the segmental metabolism of arachidonic acid by the P-450 pathway. In proximal convoluted and proximal straight tubules we will measure the effects of exogenously administered P-450 metabolites of arachidonic acid on volume absorption and organic ion transport. Specific inhibitors of P-450 pathway and induction of cytochrome P-450 will also be examined. The mechanism of action of these metabolites on transport will be defined utilizing microelectrode techniques and selective inhibitors or certain signal transduction pathways. The effects of P-450 metabolites on medullary thick ascending limb chloride transport will be tested as will the pattern of P-450 metabolites produced by medullary thick ascending limbs. Should important effects on transport be observed, the molecular mechanism and the signal transduction pathway used by these P-450 metabolites will be tested. Finally, we will extend our previous studies by determining if the P-450 epoxides and diols blunt arginine vasopressin's actin through a mechanism that differs from prostaglandin E2. The mechanism of the inhibition will be determined. We feel that the hypotheses being tested represent a focused and well founded group of experiments to test the biologic importance of the P-450 arachidonic acid metabolites in the kidney.
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