VASOMOTOR CONTROL OF DESCENDING VASA RECTA

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DESCRIPTION: (Adapted from the Investigator's Abstract) Descending vasa recta (DVR) are resistance vessels through which blood flow to the renal medulla is supplied. Angiotensin II (ANGII) appears to have tonic effects to constrict the medullary microcirculation and stimulate medullary NO production. The PI offers preliminary data suggesting that ANGII stimulates both AT1 and AT2 receptor subtypes on DVR, that ANGII-induced constriction is mediated by a cyclooxygenase product, and that the AT1-induced constriction is abrogated by AT2 receptor activation. Moreover, the DVR endothelium appears to respond to AT1 activation by increasing intracellular [Ca] only when AT2 receptors are blocked. Finally, AT2 receptor activation appears to hyperpolarize DVR endothelium, while having no effect on membrane potential of neighboring pericytes. The following specific aims are provided: Aim 1-Utilize both RT-PCR and a pharmacological assessment of responsiveness to AT2 receptor blockers to verify expression of the AT2 receptor in DVR. Aim 2-Measure ANGII-induced intracellular [Ca] responses and NO generation by DVR and interbundle nephron segments isolated from the renal outer medulla, and the effect of dietary salt loading on these responses. Aim 3-Utilize membrane permeant cyclic nucleotide analogues and guanylate or adenylate cyclase inhibitors to test the hypothesis that blockade of AT1-mediated calcium signaling in DVR endothelia by AT2 receptor activation involves cyclic nucleotides. The role of intracellular phosphorylation events and tyrosine phosphatase or kinase activity in this phenomenon will also be examined. Aim 4-Utilize patch clamp and Mn-quench of fura2 to examine the ability of AT1 and AT2 receptor activation to modulate endothelial Ca influx. Aim 5-Test the hypothesis that DVR pericytes lack L-type voltage operated Ca channels by examining the ability of nifedipine to block vasoconstriction, the effect of ANGII on pericyte membrane potential, and the effect of membrane depolarization on whole call Ca currents. Additional experiments will test the hypothesis that AT1 and AT2 stimulation modulates pericyte intracellular [Ca]. Finally, the PI hypothesizes that ANGII constricts OMDVR via thromboxane generation. This postulate will be tested by examining the ability of a PGH2-TxA2 receptor antagonist to block ANGII-induced vasoconstriction, the ability of a TxA2 analogue to cause vasoconstriction, and the ability of ANGII to stimulate TxB2 production in isolated vessels.