Ouabain, Local Ca2+ Control and Myogenic Tone

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Most rat models of salt-dependent hypertension are associated with elevated plasma levels of endogenous ouabain (an adrenocortical hormone), and chronic ouabain administration induces hypertensio n in rodents. This and other evidence suggests that ouabain-induced hypertension (OH) is a good model for salt-dependent hypertension, but the steps leading from ouabain to high blood pressure are Unknown. This project addresses the hypothesis that specific arterial smooth muscle Na + pumps with high ouabain affinity and the Na/Ca exchanger (NCX) play key roles in translating ouabain's inhibitory action on these Na+ pumps into modulation of cytosolic Ca 2+ and control of arterial contractility and myogenic tone. The four Specific Aims are: 1) To characterize the Ca 2+ (and Na+) entry mediated by store-operated channels (SOCs) and by NCX in small arteries, and to determine how these transporters participate in Ca 2+ homeostasis and the control of myogenic tone. 2) To test the hypothesis that genetically or pharmacologically reduced activity of Na+ pumps with alpha2 (but not alpha1) subunits (the two isoforms expressed in mesenteric artery myocytes) modulates intracellular Ca 2+ and myogenic tone in small arteries. We explore the idea that certain agents with anti-hypertensive activity may interfere with ouabain's action on these Na+ pump alpha2 subunits. 3) To test the hypothesis that smooth muscle NCX mediates the effects of reduced activity of Na + pumps with alpha2 subunits on cytosolic Ca 2+ and myogenic tone. We explore the idea that these effects of ouabain can be abrogated by agents with antihypertensive activity that block NCX. 4) To test the hypothesis that chronic in vivo ouabain administration (manifested as OH) and acute in vitro ouabain administration have similar effects on the mechanisms that control arterial contractility and myogenic tone in small arteries. Rat and mouse pressurized small mesenteric arteries loaded with Ca 2+ and Na + indicators will be used to study ion concentration changes (confocal and widefield microscopy), membrane potential and myogenic tone simultaneously. Transgenic mice with altered Na + pump and NCX genes, and novel anti-hypertensive agents that directly block ouabain's action or that selectively block NCX, will be used to identify specific steps in the sequence from ouabain to altered arterial contractility. These results will improve understanding of the pathogenesis of salt-dependent hypertension and will pinpoint new targets for innovative therapy.