Ouabain, Local Ca2+ Control and Myogenic Tone

大多数盐依赖型高血压大鼠模型都与血浆内源性哇巴因(一种肾上腺皮质激素)水平升高有关，而长期服用哇巴因会导致啮齿动物高血压。这和其他证据表明，哇巴因诱导的高血压(OH)是盐依赖型高血压的一个很好的模型，但从哇巴因到高血压的步骤尚不清楚。本项目提出的假设是，具有高哇巴因亲和力的特定动脉平滑肌Na+泵和Na/Ca交换器(NCX)在将哇巴因对这些Na+泵的抑制作用转化为对细胞内钙离子的调节以及对动脉收缩和肌源性张力的控制中发挥关键作用。四个具体目标是：1)表征由商店操作的通道(SOCs)和通过 NCX在小动脉中的作用，并确定这些转运体如何参与钙稳态和肌源性张力的控制。2)验证遗传或药物降低的Na+活性泵入α2(而不是α1)亚基(这两种异构体在肠系膜动脉肌细胞中表达)调节小动脉细胞内钙离子和肌源性张力的假说。我们探讨了某些具有降压活性的药物可能干扰哇巴因对这些钠泵α2亚单位的作用的想法。3)验证平滑肌NCX介导α2的Na+泵活性降低效应的假设 胞浆钙和肌源性张力的亚基。我们探讨了哇巴因的这些作用可以被具有抗高血压活性的药物阻断NCX的想法。4)验证哇巴因体内慢性给药(表现为OH)和体外急性给药在控制小动脉收缩和肌源性张力机制上的相似作用。大鼠和小鼠加压肠系膜小动脉负载钙、钠指示剂，同时研究离子浓度(共聚焦显微镜和宽场显微镜)、膜电位和肌源性张力的变化。具有改变的Na+泵和NCX基因的转基因小鼠，以及直接阻断哇巴因作用或选择性阻断NCX的新型抗高血压药物，将被用于识别从哇巴因到动脉收缩能力改变的序列中的特定步骤。这些结果将提高人们的理解 研究盐依赖型高血压的发病机制，并将为创新治疗找到新的靶点。

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.