Ouabain, Local Ca2+ Control and Myogenic Tone

大多数盐依赖性高血压大鼠模型与内源性哇巴因（一种肾上腺皮质激素）的血浆水平升高相关，长期给予哇巴因可诱导啮齿动物高血压。这和其他证据表明，哇巴因诱导的高血压（OH）是盐依赖性高血压的良好模型，但哇巴因导致高血压的步骤尚不清楚。本项目提出了一个假说，即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胞浆Ca 2+的调节以及对动脉收缩性和肌源性张力的控制中起关键作用。本研究的四个具体目的是：1）研究钙池操纵通道（SOC）介导的Ca ~（2+）（和Na ~+）内流， NCX在小动脉中的作用，并确定这些转运体如何参与Ca 2+稳态和肌张力的控制。2)为了验证以下假设，即具有α 2（而非α 1）亚单位（在肠系膜动脉肌细胞中表达的两种亚型）的Na+泵的遗传性或非遗传性降低的活性调节小动脉中的细胞内Ca 2+和肌源性张力。我们探讨的想法，某些药物的抗高血压活性可能会干扰哇巴因的行动，这些Na+泵α 2亚基。3)为了检验平滑肌NCX介导Na +泵活性降低对α 2的影响的假设， 亚基对胞浆Ca 2+和肌源性张力的影响。我们探讨的想法，哇巴因的这些影响可以废除的药物具有抗高血压活性，阻断NCX。4)为了检验以下假设：体内长期给予哇巴因（表现为OH）和体外急性给予哇巴因对控制小动脉中动脉收缩性和肌源性张力的机制具有相似的作用。大鼠和小鼠加压小肠系膜动脉加载Ca 2+和Na +指示剂将用于研究离子浓度变化（共聚焦和宽视野显微镜），膜电位和肌张力同时。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，将用于鉴定从哇巴因到改变的动脉收缩性的序列中的特定步骤。这些结果将增进对 盐依赖性高血压的发病机制，并将为创新治疗确定新的目标。

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.