Vascular Calcium Channel Expression in Hypertension

描述(申请人提供)：L型钙离子通道是一种多蛋白质复合体，包括一个成孔的11C亚基和较小的辅助亚基。辅助2亚基促进11C亚基在细胞膜表面的表达，从而增加功能钙通道的数量。在血管平滑肌细胞(VSMCs)中，钙离子通道仅有少量表达，以严格调节电压门控钙离子内流和血管收缩。然而，在高血压的发展过程中，我们已经表明，钙通道在VSMC中深刻上调，以刺激异常的血管收缩。这一修订项目的目标是确定血压升高时钙通道上调的机制。我们注意到，血管紧张素(Ang II)高血压小鼠的肠系膜循环中有一个特殊的2亚单位，23，选择性地和深刻地增加。由此产生的钙通道11C23复合体的过度丰富导致了小动脉中钙离子内流的增加和钙离子依赖性张力的异常。事实上，钙通道的药理阻断在体内显著降低了Ang II高血压小鼠的血压，但在对照组小鼠中几乎没有降压作用，提示钙通道在高血压的发病机制中起着中心作用。在审查者的要求下，这项修订的应用程序密切关注于描绘高血压小鼠血管钙通道异常的机制，以利用基因缺失模型。目的1确定钙通道11C23复合体的数量增加是否与两种高血压模型小鼠的VSMCs内钙异常内流有关。目的2将使用23个基因敲除小鼠来确定23个亚基是否是钙通道上调和高血压发展的必要贡献者。我们预测血管钙通道11C23复合体将不能对血管紧张素转换酶II或去甲肾上腺素的注射作出反应，并且高血压的发展将被严重钝化。最后，Aim 3将利用我们开发的一种新的微血管分析方法来监测单个加压小鼠肠系膜动脉VSMCs中钙通道的表达。使用这一独特的检测方法，我们将直接检验这一假说，即高腔内压抑制钙通道的周转，从而在高血压期间增加它们在VSMC表面膜上的表达。与公共健康相关：6000万美国人患有高血压，这会导致心脏病、肾脏疾病和中风。这项研究项目将确定血管肌肉细胞中的特定分子是否需要膜蛋白的异常出现，从而导致动脉过度收缩，从而使血压上升到危险水平。

DESCRIPTION (provided by applicant): The L-type Ca2+ (CaL) channels are multi-protein complexes that include a pore- forming 11C subunit and smaller ancillary subunits. The ancillary 2 subunits promote the expression of 11C subunits at the surface membrane to increase the number of functional CaL channels. In vascular smooth muscle cells (VSMCs), CaL channels are only sparsely expressed in order to tightly regulate voltage-gated Ca2+ influx and vascular contraction. However, during the development of hypertension, we have shown that CaL channels profoundly upregulate in the VSMCs to fuel abnormal vasoconstriction. The goal of this revised project is to determine the mechanism of CaL channel upregulation in response to rises in blood pressure. We have noted that a specific 2 subunit, 23, selectively and profoundly increases in the mesenteric circulation of angiotensin (Ang II) hypertensive mice. The resulting overabundance of CaL channel 11C23 complexes results in elevated Ca2+ influx and abnormal Ca2+-dependent tone in the small arteries of the affected animals. In fact, pharmacological block of CaL channels sharply reduces blood pressure in Ang II hypertensive mice in vivo, but has little antihypertensive effect in control mice, suggesting a central contribution of CaL channels to the pathogenesis of hypertension. At the reviewers' behest, this revised application is tightly focused on delineated the mechanism of vascular CaL channel abnormalities in hypertensive mice to take advantage of gene deletion models. Aim 1 will determine if an increased number of CaL channel 11C23 complexes is associated with anomalous Ca2+ influx in VSMCs of two mouse models of hypertension. Aim 2 will use 23 knockout mice to determine if the 23 subunit is a requisite contributor to CaL channel upregulation and the development of hypertension. We predict that vascular CaL channel 11C23 complexes will fail to upregulate in response to Ang II or norepinephrine infusion in 23 knockout mice, and that the development of hypertension will be severely blunted. Finally, Aim 3 will utilize a novel microvascular assay that we have developed to monitor CaL channel expression in the VSMCs of single, pressurized mouse mesenteric arteries. Using this unique assay, we will directly test the hypothesis that high intraluminal pressure inhibits the turnover of CaL channels, thereby increasing their expression at the VSMC surface membrane during hypertension. PUBLIC HEALTH RELEVANCE: Sixty million Americans have high blood pressure, which leads to heart attacks, kidney disease and stroke. This research project will determine if a specific molecule in the muscle cells of blood vessels is required for the abnormal appearance of membrane proteins that causes arteries to contract too much, thereby elevating blood pressure to dangerous levels.