Persistent Calcium Sparklets in Vascular Smooth Muscle

描述（由申请方提供）：通过二氢吡啶敏感性、电压门控L型钙通道的钙内流在动脉平滑肌兴奋性、收缩和基因表达的调节中起关键作用。我们最近发现，L型钙通道的小簇可以在“持续”门控模式下工作，在平滑肌中产生几乎连续的钙内流位点（称为“持续钙火花”），这导致了范式转变，其中这些细胞中的钙内流主要由该过程结合单个L型钙通道的罕见电压依赖性开放控制。然而，在生理和病理生理条件下，持续性钙火花对局部和全局细胞内钙的调节以及这些钙内流事件的激活和调节的分子机制的作用几乎是未知的。这项工作将使用我们小组开发的新方法来确定钙火花在脑动脉平滑肌中的生物物理特性和功能作用。我们的初步结果表明，所有提出的实验是可行的，并将提供重要的新信息。这项工作旨在测试三个新的假设。在具体目标1中，我们将检验通过钙火花的钙内流有助于局部和整体细胞内钙浓度变化的假设。具体目标2中的实验将检验Cav1.2通道是动脉平滑肌中钙火花的基础的假设。最后，在具体目标3中，我们建立在前两个具体目标的工作基础上，并测试了高血压期间持续钙火花活动增加的假设。这项工作应该提供钙火花介导的信号传导及其在调节脑动脉功能中的作用的第一个综合观点，并显着提高我们对健康和疾病中动脉功能的理解。

DESCRIPTION (provided by applicant): Calcium influx via dihydropyridine-sensitive, voltage-gated L-type calcium channels plays a crucial role in the regulation of excitability, contraction, and gene expression in arterial smooth muscle. Our recent discovery that small clusters of L-type calcium channels can operate in a "persistent" gating mode that create sites of nearly continual calcium influx (called "persistent calcium sparklets") in smooth muscle has led to a paradigm shift, whereby calcium influx in these cells is predominantly controlled by this process in combination with rare voltage-dependent openings of individual L-type calcium channels. However, the role of persistent calcium sparklets on the regulation of local and global intracellular calcium as well as the molecular mechanisms underlying the activation and modulation of these calcium influx events in smooth muscle under physiological and pathophysiological conditions is virtually unknown. The proposed work will use new methods developed by our group to define the biophysical properties and functional roles of calcium sparklets in cerebral artery smooth muscle. Our preliminary results suggest that all proposed experiments are feasible and will provide important new information. The proposed work seeks to test three novel hypotheses. In Specific Aim 1, we will test the hypothesis that calcium influx via calcium sparklets contributes to changes in local and global intracellular calcium concentration. The experiments in Specific Aim 2 will test the hypothesis that Cav1.2 channels underlie calcium sparklets in arterial smooth muscle. Finally, in Specific Aim 3, we build on the work in the previous two Specific Aims and test the hypothesis that persistent calcium sparklet activity is increased during hypertension. This work should provide the first integrated view of calcium sparklet-mediated signaling and their role in modulating cerebral arteries function and significantly enhance our understanding of arterial function in health and disease.