Regulation of Coronary Blood Flow

项目摘要 这个项目的目标是更好地了解代谢偶联的机制 心肌氧需求和氧供应之间通过冠状动脉血流量的变化。 氧化还原敏感性电压门控钾通道（即，Kv1.x）在冠状动脉血管平滑肌（CVSM）中的作用 已知对于响应于增加的心肌血流量（MBF）的增强是必不可少的。 心脏负荷CVSM中Kv 1通道活性如何响应于来自一个人的代谢信号而增强 活性心肌未知。在这里，我们提出了一个重要的作用，代谢调节Kv 1通道 通过胞质辅助Kvβ亚基的四聚体组装的活性，这些亚基是醛酮 还原酶（AKR）超家族并结合氧化和还原的吡啶核苷酸（例如，NAD（H））高 亲和力我们的初步数据与总体假设一致，即NAD（H）和 Kvβ1和Kvβ2精确控制局部MBF与心脏需氧量之间的耦合。 在目的1中，我们将描述冠状动脉Kvβ亚单位在代谢性充血调节中的作用 反应为此，我们将使用非侵入性心肌声学造影（MCE）来测量MBF 在麻醉WT和Kvβ-null动物中， 去甲肾上腺素（NE）。CVSM中Kvβ1和Kvβ2对代谢性充血的细胞特异性贡献将是 在双转基因动物中检查，在空载体中可诱导平滑肌特异性表达Kvβ， 背景动物体外电生理学和肌描记术将测试Kvβ在改变肌张力中的相对作用。 Kv 1激活和失活以及代谢性血管舒张的电压依赖性。在目标2中， 将阐明Kvβ介导的代谢偶联机制。我们将用电生理学来测量， Kvβ亚基对Kv 1活性调节的相对功能贡献， 改变CVSM中NADH：NAD+的氧化还原比的细胞代谢。我们将量化 使用遗传编码的荧光生物传感器和MALDI-MS成像，CVSM中的NADH/NAD+氧化还原，以及 通过Kvβ催化确定NAD（H）周转在调节冠状动脉血管舒张中的作用， 加强医疗保险基金。在目标3中，我们将阐明Kvβ在心血管运动适应中的作用 条件反射为了做到这一点，我们将野生型和转基因动物进行强制跑步机跑步 在测量MBF、冠状动脉血管舒张能力和Kv激活的适应性之前的运动方案， 失活特性。这一目标还将解决生理性心肌缺血的总体依赖性。 适应和增强运动能力对冠脉Kvβ依赖性MBF变化的影响。

PROJECT SUMMARY The goal of this project is to gain a better understanding of mechanisms that underlie metabolic coupling between myocardial oxygen demand and oxygen supply via changes in coronary blood flow to the heart. Redox-sensitive voltage-gated potassium channels (i.e., Kv1.x) in coronary vascular smooth muscle (CVSM) are known to be essential to the enhancement of myocardial blood flow (MBF) in response to increased cardiac workload. How Kv1 channel activity is enhanced in CVSM in response to metabolic signals from an active myocardium is unknown. Here, we propose an essential role for metabolic regulation of Kv1 channel activity by the tetrameric assembly of cytosolic auxiliary Kvβ subunits, which are members of the aldo-keto reductase (AKR) superfamily and bind oxidized and reduced pyridine nucleotides (e.g., NAD(H)) with high affinity. Our preliminary data are consistent with the global hypothesis that interactions between NAD(H) and Kvβ1 and Kvβ2 impart precise control over the coupling between regional MBF with cardiac oxygen demand. In Aim 1, we will delineate the role of coronary Kvβ subunits in the regulation of the metabolic hyperemia response. To do this, we will use non-invasive myocardial contrast echocardiography (MCE) to measure MBF as a function of cardiac workload in anesthetized WT and Kvβ-null animals upon administration of norepinephrine (NE). The cell-specific contribution of Kvβ1 and Kvβ2 in CVSM to metabolic hyperemia will be examined in double transgenic animals with inducible smooth muscle-specific expression of Kvβ in null background animals. In vitro electrophysiology and myography will test the relative roles for Kvβ in altering the voltage-dependence of Kv1 activation and inactivation and metabolic vasodilation, respectively. In Aim 2, we will elucidate the mechanism of Kvβ-mediated metabolic coupling. We will measure, using electrophysiology, the relative functional contribution of Kvβ subunits to regulation of Kv1 activity upon specific manipulations in cellular metabolism that alter the redox ratio of NADH:NAD+ in CVSM. We will quantify changes in NADH/NAD+ redox in CVSM using genetically-encoded fluorescent biosensors and MALDI-MS imaging, and determine the role of NAD(H) turnover by Kvβ catalysis in regulation of coronary vasodilation and enhancement of MBF. In Aim 3, we will clarify the role of Kvβ in cardiovascular adaptation to exercise conditioning. To do this, we will subject WT and genetically-modified animals to a forced treadmill running exercise protocol before measuring adaptations in MBF, coronary vasodilatory capacity, and Kv activation and inactivation properties. This aim will also address the overall dependence of physiological myocardial adaptations and enhancement of exercise capacity on coronary Kvβ-dependent changes in MBF.