Understanding the role of mitochondrial dysfunction in cardiac arrhythmias using a novel 3D panoramic optical mapping system

摘要 心律失常是发病和死亡的主要原因，并且由于老龄化而日益普遍 患有糖尿病、心力衰竭和高血压的人群。心房颤动（AF）和心室颤动（VF）是 混乱性心律失常，而房性心动过速 (AT)、心房扑动 (AFL) 和室性心动过速 (VT) 更有组织性、局灶性或宏观折返性心律失常。我们对具体机制的掌握 容纳有组织和/或混乱节律的心脏基质是不完整的。心律失常的致病因素 包括纤维化、晚期 Na 电流增加和反应性氧化应激 (ROS) 增加，导致增强 线粒体自噬，是消除有缺陷的线粒体以维持线粒体整体健康的过程 线粒体池。我们的方法论突破是使用 3D 全景解剖和光学测绘， 结合线粒体自噬检测来表征电激活、底物之间的相互作用 由于纤维化和线粒体自噬导致的异质性，以及动作电位持续时间（APD）异质性。我们提出的 概念是更大或更多数量的纤维化区域、APD异质性和/或线粒体自噬将允许 更混乱的房性或室性心律失常。通过单独破坏这些途径并定义 对心律失常发生的影响，我们将确定这三个过程如何共同调节或 功能上相互依赖。我们将小鼠与检测线粒体自噬的报告基因 Keima 蛋白进行杂交 两系转基因小鼠由于基因突变而患有自发性和持续性 AF、AFL、AT、VT 和 VF 人类心脏 NaV1.5 通道基因 SCN5A。该项目提出了一种综合实验方法 使用 (1) 对 Na 超载小鼠模型的全心脏进行 AF、AFL、AT、VT 和 VF 多模态成像 或心肌梗死引起的 VT/VF 了解有组织和混乱的心房和心房的机制 室性心律失常发生，(2) AAV 递送线粒体过氧化氢酶以逆转术后增加的线粒体自噬 心肌梗塞和（3）通过 AAV 将视紫红质通道蛋白 2 递送至整个小鼠心脏的光遗传学 使用聚焦光刺激来触发、预防和终止房性和室性心律失常。拟议的 实验非常重要且具有创新性，因为共同配准的 3D 全景成像将使我们能够 剖析驱动有组织和混乱心律失常的机制，这可能会导致新的和 心律失常的有效治疗策略。

ABSTRACT Cardiac arrhythmias are a major cause of morbidity and mortality, and are increasingly prevalent due to an aging population with diabetes, heart failure and hypertension. Atrial fibrillation (AF) and ventricular fibrillation (VF), are chaotic arrhythmias, whereas, atrial tachycardia (AT), atrial flutter (AFL) and ventricular tachycardia (VT) are more organized, focal or macro-reentrant arrhythmias. Our grasp of the specific mechanisms that allow for the cardiac substrate to harbor organized and/or chaotic rhythms is incomplete. Causative factors of arrhythmias include fibrosis, increased late Na+ current and increased reactive oxidative stress (ROS) causing augmented mitophagy, which is a process of eliminating defective mitochondria to maintain the overall health of the mitochondrial pool. Our methodological breakthrough is to use 3D panoramic anatomical and optical mapping, in conjunction with mitophagy detection to characterize the interplay amongst electrical activation, substrate heterogeneity due to fibrosis and mitophagy, and action potential duration (APD) heterogeneity. Our proposed concept is that larger or greater number of areas of fibrosis, APD heterogeneity and/or mitophagy will allow for more chaotic atrial or ventricular arrhythmias. By individually disrupting these pathways and defining the consequences on arrhythmogenesis, we will determine how these three processes are co-regulated or functionally inter-dependent. We crossed mice with a reporter Keima protein which detects mitophagy, together with two lines of transgenic mice with spontaneous and sustained AF, AFL, AT, VT and VF due to mutations in the human cardiac NaV1.5 channel gene SCN5A. This project presents an integrated experimental approach using (1) multi-modality imaging of whole hearts of murine models of Na+ overload with AF, AFL, AT, VT and VF or myocardial infarction induced VT/VF to understand the mechanisms of organized and chaotic atrial and ventricular arrhythmogenesis, (2) AAV delivery of mitochondrial catalase to reverse increased mitophagy after myocardial infarction and (3) optogenetics via AAV delivery of channelrhodopsin-2 into whole murine hearts and use of focused light stimulation to trigger, prevent and terminate atrial and ventricular arrhythmias. The proposed experiments are highly significant and innovative in that co-registered 3D panoramic imaging will allow us to dissect the mechanisms that drive organized and chaotic cardiac arrhythmias, which may lead to new and effective treatment strategies of cardiac arrhythmias.