CaMKII in Sinus Node Physiology and Disease

描述（由申请人提供）：最近令人兴奋的证据支持这样一种观点，即窦房结（SAN）心脏起搏器细胞通过增强肌浆网（SR） Ca2+释放和增强肌质Na+/Ca2+交换器（NCX）携带的去极化内向电流导致心率增加。在收缩的心肌细胞中，专门用于兴奋-收缩耦合和细胞Ca2+循环的蛋白质被保存在SAN细胞中，我们的研究表明，SAN细胞和心室肌细胞具有相似的SR Ca2+含量，以细胞膜电容为指标，尽管SAN细胞表现出最小的机械活性。多功能Ca2+和钙调素依赖性蛋白激酶II （CaMKII）是一种丝氨酸-苏氨酸激酶，调节sr相关的Ca2+稳态蛋白、磷蛋白（PLN）和红嘌呤受体（RyR）。我们的新数据支持CaMKII在肾上腺素能受体（AR）介导的SAN“战斗或逃跑”生理反应中起核心作用的观点。此外，在过度的病理性AR刺激条件下，CaMKII抑制可提高SAN细胞存活并防止SAN心律失常。我们假设1)CaMKII调节SR Ca2+的摄取和释放，作为生理SAN对AR刺激反应的细胞机制的一部分；2) CaMKII抑制通过防止SR Ca2+超载和减少SR Ca2+泄漏来预防SAN心律失常、SAN细胞死亡和肥大；3)向内NCX电流是SR Ca2+释放激活的主要肌层靶，导致4期细胞膜去极化增强和心率加快。我们的实验室已经适应并开发了新的方法来可靠地分离小鼠和兔子的SAN细胞，并进行体内病毒基因转移到SAN和体外病毒基因转移到培养的SAN细胞。这些创新的方法将使我们能够全面测试CaMKII， SR Ca2+和NCX在心脏起搏中的作用。公共卫生相关性：窦房结（SAN）心脏起搏器衰竭是美国发病率和死亡率的重要原因。我们每年在永久性手术植入的心脏起搏器上花费超过20亿美元。起搏器是一种成功但非常不完善的治疗方法，它不能治疗潜在疾病，容易发生机械故障、感染，需要反复手术更换。因此，改良的非侵入性治疗SAN衰竭将为美国人带来重要的公共健康益处。此外，SAN衰竭是房颤患者的常见发现，并与心力衰竭死亡率增加有关。尽管这些特化的SAN细胞具有明显的重要性，但对于理解SAN细胞信号传导以及细胞内Ca2+和SAN细胞电输出之间的耦合，仍然存在关键的知识空白。提高对SAN细胞生物学的理解是开发改进的治疗方法来调节SAN功能和延长SAN生存的必要的第一步。我们的建议侧重于CaMKII作为诱导SAN疾病的核心但被忽视的信号的作用。

DESCRIPTION (provided by applicant): Exciting recent evidence supports a view that sinoatrial node (SAN) cardiac pacemaker cells cause heart rate increases by enhancement of sarcoplasmic reticulum (SR) Ca2+ release and augmentation of depolarizing inward current carried by the sarcolemmal Na+/Ca2+ exchanger (NCX). Proteins dedicated to excitation-contraction coupling and cellular Ca2+ cycling in contracting cardiomyocytes are preserved in SAN cells, and our studies show SAN cells and ventricular myocytes have similar SR Ca2+ content, indexed for cell membrane capacitance, despite the fact that SAN cells exhibit minimal mechanical activity. The multifunctional Ca2+ and calmodulin-dependent protein kinase II (CaMKII) is a serine- threonine kinase that regulates the SR-associated Ca2+ homeostatic proteins, phospholamban (PLN) and ryanodine receptors (RyR). Our new data support a view that CaMKII plays a central role in ¿adrenergic receptor (¿AR) mediated SAN 'fight or flight' physiological responses. Furthermore, under conditions of excessive, pathological ¿AR stimulation, CaMKII inhibition enhances SAN cell survival and protects against SAN arrhythmias. We hypothesize that 1) CaMKII regulates SR Ca2+ uptake and release as part of a cellular mechanism for physiological SAN responses to ¿AR stimulation; 2) CaMKII inhibition protects against SAN arrhythmias, SAN cell death and hypertrophy by preventing SR Ca2+ overload and reducing SR Ca2+ leak; 3) inward NCX current is the major sarcolemmal target activated by SR Ca2+ release, leading to enhanced phase 4 cell membrane depolarization and faster heart rates. Our laboratory has adapted and developed new methods for reliably isolating mouse and rabbit SAN cells, and performing in vivo viral gene transfer to the SAN and in vitro viral gene transfer to cultured SAN cells. These innovative approaches will allow us to comprehensively test the role of CaMKII, SR Ca2+ and NCX in cardiac pacing. PUBLIC HEALTH RELEVANCE: Sinoatrial nodal (SAN) cardiac pacemaker failure is a significant cause of morbidity and mortality in the United States. We spend over 2 billion dollars on permanent, surgically-implanted pacemakers annually. Pacemakers are a successful but highly imperfect therapy that do not treat the underlying disease and are subject to mechanical failure, infection and require repetitive surgical replacement. Thus, improved, non-invasive therapies for SAN failure would produce important public health benefits for Americans. Furthermore, SAN failure is a frequent finding in patients with atrial fibrillation and is associated with increased mortality in heart failure. Despite the clear importance of these specialized SAN cells, critical knowledge gaps remain for understanding SAN cell signaling and the coupling between intracellular Ca2+ and SAN cell electrical outputs. Improved understanding of SAN cell biology is a necessary first step to developing improved therapies for regulating SAN function and extending SAN survival. Our proposal focuses on the role of CaMKII as a central, but neglected, signal for inducing SAN disease.