Mechanisms of Auto-Rhythmicity in Heart Development

描述（由申请人提供）：这项工作的长期目标是了解健康和疾病中心脏自律性的机制。这项建议的目的是确定离子机制的心脏起搏在发展中的心脏。心律失常在所有年龄段都很常见，经常导致心脏性猝死。大多数心律失常是心肌病的结果，其中有证据表明，胚胎蛋白质亚型，重要的节律性和收缩性，重新表达。自发性节律活动在心脏发育的早期就开始了，因为心脏新月形折叠形成中空管并开始泵血。胚胎心肌细胞的相对简单性使其在辨别胚胎和成人心脏自律性的机制方面很有用。从早期胚胎心管（包括原始心室）分离的所有肌细胞在作为单细胞分离时都是自主节律和自发搏动的。胚胎心脏节律性的基础尚不清楚。本实验将验证胚胎心脏Na/Ca交换器携带的离子电流与Ca ~（2+）、K ~+电流的活性相协调，从而建立起心脏起搏舒张期去极化的假说。其目标是：1）确定在发育中的胚胎心脏中作为心脏起搏器舒张期去极化基础的内向电流是否仅由INaCa携带; 2）确定来自Ca 2+电流、SR和其他来源的Ca 2+对心脏发育中起搏器功能的贡献; 3）确定在心脏发育中产生舒张期去极化的INaCa、If和K+电流之间的关系。膜片钳方法结合使用荧光钙和其他离子指示剂，和计算机建模将被用来确定在细胞水平上的自发性心律失常的机制，在小鼠和鸡胚胎的管状心脏的早期发育。将使用定量RT-PCR和免疫荧光共聚焦显微镜测定关键蛋白质的发育表达。从拟议的实验完成的结果将导致更好地了解人类心脏病的心律失常，并促进治疗策略的设计。

DESCRIPTION (provided by applicant): The long term objective of this work is to understand the mechanisms for cardiac autorhythmicity in health and disease. The objective of this proposal is to determine the ionic mechanisms underlying cardiac pacemaking in developing heart. Cardiac arhythmias are common to all ages and frequently lead to sudden cardiac death. Most arhythmias are a result of cardiomyopathies in which there is evidence showing that embryonic protein isoforms, important for rhythmicity and contractility, are re-expressed. Spontaneous rhythmic activity begins very early in heart development as the cardiac crescent folds to form a hollow tube and begins pumping blood. The relative simplicity of the embryonic cardiac myocyte makes it useful in discerning mechanisms responsible for autorhythmicity in the embryonic and adult heart. All of the myocytes isolated from the early embryonic heart tube, including the primitive ventricle, are autorhythmic and beat spontaneously when isolated as single cells. The basis of rhythmicity in the embryonic heart is unknown. The proposed experiments will test the hypothesis that the ionic current carried by the Na/Ca exchanger is coordinated with the activity of Ca2+, K+ currents to establish the pacemaker diastolic depolarization in embryonic heart. The aims are: 1) determine whether the inward current underlying the cardiac pacemaker diastolic depolarization is carried exclusively by INaCa in the developing embryonic heart; 2) determine the contributions of Ca2+ from Ca2+ currents, the SR, and other sources to pacemaker function in heart development; and 3) determine the relationships between lNaCa, If and K+ currents in generating the diastolic depolarization in heart development. Patch clamp methods in conjunction with the use of fluorescent Ca and other ion indicators, and computer modeling will be used to determine the mechanisms of auto-rhythmicity at the cellular level in early development of the tubular heart in both mouse and chick embryos. Developmental expression of key proteins will be determined using quantitative RT-PCR and immunoflourescence confocal microscopy. The results from the completion of the proposed experiments will lead to a greater understanding of cardiac arhythmias in human heart disease and facilitate the design of treatment strategies.