Genetic Analyst of Early Conduction System Development

描述（由申请人提供）：了解专门的心室心脏传导系统的发展对于产生心律失常的生物治疗和掌握心脏组织再生至关重要。心律失常和猝死是几种心肌病的特征，这些心肌病具有异常小梁形成或致密化不全。心室肌小梁的发育和维持是心脏器官发生的关键过程，并通过erbb 2/神经调节蛋白（nrg）信号通路与成熟心室心传导模式的正常发育相关。erbb 2/nrg通路已经被认为在传导系统的发育中起作用。然而，无论是小梁形成的过程，还是它在为发育中的传导系统奠定基础方面的作用，都还不清楚。本研究的长期目标是描述特化传导系统的早期结构和功能，并确定小梁形成与传导系统发育的关系。该项目的具体目标有两个方面。首先，我们将建立专门的斑马鱼心室心脏传导系统的分子标记。我们将测试的假设，即第一个细胞的传导系统的位置位于心肌小梁层，和小梁突变缺乏传导系统的细胞，通过分析这些标志物的表达进行评估。第二，我们将检验这一假设，即小梁形成的过程，而不仅仅是nrg/erbb 2通路，是传导系统发育所必需的。由于其遗传易处理性、半透明的胚胎发生以及与人类电生理表型和疾病的相似性，斑马鱼是这些实验的有用模式生物。为了识别传导细胞，我们将使用原位杂交和抗体染色的候选标记物的整体安装和切片斑马鱼心脏。为了评估野生型、突变型和基因改变心脏的传导系统发育，我们将使用光学标测和胚胎ECG监测。我们将评估erbb 2突变体的传导表型，以及注射心脏特异性显性负性结构的野生型胚胎，这些结构旨在阻断小梁形成中涉及的其他途径。研究发育中的心脏传导系统的结构和功能及其与小梁形成的关系将进一步加深我们对先天性和获得性心肌病中心律失常的理解，并将提供通过操纵传导细胞来帮助治疗室性心律失常和猝死的工具和知识。

DESCRIPTION (provided by applicant): Understanding development of the specialized ventricular cardiac conduction system is critical to generating biologic treatments for arrhythmia and mastering cardiac tissue regeneration. Arrhythmia and sudden death characterize several cardiomyopathies with aberrant trabeculation or noncompaction. The development and maintenance of trabeculated ventricular myocardium is a key process in heart organogenesis and appears linked to the normal development of a mature ventricular cardiac conduction pattern by way of erbb2/neuregulin (nrg) signaling. The erbb2/nrg pathway is already known to play a part in conduction system development. However, neither the process of trabeculation nor its role in setting the stage for the developing conduction system is understood. The long-term goals of this research are to describe the early structure and function of the specialized conduction system, and to determine how trabeculation relates to conduction system development. The specific aims of this project are twofold. First, we will establish molecular markers of the specialized ventricular cardiac conduction system in zebrafish. We will test the hypotheses that the location of the first cells of the conduction system are located in the trabeculating myocardial layer, and that trabeculation mutants lack conduction system cells as assessed by analyzing the expression of these markers. Second, we will test the hypothesis that the process of trabeculation, and not just the nrg/erbb2 pathway, is required for conduction system development. The zebrafish is a useful model organism for these experiments due to its genetic tractability, translucent embryogenesis, and similarity to human electrophysiological phenotypes and disease. To identify conduction cells, we will use in situ hybridization and antibody staining of candidate markers on whole-mount and sectioned zebrafish hearts. To assess conduction system development in wild-type, mutant, and genetically altered hearts, we will use optical mapping and embryonic ECG monitoring. We will assess conduction phenotypes in erbb2 mutants, as well as in wild-type embryos injected with heart-specific dominant negative constructs designed to block other pathways involved in trabeculation. Studying the structure and function of the developing cardiac conduction system and its relationship to trabeculation will further our understanding of arrhythmia in both congenital and acquired cardiomyopathies and will provide tools and knowledge that can help treat ventricular arrhythmia and sudden death by manipulating conduction cells.