5 D impulse mapping in the embryonic heart

胚胎心脏的 5D 脉冲图

• 批准号: 10572425
• 负责人: ANDREW Martin ROLLINS
• 金额: $ 71.09万
• 依托单位: CASE WESTERN RESERVE UNIVERSITY
• 依托单位国家: 美国
• 财政年份: 2023
• 资助国家: 美国
• 起止时间: 2023-01-01 至 2026-12-31
• 项目状态: 未结题
• 来源: https://reporter.nih.gov/project-details/10572425
• 关键词: 3-Dimensional; Address; Birds; Calcium; Cardiac; Cardiac Electrophysiologic Techniques; Cardiac Myocytes; Cardiac conduction system; Collaborations; Compensation; Complex; Confocal Microscopy; Congenital Abnormality; Congenital Heart Defects; Coupled; Data; Detection; Development; Dyes; Electrophysiology (science); Embryo; Embryonic Heart; Etiology; Fetal Alcohol Exposure; Fluorescent Dyes; Fluorescent in Situ Hybridization; Functional disorder; Gene Expression; Genes; Goals; Health; Heart; Image; Imaging Device; Incubated; Lead; Lighting; Longitudinal Studies; Maps; Membrane; Modeling; Molecular; Morphogenesis; Motion; Multimodal Imaging; Optics; Pathology; Pattern; Pharmaceutical Preparations; Physiological; Physiology; Productivity; Property; Quail; Qualifying; Reaction; Reporter; Role; Scanning; Signal Transduction; Structure; System; Techniques; Technology; Therapeutic; Time; Tissues; Transfection; Transgenic Organisms; Tubular formation; Viral; calcium indicator; cardiogenesis; developmental cardiology; fluorescence imaging; heart cell; heart motion; hemodynamics; image registration; in vivo; new technology; prevent; protein expression; ratiometric; reconstruction; response; serial imaging; success; technology development; tool; two photon microscopy; voltage

Project Summary/Abstract Congenital heart defects (CHDs) are among the most common and devastating of birth defects. Abnormal development of cardiac conduction often associates with CHD etiology. Investigating the development of cardiac conduction is essential for us to understand the mechanisms behind such conditions. Development of the cardiac conduction system (CCS) is a complicated transformation that comes about through interplay between molecular signaling, structural properties, and physiological function, including hemodynamics and electrophysiology. While an understanding of the molecular networks has progressed rapidly in recent decades, tools to follow the physiological factors contributing to development and differentiation of the CCS remain deficient. This gap is important to fill. Connecting molecular and functional information is key for understanding pathologies of the conduction system and for guiding potential therapeutic strategies. Optical mapping (OM) of transmembrane voltage or intracellular calcium dynamics in the heart using voltage- or calcium-sensitive fluorescent dyes is a powerful tool for studying cardiac electrophysiology, and has been adapted for imaging early embryonic hearts with great success. However currently, OM is limited to imaging excised embryonic hearts which are stilled with excitation-contraction-uncoupler drugs. Removing fragile tubular hearts from the structure and hemodynamic load of the embryo, and incubating them in dyes and drugs interferes with the normal physiology and does not allow longitudinal study over stages of development. The goal of this project is to develop technology to enable comprehensive, longitudinal imaging of the electrophysiological function of the living, beating heart of the early avian embryo, cultured under near- physiological conditions. We will target 1-2 days of active morphogenesis through the transition from homogeneous to heterogeneous conduction velocity Three key technology developments are needed to achieve this. (A) Episcopic, volumetric, fast imaging of fluorescent voltage and calcium indicators is needed to image the intact, living embryo, and to capture conduction dynamics (Aim 1). (B) Motion correction is needed to enable conduction mapping of the beating heart, without using excitation-contraction-uncoupling drugs (Aim 1). (C) Embryonic quail models with calcium and voltage reporters expressed in cardiomyocytes are needed to enable in vivo and longitudinal imaging of electrophysiology (Aim 2). The proposed technology will enable simultaneous 3D conduction mapping over two time scales, the heartbeat, and heart development (5D impulse mapping). Coupled with quantitative 3D FISH, this will allow point-to-point 3D registration between conduction data and gene/protein expression, which is not currently available, enabling studies to better understand mechanisms of conduction function, dysfunction and development (Aim 3).

项目概要/摘要 先天性心脏病（CHD）是最常见且最具破坏性的出生缺陷之一。异常 心脏传导的发展通常与 CHD 病因相关。调查发展 心脏传导对于我们了解此类情况背后的机制至关重要。 心脏传导系统（CCS）的发展是一个复杂的转变 通过分子信号传导、结构特性和生理功能之间的相互作用，包括 血流动力学和电生理学。虽然对分子网络的理解取得了迅速进展 近几十年来，追踪有助于发育和分化的生理因素的工具出现了。 CCS 仍然存在不足。填补这一空白很重要。连接分子和功能信息是关键 了解传导系统的病理学并指导潜在的治疗策略。 使用光学映射 (OM) 测量心脏中的跨膜电压或细胞内钙动态 电压或钙敏感荧光染料是研究心脏电生理学的有力工具，并且具有 已被用于早期胚胎心脏成像并取得了巨大成功。然而目前，OM仅限于成像 切除的胚胎心脏，用兴奋收缩解偶联剂药物使其静止。去除脆弱的管状体 从胚胎的结构和血流动力学负荷中观察心脏，并在染料和药物中孵育它们会产生干扰 具有正常的生理机能，不允许对发育阶段进行纵向研究。 该项目的目标是开发技术以实现对物体的全面纵向成像 近培养的早期禽胚胎活的、跳动的心脏的电生理功能 生理条件。我们的目标是通过从过渡到 1-2 天的活跃形态发生 均质到异质传导速度需要三项关键技术开发 实现这一点。 (A) 需要荧光电压和钙指示剂的落射、体积、快速成像 对完整的活胚胎进行成像，并捕获传导动力学（目标 1）。 (B) 需要运动校正 无需使用兴奋-收缩-解偶联药物即可绘制跳动心脏的传导图（Aim 1). (C) 需要具有在心肌细胞中表达的钙和电压报告基因的胚胎鹌鹑模型 实现电生理学的体内和纵向成像（目标 2）。 所提出的技术将能够在两个时间尺度上同时进行 3D 传导映射， 心跳和心脏发育（5D 脉冲映射）。与定量 3D FISH 相结合，这将允许 传导数据和基因/蛋白质表达之间的点对点 3D 配准，目前还没有 可用，使研究能够更好地了解传导功能、功能障碍和 发展（目标 3）。