HETEROGENEITY OF EMBRYONIC CHICK MYOCARDIUM

胚胎心血管系统同时调节结构和功能的戏剧性转变。该过程对于实现成熟心脏的复杂3D几何形状和功能至关重要，并且该过程中的错误/失败是先天性心血管畸形的一种拟议发病机制。该修订提案代表了一种新的方法来定义机械负荷条件的特定变化如何改变局部CV收缩性，肌膜Ca 2+电流和肌细胞被动和主动特性。具体目标一。在正常发育、重新分配前负荷（LA结扎）或增加后负荷（CT带）的第18 - 34期鸡胚中，确定机械壁应变、心肌收缩力和Cab电流之间的关系，以确定响应于改变的机械负荷而发生的功能适应的时间过程、位置和程度。假设1-1。以2D和/或3D心外膜应变测量的局部心室机械负荷增加会增加局部胚胎收缩力，以体内收缩壁应力-应变关系和收缩力（Eav-max）以及体外力-长度、力-频率和力-速度关系测量。假设1-2。增加机械负荷增加胚胎T型和L型Ca 2+电流，如通过同时测量体外力-长度关系和细胞内Ca 2+（条）;全细胞膜片钳和Ca 2+成像以及Ni 2+、米贝拉地尔、硝苯地平和ryanodine阻断（条和细胞）所定义。 具体目标2。定义机械壁应变之间的关系，肌纤维和胶原对齐，和粘弹性在第18至34期鸡胚在正常发育，重新分配前负荷（LA结扎）或增加后负荷（CT带），补充在体内和体外研究。 假设2-1。心室机械负荷增加加速和重新分配肌纤维排列。 假设2-2。 心室机械负荷增加会降低根据被动壁应力-应变关系（体外，有限元伪应变能量密度函数）计算的顺应性，并增加胶原含量和对齐。重要性：该修订提案测试了发育中心脏的机械负荷、心肌功能、肌纤维和基质结构之间的特定关系，并为未来的实验提供了关键基础，以确定发育中心肌中抑制机械力的细胞内机制（基因、蛋白质、反式激活因子）。

Embryonic cardiovascular systems regulate dramatic transitions in structure and function simultaneously. This process is critically important to achieve the complex 3D geometry and function of the mature heart and errors/failures in this process are one proposed pathogenetic mechanism for congenital cardiovascular malformations. This revised proposal represents an novel approach to define how specific changes in mechanical loading conditions alter regional CV contractility, sarcolemmal Ca2+ currents and myocyte passive and active properties. Specific Aim I. Define the relationship between mechanical wall strain, myocardial contractility, and Cab currents in stage 18 to 34 chick embryos during normal development, redistributed preload (LA ligation) or increased afterload (CT band) to define the time-course, location, and extent of functional adaptation that occurs in response to altered mechanical load. Hypothesis 1-1. Increased regional ventricular mechanical load measured as 2D and/or 3D epicardial strain increases local embryonic contractility measured as in vivo systolic wall stress- strain relations and contractility (Eav-max) and in vitro force-length, force-frequency, and force-velocity relations. Hypothesis 1-2. Increased mechanical load increases embryonic T-type and L-type Ca2+ currents as defined by simultaneous measurement of in vitro force-length relations and intracellular Ca2+ (strips); whole-cell patch clamp and Ca2+- imaging and Ni2+, mibefradil, nifedipine, and ryanodine blockade (strips and cells). Specific Aim 2. Define the relationship between mechanical wall strain, myofiber and collagen alignment, and viscoelastic properties in stage 18 to 34 chick embryos during normal development, redistributed preload (LA ligation) or increased afterload (CT band), complementing both in vivo and in vitro studies. Hypothesis 2-1. Increased ventricular mechanical load accelerates and redistributes myofiber alignment. Hypothesis 2-2. Increased ventricular mechanical load decreases compliance calculated from passive wall stress-strain relations (in vitro, finite element pseudo-strain energy density function) and increases collagen content and alignment. Significance: This revised proposal TESTS specific relationships between mechanical load, myocardial function, and myofiber and matrix architecture in the developing heart and provides a critical foundation for future experiments to identify the intracellular mechanisms (genes, proteins, trans-activating factors) that transduce mechanical forces in the developing myocardium.