L-type channel trafficking and modulation in heart

摘要 心肌L型钙通道在心肌兴奋-收缩偶联、动作电位中起关键作用 持续时间和基因表达。CAV1.2功能异常，包括长开模式增加 门控和钝化的肾上腺素能反应与心力衰竭和肥厚有关。这个 CaV1.2的激活增加，进而触发钙反应信号通路，这有助于 心力衰竭和肥厚的发病机制。将CaV1.2适当地定位到不同的表面位置，以及 荷尔蒙调节它们的活动，对正常的心脏生理至关重要。心脏中的Cav1.2与 影响渠道运输、定位、周转和功能的大型超分子复合体。大有可为 关于CaV1.2贩运和调制的基本机制的普遍教条的来源是 利用在异源表达系统中重组的重组通道的研究。不幸的是， 然而，至少一些关于贩运和监管的最关键问题-肾上腺素能 心肌细胞中的激动剂不能用异源表达来评估，可能是因为独特的 心肌细胞的细胞内环境在其他地方是不能复制的。在上一个资助期，我们 开发和实施创新方法，以调查CaV1.2贩运和β背后的决定因素 心肌细胞的肾上腺素能直接调节。使用这一方法，我们的研究发现，成人中β-Lessα1C 心肌细胞产生具有正常基础活动的CaV1.2通道，这些通道不受PKA的调节 第一次有机会探讨这种调节对交感调节的相对贡献 飞行反应。我们提出了三个目标，以我们之前的发现为基础，并旨在深化 CaV1.2在心脏中的调节机制：(1)确定Cav与1C结合在心肌细胞中的作用 在体调节心肌收缩能力；(2)确定亚基使- CaV1.2的肾上腺素能调节。(3)阐明β依赖和β非依赖的CaV1.2的作用机制 渠道贩运。这三个目的，应该提供关于监管的关键新认识 心肌细胞内钙离子的内流，与了解心脏病理和 心肌兴奋收缩偶联和肾上腺素能调节的分子机制 心脏的钙通道。

SUMMARY The cardiac L-type Ca2+ channel plays a key role in cardiac excitation-contraction coupling, action potential duration, and gene expression. Abnormalities in CaV1.2 function, including increased long-opening-mode gating and blunted adrenergic responsiveness, are associated with heart failure and hypertrophy. The increased activation of CaV1.2, in turn, triggers Ca2+-responsive signaling pathways, which contribute to the pathogenesis of heart failure and hypertrophy. Proper targeting of CaV1.2 to distinct surface sites, and hormonal regulation of their activity, is vital for normal cardiac physiology. Cav1.2 in heart is associated with large supramolecular complexes that impact on channel trafficking, localization, turnover, and function. Much of the prevailing dogma relating to mechanisms underlying CaV1.2 trafficking and modulation is derived from studies using recombinant channels reconstituted in heterologous expression systems. Unfortunately, however, at least some of the most critical questions regarding trafficking and regulation by -adrenergic agonists in cardiomyocytes cannot be assessed using heterologous expression, likely because the unique intracellular environment of cardiomyocytes is not reproduced elsewhere. In the previous funding period, we developed and implemented innovative methods to probe determinants underlying CaV1.2 trafficking and β- adrenergic regulation directly in cardiomyocytes. Using this approach, our findings that β-less α1C in adult cardiomyocytes generate CaV1.2 channels with normal basal activity that are not regulated by PKA provide the first opportunity to probe the relative contribution of this modulation to sympathetic regulation in the fight or flight response. We propose three Aims that build on our previous findings, and designed to deepen mechanistic understanding of CaV1.2 regulation in heart: (1) Determine the role of CaV binding to 1C in regulating cardiac contractility in vivo; (2) To determine the mechanism(s) by which  subunits enable - adrenergic regulation of CaV1.2. (3) To elucidate the mechanisms of β-dependent and β-independent CaV1.2 channel trafficking. The three Aims, which should provide key new understandings concerning the regulation of Ca2+ influx in cardiomyocytes, are highly relevant towards understanding cardiac pathologies and the molecular mechanisms responsible for cardiac excitation-contraction coupling and adrenergic modulation of the cardiac Ca2+ channel.