BETA ADRENERGIC SIGNALING IN NEONATAL AND ADULT MYOCYTES

我们实验室最近的研究发现了以前未被认识到的 心脏β-肾上腺素能受体亚型信号传导的复杂性， 影响深远的发育变化β/2-受体激活效应 机制等β受体亚型信号传导的至少两个方面可以 不能用传统的受体概念来解释， 激活信号通路：[1]观察到β/2受体 仅在新生儿（而非成人）心肌细胞中增加cAMP积累。 [2]新生儿β受体亚型的观察 心肌细胞显示出对抑制性 毒蕈碱胆碱能激动剂的作用。在试图了解 这些有趣的复杂性背后的机制， 受体亚型信号传导，在本项目的更新研究将 应用与分子异质性有关的新概念， β-受体亚型信号级联（G蛋白亚基，腺苷酰 环化酶同工型和PKA酶的组分）以及 将第二信使分子分隔成小窝 （专门的膜子域，作为"信号处理"， "中心"）用于促进和/或限制受体信号传导事件。 前两个具体目标将阐明分子和/或细胞 β-肾上腺素能受体亚型信号传导差异的基础 新生儿和成人心室肌细胞。第三个具体目标将建立 根据我们最近令人兴奋的观察，β/2受体影响 通过cAMP非依赖性信号传导在成年心室肌细胞中的收缩 导致HCO/3依赖性细胞内碱化的途径， 研究β受体亚型对pH/i的调节。第四特定 目的将验证交感神经支配影响 β/2-受体作用。这些研究将结合联合收割机广泛的 生物化学/分子技术，荧光显微镜与离子- 灵敏的探针，和测量空载细胞缩短;这 允许分析细胞内信号传导的功能作用 β受体亚型反应性的中间体。通过考虑 调节心肌细胞自主神经的新信号转导机制 在正常生长、发育和/或交感神经系统的情况下， 心室的神经支配，本项目的研究将有助于 该计划的长期目标，这是了解结构， 分子、生物化学和/或离子决定因素，导致不同的 儿茶酚胺依赖性电生理和收缩反应 新生儿和成人心脏

Recent studies from our laboratory identify previously unappreciated complexities in cardiac beta-adrenergic receptor subtype signaling, with profound developmental changes beta/2-receptor activation of effector mechanisms. At least two aspects of beta-receptor subtype signaling can not be explained in the context of traditional concepts of receptor- activated signaling pathways: [1] The observation that beta/2-receptors increase cAMP accumulation only in neonatal (not in adult) cardiomyocytes. [2] The observation that individual beta-receptor subtypes in neonatal cardiomyocytes display differential susceptibility to the inhibitory effects of muscarinic cholinergic agonists. In an attempt to understand the mechanisms that underlie these intriguing complexities in beta- receptor subtype signaling, studies in the renewal of this Project will apply newer concepts related to molecular heterogeneity of components of beta-receptor subtype signaling cascades (G protein subunits, adenylyl cyclase isoforms, and components of the PKA enzyme) as well as the concept that compartmentalization of second messenger molecules to caveolae (specialized membrane subdomains that serve as 'signaling processing center') serves to facilitate and/or restrict receptor signaling events. The first two Specific Aim will elucidate the molecular and/or cellular basis for differences in beta-adrenergic receptor subtype signaling in neonatal and adult ventricular myocytes. The third Specific Aim will build upon our exciting recent observation that beta/2-receptors influence contraction in adult ventricular myocytes via a cAMP-independent signaling pathway leading to HCO/3-dependent intracellular alkalinization to investigate pH/i-regulation by beta-receptor subtypes. The fourth Specific Aim will test the hypothesis that sympathetic innervation influence beta/2-receptor action. These studies will combine a broad range of biochemical/molecular techniques, fluorescence microscopy with ion- sensitive probes, and measurements of unloaded cell shortening; this permits an analysis of the functional role of intracellular signaling intermediates in beta-receptor subtype responsiveness. By considering novel signal transduction mechanisms that regulate cardiomyocyte autonomic responses in the context of normal growth, development, and/or sympathetic innervation of the ventricle, studies in this Project will contribute to the Program's long-range goal, which is to understand the structural, molecular, biochemical, and/or ionic determinants that lead to distinct catecholamine-dependent electrophysiologic and contractile responses in newborn and adult hearts.