Rybin Vitalyi O, Pak Elena, Alcott Sasha, Steinberg Susan F
Department of Pharmacology, College of Physicians and Surgeons, Columbia University, 630 West 168th Street, New York, NY 10032, USA.
Mol Pharmacol. 2003 Jun;63(6):1338-48. doi: 10.1124/mol.63.6.1338.
Cardiomyocyte beta2-adrenergic receptors (beta-ARs) provide a source of inotropic support and influence the evolution of heart failure. Recent studies identify distinct mechanisms for beta2-AR actions in neonatal and adult rat cardiomyocytes. This study examines whether ontogenic changes in cardiac beta2-AR actions can be attributed to altered Gi expression or changes in the spatial organization of the beta2-AR complex in membrane subdomains (caveolae). We show that beta2-ARs increase cAMP, calcium, and contractile amplitude in a pertussis toxin (PTX)-insensitive manner in neonatal cardiomyocytes. This is not caused by lack of Gi; Galphai expression is higher in neonatal cardiomyocytes than in those of adult rats. beta2-ARs provide inotropic support without detectably increasing cAMP, in adult cardiomyocytes. This cannot be attributed to dual coupling of beta2-ARs to Gs and Gi, because beta2-ARs do not promote cAMP accumulation in PTX-pretreated adult cardiomyocytes. Spatial segregation of beta2-ARs, Galphas/Galphai, and adenylyl cyclase to distinct membrane subdomains also is not a factor, because all of these proteins copurify in caveolin-3-enriched vesicles isolated from adult cardiomyocytes. However, these studies demonstrate that enzyme-based protocols routinely used to isolate ventricular cardiomyocytes lead to proteolysis of beta-ARs. The functional consequences of this limited beta-AR proteolysis is uncertain, because truncated beta1-ARs promote cAMP accumulation and truncated beta2-ARs provide inotropic support in adult cardiomyocytes. Collectively, these studies indicate that components of the beta2-AR signaling complex compartmentalize to restricted membrane subdomains in adult rat cardiomyocytes. Neither compartmentalization nor changes in Gi expression fully explain the ontogenic changes in beta2-AR responsiveness in the rat ventricle.
心肌细胞β2 - 肾上腺素能受体（β - ARs）提供了变力支持的来源，并影响心力衰竭的发展。最近的研究确定了β2 - AR在新生大鼠和成年大鼠心肌细胞中作用的不同机制。本研究探讨了心脏β2 - AR作用的个体发育变化是否可归因于Gi表达的改变或膜亚结构域（小窝）中β2 - AR复合物空间组织的变化。我们发现，在新生心肌细胞中，β2 - ARs以百日咳毒素（PTX）不敏感的方式增加环磷酸腺苷（cAMP）、钙和收缩幅度。这并非由于缺乏Gi；新生心肌细胞中Gαi的表达高于成年大鼠心肌细胞。在成年心肌细胞中，β2 - ARs提供变力支持，但未检测到cAMP增加。这不能归因于β2 - ARs与Gs和Gi的双重偶联，因为β2 - ARs不会促进PTX预处理的成年心肌细胞中cAMP的积累。β2 - ARs、Gαs/Gαi和腺苷酸环化酶在不同膜亚结构域的空间分隔也不是一个因素，因为所有这些蛋白质在从成年心肌细胞分离的富含小窝蛋白 - 3的囊泡中共同纯化。然而，这些研究表明，常规用于分离心室心肌细胞的基于酶的方案会导致β - ARs的蛋白水解。这种有限的β - AR蛋白水解的功能后果尚不确定，因为截短的β1 - ARs促进cAMP积累，而截短的β2 - ARs在成年心肌细胞中提供变力支持。总体而言，这些研究表明，β2 - AR信号复合物的成分在成年大鼠心肌细胞中分隔到受限的膜亚结构域。分隔和Gi表达的变化都不能完全解释大鼠心室中β2 - AR反应性的个体发育变化。
