TRANSGENIC MANIPULATION OF BETA-ARK1 MODIFIES BETA1-AR RESPONSIVENESS

激动剂诱导的β-肾上腺素能受体(β-AR)的磷酸化 通过细胞悬液中的β-肾上腺素能受体激酶1(BetaARK1) 可导致受体之间的正常连接迅速中断 和刺激性鸟嘌呤核苷酸结合蛋白Gs。要确定 β-ARK1对心肌功能的直接功能意义 我们测量了对急性β1-AR刺激的收缩反应。 从对照和转基因小鼠分离的左心室肌细胞 过表达BetaARK1(TGbetaK12)或BetaARK1抑制剂 (TGMini27)。对五种浓度(0.01至0.10_M)的收缩反应 β1-AR激动剂去甲肾上腺素(NE)+哌唑嗪(1_M) 在60秒休息后测量，即休息状态收缩(RSC)， 在稳态(SSC)时，以0.5赫兹(23℃)刺激。在基线上， RSC和SSC在对照组之间无显著差异， TGbK12或TGMini27小鼠。另一方面，无论是剂量反应还是 NE诱导的SSC/RSC反应的动力学明显不同 试验组间差异有统计学意义(P<0.001)。具体地说，最大 去甲肾上腺素诱导大鼠心肌细胞收缩反应 TGbetaK12组的反应仅为对照细胞的70%， 在TGMini27细胞中测得的反应的50%。人类社会的演变 TGMini27细胞的NE收缩反应分别快37%和67% 对照组和TGbK12细胞(p<0.03)。因此， 对Beta1-AR信令进行实质功能的BetaARK1调制 在心肌细胞中，即使在短期1-AR刺激期间也是如此。这些 结果与激动剂诱导的磷酸化和 βARK1在单个心肌细胞中对心脏β1-AR的脱敏作用 强调BetaARK1作为一项重要决定因素的潜在作用 心脏β1-AR收缩反应。这项研究的发现 具有特殊的临床相关性，因为它们为 使用βARK1抑制作为一种潜在的治疗方法来改善 慢性心力衰竭患者的心功能 已知的是，β-ARK1水平升高。

Agonist-induced phosphorylation of beta-adrenergic receptors (beta-AR's) by the beta-adrenergic receptor kinase 1 (betaARK1) in cell suspensions can result in a rapid disruption of the normal coupling between receptors and the stimulatory guanine nucleotide-binding protein Gs. To determine the direct functional significance of betaARK1 on myocardial performance, we measured the contractile responses to acute beta1-AR stimulation in left ventricular myocytes isolated from control and transgenic mice overexpressing either betaARK1 (TGbetaK12) or a betaARK1 inhibitor (TGMini27). Contractile response to five concentrations (.01 to .10 _M) of the beta1-AR agonist norepinephrine (NE) plus prazosin (1 _M) was measured after a 60 second rest, i.e. the rested state contraction (RSC), and during steady state (SSC) stimulation at 0.5 Hz (23_). At baseline, there were no significant differences in RSC or SSC between control, TGbK12 or TGMini27 mice. On the other hand, both the dose-response and kinetics for the NE-induced SSC/RSC response were significantly different among experimental groups (p lesser than 0.001). Specifically, the maximal contractile response induced by NE in myocytes isolated from the TGbetaK12 group was only 70% of the response observed in control cells, and 50% of the response measured in TGMini27 cells. The evolution of the NE contractile response in TGMini27 cells was 37% and 67% faster vs control and TGbK12 cells, respectively (p lesser than 0.03). Thus, substantial functional betaARK1 modulation of beta1-AR signalling occurs in cardiac myocytes, even during short term 1-AR stimulation. These results are consistent with a role for agonist-induced phosphorylation and desensitization of cardiac beta1-AR's by betaARK1 in single myocytes, and highlight the potential role of betaARK1 as an important determinant of the cardiac beta1-AR contractile response. The findings of this study have particular clinical relevance as they provide further support for the use of betaARK1 inhibition as a potential therapeutic approach to improve cardiac function in those humans with chronic heart failure, where betaARK1 levels are known to be elevated.