Acetylcholine inhibits the hypoxia-induced reduction of connexin43 protein in rat cardiomyocytes
Acetylcholine inhibits the hypoxia-induced reduction of connexin43 protein in rat cardiomyocytes
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DOI：
10.1254/jphs.fp0051023
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发表时间：
2006-07-01
影响因子：
3.5
通讯作者：
Sato, Takayuki
中科院分区：
文献类型：
--
作者：
Zhang, Yanan;Kakinuma, Yoshihiko;Sato, Takayuki
在最近的一项研究中，我们证明迷走神经刺激通过调节连接蛋白43（Cx43）抑制致死性心律失常来增加心肌梗死大鼠的存活率。然而，这种效应的确切机制仍有待阐明。为了研究这些机制和缝隙连接调节的信号转导，我们研究了乙酰胆碱（ACh），一种副交感神经系统的神经递质，对差距连接组件Cx43的影响使用H9c2细胞。当细胞缺氧时，总Cx43蛋白水平降低。相比之下，ACh预处理可抑制这种作用。为了研究信号转导，细胞预处理与L-NAME，一氧化氮合酶抑制剂，然后ACh和缺氧。发现L-NAME抑制ACh效应。然而，NO供体，SNAP，部分抑制缺氧诱导的Cx43减少。为了阐明缺氧条件下Cx43减少的机制，用蛋白酶体抑制剂MG 132预处理细胞。蛋白酶体抑制产生了显着的恢复缺氧下的总Cx43蛋白水平的下降。然而，与MG 132和ACh共处理没有产生任何进一步增加总Cx43蛋白水平。使用乙酰胆碱或冈田酸，磷酸酶抑制剂的功能研究表明，这两种试剂抑制缺氧引起的染料转移的减少。这些结果表明，ACh是负责恢复Cx43蛋白水平的下降，导致间隙连接的功能激活。
In a recent study, we demonstrated that vagal stimulation increases the survival of rats with myocardial infarction by inhibiting lethal arrhythmia through regulation of connexin43 (Cx43). However, the precise mechanisms for this effect remain to be elucidated. To investigate these mechanisms and the signal transduction for gap junction regulation, we investigated the effect of acetylcholine (ACh), a parasympathetic nerve system neurotransmitter, on the gap junction component Cx43 using H9c2 cells. When cells were subjected to hypoxia, the total Cx43 protein level was decreased. In contrast, pretreatment with ACh inhibited this effect. To investigate the signal transduction, cells were pretreated with L-NAME, a nitric oxide synthase inhibitor, followed by ACh and hypoxia. L-NAME was found to suppress the ACh effect. However, a NO donor, SNAP, partially inhibited the hypoxia-induced reduction in Cx43. To delineate the mechanisms of the decrease in Cx43 under hypoxia, cells were pretreated with MG132, a proteasome inhibitor. Proteasome inhibition produced a striking recovery of the decrease in the total Cx43 protein level under hypoxia. However, cotreatment with MG132 and ACh did not produce any further increase in the total Cx43 protein level. Functional studies using ACh or okadaic acid, a phosphatase inhibitor, revealed that both reagents inhibited the decrease in the dye transfer induced by hypoxia. These results suggest that ACh is responsible for restoring the decrease in the Cx43 protein level, resulting in functional activation of gap junctions.