Regulation of bradykinin-induced activation of volume-sensitive outwardly rectifying anion channels by Ca2+ nanodomains in mouse astrocytes
Regulation of bradykinin-induced activation of volume-sensitive outwardly rectifying anion channels by Ca2+ nanodomains in mouse astrocytes
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DOI：
10.1113/jphysiol.2011.208173
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发表时间：
2011-08-15
影响因子：
5.5
通讯作者：
Okada, Yasunobu
中科院分区：
文献类型：
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作者：
Akita, Tenpei;Okada, Yasunobu
体积敏感的外向整流(VSOR)阴离子通道在细胞体积调节、细胞死亡诱导和细胞间通讯等多种基本细胞功能中发挥着关键作用。我们先前证明，在培养的小鼠皮质星形胶质细胞中，VSOR通道在炎症介质缓激肽的作用下被激活，即使细胞体积没有增加。在这里，我们报道了VSOR通道的激活必须首先由高[Ca~(2+)](I)的‘纳米结构域’所介导，这些高[Ca~(2+)](I)既产生于细胞内钙释放部位，又产生于质膜上的钙进入部位。缓激肽引起[Ca~(2+)](I)升高，最初由Ca~(2+)释放引起，然后由Ca~(2+)内流引起。通过去除细胞外钙离子和耗尽钙离子储备抑制[Ca~(2+)](I)升高，以一种分级的方式抑制VSOR通道的激活。定量RT-PCR和用小干扰RNA抑制基因表达表明，ORAIL、TRPC1和TRPC3通道参与了钙离子的内流，尤其是通过TRPC1通道的内流强烈地参与了缓激肽诱导的VSOR通道的激活。此外，还发现钙依赖的蛋白激酶Cα和β通过诱导活性氧的产生来介导[Ca~(2+)](I)升高后的激活。然而，细胞内应用10 mM的慢钙离子螯合剂EGTA或1 mM的快速螯合剂BAPTA对VSOR通道的激活几乎没有影响。在10 mM处应用BAPTA可显著抑制1/3的激活。提示缓激肽诱导的VSOR通道激活受单个钙离子释放和进入通道附近钙离子的调节，为细胞体积调节和细胞间通讯的局部调控提供了基础。
Volume-sensitive outwardly rectifying (VSOR) anion channels play a key role in a variety of essential cell functions including cell volume regulation, cell death induction and intercellular communications. We previously demonstrated that, in cultured mouse cortical astrocytes, VSOR channels are activated in response to an inflammatory mediator, bradykinin, even without an increase in cell volume. Here we report that this VSOR channel activation must be mediated firstly by 'nanodomains' of high [Ca2+](i) generated at the sites of both Ca2+ release from intracellular Ca2+ stores and Ca2+ entry at the plasma membrane. Bradykinin elicited a [Ca2+](i) rise, initially caused by Ca2+ release and then by Ca2+ entry. Suppression of the [Ca2+](i) rise by removal of extracellular Ca2+ and by depletion of Ca2+ stores suppressed the VSOR channel activation in a graded manner. Quantitative RT-PCR and suppression of gene expression with small interfering RNAs indicated that Orail, TRPC1 and TRPC3 channels are involved in the Ca2+ entry and especially the entry through TRPC1 channels is strongly involved in the bradykinin-induced activation of VSOR channels. Moreover, Ca2+-dependent protein kinases C alpha and beta were found to mediate the activation after the [Ca2+](i) rise through inducing generation of reactive oxygen species. Intracellular application of a slow Ca2+ chelator, EGTA, at 10 mM or a fast chelator, BAPTA, at 1 mM, however, had little effect on the VSOR channel activation. Application of BAPTA at 10 mM suppressed significantly the activation to one-third. These suggest that the VSOR channel activation induced by bradykinin is regulated by Ca2+ in the vicinity of individual Ca2+ release and entry channels, providing a basis for local control of cell volume regulation and intercellular communications.