Muscarinic Stimulation of α1E Ca Channels Is Selectively Blocked by the Effector Antagonist Function of RGS2 and Phospholipase C-β1
Muscarinic Stimulation of α1E Ca Channels Is Selectively Blocked by the Effector Antagonist Function of RGS2 and Phospholipase C-β1
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DOI：
10.1523/jneurosci.20-19-07167.2000
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发表时间：
2000-10
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影响因子：
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通讯作者：
K. Melliti;U. Meza;B. Adams
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作者：
K. Melliti;U. Meza;B. Adams
神经元α1E钙通道亚基在哺乳动物脑中广泛表达，在那里它们被认为形成R型钙通道。最近的研究表明，R型通道有助于神经分泌和树突状钙内流，但对其调节机制知之甚少。在这里，我们表明，α1E通道通过M1毒扁豆碱受体被强烈刺激，而只有微弱的抑制。这两种形式的通道调节都是由百日咳毒素不敏感的G蛋白介导的。G蛋白信号转导调节因子2(RGS2)或磷脂酶C-β1的C末端区域(PLCβ1ct)可阻断通道刺激，这两种信号转导蛋白可作为GαQ的GTP酶激活蛋白。相反，RGS2和PLCβ1ct不能通过M1受体阻断对α1E的抑制。然而，β-肾上腺素能受体激酶1的C末端区域可以阻止抑制，该区域隔离Gβγ二聚体。因此，对α1E的刺激是由百日咳毒素不敏感的Gα亚单位(如GαQ)介导的，而抑制是由Gβγ介导的。RGS2和PLCβ1ct选择性阻断刺激的能力表明这些蛋白主要作为效应器拮抗剂发挥作用。为了支持这一解释，RGS 2阻止了非水解性鸟苷5‘-0-(3-硫代三磷酸)对α1E的刺激。我们还报道了对Rbe-II的强烈毒碱刺激，Rbe-II是一种变异的α1E钙通道，对电压依赖抑制不敏感。我们的结果预测，G-αQ偶联受体主要刺激天然的R型钙通道。受体介导的R型钙电流的增强可能对神经分泌、树突状细胞的兴奋性、基因表达或其他神经元功能产生重要影响。
Neuronal α1E Ca channel subunits are widely expressed in mammalian brain, where they are thought to form R-type Ca channels. Recent studies have demonstrated that R-type channels contribute to neurosecretion and dendritic Ca influx, but little is known concerning their modulation. Here we show that α1E channels are strongly stimulated, and only weakly inhibited, through M1 muscarinic acetylcholine receptors. Both forms of channel modulation are mediated by pertussis toxin-insensitive G-proteins. Channel stimulation is blocked by regulator of G-protein signaling 2 (RGS2) or the C-terminal region of phospholipase C-β1 (PLCβ1ct), which have been previously shown to function as GTPase-activating proteins for Gαq. In contrast, RGS2 and PLCβ1ct do not block inhibition of α1E through M1 receptors. Inhibition is prevented, however, by the C-terminal region of β-adrenergic receptor kinase 1, which sequesters Gβγ dimers. Thus, stimulation of α1E is mediated by a pertussis toxin-insensitive Gα subunit (e.g., Gαq), whereas inhibition is mediated by Gβγ. The ability of RGS2 and PLCβ1ct to selectively block stimulation indicates these proteins functioned primarily as effector antagonists. In support of this interpretation, RGS2 prevented stimulation of α1E with non-hydrolyzable guanosine 5′-0-(3-thiotriphosphate). We also report strong muscarinic stimulation of rbE-II, a variant α1E Ca channel that is insensitive to voltage-dependent inhibition. Our results predict that Gαq-coupled receptors predominantly stimulate native R-type Ca channels. Receptor-mediated enhancement of R-type Ca currents may have important consequences for neurosecretion, dendritic excitability, gene expression, or other neuronal functions.