Melatonin‐mediated inhibition of Purkinje neuron P‐type Ca2+ channels in vitro induces neuronal hyperexcitability through the phosphatidylinositol 3‐kinase‐dependent protein kinase C delta pathway
Melatonin‐mediated inhibition of Purkinje neuron P‐type Ca2+ channels in vitro induces neuronal hyperexcitability through the phosphatidylinositol 3‐kinase‐dependent protein kinase C delta pathway
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DOI：
10.1111/jpi.12218
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发表时间：
2015-04
影响因子：
10.3
通讯作者：
Y. Zhang;Hua Li;Yichen Pu;S. Gong;Chunfeng Liu;Xinghong Jiang;J. Tao
中科院分区：
文献类型：
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作者：
Y. Zhang;Hua Li;Yichen Pu;S. Gong;Chunfeng Liu;Xinghong Jiang;J. Tao
尽管褪黑激素受体在哺乳动物中枢神经系统和外周组织中广泛表达，但有关褪黑激素在小脑浦肯野细胞中的功能的数据有限。在这里，我们确定了褪黑激素在调节大鼠浦肯野神经元 P 型 Ca2+ 通道和动作电位放电中的新功能作用。 0.1 μm 的褪黑激素可逆地将峰值电流 (IBa) 降低 32.9%。这种效应依赖于褪黑激素受体 1 (MTR1)，并且与失活电压依赖性的超极化转变相关。百日咳毒素预处理、QEHA 肽的细胞内应用以及针对 Gβ 亚基的选择性抗体可阻止褪黑激素的抑制作用。用磷脂酰肌醇 3 激酶 (PI3K) 抑制剂进行预处理可消除褪黑激素诱导的 IBa 降低。令人惊讶的是，褪黑激素反应不受 Akt（PI3K 的常见下游靶标）的调节。褪黑激素治疗使蛋白激酶 C (PKC) 活性显着增加 2.1 倍。 PKC 拮抗剂（但不是蛋白激酶 A）可以消除褪黑激素诱导的 IBa 降低。褪黑素的应用增加了 PKCδ 的膜丰度，而 PKCδ 抑制（无论是药理学上还是基因上）消除了褪黑素诱导的 IBa 反应。从功能上讲，褪黑激素使自发动作电位放电增加 53.0%； MTR1 的敲除和 P 型通道的阻断消除了这种效应。因此，我们的结果表明，褪黑激素通过 MTR1 激活抑制 P 型通道，MTR1 依次与 Gi/o 蛋白的 βγ 亚基和下游 PI3K 依赖性 PKCδ 信号传导偶联。这可能有助于其生理功能，包括小脑浦肯野神经元的自发放电。
Although melatonin receptors are widely expressed in the mammalian central nervous system and peripheral tissues, there are limited data regarding the functions of melatonin in cerebellar Purkinje cells. Here, we identified a novel functional role of melatonin in modulating P‐type Ca2+ channels and action‐potential firing in rat Purkinje neurons. Melatonin at 0.1 μm reversibly decreased peak currents (IBa) by 32.9%. This effect was melatonin receptor 1 (MTR1) dependent and was associated with a hyperpolarizing shift in the voltage dependence of inactivation. Pertussis toxin pretreatment, intracellular application of QEHA peptide, and a selective antibody raised against the Gβ subunit prevented the inhibitory effects of melatonin. Pretreatment with phosphatidylinositol 3‐kinase (PI3K) inhibitors abolished the melatonin‐induced decrease in IBa. Surprisingly, melatonin responses were not regulated by Akt, a common downstream target of PI3K. Melatonin treatment significantly increased protein kinase C (PKC) activity 2.1‐fold. Antagonists of PKC, but not of protein kinase A, abolished the melatonin‐induced decrease in IBa. Melatonin application increased the membrane abundance of PKCδ, and PKCδ inhibition (either pharmacologically or genetically) abolished the melatonin‐induced IBa response. Functionally, melatonin increased spontaneous action‐potential firing by 53.0%; knockdown of MTR1 and blockade of P‐type channels abolished this effect. Thus, our results suggest that melatonin inhibits P‐type channels through MTR1 activation, which is coupled sequentially to the βγ subunits of Gi/o‐protein and to downstream PI3K‐dependent PKCδ signaling. This likely contributes to its physiological functions, including spontaneous firing of cerebellar Purkinje neurons.