Laboratory of Molecular Pharmacology, Division of Pharmaceutical Sciences, Kanazawa University Graduate School of Natural Science and Technology, Japan.
J Pharmacol Sci. 2011;116(4):350-61. doi: 10.1254/jphs.11069fp. Epub 2011 Jul 9.
Clock genes are believed to play a pivotal role in the generation and oscillation of circadian rhythm as a central clock in the hypothalamic suprachiasmatic nucleus in the mammalian brain. In this study, mRNA expression was for the first time demonstrated with clock genes in both cultured murine microglia and microglial cell line BV-2 cells. Exposure to ATP transiently increased Period-1 (Per1) mRNA expression without affecting that of other clock genes in BV-2 cells, while a similarly transient increase was shown in Per1 mRNA expression in a manner sensitive to P2X7 purinergic receptor antagonists in cultured microglia exposed to ATP. In BV-2 cells transfected with a Per1 promoter luciferase reporter plasmid, ATP significantly increased luciferase activity in a manner sensitive to a P2X7-receptor antagonist. In both microglia and BV-2 cells, a significant increase by ATP was seen in the immunocytochemical fluorescence intensity of cells expressing Per1 protein, with mRNA expression of different P2 receptors including P2X7. Per1 siRNA significantly decreased the number of cells with processes in BV-2 cells exposed to ATP. These results suggest that ATP selectively promotes Per1 expression through gene transactivation after stimulation of P2X7 purinergic receptors in microglial cells.
时钟基因被认为在生成和振荡生物钟中发挥关键作用，作为哺乳动物大脑下丘脑视交叉上核中的中央时钟。在这项研究中，首次证明了培养的鼠小胶质细胞和小胶质细胞系 BV-2 细胞中存在时钟基因的 mRNA 表达。ATP 的暴露短暂地增加了 Period-1 (Per1) mRNA 的表达，而不影响其他时钟基因的表达，而在暴露于 ATP 的培养小胶质细胞中，Per1 mRNA 的表达以对 P2X7 嘌呤能受体拮抗剂敏感的方式表现出类似的短暂增加。在转染 Per1 启动子荧光素酶报告质粒的 BV-2 细胞中，ATP 以对 P2X7 受体拮抗剂敏感的方式显著增加荧光素酶活性。在小胶质细胞和 BV-2 细胞中，ATP 均显著增加了表达 Per1 蛋白的细胞的免疫细胞化学荧光强度，同时也增加了不同的 P2 受体（包括 P2X7）的 mRNA 表达。在暴露于 ATP 的 BV-2 细胞中，Per1 siRNA 显著减少了具有突起的细胞数量。这些结果表明，ATP 通过刺激小胶质细胞中的 P2X7 嘌呤能受体后进行基因转录激活，选择性地促进 Per1 表达。
