遗传性癫痫大鼠海马组织Ca2+/CaV1.2/CaM/CaMKⅡ信号通路的异常变化

摘要

目的 研究遗传性癫痫大鼠 (tremor,TRM) 海马组织电压门控性L型钙离子通道α1C亚单位 (CaV1.2)、钙调蛋白 (CaM)、钙调蛋白依赖性蛋白激酶Ⅱ (CaMKⅡ) 和细胞内钙离子浓度 ([Ca2+]i) 的变化情况.方法应用Western blot法与免疫荧光双标法检测TRM海马CA1、CA3和DG区CaV1.2、CaM和磷酸化CaMKⅡ (p-CaMKⅡ) 的蛋白表达及分布;激光共聚焦显微镜检测TRM海马组织中[Ca2+]i.结果与正常Wistar大鼠相比,TRM海马组织中CaV1.2和CaM的蛋白表达明显升高 (P<0.01),而p-CaMKⅡ的蛋白表达明显下降 (P<0.01);免疫荧光双标法结果显示:CaV1.2、CaM、p-CaMKⅡ在CA1、CA3区的锥体细胞和DG区的颗粒细胞群表达丰富,同时CaV1.2与CaM、p-CaMKⅡ与CaM在海马各区域均存在共定位;激光共聚焦显微镜检测TRM海马细胞[Ca2+]i明显增强 (P<0.01).结论 Ca2+/ CaV1.2/CaM/CaMKⅡ通路的异常变化可能参与遗传性癫痫大鼠的癫痫发生与发展.%Aim To observe the changes of L-type voltage-dependent calcium channel α1C-class ( Cavl. 2 ), calmodulin ( CaM ), calmodulin-depend-ent protein kinase Ⅱ ( CaMK Ⅱ ) and intracellular calcium concentration ([ Ca2+]i) in hippocampus of tremor rats ( TRMs ). Methods The protein distributions and expressions of Cav 1.2, CaM and phosphoryl-ated CaMK Ⅱ ( p-CaMK Ⅱ ) in TRM hippocampus were detected by double-labeling immunofluorescence and Western blot; [ Ca2+]i was measured by confocal laser scanning microscope. Results Compared to control rats, the protein expressions of Cav 1. 2 and CaM were increased, and p-CaMK Ⅱ was decreased in TRM hippocampus ( P < 0. 01 ). Double-labeled im-munofluorescence consequences suggested that in TRMs, Cavl. 2, CaM and p-CaMK Ⅱ protein appeared throughout CA1, CA3 and DG regions of hippocampus. It was noteworthy that co-localized neurons that expressed Cav 1. 2 with CaM and p-CaMK Ⅱ with CaM were also detected. Furthermore, compared to control rats, intracellular calcium concentrations were increased ( P < 0. 01 ). Conclusion The abnormal changes of Ca2+/Cav 1. 2/CaM/CaMK Ⅱ signaling pathway might be involved in TRMs' epileptogenesi.