摘要
目的利用无镁细胞外液诱导原代培养大鼠海马神经元癫痫放电模型来检测电压门控性钙通道Ca v1.2和钙调蛋白激酶Ⅱ的表达变化。方法采用新生24h内Wistar大鼠,取海马进行神经元原代培养。体外培养至12d,无镁细胞外液处理一部分神经元3h后,应用全细胞膜片钳技术记录神经元的放电情况以及免疫印迹法检测电压门控性钙通道Ca v 1.2和钙调蛋白激酶Ⅱ的蛋白表达。无镁细胞外液处理另一部分细胞12h后检测Ca v1.2和钙调蛋白激酶Ⅱ的蛋白表达。结果在无镁细胞外液处理3h后,神经元存在自发的"癫痫样"放电,而神经元Ca v1.2和磷酸化钙调蛋白激酶Ⅱ表达不变;无镁诱导12h后,神经元电压门控性钙通道Ca v1.2表达下调,而磷酸化钙调蛋白激酶Ⅱ表达上调。结论电压门控性钙通道Ca v1.2和钙调蛋白激酶Ⅱ的表达变化可能与无镁诱导体外培养大鼠海马神经元自发异常放电的基础病理机制相关。
Objective To study the expression of voltage-gated calcium channel Cav1.2 and calcium/ calmodulin-dependent protein kinase Ⅱ （CaMKⅡ） in cultured hippocampal neurons of Mg2＋-free epileptiform discharge model Wistar rat. Methods The postnatal hippocampus was taken out from 1-day-old Wistar rats and used for primary culture. 12 days after primary culture, neurons were treated with magnesium-free extracellular fluid for 3h, and the discharge activity was recorded by patch clamp. The expression of Cav1.2 and CaMKⅡ proteins was detected by western blot after magnesium-free extracellular fluid treatment for 3h and 12h. Results Neurons showed spontaneous ＂epileptiform discharge＂ 3h after magnesium-free extracellular fluid treatment, but with no difference for the expression levels of Cav1.2 and pCaMKⅡ proteins between magnesium-free group and control group. However, the expression level of Cav1.2 was downregulated and pCaMKⅡ was upregulated in magnesium-free group compared with control group 12hs after magnesium-free extracellular fluid treatment. Conclusion The abnormal expression of Cav1.2 and CaMKⅡ might be related to the underlying mechanism of spontaneous ＂epileptiform discharge＂ in Mg2＋-free neuron model rat.
出处
《解剖科学进展》
CAS
2012年第2期122-125,共4页
Progress of Anatomical Sciences
基金
国家自然科学基金资助项目(810014291)
辽宁省教育厅项目一般项目(L2010573)
关键词
海马
膜片钳
电压门控性钙通道
钙调蛋白激酶Ⅱ
WISTAR大鼠
hippocampus
patch clamp
voltage-gated calcium channel
calcium/calmodulin-dependent protein kinase Ⅱ
Wistar rat
