早期惊厥样放电对发育中神经元NMDA受体1表达的远期影响

摘要

Objective To explore the long-term effect of early discharges on NMDAR-1 expression in developing rat cortical neurons. Methods The seizure model was recurrent epileptiform discharges induced by transient magnesium-free media treatment in the primary cultured embryo cortical neurons of rat. According to different treatment to cortical neurons at cultured day 6, cells were assigned to three groups respectively based on the mediums they were transiently exposed to. The neurons were exposed to Dulbecco's Modified Eagle's Medium (DMEM) in control group 1 (CONT1), exposed to a physiological solution (PS) in control group 2(CONT2) ,and exposed to the same medium as CONT2 except for the removal of magnesium in the magnesium-free physiological solution (MGF) group. Following a 3 h exposure, the expression of N-methyl-D-aspartate receptor 1 (NR1) mRNA and protein were measured at different culture days (7,12,17) using real-time PCR and Western blot assays. Results The expression of NR1 mRNA and protein in DMEM medium had higher value level at culture day in vitro (DIV) 7,and then they decreased gradually and had the lowest value at DIV12. They increased again with development. Compared with groups CONT1 and CONT2 respectively,expression of NR1 mRNA and protein in group MGF decreased at DIV7 and increased at DIV12 (P＜0.05) ,and there were no statistic significant differences at DIV17. Conclusion The results suggest that NR1 mRNA and protein may alter with the development and there are long-term effects of Mg2+-free induced seizure-like discharges on expression of NR1 mRNA and protein in developing rat cortical neurons.%目的 探讨早期惊厥样放电对发育中大鼠皮层神经元N-甲基-D-天冬氨酸受体1(N-methyl-D-aspartate receptor 1,NR1)表达的远期影响.材料与方法以原代培养的大鼠大脑皮层神经元无镁诱导的反复惊厥样放电为模型.根据对培养6 d皮层神经元的不同处理分为三组:正常DMEM培养液组(CONT1)、正常细胞外液组(CONT2)和无镁细胞外液组(MGF).神经元分别在上述三种液体中孵育3 h,然后恢复正常DMEM培养基继续培养,分别应用实时定量PCR与Western blot方法在培养7、12及17 d时测定皮层神经元NMDA受体1 mRNA与蛋白表达的变化.结果 在正常DMEM培养液中,NR1 mRNA与蛋白的表达在培养7 d时处于较高水平,以后逐渐下降,以培养12 d为最低.与CONT2和CONT1组相比,MGF组NR1 mRNA与蛋白的表达在培养7 d皮层神经元均明显降低,在12 d时明显增高(P＜0.05).CONT2组与CONT1组相比无统计学差异(P＞0.05).结论 发育中皮层神经元早期无镁诱导惊厥样放电对NMDA受体1 mRNA与蛋白表达具有远期影响,提供了神经元早期惊厥样放电引起远期功能损伤的可能机制.