电磁脉冲对海马神经元的损伤效应及其对[Ca^(2+)]_i的影响

The injurious effects and the changes of [Ca^(2+)]_i iuduced by electromagnetic pulse on hippocampal neurons in vitro

目的 观察电磁脉冲(EMP)对海马神经元的损伤效应及其对[Ca2+ ]i的影响,以深入探讨EMP致脑损伤的机制。方法 EMP辐射条件为6×104V/m,脉冲上升时间为20ns,脉宽为30μs,频率为2. 5脉冲/min,作用2min。原代培养的海马神经元经EMP辐射后,采用MTT法对细胞活力进行测定;FACS法检测细胞凋亡和坏死;Fluo 3 AM荧光探针负载、激光共聚焦显微镜扫描测定神经元胞内游离钙离子浓度,即[Ca2+ ]i。结果 细胞被EMP辐射后0~6h活力明显下降(P<0. 05), 12h开始回升, 24h基本接近正常水平;EMP辐射后0~12h细胞凋亡率显著升高,坏死也有增加;EMP辐射后即刻引起神经元内的Ca2+荧光强度明显增加。结论 EMP致海马神经元活力下降、凋亡增加、胞内钙超载。

Objective To investigate the injurious effects and the changes of [Ca^(2+)]_i induced by electromagnetic pulse ~EMP) on hippocampal neurons in vitro. Methods Primary hippocampal neurons in vitro were exposed to EMP^6×10~4V/m,rise time 20ns,pulse width 30μs,5 pulses in 2 minute) radiation. Then the survival rate of neurons was detected with MTT, and the apoptosis and necrosis of neurons were observed with FACS. Fluo-3-AM fluorenscent probe and laser scanning confocal microscopy were employed to detect [Ca^(2+)]_i in neurons as well. Results The survival rate of neurons decreased significantly at 0~6h after radiation(P<0.05),then increased gradually after 12h and reached normal level at 24h.The rate of apoptosis increased markedly at 0~12h after EMP and the rate of necrosis increased too. A higher intracellular free calcium ion concentration was seen after EMP. Conclusion EMP caused an overloading of Ca^(2+) in hippocampal cells in vitro. Necrosis and apoptosis of neurons could be found as well.