电磁脉冲辐照所致海马神经细胞膜穿孔的原子力显微镜研究

The Atom Force Microscopy Study on the Hypothalamus Neurons Membrane Perforate Induced by EMP

目的:应用原子力显微镜(atomicforcemicroscopy熏AFM)对电磁脉冲辐照前后培养的海马神经元细胞膜进行观察,以探讨电磁脉冲对细胞损伤的机制。材料和方法:将新生的Wistar(24小时内)乳鼠解剖镜下分离出海马组织,将海马神经细胞稀释成1×106个/ml浓度的细胞悬液;将细胞悬液种植到六孔板中,2ml/孔。培养14天后,高场强EMP模拟源穴有界波模拟源雪,场强为6×104V/m,脉冲上升时间为20ns,脉宽为30μs熏频率包含0～100MHz。以2.5次/min熏辐照2分钟。并于辐照后即刻固定细胞,应用日本岛津穴SHIMADZU雪公司的SPM-9500J3型原子力显微镜对细胞表面进行接触式连续扫描。结果:电磁脉冲辐照后即刻就可引起大鼠海马神经元细胞膜表面大小不一、类圆形和不规则形的穿孔,穿孔最大口径和深度可达281.56nm×89.90nm×17.76nm。结论:电磁脉冲辐照后可直接导致海马神经元细胞膜的穿孔,提示细胞膜可能是电磁脉冲生物效应的靶部位之一。

Objective: Using the atomic force microscopy (AFM) to observe the membrane surface changes of rat's hippocampus primary culture neurons irradiated by EMP, and to explore possible injured mechanism of EMP to neuron membrane. Methods: Hippocampus from neonatal rats(24 hours) were isolated by dissection microscope, the concentration of the dilution neurons was 1×106/ml, then the cells were cultured in 6-well culture plates. About 14 days later, the cells were irradiated by high field strength 5 EMP ( electromagnetic pulse, EMP) within 2 minutes, The electromagnetic pulse simulator which provides 5 pulses/min with a high electric field intensity 60 kv/m, 20 ns rise time and 30 μs pulse wide. The exposed cells were fixed immediately following irradiation and scanned by AFM (SPM-9500J3, SHIMADZU). Results: The results showed that there is membrane perforate with different size and shape (281.56 nm×89.90 nm×17.76 nm). Conclusion: EMP could injury the neuron membrane of hippocampus, and the membrane may be the target position of EMP.