大鼠海马神经元体外缺糖缺氧模型的建立

ESTABLISHMENT OF THE MODEL OF OXYGEN-GLUCOSE DEPRIVATION IN VITRO RAT HIPPOCAMPAL NEURONS

目的 :建立体外培养大鼠海马神经元缺糖缺氧模型。方法 :取培养 12d的海马神经元 ,在缺糖缺氧条件下分别培养 0 .5～ 4h后取出 ,换原神经元培养液 ,在常氧下继续培养 2 4h后测定培养液中乳酸脱氢酶活性。测定神经元形态变化 ,并计算神经元存活百分率。同时用原位末端标记 (TUNEL)法检测神经元凋亡。结果 :缺糖缺氧后海马神经元胞体逐渐肿胀 ,培养液中LDH释放量逐渐增多 ,细胞存活率逐渐减少。恢复糖和氧供应后 2 4h ,凋亡神经元百分率明显增多。结论 :用改进的无血清、无糖人工脑脊液成功建立了大鼠海马神经元离体缺糖缺氧模型。

Aim:To establish the model of oxygen-glucose deprivation in vitro rat hippocampal neurons. Methods: The hippocampal neurons cultured for 12 d were exposed to combined oxygen-glucose deprivation for 0.5～4 h and then cultured with original medium in normoxia for 24 h. Auto-biochemical analyzer determined LDH activity. The change of neuronal morphology and neuron survival were observed by converted contrast microscope and assessed by photography analysis system. Neuron apoptosis was detected by using the terminal deoxynucleotidyl transferese-mediated biotinylated deoxyuridine triphosphate nickel end labeling(TUNEL) method. Results:The neurons swelled, LDH release increased and neuron survival decreased after gradually oxygen-glucose deprivation. The percentage of apoptosis increased obviously 24 h after recovering the supply of oxygen and glucose. Conclusion:The model of oxygen-glucose deprivation in vitro rat hippocampal neurons is established successfully by using the modified ACSF(artificial cerebral spinal fluid) with serum and glucose free.