一种定量评价离体海马脑片缺糖/缺氧损伤的新方法  

Novel quantitative method for evaluating oxygen/glucose deprivation-induced injury of hippocampal slices

作　　者：严乐勤[1] 魏尔清[1] 胡海涛[1] 张纬萍[1] 王梦令[1] 沈建中[1] 

机构地区：[1]浙江大学医学院,浙江杭州310031

出　　处：《浙江大学学报（医学版）》2002年第2期81-85,共5页Journal of Zhejiang University(Medical Sciences)

基　　金：国家自然科学基金 ((39770 2 70 );浙江省自然科学基金 (3990 90 )资助项目

摘　　要：目的 :建立一种定量评价离体脑片缺糖 /缺氧损伤的简便、快速、灵敏的方法。方法 :离体海马脑片缺糖 /缺氧不同时间后以 2 % 2 ,3,5 -三苯基氯化四氮唑 (TTC)染色 ,用乙醇∶二甲亚砜 (5 0∶ 5 0 )抽提 TTC红色结晶产物 ,分光光度法测定 OD490 值 ,并与 LDH释放率进行比较。结果 :随着缺糖 /缺氧时间的延长 ,TTC染色 (OD490 值 )逐渐下降 ,而 LDH释放率逐渐上升 ,两结果呈负相关 (r=- 0 .933,P<0 .0 1) ,提示脑片缺血性损伤。尼莫地平、地塞米松和氯胺酮可减轻脑片缺血性损伤 ,但 ONO- 10 78无效。结论 :TTC染色法简单易行 ,且客观灵敏 ,可用于离体脑片损伤的评价及药物初筛。Objective: To establish a simple, sensitive in vitro method to evaluate oxygen/glucose deprivation (OGD) induced injury of brain hippocampal slices in rats. Methods: Rat hippocampal slices were incubated in 2% 2,3,5 triphenyltetrazolium chloride (TTC) solution after oxygen/glucose deprivation. They were then soaked in a measured volume of ethanol and dimethylsulfoxide(50∶50) to extract the TTC formazan product which was then measured by spectrophotometry. OGD induced LDH release was simultaneously measured. Results: Progressive prolongation of OGD induced hippocampal injury resulted in decreased formazan coloration as determined by spectrophotometry. There was a parallel increase in LDH release, thus a negative correlation in these two products was noted. ( r =-0.933, P <0.01). The injury was attenuated in the brain slices pre treated with nimodipine, dexa methasone, and ketamine, but not ONO 1078. Conclusion: Solvent extraction and spectrophotometric quantification of formazan represents an objective measurement of OGD induced injury of rat hippocampal slices.

关 键 词：脑损伤 OGD 分光光度法 定量评价 脑缺血 动物实验 脑片缺糖缺氧损伤 

分 类 号：R741.02[医药卫生—神经病学与精神病学]