摘要
目的 研究大鼠脑局灶性缺血再灌注模型中N 甲基 D天冬氨酸 (NMDA)受体随时间的变化。方法 采用线栓法阻断大鼠大脑中动脉 ,产生局灶性脑缺血再灌注模型 ,快速取脑 ,冰冻切片。以3H MK 80 1作为放射性配基 ,将恒温保温后的标本在室温下氚片上进行放射自显影 ,勾划脑左右两侧感兴趣区 (ROI) ,采用LETCAQ 5 5 0IWL图像分析系统测定并比较缺血侧与对侧ROI内NMDA受体的密度。在受体结合法中 ,分离出缺血区中段皮层 ,匀浆、离心后与放射性配基进行受体结合反应 ,将液闪测得的数值作Scatchard图 ,求出NMDA受体的亲和力 (Kd)及最大结合容量 (Bmax)并进行比较。结果 根据放射自显影图 ,缺血 2h组以及缺血 2h再灌 2h组同假手术组相比 ,相应的NMDA受体密度比值明显升高 ,表明NMDA受体通道大量开放。这主要是兴奋性氨基酸升高所致。再灌注2 4h组及再灌注 72h组相应ROI中NMDA受体开放明显降低 ,且两组无明显差异。各实验组中缺血及再灌注区Kd 值都未发生明显变化 ,表明缺血再灌注前后受体亲和力未发生改变。缺血 2h组及再灌注 2h组Bmax值明显升高 ,再灌 2 4h及 72h组的Bmax明显降低。结论 大鼠脑局灶缺血及再灌注过程中 ,兴奋性氨基酸明显上升并维持数小时 ;NMDA受体在 4h内被大量激活 ,通道开放 ,引起缺血损伤。
Objective To study the changes of N methyl D aspartate(NMDA) receptor in the rat brain during reperfusion after local ischemia. Methods The rat cerebral focal ischemia/reperfusion model after middle cerebral artery occlusion (MCAO) was made by the intraluminal suture method. After ischemia and reperfusion models were completed, the rat brain was removed immediately and dissected under the frozen state. After incubation with 3H MK 801which was used as a radioactive ligand, the samples were exposed on the hypersensitive film at the room temperature. Regions of interest (ROIs) were set bilaterally in each autoradiographic brain image. LEICAQ 550IWL imaging analysis system was used to detect the density of NMDA receptor. The ratios of the receptor density between affected and unaffected side in the same ROI were compared. Before receptor binding assay, the middle section of the ischemic cortex was separated, homogenated and centrifugated. After binding with the radioligand, radioactivity was measured by liquid scintillation counter. Scatchard plot was made from the liquid scintillation counter and affinity ( K d) and maximal binding (B max ) changes of NMDA receptor were calculated. The results of K d and B max were compared between groups. Results NMDA receptor density in the correspon ding ROI increased significantly during 2 h ischemia and 2 h reperfusion in comparison with the result of the sham group, which showed that the NMDA receptor channel was opened abuntantly. The opening of the channel was mainly caused by the increase of the excitatory amino acids. There was significant decrease in the degree of channel opening in the 24 h and 72 h reperfusion groups in corresponding ROIs and no difference was found between two groups. The value of K d in ischemia and reperfusion groups did not change. B max of NMDA receptor increased obviously during 2 h reperfusion and decreased obviously during 24 h and 72 h reperfusion. Conclusions During the ischemia reperfusion, the excitatory amino acids increased greatly and remained high for a few hours. NMDA receptor was activated and its ion channel opened accordingly, which produced the ischemia damage. The best time of the therapy window of brain ischemia should be within 4 h.
出处
《中华核医学杂志》
CAS
CSCD
北大核心
2000年第6期259-262,共4页
Chinese Journal of Nuclear Medicine
关键词
受体
NMDA
放射自显影
脑梗塞
局灶性脑缺血
Receptors, N methyl D aspartate
Autoradiography
Radioligand assay
Cerebral infarction
