低氧预处理降低大鼠癫痫易感性及其脑保护作用的机制初探

Preliminary study on the role of hypoxia preconditioning in decreasing the susceptibility to epilepsy and brain protection in rats

目的初步探讨间断低氧预处理（intermittent hypoxia preconditioning,IHP）对氯化锂-匹鲁卡品（lithium-pilocarpine,Li-pilo）致痫大鼠痫性发作的影响及其脑保护作用机制。方法 96只清洁级SD大鼠,随机均分为对照组,癫痫组和4个间断低氧预处理＋癫痫组。癫痫组及4个间断低氧预处理＋癫痫组（分别在5 d IHP预处理后1,3,7,14日注射）大鼠通过腹腔注射氯化锂-匹鲁卡品建立癫痫模型。随后进行240 min的发作行为学,全身性发作潜伏期及百分比量化分析并通过水迷宫实验测试大鼠认知功能。接着利用TUNEL标记和免疫印迹方法进行大鼠海马神经元凋亡及相关蛋白（BCL-2,Bax和cleaved-caspase-3）的检测。结果氯化锂-匹鲁卡品注射后50~150 min之间,癫痫发作的改良Racine评分达到峰值。间断低氧预处理后3日诱发癫痫组大鼠其平均改良Racine评分明显低于其余各组,该组全身性发作潜伏期及百分比与癫痫组相比差异亦有统计学意义（P〈0.05）。相比癫痫组和其余3个间断低氧预处理＋癫痫组大鼠,间断低氧预处理后3日诱发癫痫组大鼠逃避潜伏期（escape latency）较短,神经元凋亡计数较低,而目标象限停留时间百分比较高（P〈0.05）。间断低氧预处理后3日诱发癫痫组的水迷宫以及凋亡检测参数与对照组相比较无明显差异（P〉0.05）。结论 IHP预处理通过抑制异常凋亡降低大鼠癫痫易感性,并具有脑保护作用。

Objective To preliminarily explore the effects and brain protective mechanism of intermittent hypoxia preconditioning （IHP） on rats with seizures induced by lithium-piloearpine （Li-pilo）. Methods A total of 96 8-week old male Sprague Dawley rats （ clean grade） were randomly divided into control group, seizure group and four IHP-seizore groups. The animal model of epilepsy was established by intraperitoneal injection of Li-pilo in the seizure group and four IHP-seizure groups （Li-pilo was injected at 1 , 3, 7, or 14 days after a 5-day regimen of IHP）. Subsequent seizure behavior, the latency period and percentage of generalized seizures were quantitatively evaluated for 240 rain and the cognitive function was tested by Morris water maze task, and followed by the detection of hippocampus neuron apoptosis and related protein （BCL-2, Bax, and cleaved-caspase-3） by TUNEL labeling and Western blot, respectively. Results The induced seizure peaked on an average between 50 -150 min after Li-pilo administration, scored using a modified Racine scale. The average scores of modified Racine scale in the IHP-3d seizure group was significantly lower than that in the other groups. The latency period and percentage of generalized seizures in the IHP-3d seizure group rats were significantly different from the parameters in the seizure group rats （P 〈 0. 05）. IHP-3d seizure rats showed lower escape latency, neuronal apoptosis counts and higher percentage of time in the probe quadrant compare with the seizure group and the other three IHP-seizure groups （ P 〈 0.05 ）. Compared with the control group, the parameters of water maze and apoptosis detection in the IHP-3d seizure group showed no significant changes （P 〉 0. 05 ）. Conclusions The results indicate that IHP treatment may help to decrease the susceptibility to epilepsy by reducing abnormal apoptosis, and has a brain protective effect on the seizure rats.