介导内吞功能的dynamin1与癫痫的相关性研究

发布时间：2018-03-17 17:07

本文选题：颞叶癫痫　切入点：内吞　出处：《重庆医科大学》2014年硕士论文　论文类型：学位论文

【摘要】：目的：本研究旨在了解锂-匹鲁卡品诱导的癫痫发作大鼠模型和颞叶癫痫患者中dynamin1表达水平,并将dynamin1特异性抑制剂运用于体内和体外实验以探究dynamin1与癫痫发作的关系。方法:课题组前期收集癫痫患者及对照组人脑组织,在癫痫患者和对照组人脑组织中,用逆转录PCR检测dynamin1基因的mRNA水平,并用western blot和免疫荧光法,测定dynamin1在人脑组织的蛋白表达水平和表达部位。对于匹鲁卡品诱导建立的大鼠癫痫模型,用上述相同实验测定动物脑组织中dynamin1表达情况。侧脑室注射dynamin1特异抑制剂dynasore后再造模观察大鼠模型行为学变化,并采用膜片钳技术观察dynasore对痫样放电兴奋性的影响。结果：研究发现匹鲁卡品诱导的大鼠癫痫模型中不同时间点(1天,3天,7天)和对照组相比, dynamin1的mRNA水平和蛋白水平较对照组均增高，，且癫痫患者与对照组比较，dynamin1的mRNA水平和蛋白水平明显高于对照组。Dynamin1主要在神经元的胞浆和轴突表达,星形胶质细胞未见表达。研究发现dynasore为10μM组大鼠癫痫发作潜伏期明显延长,发作程度减轻，发作频率降低。全细胞膜片钳记录到用无镁外液诱导出持续痫样放电后，浓度为10μM的dynasore能明显减少动作电位频率。但在行为学和膜片钳实验中，一致发现dynasore浓度为1μM和160μM时与DMSO组和生理盐水组未见明显差异。结论：我们的研究发现在癫痫动物模型和癫痫患者中，dynamin1的mRNA水平和蛋白水平均升高，其特异性抑制剂dynasore在特定浓度（10μM）下具有在体内匹罗卡品/体外癫痫脑片模型抗癫痫发作作用，说明dynamin1可能和癫痫发作相关，其特异性抑制剂dynasore具有抗癫痫发作的效应，但其具体抗癫痫发作机制有待进一步研究。
[Abstract]:Aim: to investigate the expression of dynamin1 in rat model of lithium-pilocarpine induced epilepsy and temporal lobe epilepsy, and to investigate the relationship between dynamin1 and seizure by using dynamin1 specific inhibitor in vivo and in vitro. Methods: the human brain tissues of epileptic patients and control group were collected. The mRNA level of dynamin1 gene was detected by reverse transcriptase PCR (RT PCR) and western blot and immunofluorescence assay in epileptic and control human brain tissues. The protein expression level and expression site of dynamin1 in human brain tissue were determined. The expression of dynamin1 in brain tissue was measured by the same experiment mentioned above. The behavioral changes of rat model were observed after the injection of dynasore, a specific inhibitor of dynamin1, and the effect of dynasore on the excitability of epileptiform discharge was observed by patch clamp technique. Results: compared with the control group, the mRNA and protein levels of dynamin1 in the rat epilepsy model induced by pilocarpine were higher than those in the control group at different time points (1 day, 3 days and 7 days), and compared with the control group, the levels of mRNA and protein were higher than those of the control group. The levels of mRNA and protein in epileptic patients were significantly higher than those in control group. The expression of dynamin1 was mainly found in the cytoplasm and axon of neurons, but not in astrocytes. It was found that the latency of epileptic seizure was prolonged in rats with 10 渭 M dynasore. After sustained epileptiform discharge was induced by magnesium free solution, the frequency of action potential was significantly reduced by 10 渭 M dynasore. However, in behavioral and patch clamp experiments, the frequency of action potential was significantly decreased in behavioral and patch clamp experiments. It was found that there was no significant difference between DMSO group and normal saline group when the concentration of dynasore was 1 渭 M and 160 渭 M. Conclusion: our study found that the levels of mRNA and protein in epileptic animal model and epileptic patients were increased. Dynasore, a specific inhibitor, has antiepileptic effects on pilocarpine / epileptic brain tablets in vivo and in vitro at a specific concentration of 10 渭 M, suggesting that dynamin1 may be associated with epileptic seizures, and its specific inhibitor, dynasore, has an antiepileptic effect. However, the mechanism of antiepileptic seizure needs further study.
【学位授予单位】：重庆医科大学
【学位级别】：硕士
【学位授予年份】：2014
【分类号】：R742.1

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