P2X3R在难治性颞叶癫痫患者及癫痫大鼠模型脑组织中表达变化的研究

发布时间：2018-02-11 18:41

  本文关键词： P2X3R 颞叶癫痫 兴奋性 钙离子　出处：《重庆医科大学》2016年硕士论文　论文类型：学位论文

【摘要】：目的:探讨P2X3R在难治性颞叶癫痫患者颞叶皮质和氯化锂-匹鲁卡品诱导的癫痫大鼠模型脑组织中表达的变化,通过膜片钳全细胞记录观察无镁脑脊液诱发S-D鼠海马CA1区锥体神经元动作电位频率加快,对P2X3R进行干预后,观察锥体神经元动作电位频率的改变并分析其可能的机制。方法:1.收集15例难治性颞叶癫痫患者病灶组织标本和10例脑外伤手术患者相对正常的颞叶皮质标本。2.称重后腹腔注射氯化锂-匹鲁卡品诱导S-D大鼠产生急性期癫痫持续状态,并通过视频监控选出2个月内产生自发发作的S-D大鼠作为慢性癫痫模型,对照组腹腔注射生理盐水进行对照。3.运用免疫荧光技术对P2X3R在颞叶癫痫患者颞叶组织和癫痫大鼠海马及邻近皮质的表达进行定位,通过Western-blot技术和反转录聚合酶链反应(RT-PCR)测定实验组与对照组P2X3R蛋白和相应m RNA表达量的变化。4.S-D鼠麻醉后断头取脑,运用振动切片机切出含有清晰海马结构的脑片,脑片厚度为350μm,并在26℃下人工脑脊液孵育1小时,之后运用膜片钳全细胞记录技术,通过激活或抑制P2X3R受体,观察无镁状态下诱发的快速动作电位频率的变化。结果:P2X3R在颞叶癫痫患者颞叶皮质和癫痫大鼠海马及邻近皮质的神经元树突和胞质中均有表达,并且P2X3受体在癫痫患者及癫痫大鼠模型中表达升高。无镁脑脊液可诱发S-D鼠海马CA1区锥体神经元兴奋性升高,引起快速样放电,抑制P2X3R受体后,锥体神经元动作电位频率减慢,激活受体后,电活动加快。结论:P2X3R在癫痫患者及癫痫大鼠模型中表达升高并且阻断P2X3R可抑制神经元过高的兴奋性,故我们推测升高的P2X3R在癫痫的形成过程中具有一定作用。
[Abstract]:Objective: to investigate the changes of P2X3R expression in the temporal cortex of patients with refractory temporal lobe epilepsy and the brain tissue of rats with epilepsy induced by lithium-pilocarpine. The frequency of action potential of pyramidal neurons in hippocampal CA1 region of S-D rats induced by magnesium free cerebrospinal fluid (CSF) was observed by whole-cell patch clamp recording. To observe the change of action potential frequency of pyramidal neurons and to analyze its possible mechanism. Methods: 1. 15 patients with intractable temporal lobe epilepsy and 10 patients with traumatic brain surgery were collected. After weighing, S-D rats were induced by intraperitoneal injection of lithium chloride and pilocarpine to produce acute epileptic status. S-D rats with spontaneous seizures within 2 months were selected as chronic epilepsy models by video surveillance. The expression of P2X3R in temporal lobe tissues of temporal lobe epilepsy and hippocampus and adjacent cortex of epileptic rats was determined by immunofluorescence technique. The changes of P2X3R protein and corresponding m RNA expression were measured by Western-blot and reverse transcriptase polymerase chain reaction (RT-PCR). 4. After anesthesia, brain slices with clear hippocampal structure were cut out by vibration slicer. The thickness of brain slices was 350 渭 m, incubated with artificial cerebrospinal fluid at 26 鈩，

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