红藻氨酸致痫大鼠海马神经元凋亡及其与caspase-3关系的研究

目的　研究大鼠癫痫发作后海马神经元凋亡及其与天冬氨酸特异性半胱氨酸蛋白酶 -3 (cysteinylasparate-specific proteinase,caspase-3 )表达的关系。方法　采用红藻氨酸 (kainic acid,KA)诱导大鼠癫痫模型 ,以原位末端标记 (TUNEL)及透射电镜检测癫痫发作后 6h及 1、3、7d海马神经元凋亡 ;半定量 RT-PCR及免疫组化法检测 caspase-3 m RNA及 caspase-3阳性表达。结果　KA致痫后 1 d,海马 CA1、CA3及 CA4区开始出现凋亡细胞 ,3 d时明显增多 ,7d时最多。 3个时间组相应区域间凋亡神经元数比较差异均有显著性 (P<0 .0 0 1 )。透射电镜观察可见典型的凋亡细胞形态学改变。 RT-PCR结果显示 ,KA致痫后 6h,海马组织 caspase-3 m RNA表达较对照组显著增高 (P <0 .0 5 ) ,1、3、7d caspase-3 m RNA仍持续高水平表达 (P <0 .0 5 )。免疫组化结果显示 ,KA致痫后 1 d,海马 CA1、CA3、CA4区开始出现 caspase-3阳性表达 ,3 d时阳性表达进一步增强 ,7d时表达最强。结论　凋亡参与 KA致痫大鼠癫痫发作后海马神经元迟发性死亡过程 ,caspase-3可能在癫痫后神经元凋亡过程中具重要的作用。     

红藻氨酸对大鼠海马锥体细胞钙电流的影响

目的：应用膜片钳技术,观察红藻氨酸（KA）对大鼠海马锥体细胞ca2＋电流的影响,以研究癫痫的发病机制。方法：采用酶加机械分离法制备出生10～12d的大鼠海马锥体神经元标本,用全细胞膜片钳技术测定其生理学特性及观察KA对ca2＋电流的影响。结果：分离出的海马锥体细胞形态正常,有较长突起;用膜片钳技术证实,其保存了主要的离子通道活性。KA20μmol／L和100μmol／L的浓度均可使海马锥体细胞ca2＋电流峰值增大（n=8,P〈0．01）。结论：①大鼠海马锥体神经细胞具有明显的突起,细胞膜表面光洁、晕光好,适用于膜片钳实验研究;②KA使ca2＋内流增加,引起“Ca”超载”导致细胞毒性等一系列反应;③KA通过激活α-氨基羟甲基恶唑丙酸（AMPA）受体,诱发快速的兴奋性突触后电位（EPSP）,参与兴奋性突触传递。AMPA受体的激活可能是癫痫的发病机制之一。     

癫痫发作敏感大鼠T区Bcl-2蛋白表达变化

为探讨前深梨状皮层T区中Bcl- 2蛋白的表达变化在红藻氨酸诱导的癫痫发作敏感性长期增强中的作用,本研究以免疫组化方法检测癫痫发作敏感大鼠前深梨状皮层T区Bcl- 2蛋白表达变化,以硫堇染色观察神经元脱失情况,并与经蝎毒处理后癫痫发作敏感性明显降低的大鼠进行比较。结果显示:与对照组比较,癫痫发作敏感大鼠前深梨状皮层T区Bcl 2蛋白免疫反应阳性细胞数目明显减少,染色强度明显降低(P<0. 05),神经元数量明显减少;以剂量为100mg/ (kg·d)的蝎毒给予动物连续灌胃4周,可明显降低其癫痫发作敏感性(P<0. 05),脑内前深梨状皮层神经元脱失程度减轻,Bcl- 2蛋白免疫反应产物与对照组相比无明显变化。以上结果提示,前深梨状皮层T区Bcl- 2蛋白表达减少可能是癫痫发作敏感性长期增强的重要原因之一。     

代谢型谷氨酸受体配体(s)-4C3HPG对红藻氨酸诱导性癫痫发作活动的调节研究

目的　探讨代谢型谷氨酸受体配体 (s) 4 羧基 3 羟苯基甘氨酸〔(s) 4C3HPG〕对红藻氨酸 (KA)诱导性癫痫发作活动的调节作用。方法　选用KA诱导大鼠复杂部分性发作模型 ,采用侧脑室注射 (s) 4C3HPG法 ,观察其对大鼠癫痫发作行为学、脑电图放电时间以及对大鼠海马的组织病理学影响。结果　 (s) 4C3HPG +KA组大鼠行为学评分与盐水 +KA组相比明显减少 ,在KA注射 40min之后差异有显著性 (P <0 0 1) ,湿狗样运动 (WDS)数目也减少 ,于KA注射后 40、6 0、80、10 0、12 0min差异有显著性 (P <0 0 1)。 (s) 4C3HPG +KA组大鼠EEG虽有癫痫样波发放 ,但放电持续时间较对照组缩短 ,在KA注射 30min之后差异有显著性 (P<0 0 1)。(s) 4C3HPG +KA组大鼠海马CA1、CA3区正常细胞数较对照组大鼠增多 (P <0 0 1) ,死亡或损伤细胞数要减少 (P <0 0 1)。结论　 (s) 4C3HPG可减轻KA诱导性癫痫发作的程度 ,具有神经保护性作用     

红藻氨酸诱导癫痫发作小鼠脑组织中NO^—2与硫代巴比妥酸反…

了解活性介质一氧化氮与癫痫发作的关系。用比色法检测红藻氨酸诱导ＢＡＬＢ／ｃ小鼠癫痫发作后，不同时点脑匀浆上清液中亚硝酸根与硫代巴比妥酸反应物的含量。ＮＯ＾－２浓度与ＴＢＡ反应物含量的多少与ＫＡ诱导的癫痫发作时程有关。在发作初期，ＮＯ＾－２随发作持续而增多，但发作后期又迅速减少。ＮＯ＾－２浓度的升高与减低受Ｌ－精氨酸及其硝基衍生物Ｎ＾Ｇ位硝基左型精氨酸的影响。     

雌激素对癫癎大鼠中枢神经系统Fos蛋白表达的影响及其作用部位的研究

目的 :了解雌激素致作用的部位。方法 :利用红藻氨酸和 6 氟二乙酯致的两种大鼠癫模型 ,应用免疫组化法检测单纯致及给予雌激素后再致大鼠海马 ,大脑皮质 ,纹状体的Fos表达。结果 :两种模型单纯致组海马、皮质、纹状体Fos表达较正常组显著增高 (P <0 0 1)。给予雌二醇 (E2 )后再致 ,红藻氨酸模型中海马 ,皮质Fos表达较单纯致组增加 (P <0 0 1,P <0 0 5 ) ;而 6 氟二乙酯模型中无变化。结论 :E2 促进癫发作的敏感性不是普遍增加全脑神经元兴奋的结果 ,而是与海马有关     

Effects of topiramate on hippocampal neuronal apoptosis in rats after kainic acid-evoked seizures*☆

BACKGROUND： Apoptosis plays an important role in brain injury after seizures and the formation of chronic epilepsy. It is important to investigate whether topiramate exhibits either antiepileptic and/or antiapoptotic effects on hippocampal neurons. OBJECTIVE： To observe neuronal apoptosis in hippocampus of rat seizure models, and to investigate the antagonizing effect of topiramate on neuronal apoptosis after seizures.
DESIGN： An animal experiment of comparative observation.
SETTING： First Affiliated Hospital of Guangxi Medical University. MATERIALS： Sixty healthy male Sprague Dawley （SD） rats, 4-6 weeks old and weighing 160-220 g, were provided by the Experimental Animal Center of Guangxi Medical University. Main apparatus and reagents were as follows： Rat brain solid positioner （SR-6N, made in Japan）; kainic acid by Sigma （USA）; pathological image analyzer （DMR＋550） by Leica （Germany）; in situ apoptosis detection kit by Wuhan Boster Biological Technology Co., Ltd; topiramate by Xi＇an-Janssen Pharmaceutical, Ltd. The treatment on animals in the experiment was in accordance with the standards of animal ethics. METHODS： The experiments were performed at the Scientific Experimental Center of Guangxi Medical University from June to December 2006. The rats were randomly divided into a topiramate-treated group （n = 30） and a model group （n = 30）. ① After anesthesia, all rats were administered a kainic acid injection （0.2 μL, 2 g/L） into the right lateral ventricle. Grade Ⅲ and greater Racine standards were considered to be a successful model establishment. Thirty minutes after seizure , rats in the topiramate-treated group were treated with an intraperitoneal （i.p.） injection of topiramate every day （40 mg/kg/d） for 2 weeks. The rats in the model group were treated with an equal volume of saline for 2 weeks. ③ Six rats in the topiramate-treated group were sacrificed at 1 day, and 1, 2, 3, and 4 weeks after treatment, respectively. The model group ani     

Effects of D2 receptor antagonist haloperidol on hippocampal neuronal apoptosis in a rat model of temporal epilepsy

BACKGROUND： Dopamine receptors are divided into D1 and D2 subgroups. It has been reported that D2 receptors resist neural toxicity induced by excitatory amino acids and muscarine, and also alleviate epilepsy attacks following pilocarpine treatment. However, it has not yet been established whether D2 receptors regulate temporal epilepsy. OBJECTIVE： To observe the effects of the D2 antagonist haloperidol on hippocampal neuronal apoptosis and electrical brain activity in a rat model of kainic acid-induced temporal epilepsy. DESIGN, TIME AND SETTING： Randomized grouping and histopathological study were performed at the Neurology Medicine Institute of Zhujiang Hospital, Southern Medical University from August to December 2004. MATERIALS： Twenty-five adult, male, Sprague Dawley rats were selected for the present study. Kainic acid （Sigma, USA） was injected into the right lateral ventricle to establish models of temporal epilepsy. A PowerLah multiplying channel eleetrophysiolograph was provided by AD Instruments, Australia. METHODS： The rats were randomly divided into 5 groups （n = 5）： control, model, haloperidol hippocampus, haloperidol striatum, and haloperidol substantia nigra. Temporal epilepsy was established in all rats except the control group. Haloperidol was slowly injected into the hippocampus, striatum and substantia nigra, respectively, in three different injection groups. Normal saline was injected into the right lateral ventricle of the control rats. MAIN OUTCOME MEASURES： Hippocampal apoptosis was observed on the day 3 of treatment using TUNEL staining. Changes in electroencephalogram at 0, 0.5, 2, 6, and 12 hours following treatment onset were observed using a PowerLab multiplying channel electrophysiolograph. Animal behaviors were classified according to the Racine criteria. RESULTS： Twenty-five rats were included in the final analysis. Seizures did not occur in the control group. In the model group, 10 minutes after kainic acid injection to the lateral cerebral ventricle, epilep     

癫痫患儿与实验性癫痫动物生长抑素的测定及意义

为揭示生长抑素在癫痫发生发展中的作用及其与癫痫患儿行为之间的关系，采用放射免疫法测定海人酸（ＫＡ）诱发癫痫动物模型脑组织与血浆生长抑素放射免疫活性（ＳＬＩ）。对２２例癫痫患儿进行血浆ＳＬＩ的检测和事件相关诱发电位的测查，并用Ａｃｈｅｎｂａｃｈ儿童行为量表进行评价。结果显示，动物额叶皮层内ＳＬＩ在注射ＫＡ后７天显著增高，可能与ＫＡ诱发动物癫痫敏感性增高有关。癫痫患儿血浆ＳＬＩ显著高于正常对照组，其行为异常发生率为４０．９％，Ｎ１００、Ｐ３００潜伏期均明显延长，血浆ＳＬＩ与攻击行为、多动行为呈显著负相关。提示癫痫发作后血浆ＳＬＩ的变化既反映了发作后生长抑素能神经元功能的改变，又可能是癫痫患儿行为异常或认知障碍的生物化学基础之一     

红藻氨酸致痫大鼠海马区神经元凋亡的动态变化

目的探讨红藻氨酸(kainic acid,KA)诱导的大鼠癫痫状态海马神经元的形态学变化、凋亡情况及抗痫药物的神经保护作用.方法90只Wistar大鼠随机分为对照组、KA组和卡马西平(CBZ)组,后两组再按癫痫发作后1 h、4h、12h、24h、48h和72h不同时点分为6个亚组.KA注射后,观察大鼠癫痫发作后的行为学变化;采用HE染色法观察大鼠癫痫状态海马CA1、CA3区神经元形态学改变;采用原位细胞凋亡检测法观察癫痫状态海马CA1、CA3区神经元凋亡情况.结果KA注射后,大鼠出现严重的惊厥;在癫痫发作后12h,海马CA1区、CA3区开始出现凋亡细胞[CA1区:KA组(6.53±1.36)个,CBZ组(5.85±1.68)个;CA3区:KA组(9.58±1.63)个,CBZ组(7.36±1.27)个],48h凋亡细胞达到峰值(P＜0.01)[CA1区:KA组(42.263±3.28)个,CBZ组(35.39±2.36)个;CA3区:KA组(57.64±12.76)个,CBZ组(38.37±13.65)个].经CBZ干预后凋亡细胞明显减少(P＜0.05).结论癫痫发作后的迟发性神经元死亡很可能是由凋亡引起的,CBZ可抑制癫痫状态海马神经元凋亡.