【作者】 张冬青；

【导师】 孙若鹏；

【作者基本信息】 山东大学 ， 儿科学， 2008， 硕士

【摘要】 第一部分拉莫三嗪对癫痫幼鼠认知功能的影响目的癫痫是一种常见的慢性神经系统疾病,部分癫痫患者伴有认知功能的损害。很多学者认为其认知的损害与抗癫痫药物的副作用有关。随着新型抗癫痫药物拉莫三嗪在临床的广泛使用,其对认知的影响也越来越引起人们的关注。本研究利用海人酸致癫痫持续状态后形成的慢性自发性颞叶癫痫幼鼠模型,初步探讨了长期应用拉莫三嗪对发育期大脑空间学习记忆能力的影响。方法将出生后7d的雄性wistar大鼠65只随机分为海人酸(KA)组33只和对照(control)组32只,KA组给予海人酸(kainic acid,KA)1mg/kg(浓度0.5mg/ml)腹腔注射,control组只腹腔注射相同剂量生理盐水。按照Lado幼鼠癫痫发作分级标准,腹腔注射后连续观察8h,癫痫发作达5级以上癫痫持续状态的大鼠若两周后出现自发性反复惊厥发作则为造模成功。将KA组中造模成功的26只存活大鼠随机分为癫痫未治疗(EP)组13只、癫痫拉莫三嗪治疗(EP+LTG)组13只,control组随机分为生理盐水(NS)组、生理盐水拉莫三嗪治疗(NS+LTG)组16只。治疗组均于自发性发作出现一周后给予抗癫痫新药拉莫三嗪治疗,观察大鼠癫痫行为的改变,在用药5周后对大鼠进行Morris水迷宫行为测试以评价其视觉-空间学习和记忆能力。结果KA组大鼠在注射海人酸2～10min后出现连续性点头动作及交替前肢痉挛,平均潜伏时间4.3±3.1min,15～20min后出现双侧前肢痉挛、后退、摔倒及强直发作等,随后发展为癫痫持续状态,持续约1～1.5h。注射KA后8小时内大鼠死亡5只,24h后又死亡2只。注射生理盐水的对照组均未发生惊厥,也无死亡。KA组大鼠自注射海人酸2周后陆续出现SRS,LTG用药结束前一周内EP组平均每只大鼠出现SRS 5.94±1.53次,EP+LTG组有2只未见发作,平均每只大鼠出现SAS 2.18±1.16次,两组比较差异具有显著性(P＜0.001)在Morris水迷宫测试中,与NS组相比,EP组大鼠逃逸潜伏期明显延长,在目的象限停留时间缩短(P＜0.05),EP+LTG组大鼠较EP组大鼠逃逸潜伏期明显缩短,在目的象限停留时间延长(P＜0.05),LTG对未经海人酸致痫的大鼠水迷宫操作无影响(P＞0.05).结论拉莫三嗪能有效抑制海人酸致痫大鼠的反复惊厥发作,具有明显抗癫痫作用,且能改善发育期癫痫大鼠的空间学习记忆能力。第二部分拉莫三嗪对癫痫幼鼠海马中多药耐药基因表达的影响目的癫痫患者中约有四分之一对大部分抗癫痫药物耐药从而发展为难治性癫痫。多药耐药基因(multipledrug resistant gene-1,MDR1)及其基因产物P-糖蛋白(P-gp)的高表达被认为与癫痫耐药密切相关。近年学者们多以成年大鼠急性期癫痫持续状态作为研究多药耐药基因的模型,但许多研究表明,在个体发育不同阶段,癫痫的产生、维持以及对抗癫痫药物的反应不尽相同,因此我们以海人酸致癫痫持续状态后形成的慢性自发性颞叶癫痫幼年大鼠模型为研究对象,探讨多药耐药基因及其表达产物在大鼠海马中的表达及抗癫痫新药拉莫三嗪对其表达的影响。方法将出生后7d的雄性wistar大鼠65只随机分为海人酸(KA)组33只和对照(control)组32只,KA组给予海人酸(kainic acid,KA)1mg/kg(浓度0.5mg/ml)腹腔注射,control组只腹腔注射相同剂量生理盐水。按照Lado幼鼠癫痫发作分级标准,腹腔注射后连续观察8h,癫痫发作达5级以上癫痫持续状态的大鼠若两周后出现自发性反复惊厥发作则为造模成功。将KA组中造模成功的26只存活大鼠随机分为癫痫未治疗(EP)组13只、癫痫拉莫三嗪治疗(EP+LTG)组13只,control组随机分为生理盐水(NS)组、生理盐水拉莫三嗪治疗(NS+LTG)组16只。治疗组均于自发性发作出现一周后给予抗癫痫新药拉莫三嗪治疗6周,然后将所有大鼠断头取海马,用RT-PCR法测定多药耐药基因mdrla和mdrlb mRNA的表达。结果EP组、EP+LTG组的mdrla和mdrlb mRNA表达均比NS组明显增高(P＜0.001)EP+LTG组及NS+LTG组的mdrla和mdrlb mRNA表达分别较EP组及NS组增高,但无统计学意义(P＞0.05)。结论反复癫痫发作可使癫痫幼鼠海马中mdrla、mdrlb mRNA表达增加,抗癫痫新药拉莫三嗪对癫痫及正常幼鼠海马中mdrla、mdrlb mRNA的表达无明显影响。更多还原

【Abstract】 ObjectiveEpilepsy is a common chronic disease of nervous system.Some epilepsy patients have impaired cognitive abilities.Many scholars hold that the antiepileptic drugs may contribute to this impairment.With the widely clinical usage of lamotrigine,the effect of lamotrigine on connitive has attracted more and more attention.In this study,an immature murine model of chronic spontaneous temporal epilepsy,which is attributed to statural epilepticus induced by kainic acid,was used to investigate the chronic effect of lamotrigine on spatial learning and memory.MethodsThe number of 65 male young rats of 7 days old were randomly divided into experimental group and control group.Rats in the experimental group were received intraperitoneal injection of 1 mg/kg kainic acid to induce seizures,and control rats were injected with the same dose of sodium chloride.According to Lado standard classification of seizures,those young rats whose seizure degree were beyond five and became status epilepticus after intraperitoneal injection were used as successful seizure models if they caught the spontaneous seizures after two weeks.When spontaneous seizures were developed,the 26 survived epileptic rats were divided into EP group and EP+LTG group.Rats of control group were divided into NS group and NS+LTG group.Rats of EP+LTG group and NS+LTG group were treated with therapeutic dose of lamotrigine after the spontaneous seizures developed for a week.Spontaneous recurrent seizures were recorded everyday.Spatial learning and memory ability was evaluated by Morris water maze after 5 weeks drug treatment.ResultsIn KA group,the rats received intraperitoneal injection of kainic acid suffered from epileptic seizure after 2-10 min injection.The latent time was average 4.3±3.1min.Five rats died whin eight hours and two rats died after 24 hours.In control group,no rats suffered from seizures or died.Rats of KA group gradually developed spontaneous recurrent seizure (SRS)2 weeks after kainic acid injection.During the last week of Lamotrigine treatment,in EP group,the mean times of SRS was 5.94±1.53. In EP+LTG group,two rats did not suffer from SRS,and the mean times of SRS was 2.18±1.16.There was significant difference between the two groups(P＜0.05).In the test of Morris water maze,the EP group spent the most time in finding the platform but the shortest time in the target quadrant. Compared with the EP group,EP+LTG group spent less time in finding the platform and the more time in the target quadrant(P＜0.05).LTG had no effect on water maze performance to rats without KA induction.ConclusionLTG can control the spontaneous recurrent seizure of spontaneous epileptic immature rats perfectly.For developing epileptic rat brain, LTG can improve the ability of Spatial Learning and Memory. ObjectiveAbout 25%of patients with epilepsy suffer from intractable seizures,which cannot be properly controlled by antiepileptic drugs. Overexpression of multidrug resistance gene and p-glycoprotein is thought to have close relationship with Refractory Epilepsy. Recently,many scholars used the murine model of status epilepicus to study the multipledrug resistant gene,nevertheless,some research showed that the production and maintenance of epilepsy is not similar in different periods of ontogeny.In this study,childhood rat model of spontaneous seizures induced by kainic acid was used to explore whether the new antiepileptic drug-lamotrigine affects the expression of mdr1a,mdr1b and P-Glycoprotein(P-gp)in the hippocampus of spontaneous seizures rat.MethodsThe number of 65 male young rats of 7 days old were randomly divided into experimental group and control group.Rats in the experimental group were received intraperitoneal injection of 1 mg/kg kainic acid to induce seizures,and control rats were injected with the same dose of sodium chloride.According to Lado standard classification of seizures,those young rats whose seizure degree were beyond five and became status epilepticus after intraperitoneal injection were used as successful seizure models if they caught the spontaneous seizures after two weeks.When spontaneous seizures were developed,the 26 survived epileptic rats were divided into EP group and EP+LTG group.Rats of control group were divided into NS group and NS+LTG group.Rats of EP+LTG group and NS+LTG group were treated with therapeutic dose of lamotrigine after the spontaneous seizures developed for a week.All rats were killed at the 42th day of administration.Mdr1a and mdr1b mRNA in the hippocampus was measured by RT-PCR.ResultsCompared with the control groups,expression of mdr1a and mdr1b mRNA in the hippocampus increased significantly in EP and EP+LTG group(P＜0.001)Mdr1a and mdr1b mRNA expression level of EP+LTG group was higher than that of EP group,and NS+LTG group higher than NS group,but there was no statistical difference(P＞0.05).ConclusionFrequent seizures results inoverexpression of mdr1a,mdr1b mRNA in the hippocampus.Lamotrigine dose not enhance the expression of mdr1a,mdr1b mRNA in the hippocampus.更多还原