西罗莫司抗红藻氨酸诱发的未成年C57BL/6小鼠癫痫的有效剂量  

Effective dosage of sirolimus for seizure treatment of immature C57BL/6 mice induced by kainic acid

作　　者：吴美玲[1] 杨鑫杰 刘福荣[1] 王昱植 陈丹娇 吴赟[1] 朱锋[1] 曾玲晖[1] 

机构地区：[1]浙江大学城市学院医学院,浙江杭州310015

出　　处：《中国药理学与毒理学杂志》2017年第1期51-58,共8页Chinese Journal of Pharmacology and Toxicology

基　　金：国家自然科学基金(81371429);杭州市科技局重大科技创新项目(20152013A02);杭州市科技局社会公益项目(20160533B73)~~

摘　　要：目的探索西罗莫司(Sir)治疗未成年小鼠癫痫的安全有效剂量及作用。方法 10日龄的C57BL/6小鼠单次ip给予卡莫酸(红藻氨酸,KA)12.0 mg·kg-1诱导癫痫,于造模后24 h隔天分别ip给予Sir 0.3,1.0和3.0 mg·kg-1直至3 d、7 d、3周、5周和6周,采用Western蛋白印迹法检测S6蛋白表达及其磷酸化水平,确定最低有效剂量。观察Sir最低有效剂量对学习记忆和生长发育的影响,其中Doublecortin(DCX)免疫荧光染色检测海马神经元发育,Morris水迷宫测定小鼠学习记忆水平,悬尾、O迷宫和新事物认知实验测定小鼠焦虑抑郁状态。结果 Western蛋白印迹结果显示,Sir 0.3 mg·kg-1对正常小鼠和KA致痫小鼠S6蛋白磷酸化无显著影响,而1.0和3.0 mg·kg-1均可显著抑制KA致痫小鼠S6蛋白磷酸化(P<0.05)。选取Sir 1.0 mg·kg-1作为最低有效剂量。Sir 1.0 mg·kg-1对DCX表达和体质量无显著影响。Morris水迷宫结果显示,与正常对照组相比,模型组小鼠平台潜伏期和游泳距离明显延长(P<0.05),穿越平台象限次数明显下降(P<0.05),而Sir 1.0 mg·kg-1可显著逆转KA致痫模型小鼠学习记忆功能下降(P<0.05),且与正常对照组无显著差异。KA致痫模型小鼠在悬尾实验中显示悬尾不动时间明显上升(P<0.05),O迷宫实验中移动距离、开臂中停留时间以及新事物认知实验中与新事物的触碰时间、触碰频率、移动距离和速度明显缩短(P<0.05),而Sir1.0 mg·kg-1可显著逆转KA致痫模型小鼠的焦虑抑郁状态(P<0.05),且与正常对照组无显著差异。结论Sir 1.0 mg·kg-1对未成年期癫痫小鼠的m TOR信号通路异常激活和癫痫后共病形成有抑制作用,且不良反应轻微,为理想的给药剂量。OBJECTIVE To explore the safe and effective dose of sirolimus（Rapamycin,Sir） and its effect on seizure comorbidities. METHODS Immature C57BL/6 mice at postnatal 10 d of age were administered with kainic acid（KA） 12.0 mg·kg（-1）intraperitoneally by a single injection to induce acute seizure. Sir 0.3, 1.0 and 3.0 mg·kg（-1）was injected 24 h after seizure every other day until 3 d, 1 week,3 weeks, 5 weeks and 6 weeks. Western blotting analysis was used to detect the expression and phosphorylation level of S6 protein and to determine the minimum effective dose of Sir. Effect of the minimum effective dose of Sir on cognitive function and body growth was observed by several evaluations.Immunofluorescent intensity of Doublecortin（DCX） immunofluorescent staining was conducted to evaluate the development of neurons in the hippocampus. Morris water maze was used to assess the cognitive function. Tail suspension test, O maze and new object recognition test were used to study the anxiety-like behaviors of mice. RESULTS The result of Western blotting showed that Sir 0.3 mg·kg（-1） had no significant effect on the phosphorylation of S6 protein in normal mice or KA mice, whereas 1.0 and3.0 mg · kg-1could significantly inhibit the phosphorylation of S6 protein in KA mice（P〈0.05）. Sir1.0 mg·kg（-1）had no obvious effect on DCX-positive cells or body wass. Morris water maze showed that KA-induced seizure resulted in prolonged escape latency and swimming length（P〈0.05）, and a decreased crossing number of target quadrant（P〈0.05）. Sir 1.0 mg·kg（-1）significantly reversed the deficit of cognitive function of KA-induced seizure mice（P〈0.05）, whereas no significant difference was found between Sir group and normal control group. Compared with normal control group, model group showed increased freezing time in tail suspension test（P〈0.05）, decreased migration length and retention time in open arms in O maze（P〈0.05）, decreased retention time and touch frequency with new o

关 键 词：癫痫 西罗莫司 学习认知功能 蛋白表达 

分 类 号：R742.1[医药卫生—神经病学与精神病学]