耳穴区经皮电刺激对颞叶癫痫大鼠癫痫发作频率及海马区白介素-1β、肿瘤坏死因子-α表达的影响

Effects of Transcutaneous Electrostimulation of Auricular Points on Behavior and Hippocampal IL-1β and TNF-α Expression in Temporal Lobe Epilepsy Rats

目的:通过观察经皮电刺激耳迷走神经支配的耳穴区对颞叶癫痫大鼠自发癫痫发作频率及海马区炎性因子白细胞介素1β(interleukin-1β,IL-1β)、肿瘤坏死因子-α(tumor necrosis factor-α,TNF-α)蛋白及mRNA表达的影响,探讨耳穴刺激对颞叶癫痫的缓解效应及其可能的分子机制。方法:SD大鼠随机分为正常组、模型组、治疗组,每组12只。采用氯化锂-匹罗卡品腹腔注射复制慢性自发性颞叶癫痫大鼠模型,对治疗组大鼠耳迷走神经支配的"心""肺""皮质下"耳穴区进行经皮电刺激干预,每日1次,每次20min,连续6周。用长时程视频监测动物的癫痫发作情况,治疗后采用免疫荧光与荧光定量real-time PCR方法观察癫痫大鼠海马区IL-1β、TNF-α蛋白及mRNA表达情况。结果:颞叶癫痫大鼠有反复自发的癫痫发作,模型组大鼠的自发癫痫发作频率随时间延长有增加的趋势,治疗组大鼠的自发癫痫发作频率治疗后较治疗前显著减少(P<0.05)。与正常组比较,模型组大鼠海马CA 1、CA 3区IL-1β、TNF-α免疫活性增强,免疫阳性细胞数明显增多(P<0.05),海马内IL-1β、TNF-αmRNA表达水平显著上调(P<0.01);与模型组比较,治疗组大鼠海马CA 1、CA 3区IL-1β与TNF-α蛋白的免疫活性降低,免疫阳性细胞数明显减少(P<0.05),海马内IL-1β、TNF-αmRNA表达水平显著下调(P<0.05,P<0.01)。结论:耳迷走神经支配的耳穴区经皮刺激干预能显著降低颞叶癫痫大鼠反复自发的癫痫频率,此效应可能与下调海马区炎性因子IL-1β、TNF-α蛋白的免疫反应及mRNA表达水平有关,即经皮耳迷走神经刺激的抗癫痫效应可能与其海马内的抗炎作用有关。

Objective To observe the effect of transcutaneous otopoint electrostimulation（TCOES）on behavior and expression of hippocampal interleukin-1β（IL-1β）and tumor necrosis factor-α（TNF-α）expression in lithium-pilocarpine induced chronic spontaneous temporal lobe epilepsy（TLE）rats,so as to investigate its antiepileptic mechanism.Methods Thirty-six SD rats were randomly divided into control,model and TCOES groups（n=12in each group）.The epilepsy model was established by intraperitoneal injection of lithium chloride（127.2mg/kg）,scopolamine（1mg/kg,20 h after the 1st injection）and pilocarpine（10mg/kg,30 m in after scopolamine injection）.Rats of the control group were treated by injection of normal saline（i.p.i.）.Transcutaneous electrostimulation（1mA,20Hz）was applied to bilateral otopoints＂Heart＂ ＂Lung＂ and ＂Subcortex＂for 20 min,once daily for 6weeks except the weekends.The behavior reactions were observed by a video monitoring system.The expression of IL-1β and TNF-αproteins and genes in the hippocampus were determined by immunofluorescence and quantitative real-time PCR,separately.Results Behavioral observation showed that after TCOES intervention,the frequency of epilepsy attack was significantly decreased in comparison with pre-treatment（P〈0.05）.Immunofluorescence and real-time PCR showed that compared with the control group,the immunoactivity of IL-1βand TNF-αin both hippocampal CA 1and CA 3 regions and hippocampal IL-1β and TNF-α gene expression were obviously increased in the model group（P〈0.05,P〈0.01）.Following TCOES,the increased hippocampal IL-1β and TNF-αand IL-1β mRNA and TNF-α mRNA expression levels were all suppressed（P〈0.05,P〈0.01）.Conclusion TCOES intervention has an antiepileptogenic effect in temporal lobe epilepsy rats,which may be related to its effects in down-regulating expression of proinflammatory cytokine IL-1β and TNF-α in the hippocampus.