TrkB信号通路调控的研究.pdf

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1、TianjinMedJ，May2014，Vol42No5TrkB同亚型对癫痫海马神经元BDNF／TrkB信号通路调控的研究吴秋静常伟潘立平宋毅军赵文【摘要】目的探讨癫痫海马神经元中酪氨酸激酶受体B(TrkB)不同亚型对脑源性神经营养因子(BDNF)／TrkB信号通路的调控机制。方法取原代培养7d后的海马神经元，分为钙调蛋白抑制剂(ALLN)组和翻译抑制剂(An—isomycin)两大组。ALLN组又分为正常组、正常+BNDF组、癫痫组、癫痫+BDNF组、正常+ALLN组、癫痫+ALLN组和癫痫+ALLN+BDNF~；Anisomvcin组又分为正常组、正常+BDNF组、癫痫组

2、、癫痫+BDNF组、正常+Anisomycin组、癫痫+Anisomvcin组和癫痫+Anisomycin+BDNF组。免疫荧光鉴定海马神经元，无镁液处理制备癫痫模型，电生理鉴定细胞痫样放电，免疫印迹技术检测各组TrkB和磷酸化TrkB(p—TrkB)蛋白的表达变化。结果(1)ALLN组：正常+BDNF组p—TrkB／TrkB灰度值高于正常组；癫痫+BDNF组高于癫痫组，低于正常+BDNF组；癫痫+ALLN+BDNF组低于癫痫+BDNF组，与癫痫+ALLN组差异无统计学意义。(2)Anisomycin组：正常+BDNF组p-TrkB／TrkB灰度值高于正常组；癫痫+BDNF组

3、高于癫痫组，低于正常+BDNF组；癫痫+Anisomycin+BDNF组高于癫痫+BDNF组和癫痫+Anisomy—cin组。结论通过Anisomycin降低TrkB．T表达可改善癫痫状态下BDNF／TrkB受抑制的状态，通过ALLN升高TrkB．FL表达无法改善其抑制状态。【关键词】癫痫，颞叶；受体，TrkB；脑源性神经营养因子；茴香霉素；海马；神经元；钙调蛋白抑制剂【中图分类号】R742．1【文献标识码】A【DOI】10．3969／jAssn．0253—9896．2014．05．002AnExperimentalStudyoftheRegulationofBDNF／Trk

4、BSignalPathwaybyDifferentIsoformsofTrkBjnEpilepticHippocampalNeuronswuQiujing，CHANGWei，PANLiping，SONGYijun，ZHAOWenDepartmentofNeurology，theGeneralHospitalofTianjinMedicalUniversity,Tianjin300052，China【Abstract】0bjectiveToinvestigatethemechanismofbrainderivedneurotrophicfactor(BDNF)regulated

5、bydiffer—entisoformsoftyrosinekinasereceptorB(TrkB)inepileptichippocampalneurons．MethodsPrimaryhippocampalneu·ronswereculturedinvitrofor7days，anddividedintotwogroups，ALLN(calcineurininhibitor)groupandAnisomycin(trans—lationinhibitor)group．ALLNgroupincludedcontrolgroup，control+BDNFgroup，epil

6、epsygroup，epilepsy+BDNFgroup，control+ALLNgroup，epilepsy+ALLNgroupandepilepsy+ALLN+BDNFgroup．Anisomycingroupwassub—dividedintocon—trolgroup，control+BDNFgroup，epilepsygroup，epilepsy+BDNFgroup，contml+Anisomycingroup，epilepsy+Anisomycingroupandepilepsy+Anisomycin+BDNFgroup．Theimmunofluorescentt

7、echniquewasusedtoidentificatethehippocampalneurons．Epileptiformdischargesweredetectedbyelectrophysiol0gica1techniques．Westernblotassaywasusedtodeter—minetheproteinexpressionofTrkBandphosphorylatedTrkB(p—TrkB)inallcellgroups．Results(1)InALLNgroup，thegrayv