谷氨酸对小鼠胚胎中脑原代细胞培养中多巴胺能神经元损伤的时程效应  

Time-course effect of glutamate on morphological changes of dopaminergic neurons in mouse mesencephalic dissociated cultures

作　　者：贾晓晶[1] 张秦玲[1] 侯吉光[1] 翟成伟[1] 董素光[1] Wolf-Dieter Rausch 王铁君[1] 

机构地区：[1]吉林大学第二医院肿瘤放射治疗科,吉林长春130041 [2]奥地利维也纳兽医大学医学免疫化学研究所

出　　处：《吉林大学学报（医学版）》2010年第6期1075-1078,1206,共5页Journal of Jilin University：Medicine Edition

基　　金：中国科技部中奥(地利)国际交流合作项目资助课题(Ⅶ15A)

摘　　要：目的:探讨谷氨酸兴奋性损伤诱导小鼠胚胎中脑原代培养多巴胺能神经元损伤的时程效应,阐明谷氨酸兴奋性毒性在帕金森病(PD)模型中对多巴胺能神经元损伤的作用。方法:取孕14d小鼠胚胎的中脑组织制备成细胞悬液,进行原代培养,分为对照组和谷氨酸用药组。细胞在体外培养第10天,加入500μmol·L-1谷氨酸并作用15min,分别于用药后2、4、6、24和48h进行酪氨酸羟化酶(TH)免疫细胞化学染色。在显微镜下计数TH染色阳性神经元数量及每个神经元突起数量。结果:对照组中每个培养孔多巴胺能神经元的数量平均为(778.0±18.6)个,谷氨酸用药组2、4、6、24和48h多巴胺能神经元数量平均为(306.0±8.6)、(314.0±15.4)、(298.0±19.1)、(307.0±18.5)及(323.0±23.8)个。谷氨酸用药各组中多巴胺能神经元数量较对照组明显减少(P<0.05),但谷氨酸用药组2、4、6、24和48h多巴胺能神经元数量比较差异无显著性(P>0.05)。随机选取每组100个多巴胺能神经元的突起进行计数,对照组多巴胺能神经元的突起数量平均为(3.440±0.106)个,谷氨酸用药组2、4、6、24和48h时多巴胺能神经元突起数量分别为(2.240±0.105)、(2.390±0.103)、(3.070±0.105)、(3.420±0.987)和(3.420±0.923)个。谷氨酸用药后2和4h多巴胺能神经元的突起数量较其他各组均明显减少(P<0.05)。结论:谷氨酸可在体外诱导小鼠胚胎中脑原代培养中多巴胺能神经元的损伤和死亡,是一个相对急性、不可逆的损伤过程,但随着损伤后恢复时间的延长,损伤的多巴胺能神经元形态可恢复。Objective To investigate the time-course effect of glutamate on dopaminergic neurons injury in mouse mesencephalic dissociated culture,and elucidate the effect of the neurotoxicity of glutamate on dopamingergic neurons injury in Parkinson’s disease （PD）model.Methods Mouse embroysof14d wereusedto makemecsencephalic dissociated culture.The experiment was divided into control group and glutamate group.The cultures were treated with 500μmol·L-1 of glutamate for 15 min on the 10th DIV,and stained by TH immunocytochemistry 2,4,6,24and 48hafter treatment.The numbers of dopaminergic neurons and the average neurite numbers per dopaminergic neuron were evaluated by counting under microscope.Results The TH-ir neurons were counted and there were 778.0±18.6dopaminergic neurons per well in control group;there were 314.0±15.4,298.0±19.1,307.0±18.5,and 323.0±23.8dopaminergic neurons per well respectively at 2,4, 6,24and 48hin glutamate group,the TH-ir neurons in glutamate group were significantly reduced compared with control group （P0.05）.The average numbers of neurites per dopaminergic neuron were 3.440±0.106in control group,2.24 0±0.105,2.390±0.103,3.070±0.105,3.420±0.987and 3.420±0.923respectively at 2,4, 6,24and 48hin glutamate goroup.There were a significantly lower neurite numbers 2and 4hafter glutamate treatment,as compared with all the other groups（P0.05）.Conclusion The loss of dopaminergic neurons induced by glutamate is caused by a relative acute neural death and is irreversible.But the number of neurite and length of neurites are gradually improved as the recovery period is prolonged.

关 键 词：谷氨酸 帕金森病 中脑 原代培养 

分 类 号：R-332[医药卫生] R742.5