非洲爪蟾蝌蚪脑神经细胞体外培养方法的建立  

Establishment of an in Vitro Method for Culturing Brain Neurons from Xenopus laevis Tadpoles

作　　者：颜世帅[1] 徐海明[1] 李岩[1] 秦占芬[1] 

机构地区：[1]中国科学院生态环境研究中心环境化学与生态毒理学国家重点实验室,北京100085

出　　处：《生态毒理学报》2010年第5期752-756,共5页Asian Journal of Ecotoxicology

基　　金：中国科学院知识创新重要方向性项目(No.KZCX2-YW-Q-02-05;No.KZCX2-YW-420-3);国家自然科学基金项目(No.20677074)

摘　　要：体外培养的神经细胞是神经毒性机制研究和神经毒物筛查的重要材料.目前鼠体外神经细胞的应用最为广泛.鉴于近年来发育生物学的模型动物非洲爪蟾越来越多地应用于毒理学研究,论文建立了一种体外培养非洲爪蟾脑神经细胞的方法.该方法取52～53阶段的非洲爪蟾蝌蚪的脑组织,在L-15培养液中直接吹打获得分散细胞,接种于包被多聚赖氨酸的培养板中,22℃培养.培养细胞状态良好,72小时后神经元细胞初步建立神经网络.用β2tubulin7B9可对脑神经元细胞进行荧光染色,鉴别神经突的生长和神经网络的形成.与鼠脑神经元细胞体外培养方法比较,该方法具有无需胰蛋白酶分离、无需胶质细胞共培养、可同时操作多只脑、能获得大量细胞等简单快捷的特点.因此,体外培养的非洲爪蟾蝌蚪脑神经细胞可作为目前体外神经毒理研究中鼠脑神经元的补充材料,用于神经毒性机制的研究和神经毒物的筛查.Cultured neurons in vitro are important materials for studying the molecular mechanism of neurotoxicity and screening neurotoxicants.Neurons from rats and mice are the most widely used in neurotoxicology.Considering more and more application of Xenopus leavis,an animal model for developmental biology,in toxicology study,we established an in vitro method for culturing brain neurons from X.laevis tadpoles.Brains of X.laevis tadpoles at stage 52~53 were dissected and isolated in the L-15 medium by beating.Scattered cells were plated in poly-lysine-coated 24-well plate and cultured at 22℃.The neurons grew well and primary neuronal network was formed after 72h.Cultured neurons were stained by β2 tubulin 7B9 using immunofluorescence to identify neurite growth and neural network.Compared with the method for culturing rat/mouse brain neurons,this simple and rapid method for X.laevis brain neurons has some advantages,such as a separating process without trypsin,a culturing process without glia cells,and obtaining abundant neurons from several brains at the same time.Therefore,cultured brain neurons from X.laevis tadpoles could be used as an alternate material for rat/mouse brain neurons in studying neurotoxicological mechanisms and screening neurotoxicants.

关 键 词：非洲爪蟾 脑神经细胞 体外培养 神经毒性 

分 类 号：Q813.1[生物学—生物工程]