SD大鼠原代海马神经元的一种优化培养方法  

An optimized culture method of primary hippocampal neurons in SD rats

作　　者：强婷婷 胡宪文[1] 张丽 QIANG Ting-ting;HU Xian-wen;ZHANG Li(Dept of Anesthesiology and Perioperative Medicine,the Second Hospital of Anhui Medical University,Key Laboratory of Anesthesiology and Perioperative Medicine of Anhui Higher Education Institutes,Anhui Medical University,Hefei 230601,China)

机构地区：[1]安徽医科大学第二附属医院麻醉与围术期医学科,麻醉与围术期医学安徽普通高校重点实验室,安徽合肥230601

出　　处：《中国药理学通报》2022年第6期956-960,共5页Chinese Pharmacological Bulletin

基　　金：安徽省自然科学基金资助项目(No 1908085QH358);安徽医科大学第二附属医院国家自然科学基金孵育计划(No 2020GQFY01)。

摘　　要：目的建立一种稳定、高效的无血清胎鼠海马神经元原代培养方法,以此来获得纯度更高、活力更好的海马神经元。方法取孕(18±1)d SD大鼠胎鼠海马组织,经消化吹打制成细胞悬液,种植于Neurobasal或DMEM/F12培养基后分为:无血清培养组、胎牛血清培养组、胎牛血清+5-氟-2′脱氧尿苷培养组。3组细胞培养12 h后均全量换液为Neurobasal培养基,FUDR培养组神经元培养至d 3时加入FUDR抑制胶质细胞生长。观察3组海马神经元在不同培养时间点下的生长状态;检测神经元活力、细胞凋亡率以及鉴定海马神经元纯度。结果与胎牛血清培养组相比,无血清培养组培养出来的神经元活力更高,细胞凋亡率低,神经元纯度更高;然而加入FUDR使神经元活力降低,细胞凋亡率增加。结论无血清的培养方法培养出来的神经元活性好,纯度高,且不需要加入胶质细胞抑制剂,是一种可行性较强、较为优化的海马神经元原代培养方法。Aim To establish a stable and efficient method for the primary culture of hippocampal neurons from serum-free fetal rats,so as to obtain hippocampal neurons with higher purity and better vitality.Methods The hippocampal tissues of SD rat fetus rats on the(18±1)day of pregnancy were cultured by Neurobasal or DMEM/F12 medium and divided into serum-free culture group,fetal bovine serum culture group and 5-fluoro-2′deoxyuridine culture group.After cells of three groups were cultured for 12 hours,all medium was changed to Neurobasal medium.When the neurons in FUDR culture group were cultured to 3 days,FUDR was added to inhibit the growth of glial cells.The growing status of hippocampal neurons at different culture time points was observed,neuronal activity was detected,cell apoptosis was assessed,and the purity of hippocampal neurons was identified.Results Compared with 10%serum culture group,the neurons cultured in serum-free culture group showed higher viability,lower apoptosis rate and higher purity;FUDR reduced cell viability and increased cell apoptosis.Conclusions The neurons cultured by serum-free culture have good activity and high purity,instead of adding glial cell inhibitors,which is a feasible and optimized primary culture method for hippocampal neurons.

关 键 词：海马神经元 原代培养 高纯度 胶质细胞 FUDR 胎牛血清 

分 类 号：R-332[医药卫生] R322.8