中国组织工程研究

中国组织工程研究 ›› 2010, Vol. 14 ›› Issue (14): 2647-2651.doi: 10.3969/j.issn.1673-8225.2010.14.038

• 干细胞培养与分化 stem cell culture and differentiation • 上一篇    下一篇

原代培养大鼠视网膜与视皮质神经元的方法学特点

张  荻1，吴小影2，刘双珍2   

  1. 1河南省眼科研究所，河南省郑州市  450003；2中南大学湘雅医院眼科，湖南省长沙市  410008
  • 出版日期:2010-04-02 发布日期:2010-04-02
  • 通讯作者: 吴小影，教授，中南大学湘雅医院眼科，湖南省长沙市 410008
  • 作者简介:张 荻☆，女，1979年生，河南省郑州市人，汉族，2008年中南大学毕业，博士，主治医师，主要从事斜视与小儿眼科的研究。

Primary culture of rat retinal and visual cortical neurons: Methodological characteristics

Zhang Di1, Wu Xiao-ying2, Liu Shuang-zhen2   

  1. 1 Henan Eye Institute, Zhengzhou  450003, Henan Province, China; 2 Department of Ophthalmology, Xiangya Hospital of Central South University, Changsha  410008, Hunan Province, China
  • Online:2010-04-02 Published:2010-04-02
  • Contact: Wu Xiao-ying, Professor, Department of Ophthalmology, Xiangya Hospital of Central South University, Changsha 410008, Hunan Province, China wxy_64@yahoo.com.cn
  • About author:Zhang Di☆, Doctor, Attending physician, Henan Eye Institute, Zhengzhou 450003, Henan Province, China reedzhang79@yahoo.com.cn

PDF

468

摘要：

背景：神经元的原代培养是进行神经系统结构和功能研究的一种重要手段。如果能建立稳定良好的视网膜、视皮质神经细胞体外培养体系，对深入研究其各种细胞成分的生物学功能、病理改变过程与机制以及药物反应等十分重要。
目的：对比观察原代培养新生大鼠视网膜及视皮质神经元的特点，探求最佳分离和培养方法。
方法：分别采用机械分离及酶消化法分离新生大鼠视网膜及视皮质神经元，应用含体积分数为10%新生牛血清、10%F-12 Nutrient Mixtures的DMEM培养基进行接种培养，含2% B-27 Serum-Free Supplements的Neurobasal Medium进行维持培养，利用尼氏染色进行神经元鉴定。
结果与结论：培养的神经元生长良好，胞体饱满，突起长。尼氏染色示视网膜神经元比例大于90%，视皮质神经元比例大于50%。提示视网膜及视皮质神经元培养方法及生长特点存在不同，需采用不同的方法进行培养以获得高纯度的神经元。

关键词: 视网膜, 视皮质, 神经元, 原代培养, 大鼠

Abstract:

BACKGROUND: Primary culture of neurons is an important way to study the structure and functions of the nervous system. It is also important to explore pathomechanism and medicine reaction of some ophthalmology diseases.
OBJECTIVE: To explore an optimal way to the separation and culture of retinal and visual cortical neurons in new-born rats through comparative observation of different primary culture methods. 
METHODS: Retinal and visual cortical neurons isolated from new-born rats were firstly cultured in Dulbecco’s modified Eagle’s medium supplemented with 10% new-born calf serum and 10% F12 nutrient mixtures, followed by maintaining culture in Neurobasal medium containing 2% B27 serum-free supplements. Nissls staining was performed for neuron identification.
RESULTS AND CONCLUSION: Cultured neurons grew well with plump cell bodies and long processes. Nissls staining showed that the purity of retinal neurons was greater than 90% and the proportion of visual cortical neurons was higher than 50%. The results suggested that there are some differences in culturing methods and growth characteristics of retinal and visual cortical neurons of new-born rats, accordingly, different culture methods are required to obtain high purity neurons.

中图分类号: