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C. Romano, X. Luo; Effects of Media, Additives, and Serum on Neuronal Survival in Dissociated Cultures from Adult Mouse Retina . Invest. Ophthalmol. Vis. Sci. 2003;44(13):5248.
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© ARVO (1962-2015); The Authors (2016-present)
Purpose: Retinal cell cultures have been used for many years by many investigators, but those studies almost always utilize embryonic or neonatal retinas. It is critical to study the cellular physiolgy and pathology of adult neurons, particularly to understand adult-onset diseases such as most glaucoma. This is because developing neurons have very different physiological functions, patterns of gene expression, and phenotypic characteristics in general, compared to adult neurons. We have recently been using dissociated mixed neuronal cultures from adult mouse retinas, with good success. In this study, we compared media, additives, and serum, with an eye to determine optimum conditions for RGC survival. Methods: Adult (5-6 weeks) male C57BL/6 mice were used. The culture method was based on that previously described for adult pig (Luo and Hicks, IOVS 42:1096, 2001). Cell types were identified by immunolabeling for specific markers, and populations quantified by fluorescence microscopy and counting, after 7 days in vitro. Results: In adult mice dissociated cultures, DMEM-F12 with 5% fetal bovine serum gave very poor neuron survival. Neurobasal-A with B27 additives provided good survival of all retinal cells. Neurobasal-A with serum (2-10%) provided much better survival of RGC's compared with B27. Even low concentrations of serum (2%) were maximally effective. Adding B27 plus serum gave slightly better results than serum alone. Interestingly, only RGC survival was improved by serum: AC's, BC's and PR's survived equally well with B27 alone, serum, or the combination. Conclusions: Excellent survival of all classes of retinal cells is obtained with Neurobasal-A containing 2% B27. Serum contains a factor or factors that specifically promote the survival of ganglion cells. The identity and mechanism of action of this factor(s) is being investigated.
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