A novel serum free primary astrocyte culture method that mimic quiescent astrocyte phenotype

Abstract Background Primary astrocyte cultures have been used for decades to study astrocyte functions in health and disease. The current primary astrocyte cultures are mostly maintained in serum-containing medium which produces astrocytes with a reactive phenotype as compared to in vivo quiescent astrocytes. The aim of this study was to establish a serum-free astrocyte culture medium that maintains primary astrocytes in a quiescent state. New method Serum free astrocyte base medium (ABM) supplemented with basic fibroblast growth factor 2 (FGF2) and epidermal growth factor (EGF) (ABM-FGF2-EGF) or serum supplemented DMEM (MD-10%FBS) was used to culture primary astrocytes isolated from cerebral cortex of postnatal day 1 C57BL/6 mice. Results Compared to astrocytes cultured in MD-10%FBS medium, astrocytes in ABM-FGF2-EGF had higher process bearing morphologies similar to in vivo astrocytes. Western blot, immunostaining, quantitative polymerase chain reaction and metabolic assays revealed that astrocytes maintained in ABM-FGF2-EGF had enhanced glycolytic metabolism, higher glycogen content, lower GFAP expression, increased glutamine synthase, and glutamate transporter-1 mRNA levels as compared to astrocytes cultured in MD-10% FBS medium. Comparison to existing methods These observations suggest that astrocytes cultured in ABM-FGF2-EGF media compared to the usual FBS media promote quiescent and biosynthetic phenotype similar to in vivo astrocytes. Conclusion This media provides a novel method for studying astrocytes functions in vitro under physiological and pathological conditions.