Binding of 125I-epidermal growth factor (EGF) to purified populations of rat astrocytes, oligodendrocytes, and neurons was measured. Astrocytes bound 40,000-100,000 EGF molecules per cell, while oligodendrocytes bound only 6,000-10,000 EGF molecules per cell. In contrast, neurons had little or no capacity to bind 125I-EGF. EGF alone was able to stimulate incorporation of tritiated thymidine fivefold in purified astrocyte cultures incubated in serum-free medium. When EGF was added to the previously described chemically defined medium for astrocytes, incorporation of tritiated thymidine in purified astrocytes was equivalent to that observed in medium containing 10% fetal calf serum. Addition of EGF to this chemically defined medium also doubled the proliferative capability of the medium for cultured astrocytes. EGF was maximally effective in stimulating 3H-thymidine incorporation at concentrations between 1-10 ng/ml.