Ethanol suppression of astrocyte mitogenesis is well recognized but ethanol, under some conditions, has also been shown to stimulate astrocyte proliferation. This study addressed the role of protein kinase C and other mitogenic factors as mechanisms responsible for the bidirectional effects of ethanol on astrocyte DNA synthesis. Ethanol treatment inhibited astrocyte DNA synthesis both at 4 hr (short term) and 24 hr (long term) in serum free medium. In contrast, when the medium contained serum, ethanol was less effective in inhibiting DNA synthesis at 4 hr and treatment with ethanol for 24 hr increased DNA synthesis. Protein kinase C activity was increased in cells treated with ethanol for either 4 or 24 hr. Ethanol inhibition of DNA synthesis in serum free medium was not reversed by down regulating protein kinase C. In contrast, downregulating protein kinase C activity by continuous treatment with phorbol myristic acetate partially reversed the effect ethanol had on DNA synthesis. Also, directly inhibiting protein kinase C with H-7 in cells maintained and treated in the presence of serum abolished the stimulatory effect ethanol had on DNA synthesis. It appears that the negative regulation of astrocyte DNA synthesis by ethanol occurs by protein kinase C and serum independent mechanisms whereas adaptive or stimulatory effects of ethanol on astrocyte DNA synthesis requires the interaction of protein kinase C with other factors present in serum.