While several mitogens for astrocytes have been described, the signal transduction pathway(s) that mediates their proliferative effect remains unclear; in this report, a major role for the protein kinase C (PKC) system is suggested by several lines of evidence. Firstly, biologically active phorbol esters, 4 beta-phorbol-12,13-dibutyrate and phorbol-12-myristate-13-acetate, increase the proliferation of astrocytes as determined by [3H]thymidine incorporation or bromodeoxyuridine immunofluorescence; this effect is not reproduced by a phorbol ester that binds to PKC but does not activate it (4 alpha-phorbol-12,13-didecanoate). Secondly, 2 relatively selective inhibitors of PKC, H7 and staurosporine, attenuate the basal rate of proliferation of astrocytes in concentrations that were not cytotoxic to cells. Thirdly, mitogen-enhanced proliferation of astrocytes can be blocked by PKC inhibitors; this is observed for all astrocyte mitogens tested. Fourthly, measurements of PKC enzyme activity in astrocytes in response to serum-mitogenic factors, or to staurosporine, revealed a statistically significant correlation with proliferation rate. The mediation by PKC is not dependent on species- or age factors, since neonatal mouse or adult human astrocytes gave comparable results. The results have relevance to normal development and reactive gliosis post-injury, 2 conditions where astrocytes undergo proliferation, and to glioma growth.