Pretreatment of CD-1 mouse skin with prostratin (12-deoxyphorbol 13-acetate) inhibited biological response to phorbol 12-myristate 13-acetate. The three responses examined were hyperplasia, induction of ornithine decarboxylase, and edema; the characteristics of inhibition depended on the specific response. Hyperplasia is the best short-term correlate of tumor promotion. Two or more pretreatments with 2.56 mumol (1 mg) prostratin, administered at intervals of 1-4 days, almost completely blocked the hyperplasia induced by phorbol 12-myristate 13-acetate applied 15 min to 6 h after the last pretreatment. Inducibility of hyperplasia was partially restored at 2 days and recovered by 4 days. Prostratin was more potent for inhibition of ornithine decarboxylase induction (50% inhibitory dose = 25.6 nmol) than it was for hyperplasia: the inhibition was largely attained by the first application, and the recovery from inhibition was slower (8 days). Edema was partially inhibited by prostratin (dose giving 50% of maximal inhibition = 512 nmol). We have previously demonstrated that prostratin is a protein kinase C activator. Our present results show that prostratin is a functional antagonist for a class of protein kinase C mediated responses. The findings emphasize the diversity of biological outcome for protein kinase C activators, presumably driven by the extensive heterogeneity in the protein kinase C pathway.