The topical application of croton oil, benzo(a)pyrene, acetic acid, and 12-O-tetradecanoyl-phorbol-13-acetate to mouse skin caused an increase in the activity of epidermal low-affinity cyclic adenosine 3':5'-monophosphate (cyclic AMP) phosphodiesterase. The increase was most pronounced with croton oil, began between 4 and 6 hr after application of this material, and was maintained for at least 48 hr. The activity of cyclic guanosine 3':5'-monophosphate phosphodiesterase was also increased by treatment with croton oil or 12-O-tetradecanoyl-phorbol-13-acetate, but detailed time courses were not obtained. Increased activity was observed in both the soluble fractions and the washed particulate fractions of epidermis. Fractionation of soluble extracts from acetone-treated epidermis on DEAE-cellulose columns showed the presence of enzymes with specificity for both cyclic AMP and cyclic guanosine 3':5'-monophosphate, together with a peak catalyzing the hydrolysis of both cyclic AMP and cyclic guanosine 3':5'-monophosphate. The activity of this latter nonspecific activity was selectively increased following treatment with croton oil. The increase in cyclic AMP phosphodiesterase activity was partially abolished by multiple injections of cycloheximide, suggesting that new protein synthesis was involved. Injection of the alpha-receptor antagonist phentolamine abolished a croton oil-induced rise in epidermal cyclic AMP levels and decreased the induction of cyclic AMP phosphodiesterase activity. From these results it was concluded that the increase in enzyme activity was induced by cyclic AMP.