The cannabinoids have been shown to affect both membrane lipid ordering and the activities of several membrane-associated proteins. We have investigated the effects of the cannabinoids on protein kinase C, a lipid-dependent enzyme that functions as an important regulator of signal-transduction processes in the brain. The naturally occurring cannabinoid delta 9-tetrahydrocannabinol (delta 9-THC) increased the activity of protein kinase C isolated from rat forebrain at concentrations of 10 microM and above. 11-OH-delta 9-THC, cannabinol and cannabidiol also increased protein kinase C activity in the same concentration range. delta 9-THC (10 microM) decreased the Kact of protein kinase C for calcium from 28 microM to 18 microM and had no effect on the phosphatidylserine concentration-stimulation relationship. At a concentration of 30 microM, delta 9-THC increased the binding of [3H]phorbol-12,13-dibutyrate ([3H]PDBu) to protein kinase C and decreased the Kd for [3H]PDBu from 8.2 nM to 5.4 nM. delta 9-THC also had effects on lipid ordering of PS micelles, producing a significant increase in the fluorescence anisotropy of 1,6-diphenyl-1,3,5-hexatriene at a concentration of 10 microM. These data suggest that delta 9-THC activates protein kinase C via a novel mechanism, possibly as a result of effects on vesicle lipid physical characteristics.