There is increasing evidence that individual protein kinase C (PKC) isozymes mediate specific effects of ethanol on the nervous system. In addition, multiple lines of evidence suggest that the mesoaccumbens dopamine reward system is critically involved in the rewarding and reinforcing effects of ethanol. Yet little is known about the role of individual PKC isozymes in ethanol reinforcement processes or in regulation of mesolimbic systems. In this study, we report that mice lacking the epsilon isoform of PKC (PKCepsilon) show reduced operant ethanol self-administration and an absence of ethanol-induced increase in extracellular dopamine levels in the nucleus accumbens. PKCepsilon null mice exhibited a 53% decrease in alcohol-reinforced operant responses under basal conditions, as well as following ethanol deprivation. Behavioural analysis revealed that while both genotypes had the same number of drinking bouts following deprivation, PKCepsilon null mice demonstrated a 61% reduction in number of ethanol reinforcers per bout and a 57% reduction in ethanol-reinforced response rate. In vivo microdialysis experiments showed that, in contrast to wild-type mice, PKCepsilon null mice exhibited no change in extracellular levels of dopamine in the nucleus accumbens following acute administration of ethanol (1 and 2 g/kg i.p.), while mesolimbic dopamine responses to cocaine (20 mg/kg i.p.) or high potassium (100 mM) in these mice were comparable with that of wild-types. These data provide further evidence that increases in extracellular mesolimbic dopamine levels contribute to the reinforcing effects of ethanol, and indicate that pharmacological agents inhibiting PKCepsilon may be useful in the treatment of alcohol dependence.