Protein kinase G (PKG) activation has been implicated in the regulation of synaptic plasticity in the brain. This study was conducted to determine the involvement of PKG-associated dopamine D2 (D2) receptors in the regulation of dopamine release, ΔFosB expression and locomotor activity in response to repeated cocaine exposure. Repeated systemic injections of cocaine (20 mg/kg), once a day for seven consecutive days, increased cyclic guanosine monophosphate (cGMP) and extracellular dopamine concentrations in the dorsal striatum. Inhibition of neuronal nitric oxide synthase (nNOS), cGMP or PKG and stimulation of D2 receptors decreased the repeated cocaine-induced increase in dopamine concentrations. Similar results were obtained by the combining nNOS, cGMP or PKG inhibition with stimulation of D2 receptors. Parallel to these data, PKG inhibition, D2 receptor stimulation, and combining PKG inhibition with stimulation of D2 receptors decreased the repeated cocaine-induced increases in ΔFosB expression and locomotor activity. These findings suggest that control of D2 receptors by PKG activation after repeated cocaine is responsible for upregulating dopamine release and sustained long-term changes in gene expression in the dopamine terminals and gamma-aminobutyric acid neurons of the dorsal striatum, respectively. This upregulation may contribute to behavioral changes in response to repeated exposure to cocaine.