Neuropeptide Y (NPY) and protein kinase A (PKA) have been implicated in neurobiological responses to ethanol. We have previously reported that mutant mice lacking normal production of the RIIβ subunit of PKA (RIIβ−/− mice) show enhanced sensitivity to the locomotor stimulant effects of ethanol and increased behavioral sensitization relative to littermate wild-type RIIβ+/+ mice. We now report that RIIβ−/− mice also show increased NPY immunoreactivity in the nucleus accumbens (NAc) core and the ventral striatum relative to RIIβ+/+ mice. These observations suggest that elevated NPY signaling in the NAc and/or striatum may contribute to the increased sensitivity to ethanol-induced behavioral sensitization that is a characteristic of RIIβ−/− mice. Consistently, NPY−/− mice failed to display ethanol-induced behavioral sensitization that was evident in littermate NPY+/+ mice. To examine more directly the role of NPY in the locomotor stimulant effects of ethanol, we infused a recombinant adeno-associated virus (rAAV) into the region of the NAc core of DBA/2J mice. The rAAV-fibronectin (FIB)-NPY13–36 vector expresses and constitutively secretes the NPY fragment NPY13–36 (a selective Y2 receptor agonist) from infected cells in vivo. Mice treated with the rAAV-FIB-NPY13–36 vector exhibited reduced expression of ethanol-induced behavioral sensitization compared with mice treated with a control vector. Taken together, the current data provide the first evidence that NPY signaling in the NAc core and the Y2 receptor modulate ethanol-induced behavioral sensitization.