Pituitary adenylate cyclase-activating polypeptide (PACAP) has been conserved remarkably during evolution and is widely expressed in the mammalian brain. In Drosophila, mutation of the PACAP homologue results in behavioral defects, including impaired olfaction-associated learning and changes in ethanol sensitivity. Here, we report the generation of mice lacking the PACAP gene (PACAP^−/−). PACAP^−/− mice were born in the expected Mendelian ratios but had a high early-mortality rate. The surviving adult PACAP^−/− mice displayed remarkable behavioral changes; they exhibited hyperactive and explosive jumping behaviors in an open field, increased exploratory behavior, and less anxiety in the elevated plus maze, emergence, and novel-object tests. Analysis of PACAP^−/− mice brains revealed that the serotonin metabolite 5-hydroxyindoleacetic acid was slightly decreased in the cortex and striatum compared with wild-type mice. The present study provides evidence that PACAP plays a previously uncharacterized role in the regulation of psychomotor behaviors.