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The function of striatal adenosine A_2A receptors (A_2ARs) is well recognized because of their high expression levels and the documented antagonistic interaction between A_2ARs and dopamine D₂ receptors in the striatum. However, the role of extrastriatal A_2ARs in modulating psychomotor activity is largely unexplored because of the low level of expression and lack of tools to distinguish A_2ARs in intrinsic striatal versus nonstriatal neurons. Here, we provided direct evidence for the critical role of A_2ARs in extrastriatal neurons in modulating psychomotor behavior using newly developed striatum-specific A_2AR knock-out (st-A_2AR KO) mice in comparison with forebrain-specific A_2AR KO (fb-A_2AR KO) mice. In contrast to fb-A_2AR KO (deleting A_2ARs in the neurons of striatum as well as cerebral cortex and hippocampus), st-A_2AR KO mice exhibited Cre-mediated selective deletion of the A_2AR gene, mRNA, and …