Adenosine A 2A receptors (A 2AR) are located postsynaptically in striatopallidal GABAergic neurons, antagonizing dopamine D 2 receptor functions, and are also located presynaptically at corticostriatal terminals, facilitating glutamate release. To address the hypothesis that these two A 2AR populations differently control the action of psychostimulants, we characterized A 2AR modulation of cocaine-induced effects at the level of DARPP-32 phosphorylation at Thr-34 and Thr-75, c-Fos expression, and psychomotor activity using two lines of cell-type selective A 2AR knockout (KO) mice with selective A 2AR deletion in GABAergic neurons (striatum-A 2AR-KO mice), or with A 2AR deletion in both striatal GABAergic neurons and projecting cortical glutamatergic neurons (forebrain-A 2AR-KO mice). We demonstrated that striatum-A 2AR KO mice lacked A 2ARs exclusively in striatal GABAergic terminals whereas forebrain-A 2AR KO mice lacked A 2ARs in both striatal GABAergic and glutamatergic terminals leading to a blunted A 2AR-mediated facilitation of synaptosomal glutamate release. The inactivation of A 2ARs in GABAergic neurons reduced striatal DARPP-32 phosphorylation at Thr-34 and increased its phosphorylation at Thr-75. Conversely, the additional deletion of corticostriatal glutamatergic A 2ARs produced opposite effects on DARPP-32 phosphorylation at Thr-34 and Thr-75. This distinct modulation of DARPP-32 phosphorylation was associated with opposite responses to cocaine-induced striatal c-Fos expression and psychomotor activity in striatum-A 2AR KO (enhanced) and forebrain-A 2AR KO mice (reduced). Thus, A 2ARs in glutamatergic corticostriatal terminals and in GABAergic striatal neurons modulate the action of psychostimulants and DARPP-32 phosphorylation in opposite ways. We conclude that A 2ARs in glutamatergic terminals prominently control the action of psychostimulants and define a novel mechanism by which A 2ARs fine-tune striatal activity by integrating GABAergic, dopaminergic and glutamatergic signaling.