Classic theories propose opponent functions for striatal dopamine (DA) and serotonin (5-hydroxytryptamine; 5HT), with DA promoting approach and 5HT promoting patience or avoidance. How these neuromodulators regulate downstream circuits to achieve such antagonistic effects remains mysterious. Here, we mapped striatal 5HT receptor expression and recorded from genetically-identified striatal neurons to demonstrate that DA and 5HT selectively activate distinct populations of striatal neurons to exert opponent control over striatal output.