Role of dopamine neurotransmission in retrieval of drug-associated memory is not fully understood. The present study provides direct evidences that dopamine neurotransmission is differentially activated in the prefrontal cortex (PFC) and nucleus accumbens (NAc) in the cocaine conditioned place preference (CPP) paradigm in behaving mice using in vivo microdialysis. In mice that had acquired cocaine CPP, the dopamine levels in the PFC, but not in the NAc, increased in response to the cocaine-associated cue, when mice were placed in the cocaine chamber of two-separated chamber apparatus. The induction of the dopamine response and the development of cocaine CPP were mediated through activation of glutamate NMDA/AMPA receptor signaling in the PFC during conditioning. Activation of dopamine D1 or D2 receptors signaling in the PFC was required for the cocaine-induced locomotion, but not for the induction of the dopamine response or the development of cocaine CPP. Interestingly, dopamine levels in the NAc increased in response to the cocaine-associated cue, when mice were placed at the center of two-connected chamber apparatus, which requires the motivated exploratoration of cocaine reward. In summary, dopamine neurotransmission in the PFC is activated to retrieve the context-associated reward memory of cocaine, whereas dopamine neurotransmission in the NAc is activated in the process of motivated exploration of cocaine reward. The present findings suggest the nature of difference in dopamine transmission in the PFC and NAc to the psychological reward cues.