Previous research has suggested that serotonin 5-HT(2A) receptors modulate the functioning of the mesocortical dopamine (DA) pathway. However, the specific role of 5-HT(2A) receptors localized within the medial prefrontal cortex (mPFC) is not known. The present study employed in vivo microdialysis to examine the role of this receptor in the modulation of basal and K(+)-stimulated (Ca(2+)-dependent) DA release. The selective 5-HT(2A) antagonist M100,907 was infused directly into the mPFC of conscious rats. This resulted in a concentration-dependent blockade of K(+)-stimulated DA release. Intracortical application of M100,907 also blocked increases in DA release produced by the systemic administration of the 5-HT(2A/2C) agonist, 1-(2,5-dimethoxy-4-iodophenyl)-2-aminopropane (DOI). These findings demonstrate that local 5-HT(2A) antagonism has an inhibitory effect on stimulated, Ca(2+)-dependent DA release. They suggest that cortical 5-HT(2A) receptors potentiate the phasic release of mesocortical DA.