The present study was conducted to understand the effect of haloperidol, a typical antipsychotic, and clozapine, an atypical one, on the release of endogenous glutamate in nerve terminals isolated from rat prefrontal cortex using an on-line enzyme-linked fluorometric assay. We found that both haloperidol and clozapine significantly inhibited 4-aminopyridine (4AP)-evoked and veratridine-evoked but not KCl-evoked glutamate release from prefrontocortical synaptosomes. This inhibition produced by these two drugs was concentration-dependent with different potency, and associated with a reduction both in the depolarization-evoked increase in the intrasynaptosomal free Na(+) concentration ([Na(+)](i)) and in 4AP or KCl-evoked depolarization of the synaptosomal plasma membrane potential. Additionally, in the presence of calcium-free medium containing 0.2 mM EGTA, the Ca(2+)-independent component of 4AP-evoked glutamate release was also inhibited by haloperidol or clozapine. Based on these results, we suggest that haloperidol and clozapine inhibit glutamate release from rat prefrontocortical nerve terminals by affecting ion-channel activities determining nerve terminal excitability, probably as a result of Na(+) channel blockage or K(+) channel activation.