The intermediate lobe of the pituitary (IL) is a promising model system for the study of dopaminergic neurotransmission. We sought to characterize the release of dopamine (DA) from the IL using electrochemical methods which provide increased spatial and temporal resolution compared to the superfusion techniques which had previously been employed to study this system. After pretreatment with monoamine oxidase (MAO) inhibitors, we were able to routinely measure DA overflow after 30-sec, 9-Hz electrical stimulation of the pituitary stalk. This effect was blocked by tetrodotoxin and Cd++ ions confirming that it was due to synaptic activation. Intracellular recording from the postsynaptic cells (melanotrophs) showed a postsynaptic hyperpolarization with a time course that was identical to that of DA overflow. Overflow was increased and prolonged by agents that inhibited DA uptake (nomifensine, mazindol or cocaine) and increased but not prolonged by a D2 antagonist (sulpiride), presumably acting via presynaptic autoreceptors. The DA precursor 3,4-dihydroxyphenylalanine also increased DA overflow gradually over a 45-min time period. In view of reports of possible colocalization of DA and gamma-aminobutyric acid in IL, we determined the possible effects of GABAergic agents on DA release. Neither agonists (muscimol or baclofen) nor antagonists (bicuculline or 2-hydroxysaclofen) had any effect on DA overflow. By using the MAOA-specific inhibitor clorgyline and the MAOB-specific inhibitor deprenyl we were able to confirm that the MAOA form of the enzyme is involved in DA metabolism in this preparation.