The balance between GABA-mediated inhibitory and glutamate-mediated excitatory synaptic transmission represents a fundamental mechanism for controlling nervous system function, and modulators that can alter this balance may participate in the pathophysiology of neuropsychiatric disorders. Pregnenolone sulfate (PS) is a neuroactive steroid that can modulate the activity of ionotropic glutamate and GABAA receptors either positively or negatively, depending upon the particular receptor subtype, and modulates synaptic transmission in a variety of experimental systems. To evaluate the modulatory effect of PS in vivo, we infused PS into rat striatum via a microdialysis probe while monitoring local extracellular dopamine (DA) levels. The results demonstrate that PS at low nanomolar concentrations significantly increases extracellular DA levels. The PS -induced increase in extracellular DA is antagonized by the NMDA receptor antagonist, D-AP5, but not by the sigma receptor antagonist, BD 1063. The results demonstrate that exogenous PS, at nanomolar concentrations, is able to increase DA overflow in the striatum through an NMDA receptor mediated pathway.