Circumstantial evidence suggests that sensitization to the behavioral effects of d-amphetamine is mediated by increased glutamate levels in the ventral tegmental area. To test this directly, the present study examined whether increasing glutamate levels in the ventral tegmental area with a glutamate uptake inhibitor is sufficient, in the absence of d-amphetamine administration, to elicit sensitization to a subsequent d-amphetamine challenge. Rats were treated bilaterally once a day for 2 days with either intra-ventral tegmental area L-trans-pyrollidine-2,4-dicarboxylic acid (50 nmol), saline, L-trans-pyrollidine-2,4-dicarboxylic acid coadministered with the competitive N-methyl-d-aspartate antagonist (+/-)-3-(2-carboxy-piperazin-4-yl)-propyl-1-phosphonic acid; CPP, 0.5 nmol), or CPP alone (0.5 nmol; all 1.0 microl/side). Following a 2 day withdrawal period, all rats were administered systemic d-amphetamine (1 mg/kg, i.p.). Repeated intra-ventral tegmental area injection of L-trans-pyrollidine-2,4-dicarboxylic acid sensitized animals to the behavioral effects of a systemic d-amphetamine challenge, an action which was blocked by co-administration of CPP. The results directly implicate ventral tegmental area glutamate in the process of sensitization to d-amphetamine. Furthermore, they demonstrate that inhibition of glutamate uptake produces the neuroadaptations necessary to induce sensitization, adding support to the contention that d-amphetamine sensitizes by modulating glutamate uptake.