1. Physiological stimulation of neuronal activity induces an increase in extracellular lactate. Experiments were designed to determine the role of the reuptake of neuronally released glutamate in lactate delivery to the extracellular compartment. 2. In vivo microdialysis was used in freely moving rats. The lactate concentration in striatal dialysate was assayed using an enzyme-based on-line assay at 1 min intervals. Drugs were given locally through the dialysis probe. 3. The extracellular concentration of lactate, determined using the zero net flux method, was 346 +/- 21 microM. 4. Induced grooming caused a maximal increase in lactate concentration in striatal dialysate of 58 +/- 10%. 5. Administration of 100 microM glutamate caused a transient increase in dialysate lactate concentration of 72 +/- 17%. 6. A 20 min infusion of the glutamate uptake blockers beta-D,L-threohydroxyaspartate (THA) or pirrolidine-2-4-dicarboxylate (PDC) produced an increase in basal lactate, which was sustained in response to THA and transient in response to PDC. 7. Grooming induced during the infusion of PDC produced no significant increase in lactate. 8. Grooming induced after the infusion of the glutamate uptake blockers gave rise to a reduced increase in lactate. 9. These results support the hypothesis that stimulated release of lactate is dependent on the uptake of glutamate.