Studies that have attributed the discriminative stimulus effects of ±3,4-methylenedioxymethamphetamine (MDMA) to serotonergic mechanisms typically use a relatively low training dose of 1.5 mg/kg. The role of serotonin in the discriminative stimulus effects of higher doses of MDMA is, however, unknown. Separate groups of rats were trained to discriminate MDMA (1.5 or 3.0 mg/kg) from saline using a two-lever, food-reinforced drug-discrimination procedure. Generalization tests were carried out with a range of serotonin and dopamine ligands. Fluoxetine (0.3–3 mg/kg), clomipramine (1–10 mg/kg) and meta-chlorophenylpiperazine (0.3–2 mg/kg) dose-dependently substituted for the 1.5 mg/kg MDMA stimulus, but not the 3.0 mg/kg MDMA stimulus. 8-OH-DPAT (0.03–0.3 mg/kg) and RU-24969 (0.3–3 mg/kg) substituted for both the low-dose and the high-dose MDMA stimulus. The generalization dose–effect curve produced by 2,5-dimethoxy-4-iodoamphetamine (0.3–3 mg/kg) was shifted to the right for the 3.0 mg/kg MDMA-trained group. Amphetamine (0.25 and 0.5 mg/kg) and apomorphine (0.125 and 0.25 mg/kg) substituted for the 3.0 mg/kg, but not the 1.5 mg/kg MDMA stimulus. The results suggest some differences in the role of serotonin and dopamine in the discriminative stimulus effects of a low versus a higher dose of MDMA.