Although numerous studies investigated the mechanisms underlying 3,4-methylenedioxymethamphetamine (MDMA)-induced neurotoxicity, little is known about its long-term functional consequences on 5-HT neurotransmission in mice. This led us to evaluate the delayed effects of MDMA exposure on the 5-HT system, using in-vitro and in-vivo approaches in both 5-HTT wild-type and knock-out mice. Acute MDMA in-vitro application on slices of the dorsal raphe nucleus (DRN) induced concentration-dependent 5-HT release and 5-HT cell firing inhibition. Four weeks after MDMA administration (20 mg/kg b.i.d for 4 d), a 2-fold increase in the potency of the 5-HT1A receptor agonist ipsapirone to inhibit the discharge of DRN 5-HT neurons and a larger hypothermic response to 8-OH-DPAT were observed in MDMA- compared to saline-treated mice. This adaptive 5-HT1A autoreceptor supersensitivity was associated with decreases in 5-HT levels but no changes of [3H]citalopram binding in brain. Long-term MDMA treatment also induced a 30% decrease in BrdU labelling of proliferating hippocampal cells and an increased immobility duration in the forced swim test suggesting a depressive-like behaviour induced by MDMA treatment. All these effects were abolished in 5-HTT-/- knock-out mice. These data indicated that, in mice, MDMA administration induced a delayed adaptive supersensitivity of 5-HT1A autoreceptors in the DRN, a deficit in hippocampal cell proliferation and a depressive-like behaviour. These 5-HTT-dependent effects, opposite to those of antidepressants, might contribute to MDMA-induced mood disorders.