Modulation of neurotrophic factors to protect neurons from damage is proposed as a novel mechanism for the action of antidepressants. However, the effect of antidepressants on modulation of glial cell line-derived neurotrophic factor (GDNF), which has potent and widespread effects, remains unknown. Here, we demonstrated that long-term use of antidepressant treatment significantly increased GDNF mRNA expression and GDNF release in time- and concentration-dependent manners in rat C6 glioblastoma cells. Amitriptyline treatment also increased GDNF mRNA expression in rat astrocytes. GDNF release continued for 24 h following withdrawal of amitriptyline. Furthermore, following treatment with antidepressants belonging to several different classes (amitriptyline, clomipramine, mianserin, fluoxetine and paroxetine) significantly increased GDNF release, but which did not occur after treatment with non-antidepressant psychotropic drugs (haloperidol, diazepam and diphenhydramine). Amitriptyline-induced GDNF release was inhibited by U0126 (10 microM), a mitogen-activated protein kinase (MAPK)-extracellular signal-related kinase (ERK) kinase (MEK) inhibitor, but was not inhibited by H-89 (1 microM), a protein kinase A inhibitor, calphostin C (100 nM), a protein kinase C inhibitor and PD 169316 (10 microM), a p38 mitogen-activated protein kinase inhibitor. These results suggested that amitriptyline-induced GDNF synthesis and release occurred at the transcriptional level, and may be regulated by MEK/MAPK signalling. The enhanced and prolonged induction of GDNF by antidepressants could promote neuronal survival, and protect neurons from the damaging effects of stress. This may contribute to explain therapeutic action of antidepressants and suggest new strategies of pharmacological intervention.