Neurosteroids synthesized in the central nervous system exert a powerful influence on depression, mood and stress disorders. Allopregnanolone is one of the most potent endogenous positive modulators known for GABA-A receptors, acting especially on extrasynaptic delta subunit- containing receptor subtypes. Neurosteroids have been proposed to alter actions of various drugs of abuse, including alcohol and opiates, through modulation mesolimbic dopamine (DA) system. This study was conducted to provide a better understanding of the mechanism by which neurosteroids modulate the mesolimbic DA system. We have studied ganaxolone, a synthetic analog of allopregnanolone lacking nuclear steroid hormonal activity. The functional changes after acute ganaxolone in the ventral tegmental area (VTA) were assessed by whole-cell patch clamp technique. Rewarding and reinforcing properties of ganaxolone were evaluated by using conditional behavioral model in C57BL/6J mice and in GABA-A receptor delta subunit knockout mice. Ganaxolone dose-dependently induced potentiation of glutamate receptor currents in VTA DA neurons, which lasted notably for 6 days and was at least partly mediated by insertion of GluA2-subunit lacking AMPA receptors. In behavioral experiments ganaxolone demonstrated conditioned place aversion. Ganaxolone-induced potentiation of VTA DA neurons could mediate some of the behavioral effects of neurosteroids, perhaps through disinhibition of local VTA interneurons in the same way as another extrasynaptic GABA-A receptor agonist gaboxadol (Vashchinkina et al., J Neurosci 32:5310–5320, 2012). Acknowledgements: The study was supported by the Finnish Foundation for Alcohol Studies and the Academy of Finland.