Currently, accumulating studies indicated that upregulation of glycogen synthase kinase3β (GSK3β)played an important role in depression pathogenesis.Our previous study demonstrated that ammoxetine, a novel serotonin and norepinephrine uptake inhibitor, displayed antidepressant activity more potent and faster than existing antidepressants, which may be due to the increasing of hippocampal inhibitory serinephosphorylation of glycogen synthase kinase3 (GSK3).The present study was to evaluate whether activation of PI3K/Akt signaling, one of the most important pathways regulating the phosphorylation of GSK3 β, was required for ammoxetine induced antidepressant effects and upregulation of pGSK3 β.Behavioral results indicated that acute oral administration of ammoxetine at 10 mg/kg produced robust antidepressant effects in the forced swimming test and learned helpless test in mice, which were blocked totally by phosphatidylinositol (PI3)kinase (PI3K) inhibitor LY294002.Then, Western blot results demonstrated that ammoxetine induced increasing of GSK3 β phosphorylation and activation of PI3K/Akt signaling can also be antagonized at the same testing time points by LY294002.These findings suggest that activation of PI3K/Akt/GSK3β signaling is pivotal and necessary for the antidepressant effects of ammoxetine in the forced swimming test and learned helpless test in mice.