Astrocytes have become promising new agents against major depressive disorders (MDD) primarily due to the crucial role they play in the pathogenesis of such disorders. However, a simple and reliable animal model that can be used to screen for astrocyte-targeting antidepressants has not yet been developed. In this study, we utilized a repeated corticosterone (CORT) injection paradigm to develop a mouse depression model wherein we examined the occurrence of alterations in hippocampal astrocyte population by using two astrocytic markers, namely, glial fibrillary acidic protein (GFAP) and S100β. Moreover, we determined the effects of fluoxetine and diazepam on CORT-induced astrocytic alterations to assess the predictive validity. Results showed that repeated CORT injections showed no effects on the number of GFAP+ and S100β+ astrocytes, but they decreased the protrusion length of GFAP+ astrocytes and GFAP protein expression in the hippocampus. Furthermore, repeated CORT injections produced a sustained increase of S100β protein levels in the entire hippocampus of male mice. CORT-induced hippocampal astrocyte disruption was antagonized by chronic fluoxetine treatment. By contrast, the anxiolytic drug diazepam was ineffective in the same experimental setting. All these findings suggest that the repeated CORT injection paradigm produces the astrocytic alterations similar to those in MDD and can serve as a useful mouse model to screen antidepressants meant to target astrocytes. These observations can also help in further discussing the underlying mechanisms of CORT-induced astrocytic alterations.