Dysregulated microRNAs (miRNAs) are crucial regulators of cerebrovascular conditions, including ischemic stroke. Circulating miR-125a-5p is associated with ischemic stroke and may have clinical utility as an early diagnostic biomarker. This study conducted a series of experiments that were designed to elucidate the regulatory action of miR-125a-5p in ischemic brain injury and its underlying mechanisms. The results of this study found that the expression of miR-125a-5p was increased in BV2 microglial cells under oxygen-glucose deprivation/reoxygenation (OGD/R) setting, as well as in rat brain tissue after middle cerebral artery occlusion (MCAO). OGD/R triggered BV2 microglial cell apoptosis, whereas downregulation of miRNA-125a-5p suppressed the apoptosis rate in OGD/R-induced BV2 microglial cells. Subsequently, insulin-like growth factor binding protein 3 (IGFBP3) is a molecular target of miR-125a-5p, and IGFBP3 knockdown reversed the effects of miR-125a-5p inhibitor on BV2 microglial cells in vitro. These data supported the fact that miR-125a-5p silencing protects against cerebral ischemia-induced injury by targeting IGFBP3.