The aim of this study was to investigate the protective effect and potential molecular mechanism of sarsasapogenin (Sar) in cerebral ischemia/reperfusion injury (CI/R). The oxygen-glucose deprivation/reoxygenation (OGD/R)-induced mouse hippocampal neurons (HT22) were used as the in vitro model of CI/R. HT22 cells were divided into control (Con) group, OGD/R group, low-dose Sar group (2 μmol/L), medium-dose Sar group (4 μmol/L), high-dose Sar group (8 μmol/L), LY294002 group (20 μmol/L) and high-dose Sar+LY294002 group (8 μmol/L Sar+20 μmol/L LY294002). Cell proliferation activity was detected by CCK-8 method. Cysteinyl aspartate-specific protease-3 (CASP3) activity, superoxide dismutase (SOD), malondialdehyde (MDA) and glutathione (GSH) levels were detected with the kits. 2′,7′-Dichlorodihydrofluorescein diacetate (DCFH-DA) method was used to detect reactive oxygen species (ROS) level. Lipid peroxidation was detected by lipid peroxidation fluorescence probe method. The level of ferrous iron was detected by colorimetry. The protein expression was detected by Western blot. The results showed that Sar could enhance the cellular activity of HT22 after OGD/R injury. Compared with OGD/R group, ROS level, lipid superoxidation, ferrous iron and MDA levels were decreased in the Sar group, while SOD activity, GSH and glutathione peroxidase 4 protein (GPX4) levels were increased. In addition, after Sar treatment, phosphorylated-phosphoinositide 3-kinase (p-PI3K)/PI3K ratio and phosphorylated-protein kinase B (p-AKT)/AKT ratio increased. LY294002 weakened the alleviating effects of Sar on OGD/R-induced oxidative stress and ferroptosis. Overall, Sar alleviates OGD/R-induced oxidative stress and ferroptosis in hippocampal neurons by activating the PI3K-AKT signaling pathway.