AIM: To examine the protective effect of mitochondrial division inhibitor-1 (Mdivi-1) against high-altitude-induced memory impairment in mice. METHODS: C57BL/6J male mice were administered Mdivi-1 before exposure to a simulated high-altitude hypoxia environment. The novel object recognition test and Morris water maze were used to test cognitive function. Golgi staining was used to visualize dendritic spines. PCR, Western blot, and immunofluorescence were performed to detect microglial activation and synaptic phagocytosis. RESULTS: Mice exposed to short-term or long-term simulated high-altitude conditions experienced memory deficits. However, these deficits were significantly mitigated by pre-treatment with Mdivi-1. Simulated high-altitude exposure caused a reduction in synapses (dendritic spines) and the activation of microglia. Following Mdivi-1 injection, synapse density was significantly increased, and microglial activation was attenuated. Under hypoxic conditions, primary cultured microglia exhibited significantly enhanced phagocytic activity towards TRITC-Dextran or synaptosomes, which was abolished by Mdivi-1. Additionally, Mdivi-1 inhibited the HIF-1 signaling pathway and restricted the hypoxia-induced glycolytic activity in microglia. Specific inhibition of glycolysis effectively weakened the phagocytic capacity of microglia under hypoxia. CONCLUSION: Mdivi-1 dramatically mitigated memory impairment in mice induced by simulated high-altitude exposure. Mdivi-1 reduced microglial glycolysis in hypoxic conditions, thereby limiting microglial activation and preventing excessive synaptic phagocytosis. Consequently, it effectively protected memory.