The large-conductance Ca²⁺- and voltage-activated K⁺ (BK) channel is formed by four pore-forming subunits encoded by a single Slo1 gene and dually regulated by membrane voltage and intracellular Ca²⁺ levels. Recently, accumulating evidence using pharmacological methods suggests that the BK channel are associated with nociceptive sensitization. However, mechanisms underlying the effect of the BK channel on the regulation of pain signaling is still largely unknown. Here, we studied the role of the BK channel in the pain signaling in vivo using mice deficient for Slo1. In a partial sciatic nerve ligation (PSNL) model, Slo1 KO mice showed partially restored thermal hyperalgesia compared to their WT littermates. In microglia from WT mice, p38 MAPK phosphorylation was significantly increased after lysophosphatidic acid (LPA) treatment; conversely, it was not significantly upregulated in microglia from Slo1 KO mice, suggesting that the BK channel contributes to the signaling of microglial hyperactivation. Notably, intrathecal (i.t.) injection of microglia derived from WT mice into Slo1 KO mice before PSNL induced the normal development of hyperalgesia in Slo1 KO mice. These results indicate that the BK channel activation in spinal microglia, but not in neuron, contribute to the induction of neuropathic pain in this PSNL model.