Objective To investigate the effect of esketamine on astrocyte activation in the spinal dorsal horn of mice with chronic pain and evaluate its impact on Wnt / β-catenin signaling. Methods Forty C57BL / 6J mice were randomly divided into five groups: sham surgery (sham) group, pain model (model) group, low-dose esketamine (LEs) group, medium-dose esketamine (M-Es) group, and high-dose esketamine (H-Es) group. Except in the sham group, chronic pain was induced via sciatic nerve branch selective injury. Each group received the designated treatment. Mechanical withdrawal threshold (MWT) and thermal withdrawal latency (TWL) were measured. Spinal cord expression of glial fibrillary acidic protein (GFAP) was evaluated by immunohistochemistry. Immunofluorescence was used to assess astrocyte activation in the dorsal horn. Enzyme-linked immunosorbent assays measured levels of interleukin-1β(IL-1β), interleukin-6(IL-6), and tumor necrosis factor-α (TNF-α) in the spinal cord. Quantitative reverse transcription polymerase chain reaction( qRT-PCR) was performed to analyze mRNA expression patterns of Wnt3a, β-catenin, and Cyclin D1. Protein expression levels of Wnt3a, β-catenin, and Cyclin D1 in the spinal cord were evaluated by Western Blot. Results Compared with the sham group, mice in the model group showed significantly reduced MWT and TWL ( both P<0. 001). Levels of interleukin-1β, interleukin-6, and TNF-α, as well as expression of GFAP protein, were substantially increased (all P<0. 001); co-expression was evident between GFAP and Iba-1. mRNA and protein expression levels of Wnt3a, β-catenin, and Cyclin D1 were also significantly elevated (all P<0. 001). Compared with the model group, mice in all esketamine dose groups exhibited significantly increased MWT and TWL,and there was a dose-dependent effect (all P<0. 001). Levels of IL-1β, IL-6, TNF-α, and GFAP protein expression were significantly reduced (P<0. 05), co-expression of GFAP and Iba-1 was absent, and the mRNA and protein expression levels of Wnt3a, β-catenin, and Cyclin D1 were significantly decreased (P<0. 001). Conclusions Esketamine inhibits astrocyte activation and the inflammatory response, reducing chronic pain. The mechanism may involve regulation of Wnt / β-catenin signaling.