Objective To explore the influences of intraperitoneal injection of dexmedetomidine on activation of spinal glial cells and expressions of inflammatory factors in rats with diabetic neuropathic pain (DNP). Methods 32 health male SD rats, weight 180~200g, were divided into four groups (n=8) according to body with the random number table method. The four groups included the control group (group C), diabetic neuropathic pain group (group DNP), dexmedetomidine group (group DEX) and α2 receptor antagonist group (group YOH). Streptozotocin (STZ) was used to establish the DNP model in rats. Starting from 28 d after STZ injection, DEX group was given intraperitoneal injection of dexmedetomidine (50μg/kg) for consecutive 7d, while YOH group was given intraperitoneal injection of yohimbine (0.1mg/kg) 30min before intraperitoneal injection of dexmedetomidine (50μg/kg) for consecutive 7d. Mechanical withdrawal threshold (MWT) was determined in each group of rats 29 -35d after STZ injection. Rats were sacrificed after MWT detection on 35d. Immunofluorescence double-staining was used to determine activation of microglia and astrocytes in rat spinal cord of each group. ELISA was used to determine TNF-α and IL-1 in rat spinal cord of each group. Results Compared with group C, rat body weight was decreased while blood glucose was increased in group DNP, DEX and YOH (P<0.05); but there were no statistically significant differences among group DNP, DEX and YOH in rat body weight and blood glucose (P>0.05). Compared with group C, MWT at each time point was decreased in group DNP and YOH (P<0.05); while compared with group DNP, MWT at each time point after operation was increased in group DEX (P<0.05). Compared with group C, activation rate of microglia and content of TNF-α and IL-1 were all increased in rat spinal cord of group DNP and YOH (P<0.05); compared with group DNP, activation rate of microglia and content of TNF-α and IL-1 were all decreased in rat spinal cord of group DEX (P<0.05); there were no statistically significant differences among groups in activation rate of spinal astrocytes (P>0.05). Conclusion Dexmedetomidine can relieve hyperalgesia in rats with diabetic neuropathy pain, and the mechanism may be related to inhibition of microglia activation and reduction of the inflammatory reaction in the spinal cord.