Objective To observe the activation of astrocytes, microglia and expression of proinflammatory cytokines in the periaqueductal gray (PAG) by Complete Freunds Adjuvant (CFA)induced inflammatory pain. Methods Heat hyperalgesia and mechanical allodynia were observed and the expressions of mRNA, protein of glial markers， and proinflammatory cytokines in the PAG were examined following intraplantar administration CFA in right hindpaw of the rat by behavioral testing, reverse transcription polymerase chain reaction (RT-PCR ), Western blotting, and immunohistochemistry. Results CFA injection induced chronic thermal hyperalgesia and mechanical allodynia. After injection, the thermal hyperalgesia was recovered on Day 14, but the mechanical allodynia lasted more than 21 days. Microglial marker (Cluster of differentiation 14, CD14) and proinflammatory cytokines (Interleukinin-1β, tumor necrosis factorα) showed a significant increase in their expression during all phases (acute, subacute and chronic) of inflammation. GFAP was observed only in PAG at subacute and chronic phases of inflammation. Conclusion The activation of microglia and astrocytes may be involved in the initiation and maintenance of inflammatory pain. Enhanced cytokines expression might play