1. FK506 and cyclosporin A (CsA) are immunosuppressive drugs, that specifically inhibit T-cell activation via calcineurin inhibition. This study was undertaken to investigate whether calcineurin inhibitors exert analgesic actions in rat adjuvant-induced arthritis (AIA), an animal model of rheumatoid arthritis (RA). 2. AIA was induced in female Lewis rats. Single doses of FK506 and CsA were orally administered to arthritic rats 17 days after arthritis induction. Intensity of hyperalgesia was assessed by measuring the pain threshold of hind paws. Tumor necrosis factor (TNF)-alpha, IL-1beta and PGE(2) levels in paw extracts were determined by ELISA. TNF activity was measured by L929 cell cytotoxicity assay. IL-1beta and cyclooxygenase (COX) mRNA expression in arthritic paws were measured by RT-PCR. 3. Single doses of FK506 and CsA markedly reduced joint hyperalgesia 24 h after drug administration, without affecting inflammation in an advanced stage of AIA. 4. The calcineurin inhibitors partially reduced the elevated level of TNF-alpha in arthritic paws, however, the analgesic effects of these drugs were not associated with the reduction in TNF-alpha level. 5. Moreover, treatment with anti-rat TNF-alpha antibody did not affect the hyperalgesia, when TNF-alpha activity was suppressed in arthritic paws by that treatment. 6. Both calcineurin inhibitors reduced the elevated level of IL-1beta in arthritic paws to a normal level, 24 h after drug administration. 7. FK506 reduced IL-1beta and COX-2 mRNA expression and PGE(2) level in arthritic paws. 8 In conclusion, calcineurin inhibitors rapidly reduce joint hyperalgesia probably by downregulating IL-1beta, but not TNF-alpha, in AIA. Our findings may provide a new strategy for the treatment of pain in RA.