Tramadol is a centrally acting analgesic drug able to prevent nociceptor sensitization when administered into the temporomandibular joint (TMJ) of rats. The mechanism underlying the peripheral anti-inflammatory effect of tramadol remains unknown. This study demonstrated that intra-TMJ injection of tramadol (500 μg/TMJ) was able to inhibit the nociceptive response induced by 1.5% formalin or 1.5% capsaicin, suggesting that tramadol has an antinociceptive effect, acting directly on the primary nociceptive neurons activating the nitric oxide/cyclic guanosine monophosphate signaling pathway. Tramadol also inhibited the nociceptive response induced by carrageenan (100 μg/TMJ) or 5-hydroxytryptamine (225 μg/TMJ) along with inhibition of inflammatory cytokines levels, leukocytes migration and plasma extravasation. In conclusion, the results demonstrate that peripheral administration of tramadol has a potential antinociceptive and anti-inflammatory effect. The antinociceptive effect is mediated by activation of the intracellular nitric oxide/cyclic guanosine monophosphate pathway, at least in part, independently from the opioid system.