The aim of the present study was to investigate the effect of nursing cooperation on the post-operative complication of pain following total knee arthroplasty (TKA) and to explore the effects of tetramethylpyrazine (TMP) application on post-TKA pain. A total of 26 patients who received TKA between June 2014 and March 2016 were enrolled in this study. Nursing cooperation was provided to the patients during the TKA surgery, and pain was evaluated based on the visual analog scale (VAS). In addition, 40 male Sprague Dawley rats were used for the TKA model construction. The rats were randomly separated into 4 groups (sham, TKA, TKA+TMP and TKA+TMP+Interferon γ). Pain tolerance in rats was evaluated by mechanical stimulation. Inflammatory cytokine levels in TKA rat tissue were detected using ELISA. mRNA and protein expression of Janus kinase/signal transducer and activator of transcription 3 (JAK/STAT3) was detected using reverse transcription-polymerase chain reaction and western blot analysis, respectively. The results indicated that nursing cooperation serves a critical function during TKA and was associated with a lower level of pain compared with the control (P<0.05). Furthermore, TMP treatment reduced the level of inflammatory cytokines in the rat tissues, including interleukin (IL)-6, IL-10 and tumor necrosis factor-α in post-TKA (P<0.01). TMP was indicated to alleviate pain in post-TKA through suppressing the JAK/STAT3 signaling pathway. The results of the present study suggest that nursing cooperation is critical to TKA, and TMP may alleviate post-TKA pain through inhibiting the JAK/STAT3 signaling pathway.