•EA stimulation increased the PWTs and declined anxiety-like behaviors of CFA rats.•EA stimulation improved the NPS and NPSR protein expressions in ipsilateral ACC.•NPS microinjection increased the PWTs and alleviated the anxiety-like behavior.•NPSR inhibitor microinjection reversed the analgesic and anti-anxiety effects of EA. The neuropeptide S/Neuropeptide S receptor (NPS/NPSR) system is involved in the regulation of anxiety in rodents. Chronic inflammation can induce anxiety. Our lab has observed that electroacupuncture (EA) has a beneficial effect on chronic inflammatory pain and pain-related anxiety; however, the mechanism should be further clarified. In the present study, we used an inflammatory pain model to investigate the role of the NPS/NPSR system in the anterior cingulate cortex (ACC) in the analgesic and antianxiety effects of EA. In an inflammatory pain model, the paw withdrawal thresholds (PWTs) were decreased, pain-related anxiety-like behaviors were induced, and the ipsilateral protein expression of NPS and NPSR was decreased in the ACC. EA stimulation increased the PWTs, reduced pain-related anxiety-like behavior, and enhanced the ipsilateral protein expression of NPS and NPSR in the ACC. NPS microinjection increased the PWTs and decreased pain-related anxiety-like behaviors. Furthermore, an NPSR inhibitor combined with EA reversed the effect of EA on the PWTs and pain-related anxiety-like behaviors. Our results suggest that EA suppresses pain and pain-related anxiety-like behavior of chronic inflammation in rats by increasing the expression of the NPS/NPSR system in the ACC.