Electro-acupuncture at “Zusanli” （ST36） alleviates anxiety and pain in mice with chronic inflammatory pain-anxiety comorbidity

ObjectiveTo observe the effect of electro-acupuncture （EA） at different acupoints on chronic inflammatory pain-anxiety comorbidity in mice， and to investigate the effect of naloxone on EA induced activation of neurons of the anterior cingulate cortex （ACC）， medial prefrontal cortex （mPFC）， paraventricular nucleus of the thalamus （PVT）， and mediodorsal thalamus nucleus （MD）， so as to explore whether the anti-anxiety effect of EA depends on its analgesic mechanism.MethodsIn the first part of the experiment， the efficacy of EA at different acupoints was compared. Sixty male C57BL/6 mice were randomly divided into control， model， EA Zusanli （ST36）， EA Baihui （GV20）， and EA Shengmen （HT7） groups （12 mice/group）. In the second part， the impact of naloxone on the analgesic and anxiety effects of EA at ST36 was investigated. Forty male C57BL/6 mice were randomly divided into control， model， EA ST36， and EA ST36 + naloxone groups （10 mice/group）. Except for those in the control group， all the mice were subjected to a chronic inflammatory pain-anxiety comorbidity by subcutaneous injection of 20 μL emulsified complete Freund’s adjuvant （CFA） into the hind paw. Mice in the control group received an equivalent volume of PBS. EA （2 Hz/100 Hz， 0.2—0.4 mA） intervention was initiated on the 12th day after modeling， and applied to “Zusanli” （ST36， bilateral） or “Baihui”（GV20）， or “Shengmen” （HT7， bilateral）. The EA intervention was conducted for 30 min， once daily for 6 d. Naloxone was intraperitoneally injected 30 min before EA intervention. Paw withdrawal thresholds （PWTs） were measured using von Frey filaments. The anxiety-like behaviors were assessed using the open field （OF） test and elevated plus maze （EPM） test， separately. The expression of c-Fos in the ACC， mPFC， PVT， and MD was detected by immunofluorescence staining.ResultsCompared with the control group， the model group exhibited a significant decrease in the PWTs， number of entries into the open arms and time spent in the open arms in the EPM test， and in the number of entries into the central area， time spent in the central area， and distance traveled in the central area in the OF test （PPPPPPPPPPPPPConclusionEA at ST36 has a better analgesic and emotional relief effects on mice with comorbid pain and emotion compared to EA at GV20 and HT7. The underlying mechanism may involve the inhibition of neuronal excitability in the ACC， mPFC， MD and PVT. Additionally， the emotional relief effect of EA at ST36 is dependent on its analgesic efficacy， which may be related to the inhibition of neuronal excitability in the ACC and MD.