The Effect Of Electroacupuncture On Changes Of The Phosphorylation Of Capsaicin Receptor In Chronic Morphine Tolerance Rats And The Mechanism Of Electroacupuncture

Opioids have been regarded as the most effective analgesics for management of acute and chronic pain, however, opioid tolerance occurs following chronic drug exposure which limits their usefulness. So, the molecule mechanism and clinic intervene approach of morphine tolerance are aroused general interest. Capsaicin receptor TRPV1 was cloned in 1997 and was considered the conformity of various pain induced by tissue injuries and inflammation. A lot of evidence have proved that capsaicin receptor were directly involved in the formation of inflammatory hyperalgesia. Electroacupuncture(EA) is a treasure of traditional medicine, which has been proved to be an effective analgesic method. Mechanism of acupuncture analgesia is a hot topic in the field of pain research domestic and abroad. Based on this model, the effect of EA on morphine tolerance and the change of phosphorylation of TRPV1 and the mechanism of EA were explored. The results of this study are expected to provide important information for future treatment and studies associated with morphine tolerance.Part one:Effect of electroacupuncture(EA) on morphine tolerance and on the phosphorylation of capsaicin receptor(TRPVl) in dorsal root ganglion in ratsObjective:To investigate the effect of different frequent electroacupuncture(EA) on morphine tolerance and on the phosphorylation of capsaicin receptor(TRPV1) in dorsal root ganglion of rats.Method:Healthy male SD rats,250～280g, were implanted intrathecal catheters through foramen magnum. The eligible rats were injected complete Freund's adjuvant(CFA) to right hind leg ankle to made into model of adjuvant-induced arthritis, and randomized into 4 groups(n=5) 3 days later:Administered intrathecally 10μl saline for control group(group N),10μg(10μl) morphine for morphine group (group M),10μg morphine and 2Hz EA for low frequent EA group(group H),10μg morphine and 15Hz EA for high frequent EA group(group I). The drugs were administered twice daily(8:00AM and 8:00PM) for 7 days. Group H and group I were given 2mA,2Hz and 2mA and 15Hz EA for 30min a day respectively. Mechanical withdrawal threshold (MWT) and thermal paw withdrawal latency (PWL) of rats were examined, and the formation of hyperalgesia was the symbol of morphine tolerance. Lumbar 4 dorsal root ganglion of all groups were taken on the 8th day of administration for further analysis.Results:Mechanical and thermal hyperalgesia examination evinced that:the pain threshold of all groups after inflammation were decreased compared with that before inflammation; change of pain threshold of control group was not significant during the 7 days of administration; pain threshold of group M increased first, which decreased from the 3rd day of administration, and kept on the low level until the 7th day. Pain threshold of rats on the 5th day in group H and group I had no difference compared with the 3rd day, and there was no difference on the 7th day compared with that on the 5th day. Pain threshold of group H were higher than that of group I. The expression of phosphorylated TRPV1 was the highest in group M compared with the other three groups and there are significant difference between group M and other 3 groups (P<0.05). The expression of phosphorylated TRPV1 in group H was the lowest in the 4 groups and there were difference between group H and group I (P< 0.05).Conclusion:Chronic intrathecal morphine exposure can induce morphine tolerance.Low frequent EA can reduce the phosphorylation of TRPV1 and inhibit the formation of morphine tolerance, while the effect of 2Hz EA is better than that of 15Hz EA.Part two:Mechanism of EA in alleviating morphine tolerance Objective:To investigate the effect of ERK/CREB in MAPK signaling pathway on EA treatment,and explore the mechanism of EA in alleviating morphine tolerance.Method:Choose the control group(group N),morphine group(group M) and low frequent EA group (group H) mentioned in part one. P-CREB and ERK1/2 in DRG were detected by immunohistochemisty and western-blot.Results:Phosphorylation of CREB and ERK in group M were higher than the other two groups.Conclusion:ERK-CREB signaling pathway plays a role in the process of EA treatment.