| ObjectiveTo investigate the role of BDNF-mTOR signaling pathway in chronic restraint stress-induced hyperalgesia.Methods1.Sixteen male C57BL/6 mice(2 months old,20～25g)were randomly divided into two groups: control group(Sham)and chronic restraint stress group(CRS).The chronic restraint stress model started at 8:00 a.m.and ended at 2:00 p.m.once daily for 7 consecutive days.During the stress period,the eating and drinking were forbidden.The pain thresholds were tested on the day before,the third day,the fifth day,the seventh day after the model was established and the first day,the third day after the model ended.The paw withdrawal mechanical threshold(PWMT)and tail-flick latency(TFL)were measured by von Frey filaments and hot water respectively.The brain tissues(anterior cingulate cortex(ACC)and hippocampal dentate gyrus(DG))and spinal cord were collected after behavioral test.Western blot was used to detect the changes of BDNF,Trk B,p-Trk B,mTOR and p-mTOR levels in the anterior cingulate cortex,dentate gyrus of hippocampus and spinal cord.2.48 male C57BL/6 mice(2 months old,20～25g)were randomly divided into 8 groups: vehicle control group(Veh + Sham),vehicle stress group(Veh + CRS),7,8-DHF control group(DHF + Sham),7,8-DHF stress group(DHF + CRS),vehicle control group(Veh + Sham),vehicle stress group(Veh + CRS),rapamycin control group(Rapa + Sham),rapamycin stress group(Rapa + CRS).All drugs were injucted intraperitoneally on the 7th day after behavioral test,PWMT and TFL were evaluated on the 1st and 3rd day after administration,and brain tissues and spinal cord were collected immediately after behavioral test.3.48 Sprague-Dawley(SD)rats(weight,180～200 g)were implanted intrathecal catheter after anesthesia.The rats were randomly divided into 8 groups: vehicle control group(Veh + Sham),vehicle stress group(Veh + CRS),7,8-DHF control group(DHF + Sham),7,8-DHF stress group(DHF + CRS),vehicle control group(Veh + Sham),vehicle stress group(Veh + CRS),rapamycin control group(Rapa + Sham),rapamycin stress group(Rapa + CRS).All drugs were injected intrathecally on the 7th day after behavioral test,PWMT and TFL were tested on the 1st and 3rd day after administration,and brain tissues and spinal cord were collected immediately after behavioral test.Western blot was used to detect the changes of BDNF,Trk B,p-Trk B,mTOR and pmTOR levels in spinal cord.Results1.CRS significantly decreased PWMT and TFL on days 3,5,7,8,and 10(P < 0.05).BDNF and p-mTOR levels in ACC significantly altered in mice exposed to CRS than those in sham mice.In addition,BDNF level significantly decreased(P < 0.05);p-mTOR level significantly increased(P < 0.05);and p-Trk B(P > 0.05),Trk B(P > 0.05),and mTOR(P > 0.05)levels showed no significant difference.BDNF level significantly decreased,whereas p-mTOR level significantly increased in the spinal cord of mice exposed to CRS.Compared with those in the sham group,mice in the CRS group showed no significant alterations in p-Trk B(P > 0.05),Trk B(P > 0.05),and mTOR(P > 0.05)levels.Furthermore,there was no significant difference in levels of BDNF(P > 0.05),p-Trk B(P > 0.05),Trk B(P > 0.05),p-mTOR(P > 0.05)and mTOR(P > 0.05)in the dentate gyrus of the hippocampus between the sham and CRS groups.2.Compared with vehicle group,CRS significantly decreased PWMT(P < 0.05)and TFL(P < 0.05).However,intraperitoneal 7,8-DHF or rapamycin injection did not affect PWMT(P > 0.05)and TFL(P > 0.05).There was no significant difference in Veh + Sham,DHF + Shamh and Rapa + Sham groups(P > 0.05).3.Compared with vehicle group,CRS significantly decreased PWMT(P < 0.05)and TFL(P < 0.05).Intrathecal 7,8-DHF injection significantly improved CRS-induced abnormal behaviors,as indicated by PWMT and TFL(P < 0.05).There was no significant difference in Veh + Sham,DHF + Sham and Rapa + Sham groups(P > 0.05).Compared with vehicle group(Veh + CRS),following 7,8-DHF administration,p-Trk B level significantly increased(P < 0.05),whereas p-mTOR level significantly deceased(P < 0.05).In contrast,BDNF,Trk B,and mTOR levels were not affected by 7,8-DHF administration(P > 0.05).The levels of BDNF,Trk B,p-Trk B,mTOR and p-mTOR were no significant difference between Veh + Sham and DHF + Sham groups(P > 0.05).Compared with Veh + CRS group,intrathecal rapamycin injection significantly improved CRS-induced abnormal behaviors,as indicated by PWMT and TFL(P < 0.05);however,there was no significant difference between Veh + Sham and Rapa + Sham groups(P > 0.05).Compared with vehicle group(Veh + CRS),following rapamycin administration,p-mTOR level significantly deceased(P < 0.05).However,BDNF,Trk B,p-Trk B and mTOR levels were not affected by rapamycin administration(P > 0.05).In addition,the levels of BDNF,Trk B,p-Trk B,mTOR and p-mTOR were no significant difference between Veh + Sham and DHF + Sham groups(P > 0.05).Conclusion1.CRS-induced hyperalgesia is strongly associated with abnormal BDNF-mTOR signaling pathway in the spinal cord.2.Intrathecal injections of 7,8-DHF or rapamycin may offer effective therapies for CRS-induced hyperalgesia. |
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