CAJ | 학술논문

目的探讨脊髓背角自噬与大鼠吗啡耐受形成的关系.方法雄性成年SD大鼠,体重250～ 300 g,取鞘内置管成功的大鼠24只,采用随机数字表法,将其分为3组(n=8):对照组(C组)、吗啡耐受组(M组)和吗啡+自噬增强剂雷帕霉素组(MR组).采用鞘内注射吗啡20 μg,2次/d,连续7d的方法制备吗啡耐受模型.C组给予等容量生理盐水.MR组鞘内注射吗啡20 μg,2次/d,连续7d,并于第3天第2次注射吗啡同时鞘内注射雷帕霉素2.3μg,连续3d.于鞘内注射前及第1、3、5、7天第2次鞘内注射后30 min测定机械缩足反应阈(MWT).最后1次MWT测定结束后1h取L4-6段脊髓背角,采用Western blot法测定总哺乳动物雷帕霉素靶蛋白(mTOR)和磷酸化mTOR(p-mTOR)及自噬标记蛋白LC3Ⅱ的表达.以p-mTOR占总mTOR表达水平的百分比反映mTOR的活性.结果随鞘内注射时间延长,M组和MR组MWT逐渐降低(P＜0.05)；与C组比较,M组和MR组鞘内注射期间MWT升高,脊髓背角mTOR活性降低,LC3Ⅱ表达上调(P＜0.05)；与M组比较,MR组鞘内注射第3、5、7天MWT升高,脊髓背角mTOR活性降低,LC3Ⅱ表达上调(P＜0.05).结论脊髓背角自噬增强是吗啡耐受形成时机体的适应性调节机制,可延缓吗啡耐受形成.
목적 탐토척수배각자서여대서마배내수형성적관계.방법 웅성성년SD대서,체중250～ 300 g,취초내치관성공적대서24지,채용수궤수자표법,장기분위3조(n=8):대조조(C조)、마배내수조(M조)화마배+자서증강제뢰파매소조(MR조).채용초내주사마배20 μg,2차/d,련속7d적방법제비마배내수모형.C조급여등용량생리염수.MR조초내주사마배20 μg,2차/d,련속7d,병우제3천제2차주사마배동시초내주사뢰파매소2.3μg,련속3d.우초내주사전급제1、3、5、7천제2차초내주사후30 min측정궤계축족반응역(MWT).최후1차MWT측정결속후1h취L4-6단척수배각,채용Western blot법측정총포유동물뢰파매소파단백(mTOR)화린산화mTOR(p-mTOR)급자서표기단백LC3Ⅱ적표체.이p-mTOR점총mTOR표체수평적백분비반영mTOR적활성.결과 수초내주사시간연장,M조화MR조MWT축점강저(P＜0.05)；여C조비교,M조화MR조초내주사기간MWT승고,척수배각mTOR활성강저,LC3Ⅱ표체상조(P＜0.05)；여M조비교,MR조초내주사제3、5、7천MWT승고,척수배각mTOR활성강저,LC3Ⅱ표체상조(P＜0.05).결론 척수배각자서증강시마배내수형성시궤체적괄응성조절궤제,가연완마배내수형성.
Objective To investigate the relationship between autophagy in spinal dorsal horn and development of morphine tolerance in rats.Methods Twenty-four healthy male Sprague-Dawley rats,in which intrathecal (IT) catheters were successfully placed,were randomly divided into 3 groups (n =8 each):control group (group C),morphine tolerance group (group M) and morphine + rapamycin as a reinforcing agent for autophagy group (group MR).Morphine tolerance was induced with IT morphine 20 μg twice a day for 7 consecutive days.While the equal volume of normal saline was given in group C.In addition,rapamycin 2.3μg was injected intrathecally at the second injection of morphine on 3rd day lasting for 3 consecutive days in group MR.Mechanical withdrawal threshold (MWT) to yon Frey filament stimulation was measured before IT injection and 30 min after the second IT injection on 1st,3rd,5th and 7th days.The rats were sacrificed 1 h after the last MWT measurement and the L4-6 segment of the spinal cord was removed for determination of the total mammalian target of rapamycin (mTOR) and phosphorylated mTOR(p-mTOR) and autophagy marker protein LC3 Ⅱ expression in spinal dorsal horn by Western blot.The percentage of p-mTOR expression in total mTOR expression was considered as reflection of the activity.Results MWT was gradually decreased with the prolongation of time of IT injection (P ＜ 0.05).Compared with group C,MWT was significantly increased during IT injection,mTOR activity was decreased and LC3 Ⅱ expression was up-regulated in groups M and MR (P ＜ 0.05).Compared with group M,MWT was significandy increased on 3rd,5th and 7th days after IT injection,mTOR activity was decreased and LC3 Ⅱ expression was up-regulated in group MR (P ＜ 0.05).Conclusion Increased autophagy in spinal dorsal horn is the regulatory mechanism of the body during the development of morphine tolerance in rats and can delay the development of morphine tolerance.