鞘内吗啡预处理对心肌缺血再灌注大鼠脊髓背角SG区神经元兴奋性的影响

Effect of intrathecal morphine preconditioning on excitability of substantia gelatinosa neurons in dorsal horn of spinal cord in a rat model of myocardial ischemia-reperfusion

目的探讨鞘内吗啡预处理对心肌缺血再灌注大鼠脊髓背角胶状质区（SG区）神经元兴奋性的影响。方法鞘内置管成功的成年雄性SD大鼠36只，体重200～300g，采用随机数字表法分为3组（n=12）：假手术组（S组）、缺血再灌注组（I／R组）和鞘内吗啡预处理组（ITMP组）。采用结扎冠状动脉左前降支30min、再灌注120min的方法制备心肌缺血再灌注损伤模型。ITMP组于缺血前30min鞘内输注吗啡3μg／kg（10μl）5min，停止5min，重复3次；I／R组给予等容量生理盐水。随机取6只大鼠，再灌注10min时处死，急性分离T2-6脊髓，制作脊髓切片，采用全细胞膜片钳技术测定脊髓背角SG区神经元静息电位、动作电位阈值（APT）、动作电位峰值（APP）、动作电位时程，记录步阶电流40、60、80、100pA诱发的动作电位个数；随机取6只大鼠，再灌注120min时处死，取心肌组织，确定梗死区体积（IS）与缺血危险区体积（ARR），计算IS／ARR比率；Westernblot法检测T2-6脊髓背角c-fos表达水平。结果与S组比较，I／R组IS／AAR比率升高，脊髓背角c—fos表达上调，SG区神经元动作电位个数明显增加，APT降低，APP升高（P〈0．05）；与I／R组比较，ITMP组IS／AAR比率降低，脊髓背角c—fos表达下调，SG区神经元动作电位个数减少，APT升高，APP降低（P〈0．05）。结论鞘内吗啡预处理减轻大鼠心肌缺血再灌注损伤的机制与降低脊髓背角SG区神经元兴奋性，减轻伤害性刺激反应有关。

Objective To investigate the effect of intrathecal morphine preconditioning （ITMP） on the excitability of substantia gelatinosa （SG） neurons in the dorsal horn of the spinal cord in a rat model of myocardial isehemia-reperfusion （I/R）. Methods Thirty-six adult male Sprague-Dawley rats, weighing 200-300 g, in which intrathecal catheters were successfully placed without complications, were randomly divided into 3 groups （n= 12 each） using a random number table： sham operation group （group S） , group I/R, and group ITMP. Myocardial I/R injury was produced by occlusion of the left anterior descending branch of the coronary artery for 30 min followed by 120 min reperfusion. In group ITMP, the rats received intrathecal morphine 3 μg/kg （ 10 μl） by three cycles of 5 min infusions interspersed with 5 min infusion-free periods starting from 30 min before ischemia, and the equal volume of normal saline was given instead of morphine in group I/R. At 10 min of reperfusion, 6 rats randomly selected in each group were sacrificed, and the T2-6 segments of the spinal cords were acutely isolated to prepare spinal cord slices. The resting potential, threshold of action potential （APT）, peak of action potential （APP） and action potential duration in SG neurons in the dorsal horn of spinal cord slices were determined using the whole-cell patch-clamp technique, and the number of action potentials evoked by currents of 40, 60, 80 and 100 pA was recorded. At 120 min of reperfusion, 6 rats randomly selected in each group were sacrificed, and myo- cardial specimens were obtained for determination of myocardial infarct size （IS） and area at risk （AAR） , and IS/AAR ratio was calculated. The expression of c-fos in the T2-6 dorsal horns of the spinal cords was detected by Western blot. Results Compared with group S, the IS/AAR ratio was significantly increased,the expression of c-fos was up-regulated, the number of action potentials in SG neurons in dorsal horns of spinal cord was increased, APT was decreased, and APP was increased in group I/R （P〈0.05）. Compared with group I/R, the IS/AAR ratio was significantly decreased, the expression of c-fos was down-regulated, the number of action potentials in SG neurons in dorsal horns of spinal cord was decreased, APT was increased, and APP was decreased in group ITMP （P〈0.05）. Conclusion The mechanism by which ITMP attenuates myocardial I/R injury is related to decrease in the excitability of SG neurons in the dorsal horn of the spinal cord and reduction of responses to nociceptive stimuli in rats.