氯胺酮对甲醛致痛诱导大鼠脊髓背角神经元兴奋性的抑制(英文)

Ketamine inhibits the excitability induced by formalin in spinal dorsal horn neurons of rats

背景:氯胺酮是否可通过影响脊髓水平的伤害性信息的传递而发挥抗伤害作用尚不清楚;一氧化氮在脊髓水平主要参与痛觉过敏的形成和发展,可诱导Fos表达,但其是否参与了氯胺酮对痛信号的转导或调控的机制不明。目的:观察大鼠脊髓对甲醛痛刺激的反应及氯胺酮的影响。设计:均衡随机的动物实验。单位:徐州医学院附属医院麻醉科和江苏省麻醉学重点实验室。材料:实验于2000-01/03在徐州医学院江苏省麻醉学重点实验室进行。取SD大鼠30只,用均衡随机方法分为6组熏甲醛组6只,甲醛+氯胺酮组6只,氯胺酮+甲醛组6只,氯胺酮组6只,甲醛+生理盐水组3只,生理盐水组3只,各组雌雄比例相同。方法:①甲醛组:体积分数为0.05的甲醛200μL一侧前爪掌心皮下注射,刺激1h。②甲醛+氯胺酮组:甲醛痛刺激10min后腹腔注射100mg/kg氯胺酮1h。③氯胺酮+甲醛组:腹腔注射氯胺酮10min后再行甲醛痛刺激1h。④氯胺酮组:腹腔注射同等剂量氯胺酮1h。⑤甲醛+生理盐水组:甲醛痛刺激10min后腹腔注射等容(10mL/kg)的生理盐水1h。⑥生理盐水组:腹腔注射等容生理盐水1h。主要观察指标:①各组大鼠行为学表现。②取脊髓切片,用c-fos基因免疫组化法和NADPH-d组化技术染色,观察大鼠脊髓背角4层(Ⅰ～Ⅱ层,Ⅲ～Ⅳ层,Ⅴ～Ⅵ层,Ⅶ～Ⅹ层)切片Fos样免疫阳性神经元(FLI)和FLI/NOS双标记神经元的数目变化。结果:30只大鼠全部进入结果分析。①行为学变化:甲醛组及甲醛+生理盐水组大鼠注射甲醛后,出现痛反应;注射氯胺酮的大鼠,注射后数分钟内翻正反射消失,无明显的痛行为表现,而呈持续睡眠状态,至灌注时翻正反射仍未恢复。②FLI神经元表达:甲醛组及甲醛+生理盐水组大鼠注射侧脊髓背角出现大量FLI阳性神经元,主要分布在脊髓背角Ⅰ～Ⅱ层;氯胺酮+甲醛组、甲醛+氯胺酮组大鼠脊髓FLI细胞的分布与甲醛组及甲醛+生理盐水组基本相似,但FLI阳性细胞数量显著减少(P<0.01);氯胺酮组和生理盐水组大鼠脊髓未见或偶见FLI阳性细胞。③FLI/NOS双标记神经元表达:氯胺酮+甲醛组、甲醛+氯胺酮组脊髓背角Ⅰ～Ⅱ层双标记神经元数目显著少于甲醛组及甲醛+生理盐水组眼(1±1),(1±1),(7±3),(8±3)个/切片,P<0.01演,氯胺酮组和生理盐水组无表达。结论:同侧相应脊髓节段的某些神经元参与了化学性致痛信息的传导和调控,氯胺酮通过抑制这些神经元的活动而产生抗伤害作用;此作用与抑制脊髓内一氧化氮合酶阳性神经元的活性有关。

BACKGROUND： It is indistinct that whether ketamine can exert antinociceptive effect through influencing the transmission of nocuous information in spinal cord; Nitric oxide （NO） in spinal cord participates mainly in the formation and development of hyperalgesia, and it can also induce Fos protein expression. It is still controversal whether it contributes to the transmission and mediation of ketamine to pain signal. OBJECTIVE： To observe the response to formalin stimulation in spinal cord of the rats and the effect of ketamine. DESIGN： Balanced randomized animal trial. SETTING： Department of Anesthesiology, Affiliated Hospital of Xuzhou Medical Collegei Jiangsu Provincial Key Laboratory of Anesthesiology. MATERIALS： This trial was carried out in the Jiangsu Provincial Key Laboratory of Anesthesiology, Xuzhou Medical College from January to March 2000. Totally 30 Sprague-Dawley rats were chosen and balanced randomized into 6 groups： formalin group （n=6）, formalin ＋ ketamine group （n=6）, ketamine ＋ formalin group （n=6）, ketamine group （n=6）, formalin ＋ normal saline group （n=3） and normal saline group （n=3）. The gender ratio was the same in each group. METHODS： Formalin group： The rats were stimulated for one hour by subcutaneous injection of 0.05 volume fraction of 200 μL in the center of palm of unilateral fore-claw. Formalin ＋ ketamine group： The rats were stimulated for 10 minutes by formalin, then for one hour by intraperitoneal injection of 100 mg/kg ketamine. Ketamine ＋ formalin group： The rats were injected with ketamine for 10 minutes, then with formalin for one hour. Ketamine group： the same dosage of ketamine was intraperitoneally injected into the rats for one hour. Formalin ＋ normal saline group： The rats were stimulated for 10 minutes by formalin, then intraperitoneally given 10 mL/kg normal saline for one hour. Normal saline group： the same volume of normal saline was intraperitoneally injected into the rats for one hour. MAIN OUTCOME MEASURES： ① Behavioral performance of the rats in each group. ② Spinal sections were chosen, and stained with c-los genetic immunohistochemical and NADPH-d histochemical methods. The changes of the number of Fos-like immuno-positive neurons （FLI） and FLI/ultrie oxide synthase （NOS） double-labeled neurons in the 4-layer sections （layer Ⅰ-Ⅱ, layer Ⅲ-Ⅳ, layer Ⅴ-Ⅵ, layer Ⅶ-Ⅹ） of spinal dorsal horn of the rats were observed. RESULTS： All the thirty rats entered the stage of result analysis. ① Behavioral changes： The rats of formalin group and formalin ＋ normal saline group had apparent pain response; Several minutes after injection with ketamine, righting reflex disappeared and did not recover at perfusion period. Prolonged sleep was found without obvious pain response performance. ② FLI neuron expression： λ lot of FLI positive neurons were found in the spinal dorsal horn of injettion side of the rats in the formalin group and formalin ＋ normal saline group, and they distributed principally in the layer Ⅰ-Ⅱ of spinal dorsal horn. The distribution in the ketamine ＋ formalin group and formalin ＋ ketamine group was basically similar to that in the formalin group and formalin ＋ normal saline group, but positive neuron counts were significantly reduced （P 〈 0.01）. ③ The expression of FLI/ NOS double-labeled neurons： The number of double-labeled neurons in the spinal dorsal horn layer Ⅰ-Ⅱ of the rats in the ketamine ＋ formalin group and formalin ＋ ketamine group were significantly less than that in the formalin group and formalin ＋ normal saline group [（1±1）, （1±1）, （7±3）, （8±3）, P 〈 0.01]. CONCLUSION： Some neurons of ipsilateral corresponding spinal segments participate in the transmission and mediation of pain signal. Ketamine can suppress the activities of these neurons and exert antinociceptive effect. The antinococeptive function of ketamine may be caused by the activity depression of the NOS-positive neurons in spinal cord.