敲除缝隙连接蛋白Cx 43对针刺抑制内脏痛小鼠脊髓背角c-fos表达的影响

目的:探讨缝隙连接蛋白43(Cx43)影响针刺镇痛作用的可能机制。方法:将野生型(WT)和Cx43基因敲除杂合子(HT)小鼠随机分为6组:WT对照组、WT模型组、WT针刺组、HT对照组、HT模型组、HT针刺组,每组12只,腹腔注射0.6%醋酸(0.1ml/10g)制造内脏痛模型。针刺"中脘"、双侧"足三里"穴,每5min捻针30s,共30min。采用瞬时基因c-fos表达作为神经元活动的标志,用RT-PCR、免疫印迹法检测脊髓背角c-fos基因的表达。结果:HT和WT对照组小鼠脊髓背角c-fosmRNA和蛋白很少表达,两组间差异无显著性意义(P>0.05)。HT和WT内脏痛模型组小鼠脊髓背角c-fosmRNA和蛋白表达水平较对照组明显升高(P<0.01);两模型组间上述指标差异无显著性意义(P>0.05)。与模型组比较,WT针刺组小鼠脊髓背角c-fosmRNA和蛋白表达水平明显降低(P<0.01);HT针刺组小鼠c-fosmR-NA和蛋白水平虽然亦有下降趋势,但与模型组比较差异无显著性意义(P>0.05)。HT和WT针刺组两组间差异分别有显著性(P<0.05)和极显著性(P<0.01)意义。结论:针刺可缓解腹腔内脏痛,同时抑制伤害性痛反应诱发的脊髓c-fos表达;敲除Cx43基因可减弱针刺镇痛的作用,同时减弱针刺下调脊髓背角c-fos表达的作用;提示Cx43与针刺镇痛效应有着密切联系。

Effect of Connexin 43 Knockout on Acupuncture-induced Down-regulation of c-fos Expression in Spinal Dorsal Horn in Visceral Pain M ice

OBJECTIVE: To explore the effect of connexin 43 gene knockout on acupuncture analgesia. METHODS: Seventy-two wide type (WT) and connexin 43 gene knockout mice were separately and randomly divided into: WT control group, WT model group, WT acupuncture group, heterozygous (HT) control group, HT model group and HT acupuncture group, with 12 cases in each. Visceral pain model was established by intraperitoneal administration of acetic acid. "Zhongwan" (CV 12) and bilateral "Zusanli" (ST 36) were punctured with a filiform needle for 30 min and stimulated by manipulating the needle 30 s every 5 min. The expression of c-fos in the spinal dorsal horn was assayed by using RT-PCR and western blot techniques. RESULTS: There was no significant difference between HT and WT control mice in relative grey value of spinal c-fos mRNA expression (P>0.05), in which few c-fos mRNA and protein expressed. The expression of c-fos mRNA and protein was increased significantly following intraperitoneal acetic acid injection compared with control groups in both HT and WT mice (P<0.01). And no significant difference was found between HT and WT model groups in c-fos mRNA expression (P>0.05). Compared with WT model group, the expression of both c-fos mRNA and c-fos protein in WT acupuncture group was down-regulated significantly (P<0.01). In comparison with HT model group, the expression of both c-fos mRNA and protein in HT acupuncture group was down-regulated but without statistically significant difference (P>0.05). And the expression of c-fos mRNA and protein in HT acupuncture group was significantly higher than that in WT acupuncture group (P<0.05, 0.01). CONCLUSION: Acupuncture has a marked antinociceptive effect in visceral pain mice, and simultaneously suppresses the expression of c-fos mRNA and protein evoked by noxious stimulation in the spinal dorsal horn. Connexin 43 gene knockout may weaken acupuncture analgesia and reduce EA-induced down-regulation of c-fos expression, suggesting an involvement of connexin 43 in the analgesic effect of acupuncture.