龙血竭总黄酮对酸敏感离子通道的调控及其镇痛作用

Modulation of total flavonoids of Dragon′s Blood on acid-sensing ion channels and its analgesic effect

为研究龙血竭总黄酮(total flavonoids of Dragon′s Blood,TFDB)对背根神经节(dorsal root ganglion,DRG)细胞膜上酸敏感型离子通道(acid-sensing ion channels,ASICs)的影响,分析TFDB通过调控ASICs产生镇痛作用的机制,通过急性分离SD大鼠DRG细胞,用全细胞膜片钳技术,观察记录浓度为0.00001%、0.0001%、0.001%、0.01%和0.1%的TFDB对pH 5.0诱发的ASICs电流的影响;观察TFDB(20 mg·kg^(-1))对冰醋酸致小鼠扭体反应的影响及对完全弗氏佐剂(complete Freund′s adjuvant,CFA)诱导大鼠产生的炎性疼痛的影响;提取完全弗氏佐剂诱导的炎症模型大鼠DRG细胞进行免疫组化实验,观察了TFDB对ASIC3蛋白表达的影响.结果表明:TFDB(20 mg·kg^(-1))可以浓度依赖地抑制ASICs电流它对冰醋酸引起的扭体反应具有显著抑制作用(P<0.001),对完全弗氏佐剂诱导的大鼠炎性痛具有明显镇痛效果(P<0.05),同时可以显著抑制ASIC3蛋白表达(P<0.05).故TFDB能够抑制酸激活的ASICs电流以及下调ASIC3蛋白表达,并对冰醋酸及完全弗氏佐剂诱导的疼痛具有显著抑制效应,且该效应能够被ASICs受体激动剂所抵消,这些均说明TFDB对ASICs的调控可能是其产生镇痛效应的机制之一.

To study the effects of total flavonoids of Dragon′s Blood(TFDB)on the acid-sensitive ion channels in dorsal root ganglion(DRG)neurons,and to analyze the mechanism of the analgesic effect of TFDB on the ASICs,whole-cell patch clamp recordings were performed on acutely isolated rat DRG cells to observe concentrations of 0.00001%,0.0001%,0.001%,0.01%and 0.1%of TFDB against pH 5.0-induced ASICs current.The analgesic effect of TFDB(20 mg·kg^(-1))was observed in the glacial acetic acid-induced writhing in mice and in the complete Freund′s adjuvant(CFA)-induced inflammation pain in rats.DRG cells were extracted from the inflammatory model rats induced by CFA for immunohistochemistry to observe the effect of TFDB on ASIC3 protein expression.The results indicated that TFDB(20 mg·kg^(-1))could inhibit ASICs current in a concentration-dependent manner.It could significantly inhibit glacial acetic acid-induced pain in rats(P<0.001)and had obvious analgesic effect on CFA-induced inflammatory pain in rats(P<0.05).Meanwhile,it could significantly inhibit the protein expression of ASIC3(P<0.05).So TFDB can significantly inhibit the acid-induced ASICs current and down-regulate the expression of ASIC3 protein,and has a significant inhibitory effect on pain induced by glacial acetic acid and CFA,which can be resisted by ASICs receptor agonist.It is indicated that TFDB′s regulation of ASICs may be one of the mechanisms of its analgesic effect.