穴区机械敏感性Piezo通道在针刺缓解大鼠关节炎性痛启动机制中的作用

Involvement of mechanosensitive Piezo channels at acupoints in the initiation mechanism of acupuncture analgesia on rat ankle arthritis

目的:研究Piezo通道是否在针刺穴位启动机制中发挥作用。方法:雄性SD大鼠随机分为空白组、模型组、针刺治疗组、钆预处理+针刺组、钌红预处理+针刺组、GsMTx4预处理+针刺组和HC-067047预处理+针刺组,每组6~8只。左侧踝关节腔内行完全弗氏佐剂注射建立佐剂关节炎(AA)模型。造模后48h针刺患侧后三里穴,仅单次治疗20 min。各预处理组于针刺前30 min穴区注射相应抑制剂。以患侧足底痛阈和旷场自主行为测试作为评价疼痛的指标。穴区组织间隙内ATP(inters.ATP)用微透析法采集并用萤光素酶法检测。RTPCR和免疫荧光染色用以检测Piezo的表达。结果:与空白组比较,模型组AA大鼠表现出痛觉过敏现象,痛阈和自主行为均显著降低(P<0.01),在针刺后得到显著改善(P<0.01);与针刺治疗组比较,钌红、GsMTx4、HC-067047预处理+针刺组均可显著抑制针刺镇痛效应(P<0.01)。RT-PCR和免疫荧光染色均证实后三里穴区皮肤组织表达Piezo1和Piezo2两种亚型。针刺可上调穴区inters.ATP的峰值浓度和总量,HC-067047可显著抑制该上调作用(P<0.05)。结论:初步揭示穴区局部表达低机械阈值的Piezo通道;与高机械阈值的TRPV4通道类似,Piezo通道可被针刺激活,并参与针刺镇痛的启动机制。

Objective:To investigate whether Piezo channels at acupoints play a role in the initiation mechanism of acupuncture.Methods:Male SD rats were randomly divided into blank group,model group,acupuncture group,gadolinium pretreatment+acupuncture group,ruthenium red pretreatment+acupuncture group,GsMTx4 pretreatment+acupuncture group and HC-067047 pretreatment+acupuncture group,with 6-8 rats in each group.The left ankle joint cavity was injected with complete Freund’s adjuvant to establish an adjuvant arthritis(AA)model.One session of 20 min-acupuncture was applied at left Housanli acupoint 48 hours after modeling.The corresponding inhibitors were injected into acupoint 30 minutes before acupuncture.Pain thresholds in injured plantar and the open field experiment were used as indicators to evaluate the pain levels.The interstitial ATP(inters.ATP)at Housanli acupoint was collected by microdialysis technique and detected with luciferase-luciferin assay.RT-PCR and immunofluorescence staining were used to confirm the expression of Piezo channels.Results:AA rats showed hyperalgesia,decreased pain threshold and autonomic behavior by comparing with blank group(P<0.01).These two indexes were significantly improved by acupuncture(P<0.01).Comparing with acupuncture group,acupuncture with ruthenium red,GsMTx4 and HC-067047 significantly inhibited the analgesic effect of acupuncture(P<0.01).Both RT-PCR and immunofluorescence staining confirmed the expression of Piezo1 and Piezo2 in the skin at Housanli.There were the peak concentration and the total amounts of inters.ATP up-regulated by acupuncture,which was significantly inhibited by the pretreatment of HC-067047(P<0.05).Conclusion:This study preliminarily revealed that the contribution of low-threshold MS Piezo channels can be activated by acupuncture and participate in the initiation mechanism of its analgesic effect,which is similar to TRPV4 channels.