TRPV4在介导大鼠背根神经节持续受压后机械和热痛敏中的作用

The role of transient receptor potential vanilloid 4 in mediating mechanical and thermal allodynia following chronic compression of dorsal root ganglion in rats

目的:观察持续机械压迫(CCD)对瞬时感受器电位离子通道4(TRPV4)基因、蛋白表达及功能的影响,明确TRPV4是否参与CCD导致的机械和热痛敏。方法:建立CCD模型后,分别于手术前及手术后第7天、第14天及第28天取材前测量运动功能、机械刺激缩爪反应阈值和热辐射刺激缩爪反应潜伏期。为了测量TRPV4反义核苷酸干扰对机械和热痛阈值的影响,在蛛网膜下腔内注入TRPV4寡脱氧核苷酸(ODN)40μg/d,每天1次,第7天后测量大鼠行为学变化。使用实时定量RT-PCR检测TRPV4基因表达的变化,Western blot检测TRPV4蛋白质表达量的变化,激光共聚焦检测低渗溶液和佛波醇(4α-PDD)刺激背根神经节(DRG)神经元后细胞内钙离子浓度的变化。结果:所有动物在损伤前后步态均正常,持续压迫明显降低大鼠的机械和热痛阈,TRPV4干扰可部分逆转该痛敏。持续机械压迫可以明显增加TRPV4基因和蛋白的表达,手术后第7天,第14天和第28天,TRPV4mRNA的表达分别为假手术组大鼠的4.29倍、2.95倍和2.48倍,蛋白表达量分别为假手术组大鼠的4.34倍,3.88倍和2.47倍。持续机械压迫后,对低渗溶液和4α-PDD产生反应的DRG神经元的比例数增加,细胞内钙的峰值增高。这种反应被TRPV4反义ODN所抑制。结论:CCD可以上调TRPV4的基因、蛋白表达,敏化通道的功能;TRPV4参与介导CCD导致的机械和热痛敏。

Objective：To investigate the role of transient receptor potential vanilloid 4（TRPV4） in mediating mechanical and thermal allodynia in rodent models of chronic compression of dorsal root ganglion （CCD）.Method：The levels of TRPV4 mRNA and protein expressions in dorsal root ganglion （DRG） were assessed using real-time RT-PCR and Western blot analysis respectively at the 7th,14th and 28th d post-CCD.The effects of spinal administration of TRPV4 antisense oligodeoxynucleotide（ODN） and mismatch ODN on CCD-induced mechanical and thermal allodynia were evaluated.The calcium responses to hypotonic solution and 4α-phorbol 12,13-didecanoate （4α-PDD） were assessed following sham surgery,CCD,spinal application of TRPV4 antisense ODN and mismatch ODN.Result：The levels of TRPV4 mRNA and protein expression increased significantly at the 7th-28th d post-CCD with the highest level at the 7th d post-CCD when compared with the sham group.TRPV4 antisense ODN,but not mismatch ODN,partly reversed the CCD-induced mechanical and thermal allodynia.Additionally,TRPV4 antisense ODN had no effect on the baseline nociceptive response.The percentage of DRG neurons responsive to hypotonic solution and 4α-PDD and the fluorescence ratio of calcium response also enhanced significantly in both CCD group and mismatch ODN group.These increased responses were significantly inhibited by TRPV4 antisense ODN.Conclusion：TRPV4 plays a crucial role in CCD-induced mechanical and thermal allodynia.