脊髓背角神经元上调Nav1.8通道参与缺血再灌注损伤后痛觉过敏的机制

目的观察鞘内注射钠通道抑制剂619C89对脊髓缺血再灌注损伤引起的痛觉过敏大鼠的痛阈、脊髓背角神经元中Nav1.8通道表达的影响。方法 SD大鼠随机分为3组:假手术组(S组)、痛觉过敏组(H组,缺血再灌注前3 d鞘内注射30μL生理盐水)、钠通道抑制剂组(I组,缺血再灌注前3 d鞘内注射619C895μg/30μL)。S组仅暴露主动脉弓而不结扎,其他各组开胸后无创动脉夹夹闭主动脉弓14 min后再开放,建立SCIRI引起的痛觉过敏模型。各组手术前均于L_(5-6)鞘内置管并连续注射3 d。术后1、3、5、7、14 d分别测定各组大鼠的热痛阈及机械性痛阈;取第4~6节腰段脊髓,采用免疫双荧光法观察背角神经元状态及其Nav1.8的表达,Real time-PCR法检测各组大鼠脊髓组织Nav1.8表达。结果与S组相比,术后各观察点(尤以第7天为著)H组大鼠热痛阈和机械性痛阈降低,损伤后7 d脊髓组织中Nav1.8 mRNA的表达增加(P<0.05);I组大鼠热痛阈和机械性痛阈值明显提高(P<0.05),脊髓组织中Nav1.8 mRNA的表达降低(P<0.05)。免疫双荧光染色显示,损伤后7 d,H组大鼠脊髓背角Nav1.8的荧光强度明显增加,且主要表达在NeuN表达阳性的神经元的胞浆中;且与S组相比,H组中NeuN/Nav1.8双阳性的细胞数量明显增多,而I组双阳性的细胞数量减少(P<0.05)。结论脊髓背角神经元通过上调Nav1.8通道参与缺血再灌注损伤后痛觉过敏的形成。

Mechanisms of ischemia-reperfusion induced hyperalgesia via up-regulation of neu-ronal Nav1. 8 channel in spinal dorsal horn

Objective To observe the effects of intrathecal injection (IT) of Nav1. 8 channel inhibitor 619C89 on hyperalgesia and spinal cord levels of neuronal Nav1. 8 expressions in rat model of spinal cord ischemia-reperfusion injury ( SCIRI) . Methods Male Sprague-Dawley rats were randomly divided into three groups:group S, group H (SCIRI+IT NS) and Nav1. 8 channel inhibitor group (group I,SCIRI+IT 5 μg/30 μL 619C89). The lumbar intrath-ecal catheters were implanted in L5-6 of rats and SCIRI models were established by aortic arch occlusion for 14 min. The thermal and mechanical nociceptive thresholds were assessed by paw withdrawal latency ( PWL ) to radiant heat and von Frey filaments. The 619C89 was administered intrathecally for 3 days before surgery. The spinal mRNA expression of Nav1. 8 was assessed by Real time-PCR and double immunofluorescence staining was performed for examination of the distribution of neurons and Nav1. 8 and also quantification of NeuN/Nav1. 8 positive cells of dorsal horn at 1,3,5, 7 and 14 days after surgery. Results Compared with group S,animals in group H had significantly lower mechanical and thermal pain thresholds,but higher spinal mRNA expression of Nav1. 8 ( P<0. 05 ) . Rats in group I had signifi-cantly higher mechanical and thermal pain thresholds and significantly lower mRNA expression of Nav1. 8 compared with those in group H (at any observed time points after IR,but most significantly at 7 days,P<0. 05). Double fluo-rescent staining showed the distribution of increased fluorescence intensity of Nav1. 8 was similar to that of fluorescent staining of NeuN ( neuronal marker) . The number of NeuN/Nav1. 8 positive cells was greatly increased in group H, whereas the number was obviously decreased in group I ( P<0. 05 ) . Conclusion Up-regulation of neuronal Nav1. 8 channel in spinal dorsal horn plays a role in IR-induced hyperalgesia.