Molecular probing for the mechanism of neuropathic pain

描述(由申请人提供)：拟议的R21项目的长期目标是阐明周围神经病理性疼痛的病理机制。据推测，初级感觉神经元发生结构性改变，导致河豚毒素抗性钠通道Nav1.8在外周异常聚集，从而导致神经病变症状，如痛觉过敏、痛觉过敏和感觉障碍。然而，令人费解的是，在这些神经元的体细胞中，NaV1.8的mRNA水平交替地被报告为减少、不变或略有增加。Pi‘s小组最近观察到，坐骨神经卡压(SNE)损伤后，坐骨神经中NaV1.8mRNA的表达显著增加，但脊髓神经结扎(SNL)损伤后，NAV1.8mRNA的表达无明显变化。这表明，选择性操纵感觉神经元基因表达可能为神经病理发病机制的差异性研究提供了独特的机会。这项应用旨在开发一种新的非侵入性基因转移到感觉神经元，使用生物相容的阳离子明胶(CG)载体与质粒DMA复合。皮下注射CG/DNA复合体到无毛的后掌应该导致周围摄取并通过坐骨神经逆行运输到背根神经节(DRG)。初步研究获得了报告基因在L4/L5 DRG中的表达。目标1是一个技术开发项目，将建立CG/DNA神经基因转移的时空动力学。在该模型中还将建立小干扰RNA(SiRNA)的质粒表达。目的2探讨周围坐骨神经NaV1.8mRNA净增加部分参与神经病理性疼痛发病的机制假说。观察大鼠SNE模型背根神经节和坐骨神经中NaV1.8mRNA的空间分布。针对NaV1.8的siRNA将被应用于研究NaV1.8抑制对神经病理性疼痛行为的影响。该项目的结果将为未来神经病理性疼痛的研究提供一个新的切入点，并提供一种假定的治疗方式。

DESCRIPTION (provided by applicant): The long-term goal of the proposed R21 project is to elucidate a pathological mechanism underlying peripheral neuropathic pain. It has been postulated that constitutive changes occur in primary sensory neurons leading to abnormal peripheral accumulation of the tetrodotoxin-resistant sodium channel Nav1.8, with a resulting induction of neuropathy symptoms such as allodynia, hyperalgesia, and dysesthesia. It is puzzling, however, that the NaV1.8 mRNA level has alternately been reported to be reduced, unchanged, or slightly increased in the somata of these neurons. The Pi's group recently observed that NaV1.8 mRNA is significantly increased in the rat sciatic nerve after sciatic nerve entrapment (SNE) injury but not spinal nerve ligation (SNL) injury. This suggests that selective manipulation of sensory neuron gene expression may offer a unique opportunity for differential investigations of neuropathic pathogenesis. This application proposes to develop a novel non-invasive gene transfer to sensory neurons using a biocompatible cationized gelatin (CG) vector complexed with plasmid DMA. Subcutaneous CG/DNA complex injection to the glabrous hindpaw should result in peripheral uptake and retrograde transport to the dorsal root ganglion (DRG) via the sciatic nerve. Preliminary studies achieved reporter gene expression in the L4/L5 DRG. Aim 1 is a technology development project which will establish the spatiotemporal kinetics of CG/DNA neural gene transfer. Plasmid expression of small interfering RNA (siRNA) will also be established in this model. Aim 2 will explore the mechanistic hypothesis that the net increase of NaV1.8 mRNA in the peripheral sciatic nerve contributes, in part, to the pathogenesis of neuropathic pain. The rat SNE model will be examined for the spatial distribution of NaV1.8 mRNA in the DRG and sciatic nerve. NaV1.8-targeted siRNA will be applied to study effects of NaV1.8 suppression on neuropathic pain behavior. The outcome of this project will provide a novel entry point for future investigations of, and a putative therapeutic modality for neuropathic pain.