Development of isoform specific sensory neuronal sodium channel blockers

描述(由申请人提供)：电压门控钠通道是神经元兴奋性的关键决定因素。实验和临床证据清楚地表明，钠通道及其特性的改变可以在慢性疼痛和其他神经疾病中发挥关键作用。然而，至少有六种不同的钠通道亚型在外周感觉神经元中表达，而由于缺乏亚型特异性的神经元钠通道阻滞剂，研究特定的电压门控钠通道亚型在感觉神经元正常和异常电活动中的作用一直受到阻碍。这项R21拨款计划的目标是开发基于生物肽毒素的选择性钠通道阻滞剂，可用于研究特定钠通道异构体在兴奋性疾病(如疼痛)中的作用(S)。生物毒素在研究离子通道在疾病中的作用方面具有不可估量的价值，一些针对离子通道的毒素是重要的治疗药物。我们的初步数据显示，虎纹狼蛛毒液中的一种毒素抑制TTX敏感的神经性钠通道，但不抑制TTX抗性的神经性钠通道、心脏钠通道或骨骼肌钠通道。此外，这些数据表明，该毒素比Nav1.1、1.2和1.3 TTX敏感的钠通道更有效地阻断Nav1.6和Nav1.7 TTX敏感通道。这种毒素是开发异构型特异性感觉神经元钠通道阻滞剂的一个很好的候选分子。本项目有两个相关的具体目标：1.确定钠通道对狼蛛毒素敏感性的异构体特异性决定因素。2.开发对特定电压门控钠通道异构体具有更高选择性的毒素类似物。全细胞膜片钳电生理学、重组离子通道的诱变和多肽合成将被用来研究毒素与通道的相互作用。这里提出的实验将确定使用狼蛛毒素作为开发新工具的基础的可行性，以确定特定的钠通道在感觉神经元正常和异常生理中的作用。这项研究的长期目标是开发钠通道阻滞剂，作为治疗慢性疼痛等高度兴奋的神经疾病的新疗法。

DESCRIPTION (provided by applicant): Voltage-gated sodium channels are critical determinants of neuronal excitability. Experimental and clinical evidence clearly demonstrates that sodium channels and changes in the properties of these channels can play crucial roles in chronic pain and other neurological disorders. However, at least six different sodium channel isoforms are expressed in peripheral sensory neurons, and investigating the role of specific voltage- gated sodium channel isoforms in normal and abnormal electrical activity of sensory neurons has been hindered by the lack of isoform specific neuronal sodium channel blockers. The goal of this R21 grant proposal is to develop selective sodium channel blockers based on biological peptide toxins that can be used to investigate the role(s) of specific sodium channel isoforms in disorders of excitability such as pain. Biological toxins have been invaluable in researching the role of ion channels in disease and some toxins that target ion channels are important therapeutics. Our preliminary data show that a toxin in the venom from the tarantula Ornithoctonus huwena inhibits TTX-sensitive neuronal sodium channels but not TTX- resistant neuronal sodium channels, cardiac sodium channels or skeletal muscle sodium channels. Furthermore, these data indicate that this toxin is more potent at blocking Nav1.6 and Nav1.7 TTX-sensitive channels than Nav1.1, 1.2 and 1.3 TTX-sensitive sodium channels. This toxin is an excellent candidate molecule on which to base the development of isoform specific sensory neuronal sodium channel blockers. This project has two related specific aims: 1. Identify the isoform specific determinants of sodium channel sensitivity to tarantula toxins. 2. Develop toxin analogs with enhanced selectivity for specific voltage-gated sodium channel isoforms. Whole-cell patch-clamp electrophysiology, mutagenesis of recombinant ion channels and peptide synthesis will be used to investigate toxin-channel interactions. The experiments proposed here will determine the feasibility of using tarantula toxins as the basis for developing novel tools to determine the roles of specific sodium channels in normal and abnormal physiology of sensory neurons. The long-term goal of this research is to develop sodium channel blockers that are useful as novel therapeutics for the treatment of neurological disorders of hyperexcitability such as chronic pain.