Identifying Molecular Targets for the Proconvulsant Activity of TETS

确定 TETS 促惊厥活性的分子靶点

基本信息

批准号：
9905564
负责人：
Pamela J Lein
金额：
$ 18.7万
依托单位：
UNIVERSITY OF CALIFORNIA AT DAVIS
依托单位国家：
美国
项目类别：
财政年份：
2019
资助国家：
美国
起止时间：
2019-07-01 至 2022-06-30
项目状态：
已结题

来源：
https://reporter.nih.gov/project-details/9905564
关键词：
Acute Address Aminobutyric Acids Behavior Behavioral Binding Sites Biological Biophysics Cell Culture Techniques Cell Line Cells Cessation of life Chemicals Chlorides Convulsants Data Dose Electrophysiology (science)Environment Environmental Pollution Exhibits Exposure to Genes Genetic Goals Half-Life Hippocampus (Brain)Human In Vitro Individual Intoxication Larva Lethal Dose 50 Life Maps Mediating Membrane Methods Military Personnel Modeling Molecular Mechanisms of Action Molecular Target Mus Natural Products Neonatal Neurons Orthologous Gene Pattern Pentylenetetrazole Pharmaceutical Preparations Picrotoxin Plants Production Property Recombinants Risk Assessment Rodent Rodenticides Security Seizures Signal Transduction Site Site-Directed Mutagenesis Soil Specificity Status Epilepticus Structure Substance of Abuse Synapses Techniques Testing Toxicology Triazines United States Whole Organism Zebrafish base experimental study extracellular gamma-Aminobutyric Acid ground water in vivo inhibitor/antagonist insight knock-down medical countermeasure molecular modeling molecular site novel patch clamp positive allosteric modulator receptor superior colliculus Corpora quadrigemina tetramethylenedisulfotetramine zebrafish genome

项目摘要

Inhibition of γ-aminobutyric acid type A (GABAA) receptors (GABAAR) is the presumed mechanism of the seizure-inducing activity of the natural product picrotoxin (PTX), the rodenticide tetramethylenedisulfotetramine (TETS) and the high-energy explosive hexahydro-1,3,5-trinitro-1,3,5-triazine (RDX). TETS, which has been banned from production worldwide, is still readily available on the black market, and is associated with thousands of human poisonings per year. It is incredibly stable in the environment. RDX is an environmental contaminant found in both groundwater and soil due to its worldwide military and civilian use, and it is an illicit abuse substance. PTX is of toxicological concern, because like TETS and RDX, it is listed as a credible threat agent by the United States Department of Homeland Security. All three compounds can cause seizures that rapidly progress to status epilepticus and death; however, TETS is by far the most potent with a lethal dose of 7 to 10 mg in humans and an LD50 of 0.1 mg/kg in rodents, which makes it roughly ~40x more potent than PTX and ~1000x more potent than RDX. There currently is no approved medical countermeasure for individuals acutely intoxicated with these convulsant chemicals. While some information is available regarding the molecular site of action and the GABAAR subtype selectivity of PTX, practically nothing is known about the molecular mechanism of action of TETS and RDX other than that they are most probably GABAAR inhibitors. We intend to use molecular modeling, whole-cell patch-clamp electrophysiology and gene knockdown techniques in zebrafish to identify the subunit specificity of TETS and RDX interactions with GABAA receptors and to determine whether it differs from that of picrotoxin. Treatment with subtype selective compounds/drugs should allow us to confirm which GABAA receptor subunit combinations are important for the seizure activity of TETS and RDX, and whether they differ from the receptor subunit profile that mediate PTX action. Detailed understanding of the molecular mechanism(s) of action of TETS and RDX will not only provide novel insight as to the biological reasons for the toxicologic differences between these agents, but will also be important for evaluating the validity of “read across” risk assessment approaches for GABAA receptor antagonists, and for developing effective medical countermeasures for terminating SE in intoxicated individuals.

推测 A 型 γ-氨基丁酸 (GABAA) 受体 (GABAAR) 的抑制机制天然产物印防己毒素 (PTX)（杀鼠剂四亚甲基二磺四胺）的癫痫诱导活性 (TETS) 和高能爆炸物六氢-1,3,5-三硝基-1,3,5-三嗪 (RDX)。在世界范围内禁止生产，但在黑市上仍然很容易买到，并且与每年有数千人中毒，黑索金在环境中非常稳定。由于其在全球范围内的军事和民用用途，在地下水和土壤中发现了污染物，并且它是一种非法物质 PTX 受到毒理学关注，因为与 TETS 和 RDX 一样，它被列为可信威胁。美国国土安全部的代理所有这三种化合物都可能引起癫痫发作。迅速进展为癫痫持续状态和死亡；然而，TETS 是迄今为止最有效的致命剂量人体中 7 至 10 毫克，啮齿类动物中 LD50 为 0.1 毫克/千克，这使得它的效力大约是 PTX 的 40 倍比 RDX 强约 1000 倍，目前尚无批准的针对个人的医疗对策。虽然有一些关于这些令人惊厥的化学物质的信息。分子作用位点和 PTX 的 GABAAR 亚型选择性，实际上对此一无所知 TETS 和 RDX 的分子作用机制除了它们很可能是 GABAAR 抑制剂之外。我们打算使用分子建模、全细胞膜片钳电生理学和基因敲低斑马鱼技术鉴定 TETS 和 RDX 与 GABAA 受体相互作用的亚基特异性并确定其与印防己毒素的治疗是否不同。应该让我们能够确认哪些 GABAA 受体亚基组合对于癫痫发作活动很重要 TETS 和 RDX，以及它们是否与介导 PTX 作用的受体亚基谱不同。了解 TETS 和 RDX 的分子作用机制不仅将提供新的见解这些药物之间毒理学差异的生物学原因，但也很重要评估 GABAA 受体拮抗剂“跨读”风险评估方法的有效性，以及制定有效的医疗对策来终止醉酒个体的 SE。