Organophosphorus Pesticide Toxicology

描述（由申请人提供）：有机磷（OP）杀虫剂是农药相关人类中毒的主要原因。最近对毒死蜱和二嗪农使用的限制反映了对人类健康，特别是儿童健康的持续关注。OP毒性作用的两个主要靶点是急性毒性的乙酰胆碱酯酶（AChE）和OP诱导的延迟性神经病（OPIDN）的神经病变靶点酯酶（NTE）。本研究的重点是非胆碱能效应，部分基于合作者在无合子乙酰胆碱基因敲除小鼠（AChE -/-）和杂合子NTE基因敲除小鼠（NTE）上的发现。乙酰胆碱酯酶-/-小鼠对毒死蜱牛（CPO）（腹腔内LD50 0.45 mg/kg）与野生型小鼠（LD30 3 mg/kg）相比极为敏感，这表明了一种未知的非乙酰胆碱酯酶靶点对急性致死的重要性。第一个具体目标是在哺乳动物中确定这种非乙酰胆碱酯酶靶点。用[3h -乙基]CPO在体外和离体对AChE -/-型和野生型小鼠脑蛋白进行放射性标记。在AChE -/-和具有极低[3H]CPO水平的野生型小鼠中进行差异蛋白标记，可以选择、纯化和鉴定OP急性中毒的候选替代靶点。第二个目的是以小鼠为模型，建立NTE在OPIDN中的功能。NTE小鼠的NTE活性降低了40%，使它们成为毒理学研究的理想选择。研究将重点关注NTE水平与行为和神经病理改变的关系，NTE和野生型小鼠对辛基氟化膦乙酯和其他OP延迟毒性的敏感性，OPIDN小鼠模型的验证以及降低NTE诱导多动和延迟毒性的机制。第三个目的是确定信号转导途径中断的毒性机制，更具体地说，是由溶血磷脂酶、二酰基甘油脂肪酶和毒蕈碱乙酰胆碱受体介导的毒性机制。最后一个目标是确定OP农药的两个次要靶点的机制和意义：OP在亲核位点磷酸化与大麻素受体-1 （CB1）激动剂位点偶联，改变内源性大麻素的作用；犬尿氨酸甲酰胺酶（KFase）的结构和功能与脑致畸和二嗪农牛作用的关系。关于OP农药毒理学的知识也适用于作为化学战和恐怖主义制剂的OP神经毒气。

DESCRIPTION (provided by applicant): Organophosphorus (OP) insecticides are the principal cause of pesticide-related human poisonings. Recent restrictions on the use of chlorpyrifos and diazinon reflect continuing concern for human health, particularly children. The two principal targets of OP toxicant action are acetylcholinesterase (AChE) for acute toxicity and neuropathy target esterase (NTE) for OP-induced delayed neuropathy (OPIDN). This research focuses on noncholinergic effects, based in part on findings with collaborators on nullizygous AChE-knockout mice (AChE -/-) and heterozygous NTE-knockout mice (NTE ). AChE -/- mice are extremely sensitive to chlorpyrifos oxen (CPO) (intraperitoneal LD50 0.45 mg/kg) compared with their wild-type littermates (LD30 3 mg/kg), establishing the importance of an unidentified non-AChE target for acute lethality. The first specific aim is to identify this non-AChE target in mammals. Mouse brain proteins from AChE -/- and wild-type mice will be radiolabeled in vitro and ex vivo with [3H-ethyl]CPO. Differential protein labeling in the AChE -/- and wild-type mice with very low levels of [3H]CPO will allow selection, purification and identification of the candidate alternate target of OP acute poisoning. The second aim is to establish the function of NTE in OPIDN using mice as the model. The NTE mice have a 40% reduction in NTE activity, making them ideal for toxicological investigations. The studies will focus on the association of NTE levels with behavioral and neuropathological changes, the sensitivity of NTE and wild-type mice to ethyl octylphosphonofluoridate and other OP delayed toxicants, validation of the mouse model for OPIDN and the mechanism by which lowered NTE induces hyperactivity and delayed toxicity. The third aim is to define the mechanisms of toxicity from disruption of signal transduction pathways, and more specifically those mediated by lysophospholipase, diacylglycerol lipase, and the muscarinic acetylcholine receptor. The last goal is to define the mechanisms and significance of two secondary targets of OP pesticides: altered endocannabinoid action by OP phosphorylation at a nucleophilic site coupled to the cannabinoid receptor-1 (CB1) agonist site; kynurenine formamidase (KFase) structure and function relative to teratogenesis and diazinon oxen action in the brain. Knowledge gained on OP pesticide toxicology is also applicable to OP nerve gases as chemical warfare and terrorism agents.