Targeting M1/M3 Muscarinic Receptors to Treat Gestational Pesticide Poisoning

摘要 急性有机磷（OP）杀虫剂中毒，包括毒死蜱（CPF），家长的重点 R01格兰特，是很好地描述和结果从不可逆抑制乙酰胆碱酯酶（AChE），酶 水解神经递质乙酰胆碱（ACh）。同样令人担忧的是， 与暴露于OP杀虫剂水平相关的健康影响不会引起明显的AChE抑制 不会引起急性中毒这些亚急性OP暴露与增加 神经退行性疾病的患病率，包括阿尔茨海默病（AD）。早期的临床前研究 还报道了CPF及其氧磷代谢产物加重雄性转基因小鼠AD相关的神经病理学 携带与家族性AD（FAD）相关的基因突变，其占所有AD病例的<10%。 散发性AD（SAD）是该疾病的更普遍形式，占所有AD病例的> 90%。悲伤有一个 迟发性和相当复杂的遗传成分，可能有多个基因的突变和多态性 相互影响，并与环境因素相互作用。迄今为止， 以前的补充资料显示，识别记忆缺陷在10 - 8个月后变得明显， 雄性青少年豚鼠每天暴露于非乙酰胆碱酯酶抑制剂量的CPF（2.5 mg/kg/天） 并且这些缺陷伴随着海马肌醇水平的增加（a 在AD中检测到显著的神经元损失之前其水平升高的代谢物）。我们还发现， 通过定量PCR，AD相关基因的表达显著增加， 编码暴露于CPF的雄性豚鼠海马体中的tau蛋白。早期体外研究 已经表明CPF可以破坏血脑屏障（BBB）的完整性。这是至关重要的 因为据报道，A β沉积物通过穿过BBB的运输和星形胶质细胞从大脑中清除， 介导的间质液整体流动，称为胶质淋巴系统。本研究报告的目标 是：（i）分析，通过T1加权动态对比增强（DCE）成像使用钆布醇， 钆基造影剂，暴露后1个月和9个月不同脑区的BBB完整性 青春期豚鼠的非乙酰胆碱酯酶抑制CPF剂量，和（ii）分析，通过微PET扫描 使用18F-Florbetapir（Amyvid），A β积聚，特别是在海马和前额皮质中。 BBB破坏和A β积聚的程度将与认知缺陷的程度相关 通过行为测试和神经变性化学分析来测量。的结果 本研究和上一次补充中产生的结果将作为R01应用的基础 旨在确定有助于由环境因素引起的AD相关病理学的机制。 一个机制框架是迫切需要的假设驱动的研究，旨在确定有效的 治疗以预防和/或阻止这种灾难性疾病的进展。

ABSTRACT Acute poisoning with organophosphorus (OP) insecticides, including chlorpyrifos (CPF), the focus of the parent R01 grant, is well-described and results from irreversible inhibition of acetylcholinesterase (AChE), the enzyme that hydrolyzes the neurotransmitter acetylcholine (ACh). Equally worrisome, however, are the detrimental health effects associated with exposures to OP insecticide levels that do not cause marked AChE inhibition and do not trigger acute intoxication. These subacute OP exposures have been associated with increased prevalence of neurodegenerative diseases, including Alzheimer’s disease (AD). Earlier preclinical studies have also reported that CPF and its oxon metabolite aggravate AD-related neuropathology in male transgenic mice carrying gene mutations associated with familial AD (FAD), which accounts for <10% of all cases of AD. Sporadic AD (SAD) is the more prevalent form of the disease, accounting for >90% of all AD cases. SAD has a late onset and a rather complex genetic component, with mutations and polymorphisms of multiple genes likely interacting with each other and with environmental factors. Results generated to date with funds from a previous supplement revealed that recognition memory deficits become evident 7-8 months after a 10- day exposure of male adolescent guinea pigs to a non-AChE inhibiting dose of CPF (2.5 mg/kg/day) and that these deficits are accompanied by increases in hippocampal levels of myoinositol (a metabolite whose levels rise before significant neuronal loss is detected in AD). We also detected, by means of quantitative PCR, a significant increase in the expression of the AD-related gene that encodes the tau protein in the hippocampus of CPF-exposed male guinea pigs. Earlier in vitro studies have suggested that CPF can disrupt the blood brain barrier (BBB) integrity. This is of utmost relevance because Aβ deposits are reportedly cleared from the brain by transport across the BBB and by the astroglial- mediated interstitial fluid bulk flow referred to as the glymphatic system. The objectives of the present study are: (i) to analyze, by means of T1-weighted dynamic contrast-enhanced (DCE) imaging using Gadobutrol, a gadolinium-based contrast agent, the BBB integrity in different brain regions at 1 and 9 months after exposure of adolescent guinea pigs to non-AChE inhibiting CPF doses, and (ii) to analyze, by means of micro PET scan using 18F-Florbetapir (Amyvid), A accumulation, particularly in the hippocampus and the prefrontal cortex. The magnitude of BBB disruption and A accumulation will be correlated with the degree of cognitive deficits measured in behavioral tests and neurodegeneration assayed immunohistochemically. The results of the present study and those generated in the previous supplement will serve the basis for an R01 application aimed at identifying mechanisms that contribute to AD-related pathology precipitated by environmental factors. A mechanistic framework is critically needed for hypothesis-driven studies aimed at identifying effective treatments to prevent and/or stall progression of this catastrophic disease.