Targeting M1/M3 Muscarinic Receptors to Treat Gestational Pesticide Poisoning

摘要 有机磷(OP)杀虫剂急性中毒，包括毒死蜱(CPF)，父母的重点 R01 GRAT是众所周知的，是由于乙酰胆碱酯酶(AChE)不可逆转地抑制该酶的结果 它能分解神经递质乙酰胆碱(ACh)。然而，同样令人担忧的是， 暴露于不会造成明显AChE抑制的OP杀虫剂水平对健康的影响 并且不会引发急性中毒。这些亚急性OP暴露与 神经退行性疾病的流行，包括阿尔茨海默病(AD)。早期的临床前研究已经 还报道了CPF及其氧代谢产物加重了雄性转基因小鼠AD相关的神经病理 携带与家族性阿尔茨海默病(FAD)相关的基因突变，占所有AD病例的10%。 散发性阿尔茨海默病(SAD)是更常见的疾病形式，占所有AD病例的90%。悲伤有一种 发病晚，遗传成分相当复杂，可能有多个基因的突变和多态 相互作用，并与环境因素相互作用。到目前为止使用来自 之前的补充资料显示，认知记忆缺陷在10-8个月后变得明显 雄性青春期豚鼠每日暴露于非AchE抑制剂量的CPF(2.5 mg/kg/d) 这些缺陷伴随着海马肌肌醇(A)水平的增加 在阿尔茨海默病中检测到显著神经元丢失之前水平升高的代谢物)。我们还检测到， 通过定量聚合酶链式反应的手段，AD相关基因的表达显著增加 在暴露于CPF的雄性豚鼠的海马区编码tau蛋白。早期的体外研究 表明CPF可以破坏血脑屏障(BBB)的完整性。这一点至关重要。 因为据报道，Aβ的沉积是通过血脑屏障和星形胶质细胞从大脑中清除的- 介导的间质体液流动称为淋巴系统。本研究的目标 (I)通过使用加多布特洛尔的T1加权动态对比增强(DCE)成像，分析 基于Gd的造影剂，暴露后1个月和9个月不同脑区的血脑屏障完整性 对非AChE抑制CPF剂量的青春期豚鼠，以及(Ii)通过微型PET扫描进行分析 使用18F-FLOBETAPIR(AMYVID)，A积聚，特别是在海马体和前额叶皮质。 血脑屏障紊乱的程度和A积聚将与认知障碍的程度相关 用行为测试和免疫组织化学方法检测神经退行性变。评选结果 本研究和前一补充中生成的研究将作为R01应用的基础 旨在确定环境因素导致AD相关病理的机制。 对于以假设为驱动的研究来说，迫切需要一个机械的框架，目的是确定有效的 预防和/或延缓这一灾难性疾病进展的治疗。

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.