Targeting M1/M3 Muscarinic Receptors to Treat Gestational Pesticide Poisoning

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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.