Cholinesterase Inhibitors, Axonal Transport, and Memory

描述(申请人提供)：有机磷(OP)对人类健康构成持续的威胁，因为它们被广泛用作杀虫剂，并可能在恐怖袭击中使用。有机磷农药的急性毒性已经得到了广泛的研究；然而，长期或反复暴露于没有明显急性毒性迹象的有机磷农药水平的后果却知之甚少。此外，有临床证据表明，这种低水平的有机磷农药暴露会导致长期的认知障碍，尽管这种影响的机制尚不清楚。我们实验室的一个长期目标是阐明与慢性低水平OP暴露相关的长期神经行为缺陷的机制，以便开发更有效的治疗策略。我们在最初筹资期间进行的实验结果表明，低水平接触商业杀虫剂毒死蜱，会导致长时间的脉冲抑制(一种前注意加工模式)和空间学习缺陷，而不会显著影响运动功能。此外，毒死蜱与大脑区域中神经营养素受体和胆碱能蛋白的减少有关，这些区域对认知功能很重要。这些缺陷伴随着在体外测量的坐骨神经轴突运输的减少。然而，轴突运输缺陷的分子机制以及这种影响在大脑中发生的程度尚不清楚。这项应用的目的是确定轴突运输改变的机制，并进一步确定低水平OP暴露对认知功能的长期影响。我们的中心假设是，OPs共价修饰参与轴突运输的关键蛋白质，这种修饰损害了支持认知功能的神经通路的功能。为了实现我们的目标，我们提出了三个具体的目标：1)确定慢性低水平暴露于典型OP对注意力和认知灵活性的影响，2)确定长期低水平暴露于典型OP对脑内轴突运输的影响，以及3)确定OP导致轴突运输障碍的分子机制。为了达到这些目标，我们将使用五个选项的连续反应时间任务来评估持续注意，水迷宫任务来衡量消退(一种认知灵活性)，并通过立体定向注射可追踪的右旋糖苷、免疫组织化学和质谱仪来确定OP对大脑轴突运输的影响及其损害的后果。这个项目的意义及其与公共卫生的相关性在于，通过根据OPs对动物信息处理的基本组成部分的长期影响来机械地定义OPs，我们将解决我们对OPs随着时间的推移可能如何影响人类的知识中的一个根本差距。这些实验将有助于更好地了解与一类化学品有关的毒性，这类化学品继续对全球数百万人构成重大环境风险。与公共健康相关的有机磷是一种剧毒化学物质，在我们的环境中几乎无处不在，因此，它们对全球数百万人的健康构成了重大风险。虽然这些制剂的急性毒性已经得到了广泛的研究，但长期低水平接触有机磷的影响(特别是对认知和支持认知的神经过程)的影响却知之甚少。本申请中提出的实验旨在解决啮齿动物模型中的这些问题，方法是根据有机磷对轴突运输(神经元中的基本过程)和认知功能的特定领域(即注意力和认知灵活性)的长期影响来机械地定义有机磷。

DESCRIPTION (provided by applicant): Organophosphates (OPs) pose a constant threat to human health due to their widespread use as pesticides and their potential employment in terrorist attacks. The acute toxicity of OPs has been extensively studied; however, the consequences of prolonged or repeated exposure to levels of OPs that produce no overt signs of acute toxicity are poorly understood. Further, there is clinical evidence that such low-level exposures to OPs leads to prolonged deficits in cognition, although the mechanism for this effect is unknown. One long- term goal of our laboratories is to elucidate the mechanisms responsible for the prolonged neurobehavioral deficits associated with chronic low-level OP exposures such that more effective therapeutic strategies can be developed. The results of our experiments conducted during the initial funding period established that low- level exposures to the commercial pesticide, chlorpyrifos, resulted in protracted deficits in prepulse inhibition (a model of pre-attentive processing) and spatial learning without significantly affecting locomotor function. Further, chlorpyrifos was associated with decreases in neurotrophin receptors and cholinergic proteins in brain regions that are important to cognitive function. These deficits were accompanied by decreases in axonal transport measured in sciatic nerves ex vivo. However, the molecular mechanisms for the deficits in axonal transport and the extent to which such effects on axonal transport occur in the brain are unclear. The objective of this application is to identify the mechanisms responsible for alterations in axonal transport as well as to further define the long-term effects of low-level OP exposure on cognitive function. Our central hypothesis is that OPs covalently modify key proteins that are involved in axonal transport and that such modifications compromise the function of neuronal pathways that support cognitive function. To achieve our objective, we propose three specific aims: 1) Determine the consequences of chronic low-level exposure to representative OPs on attention and cognitive flexibility, 2) Determine the consequences of chronic low-level exposure to representative OPs on axonal transport in the brain, and 3) Identify the molecular mechanisms responsible for OP-induced deficits in axonal transport. To address these aims, we will use a five choice serial reaction time task to assess sustained attention, a water maze task to measure extinction (a form of cognitive flexibility) and stereotaxic injections of traceable dextrans, immunohistochemistry, and mass spectrometry to determine OP effects on axonal transport in the brain and the consequences of its impairment. The significance of this project and its relevance to public health is that by mechanistically defining OPs based on their long-term effects on essential components of information processing in animals, we will have addressed a fundamental gap in our knowledge of how OPs likely affect humans over time. The experiments will contribute to a better understanding of the toxicity associated with a class of chemicals that continues to pose a significant environmental risk to millions of people worldwide. PUBLIC HEALTH RELEVANCE Organophosphates are highly toxic chemicals that are almost ubiquitous in our environment and, accordingly, they pose a significant health risk to millions of people worldwide. While the acute toxicity of these agents has been extensively studied, the effects of chronic low-level exposures to organophosphates (especially on cognition and the neuronal processes that support cognition) are poorly understood. The experiments proposed in this application have been designed to address these issues in the rodent model by mechanistically defining organophosphates based on their long-term effects on axonal transport (a fundamental process in neurons) and specific domains of cognitive function (i.e., attention and cognitive flexibility).