Project 4: Biotransformation Gene-Environment Interactions in Coho Salmon

太平洋鲑鱼的数量在美国西部明显下降。特别令人关注的是，暴露于某些农药和微量金属的鲑鱼发生亚致死性神经损伤。这些行为影响包括捕食者检测和猎物选择的损失，改变生殖时间和归巢的损失。上述在个体中观察到的神经行为效应现在与人口影响有关。鲑鱼的嗅觉系统是环境化学物质神经毒性的敏感目标，包括在超级基金网站中常见的金属和农药。然而，对鱼类化学嗅觉神经毒性的机制知之甚少。我们第一个供资周期的研究产生了重要的发现，这些发现将在我们的竞争性续约中详细探讨。具体而言，我们知道：1）鲑鱼的嗅觉组织是化学生物转化的重要场所，特别是细胞色素P4503A和黄素单加氧酶（FMO）似乎介导有机磷生物转化的组织特异性和化合物特异性差异，对神经毒性具有潜在影响，2）超临界有机磷化合物致嗅损伤模型（毒死蜱）和金属（铜）之间的相互作用涉及嗅觉信号转导途径的中断。然而，铜主要影响G蛋白偶联嗅觉受体信号传导，可能是通过氧化应激，而毒死蜱激活参与抑制嗅觉信号转导的基因，3）转录特征可以帮助我们识别与混合物相关的独特基因靶点，以及区分金属和有机磷驱动的影响。根据我们的调查结果，竞争性更新的目标是：1）使用cDNA克隆、重组蛋白表达、微阵列分析和酶促方法来确定嗅觉CYP3A和黄素单加氧酶在鲑鱼从淡水到盐水的运动期间的有机磷神经毒性中的作用，2）利用原位杂交和免疫组织化学分析结合行为学研究来了解氧化应激在铜和镉介导的嗅觉损伤中的作用，3）使用蛋白质组学方法来识别和区分铜和毒死蜱的重要嗅觉蛋白靶标，4）使用从上述研究中产生的一套嗅觉生物标志物来评估通过超级基金站点迁移的鲑鱼的亚致死嗅觉神经毒性。

Pacific salmon populations have declined markedly in the Western United States. Of particular concern has been sublethal neurological injury occurring in salmon exposed to certain pesticides and trace metals. These behavioral impacts include loss of predator detection and prey selection, altered reproductive timing and loss of homing. These aforementioned neurobehavioral effects observed in individuals are now linked to population impacts. The salmon olfactory system is a sensitive target for the neurotoxicity of environmental chemicals, including metals and pesticides commonly found in Superfund sites. However, little is known about the mechanisms of chemical olfactory neurotoxicity in fish. Studies from our first funding cycle have produced important findings that will be explored in detail in our competitive renewal. Specifically, we know that: 1) the olfactory tissues of salmon are important site of chemical biotransformation, and in particular, cytochrome P4503A and flavin monooxygenases (FMO) appear to mediate tissue- and compound-specific differences in organophosphate biotransformation with potential impacts on neurotoxicity, 2) the olfactory injury by a model superfund organophosphate chemical (chlorpyrifos) and metal (copper) involves disruption of olfactory signal transduction pathways. However, copper primarily impacts G-protein coupled olfactory receptor signaling, likely through oxidative stress, whereas chlorpyrifos activates genes involved in the inhibition of olfactory signal transduction, and 3) transcriptional signatures can help us identify unique gene targets relevant to mixtures, as well differentiating metal- and organophosphate-driven affects. Based upon our findings, the objectives of the competing renewal are to: 1) use cDNA cloning, recombinant protein expression, microarray analysis and enzymatic approaches to determine the role of olfactory CYP3A and flavin monooxygenases in organophosphate neurotoxicity in salmon during movement from freshwater to saltwater, 2) use in situ hybridization and immunohistochemistry analyses coupled with behavioral studies to understand the role of oxidative stress in copper and cadmium-mediated olfactory injury, 3) use proteomics approaches to identify and discriminate important olfactory protein targets of copper and chlorpyrifos, 4) use a suite of olfactory biomarkers generated from the aforementioned studies to assess sublethal olfactory neurotoxicity in salmon migrating through Superfund sites.