Organophosphates insecticides (OPs) are common surface water contaminants in both urban and agricultural landscapes. Neurobehavioral effects on larval fish are known to occur at concentrations higher than those reported in the environment. The aim of this study was to perform a comparative analysis of neurobehavioral, molecular, and biochemical responses of four OPs (diazinon, dichlorvos, malathion, methyl-parathion) via the following endpoint measurements: distance traveled, velocity, gene expression (AChE, c-Fos, LINGO-1B, GRIN-1B), enzymatic acetylcholinesterase (AChE) activity, and carboxylesterase (CES) activity. OP exposures (5 hpf - 120 dpf) on embryo-larval zebrafish (Danio rerio) were assessed using a larval zebrafish behavior assay at concentrations: 0.01, 0.1, 10, and 100 μg/L. Individual OPs had varying degrees of neurotoxicity. Significant hypoactivity was observed in the 100 μg/L treatments for diazinon and malathion (p < 0.05) as compared to the controls. Diazinon-exposed larvae exhibited a 26% locomotor decrease, and hypoactivity was observed in malathion-exposed larvae at a reduction of 22% and 29% for distance traveled and velocity, respectively. Gene regulation and enzymatic activity changes were measured for both 0.1 and 100 μg/L exposures across OP treatments. Increased CES activity was observed for the 0.1 μg/L treatments of diazinon and methyl-parathion as well as the 100 μg/L treatment of dichlorvos; meanwhile, decreased CES activity was observed for 100 μg/L treatments of diazinon and malathion. Relative enzymatic activity of AChE was inhibited as compared to the control for the 0.1 μg/L diazinon. No other treatment group exhibited a significant effect on biochemical AChE activity; however, AChE upregulation was observed in the 0.1 μg/L exposure for diazinon, dichlorvos, and malathion. Methyl-parathion was observed to downregulate c-Fos at 0.1 μg/L exposure. Malathion upregulated LINGO-1B at 100 μg/L, a gene associated with neuronal regeneration; meanwhile, downregulation of LINGO-1B was observed for 0.1 μg/L exposure of methyl-parathion. Additional downregulation was observed for GRIN-1B in the 100 μg/L diazinon, 100 μg/L dichlorvos, and 0.1 μg/L methyl-parathion treatments. Exposure of ZF embryos to independent concentrations of 100 μg/L concentrations of diazinon and malathion resulted in hypoactivity and decreased CES activity at 5 dfp. No changes in swimming behavior were observed for either the 0.1 μg/L or 100 μg/L dichlorvos or methyl-parathion treatments. Observations from this study indicate that AChE inhibition may not be the most sensitive biomarker of OP pesticide exposure in zebrafish. Rather, the enzyme CES demonstrated higher sensitivity as a biomarker of OP toxicity.

中文翻译：

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比较分析有机磷酸盐对胚胎幼虫斑马鱼（Danio rerio）的神经行为，分子和酶促作用。

有机磷杀虫剂（OPs）是城市和农业景观中常见的地表水污染物。已知对幼虫鱼的神经行为影响的浓度高于环境中报道的浓度。这项研究的目的是通过以下终点测量值对四种OP（二嗪农，敌敌畏，马拉硫磷，甲基对硫磷）的神经行为，分子和生化反应进行比较分析：行进距离，速度，基因表达（AChE，c -Fos，LINGO-1B，GRIN-1B），酶促乙酰胆碱酯酶（AChE）活性和羧酸酯酶（CES）活性。使用浓度为0.01、0.1、10和100μg/ L的幼虫斑马鱼行为测定法评估胚胎幼虫斑马鱼（Danio rerio）的OP暴露量（5 hpf-120 dpf）。个别OP具有不同程度的神经毒性。与对照组相比，在100μg/ L的处理中对地嗪酮和马拉硫磷存在显着的机能减退（p <0.05）。暴露于二嗪农的幼虫的运动能力降低了26％，而马拉硫磷暴露的幼虫的活动距离和运动速度分别降低了22％和29％。在整个OP处理中，分别测量了0.1和100μg/ L的基因调节和酶活性变化。在0.1μg/ L的二嗪农和甲基对硫磷处理以及100μg/ L的敌敌畏处理中，观察到CES活性增加。同时，观察到100μg/ L的二嗪农和马拉硫磷治疗后CES活性降低。与0.1μg/ L二嗪农的对照相比，AChE的相对酶活性受到抑制。没有其他治疗组对生化AChE活性有显着影响。然而，在0.1μg/ L的二嗪农，敌敌畏和马拉硫磷暴露中，AChE上调。观察到甲基对硫磷在0.1μg/ L暴露下调c-Fos。马拉硫磷以100μg/ L上调LINGO-1B，该基因与神经元再生有关；同时，对甲基对硫磷暴露量为0.1μg/ L时，LINGO-1B表达下调。在100μg/ L二嗪农，100μg/ L敌敌畏和0.1μg/ L甲基对硫磷处理中，还观察到GRIN-1B的其他下调。将ZF胚胎暴露于浓度为100μg/ L的二嗪农和马拉硫磷的独立浓度下会导致机能减退并在5 dfp时降低CES活性。对于0，没有观察到游泳行为的变化。1μg/ L或100μg/ L敌敌畏或甲基对硫磷处理。这项研究的观察结果表明，抑制AChE可能不是斑马鱼中OP农药暴露最敏感的生物标记。相反，酶CES作为OP毒性的生物标志物表现出更高的敏感性。