The Neurophysiological Basis of Ammonia Toxicity and Tolerance in Fishes

鱼类氨毒性和耐受性的神经生理学基础

• 批准号: RGPIN-2015-04248
• 负责人: Wilkie, Michael
• 金额: $ 2.4万
• 依托单位: Wilfrid Laurier University
• 依托单位国家: 加拿大
• 项目类别: Discovery Grants Program - Individual
• 财政年份: 2016
• 资助国家: 加拿大
• 起止时间: 2016-01-01 至 2017-12-31
• 项目状态: 已结题
• 来源: https://www.nserc-crsng.gc.ca/ase-oro/Details-Detailles_eng.asp?id=612853
• 关键词: Neurophysiological; Basis; Ammonia; Toxicity; Tolerance

The overarching aim of my research is to determine how fishes produce and excrete nitrogenous wastes such as ammonia, and how they cope with potentially toxic build-ups of this waste product. Ammonia normally arises from the breakdown of excess amino acids, but has neurotoxic effects at high concentrations. Fish are generally much more tolerant to ammonia than mammals, which need to detoxify ammonia to urea to survive. Yet little is known about the underlying mechanisms for these differences. Our recent work suggests that anoxia tolerant fishes, such as goldfish (Carassius auratus) and crucian carp (Carassius carassius), that have evolved neurophysiological mechanisms to withstand O2 starvation, are also ammonia tolerant. Indeed, the cascade of events that characterize anoxia/ischemia in the brain are similar to those that occur with ammonia toxicity. In mammals, excess ammonia causes overactivation of glutamate receptors in the brain, leading to excitotoxicity characterized by the generation of reactive oxygen species (ROS) and water accumulation by astrocyte cells, culminating in potentially fatal brain swelling. Our findings suggest, however, that fish readily tolerate ammonia-induced brain swelling. I propose to use an integrated approach to test the working hypothesis that the greater ammonia tolerance of fishes compared to mammals is related to a greater ability of the brain to resist the neurotoxic effects of ammonia. I also propose to test the related hypothesis that the physiological adaptations that contribute to the anoxia tolerance of the goldfish, also explains their greater ammonia tolerance. Over the next 5 years, the objectives of my research program will be to: (I) Characterize the underlying mechanism(s) of acute ammonia toxicity and tolerance in the central nervous system of ammonia-tolerant goldfish and ammonia-sensitive trout (Oncorhynchus mykiss); (II) Determine if high tolerance to ROS results in greater ammonia tolerance in goldfish compared to trout; (III) Identify the underlying mechanisms that lead to brain swelling in fishes in response to ammonia and other aquatic stressors; (IV) Determine how prolonged sub-lethal ammonia exposure and feeding affect ammonia tolerance in goldfish and trout. The integrative approach I will use in my research will include whole animal models, cultured brain slice models, molecular techniques, immunohistochemistry, and electrophysiology methods. An improved understanding of the neural mechanism(s) of ammonia toxicity and tolerance in fishes will improve our fundamental understanding how vertebrates cope with ammonia, and shed light on the selective pressures that led to the evolution of different mechanisms of ammonia detoxification and handling in the vertebrates. This work will also have practical implications by better explaining how fishes respond to and tolerate build-ups of ammonia in aquatic ecosystems.

我的研究的总体目的是确定鱼类如何产生和排泄氮废物，例如氨，以及它们如何应对这种废品的潜在有毒堆积。氨通常是由于过量氨基酸的分解而产生的，但在高浓度下具有神经毒性作用。鱼通常比哺乳动物要耐受氨的耐受性，哺乳动物需要将氨们排毒至尿素以生存。然而，对于这些差异的基本机制知之甚少。我们最近的工作表明，耐用鱼类，例如金鱼（Carassius auratus）和克鲁西亚鲤鱼（Carassius carassius），它们已经进化出神经生理机制来承受O2饥饿，也具有氨的耐受性。实际上，表征大脑缺氧/缺血的一系列事件与氨毒性发生的事件相似。在哺乳动物中，多余的氨会导致大脑中谷氨酸受体过度活化，导致兴奋性毒性的特征是活性氧（ROS）产生（ROS）和星形胶质细胞的水积累，最终导致潜在致命的脑肿胀。但是，我们的发现表明，鱼很容易耐受氨诱导的脑肿胀。我建议使用一种综合方法来检验工作假设，即与哺乳动物相比，鱼类的较高氨耐受性与大脑抵抗氨的神经毒性作用的能力更大有关。我还建议测试相关的假设，即有助于金鱼的缺氧耐受性的生理适应性也解释了它们的较高氨耐受性。在接下来的5年中，我的研究计划的目标将是：（i）表征急性氨毒性和耐受性氨含量氨含量金鱼和氨敏感鳟鱼的基本机制（Oncorhynchus mykiss）； （ii）与鳟鱼相比，确定对ROS的高耐受性是否会导致金鱼的氨耐受性更高； （iii）确定导致鱼类脑肿胀的潜在机制，以响应氨和其他水生压力源； （iv）确定长时间的亚致死性氨的暴露和进食如何影响金鱼和鳟鱼中的氨耐受性。我将在研究中使用的综合方法包括整个动物模型，培养的脑切片模型，分子技术，免疫组织化学和电生理方法。对鱼类氨毒性和耐受性的神经机制的改进理解将改善我们的基本了解脊椎动物如何应对氨，并阐明导致脊椎动物中氨解毒和处理不同机制的选择性压力。这项工作还将通过更好地解释鱼类在水生生态系统中如何反应和容忍氨积聚的方式具有实际含义。