Neurophysiological and Metabolic Adaptations to Increased Ammonia and Oxygen Starvation in Fishes.

我研究的首要目标是确定水生脊椎动物如何应对环境压力，如体内氨和氧气饥饿的积累。氨是过量氨基酸分解产生的代谢废物，但浓度升高时具有神经毒性。因此，氨需要排泄或转化为毒性较低的含氮废物（n -废物）产品，如尿素或尿酸，这些产品由陆生脊椎动物产生。由于其高水溶性，氨是鱼类的主要氮废物，因为它可以相对容易地通过鳃排泄到水中。然而，鱼类在进食、运动或暴露于高外部氨后，经常需要应对体内氨的波动。在哺乳动物中，将氨解毒为尿素的能力受损（例如由于肝脏疾病）导致体内氨的高度升高。这将导致“兴奋性毒性”，其特征是中枢神经系统（CNS）的过度激活、活性氧的产生和脑含水量的增加，从而导致潜在的致命脑肿胀。相反，我们证明，当暴露于氨和低环境氧时，鲫鱼和金鱼的脑肿胀很容易耐受。这一观察结果表明，这些鱼类具有新的生理机制来保护中枢神经系统免受氨的侵害。在我的NSERC发现工作中，另一个值得注意的发现是，对高浓度氨的耐受性与长期缺氧的生存能力有关。在接下来的5年里，我的学生们将测试这个假设，即这种缺氧-氨交叉耐受性是由于这些鱼有能力防止兴奋性毒性、对氧化应激的高耐受性和对脑肿胀的恢复能力。我的具体研究目标将是：(1)描述在缺氧或缺氧条件下保护金鱼中枢神经系统免受兴奋性毒性细胞死亡的神经生理适应特征；（II）对比氨敏感虹鳟鱼（Oncorhynchus mykiss）和氨耐受金鱼中枢神经系统氨毒性和耐受机制；（三）确定缺氧、低氧或氨暴露的金鱼氧化应激与脑肿胀发展之间的联系；（IV）确定高氨和摄食如何影响太平洋盲鳗和海七鳃鳗这两种能够承受氨和低氧的无颌鱼类的抗氧化能力和神经生理过程。我将使用综合研究来解决这些目标，包括全动物模型，分离线粒体，培养脑切片模型，分子技术，免疫组织化学和电生理学。这项工作将最终提高我们对鱼类应对增加的内部氨和氧饥饿的生理适应的理解，并确定缺氧-氨交叉耐受的潜在机制。

The overarching goal of my research is to determine how aquatic vertebrates cope with environmental stressors such as build-ups of internal ammonia and O2 starvation. Ammonia is a metabolic waste product that arises from the breakdown of excess amino acids, but at elevated concentrations it is neurotoxic. For this reason ammonia needs to be excreted or converted to less toxic nitrogenous waste (N-waste) products such as urea or uric acid, which are produced by terrestrial vertebrates. Due to its high water solubility, ammonia is the primary N-waste product of fishes because it can be excreted with relative ease across the gills to the water. However, fishes frequently have to cope with fluctuations in internal ammonia following feeding, exercise or exposure to high external ammonia. In mammals, an impaired ability to detoxify ammonia to urea (e.g. due to liver disease) leads to highly elevated ammonia in the body. This then results in “excitotoxicity” characterized by hyper-activation of the central nervous system (CNS), reactive oxygen species generation and increased brain water content leading to potentially fatal brain swelling. In contrast, we demonstrated that that brain swelling is readily tolerated by crucian carp and goldfish when exposed to ammonia and low environmental O2. This observation suggests that these fishes possess novel physiological mechanisms to protect the CNS from ammonia. Another notable finding of my NSERC Discovery work was that tolerance to elevated ammonia was linked to an ability to survive prolonged O2 starvation. Over the next 5 years, my students will test the hypothesis that this anoxia-ammonia cross-tolerance is due to the ability of these fishes to prevent excitotoxicity, high tolerance to oxidative stress, and resilience to brain swelling. My specific research objectives will be to: (I) Characterize the neurophysiological adaptations that protect the goldfish CNS from excitotoxic cell death during exposure to hypoxia or anoxia; (II) Contrast the mechanism(s) of ammonia toxicity and tolerance in the CNS of ammonia sensitive rainbow trout (Oncorhynchus mykiss) to those of the ammonia-tolerant goldfish; (III) Identify the link(s) between oxidative stress and the development of brain swelling in goldfish subjected to anoxia, hypoxia or ammonia exposure; (IV) Determine how elevated ammonia and feeding affect antioxidant capacity and neurophysiological processes in the Pacific hagfish and sea lamprey, two jawless fish species able to withstand ammonia and low O2. I will use an integrative research to address these objectives including whole animal models, isolated mitochondria, cultured brain slice models, molecular techniques, immunohistochemistry, and electrophysiology. This work will ultimately improve our understanding of the physiological adaptations used by fishes to cope with increased internal ammonia and O2 starvation, and identify the underlying mechanisms of anoxia-ammonia cross tolerance.