NEUROTOXICITY OF AMMONIA AND SHORT-CHAIN FATTY ACIDS

高氨血症和有机酸血症是许多 脑病(如肝性脑病、丙戊酸脑病、雷伊氏病 疾病、牙买加人的呕吐病)。尽管氨水的证据 和/或短链脂肪酸(SCFA)是这些 脑病并不总是决定性的，最有效的治疗方法是 这些疾病是为了消除高氨血症和器质性疾病。 酸血症。此外，给予氨水和SCFA可诱导小鼠昏迷。 动物和由此产生的神经病理变化显示出惊人的 与雷伊氏病的脑结构异常相似，以及 肝性和丙戊酸脑病。因此，对 氨和SCFA的神经毒性应有助于阐明潜在的 生化生理机制和阐明病理生理学 和/或这些脑病的发病机制。研究概述了这一点 该提案旨在验证我们的假设，即氨和SCFA发挥其毒性 通过(1)干扰三羧酸(TCA)循环活动的影响， (2)改变神经元胞浆的结构和功能 薄膜。基于我们和其他研究的结果，我们进一步提出 这些毒素可以通过改变TCA循环中的 关键的、限速的和/或受调控的酶步骤的活性。我们会 进一步验证这一假设，通过调查氨和 关于(I)限速和/或调节(丙酮酸)活性的SCFA 脱氢酶复合体(PDHC)、柠檬酸合成酶、异柠檬酸脱氢酶 α-酮戊二酸脱氢酶复合体)和非调节型(富马酸酶， 大鼠脑线粒体提取液中的苹果酸脱氢酶)酶，(II) 柠檬酸、异柠檬酸和α-酮戊二酸通过 具有代谢能力的大鼠脑线粒体，以及(Iii) [1-14C]丙酮酸脱羧化(即通过PDHC的通量)和PDHC 完整脑线粒体的激活(即磷酸化)状态。我们 将表征神经元质膜一元酸载体 (MCC)通过研究MCC的底物和抑制剂的特性 大鼠脑突触体的摄取机制。然后，氨的影响 和SCFA对突触体丙酮酸摄取的影响 建立了剂量-效应关系。此外，这种毒素对人体的影响 突触体丙酮酸代谢([1-14C]丙酮酸脱羧化) 学习。对这些毒性机制的阐明可能有助于 为脑病设计更有益的治疗方法，其中 高氨血症和有机酸血症是持续的特征。

Hyperammonemia and organic acidemia are persistent features in many encephalopathies (e.g. hepatic and valproate encephalopathies, Reye's disease, Jamaican vomitting sickness). Although the evidence for ammonia and/or short-chain fatty acids (SCFA) as major toxins in these encephalopathies is not always definitive, the most effective treatments of these diseases are geared toward eliminating the hyperammonemia and organic acidemia. Moreover, administration of ammonia and SCFA induces coma in animals and the resultant neuropathologic changes show striking similarities to the brain structural abnormalities of Reye's disease, and hepatic and valproate encephalopathies. Thus, investigations of the neurotoxicity of ammonia and SCFA should help to elucidate the underlying biochemical and physiologic mechanisms and illuminate the pathophysiology and/or pathogenesis of these encephalopathies. The studies outline in this proposal aim to test our hypothesis that ammonia and SCFA exert their toxic effects by (1) interfering with tricarboxylic acid (TCA) cycle activity, and (2) altering the structure and function of the neuronal plasma membrane. Based on results of our and other studies, we further propose that these toxins could interfere with TCA cycle activity by altering the activities of key, rate-limiting and/or regulated enzymatic steps. We will further test this hypothesis by investigating the effects of ammonia and SCFA on (i) the activities of rate-limiting and/or regulated (pyruvate dehydrogenase complex (PDHC), citrate synthase, isocitrate dehydrogenase, Alpha-ketoglutarate dehydrogenase complex) and non-regulated (fumarase, malate dehydrogenase) enzymes in rat brain mitochondrial extracts, (ii) citrate-, isocitrate- and Alpha-ketoglutarate-supported oxygen uptake by metabolically competent rat brain mitochondria, and (iii) the [1-14C]pyruvate decarboxylation (i.e. flux through PDHC) by and the PDHC activation (i.e. phosphorylation) state in intact brain mitochondria. We will characterize the neuronal plasma membrane monocarboxylic acid carrier (MCC) by studying the substrate and inhibitor specificities of the MCC uptake mechanisms in rat brain synaptosomes. Then, the effects of ammonia and SCFA on synaptosomal pyruvate uptake will be investigated and the dose-effect relationships established. In addition, the toxin's effects on synaptosomal pyruvate metabolism ([1-14C]pyruvate decarboxylation) will be studied. The elucidation of these toxic mechanisms may facilitate the designs of more beneficial therapies for the encephalopathies in which hyperammonemia and organic acidemia are the persistent features.