Role of Tyrosine in MDMA Toxicity

被广泛滥用的苯丙胺类似物3，4-亚甲基二氧基甲基苯丙胺(MDMA，摇头丸) 选择性损伤大脑中5-羟色胺神经元的轴突终末。多条证据表明，多巴胺 (DA)助长了这种毒性。尽管DA明显会产生氧化副产物，如活性氧 和苯二酚，目前尚不清楚DA衍生的氧化物种如何对5-羟色胺末端产生选择性损伤， MDMA诱导的神经毒性的标志。更具体地说，DA如何在5-羟色胺中蓄积的机制 终端对其产生的选择性损害尚不清楚。目前提案的过激假设是，L-- 酪氨酸是多巴胺的氨基酸前体，参与神经退行性变过程。这一假设是基于 我们最近的初步数据显示，体内测量的细胞外酪氨酸浓度增加了5倍 在MDMA之后。与DA不同的是，酪氨酸从外周主动运输到大脑和神经元。而当 酪氨酸是DA神经元合成DA的天然前体，5HT神经元中有高浓度的酪氨酸 可能会产生有害的后果。这一提议的假设框架是，氧化环境 MDMA和高温引起5-羟色胺神经元酪氨酸非酶氧化为DA 前身是DOPA。芳香族氨基酸脱羧酶(AADC)，在5-羟色胺末端，然后会脱羧基 多巴胺到多巴胺，导致DA的积累，从而导致DA衍生的活性氧物种和氧化 在5HT终端内损坏。这种假定的酪氨酸作为MDMA诱导的毒性的媒介的作用是一个新的 一种机制，有效地综合了当前现有的假设，在某种程度上，不一致的观察和 对一个具有凝聚力、理论性和可测试性的框架提出了明显的警告。 具体目的是：(1)证明酪氨酸在5-羟色胺中非酶氧化为DOPA和DA 神经元，(2)L酪氨酸和酪胺在亚甲基四氢叶酸和高温氧化中的作用 应激；(3)评估随后的神经元损伤。使用培养的RN46A 5HT细胞是一种新的方法， 是唯一适合于解决MDMA引起的损害的假想机制。这些实验将直接 测量MDMA、酪氨酸、高温和细胞内形成的单独和相对影响 多巴胺对5-羟色胺神经元内氧化过程的影响以及这些变量如何影响细胞活性。对此的测试 模型与体内微透析研究相结合，提供了一种独特而有力的方法来解决谜团 以前与MDMA诱导的5HT系统神经退行性变有关的问题。

The widely abused amphetamine analogue, 3,4-methylenedioxymethamphetamine (MDMA, Ecstasy) selectively damages the axon terminals of 5-HT neurons in the brain. Several lines of evidence suggest that dopamine (DA) contributes to this toxicity. Although DA clearly produces oxidative by-products such as reactive oxygen species and quinones, it remains unclear as to how DA-derived oxidative species produce selective damage to 5HT terminals, a hallmark of MDMA-induced neurotoxicity. More specifically, the mechanism as to how DA accumulates within 5-HT terminals to produce its selective damage is unknown. The overaraching hypothesis of the current proposal is that L- tyrosine, the amino acid precursor of DA, contributes to the neurodegenerative process. This hypothesis is based on our recent preliminary data indicating a 5-fold increase in the extracellular concentration of tyrosine measured in vivo after MDMA. Unlike DA, tyrosine is actively transported from the periphery and into the brain and neurons. While tyrosine is the natural precursor for DA synthesis within DA neurons, high concentrations of tyrosine in 5HT neurons may have deleterious consequences. The hypothetical framework of this proposal is that the oxidative environment produced by MDMA and hyperthermia in 5-HT neurons causes the non-enzymatic oxidation of tyrosine to the DA precursor, DOPA. Aromatic amino acid decarboxylase (AADC), within the 5-HT terminal then would decarboxylate DOPA to DA, leading to an accumulation of DA and consequently, DA-derived reactive oxygen species and oxidative damage within 5HT terminals. This hypothesized role of tyrosine as a mediator of MDMA-induced toxicity is a novel mechanism that effectively synthesizes current existing hypotheses and to some extent, discrepant observations and apparent caveats into a cohesive, theoretical and testable framework. The Specific Aims are (1) to demonstrate the non-enzymatic oxidation of tyrosine to DOPA and DA within 5HT neurons, (2) to characterize the contributions of L-tyrosine and tyramine to MDMA and hyperthermia-induced oxidative stress and (3) to assess subsequent neuronal damage. The use of cultured RN46A 5HT cells is a novel approach that is uniquely suited to address the hypothesized mechanism of MDMA-induced damage. These experiments will directly measure the individual and relative effects of MDMA, tyrosine, hyperthermia, and the formation of intracellular dopamine on oxidative processes within 5-HT neurons and how these variables affect cell viability. The testing of this model in conjunction with in vivo microdialysis studies, provides a unique and powerful approach to address enigmatic issues previously related to MDMA-induced neurodegeneration of 5HT systems.