The Role of the Dopamine Transporter in Psychostimulant Abuse

多巴胺转运蛋白在精神兴奋剂滥用中的作用

• 批准号: 10529326
• 负责人: AURELIO GALLI
• 金额: $ 53.17万
• 依托单位: UNIVERSITY OF ALABAMA AT BIRMINGHAM
• 依托单位国家: 美国
• 财政年份: 2014
• 资助国家: 美国
• 起止时间: 2014-07-01 至 2025-11-30
• 项目状态: 未结题
• 来源: https://reporter.nih.gov/project-details/10529326
• 关键词: American; Amino Acids; Amphetamine Abuse; Amphetamines; Animal Model; BIN1 gene; Behavior; Behavioral; Behavioral Assay; Biochemistry; Biological Models; Brain; Cell membrane; Cessation of life; Cytoplasm; Data; Disease; Distal; Dopamine; Drosophila genus; Electrophysiology (science); Elements; Event; Family member; Geography; Grooming; Hospitalization; Human; Impairment; Locomotion; Measures; Mediating; Membrane Proteins; Methamphetamine; Mississippi; Modification; Molecular; Molecular Conformation; Motivation; Neurons; Outcome; Pharmaceutical Preparations; Pharmacotherapy; Phase; Phosphorylation; Phosphotransferases; Post-Translational Protein Processing; Process; Property; Regulation; Reporting; Rewards; Role; SNAP receptor; Stimulant; System; Testing; Translating; United States; United States Substance Abuse and Mental Health Services Administration; abuse liability; amphetamine use; behavioral phenotyping; behavioral study; biophysical analysis; dopamine transporter; dopaminergic neuron; extracellular; fly; molecular targeted therapies; neuropsychiatric disorder; neurotransmission; opioid epidemic; preference; psychostimulant; receptor; stimulant abuse; syntaxin 1

Project Summary/Abstract: Amphetamines (AMPHs) are psychostimulants commonly used for the treatment of neuropsychiatric disorders. They are also abused, with devastating outcomes. The abuse potential of AMPHs has been associated with their ability to cause mobilization of cytoplasmic dopamine (DA), leading to an increase in extracellular DA levels. This increase is mediated, at least in part, by the reversal of the DA transporter (DAT) function, which causes non-vesicular DA release (DA efflux). This DA efflux is thought to be essential for the psychomotor stimulant properties of AMPHs, a notion supported by evidence that specific inhibition of DA efflux impairs the ability of AMPH to cause locomotor behaviors. To date, no pharmacotherapies are available for the treatment of AMPHs abuse. Therefore, it is essential to understand: a) the molecular mechanisms targeted by AMPH to promote DA efflux; b) whether DA efflux disrupts DA functions in brain, and whether these disruptions support AMPH-induced behaviors; and c) how we can target these mechanisms to impair DA efflux to regulate AMPH behaviors. Previously, using a combination of biochemistry, electrophysiology, as well as behavioral assays, we and others have shown that the DAT N-Terminus (NT) is a critical structural domain for the ability of AMPH to promote specific behaviors. This is because AMPH stimulates DAT NT phosphorylation, which is vital for its ability to cause DA efflux. Furthermore, we provided the first evidence that the human DAT (hDAT) NT engages in direct associations with the soluble N-ethylmaleimide-sensitive factor attachment protein receptor protein, syntaxin 1 (Stx1), and that this interaction, in addition to hDAT phosphorylation, regulates AMPH-induced DA efflux and behaviors. This was the first demonstration that the interaction of a plasma membrane protein with Stx1 is essential for psychostimulant behaviors. Noteworthy is that our data suggest that the strength of this interaction is regulated by AMPH-induced phosphorylation of Stx1. We hypothesize that the functional and behavioral role of the hDAT NT phosphorylation is dictated by its interactions with the plasma membrane protein, Stx1, a process regulated by the phosphorylation status of Stx1. We propose to test this hypothesis through the following specific aims: 1) to determine how hDAT interacts with Stx1; 2) to determine the involvement of Stx1 in AMPH-induced DA efflux. The molecular discoveries of S.A. #1 and S.A. #2 will be evaluated in neurons, in isolated Drosophila brains, and behaviorally, by using Drosophila as an animal model to study AMPH actions. We are now able to “humanize” flies by expressing hDAT in DA neurons of Drosophila lacking the endogenous DAT (KO). In this system, we have established that AMPH- associated behaviors, such as locomotion and grooming, are DAT-dependent. Also, in this animal model, we can now determine preference (reward) and avoidance for AMPH. Thus, specific aim 3) is to determine whether hDAT NT-Stx1 interactions are required for AMPH-induced behaviors.

项目摘要/摘要：苯丙胺（AMPHS）是通常用于的精神刺激因素 神经精神疾病的治疗。他们也受到虐待，结果造成了毁灭性的结果。虐待潜力 AMPHS与引起动员细胞质多巴胺（DA）的能力有关，导致 细胞外DA水平的增加。这种增加至少部分由DA的逆转介导 转运蛋白（DAT）函数，会导致非维数DA释放（DA FEFFLUX）。被认为是 对AMPHS的精神运动刺激特性至关重要，该通知由特定的证据支持 DA外排的抑制会损害AMPH引起运动行为的能力。迄今为止，没有药物治疗 可用于治疗AMPHS滥用。因此，必须理解：a）分子 AMPH旨在促进DA外排的机制； b）DA流出是否会破坏大脑中的DA功能，并且 这些破坏是否支持Amph引起的行为； c）我们如何将这些机制定位为 损害DA外排来调节AMPH行为。 以前，使用生物化学，电生理学以及行为分析的组合，我们和 其他人则表明，DAT N末端（NT）是AMPH促进能力的关键结构域 特定行为。这是因为AMPH刺激DAT NT磷酸化，这对于其能力至关重要 导致DA流出。此外，我们提供了第一个证据，即人类DAT（HDAT）NT参与直接 与固体N-乙基马来酰亚胺敏感因子附着蛋白受体蛋白的关联，语法1 （STX1），除了HDAT磷酸化外，这种相互作用还调节Amph诱导的DA外排和 行为。这是质膜蛋白与STX1的相互作用的第一次证明 对于心理刺激行为必不可少的。值得注意的是我们的数据表明这种相互作用的强度 受AMPH诱导的STX1磷酸化调节。 我们假设HDAT NT辉煌的功能和行为作用由其决定 与质膜蛋白STX1相互作用，该过程由STX1的磷酸化状态调节。 我们建议通过以下特定目的检验这一假设：1）确定HDAT如何相互作用 使用STX1； 2）确定STX1参与AMPH诱导的DA外排。分子发现 S.A.＃1和S.A.＃2将在神经元，孤立的果蝇大脑中进行评估，并在行为上通过使用 果蝇作为研究AMPH作用的动物模型。我们现在能够通过表达HDAT来“人性化” 在果蝇的DA神经元中，缺乏内源性DAT（KO）。在这个系统中，我们已经确定 相关的行为（例如运动和修饰）是依赖于DAT的。另外，在这个动物模型中，我们 现在可以确定偏好（奖励）和避免AMPH的偏好。那是特定目标3）是确定是否 AMPH诱导的行为需要HDAT NT-STX1相互作用。