Phosphoproteomics of Extinction and Reconsolidation of Drug Memories

药物记忆消退和重建的磷酸蛋白质组学

• 批准号: 8460543
• 负责人: Mary M Torregrossa
• 金额: $ 15.16万
• 依托单位: UNIVERSITY OF PITTSBURGH AT PITTSBURGH
• 依托单位国家: 美国
• 财政年份: 2011
• 资助国家: 美国
• 起止时间: 2011-06-15 至 2016-04-30
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/8460543
• 关键词: Affect; Agonist; Animal Model; Award; Behavior; Behavior Therapy; Behavioral; Biochemical; Biochemistry; Biological; Brain region; Budgets; Cocaine; Core Facility; Cues; Development; Disease; Drug Addiction; Drug usage; Educational workshop; Endocannabinoids; Environment; Equipment; Extinction (Psychology); Fright; Funding; Future; Gene Transfer; Genetic; Goals; Grant; Histone Acetylation; Incentives; Individual; Institution; Intervention; Laboratories; Lead; Learning; Mass Spectrum Analysis; Mediating; Memory; Mental disorders; Mentors; Mentorship; Methodology; Methods; Molecular; Molecular Biology; NF-kappa B; National Institute of Drug Abuse; Neurobiology; Neuronal Plasticity; Nuclear; Paper; Pharmaceutical Preparations; Pharmacologic Substance; Phosphorylation; Phosphotransferases; Positioning Attribute; Process; Proteins; Proteomics; Psychiatry; Publishing; RNA Interference; Rattus; Relapse; Research; Research Personnel; Scientist; Self Administration; Signal Transduction; Signaling Molecule; Site; Societies; Structure; Students; Techniques; Technology; Testing; Therapeutic Intervention; Time; Training; Treatment Efficacy; Universities; Viral; Viral Vector; Virus; Western Blotting; Writing; addiction; base; behavioral pharmacology; calcineurin phosphatase; cannabinoid receptor; career; cocaine use; conditioned fear; cost; craving; drug addict; drug of abuse; effective therapy; experience; forgetting; inhibitor/antagonist; memory process; motivated behavior; multiple reaction monitoring; non-drug; novel; prevent; professor; protein activation; receptor; research study; responsible research conduct; skills; sound; success; therapy design; tool; transcription factor; vector

DESCRIPTION (provided by applicant): Recently, researchers have been investigating the use of behavioral interventions aimed at disrupting drug- associated memories to treat addiction. One of these interventions is exposure or extinction therapy where cues associated with drugs are repeatedly presented without the drug, so that an addict learns that those cues no longer predict drug availability and they stop producing craving and relapse. Another intervention is to disrupt memory reconsolidation, where a drug cue is presented followed by an intervention designed to make the addict "forget" that the cue was associated with drug use. These methods have been used with some success to treat other psychiatric disorders. While both of these processes sound similar, there is evidence that they actually engage distinct neurobiological processes. For example, pharmacological agents that enhance extinction are also likely to enhance rather than inhibit reconsolidation. Likewise, agents that inhibit reconsolidation are also likely to inhibit extinction. In other words, the same treatment can potentially produce opposite effects on a drug memory depending on whether extinction or reconsolidation is occurring. Therefore, there is a need to find medications that can both enhance extinction and inhibit reconsolidation simultaneously, so that one does not unintentionally increase the likelihood of relapse. In this application, the candidate will conduct studies using proteomics technology to identify proteins that are differentially activated and inhibited by drug memory reconsolidation and extinction processes. The candidate has had substantial training in the behavioral pharmacology of drug addiction. The candidate has also published several papers investigating the neurobiology of both drug memory extinction and reconsolidation. However, the candidate has had little experience with the biochemical techniques required to do proteomic studies, so this proposal includes ample opportunity for structured training in proteomics, biochemistry, and molecular biology that are necessary to do these studies. The candidate's short-term career goal is to obtain training in these techniques and in the successful management of an independent laboratory. In the long-term, the candidate plans to obtain a position as a tenure-track professor at research institution to develop a broad-based, translational laboratory that will be focused on identifying and validating novel treatments for addiction. The training environment at Yale University is excellent both for the ability to complete the proposed experiments and for the training opportunities available to the candidate. Yale University has one of the leading proteomics core facilities in the nation, and experts in the latest technologies available for identifying and quantifying differences in protein phosphorylation. This is essential for the proposed experiments, as one critical component of the hypothesis is that differential activation of proteins by changes in phosphorylation state will allow the identification of novel targets for medication development to treat addiction. In addition, there are several experts within the candidate's division in the Department of Psychiatry who can provide training in biochemical and molecular biological techniques. Training at Yale University provides the advantage of several researchers in close proximity who are experts in the use of the proposed technologies in the field of addiction. The candidate will be mentored by Drs. Jane Taylor, Angus Nairn, Ralph DiLeone, and Erol Gulcicek who all provide a wealth of expertise in the research proposed. The university also provides all of the equipment and laboratory space necessary for completion of these experiments. Finally, the university provides the support, funding, and training opportunities, such as seminars and workshops, to learn new techniques and receive training in grantsmanship, lab management, mentoring, and the responsible conduct of research. The candidate proposes to use phospho-proteomics to identify novel signaling cascades that are differentially activated/inhibited by drug-cue memory reconsolidation and extinction processes. Once targets are identified, they will be validated using secondary confirmation methods including multiple reactions monitoring (MRM) and Western blotting. The targets will then be further validated in a behavioral animal model using known pharmacological agents or RNA interference via viral-mediated gene transfer. The candidate will also determine by Western blotting whether the activity of specific proteins known to be differentially activated/inhibited by reconsolidation and extinction of fear memories are also regulated by drug memories. Then, the candidate will test pharmacological or viral manipulations of these targets, which include cannabinoid receptor 1 agonists and viruses targeting inhibition of nuclear factor kappa B (NFkB) in the animal model. These experiments will provide the candidate with the opportunity to learn several new techniques and has the potential for obtaining novel information about drug memory processes that can lead to future grants as an independent scientist. The proposed experiments could also lead to the development of novel, more effective treatments for addiction that result in a long-term reduction in craving and relapse. The candidate will also spend significant time during the training period learning important skills for managing an independent laboratory, including grant writing, managing budgets, mentoring students and technicians, and training in the responsible conduct of research.

描述（由申请人提供）：最近，研究人员一直在调查旨在破坏药物相关记忆以治疗成瘾的行为干预措施的使用。这些干预措施之一是暴露或灭绝疗法，在没有药物的情况下反复出示与药物相关的提示，因此瘾君子了解到这些提示不再预测药物的可用性，并且停止产生渴望和复发。另一种干预措施是破坏记忆重新溶解，在此提出药物提示，然后进行干预，旨在使瘾君子“忘记”该提示与吸毒有关。这些方法已成功地治疗其他精神疾病。尽管这两个过程听起来都相似，但有证据表明它们实际上参与了不同的神经生物学过程。例如，增强灭绝的药理学剂也可能增强而不是抑制重新溶解。同样，抑制重新溶解的药物也可能抑制灭绝。换句话说，相同的处理可能会对药物记忆产生相反的影响，具体取决于是否发生灭绝或重新溶解。因此，有必要找到可以同时增强灭绝和抑制重新固定的药物，以免无意间增加复发的可能性。在此应用中，候选人将使用蛋白质组学技术进行研究，以鉴定被药物记忆重新溶解和灭绝过程差异激活和抑制的蛋白质。候选人在药物成瘾的行为药理学方面进行了大量培训。该候选人还发表了几篇论文，研究了药物记忆灭绝和重新溶解的神经生物学。但是，该候选人在进行蛋白质组学研究所需的生化技术方面几乎没有经验，因此该建议包括蛋白质组学，生物化学和分子生物学的结构化培训的足够机会，这些机会是进行这些研究所必需的。候选人的短期职业目标是获得这些技术的培训和成功管理独立实验室的培训。从长远来看，候选人计划在研究机构担任终身教授的职位，以开发一个基于广泛的转化实验室，该实验室将着重于识别和验证成瘾的新颖治疗方法。耶鲁大学的培训环境非常出色，既能够完成拟议的实验以及候选人可用的培训机会。耶鲁大学是美国领先的蛋白质组学核心设施之一，并且是可用于识别和量化蛋白质磷酸化差异的最新技术专家。这对于拟议的实验至关重要，因为假设的一个关键组成部分是，通过变化磷酸化状态的差异激活将允许鉴定药物开发的新靶标的来治疗成瘾。此外，精神病学系的候选人部门内有几位专家可以提供生化和分子生物学技术的培训。耶鲁大学的培训为几位紧邻的研究人员提供了优势，这些研究人员是在成瘾领域使用拟议技术的专家。候选人将由博士指导。简·泰勒（Jane Taylor），安格斯·奈恩（Angus Nairn），拉尔夫·迪勒恩（Ralph Dileone）和埃罗尔·吉尔西克（Erol Gulcicek）在拟议的研究中都提供了丰富的专业知识。该大学还提供完成这些实验所需的所有设备和实验室空间。最后，大学提供了支持，资金和培训机会，例如研讨会和研讨会，以学习新技术并接受助学士，实验室管理，指导以及负责任的研究的培训。候选人建议使用磷酸化 - 蛋白质组学来鉴定新的信号级联反应，这些级联反应被药物证明记忆重新溶解和灭绝过程差异激活/抑制。一旦确定了目标，将使用辅助确认方法（包括多个反应监测（MRM）和Western印迹）对其进行验证。然后，将使用已知的药理学剂或通过病毒介导的基因转移来进一步验证靶标在行为动物模型中进一步验证。候选人还将通过蛋白质印迹来确定是否还通过药物记忆来调节，是否通过重新溶解和灭绝恐惧记忆的特定蛋白质的活性以及恐惧记忆的灭绝。 然后，候选人将测试这些靶标的药理或病毒操纵，其中包括大麻素受体1激动剂和靶向抑制动物模型中核因子Kappa B（NFKB）的病毒。 这些实验将为候选人提供学习几种新技术的机会，并有可能获得有关药物记忆过程的新信息，这些信息可能会导致作为独立科学家的未来赠款。提出的实验还可能导致成瘾的新型，更有效的治疗方法的发展，从而导致渴望和复发的长期减少。候选人还将在培训期间花费大量时间学习重要技能，以管理独立实验室，包括赠款写作，管理预算，指导学生和技术人员以及负责任的研究进行培训。