Neuronal subtype and circuit-specific epigenetic mechanisms in addiction

成瘾中的神经元亚型和回路特异性表观遗传机制

基本信息

批准号：
9260817
负责人：
Gregory Charles Sartor
金额：
$ 13.1万
依托单位：
UNIVERSITY OF MIAMI SCHOOL OF MEDICINE
依托单位国家：
美国
项目类别：
财政年份：
2016
资助国家：
美国
起止时间：
2016-05-01 至 2018-07-31
项目状态：
已结题

来源：
https://reporter.nih.gov/project-details/9260817
关键词：
Alpha Cell Anatomy Animal Model Area Attenuated BRD2 gene Behavior Binding Proteins Brain region Bromodomain Canine Adenoviruses Chromatin Chronic Clinical Trials Cocaine Cocaine Dependence Complex Corpus striatum structure Data Development Dissection Dopamine Drug Addiction Emotional Enterobacteria phage P1 Cre recombinase Epigenetic Process Faculty Family Fluorescence-Activated Cell Sorting Functional disorder Gene Expression Gene Transfer Gene Transfer Techniques Genes Genetic Transcription Goals Grant Gray unit of radiation dose Histone Acetylation Histones Individual Injectable Journals Lysine Macromolecular Complexes Mediating Mentors Messenger RNA Methods Modeling Modification Molecular Molecular Biology Techniques Molecular Genetics Neurons Nucleus Accumbens Pathway interactions Pharmacology Phase Play Population Positioning Attribute Presynaptic Terminals Promoter Regions Proteins Publishing Rattus Reader Records Recruitment Activity Request for Proposals Research Rewards Role Scientific Advances and Accomplishments Self Administration Societies Supervision Technical Expertise Technology Time Tracer Training United States National Institutes of Health Viral addiction behavioral pharmacology behavioral response career career development cell type chromatin modification craving drug seeking behavior economic cost effective therapy epigenetic drug epigenetic regulation global health improved innovation knock-down member nano-string neural circuit neuroadaptation neuronal cell body novel preference protein complex protein expression public health relevance receptor relating to nervous system retrograde transport scaffold skills small hairpin RNA tool

项目摘要

DESCRIPTION (provided by applicant): This proposal requests support for a comprehensive training plan that will enable the candidate, Dr. Gregory C. Sartor, to expand, develop, and refine technical skills that are necessary for a productive independent research career. The goal of the training plan is to acquire advanced molecular biology techniques through supervised hands-on training and didactic coursework in order to incorporate sophisticated methods into Specific Aims focused on elucidating cell type- and circuit-specific epigenetic mechanisms in addiction. The advanced scientific training will be focused on two key areas: 1) Fluorescence-activated cell sorting (FACS) of neuronal subtypes from addiction-related brain regions, and 2) developing viral-mediated gene transfer tools to manipulate epigenetic targets in a cell type- and connection-specific fashion. The career development activities will be mentored by Dr. Claes Wahlestedt with collaborative support from Drs. Kasahara and Gray, all of which have non-overlapping expertise in the methods proposed and have exceptional records of mentoring postdoctoral trainees. The research component will investigate acetyl lysine reader proteins, termed BET bromodomains, an emerging, drugable epigenetic target that is currently being studied in multiple clinical trials. These bromodomain-containing proteins bind acetylated histones and serve as a scaffold for the recruitment of macromolecular complexes that modify chromatin accessibility and transcriptional activity. I recently found that BET proteins are upregulated and recruited to the promoter region of Bdnf in the nucleus accumbens (NAc) following chronic cocaine administration. Furthermore, my data show that pharmacological inhibition of these BET proteins (systemic and intra-NAc) attenuates behavioral responses to cocaine. However, recent evidence indicates that cocaine-induced behaviors and associated chromatin modifications are regulated in a cell type-specific manner in the NAc. Therefore, precise examinations of BET proteins within this anatomically and functionally heterogeneous region are critically needed to fully understand the role of these proteins in complex addiction behaviors. My overall hypothesis is that modulation of epigenetic reader proteins in a cell type- and projection-dependent manner will attenuate cocaine seeking behaviors. Aim 1 will require training in FACS in order to determine the cell-type specific time course of BET mRNA/protein expression following short and long access to cocaine self-administration. Aim 2 will require proficiency in viral-mediated gene transfer techniques in order to manipulate BET proteins in specific cell types within the NAc during cocaine seeking behaviors. The R00 independent phase builds on this training to focus more specifically on connection-specific changes and manipulations of BET proteins during cocaine seeking behaviors. Together, these innovative studies will be the first to systemically explore epigenetic targets in cell type- and connection- specific manner during complex behaviors.

描述（由申请人提供）：该提案要求支持一项全面的培训计划，该计划将使候选人 Gregory C. Sartor 博士能够扩展、发展和完善富有成效的独立研究生涯所需的技术技能。培训计划的重点是通过监督的实践培训和教学课程获得先进的分子生物学技术，以便将复杂的方法纳入具体目标，重点是阐明成瘾中的细胞类型和电路特异性表观遗传机制。专注于两个关键领域：1）对成瘾相关大脑区域的神经亚型进行荧光激活细胞分选（FACS）；2）开发病毒介导的基因转移工具，以细胞类型和连接特异性的方式操纵表观遗传目标。开发活动将由 Claes Wahlestedt 博士在 Kasahara 博士和 Gray 博士的合作支持下进行指导，他们在所提出的方法方面拥有不重叠的专业知识，并且在指导博士后实习生方面拥有出色的记录。该组件将研究乙酰赖氨酸读取蛋白，称为 BET 溴结构域，这是一种新兴的可药物表观遗传靶点，目前正在多项临床试验中进行研究。这些含有溴结构域的蛋白结合乙酰化组蛋白，并作为招募修饰染色质的大分子复合物的支架。我最近发现 BET 蛋白被上调并被招募到伏隔核中 Bdnf 的启动子区域。此外，我的数据表明，这些 BET 蛋白（全身和 NAc 内）的逻辑药理学抑制会减弱对可卡因的行为反应。然而，最近的证据表明可卡因诱导的行为和相关的染色质修饰受到调节。因此，迫切需要对这个解剖学和功能异质区域内的 BET 蛋白进行精确检查，以充分了解这些蛋白在复杂成瘾行为中的作用。表观遗传阅读器蛋白以细胞类型和投射依赖性方式减弱可卡因寻找行为，目标 1 将需要 FACS 训练，以确定短期和长期接触可卡因后 BET mRNA/蛋白质表达的细胞类型特定时间过程。目标 2 需要熟练掌握病毒介导的基因转移技术，以便在可卡因寻找行为期间操纵 NAc 内特定细胞类型中的 BET 蛋白。 R00 独立阶段在此培训的基础上更具体地关注。总之，这些创新研究将首次系统地探索复杂行为期间细胞类型和连接特定方式的表观遗传目标。