Contributions of aberrant synaptic protein monoaminylation to opiate use disorder

异常突触蛋白单胺化对阿片类药物使用障碍的影响

• 批准号: 10681305
• 负责人: DAVID M DIETZ
• 金额: $ 69.25万
• 依托单位: ICAHN SCHOOL OF MEDICINE AT MOUNT SINAI
• 依托单位国家: 美国
• 财政年份: 2022
• 资助国家: 美国
• 起止时间: 2022-08-15 至 2027-05-31
• 项目状态: 未结题
• 来源: https://reporter.nih.gov/project-details/10681305
• 关键词: Abstinence; Acute; Affect; Amino Acids; Behavior; Behavioral; Binding; Brain; CREB1 gene; Calcium; Calmodulin; Cell Nucleus; Cells; ChIP-seq; Chemicals; Chronic; Corpus striatum structure; Coupled; Coupling; Cues; Data; Dependence; Deposition; Development; Discipline; Disease; Dopamine; Dopamine D2 Receptor; Drug Addiction; Enzymes; Event; Female; Future; Genetic Transcription; Glutamine; Goals; Health; Heroin; Histone H3; In Vitro; Individual; Laboratories; Learning; Link; Mass Spectrum Analysis; Measures; Mediating; Memory; Molecular; Molecular Abnormality; Neurobiology; Neuronal Plasticity; Neurons; Neurophysiology - biologic function; Neurosciences; Nuclear; Nuclear Translocation; Nucleus Accumbens; Open Reading Frames; Opiate Addiction; Opioid; Overdose; Pharmaceutical Preparations; Phenotype; Phosphorylation; Phosphotransferases; Play; Population; Post-Translational Protein Processing; Predisposition; Protein Isoforms; Proteins; Psychological reinforcement; Public Health; Rat Transgene; Rattus; Recombinants; Recurrent disease; Regulation; Relapse; Rewards; Rodent Model; Role; Self Administration; Serine; Signal Transduction; Site; Source; Sucrose; Synapses; Synaptic plasticity; Therapeutic Intervention; Threonine; Up-Regulation; Viral; Viral Genes; Virus; Western Blotting; addiction; amidation; behavioral pharmacology; behavioral plasticity; brain reward regions; calmodulin-dependent protein kinase II; cell type; drug abstinence; drug withdrawal; effective intervention; effective therapy; efficacious treatment; epigenomics; gene therapy; genetic manipulation; in vivo; induced pluripotent stem cell; inducible Cre; male; mutant; neurobiological mechanism; neurotransmission; novel; opioid abuse; opioid use; opioid use disorder; prolonged abstinence; psychostimulant; receptor; relapse prevention; response; therapeutically effective; transcription factor; transcriptome sequencing; transglutaminase 2

PROJECT SUMMARY Opiate use, dependence and addiction represent enduring public health issues, resulting in substantial financial and societal health burdens, as well as increasing numbers of overdoses. Drug addiction is characterized as a chronic, relapsing disease. However, to date, there remains insufficient data examining the molecular mechanisms underlying persistent opiate-induced neurobiological changes, which has led to a scarcity of effective therapies and interventions to treat and prevent relapse. Drug addiction has long been thought of as a disorder of dopamine (DA) signaling. However, therapeutic interventions targeting receptor mediated DA neurotransmission have not yet resulted in fully efficacious treatments. Therefore, an overarching goal of our laboratories – and the focus of this application – has been to investigate novel, non-canonical actions of DA involved in mediating addiction phenotypes. Our laboratory recently identified a novel signaling moiety for DA in brain, termed dopaminylation (dop), whereby DA acts as a donor source for the establishment of post-translational modifications (PTM) on substrate proteins (e.g., histone H3) via transamidation by the Transglutaminase 2 (TGM2) enzyme. In more recent efforts by our lab to unbiasedly identify additional substrates of these PTMs (focused now on synaptic proteins in nucleus accumbens/NAc, a key brain reward region), we developed a novel chemical tagging approach that, when coupled to mass spectrometry, allowed for the discovery of hundreds of dopaminylated proteins in brain, both in the context of normal neural function and in response to aberrant dopamine signaling following chronic heroin self-administration (SA) in rats. Among them, gCaMKII: 1) was found to be robustly dopaminylated at only a single amino acid residue located within its autoinhibitory helix [glutamine (Q)285], a site that exists only two amino acids away from a critical threonine (T) residue (287), which when phosphorylated directs Calmodulin (CaM) sequestration; 2) is upregulated in its dopaminylation following heroin SA, both during acute and prolonged abstinence, but not in response to natural rewards; and 3) represents a critical substrate involved in mediating long range signals from the synapse to nucleus in brain, ultimately promoting CREB activation and neuronal plasticity. Thus, this dopaminylation event on gCaMKII may represent a critical convergent mechanism linking altered dopaminergic signaling in response to heroin to CREB mediated transcriptional abnormalities. As such, we hypothesize that gCaMKIIQ285dop may play a direct role in mediating heroin relapse via aberrant modulation of CREB signaling in NAc. In Aim 1, we will fully characterize gCaMKIIQ285dop’s temporal effects on drug taking vs. relapse vulnerability in the context of heroin SA. In Aim 2, we will explore gCaMKIIQ285dop’s effects on CREB signaling/transcription following heroin SA, events that may precipitate relapse vulnerability. In Aim 3, we will investigate roles for gCaMKIIQ285dop mediated CREB signaling/transcription in D1 vs. D2 dopamine receptor-expressing MSNs during abstinence from heroin SA in the regulation of relapse behaviors.

项目摘要 阿片剂使用、依赖和成瘾是持久的公共卫生问题，造成大量财政损失。 和社会健康负担，以及越来越多的过量服用。药物成瘾的特征是 慢性复发性疾病然而，迄今为止，仍然没有足够的数据检查分子 持续性阿片诱导的神经生物学变化的潜在机制，这导致了缺乏 有效的治疗和干预措施，以治疗和预防复发。 长期以来，药物成瘾被认为是一种多巴胺（DA）信号传导障碍。然而，治疗 靶向受体介导的DA神经传递的干预尚未导致完全有效的 治疗。因此，我们实验室的首要目标--也是本申请的重点--是 研究DA参与介导成瘾表型的新的非经典作用。本实验室 最近发现了一种新的多巴胺在大脑中的信号部分，称为多巴胺化（dop），其中DA作为 用于在底物蛋白上建立翻译后修饰（PTM）的供体来源（例如， 组蛋白H3）通过转氨酶2（TGM 2）酶的转酰胺作用。在我们实验室最近的努力中， 无偏见地鉴定这些PTM的其他底物（现在集中在细胞核中的突触蛋白 在大脑中的一个关键的奖励区域，我们开发了一种新的化学标记方法， 结合质谱分析，在大脑中发现了数百种多巴胺化蛋白质，这两种蛋白质都存在于大脑中。 正常神经功能的背景和对慢性海洛因后异常多巴胺信号的反应 大鼠自身给药（SA）。其中，发现gCaMK II：1）仅在约200 μ mol/L时被稳健地多巴胺化。 位于其自身抑制螺旋内的单个氨基酸残基[谷氨酰胺（Q）285]，该位点仅存在两个 远离关键的苏氨酸（T）残基（287）的氨基酸，其在磷酸化时指导钙调蛋白 (CaM)螯合; 2）在海洛因SA后其多巴胺化上调，无论是在急性和慢性期， 长期禁欲，但不是对自然奖励的反应;和3）代表了一个关键的底物， 介导从突触到脑细胞核的长距离信号，最终促进CREB激活， 神经可塑性因此，gCaMKII上的多巴胺化事件可能代表了一种关键的收敛机制 将海洛因引起的多巴胺能信号改变与CREB介导的转录异常联系起来。作为 因此，我们假设gCaMKIIQ 285 dop可能通过异常的 调节NAc中的CREB信号传导。在目标1中，我们将充分表征gCaMKIIQ 285 dop的时间效应 关于海洛因SA背景下的吸毒与复吸脆弱性。在目标2中，我们将探索gCaMKIIQ 285 dop的 海洛因SA后CREB信号传导/转录的影响，可能会加速复发脆弱性的事件。在 目的3，我们将研究gCaMKIIQ 285 dop介导的CREB信号转导/转录在D1和D2中的作用， 海洛因戒断期间多巴胺受体表达MSNs SA在复吸行为调节中的作用