Novel tool development for quantitative PharmacoSTORM super-resolution imaging of the nanoscale distribution of D3 dopamine receptors

用于 D3 多巴胺受体纳米级分布定量 PharmacoSTORM 超分辨率成像的新工具开发

• 批准号: 10670438
• 负责人: Istvan Katona
• 金额: $ 18.55万
• 依托单位: TRUSTEES OF INDIANA UNIVERSITY
• 依托单位国家: 美国
• 财政年份: 2022
• 资助国家: 美国
• 起止时间: 2022-08-01 至 2024-07-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/10670438
• 关键词: Acute; Adult; Adverse effects; Affect; Agonist; American; Anatomy; Animal Model; Antibodies; Antidepressive Agents; Antipsychotic Agents; Anxiety; Axon; Binding; Binding Sites; Brain; Brain Diseases; Brain imaging; Brain region; COVID-19 pandemic; Cell membrane; Cells; Chemicals; Clinical; Communication; Confocal Microscopy; Data Analyses; Development; Diazomethane; Dissociation; Dopamine Receptor; Drug usage; Electrophysiology (science); Emotional; Exhibits; FDA approved; G-Protein-Coupled Receptors; Goals; Health; Hippocampus; Human; Image; Impairment; Incubated; Islands of Calleja; Knockout Mice; Lead; Legal; Ligand Binding; Ligands; Lighting; Measurement; Mediating; Medicine; Mental Depression; Mental Health; Mental disorders; Methodology; Molecular; Mus; National Institute of Drug Abuse; Nature; Neurobiology; Neurons; Neurosciences; Nucleus Accumbens; Pathologic; Pathologic Processes; Patients; Pharmaceutical Preparations; Pharmacology; Photoaffinity Labels; Physiological; Plasma Cells; Prefrontal Cortex; Presynaptic Terminals; Prevalence; Process; Property; Protocols documentation; Relapse; Rodent; Schizophrenia; Signal Transduction; Site; Slice; Substance Use Disorder; Surveys; System; Therapeutic; Therapeutic Effect; Time; Tissues; Ultraviolet Rays; Ventral Tegmental Area; Visualization; Withdrawal Symptom; Work; addiction; analog; antagonist; cell type; clinically significant; comorbidity; cyanine dye 5; data acquisition; density; design; dopamine D3 receptor; dopaminergic neuron; drug seeking behavior; granule cell; hippocampal pyramidal neuron; imaging approach; imaging modality; in vivo; innovation; nanoscale; neurobiological mechanism; neuropsychopharmacology; novel; opioid epidemic; patch clamp; pharmacologic; prevent; receptor; receptor-mediated signaling; small molecule; social; substance use; substance use treatment; superresolution imaging; tool; tool development; trend; ultra high resolution

Abstract Substance use disorders affect ~20 million American adults according to the National Survey on Drug Use and Health. About 8.5 million patients also suffer from a comorbid mental health disorder such as depression, anxiety, and schizophrenia. Moreover, the prevalence of illicit substance use, the misuse of legal substances, and other mental health illnesses are all rapidly increasing in association with the Covid-19 pandemic. Thus, mechanistic understanding of the pathological neurobiological processes underlying mental health and substance use disorders, and their co-morbidity is imperative for the development of more efficient therapies to reduce physical, emotional, and social harm. It is widely acknowledged that dopaminergic signaling is highly impaired in these brain disorders. While the dopaminergic system has been intensely investigated in the last decades, it’s tremendous molecular and cellular complexity represents a major challenge for the detailed understanding of why certain treatments affecting the dopaminergic system are therapeutically beneficial or cause adverse effects. For example, pharmacological tools such as partial agonists, positive and negative allosteric modulators, and antagonists of the D3 dopamine receptor have been proposed by the National Institute of Drug Abuse as highly promising therapeutic tools for the treatment of substance use disorders. However, our understanding of the adaptive and maladaptive plasticity mechanisms controlled by D3 dopamine receptors and how this G protein- coupled receptor is involved in substance use disorders and other psychiatric diseases have remained very limited. This is an important, because emerging therapeutic trends, for example the expanding use of the FDA- approved D3 receptor-preferring drug cariprazine (VraylarTM) in the treatment of depression and schizophrenia and its putative usefulness in the treatment of comorbid substance use disorders illuminates the clinical significance of D3 receptors. However, the generally very low copy numbers of D3 receptors in neurons and the current lack of sensitive and specific antibodies to visualize their distribution and reorganization in association with pathological processes renders studies aiming to better understand the function of D3 dopamine receptors very difficult. Therefore, the proposed work will deliver novel fluorescent tools that will enable cell-type- and subcellular compartment-specific quantification of D3 receptor distribution at the nanoscale level in intact brain circuits affected in substance use disorders. In preliminary studies, we developed the PharmacoSTORM super- resolution nanoscale pharmacology approach and applied fluorescent cariprazine to discover high cariprazine binding density in the Islands of Calleja. In the proposed R21 CEBRA, we will complete two specific aims: Aim 1. Develop photoaffinity-based fluorescent chemical probes for high-yield PharmacoSTORM super- resolution imaging of D3 dopamine receptors. Aim 2. Establish photoaffinity labeling for PharmacoSTORM imaging of low copy numbers of D3 dopamine receptors on specific neuronal types implicated in addiction.

摘要 根据全国药物使用和药物使用调查，物质使用障碍影响到约2000万美国成年人 健康。约850万患者还患有抑郁症、焦虑症、 和精神分裂症。此外，非法物质使用的盛行、合法物质的滥用以及其他 与新冠肺炎大流行相关的心理健康疾病都在迅速增加。因此，机械论 对精神健康和物质使用的病理神经生物学过程的理解 疾病，以及它们的共同发病率对于开发更有效的治疗方法以减少身体， 对情感和社会的伤害。人们普遍认为，在这些疾病中，多巴胺能信号严重受损。 大脑紊乱。虽然多巴胺能系统在过去的几十年里得到了深入的研究，但它 巨大的分子和细胞复杂性是详细理解 为什么某些影响多巴胺能系统的治疗方法在治疗上是有益的或会产生不良影响。 例如，部分激动剂、正负变构调节剂等药理工具，以及 D3多巴胺受体拮抗剂已被国家药物滥用研究所高度推荐 治疗物质使用障碍的有前景的治疗工具。然而，我们对 D3多巴胺受体控制的适应性和非适应性可塑性机制以及这种G蛋白是如何- 偶联受体参与物质使用障碍等精神疾病的研究 有限的。这是一个重要的问题，因为正在出现的治疗趋势，例如FDA的扩大使用- 批准的D3受体偏爱药物卡里普秦(VraylarTM)治疗抑郁症和精神分裂症 它在治疗共病物质使用障碍方面的假定有效性照亮了临床 D3受体的意义。然而，神经元中D3受体的拷贝数通常很低， 目前缺乏敏感和特异的抗体来直观地显示它们的分布和相关重组 通过病理过程进行研究，旨在更好地了解D3多巴胺受体的功能 非常困难。因此，拟议的工作将提供新的荧光工具，使细胞类型-和 亚细胞室特异性定量测定正常脑中D3受体在纳米水平的分布 受物质使用障碍影响的回路。在初步研究中，我们开发了Pharmaco STORM超级 分辨纳米药理学方法和应用荧光卡里拉津发现高卡里拉津 卡列哈群岛的结合密度。在拟议的R21 CEBRA中，我们将完成两个具体目标： 目的1.开发基于光亲和力的荧光化学探针，用于高效的药物合成 D3多巴胺受体的分辨率成像。 目的2.建立低拷贝数D3多巴胺的光亲和标记法 特定神经元类型上的受体与成瘾有关。