Multi-Scale Imaging Core (MSIC)

多尺度成像核心 (MSIC)

• 批准号: 10713091
• 负责人: HUI-CHEN LU
• 金额: $ 57.39万
• 依托单位: TRUSTEES OF INDIANA UNIVERSITY
• 依托单位国家: 美国
• 财政年份: 2023
• 资助国家: 美国
• 起止时间: 2023-08-15 至 2028-06-30
• 项目状态: 未结题
• 来源: https://reporter.nih.gov/project-details/10713091
• 关键词: Acute; Age; Anatomy; Antibodies; Astrocytes; Attention; Axon; Behavioral; Brain; Brain imaging; Brain region; CNR1 gene; Cannabinoids; Cannabis; Cell physiology; Cells; Chronic; Confocal Microscopy; Data; Development; Disease model; Distant; Dopamine; Drug Exposure; Drug abuse; Endocannabinoids; Energy Metabolism; Equipment; Exposure to; Functional Imaging; Glutamates; High Performance Computing; Human; Image; In Vitro; Indiana; Intake; International; Knowledge; Measurement; Medial; Metabolic; Methodology; Midwestern United States; Modeling; Molecular; Monitor; Morphology; Mus; National Institute of Drug Abuse; Neurons; Operative Surgical Procedures; Pattern; Perinatal; Pharmaceutical Preparations; Pharmacology; Phase; Physiological; Population; Prefrontal Cortex; Protocols documentation; Relapse; Research; Research Personnel; Rodent; Role; Scientist; Signal Transduction; Signaling Molecule; Signaling Protein; Slice; Somatosensory Cortex; Stains; Substance Use Disorder; THC exposure; Time; Universities; Visualization; Weaning; Withdrawal; addiction; antagonist; awake; career; cell type; compulsion; data analysis pipeline; dopamine D3 receptor; drug of abuse; drug withdrawal; endocannabinoid signaling; experience; fetal marijuana exposure; fluorescence lifetime imaging; in vivo; microscopic imaging; molecular scale; monoamine; nanoscale; neural circuit; novel strategies; postnatal; prevent; protein protein interaction; pup; sensor; single molecule; small molecule; superresolution imaging; tool; white matter

SPECIFIC AIMS-Multiscale Imaging Core (MSIC) Addictive substances trigger plasticity at the molecular, cellular and circuit levels that manifest as persistent behavioral changes that may cause substance use disorders. Targeting these changes may lead to novel strategies for preventing or treating substance use disorders. However, our knowledge of the molecular changes, the cellular processes and the abnormal circuit activity patterns that underlie various aspects of substance use disorders including compulsion, loss of intake control, withdrawal, and relapse is rather limited. To facilitate a better understanding of the molecular to circuit level plasticity accompanying drug abuse, the C3A multi- scale imaging core will support center investigators, affiliates from the Midwest and beyond, and trainees at different career stages to acquire the conceptual and technical know-how, and to access state-of-the-art equipment for nanoscale molecular measurements, for microscale anatomical analysis of subcellular and cellular profiles and for mesoscale physiological imaging of brain circuits. The C3A multi-scale imaging core will provide unprecedented imaging opportunities to examine models of substance use disorders at multiple levels, including: (1) molecular and cellular level imaging with internationally unique cell-type- and subcellular compartment-specific correlated STORM super-resolution imaging, and its recently developed PharmacoSTORM extension for nanoscale pharmacology; (2) circuit level 2P imaging to examine selective neural circuits and cell-type-specific dynamic physiological changes among large cell populations. Aim 1. Determine the cell- and subcellular compartment-specific nanoscale molecular and microscale cellular alterations triggered by chronic exposure to drugs of abuse. By employing fluorescent small molecule-based PharmacoSTORM single-molecule nanoscale pharmacology and antibody-based ImmunoSTORM super-resolution imaging, we and C3A-affiliated researchers will determine if chronic drug exposure and/or withdrawal elicit persistently altered nanoscale distribution and abundance of important signaling proteins in the cell types and brain circuits that are most relevant for substance use disorders. By correlating the nanoscale molecular measurements with microscale confocal microscopy data, we will also establish the associated morphological changes in identified subcellular compartments. Particular attention will be devoted to CB1 cannabinoid and D3 dopamine receptors that have essential roles in all phases of the addiction cycle and whose antagonists/negative allosteric modulators are among NIDA’s ten highest medication development priorities. Aim 2. Characterize the mesoscale circuit rewiring of long-range glutamatergic, dopaminergic and serotonergic axons induced by developmental or chronic exposure to drugs of abuse. Axon tracts connecting distant brain regions follow irregular trajectories, thus white matter morphology is difficult to evaluate by standard brain section staining. Therefore, we will exploit our experience in ScaleS methodology combined with optimized 2P imaging of the entire mouse brain. This approach will be used to determine the impact of developmental exposure to THC and other drugs on the integrity and trajectory of identified long-range axons. Because prenatal cannabis exposure modifies human neural circuits and rodent studies found that developing long-range glutamatergic axons are particularly sensitive to THC, we will initially determine the impact of perinatal THC exposure on glutamatergic axons originating from medial prefrontal cortex to various brain regions. Aim 3. Use in vitro and in vivo 2P sensor imaging to determine the mesoscale physiological changes in brain circuits elicited by chronic exposure to drugs of abuse. Recent advances in genetically encoded sensors for Ca2+, endocannabinoids, and monoamines provide excellent tools to visualize dynamic changes of these signaling molecules in a specific cell-type-specific manner in real-time. By combining our established and comprehensive methodology for Ca2+_imaging in acute brain slices or awake behaving mice (as young as ten days old) extending from the surgical procedure through the data analysis pipeline with High Performance Computing together with GRAB-eCB2.1 and GRABDA sensor imaging, we will support center and affiliated scientists to perform longitudinal 2P imaging to examine endocannabinoid, dopamine, and network activity changes in their relevant models of substance use disorders. We will also determine if perinatal THC exposure perturbs the development of endocannabinoid signaling in association with Ca2+-spike patterns in the primary somatosensory cortex of awake behaving mouse pups from early postnatal to weaning ages. Aim 4. Develop in vivo protocols for Fluorescence Lifetime Imaging Microscopy (FLIM) in addiction research. Drugs of abuse evoke substantial metabolic changes and perturb astrocyte-neuron interactions. We will use 2P-FLIM imaging to develop in vivo applications using FLIM-based sensors to monitor energy metabolism, signaling cascades, protein-protein interactions and to estimate the proximity between astrocytes and neurons in the substance use disorder models established by local and affiliate researchers of the imaging core.

具体目标-多尺度成像核心 (MSIC) 成瘾物质会在分子、细胞和电路层面引发可塑性，并表现为持久性 可能导致物质使用障碍的行为变化针对这些变化可能会带来新的结果。 预防或治疗物质使用障碍的策略然而，我们对分子变化的了解， 物质使用各个方面的细胞过程和异常回路活动模式 包括强迫、摄入失控、戒断和复发在内的疾病是相当有限的。 更好地理解伴随药物滥用的分子到电路水平的可塑性，C3A多 规模成像核心将为中心研究人员、中西部及其他地区的附属机构以及受训人员提供支持 不同的职业阶段，以获得概念和技术知识，并接触最先进的技术 用于纳米级分子测量、亚细胞和细胞的微观解剖分析的设备 细胞轮廓和脑回路的中尺度生理成像 C3A 多尺度成像核心。 将提供前所未有的成像机会来检查多个物质使用障碍模型 水平，包括：（1）具有国际独特的细胞类型和亚细胞的分子和细胞水平成像 隔室特异性相关 STORM 超分辨率成像及其最近开发的 PharmacoSTORM 纳米级药理学扩展；(2) 电路级 2P 成像以检查选择性 大细胞群之间的神经回路和细胞类型特异性动态生理变化。 目标 1. 确定细胞和亚细胞区室特异性的纳米级分子和微米级 长期接触滥用药物引发的细胞变化通过使用荧光小分子。 基于分子的 PharmacoSTORM 单分子纳米级药理学和基于抗体 通过ImmunSTORM超分辨率成像，我们和C3A附属研究人员将确定慢性药物是否 暴露和/或撤回引起持续的纳米级分布和丰富的重要性 与物质使用障碍最相关的细胞类型和大脑回路中的信号蛋白。 将纳米级分子测量与微米级共焦显微镜数据相关联，我们还将 确定已识别的亚细胞区室中相关的形态变化。 致力于 CB1 大麻素和 D3 多巴胺受体，它们在成瘾的各个阶段都发挥着重要作用 周期及其拮抗剂/负变构调节剂是 NIDA 排名最高的十种药物之一 发展重点。 目标 2. 表征长程谷氨酸能、多巴胺能和 由发育或长期暴露于滥用药物的轴突束诱导的血清素能轴突。 连接遥远的大脑区域遵循不规则的轨迹，因此白质形态难以评估 因此，我们将结合我们在 ScaleS 方法方面的经验。 这种方法将用于确定整个小鼠大脑的优化 2P 成像。 发育过程中接触 THC 和其他药物对已识别的长程轴突的完整性和轨迹的影响。 因为产前接触大麻会改变人类神经回路，而啮齿动物研究发现，发育中的大麻 长程谷氨酸轴突对 THC 特别敏感，我们将初步确定围产期的影响 THC 暴露在从内侧前额叶皮层到各个大脑区域的谷氨酸能轴突上。 目标 3. 使用体外和体内 2P 传感器成像来确定 长期接触滥用药物引起的大脑回路。基因编码的最新进展。 Ca2+、内源性大麻素和单胺传感器为可视化动态变化提供了出色的工具 这些信号分子以特定的细胞类型特异性的方式实时结合我们建立的和。 对急性脑切片或清醒行为小鼠（年仅 10 岁）进行 Ca2+_ 成像的综合方法 天前）从外科手术延伸到高性能的数据分析管道 与GRAB-eCB2.1和GRABDA传感器成像一起计算，我们将支持中心和附属机构 科学家进行纵向 2P 成像来检查内源性大麻素、多巴胺和网络活动 我们还将确定围产期 THC 暴露是否发生变化。 扰乱与原代细胞中 Ca2+-尖峰模式相关的内源性大麻素信号传导的发展 从出生后早期到断奶年龄的清醒行为幼鼠的体感皮层。 目标 4. 开发成瘾荧光寿命成像显微镜 (FLIM) 的体内协议 研究。滥用药物会引起显着的代谢变化并扰乱星形胶质细胞与神经元的相互作用。 将使用 2P-FLIM 成像来开发体内应用，使用基于 FLIM 的传感器来监测能量 代谢、信号级联、蛋白质-蛋白质相互作用以及估计星形胶质细胞之间的接近度 以及由当地和附属成像研究人员建立的物质使用障碍模型中的神经元 核。