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.

特定目标-多尺度成像核心（MSIC） 成瘾物质在分子、细胞和回路水平上触发可塑性，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导致新的 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， 构成物质使用各方面基础的细胞过程和异常回路活动模式 包括强迫症、摄入控制丧失、戒断和复发在内的疾病相当有限。以促进 更好地了解分子电路水平的可塑性伴随药物滥用，C3 A多- 规模成像核心将支持中心的研究人员，来自中西部和其他地区的分支机构，以及 不同的职业阶段，以获得概念和技术知识，并获得最先进的 用于纳米级分子测量的设备，用于亚细胞和 细胞轮廓和脑回路的中尺度生理成像。C3 A多尺度成像核心 将提供前所未有的成像机会，以检查多种物质使用障碍的模型， 水平，包括：（1）分子和细胞水平成像与国际独特的细胞类型和亚细胞 隔室特定相关STORM超分辨率成像，以及其最近开发的 用于纳米级药理学的PharmacoSTORM扩展;（2）电路级2 P成像，以检查选择性 神经回路和大细胞群中细胞类型特异性的动态生理变化。 目标1.确定细胞和亚细胞区室特异性纳米级分子和微米级 长期暴露于滥用药物引发的细胞改变。通过采用荧光小 基于分子的PharmacoSTORM单分子纳米级药理学和基于抗体的 ImmunoSTORM超分辨率成像，我们和C3 A附属研究人员将确定慢性药物 暴露和/或撤回引起持续改变的纳米级分布和丰富的重要 细胞类型和大脑回路中与物质使用障碍最相关的信号蛋白。通过 将纳米级分子测量与微尺度共聚焦显微镜数据相关联，我们还将 在鉴定的亚细胞区室中建立相关的形态学变化。将特别注意 致力于CB 1大麻素和D3多巴胺受体，这些受体在成瘾的各个阶段都起着重要作用 其拮抗剂/负变构调节剂是NIDA十大药物之一 发展优先事项。 目标2.表征长程多巴胺能、多巴胺能和 发育或慢性暴露于滥用药物诱导的轴突。轴突束 连接遥远的脑区遵循不规则的轨迹，因此白色物质形态难以评估 通过标准的脑切片染色。因此，我们将利用我们在ScaleS方法学中的经验， 对整个小鼠大脑进行优化的2 P成像。这种方法将用于确定 发育暴露于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的神经回路，啮齿动物的研究发现， 长距离神经元轴突对THC特别敏感，我们将初步确定围产期的影响。 THC暴露对源自内侧前额叶皮层至不同脑区的多巴胺能轴突的影响。 目标3。在体外和体内使用2 P传感器成像来确定中尺度生理变化， 长期暴露在滥用药物的环境中所引发的脑回路基因编码的最新进展 Ca 2+，内源性大麻素和单胺的传感器提供了很好的工具来可视化动态变化， 这些信号分子以特定的细胞类型特定的方式实时地。通过结合我们的既定和 在急性脑切片或清醒行为小鼠（10岁以下）中的Ca 2 +_2成像的综合方法学 天），从外科手术延伸到数据分析管道， 结合GRAB-eCB 2.1和GRABDA传感器成像计算，我们将支持中心和附属机构 科学家进行纵向2 P成像，以检查内源性大麻素，多巴胺和网络活动 物质使用障碍相关模型的变化。我们还将确定围产期THC暴露是否 干扰内源性大麻素信号传导的发展，与原发性高血压中的Ca 2+峰模式有关。 从出生后早期到断奶年龄的清醒行为小鼠幼仔的躯体感觉皮层。 目标4。开发成瘾中荧光寿命成像显微镜（FLIM）的体内方案 research.滥用药物引起大量的代谢变化，扰乱星形胶质细胞-神经元的相互作用。我们 将使用2 P-FLIM成像技术开发使用基于FLIM的传感器监测能量的体内应用 代谢，信号级联，蛋白质-蛋白质相互作用，并估计星形胶质细胞之间的接近度 和神经元在物质使用障碍模型建立的当地和附属研究人员的成像 核心