Integrative single-cell spatial transcriptomic, anatomical, and functional profiling of brain-wide ensembles engaged by opioid relapse

与阿片类药物复发有关的全脑整体的综合单细胞空间转录组、解剖学和功能分析

• 批准号: 10772455
• 负责人: Sam Golden
• 金额: $ 80.15万
• 依托单位: UNIVERSITY OF WASHINGTON
• 依托单位国家: 美国
• 财政年份: 2023
• 资助国家: 美国
• 起止时间: 2023-09-30 至 2028-06-30
• 项目状态: 未结题
• 来源: https://reporter.nih.gov/project-details/10772455
• 关键词: Address; Anatomy; Atlases; Bar Codes; Behavior; Behavioral; Biological Assay; Brain; Brain Mapping; Brain region; Cells; Classification; Clinical; Communities; Coupling; Cues; Data; Data Set; Drug Exposure; Electrophysiology (science); Emotional; Euphoria; FOS gene; Female; Fentanyl; Future; Gene Expression; Gene Expression Profile; Genes; Genetic; Genetic Transcription; Goals; Heroin; Image; Immediate-Early Genes; Implant; Individual; Individual Differences; Infusion procedures; Intravenous; Machine Learning; Maps; Methodology; Methods; Modeling; Molecular; Morphine; Mus; Neurons; Neurosciences; Opioid; Oral; Oral Characters; Pharmaceutical Preparations; Phenotype; Population; Process; Relapse; Reporting; Resolution; Risk; Route; Self Administration; Slice; Specificity; Technology; Time; United States; Viral; Vulnerable Populations; Withdrawal; brain cell; brain tissue; brain volume; cell type; combat; differential expression; drug seeking behavior; fentanyl abuse; fentanyl seeking; fentanyl self-administration; fentanyl use; functional adaptation; gene conservation; in vivo; individual variation; innovation; interest; learned behavior; male; multimodal data; multimodality; neural; neural circuit; novel; open source; opioid abuse; opioid mortality; opioid use; recruit; relapse risk; response; spatial integration; synthetic opioid; technology platform; transcriptome; transcriptomic profiling; transcriptomics

PROJECT SUMMARY Opioid abuse is devastating communities across the United States and is responsible for untold suffering. The synthetic opioid fentanyl, whether due to prescription or illicit use, is involved in nearly half of reported opioid- related deaths. Unlike heroin and morphine, fentanyl is commonly administered through non-intravenous routes, but we have a limited mechanistic understanding of fentanyl use and abuse vulnerability. A leading hypothesis for the transition from use to abuse, in vulnerable individuals, is the recruitment of drug-induced gene expression changes in certain brain circuits and cells following repeated drug exposure. However, identifying such circuit- and cell-type specific populations, in combination with their underlying genetic and functional adaptations, is often limited by the resolution, throughput, and data registration of current assays. The overall goal of this project is to (i) generate machine--guided behavioral characterization of oral and intravenous fentanyl self-administration and reinstatement in male and female mice that captures individual risk vulnerability, and (ii) perform brain-wide mapping by integrating new enabling technological platforms including single-cell spatial transcriptomics (Pixel- seq), whole brain cell type specific circuit connectivity, and functional distributed brain-wide neural activity recordings (Neuropixel 2.0), recently developed by PIs in our team. To realize the potential for the molecular and functional brain mapping and data registration, we have three Specific Aims: 1) perform single-cell spatial transcriptomic profiling, with circuit-specificity, in mice with varying degrees of fentanyl-seeking behavior; 2) perform single-cell whole-brain activity mapping, with cell-type and circuit-specificity, in mice with varying degrees of fentanyl-seeking behavior; 3) perform brain-wide distributed large-scale electrophysiological recordings during oral fentanyl-seeking and integrate spatially resolved transcriptomic data with recording data. In the first Aim, Pixel-seq will be used to generate cell atlases, analyze drug- and behavior-associated spatially conserved gene expression, and map neuronal connectivity by coupling with retrograde viral tracing. In the second Aim, we will perform immediate early gene-based whole-brain activity mapping, contextualized by cell type-specific afferent connectivity, after fentanyl self-administration and reinstatement. In the third Aim, we will first perform Neuropixel2.0 electrophysiological recording of the brain-wide distributed regions of interest, and then integrate and align Neuropixel2.0 and Pixel-seq data in a new assay called NeuroPixel-seq (NP-seq). The proposed project is innovative as our integrative approaches will, for the first time, generate spatial multimodal data of unprecedented depth and resolution within the context of opioid relapse risk. It is significant because our data will provide a much-needed accessible oral fentanyl self-administration model for the neuroscience community, paired with the first fentanyl cellular-resolution atlas, allowing non-specialized labs an accessible beachhead for participating in the identification of the mechanistic basis for fentanyl use and relapse in mice.

项目摘要 阿片类药物滥用是在美国毁灭性社区，并造成了不为人知的痛苦。这 合成阿片类药物芬太尼，无论是由于处方还是非法使用，都涉及近一半的阿片类药物 相关死亡。与海洛因和吗啡不同，芬太尼通常是通过非传播路线来施用的 但是，我们对芬太尼使用和滥用脆弱性的机械理解有限。主要假设 在脆弱的个体中，从用途到滥用的过渡是募集药物诱导的基因表达 反复暴露药物后，某些脑回路和细胞的变化。但是，确定这样的电路 - 和细胞类型的特定种群，结合其潜在的遗传和功能适应 通常受当前测定的分辨率，吞吐量和数据注册的限制。该项目的总体目标 是（i）生成机器 - 口服和静脉注射芬太尼自我管理的行为表征 并恢复捕获个人风险脆弱性的雄性和雌性小鼠，（ii）进行大脑 通过整合新的启用技术平台的映射，包括单细胞空间转录组学（像素 - SEQ），全脑细胞类型特异性电路连通性和功能分布式脑部神经活动 录音（Neuropixel 2.0），最近由PIS在我们的团队中开发。实现分子和 功能性大脑映射和数据注册，我们有三个特定的目的：1）执行单细胞空间 具有不同程度的芬太尼寻求行为的小鼠中具有电路特异性的转录组分析； 2） 在具有不同的小鼠中，用细胞类型和电路特异性执行单细胞全脑活性映射 寻求芬太尼的行为程度； 3）执行大脑范围的分布式大规模电生理学 在口头寻求芬太尼期间的记录和将空间分辨的转录组数据与记录数据集成在一起。 在第一个目标中，Pixel-Seq将用于生成细胞地图集，在空间上分析药物和行为相关 保守的基因表达和通过与逆行病毒追踪耦合来绘制神经元连通性。在 第二个目的，我们将执行由细胞上下文化的立即基于基因的全脑活动映射 芬太尼自我管理和恢复后，类型特定的传入连通性。在第三个目标中，我们将 首先，对脑范围的分布区域的电生理记录进行了neuropixel2.0感兴趣的区域， 然后在称为Neuropixel-Seq（NP-Seq）的新测定中集成并对齐Neuropixel2.0和Pixel-Seq数据。这 拟议的项目具有创新性，因为我们的综合方法首次产生空间多模式 在阿片类药物复发风险的背景下，前所未有的深度和分辨率的数据。这很重要，因为我们的 数据将为神经科学提供急需的口服芬太尼自我管理模型 社区与第一个芬太尼细胞分辨率地图集配对，允许非专业实验室可访问 Beachhead参与小鼠芬太尼使用和复发的机械基础的鉴定。