Mapping brain-wide opioid actions by profiling neuronal activities and in vivo cellular target engagement

摘要 阿片类药物在减轻疼痛方面非常有效，但它们成瘾和过量的可能性导致越来越多的 公共卫生危机。研究人员试图开发新的阿片类化合物， 滥用，副作用较少，但这些努力一直很困难。内源性阿片系统 多种受体和配体在身体的不同部位和细胞类型中异质表达。 大量的工作已经完成，以描绘阿片受体（OR）和配体之间的关系。 然而，配体-受体结合的特异性通常取决于预定配体-受体结合位点的相对亲和力。 目标的一般来说，大脑的体内空间和细胞异质性掩盖了阿片类药物的作用，使其 单凭受体亲和力很难预测 目前，单细胞和空间转录组学正在改变我们对大脑结构的理解。 然而，在我们如何测量阿片类药物的作用并将其与空间分辨的 脑细胞图谱勒韦林新兴的CATCH和反向活动标记（IAM）技术，我们提出 在整个小鼠大脑中具有空间和单细胞分辨率的阿片类药物作用的多模式分析。使用 三种不同的阿片类药物，我们的目标是映射这些药物的神经元活动和细胞结合 并将它们与从单细胞中识别出的细胞类型进行登记， 转录组学此外，我们将测试药物在不同OR中的亲和力是否决定其在体内的作用。 细胞和神经整体参与。这个项目不仅提供了一个电路级机制， 分子药理学对全脑阿片类药物的作用，而且还为评估和开发 新的阿片类药物，例如，通过将区域和细胞类型偏好纳入结构-活性-关系， 引导优化或通过揭示靶上和脱靶位点来引导进一步细胞类型特异性体外化学 筛选和优化。

Abstract Opioids are highly effective at reducing pain, but their potential for addiction and overdose has led to a growing public health crisis. Researchers have attempted to develop new opioid compounds that are less likely to be abused and have fewer side effects, but these efforts have been difficult. The endogenous opioid system has multiple receptors and ligands heterogeneously expressed across different parts of the body and cell types. Tremendous work has been done to delineate the relationships between opioid receptors (ORs) and ligands. However, the specificity of ligand-receptor engagement often depends on relative affinities at predetermined targets. In general, in vivo spatial and cellular heterogeneity of the brain obscure opioid actions, making them hard to predict based on receptor affinity alone. Currently, single-cell and spatial transcriptomics are transforming our understanding of brain architecture. However, there is a significant gap in how we measure opioid actions and align them with the spatially resolved cellular atlas of the brain. Levering emerging CATCH and inverse activity marker (IAM) techniques, we propose multimodal profiling of opioid actions with spatial and single-cell resolution across the entire mouse brain. Using three pharmacologically diverse opioids, we aim to map neuronal activities and cellular binding of these drugs onto the entire mouse brain in an unbiased way and register them with cell types identified from single-cell transcriptomics. Furthermore, we will test whether a drug’s affinities across different ORs determine its in vivo cell and neural ensemble engagement. Not only would this project provide a circuit-level mechanism linking the molecular pharmacology to brain-wide opioid actions, but also lay out a roadmap for evaluating and developing new opioids, e.g., by incorporating regional and cell-type preference into the structure-activity-relationship for lead optimization or by revealing on- and off-target sites to guide further cell-type specific in vitro chemical screening and optimization.