Blink, Lacrimation, and Nociception: Precision Mapping and Integrated Atlas Generation of Corneal Trigeminal Afferents

眨眼、流泪和伤害感受：角膜三叉神经传入的精确绘图和集成图谱生成

• 批准号: 10585769
• 负责人: Sina Farsiu
• 金额: $ 134.25万
• 依托单位: DUKE UNIVERSITY
• 依托单位国家: 美国
• 财政年份: 2022
• 资助国家: 美国
• 起止时间: 2022-09-30 至 2027-08-31
• 项目状态: 未结题
• 来源: https://reporter.nih.gov/project-details/10585769
• 关键词: 3-Dimensional; Ablation; Address; Anatomy; Atlases; Axon; Behavioral; Biological Assay; Blinking; Brain Stem; Catalogs; Categories; Cells; Classification; Clinical Research; Color; Complex; Computer software; Cornea; Coupled; Data; Data Set; Development; Dissection; Dry Eye Syndromes; Electrophysiology (science); Eye; Foundations; Future; Generations; Genes; Genetic; Genomics; Grant; Homeostasis; Human; Image; Image Analysis; Immunofluorescence Immunologic; Infection; Ions; Knowledge; Label; Lacrimation; Lead; Link; Machine Learning; Maintenance; Maps; Mediating; Methods; Microscopy; Modeling; Molecular; Morphology; Mouse Strains; Mus; Nature; Nerve; Nerve Fibers; Nerve Plexus; Neurons; Neuropathy; Nociception; Nociceptors; Output; Pain; Pattern; Pharmacology; Play; Population; Process; Production; Property; Pseudomonas aeruginosa; Quality of life; Reflex action; Resolution; Role; Signal Transduction; Structure; Structure of trigeminal ganglion; Tissues; Translational Research; Trigeminal System; Validation; Work; analog; automated segmentation; base; blink reflexes; density; differential expression; experimental study; genetic approach; grasp; human subject; imaging Segmentation; macrophage; multidisciplinary; multiple datasets; neuronal cell body; neuronal circuitry; new therapeutic target; novel strategies; ocular pain; ocular surface; open source; patch clamp; reflectance confocal microscopy; response; segmentation algorithm; transcriptome; transcriptome sequencing

ABSTRACT Corneal nerves that mediate pain, blink reflexes and tear production are indispensable in the proper maintenance of ocular surface homeostasis. However, the complexity of these neurons, both at the axon level in the cornea, and cell bodies in the trigeminal ganglion, has made it increasingly difficult to grasp their full nature, resulting in key knowledge gaps in the field. Consistent with the call for the current U01, we will address this barrier via comprehensive analyses of corneal nerves at the morphologic, molecular, and functional level. We have assembled a multidisciplinary team with complementary expertise, enabling integrated analyses of spatial, electrophysiologic, genomic and behavioral profiles in mice; and AI-assisted structure-function studies in human subjects, to open new inroads in the field. In Aim 1, we will anatomically and functionally map corneal-projecting trigeminal afferents in mice. Genetic approaches will be applied to rigorously profile the spatial arrangement of nociceptors in the cornea and trigeminal ganglion using seven- color immunolabeling. This spatial information will be directly linked with the electrophysiological profiling of these respective populations. In Aim 2, genomic analyses of corneal-projecting trigeminal afferents will be conducted applying a new platform for spatial RNA-seq at cellular resolution. We will use mouse strains to parse out the transcriptomes and behavioral outputs at the genetic level. Moreover, we apply two novel approaches to selectively target corneal nerves involved in tear production versus blinking. In Aim 3, we will discover morphological patterns of corneal nerves that predict blinking, lacrimation, and nociception in humans. We will image corneal nerves with in vivo confocal microscopy of subjects with differential blink, tear, and nociceptive behavioral outputs, thereby capturing functional analogs of the mouse experiments in Aims 1 and 2. With the AI-based auto-segmentation algorithm that we are developing, we will be able to apply machine learning for multidimensional profiling of nerve patterns, and then compare these with respective behavioral outputs. These AI-guided efforts will provide critical clues for understanding corneal afferent structure-function in humans. In summary, our collective studies will lead to an unprecedented cartography of corneal afferents in blink, lacrimation, and nociception. The advancements from this work will be poised to facilitate a deeper understanding of related pathobiology including neuropathic ocular pain and dry eye disease that will lay the foundation for future translational and clinical research. All genomic, imaging and electrophysiologic datasets produced will be made publicly available, and all software products for corneal nerve image segmentation will be made freely available online as open-source and easy-to-use software packages.

摘要 角膜神经介导疼痛，眨眼反射和眼泪的产生是必不可少的适当 维持眼表稳态。然而，这些神经元的复杂性，无论是在轴突水平， 角膜和三叉神经节中的细胞体，使得越来越难以完全掌握它们的功能。 自然，导致该领域的关键知识差距。与当前U 01的要求一致，我们将 通过对角膜神经的形态学、分子学和生物学方面的综合分析， 功能层面。我们组建了一支具有互补专业知识的多学科团队， 对小鼠的空间、电生理、基因组和行为特征进行综合分析; 在人类受试者的结构-功能研究，在该领域开辟新的进展。在目标1中，我们将在解剖学上 并在小鼠中功能性地绘制角膜投射三叉神经传入。遗传学方法将应用于 使用七种方法，严格描绘角膜和三叉神经节中伤害感受器的空间排列， 彩色免疫标记。这种空间信息将直接与电生理分析相关联， 这些不同的人口。在目标2中，将对角膜投射三叉神经传入神经进行基因组分析。 在细胞分辨率上应用空间RNA-seq的新平台。我们将使用小鼠品系， 在基因水平上解析出转录组和行为输出。此外，我们还应用了两种新颖的 选择性靶向涉及泪液产生与眨眼的角膜神经的方法。在目标3中，我们 发现预测人类眨眼、流泪和伤害感受的角膜神经形态模式。 我们将在活体共焦显微镜下对有不同眨眼、流泪和 伤害感受行为输出，从而捕获目的1和2中小鼠实验的功能类似物。 2.通过我们正在开发的基于AI的自动分割算法，我们将能够应用机器 学习神经模式的多维分析，然后将这些与相应的行为 产出这些人工智能引导的努力将为理解角膜传入结构-功能提供关键线索 在人类身上。总之，我们的集体研究将导致角膜传入神经的前所未有的绘图， 眨眼流泪和伤害感受这项工作的进展将有助于促进更深入的 了解相关的病理生物学，包括神经性眼痛和干眼病，将奠定 为未来的转化和临床研究奠定基础。所有基因组、成像和电生理数据集 所有用于角膜神经图像分割的软件产品都将 作为开放源码和易于使用的软件包在网上免费提供。