Molecular and spatial mapping of bladder nociceptors during development and matur

膀胱伤害感受器在发育和成熟过程中的分子和空间图谱

• 批准号: 8635404
• 负责人: JANET R KEAST
• 金额: $ 19.4万
• 依托单位: UNIVERSITY OF MELBOURNE
• 依托单位国家: 美国
• 财政年份: 2013
• 资助国家: 美国
• 起止时间: 2013-09-23 至 2015-08-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/8635404
• 关键词: Adult; Afferent Neurons; Analgesics; Anatomy; Antibodies; Area; Axon; Bioinformatics; Bladder; Bladder Control; Brain; Chemicals; Communication; Data; Development; Disease model; Embryo; Event; Family; Fluorescence-Activated Cell Sorting; Fluorescent Dyes; Functional disorder; Future; Gene Expression Profile; Generations; Genetic Models; Genitourinary system; Goals; Infection; Inflammation; Interstitial Cystitis; Ion Channel; Knowledge; Label; Large Intestine; Lower urinary tract; Maps; Mechanics; Methods; Micturition Reflex; Molecular; Molecular Profiling; Mus; Neurobiology; Neurons; Nociception; Nociceptors; Organ; Pain; Pelvic Pain; Pelvis; Peripheral; Physiology; Population; Property; RNA; Reflex action; Relative (related person); Reporter; Research; Research Personnel; Resolution; Rodent; Seeds; Sensory; Skeletal Muscle; Skin; Source; Spinal; Spinal Cord; Spinal Ganglia; Stimulus; Structure; TRPV1 gene; Techniques; Technology; Time; Tissue Sample; Tissues; Transcript; Viscera; Visceral; Visceral pain; afferent nerve; bone; central sensitization; chronic pain; deep sequencing; design; in vivo; member; nerve supply; neuronal cell body; novel; postnatal; prevent; public health relevance; receptor; research study; three-dimensional modeling

DESCRIPTION (provided by applicant): Pelvic pain conditions are debilitating and treatment options limited. A major issue contributing to this problem is the relative lack of understanding o factors underlying persistent visceral pain conditions - a much larger research effort exists in th area of somatic pain (e.g., pain of skin, skeletal muscle, bone origin). The goal of this nGUDMAP molecular anatomy project is to map the primary structural and molecular features of a major population of nociceptor neurons that innervate the urinary bladder of the mouse. This project is relevant to genitourinary pain, such as interstitial cystitis/painful bladder syndrome. The experiments will focus on nociceptive neurons, identified in a Trpv1 reporter mouse, to develop a deeper understanding of their structure and connections with the bladder, and their 'molecular fingerprint' (RNA profile). The study will be performed on developing and mature mice, to capture the periods of time when major changes occur in bladder control. In AIM 1, a topological map of Trpv1-positive axons within the embryonic and postnatal mouse bladder will be generated, providing a high-resolution positional map that can be superimposed onto existing GUDMAP data. In AIM 2, deep sequencing techniques will be applied to characterize the transcriptome of particular populations of these Trpv1 neurons. First, specific dorsal root ganglia (DRG) will be dissected and Trpv1- positive and -negative neurons separated using fluorescence activated cell sorting (FACS). This will allow their total RNA to be sequenced and characterized, building a molecular map of the development and maturation of these nociceptors. By applying the latest bioinformatics analyses this will (i) identify transcripts uniqe to DRG that contain visceral nociceptors, and (ii) by micro-injecting retrograde tracing methods prior to tissue sampling, characterize the transcriptome of bladder-projecting nociceptive neurons. This data will provide a rich source of knowledge to seed future hypothesis-driven studies on development and maturation of urogenital nociceptors, as well as potential application to studies on disease models.

描述（由申请人提供）：骨盆疼痛使人虚弱，治疗选择有限。造成这一问题的一个主要问题是相对缺乏对持续内脏疼痛条件的潜在因素的了解-在躯体疼痛（例如皮肤，骨骼肌，骨源性疼痛）领域存在更大的研究努力。这个nGUDMAP分子解剖项目的目标是绘制支配小鼠膀胱的主要伤害感受器神经元群的主要结构和分子特征。本项目与泌尿生殖系统疼痛相关，如间质性膀胱炎/膀胱疼痛综合征。这些实验将聚焦于在Trpv1报告小鼠中发现的伤害性神经元，以更深入地了解它们的结构和与膀胱的联系，以及它们的“分子指纹”（RNA图谱）。这项研究将在发育和成熟的小鼠身上进行，以捕捉膀胱控制发生重大变化的时间段。在AIM 1中，将生成胚胎和出生后小鼠膀胱内trpv1阳性轴突的拓扑图谱，提供可叠加到现有GUDMAP数据的高分辨率位置图谱。在AIM 2中，深度测序技术将被应用于表征这些Trpv1神经元特定群体的转录组。首先，解剖特定的背根神经节（DRG），并使用荧光活化细胞分选（FACS）分离Trpv1阳性和阴性神经元。这将允许对它们的总RNA进行测序和表征，建立这些伤害感受器发育和成熟的分子图谱。通过应用最新的生物信息学分析，这将(i)鉴定含有内脏伤害感受器的DRG特有的转录本，以及（ii）在组织采样之前通过微注射逆行追踪方法，表征膀胱突出的伤害感觉神经元的转录组。这些数据将提供丰富的知识来源，为未来关于泌尿生殖伤害感受器发育和成熟的假设驱动的研究提供种子，以及潜在的应用于疾病模型的研究。