Study of the Cholesystokinin (CCK) Molecular Subsets in the Mouse and Macaque Dorsal Horn

小鼠和猕猴背角胆囊收缩素 (CCK) 分子亚群的研究

• 批准号: 10019342
• 负责人: Cynthia Mary Arokiaraj
• 金额: $ 4.55万
• 依托单位: UNIVERSITY OF PITTSBURGH AT PITTSBURGH
• 依托单位国家: 美国
• 财政年份: 2019
• 资助国家: 美国
• 起止时间: 2019-07-01 至 2021-06-30
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/10019342
• 关键词: Acute; Affect; Anatomy; Anti-Inflammatory Agents; Area; Basic Science; Cells; Cholecystokinin; Clinical; Data; Dependovirus; Development; Emotional; Epidemic; Funding; Future; General Population; Genetic; Human; Hypersensitivity; In Situ Hybridization; Inflammatory; Injury; Interneurons; Laboratories; Life; Ligands; Light; MAF gene; Macaca; Mechanics; Mediating; Methods; Modeling; Molecular; Molecular Profiling; Mus; Neurons; Neuropathy; Opioid; Pain; Patients; Persistent pain; Pharmaceutical Preparations; Population; Primates; Public Health; Quality of life; Reporting; Role; SLC17A8 gene; Skin; Spinal; Spinal Cord; Stimulus; Suggestion; Surface; Target Populations; Techniques; Testing; Therapeutic; Thyrotropin-Releasing Hormone; Time; Touch sensation; Translating; Viral; Virus; Work; allodynia; base; cell type; clinically relevant; copine; design; designer receptors exclusively activated by designer drugs; dorsal horn; excitatory neuron; experience; experimental study; interest; mechanical allodynia; molecular marker; molecular subtypes; neural circuit; non-opioid analgesic; nonhuman primate; novel; pain behavior; pain patient; pain sensation; persistent symptom; postnatal development; promoter; psychologic; receptor; research study; seal; side effect; single cell sequencing; social; therapeutic target; tool; transcriptome; transmission process; virtual

Project Summary: Persistent pain is a long-term condition that adversely affects psychological, physical and social aspects of patients. Current treatment methods rely heavily on opioids and anti-inflammatory drugs which can lose efficacy over time or produce deleterious side effects. Persistent pain patients often experience static or dynamic mechanical allodynia which is a painful sensation caused by innocuous stimuli that arises from punctate indentation of the skin or light brushing across the skin surface. Mechanisms that convert light touch into pain in the setting of injury occur within the spinal dorsal horn. Identifying the neural circuitry involved and developing ways to target it therapeutically are of intense interest. Our laboratory recently identified a subset of cholecystokinin (CCK) expressing excitatory interneurons that are important for conveying both static and dynamic mechanical allodynia. These neurons reside in the low threshold mechanosensory zone (lamina IIi-IV) of the dorsal horn. Their role in both forms of mechanical allodynia induced by inflammatory and neuropathic injuries was demonstrated by acute silencing of these neurons using a virally targeted Designer Receptor Exclusively Activated by Designer Drugs (DREADD). Heat hypersensitivity in the inflammatory model was also reversed. Interestingly, neurons that transiently express the vesicular glutamate transporter 3 (VGLUT3) during the first two weeks of postnatal development are a subset of the lamina III-IV CCK population and are important only for conveying dynamic allodynia. To identify more precisely which CCK subset is responsible for the different forms of persistent pain, we are utilizing a single-cell transcriptome study of mouse dorsal horn, which reported that the CCK population is composed of three molecularly distinct subsets marked by Maf, Cpne4 and Trh. In preliminary experiments, I have identified the CCK neurons that we have targeted with the DREADD as expressing either Trh or Cpne4. Since all of the Trh+ neurons were included in the targeted population, in Aim 1a, I will determine the role of Trh+ neurons in persistent pain. Since the transient VGLUT3 neurons are a subpopulation of the CCK neurons and are required only for dynamic allodynia, in Aim 1b, I will similarly determine the identity of these cells. To better understand clinically relevant persistent pain circuitry, the laboratory would like to apply the same tools used in mouse to the macaque and eventually to human. Thus, In Aim 2, I will determine in the macaque dorsal horn, the molecular identity and laminar organization of the CCK subsets as a percentage of excitatory neurons within each lamina. Conservation of molecular profiles and anatomic distribution of cell-types between the mouse and non-human primate would be suggestive of a conservation of their functional roles. The work will provide important information for the design of novel viral strategies to target cell-types in the primate dorsal horn to study pain circuits as well as develop novel, non- opioid based therapeutics for mechanical allodynia.

项目总结： 持续性疼痛是一种长期的疾病，会对患者的心理、生理和社会方面产生不利影响。 病人。目前的治疗方法严重依赖阿片类药物和抗炎药物，这些药物可能会失效 久而久之，否则会产生有害的副作用。持续性疼痛患者通常会经历静态或动态 机械性异位痛感，是由点状产生的无害刺激引起的痛感 皮肤上的凹痕或光线掠过皮肤表面。将轻触转化为疼痛的机制 损伤发生在脊髓背角内。识别所涉及的神经回路并发展 将其作为治疗靶点的方法引起了人们的强烈兴趣。我们的实验室最近发现了一个子集 CCK表达兴奋性中间神经元，对静息和静息信号的传递起重要作用 动力性机械过敏症。这些神经元位于低阈值机械感受区(板层III-IV)。 后角的部分。它们在炎症性和神经病理性两种形式的机械性痛觉异常中的作用 损伤是通过使用病毒靶向的设计者受体急性沉默这些神经元来证明的。 由特制药物独家激活(DREADD)。炎症模型中的热过敏反应也 颠倒了。有趣的是，瞬时表达囊泡谷氨酸转运体3(VGLUT3)的神经元在 出生后头两周的发育是板层III-IV CCK种群的一个子集，并且是重要的 仅用于传递动态超感痛觉。为了更准确地确定哪个CCK子集负责不同的 对于持续性疼痛的形式，我们正在利用对小鼠背角的单细胞转录组研究，该研究报告 CCK群体由Maf、Cpne4和TRH三个分子上不同的亚群组成。在……里面 初步实验，我已经确定我们用DREADD靶向的CCK神经元是 表达TRH或Cpne4。由于所有的TRH+神经元都包括在目标群体中，在AIM中 我将确定TRH+神经元在持续性疼痛中的作用。由于瞬时VGLUT3神经元是一种 CCK神经元的亚群，只有动态痛觉异常才需要，在目标1b中，我将类似地 确定这些细胞的身份。为了更好地了解临床上相关的持续性疼痛回路， 实验室希望将老鼠身上使用的相同工具应用到猕猴身上，并最终应用到人类身上。因此，在 目的2、测定猕猴背角中CCK的分子同一性和板层结构 亚组以每层神经层内兴奋性神经元的百分比表示。分子轮廓的守恒性和 小鼠和非人灵长类动物之间细胞类型的解剖分布将暗示 保护他们的职能作用。这项工作将为新型病毒的设计提供重要信息 针对灵长类动物背角细胞类型的策略，以研究痛觉回路以及开发新的、非 以阿片为基础的治疗机械性超敏的药物。