Role of Gastrointestinal GCPII in Visceral Pain Signaling

胃肠道 GCPII 在内脏疼痛信号传导中的作用

• 批准号: 10678103
• 负责人: Manisha Pradhan
• 金额: $ 4.77万
• 依托单位: JOHNS HOPKINS UNIVERSITY
• 依托单位国家: 美国
• 财政年份: 2023
• 资助国家: 美国
• 起止时间: 2023-03-16 至 2026-03-15
• 项目状态: 未结题
• 来源: https://reporter.nih.gov/project-details/10678103
• 关键词: Abdomen; Abdominal Pain; Ablation; Accounting; Acute; Afferent Neurons; Analgesics; Biological Assay; Biological Markers; Biological Models; Biology; Biosensor; CRISPR/Cas technology; Cell Line; Cells; Central Nervous System; Chemoreceptors; Chronic; Clinical; Colon; Colorectal; Communication; Development; Disease; Electromyography; Endocrine; Enteric Nervous System; Enterobacteria phage P1 Cre recombinase; Enteroendocrine Cell; Enzyme-Linked Immunosorbent Assay; Enzymes; Epithelium; Evaluation; FOLH1 gene; Foundations; Future; Gastrointestinal Diseases; Gastrointestinal tract structure; Genetic; Glutamates; Hormones; Human; Hydrolysis; Hypersensitivity; Immunohistochemistry; Individual; Inflammatory; Inflammatory Bowel Diseases; Irritable Bowel Syndrome; Knockout Mice; Knowledge; Laboratories; LoxP-flanked allele; Measures; Mediating; Metalloproteases; Methods; Modeling; Modification; Mus; Musculoskeletal Pain; N-acetylaspartylglutamate; Neuroglia; Neurons; Neuropeptides; Neurotransmitters; Nociception; Pain; Pathway interactions; Patients; Peripheral Nervous System; Pharmaceutical Preparations; Phenocopy; Phenotype; Piezo 2 ion channel; Population; Publishing; Reaction; Reporter; Reporting; Research; Research Personnel; Rodent; Rodent Model; Role; Signal Transduction; Solid; Stains; Stimulus; Symptoms; Synapses; Synapsins; System; Testing; Time; Training; Visceral; Visceral pain; Western Blotting; Work; career; cell type; central pain; colorectal distension; drug development; gastrointestinal; gastrointestinal epithelium; glucagon-like peptide 1; glutamatergic signaling; gut-brain axis; ileum; in vivo; in vivo Model; inflammatory pain; inhibitor; knock-down; mouse development; mouse model; neurofilament; neurotransmission; new therapeutic target; novel; novel therapeutics; pain model; pain signal; painful neuropathy; peripheral pain; pre-clinical; promoter; protein expression; response; sensor; small molecule; therapeutic target

PROJECT SUMMARY There is an urgent need to develop non-addictive analgesic agents for the treatment of abdominal pain which is common in gastrointestinal disorders, such as irritable bowel syndrome (IBS) and inflammatory bowel disease (IBD). Glutamate carboxypeptidase II (GCPII) is a metallopeptidase that is expressed throughout the central and peripheral nervous systems (CNS, PNS respectively), where it regulates glutamate neurotransmission. Glutamate is the most abundant excitatory neurotransmitter in the body and is known to be a key player in nociception and pain signaling. In fact, small molecule GCPII inhibitors have been previously demonstrated to exert potent analgesic effects in numerous rodent models of inflammatory, neuropathic and musculoskeletal pain. While GCPII’s role in CNS and PNS pain pathways are well characterized, GCPII’s expression and function have not yet been studied in the enteric nervous system. We have recently discovered that GCPII is present throughout the ENS, with prominent expression in enteroendocrine cells (EEC) of the ileum and colon. EECs are a population of cells located throughout the gastrointestinal epithelium that are responsible for sensing luminal stimuli, secreting various hormones and neurotransmitters, and that have been reported to participate in vagal afferent signaling and bi-directional gut brain axis communication. Here, we will study the biology of GCPII in EEC-associated visceral pain signaling. Our central hypothesis is that GCPII regulates EEC glutamate secretion, altering subsequent vagal afferent activation and visceral pain signaling. In Aim #1, we will develop a glutamate biosensor assay to assess GCPII’s role as a modulator of EEC glutamate secretion, including evaluation of a small molecule GCPII inhibitor, CRISPR/Cas9 knockdown and administration of an exogenous GCPII substrate. In Aim #2, we will characterize GCPII expression changes over time in an established rodent abdominal pain model and will characterize the cellular markers of EEC subpopulations expressing GCPII. In Aim #3, we will create a novel mouse with targeted deletion of GCPII in EECs, and will objectively measure visceral pain reaction in response to aversive colorectal distension stimulation to study GCPII’s role in EEC- associated visceral pain signaling in an in vivo model system. Collectively, this research will expand our knowledge regarding GCPII expression and function in visceral pain, laying the preclinical foundation for future studies to develop novel therapeutics for abdominal pain.

项目摘要 迫切需要开发用于治疗腹痛的非成瘾性镇痛剂， 常见于胃肠道疾病，如肠易激综合征（IBS）和炎症性肠病 （IBD）。谷氨酸羧肽酶II（GCPII）是一种金属肽酶，其在整个中枢和中枢神经系统中表达， 外周神经系统（CNS，PNS分别），在那里它调节谷氨酸神经传递。 谷氨酸是体内最丰富的兴奋性神经递质，并且已知是调节神经元兴奋性的关键因素。 伤害感受和疼痛信号。事实上，小分子GCPII抑制剂先前已被证明 在许多炎症、神经病和肌肉骨骼的啮齿动物模型中发挥有效的镇痛作用 痛苦虽然GCPII在CNS和PNS疼痛通路中的作用已得到很好的表征，但GCPII的表达和功能与CNS和PNS疼痛通路中的作用密切相关。 尚未在肠神经系统中进行研究。我们最近发现GCPII存在于 在整个ENS中，在回肠和结肠的肠内分泌细胞（EEC）中显著表达。EECS 是位于整个胃肠上皮的细胞群，负责感知 腔刺激，分泌各种激素和神经递质，并已报告参与 迷走神经传入信号和双向肠脑轴通信。在这里，我们将研究GCPII的生物学 与内脏疼痛信号有关的研究。我们的中心假设是GCPII调节EEC谷氨酸 分泌，改变随后的迷走神经传入激活和内脏疼痛信号传导。在目标#1中，我们将开发一个 谷氨酸生物传感器测定，以评估GCPII作为EEC谷氨酸分泌调节剂的作用，包括 评估小分子GCPII抑制剂、CRISPR/Cas9敲低和外源性GCPII抑制剂的施用 GCPII底物。在目标#2中，我们将描述在已建立的啮齿动物中GCPII表达随时间的变化 腹痛模型，并将表征表达GCPII的EEC亚群的细胞标志物。在 目标#3，我们将创建一种在EEC中靶向缺失GCPII的新型小鼠，并将客观地测量 对厌恶性结直肠扩张刺激的内脏疼痛反应，以研究GCPII在EEC中的作用- 在体内模型系统中的相关内脏疼痛信号传导。总的来说，这项研究将扩大我们的 了解GCPII在内脏痛中的表达和功能，为未来的研究奠定临床前基础。 研究开发新的腹痛疗法。