Diacylglycerol kinase mediates receptor signaling by regulating lipid levels in somatosensory neurons

二酰甘油激酶通过调节体感神经元的脂质水平介导受体信号传导

• 批准号: 9328534
• 负责人: Victoria Brings Bartsch
• 金额: $ 3.63万
• 依托单位: UNIV OF NORTH CAROLINA CHAPEL HILL
• 依托单位国家: 美国
• 财政年份: 2017
• 资助国家: 美国
• 起止时间: 2017-04-15 至 2018-09-14
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/9328534
• 关键词: 2-arachidonylglycerol; Acute; Adverse effects; Affect; Afferent Neurons; Agonist; Allergens; American; Arachidonic Acids; Attenuated; Behavioral; Behavioral Assay; Biological Assay; Brain; Calcium; Cannabinoids; Cells; Coupled; DGK alpha; Data; Development; Diacylglycerol Kinase; Diglycerides; Educational process of instructing; Enzymes; Equilibrium; Esthesia; Exposure to; Fiber; Functional disorder; Future; Goals; Human; Hypersensitivity; Immune; Impairment; In Vitro; Inflammatory; Injury; Ion Channel; Knock-out; Knockout Mice; Laboratories; Lead; Lipase; Lipids; Maintenance; Maps; Mass Spectrum Analysis; Measures; Mechanics; Mediating; Mediator of activation protein; Modality; Molecular; Monitor; Monoacylglycerol Lipases; Monoglycerides; Mus; Nerve; Neurons; Neuropathy; Neurosciences Research; Pain; Pain management; Patients; Perception; Performance at work; Peripheral; Pharmacological Treatment; Phenotype; Phosphorylation; Phosphotransferases; Population; Process; Production; Protein Isoforms; Pruritus; Quality of life; Recovery; Research; Role; Sensory; Severities; Side; Signal Pathway; Signal Transduction; Signaling Molecule; Skin; Spinal Cord; Spinal Ganglia; Stimulus; TRPV1 gene; Testing; Tissues; Training; Vanilloid; Wild Type Mouse; Work; chronic itch; chronic pain; daily functioning; effective therapy; experimental study; in vivo; interest; lipoprotein lipase; novel; pain symptom; peripheral pain; receptor; receptor-mediated signaling; response; skills; skin disorder; somatosensory

Project Summary/Abstract Chronic pain and itch (pruritis) drastically impact every aspect of daily function, often hindering work performance and severely impairing quality of life. Current treatments aimed at alleviating these conditions have serious side effects and lack sufficient efficacy for long-term use. Characterizing the molecular mechanisms by which noxious stimuli are processed may reveal candidate molecules to target with novel pharmacological treatments. Pain- and itch-sensing neurons whose cell bodies lie in the dorsal root ganglia (DRG) detect algogenic (pain-causing) and pruritogenic (itch-causing) stimuli in peripheral tissues and transmit the signal to the spinal cord, then to the brain. Signaling mediators released after injury or in the context of skin disorders both activate DRG neurons, leading to the perception of pain and itch, and sensitize these neurons, causing enhanced responses upon further stimulation. The signaling molecules that contribute to this sensitization activate receptors that yield diacylglycerol (DAG) production in the DRG. DAG can be converted into monoacylglycerol (MAG). DAG and MAG affect activity of DRG neurons by directly modulating receptor activity, activating secondary messengers that alter signaling, and acting as precursors to downstream metabolites that affect pain and itch. I have discovered that diacylglycerol kinase (DGK) can phosphorylate MAG in addition to phosphorylating DAG. Altering levels of DAG, MAG, or their metabolites has been shown to cause differences in pain signaling. Therefore, I am interested in determining if DGK’s phosphorylation of DAG and MAG can alter lipid levels in peripheral sensory neurons and lead to changes in itch and pain. I am investigating the DGK isoform iota (DGKι) that I found to be highly expressed in the pain- and itch-sensing DRG neurons of mice and humans. I hypothesize that DGKι kinase activity regulates levels of signaling lipids in DRG neurons, mediating somatosensory signaling. Using a DGKι knockout mouse, I have confirmed that kinase activity on both DAG and MAG is reduced in DRG neurons. Preliminary experiments with these mice show that sensitivity to pruritogens is enhanced by the loss of DGKι. To determine how alterations in lipid levels contributes to somatosensory phenotypes, I will use mass spectrometry to compare levels of DAG, MAG, and related signaling lipids in neuronal tissue between wild type and DGKι-/- mice. I will further examine sensitivity to itch in vivo using acute exposure to pruritogens. Additionally, I will examine pain sensitivity to noxious mechanical and thermal stimuli in DGKι knockout mice. Mice will be sensitized via inflammatory and neuropathic injury, and the severity of sensitization and recovery will be measured. Furthermore, I will investigate calcium responses to algogenic and pruritogenic stimuli that activate receptors regulated by these lipid signaling molecules in the DRG. In vitro signaling experiments and in vivo behavioral assays coupled with lipidomic analyses will help me characterize the role of DGKι in somatosensation and discover the potential of this kinase as targets for pain and itch therapies.

项目摘要/摘要 慢性疼痛和瘙痒（瘙痒症）急剧影响每日功能的各个方面，通常会阻碍工作 表现和严重损害生活质量。目前旨在减轻这些条件的治疗 具有严重的副作用，缺乏足够的长期使用效率。表征分子 处理有害刺激的机制可能会揭示候选分子以新颖的方式靶向 药理治疗。细胞体的疼痛和瘙痒感应神经元位于背根神经节中 （DRG）检测外周组织中的算法（引起疼痛）和瘙痒（引起瘙痒）刺激并传播 信号向脊髓，然后是大脑。受伤后或皮肤背景下释放的信号传导介质 疾病都激活DRG神经元，导致疼痛和瘙痒感，并敏感这些神经元， 进一步刺激后会引起增强的反应。有助于此的信号分子 敏化激活受体，在DRG中产生二酰基甘油（DAG）。 DAG可以转换 进入单酰基甘油（MAG）。 DAG和MAG通过直接调节接收器来影响DRG神经元的活动 活动，激活改变信号传导的次级信使，并充当下游的前体 影响疼痛和瘙痒的代谢产物。我发现二酰基甘油激酶（DGK）可以磷酸化 除了磷酸化DAG之外。变化的DAG，MAG或其代谢物的水平已显示为 导致疼痛信号的差异。因此，我有兴趣确定DGK是否的DAG磷酸化 MAG可以改变周围感觉神经元中的脂质水平，并导致瘙痒和疼痛的变化。我是 研究我发现在疼痛和瘙痒感中高度表达的DGK同工型IOTA（DGKK） 小鼠和人类的DRG神经元。我假设DGKι激酶活性调节信号脂质的水平 在DRG神经元中，介导体感信号传导。使用DGKK淘汰鼠标，我已经确认 DRG神经元中DAG和MAG上的激酶活性均降低。这些小鼠的初步实验 表明DGKι的损失增强了对培养基的敏感性。确定脂质的变化如何 水平有助于体感表型，我将使用质谱法比较DAG的水平， MAG和野生型和DGKK与/ - 小鼠之间神经元组织中的相关信号脂质。我将进一步检查 使用急性暴露于瘙痒剂对瘙痒的敏感性。此外，我将检查对疼痛的敏感性 DGKι敲除小鼠中有害的机械和热刺激。小鼠将通过炎症和 神经性损伤以及灵敏度和恢复的严重程度将被测量。此外，我会的 研究对激活受体调节的受体的算法和培养基刺激的钙反应 DRG中的脂质信号分子。体外信号传导实验和体内行为测定法 脂肪组分析将帮助我表征DGKK在体验中的作用，并发现的潜力 这种激酶是疼痛和瘙痒疗法的靶标。