Characterization of epithelial-neural communication

上皮神经通讯的表征

• 批准号: 9240592
• 负责人: Kathryn Marie Albers
• 金额: $ 53.34万
• 依托单位: UNIVERSITY OF PITTSBURGH AT PITTSBURGH
• 依托单位国家: 美国
• 财政年份: 2016
• 资助国家: 美国
• 起止时间: 2016-04-01 至 2021-03-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/9240592
• 关键词: Action Potentials; Activation Analysis; Affect; Afferent Neurons; Avil; Behavior; Behavioral; C Fiber; Cells; Characteristics; Coculture Techniques; Code; Communication; Complex; Cutaneous; Electrophysiology (science); Epidermis; Epithelial; Epithelial Cells; Exhibits; Fiber; Freund' s Adjuvant; Ganglia; Generations; Genetic Models; Genetic Transcription; Glutamates; Growth Factor; Halorhodopsins; Heterogeneity; Hyperalgesia; Hypersensitivity; Inflammation; Inflammatory; Keratin; Knowledge; Lasers; Light; Lighting; Measures; Mechanics; Mediating; Modeling; Mus; Nature; Nerve; Nerve Fibers; Nervous system structure; Neurons; Nociceptors; Outcome; Pain; Pathologic; Pattern; Peripheral; Peripheral Nerves; Pharmacology; Phenocopy; Phenotype; Physiological; Plasticizers; Preparation; Process; Production; Property; Proteins; Reverse Transcriptase Polymerase Chain Reaction; Role; Sensory; Skin; Spinal Cord; Spinal Ganglia; Stimulus; Temperature; Testing; Touch sensation; Transgenic Mice; afferent nerve; behavior measurement; behavioral response; cell type; cutaneous sensory neurons; experimental study; genetic approach; inflammatory pain; insight; keratinocyte; mouse model; optogenetics; public health relevance; relating to nervous system; response; sensory stimulus

 DESCRIPTION (provided by applicant): The transduction of cutaneous stimuli has been previously thought to be solely a function of sensory fibers. It is now recognized that production of growth factors and neuroactivators (e.g., NGF, ATP, ACh, glutamate) by epidermal keratinocytes can have a profound effect on this process. To unravel these complex interactions and advance our understanding of the mechanisms regulating neural-keratinocyte communication, we developed optogenetic mouse models in which light activated channelrhodopsin (ChR2) is targeted to cutaneous sensory neurons. Light stimulation of the skin of these mice was found to elicit a robust nocifensive behavioral response. Electrophysiological analysis of this activation using a skin-nerve-ganglia and spinal cord ex vivo preparation showed preferential activation of C-fiber nociceptors. Thus, blue-laser light penetrates the epidermis and activates ChR2 at levels that depolarize peripheral nerve terminals. Interestingly, light activation of some neurons did not elicit response properties identical to those obtained using direct mechanical or thermal stimulation of the skin. We hypothesized this lack of a full response reflected a missing stimulus from the skin. We therefore isolated mice in which ChR2 was targeted exclusively to K14 keratin expressing keratinocytes. Remarkably, light stimulation of keratinocytes expressing ChR2 evoked changes in behavioral and electrophysiologic response properties of cutaneous sensory neurons. We also found that different subtypes of cutaneous afferents are activated at different levels suggesting heterogeneity in skin-neural communication. Using these new genetic models we propose three specific aims to advance these findings: Aim 1 experiments will examine how light-induced release of neuroactivators (e.g., ATP) from ChR2- expressing keratinocytes activates subtypes of primary sensory afferents. Aim 2 will determine how light activation of ChR2 or halorhodopsin expressed by subtypes of sensory afferents or keratinocytes affects afferent response properties. We will also determine how this activation compares to mechanical and/or thermal stimulation of the skin. Aim 3 experiments will determine the contribution of changes in keratinocytes and sensory neurons to thermal and mechanical hyperalgesia in a model of inflammatory pain. These studies will determine if hyperalgesia is caused by changes in primary afferents, skin keratinocytes or both. The ability to control activation of either keratinocytes or sensory afferents will provide new insights into how the skin and sensory nervous system communicate under normal and inflamed conditions.

 描述（由适用提供）：以前认为皮肤刺激的翻译仅是感觉纤维的函数。现在已经认识到，表皮角质形成细胞的生长因子和神经活化剂（例如NGF，ATP，ACH，谷氨酸）的产生可能会对这一过程产生深远的影响。为了揭示这些复杂的相互作用，并提高了我们对应对神经 - 甲状腺细胞通信的机制的理解，我们开发了光遗传小鼠模型，其中光激活的通道旋转蛋白（CHR2）针对皮肤感觉神经元。发现这些小鼠皮肤的光刺激引起了强大的单化行为反应。使用皮肤 - 毛线 - 脊髓和脊髓的离体制剂对这种激活进行电生理分析，显示了C纤维伤害感受器的首选激活。这是蓝色激光穿透表皮，并在去极化周围神经末端的水平上激活CHR2。有趣的是，某些神经元的光激活并未引起与使用皮肤直接机械或热刺激获得的反应特性。我们假设缺乏完全反应反映了皮肤缺失的刺激。因此，我们分离了CHR2仅靶向K14角蛋白表达角质形成细胞的小鼠。值得注意的是，表达CHR2的角质形成细胞的光刺激引起了皮肤感觉神经元行为和电生理反应特性的变化。我们还发现，不同的皮肤传入的亚型在不同的水平上激活，表明皮肤神经通信中的异质性。使用这些新的遗传模型，我们提出了三个特定的目的来推进这些发现：AIM 1实验将研究光诱导的神经激活因子（例如ATP）如何从ChR2表达角质形成细胞中激活原发性感官传递物的亚型。 AIM 2将确定由感觉传入或角质形成细胞的亚型表达的ChR2或卤代紫红蛋白的光激活如何影响传入的反应特性。我们还将确定这种激活与皮肤的机械和/或热刺激相比。 AIM 3实验将在炎症性疼痛模型中确定角质形成细胞和感觉神经元对热和机械性痛觉过敏的变化的贡献。这些研究将确定痛觉过敏是由主要传入，皮肤角质形成细胞或两者的变化引起的。控制角质形成细胞或感觉传入的激活的能力将为皮肤如何提供新的见解 感觉神经系统在正常和发炎的条件下进行沟通。