MODULATION OF TRIGEMINAL INHIBITION

三叉神经抑制的调节

• 批准号: 6613150
• 负责人: LI-YEN M HUANG
• 金额: $ 13.57万
• 依托单位: UNIVERSITY OF TEXAS MED BR GALVESTON
• 依托单位国家: 美国
• 财政年份: 2003
• 资助国家: 美国
• 起止时间: 2003-04-01 至 2005-03-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/6613150
• 关键词: biological signal transduction; calcium indicator; calcium ion; fluorescence resonance energy transfer; glycine receptors; laboratory rat; membrane proteins; molecular biology; neural inhibition; neural transmission; oral facial pain; protein protein interaction; protein quantitation /detection; technology /technique development; trigeminal nerve; voltage /patch clamp; yeast two hybrid system

DESCRIPTION (provided by applicant): Inhibitory synaptic transmission in nociceptive neurons is largely mediated by glycine receptors (GlyRs). It has been shown recently that intracellular Ca2+ causes a large potentiation GlyR-mediated responses. There is good evidence suggesting that the potentiation is mediated by a diffusible cytoplasmic protein (CytoP). The goal of this project is to develop a new technology to identify the CytoP and study its interactions with GlyRs in trigeminal neurons. We hypothesize that a diffusible CytoP binds to GlyRs at low intracellular concentrations ([Ca2+]i), keeping GlyRs in a low activity state. When [Ca2+]i rises, the protein is rapidly dissociated from GlyRs, resulting in an enhancement of channel activity. To test this hypothesis, we will combine molecular biology, patch-clamp and imaging techniques to determine these interactions in molecular details. The CytoP will be isolated by the two-hybrid method. The interaction of the CytoP with GlyRs will be determine by simultaneously monitor (i) GlyR-mediated currents using the patch clamp technique, (2) intracellular Ca2+ transients using fluorescent Ca2+indicators and (iii) dynamic interactions between GlyRs and cytoplasmic proteins using the frequency resonance energy transfer (FRET) imaging technique. We will then extend this technology to identify CytoP and study the GlyR-CytoP interaction in trigeminal neurons. A better understanding of the mechanism of GlyR modulation will provide important information about the neuronal signaling and may lead to new therapy for orofacial pain.

描述（由申请人提供）：伤害性神经元中的抑制性突触传递主要由甘氨酸受体（GlyR）介导。最近的研究表明，细胞内 Ca2+ 会显着增强 GlyR 介导的反应。有充分的证据表明增强作用是由可扩散细胞质蛋白（CytoP）介导的。该项目的目标是开发一种新技术来识别 CytoP 并研究其与三叉神经元中 GlyR 的相互作用。我们假设可扩散的 CytoP 在低细胞内浓度 ([Ca2+]i) 下与 GlyR 结合，使 GlyR 保持在低活性状态。当 [Ca2+]i 升高时，蛋白质迅速从 GlyR 解离，导致通道活性增强。为了检验这一假设，我们将结合分子生物学、膜片钳和成像技术来确定分子细节中的这些相互作用。 CytoP将通过双杂交方法分离。 CytoP 与 GlyR 的相互作用将通过同时监测（i）使用膜片钳技术的 GlyR 介导的电流，（2）使用荧光 Ca2+ 指示剂的细胞内 Ca2+ 瞬变和（iii）使用频率共振能量转移（FRET）成像技术的 GlyR 和细胞质蛋白之间的动态相互作用来确定。然后我们将扩展这项技术来识别 CytoP 并研究三叉神经元中 GlyR-CytoP 的相互作用。更好地了解 GlyR 调节机制将提供有关神经元信号传导的重要信息，并可能带来口面部疼痛的新疗法。