Differential control of 2-AG’s activity at CB1R by ABHD6 and MAGL

ABHD6 和 MAGL 对 CB1R 上 2-AG 活性的差异控制

基本信息

批准号：
10664172
负责人：
Nephi Stella
金额：
$ 19.44万
依托单位：
UNIVERSITY OF WASHINGTON
依托单位国家：
美国
项目类别：
财政年份：
2023
资助国家：
美国
起止时间：
2023-04-01 至 2025-03-31
项目状态：
未结题

来源：
https://reporter.nih.gov/project-details/10664172
关键词：
2-arachidonylglycerol ABHD6 gene Address Adverse effects Affect Agonist Applications Grants BDKRB2 gene Biological Models Biosensor Bradykinin Brain CNR1 gene Cannabinoids Cell Line Cells Chronic Corpus striatum structure Development Drug Addiction Electrophysiology (science)Endocannabinoids Enzymes Fluorescence Microscopy Goals Hydrolase MAGL inhibitor Maps Measures Mediating Molecular Monoacylglycerol Lipases Mus N-Methylaspartate Neurons Presynaptic Terminals Production Property Receptor Signaling Research Role Signal Transduction Slice Stimulus Synaptic Transmission System Testing Therapeutic Time Transfection Veratridine Work abuse liability cell type endocannabinoid signaling lipoprotein lipase nervous system disorder novel novel therapeutics pharmacologic postsynaptic presynaptic receptor sensor side effect small molecule spatiotemporal tool two photon microscopy

项目摘要

Summary The most abundant endocannabinoid (eCB) in brain, 2-arachidonoyl glycerol (2-AG), is inactivated by two enzymes: monoacylglycerol lipase (MAGL) and α/β hydrolase domain-contain 6 (ABHD6) that differ in their hydrolyzing activities and subcellular localization (presynaptic and postsynaptic, respectively). Accordingly, selective inhibition of each enzyme results in different spatiotemporal enhancement of 2-AG-CB1R signaling in the brain, and potentially synergistic therapeutic benefits. We recently gathered results showing that stimulated increase in 2-AG production is reliably measured using GRABeCB2.0, a recently developed 2-AG sensor. Remarkably, metabotropic receptor mediated increase in 2-AG is controlled by ABHD6, whereas ionotropic receptor-dependent increase in 2-AG is not controlled by ABHD6. These results raise the question of how MAGL controls stimulation-dependent 2-AG production? Demonstrating that receptor-dependent increases in 2-AG production and activity at CB1R signaling are differentially controlled by ABHD6 and MAGL would provide an additional level of mechanistic distinction between these eCB-hydrolyzing enzymes. To increase our understanding of the respective roles of ABHD6 and MAGL in controlling 2-AG-CB1R signaling in brain, we initiated an effort and have now successfully validated the GRABeCB2.0 pharmacological profile in neural cells in culture and identified several stimuli that increase 2-AG production. Based on this premise, we propose to address the following two questions in mouse neurons in primary culture and striatal slices using live-cell fluorescence microscopy, two-photon microscopy and electrophysiology: Aim 1: Which stimuli increase 2-AG production and GRABeCB2.0 signal in neurons? Aim 2: How do ABHD6 and MAGL differentially control stimuli-dependent increases in 2-AG-CB1R signaling in neurons? Completion of the work outlined in this new R21 grant proposal will provide a comprehensive understanding of the respective role of ABHD6 and MAGL in controlling 2-AG-CB1R signaling in the brain. Our long-term goal is to increase our understanding of the molecular, cellular, and system’s level differences by which ABHD6 and MAGL control eCB signaling in brain, a body of work that will help develop novel therapeutics with reduced potential for abuse and adverse effects produced by classic cannabinoid agonists.

概括大脑中最丰富的内源性大麻素（ECB），2-芳烃甘油（2-AG），被两个人灭活酶：单酰基甘油脂肪酶（MAGL）和α/β水解酶结构域6（ABHD6），它们的不同水解活性和亚细胞定位（分别为突触前和突触后）。选择性抑制每种酶会导致2-AG-CB1R信号在不同的时空增强大脑以及潜在的协同治疗益处。我们最近收集的结果表明，使用刺激的2ag产生增加是可靠的 Grabecb2.0，最近开发的2AG传感器。值得注意的是，代谢受体介导的2-AG的增加由ABHD6控制，而Ionotropic接收器依赖于2-AG的增加不受ABHD6的控制。这些结果提出了一个问题，即MAGL如何控制依赖模拟的2AG产生？证明接收器依赖性在2AG的产生和CB1R信号的活性中增加是由ABHD6和MAGL差异控制将提供额外的机械区别在这些欧洲央行 - 水解酶之间。为了增加我们对ABHD6各自角色的理解 MAGL在控制大脑中的2-AG-CB1R信号传导时，我们开始了一项努力，现在已经成功验证了培养神经细胞中的Grabecb2.0药理学特征，并确定了增加2 AG的几种刺激生产。基于此前提，我们建议在鼠标神经元中解决以下两个问题使用活细胞荧光显微镜，两光子显微镜和电生理学： AIM 1：哪些刺激会增加神经元中的2AG产生和grabecb2.0信号？ AIM 2：ABHD6和MAGL如何差异控制刺激依赖性依赖于2-AG-CB1R 神经元的信号传导？这项新的R21赠款提案中概述的工作的完成将提供全面的理解 ABHD6和MAGL在控制大脑中2-AG-CB1R信号传导中的相对作用。我们的长期目标是为了增加我们对ABHD6和ABHD6和系统水平差异的理解 MAGL控制欧洲央行信号在大脑中，这是一种有助于发展新疗法的工作体系经典大麻素激动剂产生的滥用和不良反应的潜力。