Cellular origin and function of brain 2-arachidonoylglycerol

内源性大麻素系统（ECS）是一种脂质信号系统，可调节多种生理功能，其中许多功能在体内平衡调节中很重要。这种反馈机制的核心是突触前g蛋白偶联的大麻素受体CB1 (CB1)，它也是大麻的主要精神活性成分（德尔塔9-四氢大麻酚，THC）的靶标。第二种大麻素受体CB2也在神经元中弱表达，但主要存在于外周组织和免疫细胞中。CB1和CB2激动剂2-花生四烯醇甘油（2-AG）是主要的内源性大麻素之一，由二酰基甘油脂肪酶（DAGL）产生。dagl - α和dagl - β两种异构体分别由Dagla和Daglb基因编码。第二种众所周知的内源性大麻素是花生四烯醇乙醇酰胺（AEA, anandamide）。与2-AG相比，AEA在脑内水平较低，并且AEA对CB1和CB2受体具有较低的功效和部分激动剂作用。迄今为止，所有大脑内源性大麻素信号传导的模型都是基于突触后是2-AG的主要来源的假设。然而，我们对神经元特异性Dagla敲除（KO）小鼠的初步结果表明，这种假设可能是错误的，因为我们没有发现这些条件KO小鼠中2-AG水平的任何变化-尽管Dagla表达大幅降低。然而，组成型Dagla KO动物显示大脑2-AG水平降低80%，并伴有情绪或压力相关行为增加的表型。这种表型与内源性大麻素信号调节情感行为和应激反应的概念完全一致，并参与抑郁症的病理。在大脑中，除了神经元外，星形胶质细胞和小胶质细胞也表达CB1受体并产生内源性大麻素。对于小胶质细胞，内源性大麻素的体外产量实际上是神经元的20倍。然而，这些内源性大麻素的功能及其在调节行为中的作用是完全不清楚的。因此，我们的目的是研究不同的脑细胞（神经元、星形胶质细胞和小胶质细胞）产生2-AG及其在动物行为中的作用。我们的重点将放在焦虑相关行为的调节及其对动物情感状态的影响上，这是ECS在动物和人类中的记录功能。为此，我们计划使用我们最近描述的条件Dagla KO小鼠。阐明脑2-AG的主要细胞来源将对整个大麻素研究领域具有重要意义。它将为未来的研究提供基础，超出本提案的范围，涉及生理学，细胞和分子生物学。

The endocannabinoid system (ECS) is a lipid signalling system that modulates a wide range of physiological functions, many of which are important in the context of homeostatic regulation. At the core of this feedback mechanism are presynaptic G-protein-coupled cannabinoid receptors CB1 (CB1), which are also the target of the primary psychoactive component of marijuana (delta9-tetrahydrocannabinol, THC). A second cannabinoid receptor, CB2, is also weakly expressed by neurons, but mostly present on peripheral tissues and immune cells. The full CB1 and CB2 agonist 2-arachidonoyl glycerol (2-AG), one of the main endocannabinoids, is generated by the diacylglycerol lipase (DAGL) enzymes. The two isoforms DAGL-alpha and DAGL-beta have been described, which are encoded by the Dagla and Daglb genes, respectively. The second well-known endocannabinoid is arachidonoylethanolamide (AEA, anandamide). The brain level AEA is lower compared to 2-AG, and AEA also has a lower efficacy and functions as a partial agonist at CB1 and CB2 receptors.To date, all models of endocannabinoid signalling in the brain are based on the assumption that the postsynapse is the main source of 2-AG. However, our preliminary results with neuron-specific Dagla knockout (KO) mice show that this assumption is probably wrong, because we did not find any changes in 2-AG levels in these conditional KO mice - although Dagla expression was substantial reduced. However, constitutive Dagla KO animals show an 80% reduction of brain 2-AG levels with a concomitant phenotype of increased emotional or stress-related behaviours. This phenotype is entirely consistent with the notion that endocannabinoid signalling regulates affective behaviours and stress responses, and is involved in the pathology of depression. It is further known that besides neurons, astrocytes and microglia also express CB1 receptors and produce endocannabinoids in the brain. For microglia, the in vitro production of endocannabinoids is actually about 20-fold higher compared to neurons. Yet, the function of these endocannabinoids and their role in modulating behaviour is completely unclear. Hence, we aim to investigate the production of 2-AG by different brain cells (neurons, astrocytes and microglia) and their role in animal behaviours. Our focus will be on the modulation of anxiety-related behaviours and its influence on the affective states of animals, which is a documented function of the ECS in animals and humans. For this purpose we are planning to use conditional Dagla KO mice that we have recently described. The elucidation of the main cellular source of brain 2-AG will be of fundamental importance to the entire field of cannabinoid research. It will provide a basis for future studies, outside the scope of this proposal, involving physiology, cell and molecular biology.