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Biosynthesis and metabolism of anandamide as an endogenous ligand for cannabinoid receptors

作为大麻素受体内源配体的 anandamide 的生物合成和代谢

基本信息

批准号：
08308036
负责人：
YAMAMOTO Shozo
金额：
$ 14.66万
依托单位：
The University of Tokushima
依托单位国家：
日本
项目类别：
Grant-in-Aid for Scientific Research (A)
财政年份：
1996
资助国家：
日本
起止时间：
1996 至 1998
项目状态：
已结题

项目摘要

Specific receptors for cannabinoids were earlier found in brain (CB1) and peripheral tissues (CB2). As endogenous ligands for these receptors, arachidonoylethanolamine (anandamide) and 2-arachidonoylglycerol (2-AG) were recently identified, and they are now under active investigations. The purpose of this research project is to study on the biosynthesis and metabolism of these endocannabinoids together with the developments of their assays and receptor agonists and antagonists. The following experimental results have been reported by individual members of the research group.1) Studies on the enzymes : Anandamide amidohydrolase which hydrolyzed anandamide and lost its biological activities was found in brain and liver. The porcine brain enzyme was partially purified by S.Yamamoto et al., who demonstrated a broad substrate specificity and the reversibility of the enzyme reaction. The anandamide synthesis from arachidonic acid and ethanolamine was confirmed with a rat recombinant enzyme. It should be noted that the same enzyme hydrolyzes 2-AG.It was reported by I.Yamamoto et al. that the enzyme was inhibited by various cannabinoids. Because of a high Km for ethanolamine, the physiological role of the reverse reaction of anandamide amidohydrolase is questionable. The particifation of phospholipase D was suggested by Waku et al. and Kudo et al. to hydrolyze N-arachidonoylphosphatidy lethanolamine releasing anandamide.2) Development of anandamide assay : A sensitive HPLC method was developed for fluorescent derivative of anandamide. GC-MS analysis was also performed with anandamide.3) Studies on biological activities : Waku et al. identified 2-AG as another endogenous ligand for CB1, and studied its structure-activity relationship as CB1 ligand. Kudo et al. found the inhibitory activity of anandamide on the collagen-induced aggregation of platelets. New agonists for CB2 without effect on CB1 were developed by Miyamoto et al..

大麻素的特异性受体早期发现于大脑（CB 1）和外周组织（CB 2）。作为这些受体的内源性配体，花生四烯酰乙醇胺（anandamide）和2-花生四烯酰甘油（2-AG）最近被确定，他们现在正在积极的研究。本研究项目的目的是研究这些内源性大麻素的生物合成和代谢，以及它们的检测方法和受体激动剂和拮抗剂的发展。本课题组的研究结果如下：1）酶的研究：在脑和肝中发现了花生四烯酸酰胺水解酶，它能水解花生四烯酸酰胺而失去其生物活性。猪脑酶由S.Yamamoto等人部分纯化，其证明了广泛的底物特异性和酶反应的可逆性。花生四烯酸和乙醇胺的合成花生四烯酸和乙醇胺证实了与大鼠重组酶。应该注意的是，相同的酶水解2-AG。据I.Yamamoto等人报道，该酶被各种大麻素抑制。由于乙醇胺的高Km，花生四烯酸酰胺水解酶的逆反应的生理作用是值得怀疑的。Waku等和Kudo等提出磷脂酶D参与水解N-花生四烯酸磷脂酰乙醇胺释放花生四烯酸。2）花生四烯酸含量测定方法的建立：建立了花生四烯酸荧光衍生物的高效液相色谱分析方法。3）生物活性研究：Waku等鉴定出2-AG为CB 1的另一种内源性配体，并研究了其作为CB 1配体的构效关系。Kudo等人发现花生四烯酸对胶原诱导的血小板聚集具有抑制活性。Miyamoto等人开发了对CB 1没有影响的CB 2的新激动剂。