Membrane localization of N-acylphosphatidylethanolamine in central neurons: Studies with exogenous phospholipases
Membrane localization of N-acylphosphatidylethanolamine in central neurons: Studies with exogenous phospholipases
复制标题
DOI：
10.1016/0929-7855(96)00510-x
复制
发表时间：
1996-09-01
期刊：
影响因子：
--
通讯作者：
Piomelli, D
中科院分区：
文献类型：
--
作者：
Cadas, H;Schinelli, S;Piomelli, D
我们研究了n -酰基磷脂酰乙醇胺（NAPE），一种假定的大麻素前体，在大鼠脑纹状体和皮质神经元的原代培养中的定位。我们用各种外源性磷脂酶（包括S. chromofuscus磷脂酶D (PLD)）探测完整的神经元。S. chromofuscus PLD不会渗透到神经元中（这表明i -125标记的PLD缺乏内化），也不会对神经元膜造成严重损伤（这表明PLD对[H-3] γ -氨基丁酸释放缺乏影响）。当神经元用[H-3]乙醇胺标记为同位素平衡时，与S. chromofuscus PLD孵育10分钟，大约50%的神经元NAPE被水解。这种水解伴随着n -酰基乙醇胺（NAE）家族的释放（通过高效液相色谱评估），其中包括大麻素受体激动剂，anandamide。外源性磷脂酶A(2) (PLA(2))（蜜蜂）和PLC（蜡样蜜蜂）分别动员了[H-3]花生四烯酸酯和[H-3]二酰基甘油，但在这些条件下对NAE的形成没有影响。这些实验表明，大约50%的神经元NAPE位于细胞外PLD（可能是质膜）容易接近的隔室中，在那里它也很容易在刺激下水解产生NAE。
We studied the localization of N-acyl phosphatidylethanolamine (NAPE), a putative cannabinoid precursor, in primary cultures of striatal and cortical neurons from the rat brain. We probed intact neurons with various exogenous phospholipases, including S. chromofuscus phospholipase D (PLD). S. chromofuscus PLD does not penetrate into neurons (as demonstrated by a lack of internalization of I-125-labeled PLD), and does not cause gross damage to the neuronal membrane (as demonstrated by a lack of effect of PLD on [H-3]gamma-aminobutyric acid release). When neurons, labeled to isotopic equilibrium with [H-3]ethanolamine, were incubated for 10 min with S. chromofuscus PLD, approximately 50% of neuronal NAPE was hydrolysed. This hydrolysis was accompanied by the release of a family of N-acyl ethanolamines (NAE) (assessed by high performance liquid chromatography), which included the cannabinoid receptor agonist, anandamide. Exogenous phospholipase A(2) (PLA(2)) (Apis mellifera) and PLC (B. cereus) mobilized [H-3]arachidonate and [H-3]diacylglycerol, respectively, but had no effect on NAE formation under these conditions. These experiments indicate that approximate to 50% of neuronal NAPE is localized in a compartment that is easily accessible to extracellular PLD, possibly the plasmalemma, where it would also be easily hydrolyzed upon stimulation to produce NAE.