Accumulation of N-arachidonoylethanolamine (anandamide) into cerebellar granule cells occurs via facilitated diffusion
Accumulation of N-arachidonoylethanolamine (anandamide) into cerebellar granule cells occurs via facilitated diffusion
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DOI：
10.1046/j.1471-4159.1997.69020631.x
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发表时间：
1997-08-01
影响因子：
4.7
通讯作者：
Campbell, WB
中科院分区：
文献类型：
--
作者：
Hillard, CJ;Edgemond, WS;Campbell, WB
N-花生四烯酸乙醇胺（anandamide，AEA）是大麻素受体的一种假定的内源性配体。原代培养的完整小脑颗粒神经元快速积累AEA，小脑颗粒细胞的[H-3]AEA积累依赖于孵育时间（t（1/2）为2.6 +/- 0.8 min，37 ℃）和温度，AEA的蓄积是饱和的，表观Km为41 +/- 15 μ M，V-max为0.61 +/- 0.04 nmol/min/10（6）个细胞。200 μ M根皮素（对照组的51.4 ± 4%）以非竞争性方式显著降低小脑颗粒细胞的[H-3]AEA积累。哇巴因或去除细胞外钠均不抑制[H-3]AEA积累。[H-3]AEA的积累是相当选择性的AEA在其他天然存在的N-酰基乙醇胺，只有N-油酰乙醇胺显着抑制[H-3]AEA的积累在10 μ M的浓度。棕榈酸和亚麻酸的乙醇酰胺在10 μ M时无活性。N-花生四烯酰苄胺和N-花生四烯酰丙胺，但不花生四烯酸，15-羟基-AEA，或12-羟基-AEA，AEA的积累竞争。当细胞预加载[H-3]AEA时，发生温度依赖性流出，半衰期为1.9 +/- 1.0 min，根皮素不抑制[H-3]AEA从细胞流出。这些结果表明，AEA是由小脑颗粒细胞积累的蛋白质介导的运输过程，具有易化扩散的特点。
N-Arachidonoylethanolamine (anandamide, AEA) is a putative endogenous ligand of the cannabinoid receptor. Intact cerebellar granule neurons in primary culture rapidly accumulate AEA, [H-3]AEA accumulation by cerebellar granule cells is dependent on incubation time (t(1/2) of 2.6 +/- 0.8 min at 37 degrees C) and temperature, The accumulation of AEA is saturable and has an apparent K-m of 41 +/- 15 mu M and a V-max of 0.61 +/- 0.04 nmol/min/10(6) cells. [H-3]AEA accumulation by cerebellar granule cells is significantly reduced by 200 mu M phloretin (51.4 +/- 4% of control) in a noncompetitive manner, [H-3]AEA accumulation is not inhibited by either ouabain or removal of extracellular sodium. [H-3]AEA accumulation is fairly selective for AEA among other naturally occurring N-acylethanolamines; only N-oleoylethanolamine significantly inhibited [H-3]AEA accumulation at a concentration of 10 mu M. The ethanolamides of palmitic acid and linolenic acid were inactive at 10 mu M. N-Arachidonoylbenzylamine and N-arachidonoylpropylamine, but not arachidonic acid, 15-hydroxy-AEA, or 12-hydroxy-AEA, compete for AEA accumulation. When cells are preloaded with [H-3]AEA, temperature-dependent efflux occurs with a half-life of 1.9 +/- 1.0 min, Phloretin does not inhibit [H-3]AEA efflux from cells. These results suggest that AEA is accumulated by cerebellar granule cells by a protein-mediated transport process that has the characteristics of facilitated diffusion.