Kaczocha Martin, Glaser Sherrye T, Deutsch Dale G
Departments of Biochemistry and Cell Biology and Neurobiology and Behavior, Stony Brook University, Stony Brook, NY 11794, USA.
Proc Natl Acad Sci U S A. 2009 Apr 14;106(15):6375-80. doi: 10.1073/pnas.0901515106. Epub 2009 Mar 23.
The endocannabinoid anandamide (arachidonoyl ethanolamide, AEA) is an uncharged neuromodulatory lipid that, similar to many neurotransmitters, is inactivated through its cellular uptake and subsequent catabolism. AEA is hydrolyzed by fatty acid amide hydrolase (FAAH), an enzyme localized on the endoplasmic reticulum. In contrast to most neuromodulators, the hydrophilic cytosol poses a diffusional barrier for the efficient delivery of AEA to its site of catabolism. Therefore, AEA likely traverses the cytosol with the assistance of an intracellular carrier that increases its solubility and rate of diffusion. To study this process, AEA uptake and hydrolysis were examined in COS-7 cells expressing FAAH restricted to the endoplasmic reticulum, mitochondria, or the Golgi apparatus. AEA hydrolysis was detectable at the earliest measurable time point (3 seconds), suggesting that COS-7 cells, normally devoid of an endocannabinoid system, possess an efficient cytosolic trafficking mechanism for AEA. Three fatty acid binding proteins (FABPs) known to be expressed in brain were examined as possible intracellular AEA carriers. AEA uptake and hydrolysis were significantly potentiated in N18TG2 neuroblastoma cells after overexpression of FABP5 or FABP7, but not FABP3. Similar results were observed in COS-7 cells stably expressing FAAH. Consistent with the roles of FABP as AEA carriers, administration of the competitive FABP ligand oleic acid or the selective non-lipid FABP inhibitor BMS309403 attenuated AEA uptake and hydrolysis by approximately 50% in N18TG2 and COS-7 cells. Taken together, FABPs represent the first proteins known to transport AEA from the plasma membrane to FAAH for inactivation and may therefore be novel pharmacological targets.
内源性大麻素花生四烯乙醇胺（arachidonoyl ethanolamide，AEA）是一种不带电荷的神经调节脂质，与许多神经递质类似，它通过细胞摄取及随后的分解代谢而失活。AEA 由内质网上的脂肪酸酰胺水解酶（FAAH）水解。与大多数神经调节剂不同，亲水性的胞质溶胶对 AEA 有效输送至其分解代谢位点构成了扩散屏障。因此，AEA 可能在一种细胞内载体的协助下穿过胞质溶胶，该载体可增加其溶解度和扩散速率。为研究此过程，在表达限于内质网、线粒体或高尔基体的 FAAH 的 COS - 7 细胞中检测了 AEA 的摄取和水解。在最早可测量的时间点（3 秒）即可检测到 AEA 水解，这表明通常缺乏内源性大麻素系统的 COS - 7 细胞拥有一种高效的 AEA 胞质运输机制。研究了已知在脑中表达的三种脂肪酸结合蛋白（FABP）作为可能的细胞内 AEA 载体。在过表达 FABP5 或 FABP7 而非 FABP3 后，N18TG2 神经母细胞瘤细胞中的 AEA 摄取和水解显著增强。在稳定表达 FAAH 的 COS - 7 细胞中也观察到了类似结果。与 FABP 作为 AEA 载体的作用一致，竞争性 FABP 配体油酸或选择性非脂质 FABP 抑制剂 BMS309403 的给药使 N18TG2 和 COS - 7 细胞中的 AEA 摄取和水解减弱了约 50%。综上所述，FABP 是已知的首批将 AEA 从质膜转运至 FAAH 进行失活的蛋白质，因此可能是新的药理学靶点。
