Role of fatty acid amide hydrolase in the transport of the endogenous cannabinoid anandamide.

A facilitated transport process that removes the endogenous cannabinoid anandamide from extracellular spaces has been identified. Once transported into the cytoplasm, fatty acid amide hydrolase (FAAH) is responsible for metabolizing the accumulated anandamide. We propose that FAAH contributes to anandamide uptake by creating and maintaining an inward concentration gradient for anandamide. To explore the role of FAAH in anandamide transport, we examined anandamide metabolism and uptake in RBL-2H3 cells, which natively express FAAH, as well as wild-type HeLa cells that lack FAAH. RBL-2H3 and FAAH-transfected HeLa cells demonstrated a robust ability to metabolize anandamide compared with vector-transfected HeLa cells. This activity was reduced to that observed in wild-type HeLa cells upon the addition of the FAAH inhibitor methyl arachidonyl fluorophosphonate. Anandamide uptake was reduced in a dose-dependent manner by various FAAH inhibitors in both RBL-2H3 cells and wild-type HeLa cells. Anandamide uptake studies in wild-type HeLa cells showed that only FAAH inhibitors structurally similar to anandamide decreased anandamide uptake. Because there is no detectable FAAH activity in wild-type HeLa cells, these FAAH inhibitors are probably blocking uptake via actions on a plasma membrane transport protein. Phenylmethylsulfonyl fluoride, a FAAH inhibitor that is structurally unrelated to anandamide, inhibited anandamide uptake in RBL-2H3 cells and FAAH-transfected HeLa cells, but not in wild-type HeLa cells. Furthermore, expression of FAAH in HeLa cells increased maximal anandamide transport 2-fold compared with wild-type HeLa cells. These results suggest that FAAH facilitates anandamide uptake but is not solely required for transport to occur.