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

A facilitated transport process that removes the endogenous cannabinoid ana ndamide from extracellular spaces has been identified. Once transported int o the cytoplasm, fatty acid amide hydrolase (FAAH) is responsible for metab olizing the accumulated anandamide. We propose that FAAH contributes to ana ndamide uptake by creating and maintaining an inward concentration gradient for anandamide. To explore the role of FAAH in anandamide transport, we ex amined anandamide metabolism and uptake in RBL-2H3 cells, which natively ex press FAAH, as well as wild-type Hela cells that lack FAAH. RBL-2H3 and FAA H-transfected Hela cells demonstrated a robust ability to metabolize ananda mide compared with vector-transfected HeLa cells. This activity was reduced to that observed in wild-type HeLa cells upon the addition of the FAAH inh ibitor 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 cell s showed that only FAAH inhibitors structurally similar to anandamide decre ased anandamide uptake. Because there is no detectable FAAH activity in wil d-type HeLa cells, these FAAH inhibitors are probably blocking uptake via a ctions on a plasma membrane transport protein. Phenylmethylsulfonyl fluorid e, a FAAH inhibitor that is structurally unrelated to anandamide, inhibited anandamide uptake in RBL-2H3 cells and FAAH-transfected Hela cells, but no t in wild-type HeLa cells. Furthermore, expression of FAAH in HeLa cells in creased maximal anandamide transport e-fold compared with wild-type HeLa ce lls. These results suggest that FAAH facilitates anandamide uptake but is n ot solely required for transport to occur.