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Background:

The endocannabinoid ligand anandamide (AEA) is removed from the extracellular space by a process ofcellular uptake followed by metabolism. In many cells, such as the RBL-2H3 cell line, inhibition of FAAH activity reduces theobserved uptake, indicating that the enzyme regulates uptake by controlling the intra- : extracellular AEA concentrationgradient. However, in other FAAH-expressing cells, no such effect is seen. It is not clear, however, whether these differencesare methodological in nature or due to properties of the cells themselves. In consequence, we have reinvestigated the roleof FAAH in gating the uptake of AEA.Methodology/Principal Findings: The effects of FAAH inhibition upon AEA uptake were investigated in four cell lines: AT1rat prostate cancer, RBL-2H3 rat basophilic leukaemia, rat C6 glioma and mouse P19 embryonic carcinoma cells. SemiquantitativePCR for the cells and for a rat brain lysate confirmed the expression of FAAH. No obvious expression of atranscript with the expected molecular weight of FLAT was seen. FAAH expression differed between cells, but all four couldaccumulate AEA in a manner inhibitable by the selective FAAH inhibitor URB597. However, there was a difference in thesensitivities seen in the reduction of uptake for a given degree of FAAH inhibition produced by a reversible FAAH inhibitor,with C6 cells being more sensitive than RBL-2H3 cells, despite rather similar expression levels and activities of FAAH. Thefour cell lines all expressed FABP5, and AEA uptake was reduced in the presence of the FABP5 inhibitor SB-FI-26, suggestingthat the different sensitivities to FAAH inhibition for C6 and RBL2H3 cells is not due to differences at the level of FABP-5.Conclusions/Significance: When assayed using the same methodology, different FAAH-expressing cells display differentsensitivities of uptake to FAAH inhibition.