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cbd faah

Cbd faah

Specific inhibitors of the FABPs were developed at Stony Brook such as SBFI26 that led to an increase in AEA levels in the brains of animals and had physiological effects. As shown in Figure ​ Figure2, 2 , inhibiting the FABPs will reduce the AEA delivery to FAAH and disrupt the outward/inward concentration gradient driven by FAAH. Intriguingly, the truxillic acid structure of SBFI26 is the core structure of (−)-incarvillateine, the active component from a Chinese herb used for rheumatism (Berger et al., 2012). It was found that some of the inhibitors (such as OMDM1, OMDM2, VDM11, AM1172, AM404) of the “putative” transmembrane transporter, inhibit FABPs, perhaps explaining, in part, their mechanism of action (Kaczocha et al., 2012b).

At first we mistakenly reported an enzymatic activity independent of the fatty acid amide hydrolase (FAAH) and calcium for the synthesis of AEA (Deutsch and Chin, 1993), but then followed up with collaborators to help elucidate the correct pathways. This misstep was caused by the condensation of ethanolamine with phenylmetylsulfony fluoride, whose product ran the same as AEA on thin layer chromatography (Bill Devane, personal communication circa 1994). The first demonstration of AEA synthesis via a calcium dependent N-acyl phosphatidylethanolamine-specific phospholipase D (NAPE-PLD) mechanism was reported in 1994 (Di Marzo et al., 1994) although this activity had been characterized with other phosphatidylethanolamines (Schmid et al., 1983). This enzyme was purified and cloned (Ueda et al., 2005) and subsequent papers using null mice confirmed that it was mainly responsible for the synthesis of AEA (Tsuboi et al., 2011; Leishman et al., 2016) although other minor pathways may be involved under certain conditions (Liu et al., 2008; Simon and Cravatt, 2010) depending upon the mouse construct (Leishman et al., 2016). FAAH can also mediate the reverse reaction for the synthesis of AEA and this has been implicated physiologically in liver regeneration (Devane and Axelrod, 1994; Arreaza et al., 1997; Izzo and Deutsch, 2011; Mukhopadhyay et al., 2011).

The Hydrolysis of Anandamide to Arachidonic Acid and Ethanolamine by FAAH.

The future

The author declares that the research was conducted in the absence of any commercial or financial relationships that could be construed as a potential conflict of interest.

Recently, again using computational analysis and ligand displacement assays, we showed that human FABP3, 5, and 7 bind THC and CBD and function as intracellular carriers (Elmes et al., 2015). Furthermore, we demonstrated that THC and CBD inhibit the cellular uptake and catabolism of AEA by targeting FABPs. This competition for FABPs may in part or wholly explain the increased circulating levels of AEA reported after consumption of cannabidiol (Leweke et al., 2012). These data may explain, in part, the action of CBD in modulating the endocannabinoid tone in vivo and its reported efficacy toward epilepsy and other neurological disorders (Ibeas Bih et al., 2015). The role of the FABP as carriers for endocannabinoids and particularly AEA will undoubtedly grow as illustrated in the liver where FABP1 also serves as an AEA carrier (Schroeder et al., 2016).

The author confirms being the sole contributor of this work and approved it for publication.

Anandamide synthesis

In 1993 we were the first to show, with rather rudimentary experiments, that AEA was actively taken up in neuroblastoma and glioma cells (Deutsch and Chin, 1993). In 1994 the uptake of AEA was confirmed and the mechanism was postulated to involve an ATP independent active membrane transporter (Di Marzo et al., 1994). The hypothesis of an AEA transmembrane transporter became dogma for many years and the “hunt” still goes on for this “putative” anandamide membrane transporter (AMT) also called the “putative endocannabinoid membrane transporter (EMT, Ligresti et al., 2010; Nicolussi et al., 2014; Nicolussi and Gertsch, 2015). Many of the AMT (EMT) proposals have fallen by the wayside. For example, a paper first showed uptake was FAAH independent and then a decade later it was proposed that a FAAH fragment called FLAT (FAAH-like anandamide transporter) was the transmembrane transporter (Fegley et al., 2004; Fu et al., 2012), the latter being questioned (Leung et al., 2013; Björklund et al., 2014; Fowler, 2014). The evidence for a transmembrane transporter was based on enzyme saturation kinetics in cell culture, uptake studies in cells and the physiological effects of “membrane transporter inhibitors.” Many dozens of such inhibitors were proposed. However, it was shown that the kinetics of uptake of AEA can show saturation owing to the passage of hydrophobic AEA through the water layer surrounding the cell and that many of these transport inhibitors were in fact FAAH inhibitors or FAAH substrates or bound to receptors confounding the mechanism of their physiological effects (Glaser et al., 2003; Alexander and Cravatt, 2006; Bojesen and Hansen, 2006; Nicolussi and Gertsch, 2015). Furthermore, it was demonstrated that AEA can freely pass through an artificial membrane without the aid of any protein (Figure ​ (Figure2, 2 , Bojesen and Hansen, 2005; Di Pasquale et al., 2009; Kaczocha et al., 2012a; Fowler, 2013, 2015). A transmembrane protein transporter has not been identified to date and the effects of these inhibitors appear to occur downstream and many of the so-called transporter inhibitors were in fact FAAH or FABP inhibitors.

In 1993 an enzyme we called anandamide amidase, now named called FAAH, was shown to break AEA down to arachidonic acid and ethanolamine (Figure ​ (Figure1) 1 ) in the membrane fractions of most rat tissues except in leg and heart muscle (Deutsch and Chin, 1993). This activity was reported in liver microsomes for fatty acid amides, other than anandamide (Bachur and Udenfriend, 1966; Schmid et al., 1985). This lack of breakdown activity in muscle was fortuitous for the success of the vas deferens assay that was employed in the discovery of AEA in 1992 (Devane et al., 1992; Pertwee et al., 1995). In our original assay we used thin layer chromatography with AEA radio-labeled in the arachidonate portion of the molecule, but later ethanolamine labeled AEA simplified the assay procedure by permitting measurement of radiolabel without a thin layer chromatography step (Omeir et al., 1995). Cloning of the enzyme permitted more detailed molecular studies including ones that showed uniquely two serine residues in the active site (Omeir et al., 1999; Patricelli et al., 1999) and that FAAH was localized to the endoplasmic reticulum (Cravatt et al., 1996). FAAH is the main player in AEA inactivation although other pathways have been implicated in the metabolism of AEA as well (van der Stelt et al., 2002; Rahman et al., 2014).

Cbd faah

1 1 Endocannabinoid Research Group, Istituto per la Chimica di Molecole di Interesse Biologico, Consiglio Nazionale delle Ricerche, Via Campi Flegrei 34, Comprensorio Olivetti, Fabbricato 70, 80078 Pozzuoli (Napoli), Italy

Aniello Schiano Moriello

Effect of cannabidiol (CBD) and some of its analogues on the hydrolysis of [ 14 C]-anandamide (AEA) by N18TG2 cell membranes. The effect is expressed as per cent of [ 14 C]-AEA hydrolysed in the absence of any substance, and data are means±s.e.mean of n=4 experiments. •=(+)-CBD; ○=CBD; ▴=(−)-7-hydroxy-CBD.

Affinity of CBD analogues for cannabinoid CB1 and CB2 receptors

1 1 Endocannabinoid Research Group, Istituto per la Chimica di Molecole di Interesse Biologico, Consiglio Nazionale delle Ricerche, Via Campi Flegrei 34, Comprensorio Olivetti, Fabbricato 70, 80078 Pozzuoli (Napoli), Italy