Giving a definition of the complex endogenous signaling system known as the “endocannabinoid system” is becoming an increasingly difficult task. In fact, the number of potential components of this system, which was originally identified from studies on the mechanism of action of the psychotropic ingredient of some varieties of cannabis, Δ9-tetrahydrocannabinol (THC), is increasing with the passing years, and the definition of “endocannabinoid” is also bound to change in the near future [1].

At the turn of the century, the endocannabinoid system was defined as the ensemble of 1) two 7-transmembrane-domain and G protein-coupled receptors (GPCRs) for THC—cannabinoid receptor type-1 (CB 1 R) and cannabinoid receptor type-2 (CB 2 R); 2) their 2 most studied endogenous ligands, the “endocannabinoids” N-arachidonoylethanolamine (anandamide) and 2-arachidonoylglycerol (2-AG); and 3) the 5 enzymes believed, at that time, to be uniquely responsible for endocannabinoid biosynthesis [i.e., N-acyl-phosphatidyl-ethanolamine-selective phospholipase D (NAPE-PLD) and diacylglycerol lipases (DAGL) α and β, for anandamide and 2-AG, respectively] and hydrolytic inactivation [i.e., fatty acid amide hydrolase (FAAH) and monoacylglycerol lipase (MAGL), for anandamide and 2-AG, respectively] (Fig. 1) [2]. This definition, however, presented a few semantic problems [1]: 1) of the > 80 cannabinoids naturally found in cannabis (with different relative composition depending on the cannabis variety), only THC and its less abundant propyl analogue, Δ9-tetrahydrocannabivarin (THCV), are capable of binding with high affinity to CB 1 R and CB 2 R (with agonist and antagonist activity for THC and THCV, respectively); hence, these 2 receptors should not be defined as “cannabinoid” receptors, but rather as THC/THCV receptors [alternatively the definition of “cannabinoid receptor” should also include those proteins that often bind to cannabinoids, such as the thermosensitive transient receptor potential (TRP) cation channels (thermo-TRPs) [3] (see below)]; 2) as a consequence, “endocannabinoids” should not be the endogenous ligands of CB 1 R and CB 2 R, but rather the ligands of all those “cannabinoid receptors” that uniquely and selectively bind to cannabinoids in general (thus, anandamide and 2-AG might not be the only endocannabinoids); and 3) again, as a consequence, “endocannabinoid enzymes” would not only be NAPE-PLD, the two DAGLs, FAAH, and MAGL, but also other enzymes responsible for the biosynthesis and inactivation of the other mediators to be eventually included in the list of the endocannabinoids.

Fig. 1 Complexity, redundancy, and promiscuity of the endocannabinoid system: the “endocannabinodome” and the interactions therewith of plant cannabinoids. Several often concurrent pathways underlie both the biosynthesis and the inactivation of the 2 most studied endocannabinoids, anandamide, and 2-arachidonoylglycerol (2-AG). Anandamide biosynthetic precursors, the N-arachidonoyl-phosphatidylethanolamines, are produced from the remodeling of phospholipids via the action of N-acyl-transferases (NATs). They are then converted to anandamide, either in 1 step, by N-acyl-phosphatidylethanolamine-selective phospholipase D (NAPE-PLD), or in sequential steps, i.e. by α,β -hydrolase-4 (ABHD4) followed by phosphodiesterase GDE1; or soluble phospholipase A2 (sPLA2) followed by lysophospholipase D (lyso-PLD); or by phospholipase C (PLC) enzymes followed by phosphatases such as PTPN22. The sn-2-arachidonate-containing diacylglycerols serving as biosynthetic precursors for 2-AG are in most cases produced from the action of PLCβ but can also come from phosphatidic acid (PA) via PA phosphohydrolase. However, 2-AG can be also produced from sn-1-lyso-phospholipids via the sequential action of phospholipase A1 (PLA1) and lyso-phospholipase C, or (not shown here) from the dephosphorylation of lysophosphatidic acid. These biosynthetic pathways are shared by anandamide and 2-AG with other N-acyl-ethanolamines and 2-mono-acyl-glycerols, respectively. These congeners, in most cases, do not activate directly the 2 cannabinoid receptors (denoted as CB 1 and CB 2 here, and as CB 1 R and CB 2 R in the main text), but have other targets, some of which shown here, such as orphan G-protein-coupled receptors (GPR55, GPR18, GPR119); the transient receptor potential of vanilloid-type 1 (TRPV1) channel; and peroxisome proliferator-activated nuclear receptors (PPARs). However, also anandamide and, to a lesser extent, 2-AG, have been suggested to be capable of activating some of these targets, particularly TRPV1 and GPR55. Anandamide also inhibits the transient receptor potential melastatin type-8 (TRPM8) channel (blue broken arrow). Both anandamide and 2-AG, following their cellular reuptake by cells, which might be facilitated by a yet-to-be-characterized endocannabinoid membrane transporter (EMT), are inactivated inside cells by enzymatic hydrolysis, respectively by fatty acid amide hydrolase (FAAH) and monoacylglycerol lipase (MAGL). In some cells FAAH, α,β-hydrolase-6 (ABHD6) and, even less frequently, α,β-hydrolase-12 (ABHD12), can also hydrolyze 2-AG. These enzymes are also responsible for the enzymatic hydrolysys of other N-acyl-ethanolamines and 2-mono-acyl-glycerols, respectively, although N-palmitoyl-ethanolamine is preferentially hydrolyzed by N-acyl-ethanolamine acid amidohydrolase (not shown). The 2 endocannabinoids, but not their non-polyunsaturated congeners, can also be oxidized by cyclooxygenase-2 (COX-2), and then processed by prostaglandin synthases, to produce prostamides, in the case of anandamide, and prostaglandin-glycerol esters, in the case of 2-AG. This latter endocannabinoid, via the action of MAGL or ABHD6, can also act as biosynthetic precursor for the nonphospholipase A2-mediated production of prostanoids. Apart from Δ9-tetrahydrocannabinol (THC), plant cannabinoids mentioned in the main text, such as cannabidiol (CBD), CBD acid (CBDA), cannabidivarin (CBDV), cannabigerol (CBG), Δ9-tetrahydrocannabivarin (THCV), THCV acid (THCVA), and CBDV acid (CBDVA), either activate (red solid arrows) or inhibit (red broken arrows) some of the receptors and enzymes of the “endocannabinoidome”. However, they often do so at medium–high micromolar concentrations, and the weight of such interactions in their pharmacology, as compared with others that they have also been suggested to exert, has not yet been fully assessed. Abn-CBD = abnormal cannabidiol; DAG = diacylglycerol; PLCβ = phospholipase β. Adapted from Di Marzo [58] Full size image

Although that depicted above would seem like the natural “evolution” of the definition of the “endocannabinoid system”, things are likely to be even more complicated. First, endocannabinoids, and also cannabinoids, have more molecular targets than just CB 1 R, CB 2 R, or thermo-TRPs, and these receptors appear to extend also to proteins that are targeted by other endogenous and exogenous substances. Furthermore, anandamide and 2-AG, like most other lipid mediators, have more than just 1 set of biosynthetic and degrading pathways and enzymes each (Fig. 1), which they often share with “endocannabinoid-like” mediators that may or may not be part of the extended definition of “endocannabinoids” provided above, that is, they may or may not interact with the same proteins to which non-THC cannabinoids bind. In some cases, these degrading pathways and enzymes lead to molecules, such as the prostamides and prostaglandin-glycerol esters (Fig. 1), which are not inactive but instead interact with other receptors, that is, these enzymes are “degrading” for endocannabinoids and “biosynthetic” for other mediators. Finally, some of these enzymes may also have additional completely different functions, for example participate in the chemical modification of molecules that have very little to do with endocannabinoid and cannabinoid targets.