Experimental approach: The [35S]GTPS binding assay was used to determine both the efficacy of cannabidiol and the ability of cannabidiol to antagonize cannabinoid receptor agonists (CP55940 and R-(+)-WIN55212) at the mouse CB1 and the human CB2 receptor.
Background and purpose: A nonpsychoactive constituent of the cannabis plant, cannabidiol has been demonstrated to have low affinity for both cannabinoid CB1 and CB2 receptors. We have shown previously that cannabidiol can enhance electrically evoked contractions of the mouse vas deferens, suggestive of inverse agonism. We have also shown that cannabidiol can antagonize cannabinoid receptor agonists in this tissue with a greater potency than we would expect from its poor affinity for cannabinoid receptors. This study aimed to investigate whether these properties of cannabidiol extend to CB1 receptors expressed in mouse brain and to human CB2 receptors that have been transfected into CHO cells.
Key results: This paper reports firstly that cannabidiol displays inverse agonism at the human CB2 receptor. Secondly, we demonstrate that cannabidiol is a high potency antagonist of cannabinoid receptor agonists in mouse brain and in membranes from CHO cells transfected with human CB2 receptors.
Conclusions and implications: This study has provided the first evidence that cannabidiol can display CB2 receptor inverse agonism, an action that appears to be responsible for its antagonism of CP55940 at the human CB2 receptor. The ability of cannabidiol to behave as a CB2 receptor inverse agonist may contribute to its documented anti-inflammatory properties.
Important recent findings with Δ 9 -THCV have been that it can induce both CB1 receptor antagonism in vivo and in vitro and signs of CB2 receptor activation in vitro at concentrations in the low nanomolar range. Further research is now required to establish whether this phytocannabinoid also behaves as a potent CB2 receptor agonist in vivo. Thus, a medicine that blocks CB1 receptors but activates CB2 receptors has potential for the management of certain disorders that include chronic liver disease and also obesity when this is associated with inflammation. The bases for the ligand and tissue dependency that Δ 9 -THCV displays as an antagonist of CB1/CB2 receptor agonists in vitro also warrant further research. In addition, in view of the structural similarity of Δ 9 -THCV to Δ 9 -THC, it will be important to determine the extent to which Δ 9 -THCV shares the ability of Δ 9 -THC, and indeed of CBD, to interact with pharmacological targets other than CB1 or CB2 receptors at concentrations in the nanomolar or low micromolar range. It will also be important to establish the extent to which CB1– and CB2-receptor-independent actions contribute to the overall in vivo pharmacology of each of these phytocannabinoids and give rise to differences between the in vivo pharmacology of Δ 9 -THC or Δ 9 -THCV and other cannabinoid receptor ligands such as CP55940, R-(+)-WIN55212 and SR141716A.
The extent to which and precise mechanisms through which the heterogeneity of the cannabinoid CB1 receptor population within the brain shapes the in vivo pharmacology of Δ 9 -THC and causes it to behave differently from agonists with higher CB1 or CB2 efficacy warrants further investigation. So too does the hypothesis that Δ 9 -THC may sometimes antagonize responses to endogenously released endocannabinoids, not least because there is evidence that such release can modulate the signs and symptoms of certain disorders and/or disease progression (reviewed in Pertwee, 2005b; Maldonado et al., 2006). Although this modulation often seems to be protective, there is evidence that it can sometimes produce harmful effects that, for example, give rise to obesity or contribute to the rewarding effects of drugs of dependence.
Why O-4394 behaves in vivo as a CB1 receptor antagonist at doses of 3 mg kg −1 i.v. or less but as a CB1 receptor agonist at doses of 10 mg kg −1 i.v. or more remains to be established. Since it does not display detectable CB1 receptor efficacy in vitro, at least in the [ 35 S]GTPγS-binding assay, one possibility is that O-4394 is metabolized in vivo to a compound that possesses significant efficacy as a cannabinoid receptor agonist and that the parent compound itself lacks such efficacy. Given the structural similarities between Δ 9 -THC and Δ 9 -THCV ( Figure 1 ), this hypothesis is supported by evidence first, that Δ 9 -THC exhibits markedly less potency in vivo as a CB1 receptor agonist than its 11-hydroxy metabolite (Lemberger et al., 1973; Wilson and May, 1975; Watanabe et al., 1990) and second, that Δ 9 -THCV can undergo metabolism to an 11-hydroxy metabolite (Brown and Harvey, 1988).
There is evidence that like established CB1 receptor antagonists such as SR141716A and AM251 (reviewed in Pertwee, 2005b), Δ 9 -THCV can block CB1-mediated effects of endogenously released endocannabinoids when administered in vivo. This evidence has come from recent experiments showing that eΔ 9 -THCV shares the ability of AM251 to reduce the food intake and body weight of non-fasted and fasted ‘non-obese’ mice when administered once (Robinson et al., 2007) and of dietary-induced obese mice when given repeatedly over 28 days (Cawthorne et al., 2007). It has also been found that like AM251, eΔ 9 -THCV can reduce the body fat content and plasma leptin concentration and increase the 24-h energy expenditure and thermic response to food of dietary-induced obese mice (Cawthorne et al., 2007), the data obtained suggesting that eΔ 9 -THCV produces its antiobesity effects more by increasing energy expenditure than by reducing food intake. In addition, both eΔ 9 -THCV and AM251 have been shown to reduce the time that ‘non-obese’ mice spend close to a food hopper (Robinson et al., 2007). These experiments were prompted by conclusive evidence that established CB1 receptor antagonists suppress feeding and body weight in animals and humans (reviewed in Matias and Di Marzo, 2007) and by the introduction into the clinic of SR141716A (rimonabant; Acomplia, Sanofi-Aventis, Paris, France) in 2006 as an antiobesity agent. Further research is now required to determine whether Δ 9 -THCV would also be effective as a medicine for the management of obesity, and indeed for drug-dependence therapy, experiments with drug-dependent animals and human subjects having shown that CB1 receptor blockade can reduce signs of drug dependence and the incidence of relapse after drug withdrawal (reviewed in Le Foll and Goldberg, 2005).
anandamide and R-(+)-methanandamide to bind to sites on muscarinic M1 and M4 receptors (Christopoulos and Wilson, 2001) and
In addition, there is the possibility that Δ 9 -THC may share actions that have so far only been shown to be exhibited by other CB1/CB2 receptor agonists (reviewed in Pertwee, 2004c, 2005a). These include the ability of
The cloning of the CB1 receptor was soon followed by the discovery that mammalian tissues can produce compounds that activate this receptor, and subsequently by the characterization of ligands such as Δ 9 -THC, (6aR)-trans-3-(1,1-dimethylheptyl)-6a,7,10,10a-tetrahydro-1-hydroxy-6,6-dimethyl-6H-dibenzo[b,d]pyran-9-methanol (HU-210), (−)-cis-3-[2-hydroxy-4-(1,1-dimethylheptyl)phenyl]-trans-4-(3-hydroxypropyl)cyclohexanol (CP55940) and (R)-(+)-[2,3-dihydro-5-methyl-3-(4-morpholinylmethyl)pyrrolo-[1,2,3-de]-1,4-benzoxazin-6-yl]-1-naphthalenylmethanone (R-(+)-WIN55212) as mixed CB1/CB2 receptor agonists and by the development of CB1– and CB2-selective agonists and antagonists (reviewed in Howlett et al., 2002; Pertwee, 2005a, 2006). It also soon became clear that CB1 receptors are located primarily in central and peripheral neurons and CB2 receptors predominantly in immune cells. CB1 receptors are also expressed by some non-neuronal cells, including immune cells, and CB2 receptors by some neurons both within and outside the brain (Skaper et al., 1996; Ross et al., 2001; Van Sickle et al., 2005; Wotherspoon et al., 2005; Beltramo et al., 2006; Gong et al., 2006). However, the role of neuronal CB2 receptors is currently unknown. The first endogenous cannabinoid receptor agonists (endocannabinoids) to be identified were N-arachidonoylethanolamine (anandamide) and 2-arachidonoylglycerol (Devane et al., 1992; Mechoulam et al., 1995; Sugiura et al., 1995), each of which can activate both CB1 and CB2 receptors and is synthesized on demand in response to elevations of intracellular calcium (Howlett et al., 2002; Di Marzo et al., 2005). Together with their receptors, these and other more recently discovered endocannabinoids (Pertwee, 2005b) constitute what is now usually referred to as the ‘endocannabinoid system’.
Δ 9 -THC and neurotransmission
Turning now to CBD, an important recent finding is that this cannabinoid displays unexpectedly high potency as a CB2 receptor antagonist and that this antagonism stems mainly from its ability to induce inverse agonism at this receptor and is, therefore, essentially non-competitive in nature. Evidence that CB2 receptor inverse agonism can ameliorate inflammation through inhibition of immune cell migration and that CBD can potently inhibit evoked immune cell migration in the Boyden chamber raises the possibility that CBD is a lead compound from which a selective and more potent CB2 receptor inverse agonist might be developed as a new class of anti-inflammatory agent. When exploring this possibility it will be important to establish the extent to which CBD modulates immune cell migration through other pharmacological mechanisms. There is also a need for further research directed at identifying the mechanisms by which CBD induces signs of inverse agonism not only in CB2-expressing cells but also in brain membranes and in the mouse isolated vas deferens.
The structure and stereochemistry of the phytocannabinoid, CBD, were first elucidated by Raphael Mechoulam in the 1960s who then went on to devise a method for its synthesis (reviewed in Pertwee, 2006). In contrast to Δ 9 -THC, CBD lacks detectable psychoactivity (reviewed in Pertwee, 2004b) and only displaces [ 3 H]CP55940 from cannabinoid CB1 and CB2 receptors at concentrations in the micromolar range ( Table 1 ). Since it displays such low affinity for these receptors, much pharmacological research with CBD has been directed at seeking out and characterizing CB1– and CB2-independent modes of action for this phytocannabinoid ( Table 3 ). Recently, however, evidence has emerged that in spite of its low affinity for CB1 and CB2 receptors, CBD can interact with these receptors at reasonably low concentrations. This has come from the discovery that CBD is capable of antagonizing cannabinoid CB1/CB2 receptor agonists with apparent KB values in the low nanomolar range both in mouse whole-brain membranes and in membranes prepared from Chinese hamster ovary (CHO) cells transfected with hCB2 receptors (Thomas et al., 2007).