Julie Gardener
New Member
GPR55 And The Vascular Receptors For Cannabinoids
C R Hiley1* and S S Kaup11Department of Pharmacology, University of Cambridge, Cambridge, UK
*Author for correspondence: Email: crh1@cam.ac.uk
Received July 11, 2007; Accepted July 20, 2007.
This article has been cited by other articles in PMC.
Br J Pharmacol. 2007 November; 152(5): 559—561.
Published online 2007 August 20. doi: 10.1038/sj.bjp.0707421
PMCID: PMC2190021
Copyright 2007, Nature Publishing Group
Abstract
CB1 and CB2 receptors mediate most responses to cannabinoids but not some of the cardiovascular actions of endocannabinoids such as anandamide and virodhamine, or those of some synthetic agents, like abnormal cannabidiol (abn-cbd). These agents induce vasorelaxation which is antagonised by rimonabant but only at high concentrations relative to those required to block CB1 receptors. Vasorelaxation to anandamide is sensitive to Pertussis toxin (though that to abn-cbd is not), and so is thought to be mediated by a G protein-coupled receptor through Gi/o. An orphan receptor, GPR55, apparently a cannabinoid receptor, is activated by abn-cbd, but is not the receptor mediating vasorelaxation to this agent, as the response persists in vessels from GPR55 knockout mice. However, the activity of anandamide in GPR55 knockout mice is not yet reported and so the role of GPR55 as a cannabinoid receptor mediating vascular responses has yet to be finalised.
Introduction
Central and peripheral responses to cannabinoids are most often associated with actions at one or another of two cloned receptors, designated the CB1 and the CB2 receptors (Howlett et al., 2002). However, vasorelaxation in the rat mesenteric arterial system, either in response to the endocannabinoid, anandamide (White and Hiley, 1997; Wagner et al., 1999), or to the phytocannabinoid, Δ9-tetrahydrocannabinol (THC; O'Sullivan et al., 2005a), is not consistent with action at these receptors. For example, the concentrations of rimonabant, a CB1 receptor antagonist, required to block the actions of anandamide are much higher than those needed to block CB1 receptors (White and Hiley, 1998) whereas the vasorelaxation to THC is insensitive to it (O'Sullivan et al., 2005a). Other blood vessels, such as the rat coronary artery (Ford et al., 2002) and rabbit aorta (Mukhopadhyay et al., 2002), also show vasorelaxant responses to cannabinoids that are not consistent with actions at CB1 or CB2 receptors although relaxation of rat aorta to THC is sensitive to CB2, rather than CB1, receptor blockade (O'Sullivan et al., 2005b).
Rather more substantial evidence for the existence of a novel cannabinoid receptor in the cardiovascular system has come from studies on abnormal cannabidiol (abn-cbd), a synthetic analogue of another phytocannabinoid, cannabidiol. It, like anandamide, evokes mesenteric vascular relaxation but is equally effective in wild-type and cannabinoid receptor knockout mice (Járai et al., 1999; Wagner et al., 1999). The responses to abn-cbd and anandamide have many similarities (Járai et al., 1999; Wagner et al., 1999; Ho and Hiley, 2003) and have been suggested to be due to actions on an additional cannabinoid receptor, the 'endothelial anandamide receptor' (Wagner et al., 1999; Offertáler et al., 2003). Identification of activity at this receptor is greatly aided by the fact that it is sensitive to antagonism by (−)-1,3-dimethoxy-2-(3-3,4-trans-p-menthadien-(1,8)-yl)-orcinol (O-1918), an analogue of abn-cbd (Offertáler et al., 2003). Since the actions of anandamide and abn-cbd are sensitive to inhibition by Pertussis toxin (PTX; White and Hiley, 1997; Járai et al., 1999), the novel receptor is thought to be G protein-coupled and act through Gi/o.
Abn-cbd and O-1918 therefore provide tools to evaluate candidate proteins as novel vascular cannabinoid receptors. The most promising field to search is the large number of orphan G protein-coupled receptors encoded within the human genome. One of these, GPR55, has been reported in patents to be activated by cannabinoid ligands (Baker et al., 2006), although rather little is otherwise known about it other than a brief report of its distribution in human brain (Sawzdargo et al., 1999). Its position as a cannabinoid receptor has been challenged on the basis of an in silico comparison of the 'functional fingerprint' of its putative binding pocket for agonists with those of the CB1 and CB2 receptors (Petitet et al., 2006) and so the appearance, in this issue, of a report from the laboratories of GlaxoSmithKline by Johns et al. (2007) is especially welcome. This paper fills in some of the uncertainties concerning the candidacy of GPR55 as the vascular cannabinoid receptor for anandamide and abn-cbd but also leaves a number of tantalizing questions.
Johns et al. (2007) report that GPR55, when expressed in HEK293 cells, is activated in the low nanomolar concentration range by both abn-cbd and trans-4-[3-methyl-6-(1-methylethenyl)-2-cyclohexen-1-yl]-5-methyl-1,3-benzenediol (O-1602), another cannabidiol analogue, as assessed by GTPγS binding. In contrast, a patent from AstraZeneca (Drmota et al., 2004) shows abn-cbd to be 1000-fold less potent than this in a similar assay. On the other hand both reports agree that R(+)-[2,3-dihydro-5-methyl-3-[(morpholinyl)methyl]pyrrolo[1,2,3-de]-1,4-benzoxazin-yl]-(1-naphthalenyl)-methanone (WIN 55,212-2), an agonist which is relatively non-selective between CB1 and CB2 receptors, has no effect in these assays. Together, these observations support the notion that GPR55 is a receptor for cannabinoid analogues with a distinct activation profile from that of the well-characterized CB1 and CB2 receptors.
A major strength of the paper by Johns et al. (2007) is the use of GPR55 knockout mice to characterize the cardiovascular role of this receptor. Interestingly, the knockout mice show no significant difference in basal heart rate and blood pressure but this conflicts with a report that GPR55 knockout mice are hypertensive which was given by Greasley from the AstraZeneca group in an unpublished talk to the Oxford Meeting of the British Pharmacological Society in December 2006. Johns et al. (2007) found that abn-cbd lowered systemic blood pressure in both the knockout and the wild-type mice, suggesting that this response is not mediated by GPR55. In the myograph, abn-cbd relaxed mesenteric arteries from both strains of mice with similar EC50 values (wild-type, 255±88nM; GPR55 knockout, 516±176nM) and sensitivity to O-1918 (KB: wild-type, 1.8±1.3μM−1; GPR55 knockout, 2.8±2.0μM−1); the KB values compare with one of 1.9μM−1 estimated from the data in Offertáler et al. (2003).
So, both Drmota et al. (2004) and Johns et al. (2007) show clearly that GPR55 is a receptor for cannabinoid analogues, but is it the 'endothelial anandamide receptor'? The GlaxoSmithKline group have studied abn-cbd and O-1602, but not anandamide itself either in the blood vessels from GPR55 knockout mice or in the GTPγS assay in cells transfected with GPR55. Furthermore, they have not shown whether or not either the GTPγS response or the vascular responses to abn-cbd are sensitive to PTX; this is a consensual observation for the vasorelaxation evoked by anandamide in rat mesenteric arteries (White and Hiley, 1997; Járai et al., 1999). On the other hand, in functional studies of rat mesenteric artery, vasorelaxation to abn-cbd is not sensitive to PTX (Ho and Hiley, 2003) whereas its actions on p42/44 mitogen-activated protein kinase and protein kinase B/Akt, in human umbilical vein endothelial cells are sensitive to toxin pretreatment (Offertáler et al., 2003). Perhaps these responses are mediated by different receptors, both of which are sensitive to antagonism by O-1918. One of these receptors could be the 'anandamide receptor', whereas the other is the receptor mediating the abn-cbd response in the GPR55 knockout mice. Alternatively, one receptor could couple to Gi/o and to another signalling pathway to mediate the different responses. Indeed, a GlaxoSmithKline patent concerning GPR55 reports that it couples through G12 or G13 when investigated in a yeast and this would not be expected to be PTX-sensitive (Brown and Wise, 2001).
Johns et al. (2007) have also not shown if the GTPγS response in GPR55-expressing cells to abn-cbd is sensitive to the abn-cbd antagonist which would help show up further pharmacological similarity or dissimilarity between this receptor and the vascular responses. Responses to both anandamide and abn-cbd are antagonised by O-1918 but they show slightly different sensitivity; at a concentration of 1μM, O-1918 induced a shift of approximately threefold against abn-cbd (Offertáler et al., 2003) but 10-fold against anandamide (O'Sullivan et al., 2004). This might be due to experimental variation, as not many results with O-1918 are yet reported, or it could be that the antagonist acts at two receptors for cannabinoids; further information about the actions of O-1918 will be welcome.
Johns et al. (2007) have clearly shown that GPR55 does not mediate the vascular responses to abn-cbd in the mouse mesenteric artery even though it is a potent agonist at the human receptor in a recombinant system. Although previous results with this agonist have been from the rat mesenteric bed, it seems unlikely that a species difference underlies this result. Therefore, investigations of GPR55 in the vasculature await the arrival of more ligands as well as more data from the AstraZeneca group to throw light on the apparent discrepancies between their results and those of Johns et al. (2007). The most prominent of these are the possible differences in blood pressure in GPR55 knockout mice and the large discrepancy in the reported potency of abn-cbd in cells expressing GPR55.
Abbreviations
abn-cbd abnormal cannabidiol
O-1602 trans-4-[3-methyl-6-(1-methylethenyl)-2-cyclohexen-1-yl]-5-methyl-1,3-benzenediol
O-1918 (−)-1,3-dimethoxy-2-(3-3,4-trans-p-menthadien-(1,8)-yl)-orcinol
PTX Pertussis toxin
THC Δ9-tetrahydrocannabinol
WIN 55,212-2 R(+)-[2,3-dihydro-5-methyl-3-[(morpholinyl)methyl]pyrrolo[1,2,3-de]-1,4-benzoxazin-yl]-(1-naphthalenyl)-methanone
Notes
Conflict of interest
The authors state no conflict of interest.
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Source: GPR55 and the vascular receptors for cannabinoids
