GPR55 as an anandamide receptor in the vasculature

Eilidh McNaughton, Vanessa Ho. St George's University of London, London, UK

Agonists of cannabinoid receptors (CB1 and CB2) such as the endocannabinoid, anandamide (AEA) are known to cause relaxation of blood vessels, however at least part of the AEA-induced relaxation is independent of CB1 or CB2 (1). This discovery has led to the suggestion of a novel AEA receptor, with the orphan G-protein-coupled receptor GPR55 being proposed as a candidate(2). The identification of GPR55 as a cannabinoid receptor remains controversial due to conflicting reports on the pharmacological profile of GPR55 and a lack of direct evidence that AEA induces relaxation via GPR55. Here, we investigated the vascular responses to AEA and L-α-lysophosphatidylinositol (LPI), an endogenous agonist of GPR55 (3), in male GPR55 KO mice and age-matched, wild-type mice (WT, C57BL/6J) mice (14-28 weeks).

Mice were killed by cervical dislocation and arteries were isolated and mounted on a wire (mesenteric arteries) or pin (aortae) myograph to measure isometric tension. Vessels were contracted to 3µM methoxamine and 600nM U46619 prior to cumulative (1µM, 3µM 10µM, 30µM) or single (10 µM) additions of AEA or LPI to the mesenteric artery or aorta respectively. Data are presented as % relaxation of precontracted tone (number of animals≥6) and analysed by two-way, followed by a Bonferroni post-hoc test, or by an unpaired t-test where appropriate.

In mesenteric arteries of WT mice, LPI induced small relaxations at 1µM (13.9±4.6%) and 3µM (16.9±4.5%) but contractions were seen at higher concentrations (10µM: 15.5±4.7%, 30µM: 4.8±6.5%). In KO males, slightly smaller responses were obtained (1µM: 9.3±3.4%, 3µM:14.2±3.6%, 10µM:11.8±5.1%, 30µM:-1.6±7.4%; not significant). In contrast, AEA caused concentration-dependent relaxation (maximal relaxation at 30µM: 65.6±7.6%) in WT mice, which was significantly diminished (P<0.05) in the KO (at 30µM: 30.4±3.3%), providing evidence for a role of GPR55 in AEA-mediated relaxation. Interestingly, the putative GPR55 antagonist O-1918 attenuated AEA responses in WT (at 30µM: control, 68.1±9.9% vs +O-1918, 19.2±7.1%, P<0.001), but caused a significant potentiation in KO mice (at 30µM: control, 32.4±3.6% vs +O-1918, 55.3±4.8%; P<0.001). These data are consistent with O-1918 being a GPR55 antagonist, but highlight an action on signalling pathway that predominates in the absence of GPR55. On the other hand, the impact of GPR55 activation on vascular reactivity may depend on the vascular region. This is because, in WT aorta, both AEA (10µM, -13.6±14.1%) and LPI (10µM, -21.7±6.3%) induce contractions that exhibited a trend towards reduction in aorta from KO mice (AEA: 3.2±5.8%; LPI: 10.5±6.3%; not significant).

1. In conclusion, our data reveal that relaxation to AEA is mediated by GPR55, at least in mesenteric arteries of mice. Further investigations are required to clarify the pharmacological profile of the receptor and its function in different vascular regions.ReferencesWagner et al (1999) Hypertension, 33:429

2. Johns et al (2007) BJP, 152:825

3. Oka et al (2007) Biochem Biophys Res Commun 362:928