Cannabinoid-induced mesenteric vasodilation through an endothelial site distinct from CB1 or CB2 receptors

Abstract

Cannabinoids, including the endogenous ligand arachidonyl ethanolamide (anandamide), elicit not only neurobehavioral but also cardiovascular effects. Two cannabinoid receptors, CB1 and CB2, have been cloned, and studies with the selective CB1 receptor antagonist SR141716A have implicated peripherally located CB1 receptors in the hypotensive action of cannabinoids. In rat mesenteric arteries, anandamide-induced vasodilation is inhibited by SR141716A, but other potent CB1 receptor agonists, such as HU-210, do not cause vasodilation, which implicates an as-yet-unidentified receptor in this effect. Here we show that "abnormal cannabidiol" (Abn-cbd) is a neurobehaviorally inactive cannabinoid that does not bind to CB1 receptors, yet causes SR141716A-sensitive hypotension and mesenteric vasodilation in wild-type mice and in mice lacking CB1 receptors or both CB1 and CB2 receptors. Hypotension by Abn-cbd is also inhibited by cannabidiol (20 microgram/g), which does not influence anandamide- or HU-210-induced hypotension. In the rat mesenteric arterial bed, Abn-cbd-induced vasodilation is unaffected by blockade of endothelial NO synthase, cyclooxygenase, or capsaicin receptors, but it is abolished by endothelial denudation. Mesenteric vasodilation by Abn-cbd, but not by acetylcholine, sodium nitroprusside, or capsaicine, is blocked by SR141716A (1 microM) or by cannabidiol (10 microM). Abn-cbd-induced vasodilation is also blocked in the presence of charybdotoxin (100 nM) plus apamin (100 nM), a combination of K(+)-channel toxins reported to block the release of an endothelium-derived hyperpolarizing factor (EDHF). These findings suggest that Abn-cbd and cannabidiol are a selective agonist and antagonist, respectively, of an as-yet-unidentified endothelial receptor for anandamide, activation of which elicits NO-independent mesenteric vasodilation, possibly by means of the release of EDHF.

Figure 1

Effects of HU-210 (10 ng/g; A), anandamide (4 μg/g; B), and Abn-cbd (20 μg/g; C) in the absence (●) and presence of 3 μg/g SR141716A (○) on mean arterial blood pressure in pentobarbital-anesthetized CB1^+/+ (Left) and CB1^−/− (Right) mice. Mice were injected with a bolus i.v. dose of the indicated agonist at 0 min, 10 min after the injection of vehicle or SR141716A. Points and bars represent means ± SE from 4–7 animals. * indicate significant difference (P < 0.05) from corresponding baseline (0) values.

Figure 2

Mesenteric vasodilator effect of anandamide (A) and abnormal cannabidiol (B) in the absence (●) or presence of SR141716A (1 μM ▵ or 5 μM ○) in CB1^+/+ (Left) and CB1^−/− mice (Right). Agonists were injected as an intraarterial bolus, whereas SR141716A was included in the perfusion buffer. Points and vertical bars represent means ± SE from 3–6 experiments. Each preparation was used for one agonist injection only. *, P < 0.05; **, P < 0.005 from corresponding value in the absence of SR141716A.

Figure 3

Vasodilator action of Abn-cbd and its antagonism by cannabidiol (CBD) in the rat isolated mesenteric arterial bed. Each panel represents a separate preparation, illustrating the long lasting vasodilation by 4 mg of Abn-cbd (A), and its reversal (B) or prevention (C) by 10 μM CBD. Phenylephrine (PE, 15 μM) used to precontract the preparation and CBD were included in the perfusion buffer, as indicated by the horizontal lines, and the bolus injection of 4 mg of Abn-cbd is indicated by the arrows. These experiments were replicated in 4–6 additional preparations with similar results. (D) Structures of CBD and Abn-cbd.

Figure 4

Anandamide-induced vasodilation (●) is inhibited in the presence 10 μM cannabidiol (○) in rat mesenteric arteries. Points and bars represent means ± SE from 4 or 5 experiments. Asterisks indicate statistically significant difference from corresponding control values: (*, P < 0.05; **, P < 0.005).

Figure 5

Analysis of the mechanism of the vasodilator action of Abn-cbd in the rat isolated mesenteric arterial bed preparation. Columns and bars represent the means ± SE for the vasodilator response to 4 mg of Abn-cbd under the following conditions: control (open column); endothelium-denuded preparations (Denud); endothelium-intact preparations perfused with 1 μM SR141716A (SR); 5 μM capsazepine (CPZ); 100 μM N^G-nitro-l-arginine methyl ester plus 10 μM indomethacin (L-NAME + INDO); 100 nM apamin (Ap); 100 nM charybdotoxin (CBX); 100 nM apamin plus 100 nM charybdotoxin (Ap + CBX); or 100 nM apamin plus 50 nM iberiotoxin (Ap + IBX). Numbers of preparations tested are in parentheses. For explanations, see text. **, P < 0.01; ***, P < 0.005.

Figure 6

Capsaicine-induced mesenteric vasodilation is competitively inhibited by capsazepine, but not by SR141716A. Vasodilator responses to intraarterial bolus injections of capsaicine were tested in the rat isolated mesenteric vascular bed under control conditions (●), in the presence of 5 μM capsazepine (■) or in the presence of 1 μM SR141716A (○). Points and vertical bars represent means ± SE from 3–5 experiments.