Role of Pre‐Junctional CB₁, But not CB₂, TRPV1 or GPR55 Receptors in Anandamide‐Induced Inhibition of the Vasodepressor Sensory CGRPergic Outflow in Pithed Rats

Abstract

Stimulation of the perivascular sensory outflow in pithed rats produces vasodepressor responses mediated by CGRP release. Interestingly, endocannabinoids such as anandamide (which interacts with CB 1 , CB 2 , TRPV1 and GPR55 receptors) can regulate the activity of perivascular sensory nerves in dural blood vessels by modulating CGRP release. Yet, as no publication has reported whether this mechanism is operative in the healthy systemic vasculature, this study has specifically analysed the receptors mediating the potential inhibitory effects of the cannabinoid (CB) receptor agonists anandamide (non-selective), JWH-015 (CB 2 ) and lysophosphatidylinositol (GPR55) on the rat vasodepressor sensory CGRPergic outflow (an index of systemic vasodilatation). Healthy pithed rats were pre-treated with consecutive i.v. continuous infusions of hexamethonium, methoxamine and the above agonists. Electrical spinal (T 9 -T 12 ) stimulation of the vasodepressor sensory CGRPergic outflow or i.v. injections of a-CGRP produced frequency-dependent or dose-dependent vasodepressor responses. The infusions of anandamide in a dosedependent manner inhibited the vasodepressor responses by electrical stimulation (remaining unaffected by JWH-015 or lysophosphatidylinositol), but not those by a-CGRP. After i.v. administration of antagonists, the inhibition by 3.1 lg/kg min anandamide was: (i) potently blocked by 31-100 lg/kg NIDA41020 (CB 1 ), (ii) unaffected by 180 lg/kg AM630 (CB 2 ), 31 lg/kg cannabidiol (GPR55) or 31-100 lg/kg capsazepine (TRPV1) and (iii) slightly blocked by 310 lg/kg AM630. The above doses of antagonists were enough to block their respective receptors. These results suggest that anandamide-induced inhibition of the vasodepressor sensory CGRPergic outflow is mainly mediated by pre-junctional activation of CB 1 receptors, with no pharmacological evidence for the role of CB 2 , TRPV1 or GPR55 receptors.Calcitonin gene-related peptide (CGRP) is a vasodilator neuropeptide that plays a pivotal role in cardiovascular regulation [1,2]. Indeed, resistance blood vessels are innervated by sympathetic [3] and primary sensory CGRPergic [4,5] nerves, which modulate the vascular tone. Certainly, electrical stimulation of the spinal cord (T 9 -T 12 ) in pithed rats receiving i.v. continuous infusions of hexamethonium (to block the vasopressor sympathetic outflow) and methoxamine (for a sustained increase in blood pressure) produces vasodepressor responses mainly mediated by CGRP release from perivascular sensory nerves [6]. Moreover, our group has shown that this vasodepressor CGRPergic outflow is inhibited by stimulation of a 2A/2C -adrenoceptors [7] as well as by serotonin 5-HT 1B [8] and 5-HT 1F [9] receptors located on perivascular sensory nerves.Interestingly, the endocannabinoid system has also been reported to modulate the perivascular release of CGRP. For example, the endocannabinoid anandamide (which interacts with CB 1 , CB 2 , TRPV1 and GPR55 receptors) can modulate CGRP release from rat dural and skin perivascular...

Olcegepant blocks neurogenic and non‐neurogenic CGRPergic vasodepressor responses and facilitates noradrenergic vasopressor responses in pithed rats

“…bolus injections of exogenous α‐CGRP. This weight range is optimal for eliciting vasodepressor responses under our experimental conditions, and the cardiovascular model in pithed rats has been in use for several years (Villalón et al, ; Marichal‐Cancino et al, ). Catheters were placed in: (a) the left femoral vein for the continuous infusion of methoxamine; (b) the right femoral vein for the bolus injection of gallamine or exogenous α‐CGRP; (c) the left jugular vein for the continuous infusion of hexamethonium; and (d) the right jugular vein for the bolus injections of olcegepant or its corresponding vehicle.…”

“…bolus injections of exogenous α‐CGRP (0.1, 0.18, 0.3, 0.56 and 1 μg·kg −1 ). Each response was obtained under unaltered baseline values ​​of blood pressure and heart rate, as reported previously (Villalón et al, ; Marichal‐Cancino et al, ). Set 2 (animal weight: 250–280 g; age range: 12–16 weeks; n = 20) for the analysis of the vasopressor responses induced by either electrical stimulation of the perivascular sympathetic outflow or i.v. bolus injections of exogenous noradrenaline.…”

Potential Mechanisms Involved in Palmitoylethanolamide-Induced Vasodepressor Effects in Rats

“…Together, these findings shed further light on the potential role of palmitoylethanolamide for modulating per se the systemic vascular tone (i.e., the vasopressor responses to sympathetic stimulation and exogenous noradrenaline) and not only as an "entourage" compound. It is important to note that all antagonists (used at doses high enough to completely block their respective receptors in pithed rats [8,9]) failed to modify per se the baseline diastolic blood pressure and heart rate. There- fore, any effect of a given antagonist on the vasopressor sympathoinhibition can be attributed to a direct interaction of the antagonist with its respective receptors on the vasopressor sympathetic nerves and not to physiological antagonism.…”

“…Endogenous cannabinoids and related mediators (i.e., endocannabinoids and endocannabinoid-like compounds) play a role in the modulation of a wide variety of physiological functions with potential therapeutic use [1][2][3][4][5][6][7]. In cardiovascular homeostasis, they exert actions on systemic blood pressure and local blood flow by mechanisms partially elucidated, including: (i) autonomic and sensory neural modulation [8,9]; (ii) stimulation of endothelial nitric oxide [10]; and (iii) control of smooth muscle cell activity [11]. Modulation of vascular tone by endocannabinoids involves activation of vascular and prejunctional G-protein-coupled receptors (CB 1 , CB 2 and GPR55), capsaicin receptors (TRPV1) and nuclear receptors (PPARα and PPARγ) [8,9,12].…”

“…In cardiovascular homeostasis, they exert actions on systemic blood pressure and local blood flow by mechanisms partially elucidated, including: (i) autonomic and sensory neural modulation [8,9]; (ii) stimulation of endothelial nitric oxide [10]; and (iii) control of smooth muscle cell activity [11]. Modulation of vascular tone by endocannabinoids involves activation of vascular and prejunctional G-protein-coupled receptors (CB 1 , CB 2 and GPR55), capsaicin receptors (TRPV1) and nuclear receptors (PPARα and PPARγ) [8,9,12]. The modulation of vascular tone by cannabinoids is of such importance that they induce prolonged hypotension in the nanomolar range doses [13].…”