Specific β ₂ AR Blocker ICI 118,551 Actively Decreases Contraction Through a G _i -Coupled Form of the β ₂ AR in Myocytes From Failing Human Heart

Abstract

Background-We have observed direct (noncatecholamine-blocking) negative inotropic effects of the selective ␤ 2 -adrenoceptor (AR) antagonist ICI 118,551 in myocytes from failing human ventricle. In this study we characterize the effect in parallel in human myocytes and in myocytes from animal models where ␤ 2 ARs or G i proteins are overexpressed. Methods and Results-Enzymatically isolated, superfused ventricular myocytes were exposed to ␤AR agonists and antagonists/inverse agonists, and contraction amplitude was measured. ICI 118,551 decreased contraction in ventricular myocytes from failing human hearts by 45.3Ϯ4.1% (nϭ20 hearts/31 myocytes, PϽ0.001) but had little effect in nonfailing hearts (4.9Ϯ4%, nϭ5 myocytes/3 hearts). Effects were significantly larger in patients classified as end-stage. Transgenic mice with high ␤ 2 AR number and increased G i levels had normal basal contractility but showed a similar negative inotropic response to ICI 118,551. Overexpression of human ␤ 2 AR in rabbit myocytes using adenovirus potentiated the negative inotropic effect of ICI 118,551. In human, rabbit, and mouse myocytes, the negative inotropic effects were blocked after treatment of cells with pertussis toxin to inactivate G i , and overexpression of G i ␣ 2 induced the effect de novo in normal rat myocytes. Conclusions-We hypothesize that ICI 118,551 binding directs the ␤ 2 AR to a G i -coupled form and away from the G s -coupled form (ligand-directed trafficking). ICI 118,551 effectively acts as an agonist at the G i -coupled ␤ 2 AR, producing a direct negative inotropic effect. Conditions where ␤ 2 ARs are present and G i is raised (failing human heart, TG␤ 2 mouse heart) predispose to the appearance of the negative inotropic effect.

The Third Intracellular Loop and the Carboxyl Terminus of β2-Adrenergic Receptor Confer Spontaneous Activity of the Receptor

“…It has been demonstrated that the inverse agonist ICI 118,551 can activate the ␤ 2 -AR/G i pathway, resulting in a negative inotropic effect in cardiomyocytes from failing human heart or transgenic mice overexpressing ␤ 2 -AR (TG␤2) in a PTX-sensitive manner (Gong et al, 2002). To determine whether the inhibitory effects of ICI 118,551 on basal contraction is mediated by activation of the ␤ 2 -AR/G i pathway, we compared the effects of ICI 118,551 in cells expressing ␤ 2 -AR or the chimeras in the presence and absence of PTX treatment.…”

Beyond Desensitization: Physiological Relevance of Arrestin-Dependent Signaling

“…Angiotensin AT 1A receptor signaling has been studied using receptors mutated in the conserved DRYXX(V/I)XXPL region of the amino terminal portion of the second intracellular loop: D125G/R126G/Y127A/M134A (AT 1 -i2m) and D125A/ R126A (DRY/AAY), which fail to activate G protein signaling but nonetheless support arrestin recruitment (Seta et al, 2002;Gá borik et al, 2003), and using the angiotensin II analog Sar 1 -Ile 4 -Ile 8 -Ang II, which antagonizes G q/11 coupling but promotes GRK phosphorylation, arrestin recruitment, and receptor endocytosis (Holloway et al, 2002). Analogous reagents for the ␤ 2 adrenergic receptor include the T68F/Y132G/Y219A mutant [␤ 2 AR(TYY)] (Shenoy et al, 2006) and a number of arrestin pathway-selective ␤ 2 receptor inverse agonists, including ICI118,551 and the clinically used drugs propranolol and carvedilol (Gong et al, 2002;Azzi et al, 2003;Wisler et al, 2007;Drake et al, 2008).…”

Cardiac Adrenoceptors: Physiological and Pathophysiological Relevance

“…Thus, taken together, evidence accumulated so far makes it rather doubtful whether in human heart, β 2 AR can couple to, besides G s -protein, G i -protein. In this context, however, it is interesting to note that Gong et al (21) have found that in ventricular cardiomyocytes from patients with heart failure (who exhibit increased ventricular G i -protein activity, see below), but not in cardiomyocytes from non-failing human hearts (with normal G i -protein), the β 2 AR-antagonist ICI 118,551 exerted agonistic effects evoking direct negative inotropic effects. Whether or not β 3 AR might exist in the human heart, is also still an open question.…”