Induction of β₃-Adrenergic Receptor Functional Expression following Chronic Stimulation with Noradrenaline in Neonatal Rat Cardiomyocytes

Abstract

This study aimed to characterize ␤ 3 -adrenergic receptors (ARs) in rat neonatal cardiomyocytes using the noradrenaline (NOR) properties to modulate the expression and function of the three ␤-ARs. We assessed the effect of NOR (physiological nonselective agonist), isoprenaline (ISO, ␤-nonselective agonist), dobutamine (DOB, ␤ 1 -selective agonist), and procaterol (PROC, ␤ 2 -selective agonist) on cAMP accumulation using cardiomyocytes untreated or treated with 100 M NOR for 24 h. The inhibition of forskolin-stimulated cAMP accumulation was determined using NOR, isoprenaline, and the In addition, the level of the three subtypes was determined by reverse transcription polymerase chain reaction and Western blotting. NOR pretreatment decreased the activation of cAMP induced by NOR, isoprenaline, and DOB, whereas PROC response was abolished. The inhibition of NOR response by CGP 20712A or ICI 118551 demonstrated that ␤ 1 -and ␤ 2 -ARs are down-regulated and that ␤ 2 -AR functional activity was also abolished in cardiomyocytes exposed to chronic stimulation. ␤ 3 -AR function was observed with NOR and ISO when ␤ 1 -/␤ 2 -ARs were blocked and with both ␤ 3 -selective agonists in NOR-treated cells only. This response was completely inhibited by SR 59230A and involved G i protein. Furthermore, the results from functional studies agree well with those from expression experiments. In conclusion, these data provide strong evidence that ␤ 3 -ARs are functionally upregulated and coupled to G i protein in rat neonatal cardiomyocytes following chronic exposure to NOR when ␤ 1 -and ␤ 2 -ARs are down-regulated.Three ␤-adrenergic receptor subtypes (␤ 1 -, ␤ 2 -, and ␤ 3 -AR) have been cloned and pharmacologically characterized (Strosberg, 1995). These different subtypes belong to the G protein-coupled receptor superfamily and modulate cardiac function after stimulation by the catecholamines, noradrenaline, and adrenaline. Increases in heart rate and force of contraction are mediated mainly by ␤ 1 -ARs and to a lesser extent by the ␤ 2 -AR (Dzimiri, 1999;Steinberg, 1999). These functional effects result from the sequential activation of stimulatory G s proteins, adenylyl cyclase, and protein kinase A (PKA), leading to the phosphorylation of proteins involved in cardiac contractility (troponin I, L-type Ca 2ϩ channels, and phospholamban) (Post et al., 1999). Previous studies have shown that the ␤ 2 -AR also couples to G i protein in cardiomyocytes (Xiao, 2001). In contrast to ␤ 1 -and ␤ 2 -AR, activation of the ␤ 3 -AR induces a negative inotropic effect in different species, including human, dog, guinea pig, and rat This work was supported by the British Heart Foundation Grant FS/03/ 095/16317.Article, publication date, and citation information can be found at

β3-Adrenoceptor Stimulation on Top of β1-Adrenoceptor Blockade

“…Contrary to ␤ 1 -AR (and to a lesser extent ␤ 2 -AR), the abundance of ␤ 3 -AR messenger ribonucleic acid (and proteins) increases in ventricular myocardium from patients with heart failure from ischemic or dilated cardiomyopathies (19), as well as sepsis (20). Such paradoxical regulation is consistent with in vitro up-regulation of ␤ 3 -ARs in rodent cardiomyocytes chronically exposed to catecholamines (21), as well as animal models of diabetes (22), a condition also associated with hyperadrenergism. Also, ␤ 3 -ARs are more resistant (than ␤ 1 -and ␤ 2 -ARs) to homologous desensitization (23).…”

Regulation of gap-junction protein connexin 43 by β-adrenergic receptor stimulation in rat cardiomyocytes

“…TRITC-conjugated anti-rabbit IgG were from Beijing Zhongshan Golden Bridge Biotechnology (Beijing, China). The doses of ISO, propranolol, ICI 118551, CGP20712A, PTX, H89, clenbuterol were used according to the international concentra tion [10,13] .…”

“…Responses to sympathetic activation are mediated through the action of the endogenous catecholamines norepinephrine and epinephrine on adrenergic receptors (ARs). The heart expresses all three subtypes of β-AR, as well as three types of α 1 -AR: α 1A -AR, α 1B -AR, and α 1D -AR [10] . A recent report demonstrated that α-adrenergic receptor agonist phenylephrine enhanced Cx43 expression, but not Cx40 and Cx45 expression, in neonatal rat cardiac myocytes (NRCMs), resulting in enhanced gap-junction conductance.…”

Acute Stress Differently Modulates Beta 1, Beta 2 and Beta 3 Adrenoceptors in T Cells, but Not in B Cells, from the Rat Spleen

“…It was shown that after short-term exhaustive exercise, characterized by increased plasma CAs, increased ␤ 2 -AR expression and density was observed in human peripheral lymphocytes [29,30] . In addition, ␤ 3 -ARs were shown to be upregulated in rat neonatal cardiomyocytes following chronic exposure to noradrenaline [31] . The ␤ 1 -AR overexpression has been detected in peripheral lymphocytes of hypertensive and cardiac patients [18,19] that are often associated with higher plasma CAs [32,33] .…”