Negative inotropy starts with the ?-adrenoceptor

See article by Tavernier et al. [1] (pages 288–296) in this issue. In this issue of Cardiovascular Research, Tavernier et al. describe the effects of β3-adrenergic stimulation on cardiac contractility in mice overexpressing the human β3-adrenoceptor (β3-AR) [1]. This study is a logical sequel to preceding investigations by the same research group that also provided the first evidence of the existence and functional role of this AR subtype in the human heart [2]. That time it was found that in contrast to the classical effects of β1-/β2-adrenergic system—positive inotropy and lusitropy—activation of β3-AR results in negative inotropy via a Gi protein-dependent pathway. Further studies revealed that β3-AR stimulation attenuates cardiac contractility through increased nitric oxide (NO) production and intracellular cGMP [3,4]. To further explore the properties and role of the β3-adrenergic system, Tavernier et al. [1] generated transgenic mice overexpressing the human β3-AR in cardiac cells. The human receptor was selected for two reasons: (i) compared to other species, human cardiac muscle exhibits the strongest negative inotropic effect induced by β3-AR agonists [5] and, (ii) cardiac failure in humans is associated with increased expression of β3-AR (see below). The effectiveness of gene expression was confirmed by polymerase chain reaction and Western blotting, showing human β3-AR mRNA and corresponding protein in transgenic mice, but not in wild-type mice. Then the contractility experiments were performed under the same conditions as those used earlier for human myocardium [2]. The results show that expression of β3-adrenoceptor was accompanied by decreased peak tension in response to low concentrations of β3-AR agonists (CL 316243 and SR 58611A), consistent with their β3-AR-specific action. In parallel, an increase in intracellular cGMP level was observed. Thus, the β3-ARs were functional in that they were coupled to downstream components of the NO-dependent signalling pathway, and thereby capable of reproducing the negative inotropic response qualitatively similar to that observed in human [2], dog [5], and guinea pig [6] cardiac preparations by β3-AR stimulation. These data clearly show that the role of β3-ARs in cardiac muscle is to initiate negative inotropic influence on cardiac contraction. The mechanisms of β3-AR-mediated attenuation of cardiac contractility are largely undiscovered, but the current evidence suggests the involvement of alterations in excitation–contraction coupling and transmembrane ion channels. A β3-AR agonist BRL 37344 is shown to induce a dose-dependent negative inotropic effect, accompanied by inhibition of L-Ca2+ channels and decreased amplitude of the intracellular Ca2+ transient [6,7]. These changes may be caused by increased cGMP levels that, either via activation of cGMP-dependent protein kinases or through attenuation of the cAMP levels by activation of cGMP-stimulated phosphodiesterases, suppress the current through L-Ca2+ channels [8]. NO, another mediator of the pathway, may exert a similar effect by covalent modification of the L-Ca2+ channel protein [8]. On the other hand, the finding that BRL 37344 decreases the amplitude and action potential duration in human endomyocardial biopsies [2] points to potential alterations in repolarizing currents. In line with this, the β3-AR was found to be functionally coupled to cystic fibrosis transmembrane conductance regulator (CFTR) channels. This enables activation of a repolarizing Cl− current under stimulation of the β3-adrenergic pathway [9]. The CFTR protein is found to be present and functional in murine heart [10]. Therefore, it could mediate decreased inotropy under stimulation of β3-ARs in the present study [1]. Controversial data exist regarding the role of K+ currents, however. It has been suggested that β3-ARs are coupled to the K+ channel KvLQT1/MinK [11]. This explains the shortening of the human cardiac action potential [2]. However, others have demonstrated that although the β3-AR is functionally coupled to a K+ channel, its stimulation with BRL 37344 brings about inhibition of IKs, thereby increasing the action potential duration in guinea-pig cardiomyocytes [12]. In experiments of Tavernier et al. [1], the role of IKs may be disregarded due to the lack of IKs in cardiac cells of adult mice [13]. This current functions in guinea pig and human hearts [12], however, and thus the decreased inotropy by β3-agonist could be attributed to changes in altered balance between inward and outward currents. The mechanisms of regulation of cardiac contractility by β3-AR-mediated pathways need to be further addressed in order to test the potential effects of increased cGMP and NO on troponin I, sarcoplasmic reticulum, and enzymes of energy metabolism, e.g., cytochrome oxidase and creatine kinase [8,14].