A brighter side of ROS revealed by selective activation of β-adrenergic receptor subtypes

Because of the high metabolic demands on the heart, oxidative phosphorylation predominates over the less efficient process of anaerobic glycolysis for energy production. Under basal conditions, ∼10% of the body's total oxygen supply is consumed by the heart (O’Rourke et al. 2005). This already staggering requirement rapidly increases during stress. Hence, oxygen bioavailability is a critical determinant of cardiomyocyte function and survival in health and disease. In this issue of The Journal of Physiology, Li et al. (Li et al. 2010) dissect the molecular mechanisms and cell signalling pathways underlying β-adrenergic receptor (AR)-mediated modulation of intracellular oxygen levels during the ‘fight-or-flight’ response. Specifically, these authors demonstrate that β₂- and not β₁-AR activation acutely increases intracellular oxygen levels through an inhibitory G protein (G_i) pathway, which is dependent on reactive oxygen species (ROS) also generated through this pathway. Indeed, the findings of Li et al. (2010) have far-reaching implications for our understanding of mechanisms underlying a wide variety of stress-related cardiovascular disorders, and can help explain the benefits and pitfalls associated with current therapies for these disorders.

ROS: the good, the bad and the ugly

Energy production requires the transfer of electrons from donors to acceptors in a series of redox reactions that occur within the electron transport chain (ETC). During this process a small percentage of electrons are continuously leaked out of the ETC, combining with molecular oxygen to produce ROS. As by-products of metabolism, ROS exert deleterious effects on cellular structure and function. In fact, excessive ROS formation and/or reduced ROS scavenging causes oxidative stress, a hallmark of cardiovascular, neurological and other age-related disorders. This ‘ugly’ side of ROS is normally countered by sophisticated cellular antioxidant defences. However, the notion that ROS are only harmful by-products of metabolism is inaccurate, since they also have critical cell signalling functions. Li et al. (2010) demonstrate the dependence of β₂-AR signalling on ROS. Because of the established benefits of β₂ signalling on cardiomyocyte survival, increased ROS levels through the β₂ pathway can therefore be considered beneficial, not harmful. As such, it is becoming quite clear that ‘not all ROS are created equal’.

Acute versus chronic activation of β-adrenergic receptors

Upon ligand binding, β-ARs are physically coupled to heterotrimeric G proteins, which are classified as stimulatory (G_s) or inhibitory (G_i). In cardiomyocytes, coupling of the receptors with G_s results in the acute activation of the adenylyl cyclase (AC)–cAMP–PKA pathway. This causes the phosphorylation of downstream components critical for excitation–contraction coupling (ECC) and electrophysiological function. Specifically, PKA-mediated phosphorylation of L-type calcium channels and phospholamban (PLB) enhances Ca²⁺ influx from the extracellular space and the rate of Ca²⁺ sequestration into the sarcoplasmic reticulum (SR) by sarco(endo)plasmic reticulum Ca²⁺-ATPase 2a (SERCA2a), respectively. Enhanced ECC underlies the acute positive inotropic and lusitropic effects of β-AR activation. Moreover, PKA-dependent phosphorylation of ion channels enhances heart rate accounting for a positive chronotropic response. While these acute effects provide an immediate increase in cardiac output during the ‘fight-or-flight’ response, the beneficial effects of β₁-AR stimulation are short lived. In fact, β₁-AR-mediated phosphorylation of ryanodine receptor RYR2 by Ca²⁺–calmodulin-dependent kinase II increases its open probability, enhances diastolic Ca²⁺ leak from the SR, and causes triggered arrhythmias (Ai et al. 2005).

Prolonged exposure to stress leads to eventual desensitization of β₁-ARs and their disrupted coupling to G proteins (El-Armouche & Eschenhagen, 2009). As a result, reduced PKA activity leads to hypo-phosphorylation of PLB, unveiling its inhibitory effect on SERCA2a. This leads to cytosolic Ca²⁺ overload and associated contractile and electrical dysfunction (El-Armouche & Eschenhagen, 2009).

It is now well established that β-AR agonists achieve stronger contractility by enhancing activator calcium at the expense of increasing mortality in animal models and patients with heart failure. Moreover, chronic β₁-AR activation increases ROS formation from mitochondrial and cellular oxidases while transcriptionally downregulating cytosolic superoxide dismutase. This causes ROS-mediated oxidative stress (Srivastava et al. 2007), mitochondrial dysfunction, permeability transition pore opening and apoptosis (Singh et al. 2001). Altered mitochondrial stability impairs excitability at the cellular and tissue levels, leading to metabolic sink block and arrhythmias (Akar et al. 2005; O’Rourke et al. 2005).

Like β₁-, β₂-ARs also couple to G_s, leading to acute activation of the AC–PKA pathway. Interestingly, β₂-ARs also couple to the pertussis toxin-sensitive G_i pathway which eventually inhibits G_s. G_i promotes cardiomyocyte survival through a protective PI3K–PKB pathway (Santos & Spadari-Bratfisch, 2006). Interestingly, G_i-mediated ROS are required for the stability of the active conformation of β₂-ARs, which probably facilitates their beneficial cell signalling effects, including modulation of intracellular oxygen availability, by preventing their internalization and desensitization (Moniri & Daaka, 2007).

Therapeutic implications

β-ARs are prime targets for heart failure treatment. β-Agonists provide acute inotropic support by increasing activator calcium, but result in sudden death due to ventricular tachyarrhythmias. In sharp contrast, β-blockers are a mainstay for heart failure therapy as they clearly improve clinical outcome, but fail to enhance contractility in the short term.

An ideal therapeutic approach entails the suppression of ‘bad’ ROS from the β₁-AR pathway while promoting the generation of ‘good’ ROS from the β₂-AR pathway. The use of selective antagonists seems appropriate in this regard. This, however, is complicated by strong feedback mechanisms whereby blocking the activity of a given subtype potentiates the signalling associated with the other. More importantly, the downstream effects of activating either subtype are neither purely beneficial nor detrimental due to complex feedback mechanisms, and a dynamically changing sub-cellular localization and activity of these receptors during disease development. Therefore, optimal therapeutic approaches should allow the careful titration of the intensity, timing and duration of β-AR subtype activation/blockade during an ever-changing sympatho-adrenergic state. Clearly, genetic manipulation of β-AR densities may not provide such leniency (El-Armouche & Eschenhagen, 2009). Pharmacotherapies, on the other hand, may offer greater control in terms of varying drug dosage and timing. A more comprehensive understanding of mechanisms by which β-AR subtypes alter cell survival and death pathways by modulating cellular ROS and oxygen levels is required for developing effective and safe therapies. The paper by Li et al. (2010) represents a discrete step in this direction.