Chronic hyperglycemia in type-1 diabetes mellitus is associated with oxidative stress (OS) and sudden death. Mechanistic links remain unclear. We investigated changes in electrophysiological (EP) properties in a model of chronic hyperglycemia before and after challenge with OS by GSH oxidation and tested reversibility of EP remodeling by insulin. Guinea pigs survived for 1 mo following streptozotocin (STZ) or saline (sham) injection. A treatment group received daily insulin for 2 wk to reverse STZ-induced hyperglycemia (STZ + Ins). EP properties were measured using high-resolution optical action potential mapping before and after challenge of hearts with diamide. Despite elevation of glucose levels in STZ compared with sham-operated (P = 0.004) and STZ + Ins (P = 0.002) animals, average action potential duration (APD) and arrhythmia propensity were not altered at baseline. Diamide promoted early (<10 min) formation of arrhythmic triggers reflected by a higher arrhythmia scoring index in STZ (P = 0.045) and STZ + Ins (P = 0.033) hearts compared with sham-operated hearts. APD heterogeneity underwent a more pronounced increase in response to diamide in STZ and STZ + Ins hearts compared with sham-operated hearts. Within 30 min, diamide resulted in spontaneous incidence of ventricular tachycardia and ventricular fibrillation (VT/VF) in 3/6, 2/5, 1/5, and 0/4 STZ, STZ + Ins, sham-operated, and normal hearts, respectively. Hearts prone to VT/VF exhibited greater APD heterogeneity (P = 0.010) compared with their VT/VF-free counterparts. Finally, altered EP properties in STZ were not rescued by insulin. In conclusion, GSH oxidation enhances APD heterogeneity and increases arrhythmia scoring index in a guinea pig model of chronic hyperglycemia. Despite normalization of glycemic levels by insulin, these proarrhythmic properties are not reversed, suggesting the importance of targeting antioxidant defenses for arrhythmia suppression.

Non‐Technical Summary Oxidative stress is a hallmark of various cardiovascular disorders that results in cellular dysfunction and death. Reactive oxygen species (ROS)‐induced ROS release (RIRR) is a fundamental mechanism which amplifies ROS levels within the cardiomyocyte resulting in cellular oxidative stress. Despite elegant studies describing the phenomenon of RIRR in isolated myocytes, its biophysical properties and functional consequences in intact myocardium remain unclear. Here, we use ROS imaging to extend the concept of RIRR to the level of the intact heart. We establish regenerative superoxide production as the mediator of RIRR‐related arrhythmias and reveal their strong dependence on a key mitochondrial channel, known as the inner membrane anion channel (IMAC). We demonstrate the efficacy of suppressing RIRR and related arrhythmias either by pharmacologically blocking IMAC or scavenging ROS using a synthetic superoxide dismutase/catalase mimetic.

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 (Gi) 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. 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’. Upon ligand binding, β-ARs are physically coupled to heterotrimeric G proteins, which are classified as stimulatory (Gs) or inhibitory (Gi). In cardiomyocytes, coupling of the receptors with Gs 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 Gs, leading to acute activation of the AC–PKA pathway. Interestingly, β2-ARs also couple to the pertussis toxin-sensitive Gi pathway which eventually inhibits Gs. Gi promotes cardiomyocyte survival through a protective PI3K–PKB pathway (Santos & Spadari-Bratfisch, 2006). Interestingly, Gi-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). 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. F.G.A. is supported by grants from the NIH NHLBI (HL091923), the American Heart Association (0830126N), and the Irma T. Hirschl and Monique Weill Caullier Trusts.