Digitalis and Na/Ca exchange: Old dog learns new mitochondrial tricks

Digitalis glycosides (like ouabain or digoxin) are known to specifically inhibit the Na/K-ATPase and have been used to enhance cardiac contractility in heart failure (HF) for more than 200 years.1,2 The main and generally accepted mechanism of action is Na/K-ATPase inhibition causes a rise in intracellular [Na] ([Na]i) which limits Ca extrusion (and may even favor Ca entry) by the cardiac sarcolemmal Na/Ca exchange. This can increase diastolic [Ca]i, sarcoplasmic reticulum (SR) Ca content and the amplitude of the Ca transient amplitude, leading to a positive inotropic effect. However, the therapeutic range is limited because if SR Ca content gets too high (glycoside toxicity), spontaneous SR Ca releases during diastole can activate inward Na/Ca exchange current leading to delayed after depolarizations and triggered arrhythmias. There is also evidence suggesting that ouabain binding to the Na/K-ATPase can trigger activation of protein kinase signaling pathways,3 but these signaling pathways may be more relevant to longer-term changes in cardiac function. The mitochondrial inner membrane also expresses a Na/Ca exchanger (a different molecular entity than the sarcolemmal Na/Ca exchange) and this is the major pathway for Ca efflux from mitochondria. This Ca efflux pathway and Ca influx via the mitochondrial Ca uniporter are the key regulators of normal intra-mitochondrial free [Ca] ([Ca]Mito). With increasing myocardial workloads (due to increased heart rate or larger Ca transients) there is a consequent rise in [Ca]Mito which causes enhancement of mitochondrial energy production because (in part) there are Ca-dependent dehydrogenases (pyruvate, isocitrate, and α-ketoglutarate dehydrogenase) that increase NADH production to fuel oxidative phosphorylation and ATP generation.4,5 In this way, elevated [Ca]i which is causative for stronger contractions and energy consumption by myosin- and Ca-ATPases inherently increases ATP production mediated by a feed-forward enhancement of [Ca]Mito. Measurement of [Ca]Mito in intact myocytes is challenging, especially with respect to the dissociation of [Ca]i from [Ca]Mito, but O’Rourke and colleagues have produced compelling myocyte [Ca]Mito data in a series of studies that culminate with a paper related to ouabain effects in this issue of the Journal of Molecular and Cellular Cardiology.6–9 A central and convincing working hypothesis of these studies is that elevated [Na]i (as occurs in HF or ouabain toxicity) lowers [Ca]Mito and thus limits the ability of elevated [Ca]Mito with workload to help energy supply meet energy demands. This may create an intrinsic limitation in energetic reserve when [Na]i is elevated in HF or with digitalis therapy. They show also that the fall in NADH levels as a consequence of reduced [Ca]Mito may also enhance production of mitochondrial reactive oxygen species (ROS)7 that can have secondary deleterious effects on numerous downstream targets that can lead to long term damage. So part of the classical digitalis toxicity may be due to altered mitochondrial Ca regulation and negative consequences of reduced [Ca]Mito and increased ROS production. This could certainly be additive with the classical view regarding SR Ca overload and spontaneous SR Ca release in initiating triggered arrhythmias. Indeed, altered function of some likely ROS targets such as the SR Ca release channel (RyR), voltage-gated sarcolemmal ion channels (e.g. Na channels) and Ca-Calmodulin dependent protein kinase10 (which itself alters RyR, Na channels and K channels) could worsen the intrinsic arrhythmogenic potential.11 While ouabain can cause inotropic effects due to elevated [Na]i and sarcolemmal Na/Ca exchange function, it does so with apparently three costs: 1) increased arrhythmogenesis, 2) reduced energetic reserve and 3) enhanced ROS production (which may indirectly enhance arrhythmogenesis). It is an interesting point that increased [Na]i enhances cytosolic [Ca]i and Ca transients, but has the opposite effect of diminishing [Ca]Mito and mitochondrial Ca transients (and both are mediated by different Na/Ca exchangers). In particular this opposite effect on [Ca]i and [Ca]Mito completely undermines the elegant natural control system in which physiological increases in [Ca]i (due to increased heart rate or enhanced Ca influx and Ca transients by β-adrenergic agonists) cause increases in [Ca]Mito which help the myocyte match ATP production with the increase in workload that usually accompanies enhanced cardiac inotropy or chronotropy. The implication is that the control of [Na]i in healthy cardiac myocytes is in a delicate balance, where it allows a dynamic balance of inotropic state (via sarcolemmal Na/Ca exchange) and mitochondrial energy and ROS production (via mitochondrial Na/Ca exchange). Regulation of [Na]i in myocytes has not generally been appreciated as much as have [Ca]i or pH regulation, except that [Na]i is a direct modulator of [Ca]i and pH via Na/Ca exchange and Na/H exchange, and that [Na]o – [Na]i gradient provides the chemical driving force for the function of Na channels and other transporters. The present study9 and prior work by this group6–8 has made it especially clear how dysregulation of [Na]i within the physiological range has tremendous and underappreciated consequences for cardiac mitochondrial function. And these [Na]i-dependent alterations in mitochondrial function have far-reaching effects beyond energetics to excitation-contraction coupling, arrhythmias and oxidative stress. Since [Na]i is already elevated in HF and the propensity for ROS production may also be enhanced in HF,12 this problem may be exacerbated with respect to glycoside use in HF. However, this paper by Liu et al.9 offers a ray of hope regarding enhanced utility of digitalis in HF. That is, by combining digitalis (a Na/K-ATPase inhibitor) with a selective mitochondrial Na/Ca exchange inhibitor (e.g. CGP-37157), one may hope to get enhanced inotropic effects, while limiting the negative energetic and arrhythmogenic effects of digitalis. Indeed, it is remarkable that partial inhibition of the mitochondrial Na/Ca exchange by CGP-37157 here and in HF myocytes6,9 can restore [Ca]Mito without overshooting the mark (which could also be deleterious), prevent NADH oxidation and ROS production, and also limit ouabain-induced arrhythmias (while enhancing ouabain- and isoproterenol-induced inotropic state). Of course, one must not completely block the mitochondrial Na/Ca exchange, because then [Ca]Mito would progressively rise to levels where the mitochondrial permeability transition pore (mPTP) would open and cause loss of mitochondrial membrane potential, ability to generate ATP and eventual mitochondrial and cell death. Therefore this intriguing new potential therapeutic opportunity must be studied in more detail to determine its true therapeutic potential. Regardless of the eventual outcome, these studies have opened a valuable new window into understanding in the cardiac myocyte environment how [Na]i regulates mitochondrial function and dysfunction.