A Leakage Leads to Failure
Roles of Sarcoplasmic Reticulum Ca2+ Leak via RyR2 in Heart Failure Progression
Cytosolic-free calcium (Ca2+) is a multifunctional intracellular messenger that regulates many different cellular processes in cardiac myocytes.1 A transient rise in intracellular free Ca2+ concentrations ([Ca2+]i) during excitation-contraction (E-C) coupling is required for initiating contraction of cardiac muscle. Membrane depolarization activates voltage-gated L-type Ca2+ channels within the transverse tubules (invaginations of the sarcolemma), and extracellular Ca2+ enters cardiac myocytes. The increased [Ca2+]i in the junctional space between transverse tubules and the sarcoplasmic reticulum (SR) triggers the Ca2+-induced Ca2+ release from the SR via ryanodine receptor (RyR2)/intracellular Ca2+ release channels. The resultant transient rise in global [Ca2+]i activates the myofilaments to produce cardiac contraction. Myocyte relaxation occurs when [Ca2+]i levels decline quickly through transport by the Ca2+ recycling proteins, such as SR Ca2+-ATPase (SERCA), which pumps Ca2+ back into SR, and the Na+/Ca2+ exchanger, which extrudes Ca2+ out of myocytes.1 In addition to its pivotal role in cardiac E-C coupling, [Ca2+]i is also a critical regulator of multiple signaling transduction pathways, including activation of protein kinases or protein phosphatases and modulation of gene transcription and expression (Figure).2
SR Ca2+ content reflects the balance between Ca2+ uptake (via SERCA) and Ca2+ efflux (via RyR2). A normal SR Ca2+ content is the key for the maintenance of physiological [Ca2+]i levels and, thus, normal contractile function of cardiac myocytes. Under conditions of persistent pathological stress on the heart, such as pressure overload–induced myocardial hypertrophy and heart failure (HF), SR Ca2+ content is reduced, and [Ca2+]i is increased. Both reduced Ca2+ pumping by SERCA and increased SR Ca2+ leak via RyR2s have been shown to contribute to the reduced SR Ca2+ content and increased [Ca2+]i.3 Other possible sources of the hypertrophy-associated increase in [Ca2+]i may be from increases in influx of extracellular Ca2+ through the voltage-gated L-type Ca2+ channel or store-operated Ca2+ channels and release of stored Ca2+ from the nucleus via the inositol 1,4,5-trisphosphate receptors (IP3Rs; Figure).1,2 Sustained elevation of [Ca2+]i drives a complex pattern of remodeling of Ca2+ handling proteins and plays a key role in the activation of several hypertrophic signaling pathways.1,2 These include the Ca2+/calmodulin (CaM)-calcineurin-nuclear factor of activated T cells (NFAT)4 and the Ca2+-CaM–dependent kinase II (CaMKII)-histone deacetylase (HDAC) pathways.5 However, it remains a mystery exactly which subcellular Ca2+ pools initiate these hypertrophic signaling and how these Ca2+-dependent signaling pathways are regulated in contracting cardiac myocytes given the highly specialized manner in which Ca2+ concentration rhythmically cycles in E-C coupling.6 It has been demonstrated previously that IP3R-mediated nuclear envelope Ca2+ fluxes may activate Ca2+-CaM-CaMKII-HDAC5 signaling in ventricular myocytes, and the enhanced store-operated Ca2+ entry into myocytes with overexpressed transient receptor protein C3 may increase NFAT transcriptional activity.7 It was suggested that enhanced Ca2+ release through IP3Rs may cause sensitization of RyR2s and a further increase in diastolic [Ca2+]i. However, it is very controversial at present whether altered patterns of Ca2+ release and reuptake associated with E-C coupling may affect hypertrophic signaling pathways and whether the diastolic SR Ca2+ leak might also activate Ca2+-dependent hypertrophic signaling pathways under pathological conditions.2,6
Previous evidence from HF patients and animal models supports a functional role for enhanced diastolic Ca2+ leak from SR through RyR2s in the development of contractile dysfunction.8 In lipid bilayers, RyR2 phosphorylation caused FK506-binding protein 12.6 dissociation from the RyR2 and increased overall open probability of RyR2 channels. A variety of alterations in the subunits of the RyR2 macromolecular complex have been found in HF patients, including decreased levels of FK506-binding protein 12.6 (or calstabin2), protein phosphatases (1A and 2A), and phosphodiesterase 4D3.8 In addition, changes in RyR2 posttranslational modifications, such as oxidation, S nitrosylation, and phosphorylation, have been shown in HF patients and animal models.8 It was found that RyR2s were “hyperphosphorylated” by protein kinase A possibly because of the hyperadrenergic state and loss of RyR2-associated phosphatases (despite increased global phosphatases in cardiac myocytes). The combination of these alterations of RyR2 may lead to a decreased ability of the channel to remain closed during diastole, resulting in a net increase in diastolic SR Ca2+ leak, a reduced SR Ca2+ content, and an increased [Ca2+]i. Therefore, it has been proposed that enhanced SR Ca2+ leak through “leaky” RyR2 Ca2+ release channels during diastole may underlie contractile dysfunction. However, this attractive hypothesis has been seriously challenged by controversial findings on the effect of protein kinase A–dependent RyR phosphorylation during E-C coupling and the lack of supporting data in intact myocytes or whole-animal models.3
In this issue of Hypertension, van Oort et al9 used knockin (gain-of-function) mice heterozygous for mutation R176Q in RyR2 (R176Q/+), which have been shown previously to increase SR Ca2+ release in atrial and ventricular myocytes after catecholaminergic stimulation to specifically address the question of whether enhanced SR Ca2+ leak through RyR2 accelerates the development of cardiac hypertrophy through the activation of Ca2+-dependent signaling pathways. They found that 8 weeks of transverse aortic constriction decreased systolic and diastolic heart functions and increased ventricular dimensions to a significantly larger extent in R176Q/+ mice when compared with wild-type mice. R176Q/+ mice displayed an enhanced hypertrophic response compared with wild-type mice as assessed by heart weight:body weight ratios and cardiomyocyte cross-sectional areas after transverse aortic constriction. Transverse aortic constriction pressure overload also resulted in an increased SR Ca2+ leak, associated with higher expression levels of the exon 4 splice form of regulator of calcineurin 1 in R176Q/+ mice compared with wild-type mice. Therefore, the authors concluded that RyR2-dependent SR Ca2+ leak may activate the prohypertrophic calcineurin/NFAT pathway during pressure overload (Figure). These findings provide convincing in vivo evidence that increased RyR2-mediated Ca2+ release from the SR is able to activate Ca2+-dependent hypertrophic signaling pathways, preferentially the calcineurin/NFAT signaling pathway, under conditions of pressure overload. These novel findings are consistent with previous reports suggesting that increased diastolic RyR2 Ca2+ leak impairs cardiac contractility because of a secondary decrease in SR Ca2+ loading. Recent clinical studies provided further evidence that genetic defects in the RyR2 gene may predispose patients toward the development of hypertrophic cardiomyopathy and HF.10 Therefore, defective Ca2+ release from the SR via mutant RyR2 may indeed adversely affect cardiac remodeling.
It should be kept in mind, however, that HF is heterogeneous and complex, and multiple mechanisms may contribute to increased [Ca2+]i (reduced SERCA function and enhanced SR Ca2+ leak) and decreased SR Ca2+ content (increased Na+/Ca2+ exchanger function) in HF. However, relative contributions of these mechanisms may vary among species (mouse versus human), pathological stimuli (pressure overload versus volume overload), and disease stages (compensated versus dyscompensented) of HF. It is clear that more studies of SR Ca2+ leak in the context of signalosome of remodeling in myocardial hypertrophy and HF2 are warranted to better understand whether this pathway is causative of hypertrophy or HF and whether targeting RyR2 to reduce the diastolic SR Ca2+ leakage may serve as a novel therapeutic strategy for the treatment of HF.
Sources of Funding
D.D.D. is supported by the National Institutes of Health, National Center for Research Resources P-20 RR-15581, and National Heart, Lung, and Blood Institute grant HL63914.
The opinions expressed in this editorial are not necessarily those of the editors or of the American Heart Association.
Bers DM, Eisner DA, Valdivia HH. Sarcoplasmic reticulum Ca2+ and heart failure: roles of diastolic leak and Ca2+ transport. Circ Res. 2003; 93: 487–490.
Molkentin JD. Calcineurin-NFAT signaling regulates the cardiac hypertrophic response in coordination with the MAPKs. Cardiovasc Res. 2004; 63: 467–475.
Ling H, Zhang T, Pereira L, Means CK, Cheng H, Gu Y, Dalton ND, Peterson KL, Chen J, Bers D, Heller BJ. Requirement for Ca2+/calmodulin-dependent kinase II in the transition from pressure overload-induced cardiac hypertrophy to heart failure in mice. J Clin Invest. 2009; 119: 1230–1240.
Houser SR, Molkentin JD. Does contractile Ca2+ control calcineurin-NFAT signaling and pathological hypertrophy in cardiac myocytes? Sci Signal. 2008; 1: e31.
van Oort RJ, Respress JL, Li N, Reynolds C, De Almeida AC, Skapura DG, De Windt LJ, Wehrens XHT. Accelerated development of pressure overload–induced cardiac hypertrophy and dysfunction in an RyR2-R176Q knockin mouse model. Hypertension. 2010; 55: 932–938.