Satyam Sarma

Calmodulin kinase II is required for fight or flight sinoatrial node physiology

The best understood “fight or flight” mechanism for increasing heart rate (HR) involves activation of a cyclic nucleotide-gated ion channel (HCN4) by β-adrenergic receptor (βAR) agonist stimulation. HCN4 conducts an inward “pacemaker” current (If) that increases the sinoatrial nodal (SAN) cell membrane diastolic depolarization rate (DDR), leading to faster SAN action potential generation. Surprisingly, HCN4 knockout mice were recently shown to retain physiological HR increases with isoproterenol (ISO), suggesting that other If-independent pathways are critical to SAN fight or flight responses. The multifunctional Ca2+ and calmodulin-dependent protein kinase II (CaMKII) is a downstream signal in the βAR pathway that activates Ca2+ homeostatic proteins in ventricular myocardium. Mice with genetic, myocardial and SAN cell CaMKII inhibition have significantly slower HRs than controls during stress, leading us to hypothesize that CaMKII actions on SAN Ca2+ homeostasis are critical for βAR agonist responses in SAN. Here we show that CaMKII mediates ISO HR increases by targeting SAN cell Ca2+ homeostasis. CaMKII inhibition prevents ISO effects on SAN Ca2+ uptake and release from intracellular sarcoplasmic reticulum (SR) stores that are necessary for increasing DDR. CaMKII inhibition has no effect on the ISO response in SAN cells when SR Ca2+ release is disabled and CaMKII inhibition is only effective at slowing HRs during βAR stimulation. These studies show the tightly coupled, but previously unanticipated, relationship of CaMKII to the βAR pathway in fight or flight physiology and establish CaMKII as a critical signaling molecule for physiological HR responses to catecholamines.

Association between diabetes mellitus and post-discharge outcomes in patients hospitalized with heart failure: findings from the EVEREST trial.

We evaluated the impact of diabetes mellitus (DM) and diabetic therapy on outcomes in patients with reduced ejection fraction (EF) after hospitalization for heart failure (HF). DM is prevalent in patients hospitalized with HF, yet inconclusive data exist on the post‐discharge outcomes of this patient population.

Targeting myocardial substrate metabolism in heart failure: potential for new therapies

The incidence and prevalence of heart failure have increased significantly over the past few decades. Available data suggest that patients with heart failure independent of the aetiology have viable but dysfunctional myocardium that is potentially salvageable. Although a great deal of research effort has focused on characterizing the molecular basis of heart failure, cardiac metabolism in this disorder remains an understudied discipline. It is known that many aspects of cardiomyocyte energetics are altered in heart failure. These include a shift from fatty acid to glucose as a preferred substrate and a decline in the levels of ATP. Despite these demonstrated changes, there are currently no approved drugs that target metabolic enzymes or proteins in heart failure. This is partly due to our limited knowledge of the mechanisms and pathways that regulate cardiac metabolism. Better characterization of these pathways may potentially lead to new therapies for heart failure. Targeting myocardial energetics in the viable and potentially salvageable tissue may be particularly effective in the treatment of heart failure. Here, we will review metabolic changes that occur in fatty acid and glucose metabolism and AMP‐activated kinase in heart failure. We propose that cardiac energetics should be considered as a potential target for therapy in heart failure and more research should be done in this area.

PKC inhibition ameliorates the cardiac phenotype in a mouse model of myotonic dystrophy type 1

Cardiac complications are a common cause of death in individuals with the inherited multisystemic disease myotonic dystrophy type 1 (DM1). A characteristic molecular feature of DM1 is misregulated alternative splicing due to disrupted functioning of the splicing regulators muscleblind-like 1 (MBNL1) and CUG-binding protein 1 (CUGBP1). CUGBP1 is upregulated in DM1 due to PKC pathway activation and subsequent CUGBP1 protein hyperphosphorylation and stabilization. Here, we blocked PKC activity in a heart-specific DM1 mouse model to determine its pathogenic role in DM1. Animals given PKC inhibitors exhibited substantially increased survival that correlated with reduced phosphorylation and decreased steady-state levels of CUGBP1. Functional studies demonstrated that PKC inhibition ameliorated the cardiac conduction defects and contraction abnormalities found in this mouse model. The inhibitor also reduced misregulation of splicing events regulated by CUGBP1 but not those regulated by MBNL1, suggesting distinct roles for these proteins in DM1 cardiac pathogenesis. The PKC inhibitor did not reduce mortality in transgenic mice with heart-specific CUGBP1 upregulation, indicating that PKC inhibition did not have a general protective effect on PKC-independent CUGBP1 increase. Our results suggest that pharmacological blockade of PKC activity mitigates the DM1 cardiac phenotype and provide strong evidence for a role for the PKC pathway in DM1 pathogenesis.

Genetic inhibition of PKA phosphorylation of RyR2 prevents dystrophic cardiomyopathy.

Aberrant intracellular Ca2+ regulation is believed to contribute to the development of cardiomyopathy in Duchenne muscular dystrophy. Here, we tested whether inhibition of protein kinase A (PKA) phosphorylation of ryanodine receptor type 2 (RyR2) prevents dystrophic cardiomyopathy by reducing SR Ca2+ leak in the mdx mouse model of Duchenne muscular dystrophy. mdx mice were crossed with RyR2-S2808A mice, in which PKA phosphorylation site S2808 on RyR2 is inactivated by alanine substitution. Compared with mdx mice that developed age-dependent heart failure, mdx-S2808A mice exhibited improved fractional shortening and reduced cardiac dilation. Whereas application of isoproterenol severely depressed cardiac contractility and caused 95% mortality in mdx mice, contractility was preserved with only 19% mortality in mdx-S2808A mice. SR Ca2+ leak was greater in ventricular myocytes from mdx than mdx-S2808A mice. Myocytes from mdx mice had a higher incidence of isoproterenol-induced diastolic Ca2+ release events than myocytes from mdx-S2808A mice. Thus, inhibition of PKA phosphorylation of RyR2 reduced SR Ca2+ leak and attenuated cardiomyopathy in mdx mice, suggesting that enhanced PKA phosphorylation of RyR2 at S2808 contributes to abnormal Ca2+ homeostasis associated with dystrophic cardiomyopathy.

CaMKIIδ mediates β-adrenergic effects on RyR2 phosphorylation and SR Ca2 + leak and the pathophysiological response to chronic β-adrenergic stimulation

Chronic activation of Ca(2+)/calmodulin-dependent protein kinase II (CaMKII) has been implicated in the deleterious effects of β-adrenergic receptor (β-AR) signaling on the heart, in part, by enhancing RyR2-mediated sarcoplasmic reticulum (SR) Ca(2+) leak. We used CaMKIIδ knockout (CaMKIIδ-KO) mice and knock-in mice with an inactivated CaMKII site S2814 on the ryanodine receptor type 2 (S2814A) to investigate the involvement of these processes in β-AR signaling and cardiac remodeling. Langendorff-perfused hearts from CaMKIIδ-KO mice showed inotropic and chronotropic responses to isoproterenol (ISO) that were similar to those of wild type (WT) mice; however, in CaMKIIδ-KO mice, CaMKII phosphorylation of phospholamban and RyR2 was decreased and isolated myocytes from CaMKIIδ-KO mice had reduced SR Ca(2+) leak in response to isoproterenol (ISO). Chronic catecholamine stress with ISO induced comparable increases in relative heart weight and other measures of hypertrophy from day 9 through week 4 in WT and CaMKIIδ-KO mice, but the development of cardiac fibrosis was prevented in CaMKIIδ-KO animals. A 4-week challenge with ISO resulted in reduced cardiac function and pulmonary congestion in WT, but not in CaMKIIδ-KO or S2814A mice, implicating CaMKIIδ-dependent phosphorylation of RyR2-S2814 in the cardiomyopathy, independent of hypertrophy, induced by prolonged β-AR stimulation.

Effect of gravity and microgravity on intracranial pressure

Astronauts have recently been discovered to have impaired vision, with a presentation that resembles syndromes of elevated intracranial pressure on Earth. Gravity has a profound effect on fluid distribution and pressure within the human circulation. In contrast to prevailing theory, we observed that microgravity reduces central venous and intracranial pressure. This being said, intracranial pressure is not reduced to the levels observed in the 90 deg seated upright posture on Earth. Thus, over 24 h in zero gravity, pressure in the brain is slightly above that observed on Earth, which may explain remodelling of the eye in astronauts.

Enhancing the metabolic substrate: PPAR-alpha agonists in heart failure

The prognosis for patients diagnosed with heart failure has significantly improved over the past three decades; however, the disease still confers a high degree of morbidity and mortality. Current treatments for chronic heart failure have focused primarily on blocking neurohormonal signaling and optimizing hemodynamic parameters. Although significant resources have been devoted toward the development of new pharmaceutical therapies for heart failure, few new drugs have been designed to target myocardial metabolic pathways despite growing evidence that on a fundamental level chronic heart failure can be characterized as an imbalance between myocardial energy demand and supply. Disruptions in myocardial energy pathways are evident as the myocardium is unable to generate sufficient amounts of ATP with advancing stages of heart failure. Down-regulation of fatty acid oxidation likely contributes to the phenotype of the “energy starved” heart. Fibrates are small molecule agonists of PPARα pathways that have been used to treat dyslipidemia. Although never used therapeutically in clinical heart failure, PPARα agonists have been shown to enhance fatty acid oxidation, improve endothelial cell function, and decrease myocardial fibrosis and hypertrophy in animal models of heart failure. In light of their excellent clinical safety profile, PPARα agonists may improve outcomes in patients suffering from systolic heart failure by augmenting myocardial ATP production in addition to targeting maladaptive hypertrophic pathways.

Sudden Infant Death Syndrome in Mice with an Inherited Mutation in RyR2

Background—Mutations in the cardiac ryanodine receptor gene (RyR2) have been recently identified in victims of sudden infant death syndrome. The aim of this study was to determine whether a gain-of-function mutation in RyR2 increases the propensity to cardiac arrhythmias and sudden death in young mice. Methods and Results—Incidence of sudden death was monitored prospectively in heterozygous knock-in mice with mutation R176Q in RyR2 (R176Q/+). Young R176Q/+ mice exhibited a higher incidence of sudden death compared with wild-type littermates. Optical mapping of membrane potentials and intracellular calcium in 1- to 7-day-old R176Q/+ and wild-type mice revealed an increased incidence of ventricular ectopy and spontaneous calcium releases in neonatal R176Q/+ mice. Surface ECGs in 3- to 10-day-old mice showed that R176Q/+ mice developed more ventricular arrhythmias after provocation with epinephrine and caffeine. Intracardiac pacing studies in 12- to 18-day-old mice revealed the presence of an arrhythmogenic substrate in R176Q/+ compared with wild-type mice. Reverse transcription–polymerase chain reaction and Western blotting showed that expression levels of other calcium handling proteins were unaltered, suggesting that calcium leak through mutant RyR2 underlies arrhythmogenesis and sudden death in young R176Q/+ mice. Conclusions—Our findings demonstrate that a gain-of-function mutation in RyR2 confers an increased risk of cardiac arrhythmias and sudden death in young mice and that young R176Q/+ mice may be used as a model for elucidating the complex interplay between genetic and environmental risk factors associated with sudden infant death syndrome.

Serum Neuregulin-1β as a Biomarker of Cardiovascular Fitness

PURPOSE: Neuregulins (NRG) are growth factors that bind to receptors of the erbB family, and are known to mediate a number of processes involved in diverse tissues. Neuregulin-1beta is expressed in skeletal muscle and is activated by exercise. We hypothesized that NRG-1beta might circulate in the bloodstream and increase as a consequence of physical activity. A study was conducted in healthy subjects to determine if NRG-1beta is immunodetectable in human serum, and if so whether levels relate acutely or chronically to exercise. METHODS: Nine healthy men underwent three bouts of exercise of varying degrees of intensity on a bicycle ergometer over a period of three weeks. Cardio-respiratory fitness was determined by measurement of maximal oxygen uptake (VO(2)max). Serum was sampled prior to and immediately after each session (up to 30 minutes post) and serum NRG-1beta was quantified utilizing an indirect sandwich ELISA assay developed in our lab. RESULTS: Across subjects, mean serum NRG-1beta levels ranged from 32 ng/mL to 473 ng/mL. Individual subjects showed relatively stable levels during the study period that did not change acutely after exercise. Serum NRG-1beta demonstrated a positive correlation with VO(2)max (r2=0.49, p =.044). CONCLUSIONS: These preliminary observations suggest that at least in healthy men, serum NRG-1beta is an indicator of cardio-respiratory fitness and does not change acutely with exercise.