Cardiac beta-Adrenergic Signaling: From Subcellular Microdomains to Heart Failure (original) (raw)
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American Journal of Physiology-Heart and Circulatory Physiology, 1999
Failing human myocardium has been associated with decreased sarcoplasmic reticulum (SR) Ca2+-ATPase activity. There remains controversy as to whether the regulation of SR Ca2+-ATPase activity is altered in heart failure or whether decreased SR Ca2+-ATPase activity is due to changes in SR Ca2+-ATPase or phospholamban expression. We therefore investigated whether alterations in cAMP-dependent phosphorylation of phospholamban may be responsible for the reduced SR Ca2+-ATPase activity in human heart failure. Protein levels of phospholamban and SR Ca2+-ATPase, detected by Western blot, were unchanged in failing compared with nonfailing human myocardium. There was decreased responsiveness to the direct activation of the SR Ca2+-ATPase activity by either cAMP (0.01–100 μmol/l) or protein kinase A (1–30 μg) in failing myocardium. Using the backphosphorylation technique, we observed a decrease of the cAMP-dependent phosphorylation level of phospholamban by 20 ± 2%. It is concluded that the i...
Cardiovascular Research, 2004
Decreased amplitude and slower kinetics of cardiomyocyte intracellular calcium (Ca 2 + i ) transients may underlie the diminished cardiac function observed in heart failure. These alterations occur in humans and animals with heart failure, including the TNF1.6 mouse model, in which heart failure arises from cardiac-specific overexpression of tumor necrosis factor a (TNFa). Objective: Since ablation of phospholamban expression (PLBKO) removes inhibition of the sarcoplasmic reticulum (SR) Ca 2 + pump, enhances SR Ca 2 + uptake and increases contractility, we assessed whether ablation of phospholamban expression could improve cardiac function, limit remodeling, and improve survival in the TNF1.6 model of heart failure. Methods: We bred PLBKO with TNF1.6 mice and characterized the progeny for survival, cardiac function (echocardiography), cardiac remodeling (hypertrophy, dilation, fibrosis), and Ca 2 + i transients and contractile function of isolated cardiomyocytes. Results: PLB ablation did not improve survival, cardiac function, or limit cardiac chamber dilation and hypertrophy in TNF1.6 mice (TKO mice). However, contractile function and Ca 2 + i transients (amplitude and kinetics) of isolated TKO cardiomyocytes were markedly enhanced. This discordance between unimproved cardiac function, and enhanced Ca 2 + i cycling and cardiomyocyte contractile parameters may arise from a continued overexpression of collagen and decreased expression of gap junction proteins (connexin 43) in response to chronic TNFa stimulation. Conclusions: Enhancement of intrinsic cardiomyocyte Ca 2 + i cycling and contractile function may not be sufficient to overcome several parallel pathophysiologic processes present in the failing heart.
Journal of Molecular and Cellular Cardiology, 2006
Reduced Ca 2+ release from the sarcoplasmic reticulum (SR) and a negative force-frequency relation characterize end-stage human heart failure. The MLP-/mouse with dilated cardiomyopathy is used as a model to explore novel therapeutic interventions but the alterations in Ca 2+ handling in MLP-/remain incompletely understood. We studied [Ca 2+ ] i in left ventricular myocytes from MLP-/and WT mice (3-4 months old; whole-cell voltage clamp, 30°C). At 1 Hz stimulation, the amplitude of [Ca 2+ ] i transients was similar. However, in contrast to WT, at higher frequencies the [Ca 2+ ] i transient amplitude declined in MLP-/and there was no increase in SR Ca 2+ content. Unexpectedly, the decline of [Ca 2+ ] i was faster in MLP-/than in WT (at 1 Hz, τ of 80 ± 9 vs. 174 ± 29 ms, P < 0.001) and the frequency-dependent acceleration of the decline was abolished suggesting an enhanced basal SERCA activity. Indeed, the Ca 2+ affinity of SR Ca 2+ uptake in homogenates was higher in MLP-/-, with the maximal uptake rate similar to WT. Phosphorylation of phospholamban in MLP-/was increased (2.3-fold at Ser 16 and 2.9-fold at the Thr 17 site, P < 0.001) with similar SERCA and total phospholamban protein levels. On increasing stimulation frequency to 4 Hz, WT, but not MLP-/-, myocytes had a net gain of Ca 2+ , suggesting inadequate Ca 2+ sequestration in MLP-/-. In conclusion, increased baseline phosphorylation of phospholamban in MLP-/leads to a reduced reserve for frequency-dependent increase of Ca 2+ release. This represents a novel paradigm for altered Ca 2+ handling in heart failure, underscoring the importance of phosphorylation pathways.
Circulation Research, 2004
Phosphoinositide 3-kinase (PI3K) has been implicated in  2-adrenergic receptor ( 2-AR)/G i-mediated compartmentation of the concurrent G scAMP signaling, negating  2-AR-induced phospholamban phosphorylation and the positive inotropic and lusitropic responses in cardiomyocytes. However, it is unclear whether PI3K crosstalks with the  1-AR signal transduction, and even more generally, with the cAMP/PKA pathway. In this study, we show that selective  1-AR stimulation markedly increases PI3K activity in adult rat cardiomyocytes. Inhibition of PI3K by LY294002 significantly enhances  1-AR-induced increases in L-type Ca 2ϩ currents, intracellular Ca 2ϩ transients, and myocyte contractility, without altering the receptor-mediated phosphorylation of phospholamban. The LY294002 potentiating effects are completely prevented by ARK-ct, a peptide inhibitor of -adrenergic receptor kinase-1 (ARK1) as well as G ␥ signaling, but not by disrupting G i function with pertussis toxin. Moreover, forskolin, an adenylyl cyclase activator, also elevates PI3K activity and inhibition of PI3K enhances forskolin-induced contractile response in a ARK-ct sensitive manner. In contrast, PI3K inhibition affects neither the basal contractility nor high extracellular Ca 2ϩ-induced increase in myocyte contraction. These results suggest that  1-AR stimulation activates PI3K via a PKA-dependent mechanism, and that G ␥ and the subsequent activation of ARK1 are critically involved in the PKA-induced PI3K signaling which, in turn, negates cAMP-induced positive inotropic effect via inhibiting sarcolemmal Ca 2ϩ influx and the subsequent increase in intracellular Ca 2ϩ transients, without altering the receptor-mediated phospholamban phosphorylation, in intact cardiomyocytes.
A-Kinase Anchoring Proteins in Cardiac Myocytes and Their Roles in Regulating Calcium Cycling
Cells
The rate of calcium cycling and calcium transient amplitude are critical determinants for the efficient contraction and relaxation of the heart. Calcium-handling proteins in the cardiac myocyte are altered in heart failure, and restoring the proper function of those proteins is an effective potential therapeutic strategy. The calcium-handling proteins or their regulators are phosphorylated by a cAMP-dependent kinase (PKA), and thereby their activity is regulated. A-Kinase Anchoring Proteins (AKAPs) play a seminal role in orchestrating PKA and cAMP regulators in calcium handling and contractile machinery. This cAMP/PKA orchestration is crucial for the increased force and rate of contraction and relaxation of the heart in response to fight-or-flight. Knockout models and the few available preclinical models proved that the efficient targeting of AKAPs offers potential therapies tailor-made for improving defective calcium cycling. In this review, we highlight important studies that iden...