Targeted Overexpression of the Sarcoplasmic Reticulum Ca ²⁺ -ATPase Increases Cardiac Contractility in Transgenic Mouse Hearts

Abstract

Abstract-Cardiac hypertrophy and heart failure are known to be associated with a reduction in Ca 2ϩ -ATPase pump levels of the sarcoplasmic reticulum (SR). To determine whether, and to what extent, alterations in Ca 2ϩ pump numbers can affect contraction and relaxation parameters of the heart, we have overexpressed the cardiac SR Ca 2ϩ -ATPase specifically in the mouse heart using the ␣-myosin heavy chain promoter. Analysis of 2 independent transgenic lines demonstrated that sarco(endo)plasmic reticulum Ca 2ϩ -ATPase isoform (SERCA2a) mRNA levels were increased 3.88Ϯ0.4-fold and 7.90Ϯ0.2-fold over those of the control mice. SERCA2a protein levels were increased by 1.31Ϯ0.05-fold and 1.54Ϯ0.05-fold in these lines despite high levels of mRNA, suggesting that complex regulatory mechanisms may determine the SERCA2a pump levels. The maximum velocity of Ca 2ϩ uptake (V max ) was increased by 37%, demonstrating that increased pump levels result in increased SR Ca 2ϩ uptake function. However, the apparent affinity of the SR Ca 2ϩ -ATPase for Ca 2ϩ remains unchanged in transgenic hearts. To evaluate the effects of overexpression of the SR Ca 2ϩ pump on cardiac contractility, we used the isolated perfused work-performing heart model. The transgenic hearts showed significantly higher myocardial contractile function, as indicated by increased maximal rates of pressure development for contraction (ϩdP/dt) and relaxation (-dP/dt), together with shortening of the normalized time to peak pressure and time to half relaxation. Measurements of intracellular free calcium concentration and contractile force in trabeculae revealed a doubling of Ca 2ϩ transient amplitude, with a concomitant boost in contractility. The present study demonstrates that increases in SERCA2a pump levels can directly enhance contractile function of the heart by increasing SR Ca 2ϩ transport. (Circ Res. 1998;83:1205-1214.)Key Words: sarcoplasmic reticulum Ca 2ϩ -ATPase Ⅲ transgenic mice Ⅲ Ca 2ϩ uptake Ⅲ working heart model T he sarcoplasmic reticulum (SR) plays a central role in the contraction-and-relaxation cycle of the heart by regulating intracellular calcium (Ca 2ϩ ) concentrations (reviewed in Reference 1). Ca 2ϩ release from the SR via the ryanodine receptor initiates muscle contraction, whereas Ca 2ϩ reuptake into the lumen of the SR leads to muscle relaxation. The Ca 2ϩ uptake function of the SR is driven by an ATP-dependent Ca 2ϩ transport pump, the sarco(endo)plasmic reticulum Ca 2ϩ -ATPase (SERCA). Molecular cloning analyses have identified a family of SERCA pumps encoded by 3 highly homologous genes (SERCA1, SERCA2, and SERCA3). [2][3][4][5][6][7][8] The SERCA2 gene encodes 2 isoforms, SERCA2a and SERCA2b, which differ at the COOH terminus as a result of alternative splicing (SERCA2a comprises 4 amino acids, and SERCA2b comprises 49 amino acids). [5][6][7] SERCA2a is the primary SERCA isoform expressed in the heart and is also present in slow-twitch skeletal muscle, smooth muscle, and fetal fast-twitch muscle. 9,10 In the rat heart, SERCA2a expr...

Adenoviral gene transfer of Akt enhances myocardial contractility and intracellular calcium handling

“…In fact, SERCA-2 overexpression is known to increase Ca 2+ transport and cardiac function, and is mainly characterized by increased systolic Ca 2+ levels, shorter transients, enhanced contractility with accelerated rates of contraction and relaxation, and no evidence of diastolic Ca 2+ overload consistent with the enhanced capacity of the SR to sequester calcium. 25,26 The mechanisms of SERCA-2 are likely complex, but do not involve significant changes of phospholamban, calsequestrin, and ryanodine receptor protein and transcripts levels, confirming previous data obtained in Akt-overexpressing mice. 11 However, the possibility of a change in the extent of phospholamban phosphorylation and its effect on SERCA-2 cannot be excluded.…”

Determinants of frequency-dependent contraction and relaxation of mammalian myocardium

“…A decrease in SERCA protein and PLB phosphorylation has been correlated with a negative force frequency relationship in human hearts [24]. The role of the SERCA pump has been studied by several laboratories both in genetically manipulated animal models [28][29][30] or gene transfer either in vivo or in cultured myocytes [21,[31][32][33], as well as in the context of human heart failure [34]. In general, a decrease in SERCA pump levels is correlated with a negative FFR.…”

Excessive Sarcoplasmic/Endoplasmic Reticulum Ca 2+ -ATPase Expression Causes Increased Sarcoplasmic Reticulum Ca 2+ Uptake but Decreases Myocyte Shortening

“…15 Myocytes were counted, and adenoviral transfection was performed at the indicated multiplicity of infection (MOI) while the cells were plated on laminin-coated 35-mm dishes at a density of 5ϫ10 5 rod-shaped myocytes/dish. This study was designed and performed in accordance with institutional guidelines for the care and use of animals.…”

“…Accordingly, transgenic mice overexpressing SERCA2a exhibited improved cardiac function and Ca 2ϩ handling. 4,5 In neonatal rat cardiomyocytes with normal and depressed SERCA2a expression, adenovirus-mediated overexpression of SERCA2a resulted in enhanced SR Ca 2ϩ uptake and accelerated decay of Ca 2ϩ transients. 6,7 Furthermore, catheter-based transfection with an adenovirus encoding SERCA2a restored cardiac function in rats in transition to heart failure 8 and improved survival.…”