Published Online
on October 25, 2004

A more recent version of this article appeared on December 7, 2004

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Submitted on June 20, 2003
Revised on March 30, 2004
Accepted on April 4, 2004

Excessive Sarcoplasmic/Endoplasmic Reticulum Ca²⁺-ATPase Expression Causes Increased Sarcoplasmic Reticulum Ca²⁺ Uptake but Decreases Myocyte Shortening

Nils Teucher , Juergen Prestle PhD, Tim Seidler MD, Susan Currie PhD, Elspeth B. Elliott , Deborah F. Reynolds PhD, Peter Schott MD, Stefan Wagner , Harald Kogler MD, Giuseppe Inesi PhD, Donald M. Bers PhD, Gerd Hasenfuss MD^*, and Godfrey L. Smith PhD

From the Department of Cardiology and Pneumology, University of Göttingen, Göttingen, Germany (N.T., J.P., T.S., P.S., S.W., H.K., G.H.); Biochemistry and Molecular Biology, University of Maryland, Baltimore (G.I.); the Department of Physiology, Loyola University Chicago, Stritch School of Medicine, Maywood, Ill (D.M.B.); and the Institute of Biomedical and Life Sciences, University of Glasgow, Glasgow, UK (S.C., E.B.E., D.F.R., G.L.S.).

^* To whom correspondence should be addressed. E-mail: hasenfus{at}med.uni-goettingen.de.

Background--Increasing sarcoplasmic/endoplasmic reticulum (SR) Ca²⁺-ATPase (SERCA) uptake activity is a promising therapeutic approach for heart failure. We investigated the effects of different levels of SERCA1a expression on contractility and Ca²⁺ cycling. We tested whether increased SERCA1a expression levels enhance myocyte contractility in a gene-dose-dependent manner.

Methods and Results--Rabbit isolated cardiomyocytes were transfected at different multiplicities of infection (MOIs) with adenoviruses encoding SERCA1a (or -galactosidase as control). Myocyte relaxation half-time was decreased by 10% (P=0.052) at SERCA1a MOI 10 and by 28% at MOI 50 (P<0.05). Myocyte fractional shortening was increased by 12% at MOI 10 (P<0.05) but surprisingly decreased at MOI 50 (-22%, P<0.05) versus control. SR Ca²⁺ uptake (in permeabilized myocytes) demonstrated a gene-dose-dependent decrease in K_m by 29% and 46% and an increase in V_max by 37% and 72% at MOI 10 and MOI 50, respectively (all P<0.05 versus control). Ca²⁺ transient amplitude was increased in Ad-SERCA1a-infected myocytes at MOI 10 (by 121%, P<0.05), but at MOI 50, the Ca²⁺ transient amplitude was not significantly changed. Caffeine-induced Ca²⁺ transients indicated significantly increased SR Ca²⁺ content in Ad-SERCA1a-infected cells, by 72% at MOI 10 and by 87% at MOI 50. Mathematical simulations demonstrate that the functional increase in SR Ca²⁺-ATPase uptake activity at MOI 50 (and increased cytosolic Ca²⁺ buffering) is sufficient to curtail the Ca²⁺ transient amplitude and explain the reduced contraction.

Conclusions--Moderate SERCA1a gene transfer and expression improve contractility and Ca²⁺ cycling. However, higher SERCA1a expression levels can impair myocyte shortening because of higher SERCA activity and Ca²⁺ buffering.

Key words: heart failure • gene therapy • calcium • sarcoplasmic reticulum

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