Overexpression of the Sarcolemmal Calcium Pump in the Myocardium of Transgenic Rats

From the Department of Medicine, University of Würzburg (A.H., S.O.-M., T.R., K.N., K.S., C.B., L.N.); Department of Physiology, University of Bonn (F.G., K.W.L., R.M.); Department of Medicine, Cardiology and Angiology, Medical Faculty, Mannheim, University of Heidelberg (K.H., H.H., P.G., G.E.); Max Delbrück Center for Molecular Medicine, Berlin (S.H., U.G., R.V.); and the Institute of Physiology, University of Leipzig (H.G.Z.), Germany. Present affiliation for A.H. is Developmental Genetics, Max Delbrueck Center for Molecular Medicine, Berlin, Germany. Present affiliation for R.V. is Institute of Clinical Pharmacology and Toxicology, Benjamin Franklin Medical Center, Berlin, Germany. Present affiliation for C.B. is Department of Anesthesiology, University of Würzburg, Germany.

Correspondence to Ludwig Neyses, MD, Department of Medicine, University of Würzburg, Josef-Schneider-Str 2, D-97080 Würzburg, Germany.

Abstract—The plasma membrane calmodulin–dependent calcium ATPase (PMCA) is a calcium-extruding enzyme controlling Ca²⁺ homeostasis in nonexcitable cells. However, its function in the myocardium is unclear because of the presence of the Na⁺/Ca²⁺ exchanger. We approached the question of the physiological function of the calcium pump using a transgenic "gain of function" model. Transgenic rat lines carrying the human PMCA 4 cDNA under control of the ventricle-specific myosin light chain-2 promoter were established, and expression in the myocardium was ascertained at the mRNA, protein, and functional levels. In vivo hemodynamic measurements in adult homozygous animals showed no differences in baseline and increased cardiac performance recruited by volume overload compared with controls. No differences between transgenic and control cardiomyocytes were found in patch clamp voltage dependence, activation/inactivation behavior of the L-type Ca²⁺ current, or fast [Ca²⁺]_i transients (assessed by the Fura-2 method). To test whether the PMCA might be involved in processes other than beat-to-beat regulation of contraction/relaxation, we compared growth processes of neonatal transgenic and control cardiomyocytes. A 1.6- and 2.3-fold higher synthesis rate of total protein was seen in cells from transgenic animals compared with controls on incubation with 2% FCS for 24 hours and 36 hours, respectively. An effect of similar magnitude was observed using 20 µmol/L phenylephrine. A 1.4-fold– and 2.0-fold–higher protein synthesis peak was seen in PMCA-overexpressing cardiomyocytes after stimulation with isoproterenol for 12 hours and 24 hours, respectively. Because pivotal parts of the - and ß-adrenergic signal transduction pathways recently have been localized to caveolae, we tested the hypothesis that the PMCA might alter the amplitude of - and ß-adrenergic growth signals by virtue of its localization in caveolae. Biochemical as well as immunocytochemical studies suggested that the PMCA in large part was colocalized with caveolin 3 in caveolae of cardiomyocytes. These results indicate that the sarcolemmal Ca²⁺-pump has little relevance for beat-to-beat regulation of contraction/relaxation in adult animals but likely plays a role in regulating myocardial growth, possibly through modulation of caveolar signal transduction.

Key Words: plasma membrane Ca²⁺-ATPase • myocardium • transgenic rat • contraction • cardiac growth

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