title: Regulation of in vivo cardiac contractility by phospholemman: role of Na+/Ca2+ exchange.
abstract: Phospholemman (PLM), when phosphorylated at serine 68, relieves its inhibition on Na(+)-K(+)-ATPase but inhibits Na(+)/Ca(2+) exchanger 1 (NCX1) in cardiac myocytes. Under stress when catecholamine levels are high, enhanced Na(+)-K(+)-ATPase activity by phosphorylated PLM attenuates intracellular Na(+) concentration ([Na(+)](i)) overload. To evaluate the effects of PLM on NCX1 on in vivo cardiac contractility, we injected recombinant adeno-associated virus (serotype 9) expressing either the phosphomimetic PLM S68E mutant or green fluorescent protein (GFP) directly into left ventricles (LVs) of PLM-knockout (KO) mice. Five weeks after virus injection, ∼40% of isolated LV myocytes exhibited GFP fluorescence. Expression of S68E mutant was confirmed with PLM antibody. There were no differences in protein levels of α(1)- and α(2)-subunits of Na(+)-K(+)-ATPase, NCX1, and sarco(endo)plasmic reticulum Ca(2+)-ATPase between KO-GFP and KO-S68E LV homogenates. Compared with KO-GFP myocytes, Na(+)/Ca(2+) exchange current was suppressed, but resting [Na(+)](i), Na(+)-K(+)-ATPase current, and action potential amplitudes were similar in KO-S68E myocytes. Resting membrane potential was slightly lower and action potential duration at 90% repolarization (APD(90)) was shortened in KO-S68E myocytes. Isoproterenol (Iso; 1 μM) increased APD(90) in both groups of myocytes. After Iso, [Na(+)](i) increased monotonically in paced (2 Hz) KO-GFP but reached a plateau in KO-S68E myocytes. Both systolic and diastolic [Ca(2+)](i) were higher in Iso-stimulated KO-S68E myocytes paced at 2 Hz. Echocardiography demonstrated similar resting heart rate, ejection fraction, and LV mass between KO-GFP and KO-S68E mice. In vivo closed-chest catheterization demonstrated enhanced contractility in KO-S68E compared with KO-GFP hearts stimulated with Iso. We conclude that under catecholamine stress when [Na(+)](i) is high, PLM minimizes [Na(+)](i) overload by relieving its inhibition of Na(+)-K(+)-ATPase and preserves inotropy by simultaneously inhibiting Na(+)/Ca(2+) exchanger.
authors.affiliation: Division of Nephrology and Center of Translational Medicine, Department of Medicine, Jefferson Medical College, Thomas Jefferson University, Philadelphia, Pennsylvania, USA.
authors.forename: Jufang
authors.initials: J
authors.lastname: Wang
authors.forename: Erhe
authors.initials: E
authors.lastname: Gao
authors.forename: Joseph
authors.initials: J
authors.lastname: Rabinowitz
authors.forename: Jianliang
authors.initials: J
authors.lastname: Song
authors.forename: Xue-Qian
authors.initials: XQ
authors.lastname: Zhang
authors.forename: Walter J
authors.initials: WJ
authors.lastname: Koch
authors.forename: Amy L
authors.initials: AL
authors.lastname: Tucker
authors.forename: Tung O
authors.initials: TO
authors.lastname: Chan
authors.forename: Arthur M
authors.initials: AM
authors.lastname: Feldman
authors.forename: Joseph Y
authors.initials: JY
authors.lastname: Cheung
mesh_terms: D000818:Animals; D002316:Cardiotonic Agents / Q000494:pharmacology; D002478:Cells, Cultured; D006321:Heart / Q000187:drug effects / Q000502:physiology; D007545:Isoproterenol / Q000494:pharmacology; D008564:Membrane Potentials / Q000187:drug effects / Q000502:physiology; D008565:Membrane Proteins / Q000378:metabolism / Q000502:physiology*; D051379:Mice; D008810:Mice, Inbred C57BL; D018345:Mice, Knockout; D009200:Myocardial Contraction / Q000187:drug effects / Q000502:physiology*; D032383:Myocytes, Cardiac / Q000187:drug effects / Q000502:physiology; D010750:Phosphoproteins / Q000378:metabolism / Q000502:physiology*; D053498:Sarcoplasmic Reticulum Calcium-Transporting ATPases / Q000378:metabolism / Q000502:physiology; D019831:Sodium-Calcium Exchanger / Q000378:metabolism / Q000502:physiology*; D000254:Sodium-Potassium-Exchanging ATPase / Q000378:metabolism / Q000502:physiology
chemical_list: D002316:Cardiotonic Agents; D008565:Membrane Proteins; C499849:NCX1 protein, mouse; D010750:Phosphoproteins; D019831:Sodium-Calcium Exchanger; C069141:phospholemman; D053498:Sarcoplasmic Reticulum Calcium-Transporting ATPases; D000254:Sodium-Potassium-Exchanging ATPase; D007545:Isoproterenol