Altered contractility and [Ca2+]ihomeostasis in phospholemman-deficient murine myocytes: role of Na+/Ca2+exchange

Abstract

Phospholemman (PLM) regulates contractility and Ca2+homeostasis in cardiac myocytes. We characterized excitation-contraction coupling in myocytes isolated from PLM-deficient mice backbred to a pure congenic C57BL/6 background. Cell length, cell width, and whole cell capacitance were not different between wild-type and PLM-null myocytes. Compared with wild-type myocytes, Western blots indicated total absence of PLM but no changes in Na+/Ca2+exchanger, sarcoplasmic reticulum (SR) Ca2+-ATPase, α1-subunit of Na+-K+-ATPase, and calsequestrin levels in PLM-null myocytes. At 5 mM extracellular Ca2+concentration ([Ca2+]o), contraction and cytosolic [Ca2+] ([Ca2+]i) transient amplitudes and SR Ca2+contents in PLM-null myocytes were significantly ( P < 0.0004) higher than wild-type myocytes, whereas the converse was true at 0.6 mM [Ca2+]o. This pattern of contractile and [Ca2+]itransient abnormalities in PLM-null myocytes mimics that observed in adult rat myocytes overexpressing the cardiac Na+/Ca2+exchanger. Indeed, we have previously reported that Na+/Ca2+exchange currents were higher in PLM-null myocytes. Activation of protein kinase A resulted in increased inotropy such that there were no longer any contractility differences between the stimulated wild-type and PLM-null myocytes. Protein kinase C stimulation resulted in decreased contractility in both wild-type and PLM-null myocytes. Resting membrane potential and action potential amplitudes were similar, but action potential duration was much prolonged ( P < 0.04) in PLM-null myocytes. Whole cell Ca2+current densities were similar between wild-type and PLM-null myocytes, as were the fast- and slow-inactivation time constants. We conclude that a major function of PLM is regulation of cardiac contractility and Ca2+fluxes, likely by modulating Na+/Ca2+exchange activity.