Chronic SR Ca 2+ -ATPase Inhibition Causes Adaptive Changes in Cellular Ca 2+ Transport

Abstract

Phospholamban, the critical regulator of the cardiac SERCA2a Ca 2+ affinity, is phosphorylated at Ser 16 and Thr 17 during β-adrenergic stimulation (eg, isoproterenol). To assess the impact of nonphosphorylatable phospholamban, a S16A, T17A double-mutant (DM) was introduced into phospholamban knockout mouse hearts. Transgenic lines expressing DM phospholamban at levels similar to wild types (WT) were identified. In vitro phosphorylation confirmed that DM phospholamban could not be phosphorylated, but produced the same shift in EC 50 of SERCA2a for Ca 2+ as unphosphorylated WT phospholamban. Rates of basal twitch [Ca 2+ ] i decline were not different in DM versus WT cardiomyocytes. Isoproterenol increased the rates of twitch [Ca 2+ ] i decline in WT, but not DM myocytes, confirming the prominent role of phospholamban phosphorylation in this response. Increased L-type Ca 2+ current ( I Ca ) density, with unaltered characteristics, was the major compensation in DM myocytes. Consequently, the normal β-adrenergic–induced increase in I Ca caused larger dynamic changes in absolute I Ca density. Isoproterenol increased Ca 2+ transients to a comparable amplitude in DM and WT. There were no changes in myofilament Ca 2+ sensitivity, or the expression levels and Ca 2+ removal activities of other Ca 2+ -handling proteins. Nor was there evidence of cardiac remodeling up to 10 months of age. Thus, chronic inhibition of SERCA2a by ablation of phospholamban phosphorylation (abolishing its adrenergic regulation) results in a unique cellular adaptation involving greater dynamic I Ca modulation. This I Ca modulation may partly compensate for the loss in SERCA2a responsiveness and thereby partially normalize β-adrenergic inotropy in DM phospholamban mice.