Role of phosphorylation of Thr17 residue of phospholamban in mechanical recovery during hypercapnic acidosis

Objectives: To assess the time course of phosphorylation of phospholamban residues, the underlying mechanisms determining these phosphorylations, and their functional impact on the mechanical recovery during acidosis. Methods: Langendorff perfused rat hearts were Submitted to 30 min of hypercapnic acidosis. Contractility, relaxation, and phosphorylation of phospholamban residues, immunodetected by specific antibodies, were determined. Results: Acidosis produced a mechanical impairment followed by a spontaneous recovery, most of which occurred within the first 3 min of acidosis (early recovery), During this period, contractility and relaxation recovered by 67 +/- 9% and 77 +/- 11%, respectively, from its maximal depression, together with an increase in the Ca2+-calmodulin-dependent protein kinase II (CaMKII)-dependent phosphorylation of Thr(17). The CaMKII inhibitor KN-93, at 1, 5 and 10 mu M, decreased Thr(17) phosphorylation to basal levels and produced a similar impairment of the early relaxation recovery (50%). However, only 5 and 10 mu M KN-93 inhibited the early contractile recovery and completely blunted the late mechanical recovery. Inhibition of the reverse mode of the Na+/Ca2+ exchanger by KB-R7943 decreased Thr(17) phosphorylation but accelerated the early contractile recovery. Conclusions: CaMKII-dependent Thr(17) phosphorylation significantly increased at the beginning of acidosis, is responsible for 50% of the early relaxation recovery, and is linked to the activation of the reverse Na+/Ca2+ mode. The early contractile recovery and the late mechanical recovery are dependent on CaMKII but independent of the phosphorylation of the Thr(17) residue of phospholamban. The reverse Na+/Ca2+ mode has an additional negative effect that opposes the early mechanical recovery. (c) 2005 European Society of Cardiology. Published by Elsevier B.V. All rights reserved.