Myocardial stunning describes the reversible decrease in myocardial contractility that follows a brief ischemic insult clinically manifested as sluggish recovery of the pump function after revascularization. Experimental evidence indicates that sarcoplasmic reticulum (SR) function is altered in the myocardial stunning. A decrease in the activity of the SR Ca2+ pump (SERCA2), and/or in the rate of Ca2+ reuptake by the SR have been described in several species, including patients submitted to moderate and reversible injury during cardiac surgery. The function of SERCA2 is regulated by the state of phosphorylation of another SR protein, named phospholamban (PLB). In the dephosphorylated state, PLB inhibits the activity of SERCA2 and the SR Ca2+ transport. Phosphorylation of PLB at Ser16 and Thr17 residues removes its inhibitory effect on SERCA2, thereby accelerating Ca2+ uptake by the SR and increasing the Ca2+ available for being released. The parent grant focuses on determining the functional role of PLB and on discriminating the significance of dual site PLB phosphorylation under physiological conditions, like beta-adrenergic stimulation. The main goal of the present collaboration constitutes an extension of the aims of the parent project to cardiac disease. Although there is evidence showing that both PLB residues become phosphorylated during ischemia and reperfusion, the functional significance of each PLB phosphorylation site in the ischemia-reperfused hearts is not known. This application proposes the study of the role of site-specific PLB phosphorylation on the recovery of cardiac contractility and relaxation in the stunned heart, and on the mechanisms by which brief ischemic insults protect from the cardiac injury (preconditioning), as well as the determination of the signaling cascade involved in these phosphorylations. To that purpose, transgenic models with specific mutation of a single site or simultaneous mutation of both phosphorylatable PLB sites, developed during the course of the parent grant, in combination with biochemical determination (immunodetection of PLB phosphorylation sites, kinase and phosphatase activities) and mechanical measurements of the cardiac activity, will be used. This experimental strategy will allow us to define the significance of PLB phosphorylation and to discriminate the role of each PLB phosphorylation site during stunning and preconditioning, which may lead to novel therapeutic approaches. This research will be done primarily in Argentina as an extension of NIH # HL26057.
| Valverde, Carlos A; Mundina-Weilenmann, Cecilia; Reyes, Mariano et al. (2006) Phospholamban phosphorylation sites enhance the recovery of intracellular Ca2+ after perfusion arrest in isolated, perfused mouse heart. Cardiovasc Res 70:335-45 |
| Mattiazzi, A; Mundina-Weilenmann, C; Vittone, L et al. (2006) The importance of the Thr17 residue of phospholamban as a phosphorylation site under physiological and pathological conditions. Braz J Med Biol Res 39:563-72 |
| Said, M; Vittone, L; Mundina-Weilenmann, C et al. (2003) Role of dual-site phospholamban phosphorylation in the stunned heart: insights from phospholamban site-specific mutants. Am J Physiol Heart Circ Physiol 285:H1198-205 |
