title: Phospholamban phosphorylation sites enhance the recovery of intracellular Ca2+ after perfusion arrest in isolated, perfused mouse heart.
abstract: OBJECTIVE
  To investigate the importance of the phosphorylation of Ser16 and Thr17 sites of phospholamban (PLN) on intracellular Ca2+ (Cai2+) handling and contractile recovery of the stunned myocardium.
  
  
  METHODS
  Cai2+ (Rhod-2, pulsed local-field fluorescence microscopy) and contractility (isovolumic left ventricular developed pressure, LVDP) were simultaneously measured in Langendorff perfused hearts from transgenic mice expressing either intact PLN (PLN-WT) or PLN with both phosphorylation sites mutated to Ala (PLN-DM), subjected to 12 min of global ischemia followed by a reperfusion period of 30 min.
  
  
  RESULTS
  Pre-ischemic values of Cai2+ and LVDP were similar in both groups. In PLN-WT, a transient increase in Thr17 phosphorylation at early reperfusion preceded a recovery of Ca2+ transient amplitude, virtually completed by the end of reperfusion. LVDP at 30 min reperfusion was 67.9+/-7.6% of pre-ischemic values, n=14. In contrast, in PLN-DM, there was a poor recovery of Cai2+ transient amplitude and LVDP was significantly lower (28.3+/-6.7%, n=11, 30 min reperfusion) than in PLN-WT hearts. Although myofilament Ca2+ responsiveness and troponin I (TnI) degradation did not differ between groups, the episodes of mechanical alternans, typical of Cai2+ overload, were significantly prolonged in PLN-DM vs. PLN-WT hearts.
  
  
  CONCLUSIONS
  PLN phosphorylation appears to be crucial for the mechanical and Cai2+ recovery during stunning and protective against the mechanical abnormalities typical of Cai2+ overload. The importance of PLN phosphorylation would primarily reside in the Thr17 residue, which is phosphorylated during the critical early phase of reperfusion. Our results emphasize that, although ablation of PLN phosphorylation does not affect basal contractility, it does alter Ca2+ handling and mechanical performance under stress situations.
authors.affiliation: Centro de Investigaciones Cardiovasculares, Facultad de Ciencias Médicas, 60 y 120, (1900) La Plata, Argentina. valverdeca@atlas.med.unlp.edu.ar
authors.forename: Carlos A
authors.initials: CA
authors.lastname: Valverde
authors.forename: Cecilia
authors.initials: C
authors.lastname: Mundiña-Weilenmann
authors.forename: Mariano
authors.initials: M
authors.lastname: Reyes
authors.forename: Evangelia G
authors.initials: EG
authors.lastname: Kranias
authors.forename: Ariel L
authors.initials: AL
authors.lastname: Escobar
authors.forename: Alicia
authors.initials: A
authors.lastname: Mattiazzi
mesh_terms: D000818:Animals; D015153:Blotting, Western; D002118:Calcium / Q000378:metabolism*; D002135:Calcium-Binding Proteins / Q000235:genetics / Q000378:metabolism*; D004591:Electrophoresis, Polyacrylamide Gel; D005456:Fluorescent Dyes; D007424:Intracellular Fluid / Q000378:metabolism*; D051379:Mice; D008822:Mice, Transgenic; D009154:Mutation; D009200:Myocardial Contraction; D009206:Myocardium / Q000378:metabolism*; D010477:Perfusion; D010766:Phosphorylation; D000099211:Phospholamban
chemical_list: D002118:Calcium; D002135:Calcium-Binding Proteins; D005456:Fluorescent Dyes; D000099211:Phospholamban