title: Adenoviral SERCA1a gene transfer to adult rat ventricular myocytes induces physiological changes in calcium handling.
abstract: OBJECTIVE
  We examined the functional consequences of expressing adult rabbit fast skeletal sarcoplasmic reticulum (SR) Ca(2+)-ATPase (SERCA1a) in isolated adult rat ventricular myocytes.
  
  
  METHODS
  Myocytes were infected with a recombinant adenovirus harboring SERCA1a. Then 2 days after myocyte infection, protein expression was estimated using Western blot and SDS-PAGE analysis. We also measured the ATP-dependent oxalate-facilitated Ca(2+) uptake of myocyte homogenates and monitored Ca(2+) transient in myocytes loaded with the Ca(2+) dye, indo-1.
  
  
  RESULTS
  SERCA1a gene expression resulted in a 36% increase in the total SERCA protein level in infected myocytes compared to controls (P<0.01), while SERCA2 and phospholamban levels did not change. This increase was associated with a 42% rise in SR Ca(2+) uptake (P<0.01), while tau (the time constant of Ca(2+) transient decay), and the time to peak fell by 32% (P<0.01) and 38% (P<0.001), respectively. Increasing the frequency of stimulation from 0.2 to 2 Hz decreased tau in both cell types (P<0.01). However, the decrease was much smaller in infected (P<0.01) than in uninfected cells (P<0.001). Isoproterenol (1 microM) further decreased tau in infected myocytes by 23% (P<0.05). In these cells, the diastolic [Ca(2+)](i) decreased by 50% (P<0.05) while the systolic [Ca(2+)](i) increased by 19% (P<0.05). No difference was found in the speed of SR Ca(2+) reloading after caffeine washout between the two cell types.
  
  
  CONCLUSIONS
  Adenovirus-mediated SERCA1a gene transfer to adult rat ventricular myocytes enhances SR Ca(2+) handling to a degree similar to that observed following physiological stimulation.
authors.affiliation: INSERM U 460, Faculté de Médecine Xavier Bichat, 16 rue Henri Huchard, 75018, Paris, France.
authors.forename: N
authors.initials: N
authors.lastname: Chossat
authors.forename: F
authors.initials: F
authors.lastname: Griscelli
authors.forename: P
authors.initials: P
authors.lastname: Jourdon
authors.forename: D
authors.initials: D
authors.lastname: Logeart
authors.forename: T
authors.initials: T
authors.lastname: Ragot
authors.forename: M
authors.initials: M
authors.lastname: Heimburger
authors.forename: M
authors.initials: M
authors.lastname: Perricaudet
authors.forename: A
authors.initials: A
authors.lastname: Lompré
authors.forename: S
authors.initials: S
authors.lastname: Hatem
authors.forename: J
authors.initials: J
authors.lastname: Mercadier
mesh_terms: D000256:Adenoviridae / Q000235:genetics*; D000704:Analysis of Variance; D000818:Animals; D015153:Blotting, Western; D002118:Calcium / Q000378:metabolism*; D000252:Calcium-Transporting ATPases / Q000032:analysis / Q000235:genetics* / Q000378:metabolism; D002478:Cells, Cultured; D018014:Gene Transfer Techniques; D005822:Genetic Vectors / Q000008:administration & dosage*; D008297:Male; D018613:Microscopy, Confocal; D009206:Myocardium / Q000201:enzymology / Q000378:metabolism*; D051381:Rats; D017208:Rats, Wistar; D012519:Sarcoplasmic Reticulum / Q000201:enzymology*; D053498:Sarcoplasmic Reticulum Calcium-Transporting ATPases
chemical_list: C506948:Atp2a2 protein, rat; D053498:Sarcoplasmic Reticulum Calcium-Transporting ATPases; D000252:Calcium-Transporting ATPases; D002118:Calcium