Diminished basal phosphorylation level of phospholamban in the postinfarction remodeled rat ventricle: role of beta-adrenergic pathway, G(i) protein, phosphodiesterase, and phosphatases.
Three weeks after myocardial infarction (MI) in the rat, remodeled hypertrophy of noninfarcted myocardium is at its maximum and the heart is in a compensated stage with no evidence of heart failure. Our hemodynamic measurements at this stage showed a slight but insignificant decrease of +dP/dt but a significantly higher left ventricular end-diastolic pressure. To investigate the basis of the diastolic dysfunction, we explored possible defects in the beta-adrenergic receptor-G(s/i) protein-adenylyl cyclase-cAMP-protein kinase A-phosphatase pathway, as well as molecular or functional alterations of sarcoplasmic reticulum Ca(2+)-ATPase and phospholamban (PLB). We found no significant difference in both mRNA and protein levels of sarcoplasmic reticulum Ca(2+)-ATPase and PLB in post-MI left ventricle compared with control. However, the basal levels of both the protein kinase A-phosphorylated site (Ser16) of PLB (p16-PLB) and the calcium/calmodulin-dependent protein kinase-phosphorylated site (Thr17) of PLB (p17-PLB) were decreased by 76% and 51% in post-MI myocytes (P<0.05), respectively. No change was found in the beta-adrenoceptor density, G(salpha) protein level, or adenylyl cyclase activity. Inhibition of phosphodiesterase and G(i) protein by Ro-20-1724 and pertussis toxin, respectively, did not correct the decreased p16-PLB or p17-PLB levels. Stimulation of beta-adrenoceptor or adenylyl cyclase increased both p16-PLB and p17-PLB in post-MI myocytes to the same levels as in sham myocytes, suggesting that decreased p16-PLB and p17-PLB in post-MI myocytes is not due to a decrease in the generation of p16-PLB or p17-PLB. We found that type 1 phosphatase activity was increased by 32% (P<0.05) with no change in phosphatase 2A activity. Okadaic acid, a protein phosphatase inhibitor, significantly increased p16-PLB and p17-PLB levels in post-MI myocytes and partially corrected the prolonged relaxation of the [Ca(2+)](i) transient. In summary, prolonged relaxation of post-MI remodeled myocardium could be explained, in part, by altered basal levels of p16-PLB and p17-PLB caused by increased protein phosphatase 1 activity.
Huang
B
B
Cardiology Division, Department of Medicine, State University of New York Health Science Center and Veterans Affairs Medical Center, Brooklyn, NY 11203, USA.
Wang
S
S
Qin
D
D
Boutjdir
M
M
El-Sherif
N
N
D000818:Animals; D002135:Calcium-Binding Proteins / Q000378:metabolism*; D019204:GTP-Binding Proteins / Q000378:metabolism*; D006352:Heart Ventricles / Q000378:metabolism* / Q000473:pathology / Q000503:physiopathology; D009203:Myocardial Infarction / Q000378:metabolism* / Q000473:pathology / Q000503:physiopathology; D010749:Phosphoprotein Phosphatases / Q000378:metabolism*; D010727:Phosphoric Diester Hydrolases / Q000378:metabolism*; D010766:Phosphorylation; D054645:Protein Phosphatase 1; D054648:Protein Phosphatase 2; D051381:Rats; D017207:Rats, Sprague-Dawley; D011943:Receptors, Adrenergic, beta / Q000502:physiology*; D012519:Sarcoplasmic Reticulum / Q000378:metabolism; D000099211:Phospholamban
D002135:Calcium-Binding Proteins; D019204:GTP-Binding Proteins; D010749:Phosphoprotein Phosphatases; D010727:Phosphoric Diester Hydrolases; D054645:Protein Phosphatase 1; D054648:Protein Phosphatase 2; D011943:Receptors, Adrenergic, beta; D000099211:Phospholamban