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 hypertro phy of noninfarcted myocardium is at its maximum and the heart is in a comp ensated stage with no evidence of heart failure. Our hemodynamic measuremen ts at this stage showed a slight but insignificant decrease of +dP/dt but a significantly higher left ventricular end-diastolic pressure. To investiga te the basis of the diastolic dysfunction, we explored possible defects in the beta-adrenergic receptor-G(s/i) protein-adenylyl cyclase-cAMP-protein k inase A-phosphatase pathway, as well as molecular or functional alterations of sarcoplasmic reticulum Ca2+-ATPase and phospholamban (PLB). We found no significant difference in both mRNA and protein levels of sarcoplasmic ret iculum Ca2+-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 kina se-phosphorylated site (Thr17) of PLB (p17-PLB) were decreased by 76% and 5 1% in post-MI myocytes (P < 0.05), respectively. No change was found in the P-adrenoceptor density, G(s alpha) protein level, or adenylyl cyclase acti vity. Inhibition of phosphodiesterase and G(i) protein by Ro-20-1724 and pe rtussis toxin respectively, did not correct the decreased p16-PLB or p17-PL B levels. Stimulation of beta-adrenoceptor or adenylyl cyclase increased bo th p16-PLB and p17-PLB in post-MI myocytes to the same levels as in sham my ocytes, suggesting that decreased p16-PLB and p17-PLB in post-MI myocytes i s not due to a decrease in the generation of p16-PLB or p17-PLB. We found t hat type I phosphatase activity was increased by 32% (P < 0.05) with no cha nge in phosphatase 2A activity. Okadaic acid, a protein phosphatase inhibit or, significantly increased p16-PLB and p17-PLB levels in post-MI myocytes and partially corrected the prolonged relaxation of the [Ca2+](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.