Ser16-, but not Thr17-phosphorylation of phospholamban influences frequency-dependent force generation in human myocardium

beta-Adrenoceptor/cAMP-dependent Ser(16)-phosphoryation as well as Ca2+-dependent Thr(17)-phosphorylation of phospholamban (PLB) influences SERCA 2a activity and thus myocardial contractility. To determine the cross-signaling between Ca2+ and cAMP pathways, the phosphorylation of Ser(16)-PLB and Thr(17)-PLB was studied at increasing stimulation frequencies as well as in the presence of beta-adrenergic stimulation in isolated ventricular trabeculae from failing (dilative cardiomyopathy, DCM, heart transplants, n=9) and non-failing human myocardium (donor hearts, NF, n=9). In addition, we measured the intracellular Ca2+-transient (fura-2) at increasing stimulation frequencies (0.5-3.0 Hz). Protein expression of SERCA 2a and phospholamban was similar in DCM and NF. In DCM, diastolic [Ca2+](i) was increased and systolic [Ca2+](i) as well as Ser(16) PLB-phosphorylation were decreased as compared to NF at 0.5 Hz. The positive force-frequency relationship in human non-failing myocardium was accompanied by a frequency-dependent increase in Ser(16)-PLB, but not Thr(17)-PLB phosphorylation. In DCM, Ser(16)-PLB as well as Thr(17)-PLB phosphorylation were not altered at higher stimulation frequencies. After application of isoprenaline (1 muM), a profound increase in Ser(16)-PLB phosphorylation was accompanied by a small increase in Thr(17)-PLB phosphorylation, only in NF. The frequency-dependent phosphorylation of Ser(16)-PLB may favor an increase in Ca2+ transient and force generation in humans. Cross talk signaling of Ser(16)/Thr(17)-PLB phosphorylation after beta-adrenergic stimulation exists in non-failing, but not in failing human myocardium. The Ca2+-dependent CaM-kinase activity may be altered in human heart failure.