A Low-Dose Beta 1-Blocker In Combination With Milrinone Improves Intracellular Ca2+ Handling In Failing Cardiomyocytes By Inhibition Of Milrinone-Induced Diastolic Ca2+ Leakage From The Sarcoplasmic Reticulum

ObjectivesThe purpose of this study was to investigate whether adding a low-dose beta 1-blocker to milrinone improves cardiac function in failing cardiomyocytes and the underlying cardioprotective mechanism.BackgroundThe molecular mechanism underlying how the combination of low-dose beta 1-blocker and milrinone affects intracellular Ca2+ handling in heart failure remains unclear.MethodsWe investigated the effect of milrinone plus landiolol on intracellular Ca2+ transient (CaT), cell shortening (CS), the frequency of diastolic Ca2+ sparks (CaSF), and sarcoplasmic reticulum Ca2+ concentration ({Ca2+}(SR)) in normal and failing canine cardiomyocytes and used immunoblotting to determine the phosphorylation level of ryanodine receptor (RyR2) and phospholamban (PLB).ResultsIn failing cardiomyocytes, CaSF significantly increased, and peak CaT and CS markedly decreased compared with normal myocytes. Administration of milrinone alone slightly increased peak CaT and CS, while CaSF greatly increased with a slight increase in {Ca2+}(SR). Co-administration of beta 1-blocker landiolol to failing cardiomyocytes at a dose that does not inhibit cardiomyocyte function significantly decreased CaSF with a further increase in {Ca2+}(SR), and peak CaT and CS improved compared with milrinone alone. Landiolol suppressed the hyperphosphorylation of RyR2 (Ser2808) in failing cardiomyocytes but had no effect on levels of phosphorylated PLB (Ser16 and Thr17). Low-dose landiolol significantly inhibited the alternans of CaT and CS under a fixed pacing rate (0.5 Hz) in failing cardiomyocytes.ConclusionA low-dose beta 1-blocker in combination with milrinone improved cardiac function in failing cardiomyocytes, apparently by inhibiting the phosphorylation of RyR2, not PLB, and subsequent diastolic Ca2+ leak.