0039: Phosphodiesterase type 4 inhibition enhances pro-arrhythmic spontaneous Ca2+ waves induced by β-adrenergic stimulation: respective role of protein kinase A and Ca2+/calmodulin kinase II

β-adrenoceptors (β-AR) increase cardiac function by rising cAMP and activating protein kinase A (PKA) which phosphorylates L-type Ca2+ channels, ryanodine receptors (RyR) and phospholamban (PLB). These proteins are also targets of Ca2+/calmodulin kinase II (CaMKII), which is also activated upon β-AR stimulation. Multiple cyclic nucleotide phosphodiesterases (PDEs) regulate cAMP, among which the PDE4 family is overriding in rodent heart. We and others have shown that PDE4 genetic ablation in mice leads to enhanced arrhythmias. Here, we investigated the mechanisms underlying the proarrhythmic effects of PDE4 inhibition. Adult rat ventricular myocytes loaded with 1μM Fura-2AM were stimulated at 1Hz and Ca2+ transients recorded using an Ionoptix® system. Spontaneous calcium waves (SCWs) occurred upon non selective β-AR stimulation with isoproterenol (Iso 1nM) during a 10s pause of electrical stimulation and increased from 0.4±0.3 (n=10) to 2.3±0.2 SCWs/10s (n=15; p<0.001) in the presence of the specific PDE4 inhibitor Ro20-1724 (Ro 10μM). Ro also potentiated the positive inotropic effect of Iso as well as the sarcoplasmic reticulum (SR) Ca2+ load (n=11; p<0.05) and SR Ca2+ leak measured in a 0Na+,0Ca2+ solution ±1mM tetracaine (n=11; p<0.001). Ro potentiated Iso-induced phosphorylation of PLB and RyR by PKA and CaMKII. PKA inhibition with 10μM H-89 abolished SR Ca2+ leak, SCWs and the inotropic effect of Iso±Ro (n=8; p<0.001). The CaMKII inhibitor KN-93 (10μM), but not its inactive analogue KN-92, diminished SCWs by 72% (n=8; p<0.001) and SR Ca2+ leak by 85% (n=16; p