Sustained Epac activation induces calmodulin dependent positive inotropic effect in adult cardiomyocytes

Cardiac actions of Epac (exchange protein directly activated by cAMP) are not completely elucidated. Epac induces cardiomyocytes hypertrophy, Ca2 +/calmodulin protein kinase II (CaMKII) and excitation–transcription coupling in rat cardiac myocytes. Here we aimed to elucidate the pathway cascade involved in Epac sustained actions, as during the initiation of hypertrophy development, where β-adrenergic signaling is chronically stimulated. Rats were treated with the Epac selective activator 8-pCPT during 4 weeks and Ca2 + signaling was analyzed in isolated cardiac myocytes by confocal microscopy. We observed a positive inotropic effect manifested by increased [Ca2 +]i transient amplitudes. In order to further analyze these actions, we incubated adult cardiomyocytes in the presence of 8-pCPT. The effects were similar to those obtained in-vivo and are blunted by Epac1 knock down. Interestingly, the increase in [Ca2 +] transients was abolished by protein synthesis blockade or when the downstream effectors of calmodulin (CaMKII or calcineurin) were inhibited, pointing to calmodulin (CaM) as an important downstream protein in Epac sustained actions. In fact, CaM expression was enhanced by 8-pCPT treatment in isolated cells, as found by Western blots. Moreover, the 8-pCPT-induced, PKA-independent, positive inotropic effect was favored by enhanced extracellular Ca2 + influx via L-type Ca2 + channels. However, 8-pCPT also induced aberrant Ca2 + release as Ca2 + waves and extra [Ca2 +]i transients, suggesting proarrhythmic effect. These results provide new insights regarding Epac cardiac actions, suggesting an important role in the initial compensation of the heart to pathological stimuli during the initiation of cardiac hypertrophy, favoring contraction but also arrhythmia risk.

Research highlights
► Sustained Epac activation by 8-pCPT induces a positive inotropic effect.
► Epac increased [Ca2+]i transients and SR Ca2+ load.
► Epac increased Ca2+ entry by enhancing ICa,L window current.
► Epac decreased Ca2+ extrusion and accelerated Ca2+ reuptake though SERCA.
► Effects depended on calmodulin expression and calcineurin and CaMKII activation.