The role of Epac in the regulation of intracellular Ca
The role of Epac in the regulation of intracellular Ca homeostasis and contractility is still matter of debate. In rat adult cardiomyocytes, acute Epac stimulation decreased the amplitude of Ca transients , ,  with either no changes  or increments  in cell shortening, suggesting an enhancement of myofilament Ca sensitivity. This was confirmed in the latter study and supported by the finding of the increased CaMKII-phosphorylation of MyBPC and TnI . The diminished Ca transient was paralleled by a decrease in the amount of Ca stored in the SR, attributed to the increased SR Ca leak induced by the CaMKII-dependent phosphorylation of RyR2 . In RG2833 to rat myocytes, an increase ,  or no change in Ca transient  were detected in mice myocytes after acute stimulation of Epac. As in the rat, 8-CPT induced an enhancement of the CaMKII-dependent phosphorylation of RyR2 and PLN . Furthermore, the Epac-specific agonist caused spontaneous triggered activity in intact perfused murine hearts, associated with increased incidence of spontaneous Ca transients and propensity to the generation of Ca waves at the myocyte level . Such arrhythmogenic features were also observed in rat myocytes but after sustained Epac activation . In this case, rat myocytes showed an increase in Ca transient, cell shortening and SR Ca content, favored by enhanced Ca influx through the L-type Ca channels. The development of KO mice did not help to clarify the specific involvement of Epac in cardiac contractile behavior. Pereira et al. 2013  showed unaltered basal cardiac function and Ca handling in KO mice of either Epac1 or Epac2 and double KO mice. Moreover Epac2 and not Epac1, was shown to be essential for 8-CPT-induced RyR2 activation, enhanced Ca leak and decreased Ca transient . However, Okumura et al. 2014  found that loss of Epac1 decreased basal cardiac contractility, reduced Ca transient and diminished SR Ca storage. Overall, the effects of Epac in intracellular Ca handling and contractility remain controversial. The apparent discrepancy may depend on experimental conditions. For instance, genetic background in the KO models, acute vs. chronic effects of 8-CPT, species and/or different extracellular Ca ([Ca]o) to which the myocytes are exposed. Related to the latter, it is important to consider that Ca supply to the cell alters intracellular Ca, dynamically adjusting the balance between SR Ca uptake and leak.
The aim of the present study was to elucidate if the acute effects of Epac stimulation depend on the SR Ca handling state. In determining this, we varied [Ca]o in order to shift the SR balance from net Ca accumulation to net Ca release and we focused on the relevance of CaMKII-dependent phosphorylation of PLN and RyR2 in the response to Epac stimulation, through the use of transgenic mice with non-phosphorylatable CaMKII sites.
Materials and methods
Discussion The discovery of Epac proteins expanded the range of cAMP effectors and many of the mechanisms believe to be PKA-mediated proved to be Epac-dependent. At the level of the myocardium, the currently understanding of the effects of Epac activation remains controversial. Inconsistent results have been reported regarding the impact of Epac on Ca homeostasis, contractility, apoptosis, hypertrophy and fibrosis , . The present study was carried out in order to clarify some of these discrepancies. We particularly focused on the effects of Epac activation on cardiomyocyte contractility and SR Ca handling. The results obtained reveal that: 1) The activation of Epac promotes different effects on Ca handling and contractility in cardiac myocytes, which do not depend on the species but on the experimental conditions that alter the fine-tuning of the SR Ca handling; 2) Epac stimulation regulates both the uptake and release of Ca from the SR; 3) CaMKII is an essential element in the downstream cascade of the Epac signaling, which through PLN phosphorylation determines the positive inotropic effect of Epac at low [Ca]o and through RyR2 phosphorylation is responsible for the arrhythmogenic effects of Epac activation at high [Ca]o and 4) PKC is involved in the Epac-induced CaMKII activation.