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, Ca(2+)/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 Ca(2+) signaling was analyzed in isolated cardiac myocytes by confocal microscopy. We observed a positive inotropic effect manifested by increased [Ca(2+)](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 [Ca(2+)] 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 Ca(2+) influx via L-type Ca(2+) channels. However, 8-pCPT also induced aberrant Ca(2+) release as Ca(2+) waves and extra [Ca(2+)](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.
Inserm, U769, Univ. Paris Sud, IFR141, Labex Lermit, Châtenay-Malabry, France.
Gema
G
Ruiz-Hurtado
Alejandro
A
Domínguez-Rodríguez
Laetitia
L
Pereira
María
M
Fernández-Velasco
Cécile
C
Cassan
Frank
F
Lezoualc'h
Jean-Pierre
JP
Benitah
Ana M
AM
Gómez
D000818:Animals; D002110:Caffeine / Q000494:pharmacology; D020746:Calcium Channels, L-Type / Q000378:metabolism; D020013:Calcium Signaling / Q000187:drug effects; D002147:Calmodulin / Q000037:antagonists & inhibitors / Q000235:genetics / Q000378:metabolism*; D006332:Cardiomegaly / Q000378:metabolism / Q000473:pathology; D002478:Cells, Cultured; D000242:Cyclic AMP / Q000031:analogs & derivatives / Q000494:pharmacology; D004789:Enzyme Activation; D020536:Enzyme Activators / Q000494:pharmacology; D055785:Gene Knockdown Techniques; D020662:Guanine Nucleotide Exchange Factors / Q000235:genetics / Q000378:metabolism*; D008297:Male; D018613:Microscopy, Confocal; D009200:Myocardial Contraction; D032383:Myocytes, Cardiac / Q000378:metabolism / Q000502:physiology*; D018408:Patch-Clamp Techniques; D034741:RNA, Small Interfering / Q000235:genetics; D051381:Rats; D017208:Rats, Wistar; D015533:Transcriptional Activation
C473217:8-(4-chloro-phenylthio)-2'-O-methyladenosine-3'-5'-cyclic monophosphate; D020746:Calcium Channels, L-Type; D002147:Calmodulin; D020536:Enzyme Activators; D020662:Guanine Nucleotide Exchange Factors; D034741:RNA, Small Interfering; C526973:Rapgef3 protein, rat; D002110:Caffeine; D000242:Cyclic AMP