Signaling pathway for endothelin-1- and phenylephrine-induced cAMP response element binding protein activation in rat ventricular myocytes: Role of inositol 1, 4, 5-trisphosphate receptors and CaMKII

Abstract

Background/Aims: Endothelin-1 (ET-1) and the α 1 -adrenoceptor agonist phenylephrine (PE) activate cAMP response element binding protein (CREB), a transcription factor implicated in cardiac hypertrophy. The signaling pathway involved in CREB activation by these hypertrophic stimuli is poorly understood. We examined signaling pathways for ET-1- or PE-induced cardiac CREB activation. Methods: Western blotting was performed with pharmacological and genetic interventions in rat ventricular myocytes. Results: ET-1 and PE increased CREB phosphorylation, which was inhibited by blockade of phospholipase C, the extracellular-signal-regulated kinase 1/2 (ERK1/2) pathway, protein kinase C (PKC) or Ca 2+ -calmodulin-dependent protein kinase II (CaMKII). Intracellular Ca 2+ buffering decreased ET-1- and PE-induced CREB phosphorylation by ≥80%. Sarcoplasmic reticulum Ca 2+ pump inhibitor, inositol 1, 4, 5-trisphosphate receptor (IP 3 R) blockers, or type 2 IP 3 R (IP 3 R2) knock-out abolished ET-1- or PE-induced CREB phosphorylation. ET-1 and PE increased phosphorylation of CaMKII and ERK1/2, which was eliminated by IP 3 R blockade/knock-out or PKC inhibition. Activation of CaMKII, but not ERK1/2, by these agonists was sensitive to Ca 2+ buffering or to Gö6976, the inhibitor of Ca 2+ -dependent PKC and protein kinase D (PKD). Conclusion: CREB phosphorylation by ET-1 and PE may be mainly mediated by IP 3 R2/Ca 2+ -PKC-PKD-CaMKII signaling with a minor contribution by ERK1/2, linked to IP 3 R2 and Ca 2+ -independent PKC, in ventricular myocytes.