Mechanisms underlying the activation of L-type calcium channels by urocortin in rat ventricular myocytes

Aims The aim of this study was to elucidate the signalling pathways implicated in the modulation of cardiac L-type Ca2+ channels by urocortin (Ucn) in ventricular myocytes. Methods and results Adult rat ventricular myocytes were stimulated in vitro with Ucn for 20–40 min. L-type calcium currents ( I CaL) were measured with the patch-clamp technique, whereas quantification of activation of extracellular signal-regulated kinases 1/2 (ERK1/2) was assessed by sandwich-ELISA. Ucn induced a significant increase in I CaL density that was not prevented by the protein kinase A (PKA) inhibitor KT-5720 or the non-selective antagonist of guanine nucleotide exchange factor brefeldin A. The Ucn effect was antagonized by astressin, a corticotropin-releasing factor receptor-2 (CRF-R2) antagonist, and significantly reduced by protein kinase C (PKC) and ERK1/2 inhibitors. The cyclic AMP (cAMP) analogue 8-pCPT-2′OMe-cAMP, which selectively activates the exchange protein activated by cAMP (Epac), was ineffective in modifying I CaL. Analysis of phospho-ERK1/2 showed that Ucn induced a significant activation of the ERK1/2 pathway in ventricular myocytes and this effect was prevented by pre-incubation with PKC inhibitors. Conclusion The present study provides evidence of new mechanisms involved in the modulation of L-type Ca2+ channels by Ucn in adult ventricular myocytes. We propose that the marked increase in I CaL density induced by Ucn is mediated through CRF-R2 and involves PKC-dependent activation of the ERK1/2 pathway, whereas PKA and Epac signalling are not implicated.