The calcineurin—nuclear factor of activated T cells signaling pathway mediates the effect of corticotropin releasing factor and urocortins on catecholamine synthesis

The biological effects of the Corticotropin‐releasing factor (CRF) family of neuropeptides are mediated by mobilization of [Ca 2+ ]. Aim of the current work was to examine if the calcineurin/NFAT (nuclear factor of activated T‐cells) signaling pathway is involved in the effect of CRF peptides in catecholamine synthesis and secretion from PC12 rat pheochromocytona cells, a model for the study of adrenal catecholamine production. PC12 cells express both types of CRF receptors. Our data are as follows: (a) The calcineurin inhibitor cyclosporine A (CsA) blocked norepinephrine secretion induced by ligands of either CRF type 1 (CRF 1 ) or 2 (CRF 2 ) receptors on PC12 cells. (b) Silencing NFAT2 expression using a selective NFAT2 siRNA blocked CRF 1 and CRF 2 ‐induced NE production. (c) CRF ligands induced NFAT transcriptional activity in cells transfected with a luciferase reporter construct controlled by NFAT binding elements (NFAT‐Luc). (d) CsA completely blocked the stimulatory effect of CRF 1 and CRF 2 ligands on NFAT activity in NFAT‐Luc transfected cells. (e) PKA, PKC, p38‐MAPK, Tpl2, Ha‐Ras, and AKT1 were crucial intermediates for both CRF 1 and CRF 2 ‐induced NFAT activation. Interestingly, MEK1/2 and ERK1/2 were crucial only for the CRF 2 ‐induced NFAT activation. (f) p38‐MAPK and Tpl2 were crucial intermediates for both CRF 1 and CRF 2 ‐induced norepinephrine production, while AKT1 affected only CRF 2 ‐induced norepinephrine production. In conclusion, our data suggest that CRF 1 and CRF 2 ligands activate the transcription factor NFAT and its activation is prerequisite for CRF‐induced catecholamine production from chromaffin cells. J. Cell. Physiol. 227: 1861–1872, 2012. © 2011 Wiley Periodicals, Inc.