Involvement of the Na+/Ca2+ exchanger isoform 1 (NCX1) in NGF-induced Neuronal Differentiation through Ca2+-Dependent Akt Phosphorylation [Signal Transduction]
November 21st, 2014 by Secondo, A., Esposito, A., Sirabella, R., Boscia, F., Pannaccione, A., Molinaro, P., Cantile, M., Ciccone, R., Sisalli, M. J., Scorziello, A., Di Renzo, G., Annunziato, L.
Neuronal growth factor (NGF) induces neuronal differentiation by modulating intracellular Ca2+ concentrations ([Ca2+]i). However, the role of the three isoforms of the main Ca2+-extruding system, the Na+/Ca2+ exchanger (NCX), in NGF-induced differentiation remains unexplored. We investigated whether NCX1, NCX2 and NCX3 isoforms could play a relevant role in neuronal differentiation through the modulation of [Ca2+]i and Akt pathway. NGF caused a progressive neurite elongation, a significant increase of the well known marker of growth cones GAP-43, an enhancement of endoplasmic reticulum (ER) Ca2+ content, and of Akt phosphorylation through an early activation of ERK1/2. Interestingly, during NGF-induced differentiation NCX1 protein level increased, NCX3 decreased, whereas NCX2 remained unaffected. At the same time, NCX total activity increased. Moreover, NCX1 colocalized and co-immunoprecipitated with GAP-43 and NCX1 silencing prevented NGF-induced effects on GAP-43 expression, Akt phosphorylation and neurite outgrowth. On the other hand, the overexpression of its neuronal splicing isoform NCX1.4, even in the absence of NGF, induced an increase in Akt phosphorylation, and GAP-43 protein expression. Interestingly, TTX-sensitive Na+ currents and SBFI-detected [Na+]i significantly increased in cells over-expressing NCX1.4 as well as ER Ca2+ content. This latter effect was prevented by TTX. Furthermore, either the [Ca2+]i chelator BAPTA-AM or the PI3-K inhibitor LY 294002 prevented Akt phosphorylation and GAP-43 protein expression rise in NCX1.4 over-expressing cells. Moreover, in primary cortical neurons, NCX1 silencing prevented Akt phosphorylation, GAP-43 and MAP2 over-expression as well as neurite elongation. Collectively, these data showed that NCX1 participates in neuronal differentiation through the modulation of ER Ca2+ content and PI3-K signaling.