There is substantial evidence supporting a hypertrophic action of serotonin [5-hydroxytryptamine (5-HT)] in cardiomyocytes. However, little is known about the mechanisms involved. We previously demonstrated that 5-HT-induced hypertrophy depends, in part, on the generation of reactive oxygen species by monoamine oxidase-A (MAO-A) (see Ref. 3). Cardiomyocytes express 5-HT(2) receptors, which may also participate in hypertrophy. Here, we analyzed the respective contribution of 5-HT(2) receptors and MAO-A in H9C2 cardiomyoblast hypertrophy. 5-HT induced a dose-dependent increase in [(3)H]leucine incorporation and stimulation of two markers of cardiac hypertrophy, ANF-luc and alphaSK-actin-luc reporter genes. Experiments using 1 microM 5-HT showed that hypertrophic response occurred independently from MAO-A. Using pharmacological inhibitors (M100907 and ketanserin), we identified a novel mechanism of action involving 5-HT(2A) receptors and requiring Ca(2+)/calcineurin/nuclear factor of activated T-cell activation. The activation of this hypertrophic pathway was fully prevented by 5-HT(2A) inhibitors and was unaffected by MAO inhibition. When 10 microM 5-HT was used, an additional hypertrophic response, prevented by the MAO inhibitors pargyline and RO 41-1049, was observed. Unlike the 5-HT(2A)-receptor-mediated H9C2 cell hypertrophy, MAO-A-dependent hypertrophic response required activation of extracellular-regulated kinases. In conclusion, our results show the existence of a dose-dependent shift of activation of distinct intracellular pathways involved in 5-HT-mediated hypertrophy of cardiac cells.