Akt plays a central role in multicellular signaling pathways. Using transgenic (Tg) mice, we have previously demonstrated that overexpression of constitutively active Akt (CA-Akt) in the heart exhibits enhanced cardiac contractility, which is a novel function of Akt mediated by enhanced by L-type Ca2+ current density and upregulation of SERCA2a protein. To further elucidate the supporting mechanisms of enhanced contractility, we examined the effect of thapsigargin, a specific inhibitor of SERCA2a, on % contraction in cardiomytes from CA-Akt Tg and wild type (WT) mice. The dose of thapsigargin to inhibit %contraction into half (IC50) was 2.6 fold higher in CA-Akt Tg than in WT mice, which is consistent with a notion that enhanced SERCA2a plays an important role in mediating increased contractility of CA-Akt Tg myocytes. We next examined whether SERCA2a is also upregulated at the level of mRNA. Semi-quantitative RT-PCR indicated that there was no significant difference in SERCA2a mRNA levels between CA-Akt Tg and WT mice. Thus, the upregulation of SERCA2a protein in CA-Akt Tg mice is caused by post-transcriptional mechanisms. We also evaluated the effect of adenovirus tranduction of CA-Akt upon L-type Ca2+ channel currents using neonatal rat cardiomyocytes. 24 hours after adenovirus transnduction, the peak ICa amplitude was significantly increased in CA-Akt transduced myocytes than in control virus transduced myocytes. This suggests that enhanced L-type Ca2+ channel currrents can be the direct consequence of Akt activation. Finally, we prepared adult rat cardiac myocyte culture and transduced adenovirus of CA-Akt and LacZ (control). Two days after tranduction, CA-Akt myocytes exhibited significantly enhanced contractility (1.4 fold in %contraction, p<0.01), increased Fura-2 Ca2+ transient (1.9 fold, p<0.01) compared with control myocytes. SERCA2a protein expression was also increased (4.8 fold, p<0.01). These findings were consistent with those in CA-Akt Tg mice. These results suggest that overexpression of Akt enhances cardiac function. The underlying cellular mechanism(s) include potentiation of L-type Ca2+ channel function and upregulation of SERCA2a, which indicate that Akt directly affects Ca2+ handling.