Cyclic AMP protects against hepatocyte apoptosis by a protein kinase A independent cAMP-GEF/phosphoinositide-3-kinase (PI3K)/Akt signaling pathway. However, the signaling pathway coupling cAMP-GEF with PI3K is unknown. The aim of this study was to investigate the role of Src tyrosine kinases (Src-TYK) and PI3K-p110 isoforms in this pathway. Studies were done in rat hepatocytes using the hydrophobic bile acid, glycochenodeoxycholic acid (GCDC) to induce apoptosis. Cyclic AMP-GEFs were selectively activated using, 4-(4-chloro-phenylthio)-2'-O-methyladenosine-3'-5'-cyclic monophosphate, (CPT-2-Me-cAMP), which sequentially phosphorylated Src-TYK (within 1 min) followed by Akt (within 5 min). Src inhibitors, PP2 and SU6656, inhibited basal and CPT-2-Me-cAMP -mediated Src and Akt phosphorylation. These inhibitors had no effect on CPT-2-Me-cAMP mediated activation of Rap GTPases. CPT-2-Me-cAMP induced transient Src dependent autophosphorylation of the epidermal growth factor receptor (EGFR). Inhibition of the EGFR with AG 1478 partially inhibited the ability of CPT-2-Me to phosphorylate Akt. While PP2 completely abolished the protective effect of CPT-2-Me-cAMP in GCDC induced apoptosis, AG 1478 partially inhibited the cytoprotective effect. CPT-2 Me-cAMP treatment resulted in Src dependent activation of the p110 and subunits of PI3K, but only the latter was sensitive to inhibition with AG 1478. Conclusion: Activation of cAMP-GEFs results in phosphorylation of Src-TYK and Akt and activation of the p110/ subunits of PI3K. Maximal cAMP-GEF mediated Akt phosphorylation as well as protection from bile acid induced apoptosis requires activation of Src-TYK and the EGFR. These studies support the existence of 2 pathways: cAMP-GEF/ Rap/ Src/PI3K/Akt and cAMP-GEF/Rap/Src/EGFR/PI3K/Akt both of which are necessary for maximal cytoprotective effect of cAMP-GEFs in hepatocytes.