Glucose-dependent insulinotropic polypeptide (GIP) potentiates glucose-stimulated insulin secretion, insulin biosynthesis, and beta-cell proliferation and survival. In previous studies GIP was shown to promote beta-cell survival by modulating the activity of multiple signaling modules and regulating gene transcription of pro- and anti-apoptotic bcl-2 family proteins. We have now evaluated the mechanisms by which GIP regulates the dynamic interactions between cytoplasmic bcl-2 family members and the mitochondria in INS-1 cells during apoptosis induced by treatment with staurosporine (STS), an activator of the mitochondria-mediated apoptotic pathway. STS induced translocation of bad and bimEL, activation of mitochondrial bax, release of mitochondrial cytochrome c, cleavage of caspase-3, and apoptosis. Each response was significantly diminished by GIP. Using selective enzyme inhibitors, overexpression of dominant-negative Akt, and Akt siRNA, it was demonstrated that GIP promoted beta-cell survival via Akt-dependent suppression of p38 MAPK and JNK and that combined inhibition was sufficient to explain the entire pro-survival responses to GIP during STS treatment. This signaling pathway also explained the pro-survival effects of GIP on INS-1 cells exposed to two other promoters of stress: thapsigargin (endoplasmic reticulum stress) and etoposide (genotoxic stress). Importantly, we discovered that GIP suppressed p38 MAPK and JNK via Akt-mediated changes in the phosphorylation state of the apoptosis signal-regulating kinase 1 in INS-1 cells and human islets, resulting in inhibition of its activity. Inhibition of apoptosis by GIP is therefore mediated via a key pathway involving Akt-dependent inhibition of apoptosis signal-regulating kinase 1, which subsequently prevents the pro-apoptotic actions of p38 MAPK and JNK.