The insulin/insulin-like growth factor (IGF) signaling pathway plays a critical role in the regulation of islet cell biology. However, the signaling pathway(s) utilized by insulin to directly modulate -cells is unclear. To interrogate whether insulin exerts endocrine effects in regulating proteins in the insulin/IGF-1 signaling cascade in vivo in physiological states via the insulin receptor, we designed two experimental approaches: 1) glucose gavage and 2) hyperinsulinemic intravenous infusion, for studies in either -cell specific insulin receptor knock-out (IRKO) or control mice. Immunostaining of sections of pancreas (collected immediately after glucose gavage or insulin infusion) from controls showed significant increases in pAKT+, p-p70S6K+, and pERK+ -cells and a significant decrease in % nuclear FoxO1+ -cells compared with corresponding vehicle-treated groups. In contrast, in IRKOs, we observed no significant changes in pAKT+ or p-p70S6K+ -cells in either experiment; however, pERK+ -cells were significantly increased, and an attenuated decrease in % nuclear FoxO1+ cells was evident in response to glucose gavage or insulin infusion. Treatment of control and IRKO -cell lines with glucose or insulin showed significantly decreased % nuclear FoxO1+ -cells suggesting direct effects. Furthermore, blocking MAPK signaling had virtually no effect on FoxO1 nuclear export in controls, in contrast to attenuated export in IRKO -cells. These data suggest insulin acts on -cells in an endocrine manner in the normal situation; and that in -cells lacking insulin receptors, insulin and glucose minimally activate the Akt pathway, while ERK phosphorylation and FoxO1 nuclear export occur independently of insulin signaling.