Orexin-A (OXA) regulates food intake and energy homeostasis. It increases insulin secretion in vivo, and in vitro, while controversial effects of OXA on plasma glucagon are reported.

We characterize the effects of OXA on glucagon secretion and identify intracellular target molecules in glucagon-producing cells.

Glucagon secretion from in situ perfused rat pancreas, isolated rat pancreatic islets and clonal pancreatic A-cells (InR1-G9) was measured by RIA. The expression of orexin receptor 1 (OXR1) was detected by western blot and immunofluorescence. The effects of OXA on cyclic AMP, AKT, PDK-1, Foxo1 and CREB were measured by ELISA and western blot. Intracellular calcium (Ca²⁺_i) concentration was detected by Fura-2, glucagon expression by real-time PCR. Foxo1 was silenced in InR1-G9 cells by transfecting cells with short interfering RNA.

OXR1 was expressed on pancreatic A-cells and InR1-G9 cells. OXA reduced glucagon secretion from perfused rat pancreas, isolated rat pancreatic islets and InR1-G9 cells. OXA inhibited proglucagon gene expression via the PI-3-kinase dependent pathway. OXA decreased cyclic AMP and [Ca²⁺]_i concentration, and increased AKT, PDK-1 and Foxo1 phosphorylation. Silencing of Foxo1 caused a reversal of the inhibitory effect of OXA on proglucagon gene expression.

Our study provides the first in vitro evidence for the interaction of OXA with pancreatic A-cells. OXA inhibits glucagon secretion and reduces intracellular cAMP and Ca²⁺_i concentration. OXA increases AKT/PDK-1 phosphorylation and inhibits proglucagon expression via PI3K- and Foxo-1-dependent pathway. As a physiological inhibitor of glucagon secretion OXA may have a therapeutic potential to reduce hyperglucagonemia in type 2 diabetes.