ObjectiveTo investigate a possible direct action of vasoactive intestinal polypeptide (VIP) on adrenal cortisol secretion and to define its mechanism of action.DesignThe human adrenocortical carcinoma cell line NCI H295, which is not contaminated by medullary chromaffin cells, was used to aid distinction between a direct action of VIP on adrenocortical cells and an indirect mechanism involving VIP-stimulated release of catecholamines.MethodsNCI H295 cells were challenged with 10−11-10−7mol/l VIP for 4 h, with or without prior exposure for 72 h to 10 mmol/l forskolin. Cortisol and cyclic AMP contents of the overlying media were measured using in-house radioimmunoassays. Cells were treated with 10−8-10−6mol/l adrenaline or 3.3 × 10−8mol/l VIP with and without 10−8-10−6mol/l propranolol to exclude the possibility that an indirect mechanism of action involving β-adrenoceptors was operating.ResultsVIP treatment produced an increase in cortisol secretion without pre-incubation, but this was markedly enhanced by prior exposure of cells to forskolin. VIP was potent, with a threshold of 10−11mol/l (n = 4), reaching a maximum 3.9 ± 0.9-fold increase in effect on cells pre-exposed to forskolin (n = 4) by 3.3 × 10−8mol/l. This increase matched the 4 h response to 10 μmol/l forskolin. Cortisol secretion was accompanied by a parallel, dose-dependent increase in accumulation of cAMP.ConclusionsVIP potently and directly stimulates secretion of cortisol from these adrenocortical cells of human origin via an adenylate cyclase-coupled VIP receptor. These findings raise the possibility of a significant and direct effect of VIP in the control of steroid secretion from the adrenal cortex in humans.