Oral administration of grape seed extract (MegaNatural®-BP Patent pending) (GSE) lowered the blood pressure in human subjects with the metabolic syndrome. We tested the hypothesis that GSE caused an endothelium dependent relaxation (EDR) in aortic rings from New Zealand White rabbits. The rings were suspended in organ baths containing oxygenated Krebs buffer at 37C. EDR evoked by acetylcholine (Ac) and GSE were measured after pre-contracting the rings with 10-5 M noradrenalin. The tissues were also tested after removing the endothelium and after incubation with L-NAME, Wotmannin (Wot) and LY 294002 (LY) and SU5416 (SU) to examine the mechanism involved in the relaxation. We also investigated the phosphorylation of e-NOS (Ser-1177) by GSE in- vitro in HUVECs. Relaxation to GSE was abolished in de-endothelialized and L-NAME treated tissues. The EDR was inhibited by PI3 kinase inhibitors (Wot and Ly) but not by SU (VEGFR2 inhibitor). Involvement of e-NOS was confirmed by phosphorylation of e-NOS (Ser-1177) in HUVECs after exposure to GSE. It is concluded that GSE caused EDR by activation of the AKT/PI3 kinase pathway via a mechanism which does not involve VEGFR2.