Portal hypertension (PHT) is characterized by splanchnic hyperemia due to a reduction in mesenteric vascular resistance. The reasons for the decreased resistance include an increased responsiveness to a vasodilator substance. Because the activation of an inhibitory guanine nucleotide regulatory (Gi) protein can result in endothelium-dependent relaxation, we tested the hypothesis that exaggerated Gi-protein induced relaxation via a nitric oxide (NO)-dependent pathway partly reflects the enhanced Gi-protein expression in PHT vessels. PHT was created in Sprague-Dawley rats by a partial portal-vein ligation. Control animals were sham operated. Using isolated vascular rings in the absence or presence of an intact endothelium, N(G)-nitro-L-arginine methyl ester (L-NAME), and pertussis toxin, dose response relationships for sodium fluoride (NaF; range, 0.1-4 mmol/L), a Gi protein activator, were determined in a cumulative manner. Gi-protein expression was determined by Western blotting. NaF caused a dose-dependent relaxation in both sham and portal hypertensive pre-contracted vessels, an effect that was significantly inhibited by pertussis toxin, endothelial denudation, and L-NAME. Concentrations of NaF greater than 4 mmol/L caused contractions, an effect that was unaffected by L-NAME. The NaF-induced relaxation response was significantly greater in PHT vessels as compared with sham concomitant with increased Gi-protein expression in PHT vessels. These data suggest that the enhanced endothelial Gi-protein-induced relaxation in PHT vessels may partly reflect enhanced expression of Gi-proteins in PHT vessels and may, thus, represent an important mechanism for exaggerated NO-dependent relaxation in the PHT vasculature.