Mechanisms underlying the vascular and hypotensive actions of the labdane ent-3-acetoxy-labda-8(17),13-dien-15-oic acid.

We investigated the mechanisms underlying the vasorelaxant and hypotensive actions of the labdane-type diterpene ent-3-acetoxy-labda-8(17),13-dien-15-oic acid (labda-15-oic acid). Vascular reactivity experiments were performed in aortic rings isolated from male Wistar rats. cAMP and cGMP were measured by enzyme immunoassay (EIA) whereas nitrate measurement was performed by chemiluminescence. Nitric oxide (NO) concentration ([NO]c) was measured in endothelial cells by flow cytometry. The cytosolic calcium concentration ([Ca2+]c) in vascular smooth muscle cells (VSMC) was measured by confocal microscopy. Blood pressure measurements were performed in conscious rats. Labda-15-oic acid inhibited the contraction induced by phenylephrine and serotonin in either endothelium-intact or endothelium-denuded rat aortic rings. The labdane significantly reduced CaCl2-induced contraction in a Ca2+-free solution containing KCl or phenylephrine. Labda-15-oic acid (0.1–300 μmol/l) concentration-dependently relaxed endothelium-intact and endothelium-denuded aortas pre-contracted with either phenylephrine or KCl. In endothelium-intact rings, the relaxation induced by labda-15-oic acid was affected by L-NAME, 7-nitroindazole, ODQ, hemoglobin, Rp-8-Br-Pet-cGMPS and thapsigargin. Blockade of K+ channels with 4-aminopyridine, apamin, charybdotoxin and glibenclamide affected the relaxation induced by labda-15-oic acid. The labdane increased cGMP and nitrate levels but did not affect cAMP levels in endothelium-intact aortas. Labda-15-oic acid increased [NO]c in endothelial cells and decreased [Ca2+]c in VSMC. The hypotension induced by intravenous administration of labda-15-oic acid (0.3–3 mg/kg) was partially reduced by L-NAME. In conclusion, the mechanisms underlying the cardiovascular actions of the labdane involve the activation of the endothelial NO-cGMP pathway, the opening of K+ channels and the alteration on Ca2+ mobilization.