Atividade espasmolítica não seletiva da 5,7-dimetoxiflavona isolada de Zornia brasiliensis Vogel (Leguminosae) e seus possíveis mecanismos de ação em aorta de rato.

Abstract

Zornia brasiliensis Vogel is popularly known as "urinaria", "urinana" or "carrapicho" and the crude ethanolic extract obtained from its aerial parts, solubilized in Cremophor, showed spasmolytic activity in ileum and trachea of guinea pig, rat uterus, cavernous body and aorta. Among other secondary metabolites, this species has flavonoids, which are reported in the literature because they have an effect on the smooth muscle. It has been reported that the 5,7-dimethoxyflavone (5,7-DMF) flavonoid, solubilized in dimethyl sulfoxide (DMSO), presents vasorelaxant activity in the rat aorta. The aim of this study was to investigate whether the flavonoid 5,7-DMF isolated from Z. brasiliensis would present a spasmolytic effect in other models of phasic and tonic smooth muscle, in addition to characterize the mechanism of action of the one that had greater pharmacological potency, compared to those already described for this flavonoid. Isotonic and isometric contractions were evaluated. All experimental protocols were approved by CEUA/UFPB (certificate: 129/16). In guinea pig ileum, flavonoid more potently antagonized the phasic contractions induced by histamine (IC50 = 9.9 ± 1.4 x 10-6 M) than by carbachol (CCh) (IC50 = 3.2 ± 0.5 x 10-5 M). 5,7-DMF also relaxed equipotently the guinea pig trachea pre-contracted with CCh both in the presence (EC50 = 2.4 ± 0.4 x 10-5 M) and in the absence (EC50 = 2.6 ± 0.4 x 10-5 M) of functional epithelium. Different, in isolated rat aorta, 5,7-DMF relaxed the pre-contracted aorta with phenylephrine with greater potency in the presence (EC50 = 4.5 ± 0.7 x 10-6 M) than in absence (EC50 = 1.7 ± 0.3 x 10-5 M) of functional endothelium, indicating the participation of the endothelium-derived relaxing factors in the 5,7-DMF vasorelaxant action. Because of the greater relaxing potency in the rat aorta between the organs tested, along with the fact that flavonoid was more effective and potent when it was solubilized in Cremophor® than in DMSO, we decided to investigate the mechanism of action of 5,7-DMF in rat aorta. It can be observed that in the presence of atropine (EC50 = 3.8 ± 0.7 x 10-6 M), the relaxation control curve of 5,7-DMF was not displaced, discarding the participation of endothelial muscarinic receptors in its vasorelaxant effect. To confirm the participation of the nitric oxid (NO) pathway, one of the main endothelium-derived relaxing factors, L-NAME an NO synthase inhibitor (NOS), was used and the curve was shifted to right in the presence of this inhibitor (EC50 = 2.0 ± 0.3 x 10-5 M) with reduction of the relaxing potency. This inhibitory effect of L-NAME was reversed in the presence of high concentrations of L-arginine (EC50 = 3.4 ± 0.8 x 10-6 M). In addition, the relaxation curve of 5,7-DMF was shifted to the left in the presence of only Larginine, with increasing relaxation potency (EC50 = 2.3 ± 0.5 x 10-6 M). Following the downstream pathway of NO, it can be observed that in the presence of PKG inhibitor, Rp-8-Br-PET-cGMPS, the relaxation control curve of 5,7-DMF was shifted to right, with a decrease in vasorelaxant potency (EC50 = 1.1 ± 0.1 x 10-5 M). In view of these results it is concluded that the NO/PKG pathway is involved in the vasorelaxant effect of flavonoid. As some of the targets of PKG are the K+ and Ca2+ channels, it was decided to evaluate their participation in the vasorelaxant effect of 5,7-DMF in moderate or high K+ increases and an equipotency can be observed in relaxing the aorta contracted with KCl 30 (EC50 = 1.5 ± 0.3 x 10-5 M) or 80 mM (EC50 = 2.2 ± 0.2 x 10-5 M), indicating a possible participation of the voltage-dependent Ca2+ channels (Cav). This hypothesis was confirmed by observation of a shift to the right of the cumulative control curves with CaCl2 in depolarizing medium, in a non-parallel manner and with reduction in Emax. In addition, 5,7-DMF relaxed the pre-contracted aorta with S-(-)-Bay K8644, a CaV1 activator (EC50 = 3.5 ± 0.7 x 10-6 M), suggesting blockage of these channels. In view of these results, it was concluded that 5,7-DMF presents a non-selective spasmolytic effect in the organs tested, and in the rat aorta its effect was potentiated when solubilized in Cremophor®, and its mechanism of action is by positive modulation of the NO/PKG pathway and negative modulation of CaV1.