A ação relaxante do flavonoide 4',5,7-triidroxi-3,6-dimetoxiflavona, isolado de Piptadenia stipulacea (Benth.) Ducke, envolve modulação positiva de canais de potássio e redução dos níveis citosólicos de cálcio em íleo de cobaia

• Autor(a) principal: Vasconcelos, Luiz Henrique César
• Data de Publicação: 2013
• Tipo de documento: Dissertação
• Idioma: por
• Título da fonte: Biblioteca Digital de Teses e Dissertações da UFPB
• Texto Completo: https://repositorio.ufpb.br/jspui/handle/tede/8066
• topic: Piptadenia stipulacea; Piptadenia stipulacea; 4’,5,7-triidroxi-3,6-dimetoxiflavona; Ação relaxante; Cálcio; Canais iônicos; 4’,5,7-triidroxi-3,6-dimetoxiflavone; Relaxant action; Ion channels; Calcium; CIENCIAS BIOLOGICAS::FARMACOLOGIA

Piptadenia stipulacea (Benth.) Ducke (Fabaceae) is a typical tree of Caatinga, popularly known as “jurema-branca”, “jurema-malícia-da-serra”, “carcará” and “calumbi” and is popularly used as heling agent and antiinflamatory. From its aerial parts was isolated the flavonoid 4’,5,7-triidroxi-3,6-dimetoxiflavona (FGAL) that, in previous studies, inhibited both CCh- and histamine-induced phasic contractions on guinea pig ileum. Thus, the aim of this work was to characterize its relaxant mechanism of action. Isotonic and isometric contractions were recorded to determine and compare the relative efficacy and potency. The myocites viability was measured by the MTT assay, and the cytosolic Ca2+ levels by the analysis of fluorescence of fluo-4. The flavonoid relaxed the ileum pre-contracted with KCl 40 mM (EC50 = 2.6 ± 0.5 x 10-6 M) or CCh 10-5 M (EC50 = 1.8 ± 0.4 x 10-6 M), being more potent when the ileum was pre-contracted with histamine 10-6 M (EC50 = 1.9 ± 0.4 x 10-7 M). In addition, the flavonoid righward shifted the cumulative concentration-response curves of histamine in a non-parallel manner, with maximum effect (Emax) reduction, presenting a profile of non-competitive pseudoirreversible antagonism. To verify if FGAL would inhibit the Ca2+ influx through the voltage-sensitive Ca2+ channels (CaV), cumulative concentration-response curves of CaCl2 in depolarizing medium (70 mM KCl) nominally without Ca2+ were obtained in both the absence (control) and presence of different concentrations of FGAL. The flavonoid righward shifted the CaCl2 contraction curves in a non-parallel manner, with Emax reduction. Moreover, FGAL relaxed the pre-contracted ileum with S-(-)-Bay K8644 (3 x 10-7 M), a CaV1 agonist, but with lower potency than with KCl or histamine, indicating an indirect blockade of these channels. Then, in order to verify whether FGAL would be positivelly modulating the K+ channels to, indirectally, block the CaV1, it was employed CsCl, a non-selective K+ channels blocker. The relaxant potency of FGAL was attenauted in the presence of CsCl (EC50 = 1.1 ± 0.3 x 10-6 M) suggesting the involvement of these channels on this relaxant effect. In contrast, the relaxant potency of FGAL was not modified in the presence of apamin, SKCa blocker (EC50 = 1.6 ± 0.3 x 10-7 M), or TEA+ 1 mM, BKCa blocker (EC50 = 2.0 ± 1.0 x 10-7 M), discarding the participation of these subtypes of K+ channels. However, in the presence of 4-AP, KV blocker (EC50 = 1.8 ± 0.2 x 10-6 M), and glibenclamide, KATP blocker (EC50 = 1.5 ± 0.5 x 10-6 M), the relaxant potency of FGAL was attenuated about 10 and 8 times, respectively, confirming that FGAL positivelly modulates these subtypes of K+ channels to relax the guinea pig ileum. In the cellular experiments, the viability of intestinal myocytes was not altered in the presence of FGAL (10-4 M). Furthermore, the fluorescence intensity emmited by fluo-4 of myocytes stimulated with histamine was attenuated by FGAL as a result of [Ca2+]c reduction. Therefore, the relaxant mechanism of action of FGAL on guinea pig ileum involves the positive modulation of KV and KATP, which, indirectly, reduces the Ca2+ influx through CaV1, leading to the reduction of the cytosolic levels of this ion.