RESUMO
The objective of this study was to determine the vasodilating effect of 3beta-hydroxy-5-spirostene (diosgenin), a phytoestrogen found in wild yams, using porcine resistance left anterior descending coronary artery. In 5-hydroxytryptamine (3 microM) pre-contracted preparation, diosgenin caused a concentration-dependent (0.01 to 1 microM), endothelium-independent relaxation, with a maximum relaxation of approximately 72% at 1 microM. No apparent effect was observed with 17beta-oestradiol and progesterone with concentrations < or =0.3 microM, and a relaxation of approximately 15% and approximately 23% caused by 17beta-oestradiol (1 microM) and progesterone (1 microM), respectively. Diosgenin-elicited relaxation was not altered by 7alpha,17beta-[9[(4,4,5,5,5-pentafluoropentyl)sulfinyl]nonyl]estra-1,3,5(10)-triene-3,17-diol (ICI 182,780), mifepristone, (+)-bicuculline, cis-N-(2-phenylcyclopentyl)azacyclotridec-1-en-2-amine (MDL 12330A), glibenclamide and scavengers of reactive oxygen species. The iberiotoxin-sensitive, Ca2+-activated K+ (BK(Ca)) current of single vascular myocytes recorded, using patch-clamp techniques, was markedly enhanced by diosgenin, 17beta-oestradiol and progesterone. Application of (9S, 10R, 12R)-2,3,9,10,11,12-hexahydro-10-methoxy-2,9-dimethyl-1-oxo-9,12-epoxy-1H-diindolo[1,2,3-fg:3',2',1'-kl]pyrrolo[3,4-i][1,6]benzodiazocine-10-carboxylic acid methyl ester (KT 5823, 300 nM) eradicated the enhancement of BK(Ca) amplitude. Diosgenin, 17beta-oestradiol and progesterone did not affect whereas phloretin, biochanin A and zearalanone (1 microM each) significantly suppressed [Ca2+]o-induced contraction. In oestrogen competition essay using human breast cancer cell (MCF-7 cells), diosgenin (0.001 nM to 10 microM) did not interact with oestrogen receptor-alpha, and no displacement of [3H]17beta-oestradiol was observed. In oestrogen receptor alpha- and beta-fluorescence polarization competitor assay, diosgenin (100 microM) demonstrated a greater competition with the beta-isoform of oestrogen receptor. These results suggest that diosgenin caused an acute, endothelium-independent coronary artery relaxation via protein kinase G signalling cascade and an activation of BK(Ca) channel of arterial smooth muscle cells. The oestrogen receptor (alpha and beta-isoforms) and progesterone receptor are probably not involved.
Assuntos
Vasos Coronários/efeitos dos fármacos , Diosgenina/farmacologia , Peptídeos/farmacologia , Canais de Potássio Cálcio-Ativados/fisiologia , Potenciais de Ação/efeitos dos fármacos , Potenciais de Ação/fisiologia , Animais , Vasos Coronários/fisiologia , Diosgenina/química , Relação Dose-Resposta a Droga , Feminino , Técnicas In Vitro , Masculino , Suínos , Vasoconstrição/efeitos dos fármacos , Vasoconstrição/fisiologia , Vasodilatação/efeitos dos fármacos , Vasodilatação/fisiologiaRESUMO
In this study, we have evaluated the underlying mechanisms responsible for the relaxation response of ligustrazine (2,3,5,6-tetra-methyl-pyrazine; 2,3,5,6-MP) and its structural analogues (2-methyl-pyrazine (2-MP); ethyl-pyrazine (EP); 2,3-di-methyl-pyrazine (2,3-MP); 2,5-di-methyl-pyrazine (2,5-MP); 2,6-di-methyl-pyrazine (2,6-MP) and 2,3,5-tri-methyl-pyrazine (2,3,5-MP)) in porcine left anterior descending coronary artery (tertiary branch, O.D. =1 mm). In 5-hydroxytryptamine (3 microM) precontracted preparations, cumulative administration (0.1-300 microM) of all pyrazine analogues caused an endothelium-independent, concentration-dependent relaxation. The relative inhibitory potency, as compared at concentration with which 50% relaxation occurred, was 2,3,5,6-MP>2,3,5-MP>EP>2,5-MP>/=2,6-MP>/=2,3-MP>2-MP. Besides, salbutamol and forskolin caused an endothelium-independent relaxation. The relaxation response of ligustrazine, salbutamol and forskolin was blunted in the presence of cis-N-(2-phenylcyclopentyl) azacyclotridec-1-en-2-amine (MDL 12330A) (10 microM, an adenylate cyclase inhibitor) and N-[2-((bromocinnamyl)amino)ethyl]-5-isoquinoline-sulphonamide (H-89, a protein kinase A inhibitor, 3 microM). Patch-clamp, whole-cell electrophysiological studies using single smooth muscle cells of the left anterior descending coronary artery revealed that ligustrazine (300 microM), salbutamol (30 microM) and forskolin (1 microM) inhibited the nifedipine-sensitive L-type Ca(2+) channels, and the inhibitory effect was eradicated by MDL 12330A (10 microM) and H-89 (1 microM). However, neither the Ca(2+)-dependent K(+) channel nor the ATP-dependent K(+) channel was modified by ligustrazine (300 microM). In conclusion, our results indicate that ligustrazine-mediated left anterior descending coronary artery relaxation is due to the activation of adenylate cyclase/protein kinase A cascade and the subsequent inhibition of nifedipine-sensitive, voltage-dependent L-type Ca(2+) channels. However, opening of K(+) channels seems to play no role in mediating the relaxation effect of ligustrazine.