Effects of pinacidil on contractile proteins in high K(+)-treated intact, and in beta-escin-treated skinned smooth muscle of the rabbit mesenteric artery

Abstract

1. The effects of pinacidil were investigated on changes in cellular Ca2+ concentration ([Ca2+]i) and tension in intact and chemically skinned smooth muscle strips of the rabbit mesenteric artery. 2. High K+ (128 mM) produced a large phasic followed by a tonic increase in [Ca2+]i and tension in intact muscle strips. Pinacidil at 10 microM but not 1 microM, inhibited the phasic and tonic contractions induced by 128 mM K+ without a corresponding change in [Ca2+]i. 3. In beta-escin-treated skinned smooth muscle, the minimum Ca2+ concentration that produced contraction was 0.1 microM and the maximum contraction was obtained at 10 microM. Pinacidil at 10 microM but not 1 microM, shifted the pCa-tension relation curve to the right and also inhibited the maximum contraction induced by Ca2+. The concentrations of Ca2+ required for half maximal tension were 0.9 microM in control and 1.5 microM in the presence of 10 microM pinacidil. Calmodulin (2 microM) increased the contraction induced by 0.3 microM Ca2+ (but not by 10 microM Ca2+) in the skinned strips. Pinacidil (10 microM) inhibited the contraction induced by 0.3 microM or 10 microM Ca2+ in the presence of 2 microM calmodulin. 4. Noradrenaline (NA, 10 microM) with guanosine triphosphate (GTP, 3 microM), guanosine 5'-O-(3-thiotriphosphate) (GTP gamma S, 3 microM) or 12-O-tetradecanoylphorbol-13-acetate (TPA, 0.1 microM) all enhanced the contraction induced by 0.3 microM Ca2+. Pinacidil (10 microM) inhibited the contraction induced by 0.3 microM Ca2+ more strongly in the presence of the above agents than in their absence. 5. Following application of 2 mm adenosine-5'-O-(3-thiotriphosphate) (ATPyS) with 0.3 microM Ca2", 4mM MgATP produced contraction in skinned strips in Ca2 -free solution containing 4mM EGTA ('Ca2+- independent contraction'). The amplitude of the Ca2 +-independent contraction was almost the same as that obtained with 1O microM Ca2 . Pinacidil (1O microM) had no effect on the amplitude of the Ca2+-independent contraction nor did it have any effect on the contraction induced by a solution containing no MgATP ('rigor contraction'). 6. It is concluded that pinacidil (10 microM) acts directly on the contractile apparatus to inhibit Ca2'-induced contraction in smooth muscle of the rabbit mesenteric artery. The site of action of pinacidil may be between Ca2 +-calmodulin complex formation and phosphorylation of the myosin light chain.