Tsunobuchi-Ushijima H, Okuno H, Gomi Y
Department of Pharmacology, Faculty of Pharmaceutical Sciences, Kanazawa University, Japan.
Naunyn Schmiedebergs Arch Pharmacol. 2000 Sep;362(3):290-4. doi: 10.1007/s002100000271.
We have reported that noradrenaline but not K+ induced a sustained and dose-dependent contraction without extracellular Ca2+ and Mn2+ in Ca2+-depleted Mn2+loaded vas deferens from the guinea pig. The Mn2+dependent noradrenaline-induced contractions developed without an increase in phosphorylation of 20-kDa myosin light chain. To clarify whether such an unusual Mn2+dependent contraction was induced only by noradrenaline or not, we examined effects of acetylcholine on Ca2+-depleted Mn2+-loaded guinea pig vas deferens. Acetylcholine (1 microM(-1) mM) induced a sustained dose-dependent contraction without extracellular Ca2+ or Mn2+. W-7 (10 microM or 100 microM) and wortmannin (1 microM) both reduced the Mn2+dependent acetylcholine-induced contractions similarly to Ca2+-dependent acetylcholine-induced contractions in isolated vas deferens without either Ca2+ depletion or Mn2+ loading. However, the Mn2+-dependent acetylcholine-induced contractions developed without a significant increase in the phosphorylation of the myosin light chain determined by urea-glycerol polyacrylamide gel electrophoresis and immunoblotting. These results indicate that acetylcholine as well as noradrenaline induces Mn2+-dependent contraction and are consistent with our previous assumption that Mn2+ may preferentially support receptor-mediated contractions in the guinea pig isolated vas deferens. The results also suggest that the activation of myosin light chain kinase is essential for the development of Mn2+-dependent acetylcholine-induced contractions, and that Mn2+ may accelerate formation of non-phosphorylated attached cross-bridges during receptor activation.
我们曾报道，在去除细胞外钙并加载锰的豚鼠输精管中，去甲肾上腺素而非钾离子能诱导出持续且剂量依赖性的收缩，且无需细胞外钙和锰。依赖锰的去甲肾上腺素诱导的收缩在20 kDa肌球蛋白轻链磷酸化未增加的情况下发生。为了阐明这种不寻常的依赖锰的收缩是否仅由去甲肾上腺素诱导，我们研究了乙酰胆碱对去除细胞外钙并加载锰的豚鼠输精管的影响。乙酰胆碱（1微摩尔/升 - 1毫摩尔/升）在无细胞外钙或锰的情况下诱导出持续的剂量依赖性收缩。W - 7（10微摩尔或100微摩尔）和渥曼青霉素（1微摩尔）在未去除钙或加载锰的离体输精管中，与依赖钙的乙酰胆碱诱导的收缩类似，都同样降低了依赖锰的乙酰胆碱诱导的收缩。然而，通过尿素 - 甘油聚丙烯酰胺凝胶电泳和免疫印迹测定，依赖锰的乙酰胆碱诱导的收缩在肌球蛋白轻链磷酸化未显著增加的情况下发生。这些结果表明，乙酰胆碱与去甲肾上腺素一样能诱导依赖锰的收缩，这与我们之前的假设一致，即锰可能优先支持豚鼠离体输精管中受体介导的收缩。结果还表明，肌球蛋白轻链激酶的激活对于依赖锰的乙酰胆碱诱导的收缩的发生至关重要，并且锰可能在受体激活过程中加速非磷酸化附着横桥的形成。
