Department of Physiological Science and Molecular Biology, Fukuoka Dental College, Fukuoka, Japan; Fukuoka School of Medical Sciences, Fukuoka, Japan.
Eur J Pharmacol. 2013 Nov 15;720(1-3):255-63. doi: 10.1016/j.ejphar.2013.09.076. Epub 2013 Oct 22.
We examined the endothelin-1 (ET-1)-induced increase in the intracellular free Ca(2+) concentration ([Ca(2+)]i) in fura-2-loaded rat pulmonary small arteries. ET-1 (30 nM) elicited a long-lasting increase in [Ca(2+)]i in physiological salt solution (PSS). In subsequent experiments, arteries were pretreated with BQ-788, an ETB-specific blocker, to allow us to focus on responses mediated via the ETA receptor, the existence of which was confirmed by immunohistochemistry. In Ca(2+)-free PSS, ET-1 evoked a small transient increase in [Ca(2+)]i, indicating Ca(2+) release from the SR (sarcoplasmic reticulum). After a switch to PSS (containing 2mM CaCl2), ET-1 elicited a long-lasting increase in [Ca(2+)]i that was not inhibited by 1 μM nicardipine, an L-type Ca(2+)-channel inhibitor, suggesting involvement of a Ca(2+)-influx pathway independent of that channel. In arteries preincubated with 30 μM cyclopiazonic acid (CPA) or 2 μM thapsigargin (TG), the ET-1-induced Ca(2+)-release was greatly reduced, and the induced Ca(2+)-influx was attenuated. U-73122, a phospholipase C (PLC) inhibitor, had inhibitory effects similar to those of CPA and TG on the ET-1-induced Ca(2+)-release and Ca(2+)-influx, whereas U-73343, an inactive analogue of U-73122, had no such effects. Two putative membrane-permeable IP3-receptor blockers, 2-aminoethoxydiphenyl borate (2APB, 50 μM) and Xestospongin C (20 μM), (a) almost completely inhibited the ET-1-induced Ca(2+)-release and Ca(2+)-influx, and (b) reduced the ET-1-induced contraction. These results indicate that in rat pulmonary small arteries, ET-1 induces receptor-operated Ca(2+) influx via the ETA receptor, and that this influx interacts with InsP3-receptor activation.
我们检测了 ET-1 诱导的 fura-2 负载的大鼠肺小动脉细胞内游离 Ca(2+)浓度 ([Ca(2+)]i)的增加。ET-1（30 nM）在生理盐溶液（PSS）中引起 [Ca(2+)]i 的持久增加。在随后的实验中，用 BQ-788 预处理动脉，BQ-788 是一种 ETB 特异性阻断剂，可使我们专注于通过 ETA 受体介导的反应，免疫组织化学证实了 ETA 受体的存在。在无 Ca(2+)的 PSS 中，ET-1 引起 [Ca(2+)]i 的短暂增加，表明 Ca(2+)从肌浆网（SR）中释放。当切换到含有 2mM CaCl2 的 PSS 后，ET-1 引起 [Ca(2+)]i 的持久增加，1 μM 尼卡地平（一种 L 型 Ca(2+)通道抑制剂）不能抑制这种增加，提示存在一种不依赖于该通道的 Ca(2+)内流途径。在预先用 30 μM 环匹阿尼酸（CPA）或 2 μM 他帕司琼（TG）孵育的动脉中，ET-1 诱导的 Ca(2+)释放大大减少，诱导的 Ca(2+)内流减弱。PLC 抑制剂 U-73122 对 ET-1 诱导的 Ca(2+)释放和 Ca(2+)内流的抑制作用与 CPA 和 TG 相似，而 U-73122 的无活性类似物 U-73343 则没有这种作用。两种潜在的膜通透性 IP3 受体阻断剂，2-氨基乙氧基二苯硼酸盐（2APB，50 μM）和 Xestospongin C（20 μM），（a）几乎完全抑制 ET-1 诱导的 Ca(2+)释放和 Ca(2+)内流，（b）减少 ET-1 诱导的收缩。这些结果表明，在大鼠肺小动脉中，ET-1 通过 ETA 受体诱导受体操纵的 Ca(2+)内流，并且这种内流与 InsP3 受体的激活相互作用。
