METHODS: We used wire myograph to study NaCl-induced vasorelaxation of mesenteric arteries isolated from mice. Cl-, Ca2+ and K+ imaging was performed in human vascular endothelial cells pre-treated with pharmacological agents.
RESULTS: The Cl- concentration-dependently induced vasorelaxation of mesenteric arteries likely through EDH. The Cl--induced vasorelaxation was attenuated in TRPV4 KO mice and inhibited by selective blockers of Na+-K+-2Cl- cotransporter 1 (NKCC1) (bumetanide, 10 μM), transient receptor potential vanilloid 4 (TRPV4) (RN-1734, 40 μM), and small conductance Ca2+-activated K+ channels (SKCa) (apamin, 3 μM)/ intermediate conductance Ca2+-activated K+ channels (IKCa) (TRAM-34, 10 μM) and myoendothelial gap junction (18α-glycyrrhetinic acid, 10 μM), but enhanced by a selective activator of IKCa/SKCa (SKA-31, 0.3 μM). Cl- decreased intracellular K+ concentrations in endothelial cells, which was reversed by apamin (200 nM) plus TRAM-34 (500 nM). Extracellular Cl- raised intracellular Cl- concentrations in endothelial cells, which was attenuated by bumetanide (10 μM). Finally, Cl- induced a transient Ca2+ signaling via TRPV4 in endothelial cells, which became sustained when the Ca2+ exit mode of Na+-Ca2+ exchanger (NCX) was blocked.
CONCLUSIONS: Cl- induces a pure EDH-mediated vasorelaxation of mesenteric arteries through activation of endothelial NKCC1/TRPV4/NCX axis. We have provided a novel insight into the role of Cl--induced vasorelaxation via EDH mechanism.
方法:我们使用钢丝肌电图研究NaCl诱导的小鼠肠系膜动脉的血管舒张。Cl-,在用药物预处理的人血管内皮细胞中进行Ca2和K成像。
结果:Cl-浓度依赖性诱导肠系膜动脉血管舒张可能通过EDH。在TRPV4KO小鼠中,Cl-诱导的血管舒张作用减弱,并被Na-K-2Cl-协同转运蛋白1(NKCC1)(布美他尼,10μM),瞬时受体电位香草酸4(TRPV4)(RN-1734,40μM),和小电导Ca2+激活的K+通道(SKCa)(阿帕明,3μM)/中等电导Ca2激活的K通道(IKCa)(TRAM-34,10μM)和肌内皮缝隙连接(18α-甘草次酸,10μM),但通过IKCa/SKCa的选择性激活剂(SKA-31,0.3μM)增强。Cl-降低内皮细胞细胞内K+浓度,其被阿帕明(200nM)加TRAM-34(500nM)逆转。细胞外Cl-升高内皮细胞中的细胞内Cl-浓度,通过布美他尼(10μM)减毒。最后,Cl-通过TRPV4在内皮细胞中诱导短暂的Ca2信号传导,当Na+-Ca2+交换剂(NCX)的Ca2+退出模式被阻断时,其变得持续。
结论:Cl-通过激活内皮NKCC1/TRPV4/NCX轴诱导肠系膜动脉的纯EDH介导的血管舒张。我们通过EDH机制对Cl诱导的血管舒张的作用提供了新的见解。