Abstract：
OBJECTIVE: Although absorbed NaCl increases intestinal blood flow to facilitate absorption and transportation, it is unclear if it can directly mediate mesenteric arterial relaxation. We aimed to investigate and test our hypothesis that Cl- induces mesenteric arterial vasorelaxation via endothelium-dependent hyperpolarization (EDH).
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.
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.
摘要：
目的：尽管吸收的NaCl会增加肠血流量以促进吸收和运输，目前尚不清楚它是否能直接介导肠系膜动脉舒张。我们旨在研究和检验我们的假设，即Cl-通过内皮依赖性超极化（EDH）诱导肠系膜动脉血管舒张。
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。
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。
