Angiotensin II activates two cation conductances with distinct TRPC1 and TRPC6 channel properties in rabbit mesenteric artery myocytes
Angiotensin II activates two cation conductances with distinct TRPC1 and TRPC6 channel properties in rabbit mesenteric artery myocytes
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DOI：
10.1113/jphysiol.2006.119305
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发表时间：
2006-12-01
影响因子：
5.5
通讯作者：
Large, W. A.
中科院分区：
文献类型：
--
作者：
Saleh, S. N.;Albert, A. P.;Large, W. A.
血管紧张素II（Ang II）是一种有效的血管收缩剂，在控制血压中具有重要作用;然而，关于Ang II引起的血管收缩反应的细胞机制的信息很少。本研究旨在应用膜片钳技术在单通道水平研究血管紧张素Ⅱ对新鲜分离的兔肠系膜动脉心肌细胞阳离子电导的影响。在细胞贴附贴片中，低浓度Ang II（1 nM）的浴应用激活了电导状态约为15、30和45 pS的阳离子通道电流（I-cat 1）。在相对较高的浓度下，Ang II（100 nM）抑制I-cat 1，但诱发另一个电导约为2 pS的阳离子通道（I-cat 2）。血管紧张素II诱导的I-cat 1和I-cat 2可被AT（1）受体拮抗剂氯沙坦和磷脂酶C（PLC）抑制剂U 73122抑制。二酰基甘油（DAG）脂肪酶抑制剂RHC 80267最初诱导I-cat 1，随后被抑制，显示I-cat 2。DAG类似物1-油酰基-2-乙酰基-sn-甘油（1 μ M）激活I-cat 1和I-cat 2，但肌醇1，4，5-三磷酸没有引起任何电导。蛋白激酶C（PKC）抑制剂白屈菜红碱（3 μ M）增强血管紧张素II诱导的I-cat 1和抑制I-cat 2，而PKC激活剂佛波醇-12，13-二丁酸酯（1 μ M）减少血管紧张素II诱导的I-cat 1，但激活I-cat 2。此外，在用白屈菜红碱预处理的细胞贴附贴片中，应用100 nM Ang II激活I-cat 1。这些数据表明，PKC抑制I-cat 1，但刺激I-cat 2。消耗细胞内Ca 2+储存的试剂也激活具有与I-cat 2相似性质的阳离子通道电流。将抗TRPC 6和抗TRPC 1抗体浴施用于由内而外的贴片，分别抑制I-cat 1和I-cat 2。此外，氟芬那酸和零外部Ca 2+浓度，分别增强和减少血管紧张素II诱导的I-cat 1。免疫细胞化学研究显示TRPC 6和TRPC 1的表达，TRPC 6优先分布在质膜和TRPC 1的表达位于整个肌细胞。这些结果表明，血管紧张素Ⅱ激活两个不同的阳离子电导在肠系膜动脉肌细胞的AT（1）受体的刺激连接PLC。I-cat 1通过PKC非依赖性机制被DAG激活，而I-cat 2涉及通过PKC依赖性途径起作用的DAG。高浓度的Ang II通过激活PKC的抑制作用抑制I-cat 1。TRPC 6和TRPC 1通道蛋白分别是Ang II诱导的I-cat 1和I-cat 2的重要组成部分。
Angiotensin II (Ang II) is a potent vasoconstrictor with an important role in controlling blood pressure; however, there is little information on cellular mechanisms underlying Ang II-evoked vasoconstrictor responses. The aim of the present study is to investigate the effect of Ang II on cation conductances in freshly dispersed rabbit mesenteric artery myocytes at the single-channel level using patch-clamp techniques. In cell-attached patches, bath application of low concentrations of Ang II (1 nM) activated cation channel currents (I-cat1) with conductances states of about 15, 30 and 45 pS. At relatively high concentrations, Ang II (100 nM) inhibited I-cat1 but evoked another cation channel (I-cat2) with a conductance of approximately 2 pS. Ang II-evoked I-cat1 and I-cat2 were inhibited by the AT(1) receptor antagonist losartan and the phospholipase C (PLC) inhibitor U73122. The diacylglycerol (DAG) lipase inhibitor RHC80267 initially induced I-cat1 which was subsequently inhibited to reveal I-cat2. The DAG analogue 1-oleoyl-2-acetyl-sn-glycerol (1 mu M) activated I-cat1 and I-cat2 but inositol 1,4,5-trisphosphate did not evoke either conductance. The protein kinase C (PKC) inhibitor chelerythrine (3 mu M) potentiated Ang II-evoked I-cat1 and inhibited I-cat2 whereas the PKC activator phorbol-12,13-dibutyrate (1 mu M) reduced Ang II-induced I-cat1 but activated I-cat2. Moreover in cell-attached patches pretreated with chelerythrine, application of 100 nM Ang II activated I-cat1. These data indicate that PKC inhibits I-cat1 but stimulates I-cat2. Agents that deplete intracellular Ca2+ stores also activated cation channel currents with similar properties to I-cat2. Bath application of anti-TRPC6 and anti-TRPC1 antibodies to inside-out patches inhibited I-cat1 and I-cat2, respectively. Also flufenamic acid and zero external Ca2+ concentration, respectively, potentiated and reduced Ang II-evoked I-cat1. Immunocytochemical studies showed TRPC6 and TRPC1 expression with TRPC6 preferentially distributed in the plasma membrane and TRPC1 expression located throughout the myocyte. These results indicate that Ang II activates two distinct cation conductances in mesenteric artery myocytes by stimulation of AT(1) receptors linked to PLC. I-cat1 is activated by DAG via a PKC-independent mechanism whereas I-cat2 involves DAG acting via a PKC-dependent pathway. Higher concentrations of Ang II inhibit I-cat1 by activating an inhibitory effect of PKC. It is proposed that TRPC6 and TRPC1 channel proteins are important components of Ang II-induced I-cat1 and I-cat2, respectively.