Epoxyeicosatrienoic acids (EETs) are endothelium-derived arachidonic acid metabolites of cytochrome P450. They dilate coronary arteries, open K+ channels, and hyperpolarize vascular smooth muscles. However, the mechanisms of these smooth muscle actions remain unknown. This study examined the effects of EETs on the large-conductance Ca(2+)-activated K+ channel (KCa) in smooth muscle cells of small bovine coronary arteries. In cell-attached patch-clamp experiments, 11,12-EET produced a 0.5- to 10-fold increase in the activity of the KCa channels when added in concentrations of 1, 10, and 100 nmol/L. In the inside-out excised membrane patch mode, 11,12-EET was without effect on the activity of the KCa channel unless GTP (0.5 mmol/L) or GTP and ATP (1 mmol/L) were added to the bath solution. In the presence of GTP and ATP, the increase in the KCa channel activity with 11,12-EET in inside-out patches was comparable to that in cell-attached patches. This effect of 11,12-EET in inside-out patches was blocked by the addition of GDP-beta-S (100 mumol/L). In outside-out patches, 11,12-EET also increased the KCa channel activity when GTP and ATP were added to the pipette solution. The addition of a specific anti-Gs alpha antibody (100 nmol/L) in the pipette solution completely blocked the activation of the KCa channels induced by 11,12-EET. An anti-G beta gamma or anti-Gi alpha antibody was without effect. We conclude that 11,12-EET activates the KCa channels by a Gs alpha-mediated mechanism. This mechanism contributes to the effects of EETs as endothelium-derived hyperpolarizing factors to hyperpolarize and relax arterial smooth muscle.

译文

环氧二十碳三烯酸(EET)是内皮细胞色素P450的花生四烯酸代谢产物。它们扩张冠状动脉,打开K通道,并使血管平滑肌超极化。但是,这些平滑肌动作的机制仍然未知。这项研究检查了EETs对小牛冠状动脉平滑肌细胞中大电导Ca(2)激活的K通道(KCa)的影响。在贴有细胞的膜片钳实验中,当以1,10和100 nmol / L的浓度添加时,11,12-EET使KCa通道的活性增加0.5至10倍。在由内而外的切膜模式下,除非将GTP(0.5 mmol / L)或GTP和ATP(1 mmol / L)添加到浴液中,否则11,12-EET对KCa通道的活性没有影响。在存在GTP和ATP的情况下,由内而外的贴片中11,12-EET的KCa通道活性的增加与细胞附着的贴片中的KCa通道活性的增加相当。通过添加GDP-β-S(100 mumol / L),阻止了由内而外的11,12-EET的这种作用。在外向斑块中,将GTP和ATP添加到移液器中时,11,12-EET也增加了KCa通道活性。在移液器中添加特异性抗Gsα抗体(100 nmol / L)完全阻断了11,12-EET诱导的KCa通道的激活。抗Gβγ或抗Giα抗体无效。我们得出的结论是11,12-EET通过Gs alpha介导的机制激活了KCa通道。这种机制有助于将EETs作为内皮源的超极化因子来使动脉平滑肌超极化和松弛。

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