From the Robert M. Berne Cardiovascular Research Center (K.H., E.L.C., L.J.D., C.M., B.E.I., S.K.S.), Department of Molecular Physiology and Biological Physics (C.M., B.E.I., S.K.S.), and Department of Pharmacology (L.J.D., S.K.S), University of Virginia-School of Medicine, Charlottesville.
Arterioscler Thromb Vasc Biol. 2018 Mar;38(3):542-554. doi: 10.1161/ATVBAHA.117.310038. Epub 2018 Jan 4.
Several physiological stimuli activate smooth muscle cell (SMC) GPCRs (G protein-coupled receptors) to cause vasoconstriction. As a protective mechanism against excessive vasoconstriction, SMC GPCR stimulation invokes endothelial cell vasodilatory signaling. Whether Ca influx in endothelial cells contributes to the regulation of GPCR-induced vasoconstriction remains unknown. Ca influx through TRPV4 (transient receptor potential vanilloid 4) channels is a key regulator of endothelium-dependent vasodilation. We hypothesized that SMC GPCR stimulation engages endothelial TRPV4 channels to limit vasoconstriction.
Using high-speed confocal microscopy to record unitary Ca influx events through TRPV4 channels (TRPV4 sparklets), we report that activation of SMC αARs (alpha-adrenergic receptors) with phenylephrine or thromboxane A receptors with U46619 stimulated TRPV4 sparklets in the native endothelium from mesenteric arteries. Activation of endothelial TRPV4 channels did not require an increase in Ca as indicated by the lack of effect of L-type Ca channel activator or chelator of intracellular Ca EGTA-AM. However, gap junction communication between SMCs and endothelial cells was required for phenylephrine activation or U46619 activation of endothelial TRPV4 channels. Lowering inositol 1,4,5-trisphosphate levels with phospholipase C inhibitor or lithium chloride suppressed phenylephrine activation of endothelial TRPV4 sparklets. Moreover, uncaging inositol 1,4,5-trisphosphate profoundly increased TRPV4 sparklet activity. In pressurized arteries, phenylephrine-induced vasoconstriction was followed by a slow, TRPV4-dependent vasodilation, reflecting activation of negative regulatory mechanism. Consistent with these data, phenylephrine induced a significantly higher increase in blood pressure in TRPV4 mice.
These results demonstrate that SMC GPCR stimulation triggers inositol 1,4,5-trisphosphate-dependent activation of endothelial TRPV4 channels to limit vasoconstriction.
几种生理刺激激活平滑肌细胞（SMC）G 蛋白偶联受体（GPCR），引起血管收缩。作为防止过度血管收缩的保护机制，SMC GPCR 刺激引发内皮细胞血管舒张信号。内皮细胞中钙内流是否有助于调节 GPCR 诱导的血管收缩尚不清楚。通过瞬时受体电位香草酸 4（TRPV4）通道的钙内流是内皮依赖性血管舒张的关键调节剂。我们假设 SMC GPCR 刺激使内皮 TRPV4 通道参与限制血管收缩。
使用高速共聚焦显微镜记录 TRPV4 通道（TRPV4 火花）的单元钙内流事件，我们报告说，用苯肾上腺素激活 SMC αAR（α-肾上腺素能受体）或用 U46619 激活血栓烷 A 受体可刺激肠系膜动脉中天然内皮的 TRPV4 火花。内皮 TRPV4 通道的激活不需要钙的增加，这表明 L 型钙通道激活剂或细胞内钙螯合剂 EGTA-AM 没有作用。然而，SMC 和内皮细胞之间的缝隙连接通讯对于苯肾上腺素激活或 U46619 激活内皮 TRPV4 通道是必需的。用磷脂酶 C 抑制剂或氯化锂降低肌醇 1,4,5-三磷酸水平可抑制苯肾上腺素激活的内皮 TRPV4 火花。此外，肌醇 1,4,5-三磷酸的光解极大地增加了 TRPV4 火花的活性。在加压动脉中，苯肾上腺素引起的血管收缩后是缓慢的、TRPV4 依赖性的血管舒张，反映了负调节机制的激活。与这些数据一致，苯肾上腺素在 TRPV4 小鼠中引起血压显著升高。
这些结果表明，SMC GPCR 刺激触发肌醇 1,4,5-三磷酸依赖性激活内皮 TRPV4 通道，以限制血管收缩。
