NADPH oxidase signaling, blood pressure sensing and the development of myogenic tone

在动态条件下，动脉张力受到多种刺激的调节，包括神经元活动、组织代谢和血管内压力。一个世纪前，贝利斯首次观察到动脉可以对血压的变化做出反应。“肌源性反应”在各种血管床中都有记载，它依赖于平滑肌[Ca2+]的增加。升高的压力通过激活信号转导通路来增强平滑肌[Ca2+]: 1)使平滑肌去极化并提高Ca2+通道活性；2)调动内部Ca2+储存。虽然已经确定了一些驱动肌原性张力发展的离子通道，但尚未确定能够调节压力的信号级联。目前的工作主要集中在标准的g蛋白偶联机制上，很少有人冒险“跳出这个信号框”去考虑新的分子。这包括活性氧，其产生可以由NADPH氧化酶（NOX）促进，已知与机械敏感蛋白相关的酶。***与我们对肌原性反应的长期兴趣一致，NSERC的提案将探索脑循环中血管内压力、氮氧化物信号和肌原性张力之间的联系。三个目的是：***1)确定NOX是否促进肌张力。***2)确定NOX是否通过机电耦合促进肌张力。***3)确定NOX是否调节对机械刺激敏感的离子通道。***我们的总体假设表明，NOX（特别是NOX1）通过产生H2O2和激活PKC来增强血管平滑肌中TRPC6和/或TRPM4的活性。这种去极化电流的增加将促进压力诱导的去极化，这一事件将提高细胞质[Ca2+]和肌原性张力。我们的实验方法将协同使用：1)血管肌图评估张力、膜电位和细胞质[Ca2+]， 2) Q-PCR和免疫组织化学确定NOX的表达和定位；3)电生理测定离子通道活性。实验将在大鼠、NOX1-/-小鼠和人类分离的脑动脉中进行。我们对压力传感和氮氧化物信号的关注将促进对肌生成反应及其对血压和血流调节的生物学意义的更深层次的机制欣赏。

Under dynamic conditions, arterial tone is regulated by multiple stimuli including neuronal activity, tissue metabolism and intravascular pressure. A century ago, Bayliss first observed that arteries could respond to changes in blood pressure. The “myogenic response” has been documented in various vascular beds and is dependent upon a rise in smooth muscle [Ca2+]. Elevated pressure augments smooth muscle [Ca2+] by activating signal transduction pathways that: 1) depolarize smooth muscle and elevate Ca2+ channel activity; and 2) mobilize internal Ca2+ stores. While some of the ion channels driving myogenic tone development have have been identified, the signaling cascades which enable pressure modulation have not. Present day work typically focuses on standard G-protein coupled mechanisms and few venture “outside this signaling box” to consider novel molecules. This includes reactive oxygen species whose production could be facilitated by NADPH oxidases (NOX), enzymes known to associate with mechanosensitive proteins.***Consistent with our long term interests in the myogenic response, this NSERC proposal will explore the linkage between intravascular pressure, NOX signaling and myogenic tone in the cerebral circulation. The three aims will:***1) Determine whether NOX facilitates myogenic tone. ***2) Determine whether NOX facilitates myogenic tone via electromechanical coupling.***3) Determine whether NOX modulates ion channels sensitive to mechanical stimuli.***Our overarching hypothesis states that NOX (in particular NOX1) augments TRPC6 and/or TRPM4 activity in vascular smooth muscle by generating H2O2 and activating PKC. This increase in depolarizing current will facilitate pressure-induced depolarization, an event that will elevate cytosolic [Ca2+] and myogenic tone. Our experimental approach will synergistically employ: 1) vessel myography to assess tone, membrane potential, and cytosolic [Ca2+], 2) Q-PCR and immunohistochemistry to determine NOX expression and localization; and 3) electrophysiology to ascertain ion channel activity. Experiments will be performed in cerebral arteries isolated from rat, NOX1-/- mice and humans. Our focus on pressure-sensing and NOX signaling will foster a deeper mechanistic appreciation of the myogenic response and its biological significance to blood pressure and blood flow regulation.