Mechanosensation and the basis of myogenic tone development

在动态条件下，动脉张力受多种刺激调节，包括神经元活动、组织代谢和血管内压力。 一个世纪前，贝利斯首次推测动脉可以对血压的变化做出反应。 在各种血管床中可观察到“肌源性反应”，并依赖于平滑肌[Ca 2 +]的升高。 升高的压力通过增强以下各项来增强平滑肌[Ca 2 +]：1）通过动脉去极化驱动的机制的Ca 2+通道活性;和2）来自肌浆网（SR）的内部Ca 2+释放。 虽然过去的研究已经澄清了哪些离子通道和商店参与，但问题仍然是平滑肌细胞如何实际上“感知”血管内压力以促进胞质[Ca 2 +]升高。 为了与这个关键问题保持一致，NSERC的提案将解决粘附复合物蛋白及其下游信号通路是否能够使平滑肌细胞感知机械力并升高胞质[Ca 2 +]。 在接下来的5年里，申请人将研究以下方面的作用： 1. 整合素受体对细胞溶质Ca 2+动力学和肌原性张力的影响;以及 2. 粘着斑激酶（FAK）、Src酪氨酸激酶和磷脂酶C γ（PLC γ 1）对Ca 2+动力学和肌源性张力的影响。 我们的实验方法将协同采用：1）血管肌造影术，以评估血管功能; 2）蛋白质印迹分析，以确定信号蛋白的磷酸化状态; 3）共聚焦显微镜，以测量Ca 2+动力学;和4）膜片钳电生理学，以监测离子通道活性。 整合素受体和下游信号传导蛋白将使用已建立的药理学工具或siRNA方法进行主动操纵。 我们对压力敏感和粘附复合物蛋白的关注将促进对肌源性反应及其对心血管功能的生物学意义的更深层次的机械评价。

Under dynamic conditions, arterial tone is regulated by multiple stimuli including neuronal activity, tissue metabolism and intravascular pressure. A century ago, Bayliss first surmised that arteries could respond to changes in blood pressure. The "myogenic response" is observable in a variety of vascular beds and is dependent upon a rise in smooth muscle [Ca2+]. Elevated pressure augments smooth muscle [Ca2+] by augmenting: 1) Ca2+ channel activity through a mechanism driven by arterial depolarization; and 2) internal Ca2+ release from the sarcoplasmic reticulum (SR). While past studies have clarified which ion channels and stores participate, questions remains as to how smooth muscle cells actually "sense" intravascular pressure to facilitate the cytosolic [Ca2+] rise. In keeping with this key question, this NSERC proposal will address whether adhesion complex proteins and their downstream signaling pathways enable smooth muscle cells to sense mechanical forces and elevate cytosolic [Ca2+]. Over the next 5 years, the applicant will examine the role of: 1. Integrin receptors on cytosolic Ca2+ dynamics and myogenic tone; and 2. Focal adhesion kinase (FAK), Src tyrosine kinases and phospholipase C gamma (PLCy1) on Ca2+ dynamics and myogenic tone. Our experimental approach will synergistically employ: 1) vessel myography to assess vessel function; 2) western analysis to determine the phosphorylation state of signaling proteins; 3) confocal microscopy to measure Ca2+ dynamics; and 4) patch clamp electrophysiology to monitor ion channel activity. Integrin receptors and downstream signaling proteins will be actively manipulated using established pharmacological tools or a siRNA approach. Our focus on pressure-sensing and adhesion complex proteins will foster a deeper mechanistic appreciation of the myogenic response and its biological significance to cardiovascular function.