CHANNEL REGULATION IN NORMAL/HYPERTENSIVE SMOOTH MUSCLE

本研究的总体目标是表征 传导通路，并阐明其调节在光滑 从正常和高血压动物中分离的肌细胞 模型 大多数拟议的研究将在单一的 从大鼠和兔分离的动脉平滑肌细胞 肠系膜动脉或主动脉夹层环，这可能是 少量获得，但保持双极松弛 形态学和激动剂诱导的收缩。 膜片钳研究 （全细胞和单通道记录）设计用于 表征单个Ca2+、Na+和K+通道， 来自正常和高血压动物的肌肉细胞。 的 通过血管活性受体激动剂调节离子电流， 受体、第二信使、G 将阐明蛋白质和蛋白激酶和磷酸酶。 荧光指示剂的荧光检测和成像 相同细胞将阐明细胞的时间进程和均一性 细胞质Ca 2+和H+的反应。 这些 方法将扩展到表征离子电导 在维持离体平滑肌期间发生的通路 增殖和非增殖组织培养下的细胞 条件 该提案的主要优势在于应用 一个系统的电生理学研究， 生物化学、生物物理和细胞生物学技术， 从病理学角度研究新鲜分离的组织 培养的动脉平滑肌细胞， 高血压动物模型如自发性SHR京都 高血压模型及血管紧张素诱导的继发性高血压 高血压模型。 该应用程序的长期目标是 了解生理机制和病理偏差 肌膜离子通道在调节血管紧张素转换酶活性中的作用 平滑肌张力通过来自神经元， 体液和局部产生的信号传导元件，和B）评估 如果发生离子传导途径的表型调节 在高血压下，一种已知涉及血管增生的疾病， 平滑肌细胞与研究的细胞相似。

The overall objective of this investigation is to characterize ion conductance pathways and elucidate their regulation in smooth muscle cells isolated from normal and hypertensive animal models. Most of the proposed studies will be carried out on single arterial smooth muscle cells isolated from rat and rabbit mesenteric artery or dissected rings of aorta, which can be obtained in small number but which maintain bipolar relaxed morphology and agonist-induced contraction. Patch clamp studies (whole cell and single channel recording) are designed to characterize individual Ca2+, Na+ and K+ channels in smooth muscle cells from normal and hypertensive animals. The regulation of ionic currents by vasoactive receptor agonists and the modulation of channels by receptors, second messengers, G proteins and protein kinases and phosphatases will be elucidated. Flourometric detection and imaging of fluorescent indicators in the same cells will elucidate time courses and homogeneity of cell response with respect to cytosolic Ca2+ and H+. These approaches will be extended to characterize the ion conductance pathways occurring during maintenance of isolated smooth muscle cells under proliferative and non-proliferative tissue culture conditions. The major strength of the proposal is the application of a systematic electrophysiological investigation coupled with biochemical, biophysical and cell biology techniques and a pathological perspective to the study of freshly isolated and tissue cultured arterial smooth muscle cells from normal and hypertensive animal models such as the SHR Kyoto spontaneous hypertensive model and the angiotensin induced secondary hypertensive models. The long term goal of the application is to understand physiological mechanisms and pathological deviations of a) the role of sarcolemma ion channels in regulating vascular smooth muscle tone in situ by multiple input from neuronal, humoral and locally produced signalling elements and b) to assess if phenotypic modulation of ion conductance pathways occurs under hypertension, a condition known to involve hyperplasia of smooth muscle cells similar to those studied.