Zhu Z, Arendshorst W J
Department of Physiology, University of North Carolina at Chapel Hill 27599-7545, USA.
Am J Physiol. 1996 Dec;271(6 Pt 2):F1239-47. doi: 10.1152/ajprenal.1996.271.6.F1239.
This study provides an initial characterization of basic morphological properties of cultures of vascular smooth muscle cells (VSMC) from rat preglomerular resistance vessels and of the functional coupling of angiotensin II (ANG II) receptors to cytosolic free calcium concentration ([Ca2+]i (fura 2 fluorescence photometry). Renal VSMC were isolated from interlobular arteries and afferent arterioles (< 50 microns) using an iron oxide sieving method and compared with rat aortic VSMC cultured under similar conditions. Quiescent monolayers maintained uniform morphology and [Ca2+]i signaling profile between passages 3 and 10. Arteriolar and aortic VSMC were spindle shaped and expressed smooth muscle-specific alpha-actin and myosin heavy chains SM-1 and SM-2. ANG II caused a rapid increase in [Ca2+]i, followed by a sustained plateau phase at 50-60% of the peak value. The initial maximum [Ca2+]i responses were dose dependent and of similar magnitude in renal arteriolar and aortic VSMC. ANG II (10(-7) M) increased [Ca2+]i from 50 to 240 nM in arteriolar and from 57 to 201 nM in aortic VSMC (P < 0.001 for both). Inhibition of ANG II effects on [Ca2+]i revealed significant signaling through distinct AT-receptor subtypes (losartan and PD-123319 sensitive) in renal arteriolar VSMC. In contrast, only losartan was effective in aortic VSMC. The AT2-receptor ligand CGP-42112 had no effect in either vessel type. Our results demonstrate that cultured arteriolar VSMC have anatomical similarities to aortic VSMC and functional differences in AT-receptor signaling in response to ANG II. This novel preparation should provide a useful approach with which to investigate cellular mechanisms concerning receptor coupling to signaling pathways involved in vascular reactivity of arteriolar VSMC in the microcirculation in general and the kidney in particular.
本研究初步描述了大鼠球前阻力血管的血管平滑肌细胞（VSMC）培养物的基本形态学特性，以及血管紧张素II（ANG II）受体与胞质游离钙浓度（[Ca2+]i，采用fura 2荧光光度法测定）之间的功能偶联。使用氧化铁筛分法从肾小叶间动脉和传入小动脉（<50微米）分离出肾VSMC，并与在相似条件下培养的大鼠主动脉VSMC进行比较。在第3代至第10代之间，静止的单层细胞保持着均匀的形态和[Ca2+]i信号特征。小动脉和主动脉VSMC呈纺锤形，表达平滑肌特异性α-肌动蛋白以及肌球蛋白重链SM-1和SM-2。ANG II使[Ca2+]i迅速升高，随后在峰值的50%-60%维持平台期。最初的最大[Ca2+]i反应呈剂量依赖性，且在肾小动脉和主动脉VSMC中幅度相似。ANG II（10(-7) M）使小动脉VSMC中的[Ca2+]i从50 nM升高至240 nM，使主动脉VSMC中的[Ca2+]i从57 nM升高至201 nM（两者P均<0.001）。对ANG II对[Ca2+]i作用的抑制显示，肾小动脉VSMC通过不同的AT受体亚型（对氯沙坦和PD-123319敏感）存在显著信号传导。相比之下，只有氯沙坦对主动脉VSMC有效。AT2受体配体CGP-42112对两种血管类型均无作用。我们的结果表明，培养的小动脉VSMC在解剖学上与主动脉VSMC相似，但在对ANG II的AT受体信号传导方面存在功能差异。这种新的制备方法应为研究与一般微循环尤其是肾脏中小动脉VSMC血管反应性所涉及的信号通路的受体偶联相关的细胞机制提供一种有用的方法。
