Spatial and temporal resolution of serotonin-induced changes in intracellular calcium in a cultured arterial smooth muscle cell line.

Blood vessels Pub Date : 1991-01-01 DOI:10.1159/000158870
W F Goldman
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引用次数: 10

Abstract

Ca2+ transients (1-2 microM) evoked by serotonin (5-HT) in cultured A7r5 cells were studied using fura-2 and digital imaging microscopy. Fura-2 was introduced into cells either by incubation with its acetoxymethyl ester analogue fura-2/AM or by transient ATP-induced permeabilization of the sarcolemma such that the free fura-2 entered the cell directly. The distribution of cytoplasmic Ca2+ in unstimulated cells loaded by the former method was heterogeneous, reflecting, in part, separate pools of Ca2+ in the cytosol and sarcoplasmic reticulum (SR). In contrast, the distribution of Ca2+ was uniform in cells loaded with fura-2 by transient permeabilization; this reflected the restriction of fura-2 to the cytosol. Average Ca2+ in these cells was substantially lower than that in fura-2/AM-loaded cells, because SR Ca2+ influences the fura-2 signal from fura-2/AM-loaded cells, but not from cells loaded with free fura-2. The differences in the Ca2+ distribution measured by the two loading methods were also evident during the course of 5-HT-evoked Ca2+ transients. Spatial and temporal resolution of the rising phase of 5-HT-evoked Ca2+ transients in fura-2/AM-loaded cells revealed that the onset of the Ca2+ transients was first manifested as small regions of elevated Ca2+ that subsequently expanded until peak apparent intracellular Ca2+ levels were present in virtually all of the nonnuclear regions of the cells. The rate of rise of Ca2+ varied in different cell regions with the nucleus responding the slowest.

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5 -羟色胺诱导的动脉平滑肌细胞系细胞内钙变化的时空分辨率。
利用fura-2和数字成像显微镜研究了血清素(5-HT)在培养的A7r5细胞中引起的Ca2+瞬态(1-2微米)。Fura-2通过与其乙酰氧基甲酯类似物Fura-2 /AM的孵育或通过atp诱导的肌膜的瞬时渗透使游离的Fura-2直接进入细胞而被引入细胞。通过前一种方法负载的未刺激细胞的细胞质Ca2+分布是不均匀的,部分反映了细胞质和肌浆网(SR)中Ca2+的分离池。相比之下,Ca2+在fura-2负载的细胞中通过瞬态渗透分布均匀;这反映了fura-2对细胞质的限制。这些细胞中的平均Ca2+明显低于加载fura-2/ am的细胞,因为SR Ca2+影响加载fura-2/ am的细胞的fura-2信号,但不影响加载游离fura-2的细胞。在5- ht诱导的Ca2+瞬态过程中,两种加载方法测量的Ca2+分布差异也很明显。在fura-2/ am负载的细胞中,5- ht诱发的Ca2+瞬态的上升期的空间和时间分辨率显示,Ca2+瞬态的发作首先表现为Ca2+升高的小区域,随后扩大,直到几乎所有细胞的非核区域都存在明显的细胞内Ca2+水平峰值。Ca2+在不同细胞区域的上升速率不同,细胞核反应最慢。
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