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      Spatial and Temporal Resolution of Serotonin-Induced Changes in Intracellular Calcium in a Cultured Arterial Smooth Muscle Cell Line

      Journal of Vascular Research

      S. Karger AG

      Fluorescence microscopy, Calcium, Vascular smooth muscle, Serotonin, Digital imaging

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          Ca<sup>2+</sup> transients (1–2 µ M) evoked by serotonin (5-HT) in cultured A<sub>7</sub>r<sub>5</sub> 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 Ca<sup>2+</sup> in unstimulated cells loaded by the former method was heterogeneous, reflecting, in part, separate pools of Ca<sup>2+</sup> in the cytosol and sarcoplasmic reticulum (SR). In contrast, the distribution of Ca<sup>2+</sup> was uniform in cells loaded with fura-2 by transient permeabilization; this reflected the restriction of fura-2 to the cytosol. Average Ca<sup>2+</sup> in these cells was substantially lower than that in fura-2/AM-loaded cells, because SR Ca<sup>2+</sup> influences the fura-2 signal from fura-2/AM-loaded cells, but not from cells loaded with free fura-2. The differences in the Ca<sup>2+</sup> distribution measured by the two loading methods were also evident during the course of 5-HT-evoked Ca<sup>2+</sup> transients. Spatial and temporal resolution of the rising phase of 5-HT-evoked Ca<sup>2+</sup> transients in fura-2/AM-loaded cells revealed that the onset of the Ca<sup>2+</sup> transients was first manifested as small regions of elevated Ca<sup>2+</sup> that subsequently expanded until peak apparent intracellular Ca<sup>2+</sup> levels were present in virtually all of the nonnuclear regions of the cells. The rate of rise of Ca<sup>2+</sup> varied in different cell regions with the nucleus responding the slowest.

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          Author and article information

          J Vasc Res
          Journal of Vascular Research
          S. Karger AG
          23 September 2008
          : 28
          : 1-3
          : 252-261
          Department of Physiology, University of Maryland School of Medicine, Baltimore, Md., USA
          158870 Blood Vessels 1991;28:252–261
          © 1991 S. Karger AG, Basel

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          Page count
          Pages: 10
          Application of New Techniques in Vascular Neuroeffector Research


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