Carbachol-activated calcium entry into HT-29 cells is regulated by both membrane potential and cell volume.
Carbachol-activated calcium entry into HT-29 cells is regulated by both membrane potential and cell volume.
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卡巴胆碱激活的钙进入 HT-29 细胞受膜电位和细胞体积的调节。
DOI：
10.1073/pnas.89.4.1438
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发表时间：
1992
影响因子：
11.1
通讯作者：
Machen,TE
中科院分区：
文献类型：
--
作者：
Fischer,H;Illek,B;Negulescu,PA;Clauss,W;Machen,TE
用Ca2+探针fura-2和数字成像显微镜测量单Cl（-）分泌的HT-29/B6结肠癌细胞内Ca2+ ([Ca2+]i)。静息[Ca2+]i为63 +/- 3 nM （n = 62）。在使用毒蕈碱激动剂carbachol处理期间，[Ca2+]i迅速增加到901 +/- 119 nM，随后达到309 +/- 23 nM的稳定水平，这取决于Ca2+从细胞外溶液进入细胞。本研究的目的是表征Ca2+通过细胞膜的进入途径，其依赖于膜电位和细胞体积。在静息条件下[Ca2+]i对电位和细胞体积没有明显的依赖性。在用carbachol（100微米）刺激Ca2+进入后，[Ca2+]i随着细胞的超极化（低k +或valinomycin处理）而增加，并随着去极化（高k +或gramicidin处理）而减少，正如Ca2+进入驱动力的变化所预期的那样。在受刺激的细胞中，低渗溶液导致[Ca2+]i增加，而高渗溶液阻止Ca2+进入。当缬霉素增加膜电位时，收缩引起的[Ca2+]i下降仅受到轻微影响，这表明收缩直接影响了碳甾醇激活的Ca2+电导。相比之下，在缬霉素处理的细胞中，肿胀诱导的[Ca2+]i的增加明显减少，表明间接依赖于肿胀激活的K+电导。因此，碳水化合物刺激的Ca2+进入受到膜电位和细胞体积的双重控制。这种机制可能作为一种调节影响，决定胆碱能刺激期间Ca2+内流的程度。
Intracellular Ca2+ ([Ca2+]i) was measured in single Cl(-)-secretory HT-29/B6 colonic carcinoma cells with the Ca2+ probe fura-2 and digital imaging microscopy. Resting [Ca2+]i was 63 +/- 3 nM (n = 62). During treatment with the muscarinic agonist carbachol, [Ca2+]i rapidly increased to 901 +/- 119 nM and subsequently reached a stable level of 309 +/- 23 nM, which depended on Ca2+ entry into the cells from the extracellular solution. The goal of this study was to characterize the Ca2+ entry pathway across the cell membrane with respect to its dependence on membrane potential and cell volume. Under resting conditions [Ca2+]i showed no apparent dependence on either potential or cell volume. After stimulating Ca2+ entry with carbachol (100 microM), [Ca2+]i increased with hyperpolarization (low-K+ or valinomycin treatment) and decreased with depolarization (high-K+ or gramicidin treatment) of the cell, as expected from changes in driving force for Ca2+ entry. In stimulated cells, hypotonic solutions caused [Ca2+]i to increase, whereas hypertonic solutions blocked Ca2+ entry. The shrinkage-induced decreases in [Ca2+]i were only slightly affected when the membrane potential was increased with valinomycin, suggesting that shrinkage directly affects the carbachol-activated Ca2+ conductance. In contrast, the swelling-induced increase in [Ca2+]i was significantly reduced in valinomycin-treated cells, suggesting an indirect dependence on a swelling-activated K+ conductance. Thus, carbachol-stimulated Ca2+ entry is under the dual control of membrane potential and cell volume. This mechanism may serve as a regulatory influence that determines the extent of Ca2+ influx during cholinergic stimulation.