Location-dependent photogeneration of calcium waves in HeLa cells
Location-dependent photogeneration of calcium waves in HeLa cells
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DOI：
10.1385/cbb:45:2:167
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发表时间：
2006-01-01
影响因子：
2.6
通讯作者：
Kawata, Satoshi
中科院分区：
文献类型：
--
作者：
Iwanaga, Shigeki;Kaneko, Tomoyuki;Kawata, Satoshi
细胞中的钙离子（Ca 2+）浓度负责控制重要的细胞功能，并且已经被广泛研究作为研究和控制细胞活性的手段。在这里，我们证明了在HeLa细胞中产生的飞秒激光照射的Ca 2+波，并显示出意想不到的性能的Ca 2+的释放和传播。当激光聚焦在细胞质中时，Ca 2+释放独立于外部Ca 2+内流和磷酸肌醇磷脂酶C（PLC）信号通路。细胞核不是激光诱导的Ca 2+释放的敏感靶点，而质膜的照射产生了瞬时穿孔的证据，细胞外溶液可以通过该穿孔进入细胞。通过螯合细胞外Ca ~（2+），我们发现，激光诱导的乙二醇四乙酸（EGTA）内流可以与钙诱导的钙释放竞争，并显着延迟或抑制靶细胞中Ca ~（2+）波的开始。细胞间Ca 2+的传播是腺苷三磷酸依赖性的，即使当EGTA的流入抑制靶细胞胞浆Ca 2+的上升，也可以观察到。还测试了对总细胞活力的辐照效应，发现其较低（在60 mW平均功率辐照后6小时为85%）。通过控制曝光量和焦点位置可以可靠地产生激光诱导的Ca 2+波，并且不需要笼状Ca 2+的存在。该技术有可能取代其他Ca 2+刺激方法，这些方法要么需要细胞中额外的笼状分子，要么没有很好的局部化相互作用。
The calcium ion (Ca2+) concentrations in a cell are responsible for the control of vital cellular functions and have been widely studied as a means to investigate and control cell activities. Here, we demonstrate Ca2+ wave generation in HeLa cells by femtosecond laser irradiation and show unexpected properties of the Ca2+ release and propagation. When the laser was focused in the cell cytoplasm, Ca2+ release was independent of both external Ca2+ influx and the phosphoinositide-phospholipase C (PLC) signaling pathway. The nucleus was not a susceptible target for laser-induced Ca2+ release, whereas irradiation of the plasma membrane produced evidence of transient poration, through which the extracellular solution could enter the cell. By chelating extracellular Ca2+, we found that laser-induced influx of ethylene glycol tetra-acetic acid (EGTA) can compete with calcium-induced calcium release and significantly delay or suppress the onset of the Ca2+ wave in the target cell. Intercellular Ca2+ propagation was adenosine triphosphate-dependent and could be observed even when the target cell cytosolic Ca2+ rise was suppressed by influx of EGTA. The irradiation effect on overall cell viability was also tested and found to be low (85% at 6 h after irradiation by 60 mW average power). Laser-induced Ca2+ waves can be reliably generated by controlling the exposure and focal position and do not require the presence of caged Ca2+. The technique has the potential to replace other methods of Ca2+ Stimulation, which either require additional caged molecules in the cell or do not have an interaction that is as well localized.