Kerr R, Lev-Ram V, Baird G, Vincent P, Tsien R Y, Schafer W R
Department of Biology, University of California, San Diego, La Jolla 92093, USA.
Neuron. 2000 Jun;26(3):583-94. doi: 10.1016/s0896-6273(00)81196-4.
Electrophysiology and optical indicators have been used in vertebrate systems to investigate excitable cell firing and calcium transients, but both techniques have been difficult to apply in organisms with powerful reverse genetics. To overcome this limitation, we expressed cameleon proteins, genetically encoded calcium indicators, in the pharyngeal muscle of the nematode worm Caenorhabditis elegans. In intact transgenic animals expressing cameleons, fluorescence ratio changes accompanied muscular contraction, verifying detection of calcium transients. By comparing the magnitude and duration of calcium influx in wild-type and mutant animals, we were able to determine the effects of calcium channel proteins on pharyngeal calcium transients. We also successfully used cameleons to detect electrically evoked calcium transients in individual C. elegans neurons. This technique therefore should have broad applications in analyzing the regulation of excitable cell activity in genetically tractable organisms.
电生理学和光学指示剂已被用于脊椎动物系统中，以研究可兴奋细胞的放电和钙瞬变，但这两种技术都很难应用于具有强大反向遗传学的生物体中。为了克服这一限制，我们在秀丽隐杆线虫的咽部肌肉中表达了钙黄绿蛋白（cameleon proteins），即基因编码的钙指示剂。在完整的表达钙黄绿蛋白的转基因动物中，荧光比率变化伴随着肌肉收缩，证实了钙瞬变的检测。通过比较野生型和突变型动物钙内流的幅度和持续时间，我们能够确定钙通道蛋白对咽部钙瞬变的影响。我们还成功地使用钙黄绿蛋白检测了单个秀丽隐杆线虫神经元中电诱发的钙瞬变。因此，这项技术在分析遗传易处理生物体中可兴奋细胞活动的调节方面应该有广泛的应用。
