Imaging neuronal activity during zebrafish behavior with a genetically encoded calcium indicator
Imaging neuronal activity during zebrafish behavior with a genetically encoded calcium indicator
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DOI：
10.1152/jn.00576.2003
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发表时间：
2003-12-01
影响因子：
2.5
通讯作者：
Fetcho, JR
中科院分区：
文献类型：
--
作者：
Higashijima, S;Masino, MA;Fetcho, JR
遗传编码的钙指示剂，如cameleon，提供了非侵入性监测神经元活动的承诺，但没有人证明这些指示剂是否可以报告行为脊椎动物神经元中的钙瞬变。我们表明，cameleon可以在斑马鱼的感觉和脊髓神经元在高水平表达，通过使用神经特异性启动子在瞬时表达实验和稳定的转基因系。使用标准的共聚焦显微镜，钙瞬变在确定的运动神经元和脊髓中间神经元可以检测到在逃避行为所产生的触摸的鱼的头部。小动作的限制鱼的行为过程中并不代表一个主要的问题，因为cameleon的比率性质的钙反应分析。我们的结论是，cameleon可用于非侵入性研究神经元的活动在一个完整的，行为的脊椎动物。引入遗传学指标的能力允许研究局部中间神经元的功能作用，而这些功能作用不能用其他方法容易地监测。转基因系，如我们产生的一个也可以交叉到突变系的鱼，研究结构和功能的突变后果。
Genetically encoded calcium indicators, such as cameleon, have offered the promise of noninvasively monitoring activity of neurons, but no one has demonstrated whether these indicators can report calcium transients in neurons of behaving vertebrates. We show that cameleon can be expressed at high levels in sensory and spinal cord neurons in zebrafish by using neural-specific promoters in both transient expression experiments and in a stable transgenic line. Using standard confocal microscopy, calcium transients in identified motoneurons and spinal interneurons could be detected during escape behaviors produced by a touch on the head of the fish. Small movements of the restrained fish during the behavior did not represent a major problem for analyzing the calcium responses because of the ratiometric nature of cameleon. We conclude that cameleon can be used to noninvasively study the activity of neurons in an intact, behaving vertebrate. The ability to introduce an indicator genetically allows for studies of the functional roles of local interneurons that cannot easily be monitored with other approaches. Transgenic lines such as the one we generated can also be crossed into mutant lines of fish to study both structural and functional consequences of the mutations.