Chai Cynthia M, Cronin Christopher J, Sternberg Paul W
Division of Biology and Bioengineering, California Institute of Technology; Howard Hughes Medical Institute, California Institute of Technology;
Division of Biology and Bioengineering, California Institute of Technology; Howard Hughes Medical Institute, California Institute of Technology.
J Vis Exp. 2017 Jul 13(125):55963. doi: 10.3791/55963.
The nematode, Caenorhabditis elegans' compact nervous system of only 302 neurons underlies a diverse repertoire of behaviors. To facilitate the dissection of the neural circuits underlying these behaviors, the development of robust and reproducible behavioral assays is necessary. Previous C. elegans behavioral studies have used variations of a "drop test", a "chemotaxis assay", and a "retention assay" to investigate the response of C. elegans to soluble compounds. The method described in this article seeks to combine the complementary strengths of the three aforementioned assays. Briefly, a small circle in the middle of each assay plate is divided into four quadrants with the control and experimental solutions alternately placed. After the addition of the worms, the assay plates are loaded into a behavior chamber where microscope cameras record the worms' encounters with the treated regions. Automated video analysis is then performed and a preference index (PI) value for each video is generated. The video acquisition and automated analysis features of this method minimizes the experimenter's involvement and any associated errors. Furthermore, minute amounts of the experimental compound are used per assay and the behavior chamber's multi-camera setup increases experimental throughput. This method is particularly useful for conducting behavioral screens of genetic mutants and novel chemical compounds. However, this method is not appropriate for studying stimulus gradient navigation due to the close proximity of the control and experimental solution regions. It should also not be used when only a small population of worms is available. While suitable for assaying responses only to soluble compounds in its current form, this method can be easily modified to accommodate multimodal sensory interaction and optogenetic studies. This method can also be adapted to assay the chemosensory responses of other nematode species.
线虫秀丽隐杆线虫仅由302个神经元组成的紧凑神经系统是其多样行为表现的基础。为了便于剖析这些行为背后的神经回路，开发强大且可重复的行为分析方法是必要的。以往秀丽隐杆线虫的行为研究使用了“滴落试验”“趋化性分析”和“记忆试验”的不同变体来研究秀丽隐杆线虫对可溶性化合物的反应。本文所述方法旨在结合上述三种分析方法的互补优势。简要来说，每个分析板中间的一个小圆圈被分成四个象限，对照溶液和实验溶液交替放置。加入线虫后，将分析板放入行为室，显微镜摄像头记录线虫与处理区域的接触情况。然后进行自动视频分析，并为每个视频生成一个偏好指数（PI）值。该方法的视频采集和自动分析功能最大限度地减少了实验者的参与及任何相关误差。此外，每次分析使用的实验化合物量极少，行为室的多摄像头设置提高了实验通量。该方法对于进行遗传突变体和新型化合物的行为筛选特别有用。然而，由于对照溶液区域和实验溶液区域靠得很近，该方法不适用于研究刺激梯度导航。当只有少量线虫可用时，也不应使用该方法。虽然目前这种形式仅适用于检测对可溶性化合物的反应，但该方法可以很容易地进行修改，以适应多模态感官相互作用和光遗传学研究。该方法也可适用于检测其他线虫物种的化学感应反应。
