Section of Neurobiology and Institute for Neuroscience, The University of Texas at Austin, Austin, Texas, United States of America.
PLoS One. 2010 Dec 30;5(12):e15259. doi: 10.1371/journal.pone.0015259.
Drosophila melanogaster larvae are model systems for studies of development, synaptic transmission, sensory physiology, locomotion, drug discovery, and learning and memory. A detailed behavioral understanding of larvae can advance all these fields of neuroscience. Automated tracking can expand fine-grained behavioral analysis, yet its full potential remains to be implemented for the larvae. All published methods are unable to track the larvae near high contrast objects, including the petri-dish edges encountered in many behavioral paradigms. To alleviate these issues, we enhanced the larval contrast to obtain complete tracks. Our method employed a dual approach of optical-contrast boosting and post-hoc image processing for contrast enhancement. We reared larvae on black food media to enhance their optical contrast through darkening of their digestive tracts. For image processing we performed Frame Averaging followed by Subtraction then Thresholding (FAST). This algorithm can remove all static objects from the movie, including petri-dish edges prior to processing by the image-tracking module. This dual approach for contrast enhancement also succeeded in overcoming fluctuations in illumination caused by the alternating current power source. Our tracking method yields complete tracks, including at the edges of the behavioral arena and is computationally fast, hence suitable for high-throughput fine-grained behavioral measurements.
黑腹果蝇幼虫是研究发育、突触传递、感觉生理学、运动、药物发现以及学习和记忆的模型系统。对幼虫进行详细的行为理解可以推进所有这些神经科学领域的发展。自动化跟踪可以扩展精细的行为分析，但它的全部潜力仍有待于在幼虫中实现。所有已发表的方法都无法在高对比度物体附近跟踪幼虫，包括在许多行为范式中遇到的培养皿边缘。为了解决这些问题，我们增强了幼虫的对比度以获得完整的轨迹。我们的方法采用了光学对比度增强的双重视角和事后图像处理的方法。我们在黑色食物培养基上饲养幼虫，通过使它们的消化道变暗来增强它们的光学对比度。对于图像处理，我们进行了帧平均化，然后进行减法和阈值处理（FAST）。该算法可以从电影中去除所有静态物体，包括在图像处理模块处理之前的培养皿边缘。这种对比度增强的双重方法还成功地克服了交流电源引起的光照波动。我们的跟踪方法可以生成完整的轨迹，包括在行为场的边缘，并且计算速度快，因此适合于高通量的精细行为测量。
