Werner Reichardt Centre for Integrative Neuroscience and Institute of Neurobiology, University of Tübingen, Tübingen, Germany.
Werner Reichardt Centre for Integrative Neuroscience, Central Office System Administration, University of Tübingen, Tübingen, Germany.
Nat Protoc. 2018 Jul;13(7):1539-1568. doi: 10.1038/s41596-018-0002-0.
Reliable measurement of spontaneous and evoked eye movement is critical for behavioral vision research. Zebrafish are increasingly used as a model organism for visual neural circuits, but ready-to-use eye-tracking solutions are scarce. Here, we present a protocol for automated real-time measurement of angular horizontal eye position in up to six immobilized larval fish using a custom-built LabVIEW-based software, ZebEyeTrack. We provide its customizable source code, as well as a streamlined and compiled version, ZebEyeTrack Light. The full version of ZebEyeTrack controls all required hardware and synchronizes six essential aspects of the experiment: (i) stimulus design; (ii) visual stimulation with moving bars; (ii) eye detection and tracking, as well as general motion detection; (iv) real-time analysis; (v) eye-position-dependent closed-loop event control; and (vi) recording of external event times. This includes optional integration with external hardware such as lasers and scanning microscopes. Once installation is complete, experiments, including stimulus design, can be completed in <10 min, and recordings can last anywhere between seconds and many hours. Results include digitized angular eye positions and hardware status, which can be used to compute tuning curves, optokinetic gain, and other custom data analysis. After the experiment, or based on existing videos, optokinetic response (OKR) performance can be analyzed semi-automatically via the graphical user interface, and results can be exported. ZebEyeTrack has been used successfully for psychophysics experiments, for optogenetic stimulation, and in combination with calcium imaging.
可靠的自发和诱发眼球运动测量对于行为视觉研究至关重要。斑马鱼越来越多地被用作视觉神经回路的模型生物，但现成的眼动追踪解决方案却很少。在这里，我们提出了一个使用基于自定义 LabVIEW 的软件 ZebEyeTrack 自动实时测量多达 6 条固定幼鱼水平角度眼球位置的方案。我们提供了它的可定制源代码，以及一个简化和编译的版本 ZebEyeTrack Light。ZebEyeTrack 的完整版本控制所有必需的硬件，并同步实验的六个关键方面：（i）刺激设计；（ii）用移动棒进行视觉刺激；（iii）眼睛检测和跟踪，以及一般运动检测；（iv）实时分析；（v）基于眼位置的闭环事件控制；以及（vi）外部事件时间的记录。这包括与外部硬件（如激光器和扫描显微镜）的可选集成。安装完成后，实验（包括刺激设计）可以在 10 分钟内完成，记录可以持续数秒到数小时。结果包括数字化的角度眼位置和硬件状态，可用于计算调谐曲线、视动增益和其他自定义数据分析。实验结束后，或基于现有的视频，可通过图形用户界面半自动地分析视动反应（OKR）性能，并可导出结果。ZebEyeTrack 已成功用于心理物理学实验、光遗传学刺激以及与钙成像的结合。
