Abstract
Fast object tracking in real time allows convenient tracking of very large numbers of animals and closed-loop experiments that control stimuli for many animals in parallel. We developed MARGO, a MATLAB-based, real-time animal tracking suite for custom behavioral experiments. We demonstrated that MARGO can rapidly and accurately track large numbers of animals in parallel over very long timescales, typically when spatially separated such as in multiwell plates. We incorporated control of peripheral hardware, and implemented a flexible software architecture for defining new experimental routines. These features enable closed-loop delivery of stimuli to many individuals simultaneously. We highlight MARGO's ability to coordinate tracking and hardware control with two custom behavioral assays (measuring phototaxis and optomotor response) and one optogenetic operant conditioning assay. There are currently several open source animal trackers. MARGO's strengths are 1) fast and accurate tracking, 2) high throughput, 3) an accessible interface and data output and 4) real-time closed-loop hardware control for for sensory and optogenetic stimuli, all of which are optimized for large-scale experiments.
Highlights
Automated animal tracking methods have become commonplace in the study of behavior
We show that traces acquired in MARGO are comparably accurate to those of other trackers and are robust to noisy images and changing imaging conditions
Overall we found a high degree of agreement between traces acquired in MARGO and Ctrax (Fig 2E and 2F)
Summary
Automated animal tracking methods have become commonplace in the study of behavior. They enable large sample sizes, high statistical power, and more rapid inference of mechanisms giving rise to behavior. MARGO (Massively Automated Real-time GUI for Object-tracking), a platform for high-throughput ethology permit a diversity of organisms, tracking modes, experimental paradigms, and behavioral arenas, 3) integration of peripheral hardware to enable closed-loop sensory and optogenetic stimuli, and 4) a user-friendly interface and data output format. MARGO (Massively Automated Real-time GUI for Object-tracking), a platform for high-throughput ethology correlated (r = -0.38 and r = -0.63 respectively) between the two conditions, suggesting that both individual phototactic bias and locomotor handedness bias affect each choice. From the position of these dots in camera coordinates, we constructed a registration mapping from the camera FOV to the projector display field Using this mapping, we programmed MARGO to use the real-time positions of flies to project pinwheel stimuli independently to 48 freely moving individuals simultaneously (Fig 5B, S8 Video). All flies in B were fed with all-trans-retinal
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