High-resolutionin vivokinematic tracking with injectable fluorescent nanoparticles

Abstract

Behavioral quantification is a cornerstone of many neuroscience experiments. Recent advances in motion tracking have streamlined the study of behavior in small laboratory animals and enabled precise movement quantification on fast (millisecond) timescales. This includes markerless keypoint trackers, which utilize deep network systems to label positions of interest on the surface of an animal (e.g., paws, snout, tail, etc.). These approaches mark a major technological achievement. However, they have a high error rate relative to motion capture in humans and are yet to be benchmarked against ground truth datasets in mice. Moreover, the extent to which they can be used to track joint or skeletal kinematics remains unclear. As the primary output of the motor system is the activation of muscles that, in turn, exert forces on the skeleton rather than the skin, it is important to establish potential limitations of techniques that rely on surface imaging. This can be accomplished by imaging implanted fiducial markers in freely moving mice. Here, we present a novel tracking method called QD-Pi (Quantum Dot-based Pose estimation in vivo), which employs injectable near-infrared fluorescent nanoparticles (quantum dots, QDs) immobilized on microbeads. We demonstrate that the resulting tags are biocompatible and can be imaged non-invasively using commercially available camera systems when injected into fatty tissue beneath the skin or directly into joints. Using this technique, we accurately capture 3D trajectories of up to ten independent internal positions in freely moving mice over multiple weeks. Finally, we leverage this technique to create a large-scale ground truth dataset for benchmarking and training the next generation of markerless keypoint tracker systems.