Computational protocol: A Neonatal Mouse Spinal Cord Injury Model for Assessing Post-Injury Adaptive Plasticity and Human Stem Cell Integration

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Protocol publication

[…] A non-weightbearing air stepping test (modified from ) was used to quantify limb motility in sham control, SCC and SCT mice at different times after surgery/injury (, total n = 35). Tests were performed under a heating lamp to avoid hypothermia, especially at the earliest ages (1 and 4 days after surgery/injury). Animals were suspended dorsal up from a horizontal bar in a soft harness that secured the head, body and tail without restraining limb movements. The harness was adjusted to secure the animal so that there was no obvious suffering or discomfort. Typically, once restrained, the animal was quiescent and did not struggle unless stimulated. The paws were marked in black to facilitate video kinematic tracking. A mirror was mounted below the animal at a 45-degree angle to permit recording of side and ventral views simultaneously. After a short period of quiescence the nose was touched with a smooth metal prod, which led to a series of what appeared to be escape movements involving alternating flexion and extension of each limb (air stepping) with variable interlimb coordination. Video sequences of 1–3 min were taken with a 25 Hz camera (JVC Everio), and at the end of this session the animal was returned to the litter. Raw interlaced video images were reformatted into real images using the Avidemux video editor (, sequences of these were imported into ImageJ and the trajectory of each paw was tracked frame-by-frame using a “manual tracking” plug-in . The trajectories (movements of the paws in the x-y plane), trajectory amplitudes (maximal distance traversed in the x-y plane) and instantaneous velocities (interframe distance covered divided by interframe duration) were measured.A single-track locomotion test was used to quantify gait parameters in sham control and SCC mice (n = 3 for each group) 24 days after surgery/injury. For gait analyses, mice were placed in a transparent, rectangular, ceilingless plexiglass corridor (10 cm wide×50 cm high). Mice placed at one end walked readily along the corridor following a gentle push from behind, and were videofilmed at 25 Hz through the floor of the corridor. A frame-by-frame analysis was performed as above and stance durations were determined by measuring the time from initial contact to final lift of each paw during successive placements. […]

Pipeline specifications

Software tools ImageJ, Manual Tracking
Applications Laser scanning microscopy, Microscopic phenotype analysis
Organisms Mus musculus, Homo sapiens
Diseases Spinal Cord Compression, Spinal Cord Injuries