Catwalk system

The locomotive function of hindlimb was evaluated at 1 day, 3 days, 5 days, 7 days, 14 days, 28 days, 42 days, and 56 days post-SCI using the Basso Mouse Scale (BMS) [34 (link)]. Automated gait analysis was also performed pre-surgery and 1, 2, 4, and 8 weeks post-surgery by using a “CatWalk” system (Noldus) [35 (link)]. All experiments were performed during the same period of the day (1:00 PM to 4:00 PM). For BMS, at least two examiners were blinded to the experimental group observed each mouse for 5 min. For Catwalk test, each mouse was trained to cross the Catwalk walkway daily for 7 days before SCI or control operation.

We used the Noldus CatWalk system to quantify gait characteristics 72 h after the impact. Individual animals were placed on the glass platform and the animals were allowed to freely walk up and down the CatWalk. A camera from below captured the animal’s footprint for automated analysis. The following parameters were used for a successful run: minimum duration of 0.5 s, maximum duration of 10 s, and during each run the animal’s speed could not change >65%. Each run was classified and the following characteristics were analyzed to assess for gait abnormalities: print area, swing duration, and stride length. Print area was calculated by averaging the area of all the footprints for each individual paw. Swing duration was considered to be the time between the lifting and replacing of the same paw. The stride length was calculated by the distance from the heel of one footprint to the heel of the next footprint of the same paw.

The CatWalk system (Noldus Inc., Wageningen, The Netherlands) was used to analyze the gait recovery of the animals (n = 5). In this protocol, the rats crossed a walkway (100 cm length, 5 cm width, and 0.6 cm thickness) with a glass floor illuminated from the long edge in a dark room. Data acquisition was performed with a high-speed camera (Pulnix TM-765E CCD), and the paw prints were automatically classified by the software. The paw prints were obtained during the 3 days before the partial transection of the tendons to assess the normal standard gait of the animals, and they were collected again after the lesions. Post-operative data were assessed on the 1st, 3rd, 5th, 7th, 9th, 11th, and 13th days following surgical lesion. The parameters used herein were “Max Contact Intensity”, corresponding to the pressure exerted by the paw on the glass floor during gait. The intensity of magnification can vary from 0 to 255 pixels.

For the gait recovery analysis, the CatWalk system (Noldus Inc., The Netherlands; was used. In this set up, the animal crosses a walkway with an illuminated glass floor. A high-speed video camera Gevicam (GP-3360, USA) equipped with a wide-angle lens (8.5 mm, Fujicon Corp., China) is positioned underneath the walkway and the paw prints are automatically recorded and classified by the software. The paw prints from each animal were obtained before and after the VRA. Post-operative CatWalk data were collected twice a week for 12 weeks. The peroneal functional index (PFI) was calculated as the distance between the third toe and hind limb pads (print length) and the distance between the first and fifth toes (print width). Measurements of these parameters were obtained from the right (lesioned) and left (unlesioned) paw prints, and the values were calculated using the following formula [27] (link).
Where N: normal, or non-operated side; E: experimental, or operated; PL: print length; TS: total toe spread, or distance between first to fifth toe.
The pressure exerted on the platform by individual paws was also evaluated. The Catwalk data from each day were expressed as an ipsi-/contralateral ratio.

For the gait recovery analysis, the CatWalk system (Noldus Inc, The Netherlands) was used. In this method, the animal crosses a walkway with a glass floor illuminated from the long edge. Data acquisition was carried out by a high-speed camera and the paw prints were automatically classified by the software. The paw prints from each animal (minimum of n = 8 per group) were obtained before and after the sciatic crushing. Post-operative data were assessed on the third, fourth and fifth days following surgical intervention, and then twice a week until 8 weeks post-lesion.
The parameters used herein to calculate the sciatic functional index were the distance between the third toe and hind limb pads (print length, PL), and the distance between the first and fifth toes (toe spread, TS). Measurements of the parameters were obtained from the right (normal) and left (experimental) paw prints and the values were used to calculate the sciatic functional index (SFI), according to the following formula [16 (link)]: SFI = 118.9 (ETS - NTS/NTS) - 51.2 (EPL - NPL/NPL) - 7.5 (E = experimental side, N = normal side). In addition, the pressure exerted by the individual paws during contact with the platform was evaluated. The data were expressed as the lesioned/unlesioned ratio for each day of training.