All procedures involving animal models have been reviewed by the local institutional animal care committee and the JoVE veterinary review board.
1. To enable the rabbits to acclimate in the restrainers and for the investigator to objectively confirm stabilization of cardiorespiratory rates, instrument all rabbits with the cardiorespiratory and neuronal sensors and perform continuous video monitoring for > 1 hour, 1 - 3 times per animal.
2. Intravenous medication experiment (Pentylenetetrazol, PTZ)
- To visualize the marginal ear vein, shave the posterior surface of the rabbit's ear. Use a 70% ethanol wipe to disinfect the site and dilate the marginal ear vein. This is indicated by the black dashed oval in Figure 1F.
- At this point, have one experimenter cover the rabbit's face with their hand to decrease the stress of the procedure to the rabbit. A second experimenter carefully cannulates the marginal ear vein with a sterile 25-G angiocatheter.
- Once the catheter is in the vein, place a sterile injection plug at the end of the catheter so that a needle can introduce medication intravenously. The location of the injection plug is indicated by a blue circle in Figure 1G.
- Make a splint by wrapping 4 x 4 inch gauze with tape so that it forms a tube shape and placing it inside the rabbit's ear. Then tape the splint to the ear so that the catheter is secured in place and remains upright, similar to the non-catheterized ear.
- Inject 1 mL of 10 USP units per mL of heparinized saline.
NOTE: The catheter and vessel should be visibly cleared of air and remain patent. If the catheter is not in the vessel, the syringe will not push easily and there will be accumulation of saline in the subcutaneous tissue.
- Give rabbits incremental doses of PTZ intravenously from 1 mg/kg to 10 mg/kg in 1 mg/kg increments every 10 min. Make a note at the start of each dose to indicate which animal is being injected and the concentration of the medication.
NOTE: This enables assessments of the acute and additive effects of PTZ administration. Alternatively, to further assess the chronic effects of low dose PTZ, the rabbit is given repeated doses at each low dose concentration, 7 doses at 2 mg/kg, 3 doses at 5 mg/kg, then 3 doses at 10 mg/kg, each dose is separated by 10 min.
- After each dose, carefully monitor the video-EEG-ECG-capnography-oximetry for any neuro-cardiac electrical and respiratory abnormalities and visual evidence of epileptiform activity. Note these changes in real-time and during post-analysis.
NOTE: Seizure activity often begins within 60 s of PTZ administration.
