Experiments
- All animals were fed ad libitum with standard rodent chow and housed in a humidity-controlled room with a 12-h light/dark cycle. All procedures were approved by the Health Sciences Animal Care Committee of the University of Calgary.
- Whole-brain mitochondria were acutely isolated from rodents using differential centrifugation, nitrogen disruption, and a Ficoll gradient.
- A Seahorse Bioscience XFe24 extracellular flux analyzer was used to measure mitochondrial function in cultured cells or intact isolated mitochondria. BE(2)-M17 neuroblastoma cells were cultured in microplates under normal conditions for 48 h and treated with CBD for 24 h. Acutely isolated brain mitochondria were diluted in respiration buffer and treated with different concentrations of CBD solution.
- Mitochondria were placed in 2 mL of respiration buffer in a cuvette with CaG5N and TMRE. CaG5N was used to monitor extramitochondrial Ca2+, and TMRE was used to monitor changes in Δψm.
- Patch-clamp recordings on mitoplasts prepared from acutely isolated Wistar rat brain and CB1 KO mouse brain mitochondria were performed. Transverse hippocampal slices were prepared and a high-frequency LTP protocol was implemented.
- Male Wistar rats were treated with CBD at three different daily doses intraperitoneally for 12 days. The Morris water maze testing began on day 8 of CBD treatment. A white circular pool was filled with water to a level above a hidden platform. Each training trial lasted 60 sec or until the rat located the hidden platform.
Animals
Items
| Material | Material Usage | Supplier |
| Wistar rats | N/A | Charles River |
| Cannabinoid receptor type 1 knockout (CB1 KO) mice | N/A | University of Calgary |
| Standard rodent chow | N/A | LabDiet, St. Louis, MO |
Instructions
- N/A
Methodologies
Mitochondrial Isolation
Instructions
- N/A
Methodologies
Mitochondrial Bioenergetic Assay
Items
| Material | Material Usage | Supplier |
| Seahorse Bioscience XFe24 extracellular flux analyzer | To measure mitochondrial function | N/A |
| BE(2)-M17 neuroblastoma cells | N/A | N/A |
| Oligomycin | N/A | N/A |
| FCCP (carbonyl cyanide 4-(trifluoromethoxy) phenylhydrazone) | N/A | N/A |
| Antimycin A | N/A | N/A |
| CBD solution | N/A | Sigma-Aldrich, St. Louis, MO |
Instructions
- N/A
Methodologies
mPT Assay
Items
| Material | Material Usage | Supplier |
| Respiration buffer | N/A | N/A |
| CaG5N | To monitor extramitochondrial Ca2+ | N/A |
| TMRE | To monitor changes in Δψm | N/A |
Instructions
- N/A
Methodologies
Electrophysiology
Instructions
- N/A
Methodologies
Morris Water Maze Testing
Instructions
- N/A
Methodologies
