Development of a Preclinical Inhalation Model to Test Vaporized Cannabis Distillates

We explore how inhaled cannabis vape products impact lung physiology, by combining traditional molecular biology techniques and emerging computational techniques to gain insights on how vaping changes lung function. We have shown that cannabis vapor elicits transcriptional changes in lung epithelial cells, similar to cannabis smoke, suggestive of damage. Also, mechanistic insights into vaping related lung injury are emerging, particularly with THC products containing vitamin E acetate. Most studies on THC containing products rely on non-inhalation methods, such as oral gavage and injections, which offer valuable insights, but differ from typical human use. Our goal is to model cannabis exposure in a more physiologically relevant manner, with inhalation. The potency of new cannabis products provides challenges in dosing. We want to mimic human use patterns, and achieve physiologically relevant doses without adverse effects in the mice. We establish a standardized exposure dose that delivers physiologically relevant doses of THC to the mice without adverse behavioral effects.

[Instructor] To begin, assemble all components of the system. Insert a closed tube into the buffer chamber of the puffing pump to prevent leakage. Turn on the system and launch the software. Select the experimentation sessions module. Select new study to begin a new experimental session. Define the experimental groups and subjects to be studied. Then choose the experimental template that matches the desired exposure regimen. In the session properties window, complete the operator section to document the equipment usage. For calibration, select the desired channel in the software. Follow the steps outlined in the operating software. Next, select the desired pump in the software. Perform the system flow test by following the operating software instructions. Confirm that flow is directed toward the rotameter. Cancel any prompts to start data recording if the experiment is not ready to begin. For animal preparation, acclimate male and female C57 BL6 gradually over three days to the exposure system. To begin acclimation, place each mouse in a soft restraint. Initiate a two liters per minute bias airflow with room air for a duration equivalent to their experimental exposure of 10, 20, or 30 minutes. Fully retract the mesh component of the restraint. Then hold the entire restraint in front of the mouse, and allow the mouse to enter the plunger component on its own. Verify that the mouse's nose is visible in the plunger section. Secure a binder clip just behind the mouse to prevent backward movement. When handling mice of different sexes or genotypes, use color coded binder clips to identify and distinguish them. Place six restrained mice into the nose-only exposure tower of the system. Initiate the bias flow at two liters per minute with room air to ensure sufficient airflow through the chamber. In the task docker, right click on the e-cigarette profile created and select task properties. Under puff frequency, enter the desired puff interval, such as 30 seconds for a two puffs per minute regime. Click OK when prompted to confirm the changes. Follow the prompts at the end of the session to save the puff regime as a template for future use. When ready to begin the exposure, double click on the modified e-cigarette profile to start it. Start a timer simultaneously to track the exposure duration. Test varying intensity by maintaining a 10 minute exposure and increasing puff frequency to one, two, and four puffs per minute. After the exposure ends, restart the bias flow with room air. To release animals from the restraints, remove the binder clip first. Fully retract the mesh toward the plunger, and allow the animal to exit the restraint on its own. If needed, gently tug the tail to encourage backward movement. Serum THC-COOH concentrations significantly increased in a time dependent manner with 10, 20, and 30 minutes of exposure to THC vapor at one puff per minute. Increasing puff frequency during a 10 minute exposure resulted in a dose dependent rise in serum THC-COOH levels for one, two, and four puffs per minute. Mice exposed to THC vapor for 10 minutes at two puffs per minute and four puffs per minute exhibited significant hypolocomotion, with distance traveled reduced to 0.29 and 0.05 meters respectively.