A Method to Inflict Closed Head Traumatic Brain Injury in Drosophila

Take anesthetized adult Drosophila flies in an empty vial.

Insert a cotton ball into the vial to confine the flies at the bottom, ensuring a stronger impact when inflicting injury.

Once the flies recover from anesthesia, attach the vial to the spring of the high-impact trauma device.

Rest the end of the vial on a raised platform. Then, pull back the spring to an angle and release it.

The vial strikes the platform, causing the flies to collide against the vial walls repeatedly.

The drosophila brain strikes against the head capsule, mimicking closed-head traumatic brain injury, or TBI, without causing external damage.

Allow the flies to recover, and transfer them to a vial with food.

The injury causes damage to neuron membranes, impairing neuronal function.

Signaling molecules from the damaged neurons recruit immune cells, leading to progressive degradation of brain tissue, eventually resulting in the flies' death.

Place between 1 and 60 carbon dioxide anesthetized flies in an empty vial, and stopper the vial using a tight fitting cotton ball. Then, draw a line on the vial, 1 inch from its base. Knock the flies down from the sides of the vial and push in the cotton ball until it matches up with the newly drawn line.

Wait five minutes for the flies to recover mobility from the carbon dioxide. Next, insert the end of the spring into the vial until the inner edge of the Velcro is flush with the top of the vial. This secures the vial to the spring by creating a tight compression fitting. With the spring lying flat, ensure that 1 inch of the vial overlaps the raised region of the ice bucket cover.

At this time, place on hearing protection because the impact of the vial against the ice bucket cover produces a loud noise. Next, hold the vial at the Velcro region using the thumb and forefinger of the left hand. With the right hand, hold the board tight to the bench top.

Next, pull the spring perfectly straight back to the desired angle, and then release the spring. Wait for the spring to come to a complete stop, and then remove the vial from the spring. Set the vial on the bench top and allow the flies to recover for at least five minutes. During this time the flies will slowly regain their normal motion. Next, transfer the flies to a vial with fly food for later analysis.