To begin, place a platinum harp in a weighboat near the MEA platform. Then cover the harp with about three milliliters of aCSF to reduce its hydrophobic tendencies. Use scissors to trim about 1.5 inches off the narrow tip of a transfer pipette.
Next, use the modified pipette to collect a brain slice from the slice-holding chamber. Gently dispense the brain slice and any solution in the pipette into the chip well. To position the slice properly, use a soft paintbrush to create a current in the solution that pushes the brain slice onto recording electrodes.
Using forceps, gently place the harp over the brain slice with the threads downwards to press the slice onto the recording electrodes. Orient the harp so that the side without a frame faces toward the inflow needle and the frame of the harp does not contact any of the recording electrodes. Now take a discard pipette and remove excess aCSF.
Then take an antistatic wipe, twist a corner to create a tip, and use it to soak up the remaining aCSF surrounding the recording electrodes without touching the recording electrodes, brain slice, or harp. Using a designated aCSF pipette, quickly add about two milliliters of carbogenated aCSF to cover the brain slice. Fill the well with about three milliliters carbogenated aCSF until it is roughly three quarters full.
Then use a microscope or camera to take a high-resolution picture of the brain slice on the MEA chip. Place the inflow and outflow tubes into the beaker filled with aCSF. Place the inflow needle close to the bottom of the chip well, just outside the recording electrodes.
Then place the outflow needle close to the top of the chip well, towards the edge, so that the liquid rises almost to the brim of the chip well, about four milliliters. Now set the perfusion inflow to five milliliters per minute and the perfusion outflow to seven milliliters per minute. Turn on the inflow and outflow.
Remove the inflow needle from the chip well until it begins to output solution instead of air. After that, place the needle back inside the chip well. Then use a solution heater to keep the solution at or near physiological temperature, around 34 to 37 degrees Celsius.
Let the aCSF perfuse over the brain slice for 10 minutes. After 10 minutes have elapsed, move the outflow tube to the discard beaker. Then move the inflow tube to the beaker containing the pro-convulsant solution.
Allow the non-convulsant aCSF to be flushed out of the perfusion system into the discard beaker for 10 minutes. Finally, transfer the outflow tube into the beaker containing the pro-convulsant solution and allow it to cycle until the experiment finishes. Powerful electrographic seizure-like activity was frequently observed in the neocortical regions under both the zero magnesium and 4-aminopyridine paradigms.
The hippocampal regions displayed more variability between brain slices, with some demonstrating seizure-like activity and others showing transient discharges. Neocortical activity under the zero magnesium paradigm showed higher power in multiple frequency bands compared to the 4-aminopyridine paradigm. Hippocampal activity in the zero magnesium paradigm demonstrated higher power in high gamma frequency bands compared to the neocortex and both brain regions in the 4-aminopyridine paradigm.
