EEG recordings and PTZ seizure induction

Mice were implanted with wireless telemetry units (PhysioTel ETA-F10; DSI, Data Sciences International) under sterile techniques per laboratory protocol as described above. Under anesthesia, a transmitter was placed intraperitoneally, and electrodes were threaded subcutaneously to the cranium. After skull exposure, haemostasis, and identification of the cranial sutures bregma and lambda, two 1-mm diameter burr holes were drilled over the right olfactory bulb (reference) and left occipital cortex (active). The epidural electrodes of the telemetry units, connected to the leads of the transmitter, were placed into the burr holes, and secured using stainless steel skull screws. Once in place, the skull screws were covered with dental cement. Mice were subcutaneously injected 0 and 24 hours post-operatively with 5 mg/kg meloxicam for analgesia. After 1 week of recovery, mice were individually housed in their home cages in a 12/12 light/dark cycle, within a temperature- and humidity-controlled chamber with ad libitum access to food and water.

After a 24-hour acclimation period, one-channel EEG was recorded differentially between the reference (right olfactory bulb) and active (left occipital lobe) electrodes using the Ponemah acquisition platform (DSI). EEG, core-body temperature, and locomotor activity signals were continuously sampled from all mice for 48 hours along with time-registered videos. At the end of baseline acquisition, all mice were provoked with a convulsive dose (60 mg/kg; i.p.) of the GABA_a receptor antagonist pentylenetetrazole (PTZ; Sigma-Aldrich, Co.) to measure seizure susceptibility and evaluate seizure thresholds^46,76–78. Mice were continuously monitored for clinical and electrographic seizure activity for 20 minutes.

All data were processed and analyzed using Neuroscore software (DSI). Baseline EEG was analyzed for spontaneous seizure activity, circadian biometrics, and spectral power band analysis^76,77. Relative spectral power in delta (1–4 Hz), theta (4–8 Hz), alpha (8–12 Hz), beta (12–30 Hz), low gamma (30–60 Hz) and high gamma (60–90 Hz) frequency bands of the baseline EEG were calculated using the fast Fourier transform (FFT) technique.

PTZ-induced seizure activity was broadly scored on a modified Raccine’s scale as electrographic spikes (score: 1), myoclonic seizures (score: 3), generalized tonic-clonic seizures (GTC; score: 5) and death (score: 6). Per mouse, number of myoclonic seizures, latency and incidence of GTC seizures, number of GTCs, and total duration of GTC were recorded. Mice without seizures were assigned a time of 20 min at the end of the PTZ challenge observation period.