A Reproducible Cortical Model of Focal Epileptiform Activity with a Multi-Biomarker Analysis Pipeline

The development of emerging optical and epicortical neuromodulatory strategies requires experimental models in which focal pathological activity can be generated and monitored within an accessible cortical region. Here, we established an acute cortical model of focal epileptiform activity in anesthetized mice based on local microinjection of 4-aminopyridine (4-AP) into the barrel cortex. This preparation provides precise spatial and temporal control of the epileptiform focus while enabling simultaneous multichannel electrophysiological recording. To objectively characterize the induced activity, we implemented a Python-based analysis pipeline integrating complementary electrophysiological biomarkers, including frequency-specific power, multi-unit activity (MUA), pathological fast ripples (250–650 Hz), and epileptiform bursts. Local 4-AP administration produced marked increases in high-frequency (HF; 100–250 Hz) power, multi-unit spike rate, fast ripple rate, and cumulative burst duration. Pharmacological sensitivity of the framework was evaluated using the antiseizure drug carbamazepine, which consistently attenuated these 4-AP-induced electrophysiological alterations, bringing values close to those observed in vehicle-treated animals. Together, this study provides a reproducible experimental and analytical framework for investigating focal cortical epileptiform activity and for the future evaluation of pharmacological, optical, and electrical interventions targeting superficial cortical networks.