Protocol for recording of neural activity in 3D midbrain organoids using a multiple-electrode array

Here, we present a protocol for electrophysiological recording from human three-dimensional (3D) organoids using substrates designed for two-dimensional cultures. We describe steps for generating midbrain 3D organoids from human induced pluripotent stem cells (hiPSCs) utilizing 6-well multi-electrode array (MEA) plates. We then detail procedures for recording neuronal activity via the neural module for Axion Maestro system. Our approach provides neural recording in organoids with high reliability and reproducibility, opening new avenues for research into human neurodevelopment and disease modeling.