Generation of glutamatergic/GABAergic neuronal co-cultures derived from human induced pluripotent stem cells for characterizing E/I balance in vitro

Obtaining mechanistic insights into the disruptions of neuronal excitation and in-hibition (E/I) balance in brain disorders has remained challenging. Here, we pre-sent a protocol for in vitro characterization of E/I balance. Using human induced pluripotent stem cells, we describe the generation of glutamatergic excitatory/ GABAergic inhibitory neuronal co-cultures at defined ratios, followed by analyzing E/I network properties using immunocytochemistry and multi -elec-trode array recording. This approach allows for studying cell-type-specific contri-bution of disease genes to E/I balance in human neurons. For complete details on the use and execution of this protocol, please refer to Mossink et al. (2022)1 and Wang et al. (2022).2