IMPACT OF ELECTRICAL STIMULATION EMULATING INPUT ACTIVITY ON THE MATURATION OF RAT CORTICAL NETWORKS IN VITRO

In vitro models of cortical networks, from dissociated cultures to organoids, commonly show hypersynchronous bursting, a pattern largely absent in in vivo recordings and thought to reflect immature dynamics. We hypothesized that this in vitro hypersynchrony arises from the absence of early developmental inputs (via self‑amplification mechanisms that boost basal activity levels), and that providing patterned electrical stimulation (ES) mimicking endogenous activity can drive rat cortical networks toward more physiological, in vivo–like states.

We test this by chronically stimulating primary rat cortical cultures during several days of development (DIV 1 to DIV 7). This is achieved using microelectrode arrays (MEAs), which allow continuous recording and stimulation inside an incubator. We explore developmental‑like ES protocols, namely 40 Hz gamma‑like bursts (5 pulses at 40 Hz, every 30 s) and 10 Hz spindle‑burst‑like trains (10 pulses at 10 Hz, every 10 s), and compare to non-stimulated controls. We also perform sparse AAV labeling (e.g., inhibitory-specific) and confocal imaging to assess effects on morphological maturation, axodendritic complexity, and synaptic density in a cell-type-specific way.

The impact of different “input landscapes” was assessed by comparing baseline activity and network statistics between stimulated and non-stimulated cultures. In parallel, we studied how chronic ES influences morphological maturation, including axodendritic organization and synaptic density, and whether effects differ between excitatory and inhibitory neurons. These experiments are designed to clarify the relation between in vitro hypersynchrony and lack of developmental inputs, and determine how patterned activity shapes cortical network maturation in culture.