Microfluidic co-culture system for synaptically segregated neural networks to explore astrocyte-driven neural pathology.

Investigating astrocyte-neuron communication in the absence of neuron-to-neuron signalling is challenging using traditional culture systems due to the complexity of synaptic networks. To address this, we designed a three-compartment microfluidic co-culture device that fluidically isolates two neuronal populations while permitting astrocyte growth throughout. This design enables assessment of astrocyte-specific contributions to neuropathology between synaptically segregated neurons. The device incorporates ten microchannel banks forming maze-like structures that restrict neurite extension and fluid exchange, while allowing an astrocyte monolayer to infiltrate all compartments. Using this platform, we exposed one neuron-astrocyte population to the excitotoxin kainic acid (KA) and observed neurite degeneration in the adjacent, fluidically isolated neurons connected only via astrocytes. Pre-treatment of astrocytes with the membrane-permeable chelator BAPTA-AM markedly attenuated this effect, implicating calcium in astrocyte-mediated excitotoxicity. This microfluidic system provides a controllable in vitro model of neuron-astrocyte networks, enabling directional connectivity and mechanistic studies of circuit behaviour. Our findings highlight the utility of this platform for exploring intercellular signalling pathways relevant to neurodegenerative disease.