Engineering a functional neuro-muscular junction model in a chip

Healthy bi-directional intracellular transport along the axons between the somatodendritic and synaptic terminals is crucial to maintain the function and viability of neurons. When misbalanced, there is neuronal homeostasis failure that compromises its function and viability. In fact, several neurodegenerative diseases originate from misbalanced axonal transport and function. Thus numerous techniques have been developed to establish and maintain neuronal cultures in compartmented microfluidic devices to better understand these processes mimicking neuronal polarization. Although useful, these in vitro platforms do not allow for a full specific and temporal analysis in a completely monitored way. In this study, we have utilized a microfluidic system with large open cell culture reservoirs to precisely control neuronal microenvironments, capable of mimicking axon transport and synapse formation and to facilitate their analysis. We demonstrate using this lab-on-a-chip system for long-term motoneuron co-culture with C2C12-derived myotubes to mimic neuro-muscular junction (NMJ) formation. Furthermore, by integration with a calcium (Ca²⁺) imaging technique, we have proved the NMJ functionality in-chip through KCl-induced Ca²⁺ transient in connected myotubes. This platform can potentially become a useful tool as a straightforward, reproducible, and high-throughput in vitro model for basic NMJ research, and for high-throughput drug screening.