Understanding the complex communication between different cell populations and their interaction with the microenvironment in the central and peripheral nervous systems is fundamental in neuroscience research. Due to the lack of suitable animal models capable of faithfully reproducing the physio-pathological mechanisms of neurodegenerative diseases (NDDs), the development of appropriate in vitro approaches and tools has become crucial. The rising technology of Organ-on- Chip (OoC) can be useful for a better understanding of NDDs pathogenic mechanisms towards the development of patient-specific treatments. In this research, we develop custom-designed microfluidic devices suitable for culturing up to three cell populations to investigate the interaction between the human iPSC-derived cells of the neuromotor unit (i.e. motoneurons, Schwann cells and muscular cells) in the Amyotrophic Lateral Sclerosis (ALS). The implemented OoC and iPSC technologies allow to study specific pathological mechanisms behind the ALS and represent a robust platform for advanced in vitro drug testing.