Mechanical Actuation of Organoids in Synthetic Microenvironments

Organoids are a powerful model system to explore the role of mechanical forces in sculpting emergent tissue cytoarchitecture. The modulation of the mechanical microenvironment is most readily performed using synthetic extracellular matrices (ECM); however, such materials provide passive, rather than active force modulation. Actuation technologies enable the active tuning of mechanical forces in both time and magnitude. Using such instruments, our group has shown that extrinsically imposed stretching on human neural tube organoids (hNTOs) enhanced patterning of the floor plate domain. Here, we provide a detailed protocol on the implementation of mechanical actuation of organoids embedded in synthetic 3D microenvironments, with additional details on methods to characterize organoid fate and behavior. Our protocol is easy to reproduce and is expected to be broadly applicable to investigate the role of active mechanics with in vitro model systems.