Human cerebral tissues created via active cellular reaggregation produce functionally interconnected 3D neuronal network to mimic pathological circuit disturbance

Various characteristics of a developing central nervous system, including sensory input-output1,2, neuronal migration3,4, and regionalization3,5, have been recapitulated through recent advances in the culture of brain organoids. These organoids can model the fundamental processes in brain development and disease. A remaining critical challenge, however, is to achieve complex neuronal networks with functional interconnectivity as in native brain tissue. Generation of current organoid models originates from classic dissociation-reaggregation paradigms6, often relying on mechanically-enforced quick reaggregation of pluripotent stem cells7. Here we describe an alternative method that promotes matrix-supported active (migrative) reaggregation of cells (MARC), reminiscent of in vivo developmental morphing processes, to engineer multi-regional brain tissues in vitro. Measurements of neuronal activity in intact 3D tissues revealed functional interconnectivity, characteristic of cerebral neuronal networks. As a proof of concept, we show that interconnected cerebral tissues produced using this approach can mimic propagation of epileptiform discharges in a custom-built in-vitro platform.