Functional Characterization of Three-Dimensional Cortical Cultures for In Vitro Modeling of Brain Networks

Three-dimensional (3D) in vitro cultures recapitulate key features of the brain including morphology, cell-cell and cell-extracellular matrix interactions, gradi-ents of factors, and mechanical properties. However, there remains a need for experimental and computational tools to investigate network functions in these 3D models. To address this need, we present an experimental system based on 3D scaffold-based cortical neuron cultures in which we expressed the genetically encoded calcium indicator GCaMP6f to record neuronal activity at the millimeter -scale. Functional neural network descriptors were computed with graph-theory -based network analysis methods, showing the formation of functional networks at 3 weeks of culture. Changes to the functional network properties upon pertur-bations to glutamatergic neurotransmission or GABAergic neurotransmission were quantitatively characterized. The results illustrate the applicability of our 3D experimental system for the study of brain network development, function, and disruption in a biomimetic microenvironment.