Mouse brain organoids model in vivo neurodevelopment and function and capture differences to human

Abstract

In the last decade since their emergence, brain organoids have offered an increasingly popular and powerful model for the study of early development and disease in humans. These 3D stem cell-derived models exist in a newer space at the intersection of in vivo and 2D in vitro models. Functional benchmarking has so far remained largely uncharacterised however, leaving the extent to which these models may accurately portray in vivo processes still yet to be fully realised. Here we present a standardised unguided protocol to generate brain organoids from mice, the most commonly-used in vivo mammalian model; and in parallel establish a guided protocol for generating region-specific choroid plexus mouse organoids. Both unguided and guided mouse organoids progress through neurodevelopmental stages with an in vivo-like tempo and recapitulate species-specific characteristics of neural and choroid plexus development, respectively. Neuroepithelial cells generate neural progenitors that give rise to different neural subtypes including deep-layer neurons, upper-layer neurons, and glial cells. We further adapted protocols to prolong mouse cerebral organoid (CO) cultures as slices at the air-liquid interface (ALI), enhancing accessibility for long-term studies and functional investigations. In mature mouse ALI-COs, we observed mature glia, as well as synaptic structures and long-range axon tracts projecting to distant regions, suggesting an establishment and maturation of neural circuitry. Indeed, functional analyses with high-density multi-electrode arrays (HD-MEAs) indicate comparable activity to ex vivo organotypic mouse brain slices. Having established protocols for both region-specific and unpatterned mouse brain organoids, we demonstrate that their neurodevelopmental trajectories, and resultant mature features, closely mimic the in vivo models to which they are benchmarked across multiple biochemical, morphological, and functional read-outs. We propose that mouse brain organoids are a valuable model for functional studies, and provide insight into how closely brain organoids of other species, such as human, may recapitulate their own respective in vivo development.