UC San Diego

Autism spectrum disorder (ASD) is a multifactorial disease, suspected to originate during early fetal cortical development. Due to its symptomology and complex genetic etiology, developmental studies of autism largely rely on analysis of postmortem tissue. Recently, cortical organoids have impacted ASD research by providing highly organized tissue models that recapitulate early fetal human development. Here, forebrain cortical organoids (FBOs) derived from human induced pluripotent stem cells (iPSCs) are employed in a novel assembloid technique to model the basic organization principles of the cortical connectome. In vitro fusion of two FBOs results in a single assembloid unit that demonstrates key developmental processes: axon fasciculation and pathfinding. Characterization of the assembloid system was performed using immunohistochemistry and identified fascicules (“tracts”) which projected into the adjacent cortical unit and strongly correlated with an axonal identity. Additionally, electrophysiological recordings and correlative electron microscopy provide evidence of functional activity and reciprocal connectivity between assembloid “hemispheres”. Using the assembloid model, knock down of two high confidence ASD risk genes was assessed. Live cell imaging was used to capture short-term (seven days) and long-term changes (six weeks) in axon tracts. Findings from the project demonstrated that disruption of Contactin-associated protein-like 2 (CNTNAP2, Caspr2) or immunoglobulin superfamily containing leucine-rich repeat 2 (ISLR2, Linx) impact axon pathfinding and fasciculation. Knock down of CNTNAP2 resulted in absence of axon tracts during the early developmental timepoint, while knock down of ISLR2 resulted in a loss of fasciculation but not outgrowth, at both time points assayed. These findings elucidate the role of two key ASD risk genes during development and their putative impact on human cortical connectivity.