A 3D Amphioxus Brain Atlas Illuminates the Blueprint of the Ancestral Chordate Brain and Highlights Unique Vertebrate Innovations

To reconstruct the cellular and molecular origins of brain architecture in chordates, we combined single-nucleus RNA-seq with spatial transcriptomics to generate a three-dimensional cell atlas of the central nervous system (CNS) in the early-branching chordate amphioxus.

This atlas reveals a tripartite anterior-posterior organization comprising a rostral retinal/hypothalamic region, a di-mesencephalic domain, and a caudal hindbrain/spinal cord. Challenging previous views, we identify cell clusters resembling the vertebrate zona limitans intrathalamica and midbrain-hindbrain boundary regions that demarcate major brain compartments. Conversely, the absence of telencephalic marker expression in the amphioxus rostral forebrain suggests that the telencephalon is a vertebrate innovation.