Fig. 5. Class-B neurons expanded in human lateral nucleus.

(A) Workflow for human snRNAseq. The three cerebellar nuclei are separately dissected from frozen tissue. (B) Marker expression for all neurons. N = 3 donors. Dashed line divides excitatory and inhibitory neurons. (C) Clustering results of human excitatory neurons, colored by cluster assignment and dissection. Dissection labels are imperfect, owing to close apposition of individual cerebellar nuclei. Med/Int indicates a mixed dissection. (D) Correlation matrix of mouse and human excitatory cell types. Medial and interposed nuclei contain cell types that correlate with both mouse Class-A and Class-B cell types. Lateral nucleus neurons only correlate with Class-B neurons. Dots indicate significant correlations. (E) Seurat integration of excitatory neurons from three species, colored by class (left), clustering results in integrated space (middle), and species (right). (F) Quantification of membership to the integrated clusters of Class-A and Class-B cells. Across species, Class-A and Class-B cells fall into the same clusters. (G) Hierarchical clustering of excitatory neurons averaged by class, showing conservation of excitatory cell classes across amniotes. Grayscales of line and numbers indicate bootstrapping-based branch confidence. (H) Clustering results of human inhibitory neurons. Cells are colored by cluster assignment. Marker expression is indicated in the inset. (I) Correlation matrix of mouse and human inhibitory neurons, showing one-to-one correspondences. Dots indicate significant correlations. (J) Hierarchical clustering of inhibitory cell types across all three species (color coded), showing conservation of three inhibitory classes across amniotes. Grayscales of line and numbers as above. (K) Schematic illustrating the proposed model of subnucleus duplication-and-divergence (left) and biased expansion of Class-B excitatory neurons in human lateral nucleus (right).