Fig. 7

Variability in the arrangement of Purkinje cells (PCs) in the squamate cerebellum. a, b Representative light-sheet microscopy imaging of cleared whole-cerebella showing the 3D distribution and arrangement of calbindin 1 (CALB1)-immunolabelled PCs in two representative species with different locomotor modes (see colour code and symbols in top left corner): Pogona vitticeps (a) and Boaedon fuliginosus (b). The boxed areas in the coronal 3D-rendered cerebellar views (top panels) are shown at higher magnifications in coronal (left) and sagittal (right) views in the lower panels. c, Violin plot showing the quantitative distribution of CALB1-immunolabelled PCs in the cerebellar cortex of selected squamate species with similar or different locomotor modes (colour code and symbols as above). Due to intra- and interspecies heterogeneity in molecular layer (ML) thickness, the position of individual cells (n = 250–750 per species) was calculated as the distance (in %) from the granule cell layer (GCL) to the outer border (pial surface) of the ML, and error bars represent the standard deviation. Four major positioning patterns containing three or four squamate species and reflecting the increased scattering of PCs (from I to IV) were identified based on Kruskal-Wallis statistics. Immunohistochemistry with CALB1 marker (red staining) on sagittal sections of the cerebellar cortex in selected representative species are shown at low and high magnifications (insets) for each pattern: Pogona vitticeps (I), Eryx colubrinus (II), Pseudopus apodus (III), Dasypeltis gansi (IV). Source data are provided as a Source Data file. Scale bars: 30 μm (a, b), 100 μm (c).