Avian species included in a comparative MRI dataset.

Overview of the 16 avian species included in this study, representing major avian clades and diverse ecological niches. (A) Phylogenetic relationships among the sampled species, grouped into four major avian lineages. (B) Representative coronal T2-weighted MRI sections and corresponding photographs of each species. These images illustrate the diversity of brain sizes and overall morphologies represented in the dataset and demonstrate sufficient internal contrast to identify major anatomical landmarks across taxa. All MRI datasets were processed using a standardized preprocessing workflow to enable direct cross-species comparisons.

Image sources for animal photographs used in Figure 1.

Whole-brain T2-weighted MRI highlights region-specific interspecific diversity in birds.

(A) Representative coronal T2-weighted images from multiple species reveal pronounced differences in the relative size and morphology of major brain divisions, as well as overall head morphology. The anterior commissure (AC), the principal interhemispheric commissure in birds, is readily identifiable across taxa (red arrows). (B) Quantification of AC size relative to whole-brain area across species. AC area scales approximately with brain size, indicating that apparent differences in AC prominence largely reflect variation in overall brain size rather than substantial differences in relative commissural investment. (C) Representative sagittal T2-weighted images allowing qualitative comparison of cerebellar morphology across species. Cerebellar foliation and overall cerebellar expansion vary markedly among taxa, with particularly elaborate folial patterns observed in penguins, toucans, and crows. (D) Representative cerebellar histological sections from selected species illustrating interspecific differences in folial branching and structural complexity. (E) Quantification of cerebellar folial branching. The number of basal branches (A) exhibits relatively limited interspecific variation, whereas the number of terminal branches (B) varies more extensively and is elevated in species with more highly elaborated cerebella. Together, these findings support a pattern of mosaic, region-specific diversification in avian brain organization.

Template-based registration enables standardized 3D morphometry and mosaic scaling quantification.

(A) Three-dimensional (3D) reconstructions of brains from 16 avian species, rendered from anterior-left (left) and posterior-right (right) viewpoints. The fidelity of the reconstructed surfaces reflects image acquisition resolution, with higher-resolution datasets producing smoother and more detailed reconstructions. (B) Schematic overview of the morphometric workflow, including species-specific template construction, registration into a common comparative space, and template-guided segmentation of major brain compartments. Representative overlays demonstrate the correspondence between MRI data and segmented 3D reconstructions. (C) Volumetric quantification of major brain divisions based on segmentation-derived voxel counts, expressed as proportions of total brain volume to facilitate comparison of relative regional investment across species. This workflow enables standardized evaluation of mosaic scaling across diverse avian lineages.

Histological validation links MRI-derived boundaries and diffusion-related measures to tissue architecture.

(Left) Representative Nissl-stained sections across selected taxa illustrating cytoarchitectonic transitions that align with major MRI-defined anatomical boundaries used for parcellation. (Middle) Luxol Fast Blue (LFB) staining highlights white-matter architecture and major fiber-rich compartments corresponding to MRI-defined white matter. Together, these histological observations support the cross-species consistency of the segmentation framework and provide tissue-level validation for the interpretation of MRI- and diffusion-derived metrics.

Diffusion MRI tractography and anisotropy mapping reveal conserved and divergent long-range organization across species.

(A) Region-seeded diffusion MRI tractography in chick, gentoo penguin, and large-billed crow. Streamlines were generated from four regions of interest (ROIs)—the optic lobe, dorsal cortex, cerebellum, and anterior/temporal cortex—and subsequently clustered to identify the major trajectory groups. (B) Whole-brain fractional anisotropy (FA) maps from representative species illustrating interspecific differences in regional microstructural organization. (C) Quantitative comparison of tractography- and FA-derived regional metrics in chick, gentoo penguin, and Egyptian rousette bat. The upper and lower tables summarize relative regional distributions derived from reconstructed trajectories and FA values, respectively. Together, these analyses indicate that species share broad organizational features while differing substantially in inferred long-range routing patterns and regional microstructural profiles.

Serial coronal and sagittal T2-weighted MRI sections of the mallard.

Representative serial T2-weighted MRI sections of the mallard brain are shown to provide an overview of whole-brain morphology. Consecutive coronal sections are shown together with sagittal sections to illustrate the anatomical continuity of the forebrain, midbrain, cerebellum, and brainstem. Scale bars, 10 mm.

Serial coronal and sagittal T2-weighted MRI sections of the mandarin duck.

Representative serial T2-weighted MRI sections of the mandarin duck brain are shown to provide an overview of whole-brain morphology. Consecutive coronal sections are shown together with sagittal sections to illustrate the anatomical continuity of the forebrain, midbrain, cerebellum, and brainstem. Scale bars, 10 mm.

Serial coronal and sagittal T2-weighted MRI sections of the crested partridge.

Representative serial T2-weighted MRI sections of the crested partridge brain are shown to provide an overview of whole-brain morphology. Consecutive coronal sections are shown together with sagittal sections to illustrate the anatomical continuity of the forebrain, midbrain, cerebellum, and brainstem. Scale bars, 10 mm.

Serial coronal and sagittal T2-weighted MRI sections of the domestic chicken.

Representative serial T2-weighted MRI sections of the domestic chicken brain are shown to provide an overview of whole-brain morphology. Consecutive coronal sections are shown together with sagittal sections to illustrate the anatomical continuity of the forebrain, midbrain, cerebellum, and brainstem. Scale bars, 10 mm.

Serial coronal and sagittal T2-weighted MRI sections of the pigeon.

Representative serial T2-weighted MRI sections of the pigeon brain are shown to provide an overview of whole-brain morphology. Consecutive coronal sections are shown together with sagittal sections to illustrate the anatomical continuity of the forebrain, midbrain, cerebellum, and brainstem. Scale bars, 10 mm.

Serial coronal and sagittal T2-weighted MRI sections of the black-necked stilt.

Representative serial T2-weighted MRI sections of the black-necked stilt brain are shown to provide an overview of whole-brain morphology. Consecutive coronal sections are shown together with sagittal sections to illustrate the anatomical continuity of the forebrain, midbrain, cerebellum, and brainstem. Scale bars, 10 mm.

Serial coronal and sagittal T2-weighted MRI sections of the blacksmith lapwing.

Representative serial T2-weighted MRI sections of the blacksmith lapwing brain are shown to provide an overview of whole-brain morphology. Consecutive coronal sections are shown together with sagittal sections to illustrate the anatomical continuity of the forebrain, midbrain, cerebellum, and brainstem. Scale bars, 10 mm.

Serial coronal and sagittal T2-weighted MRI sections of the gentoo penguin.

Representative serial T2-weighted MRI sections of the gentoo penguin brain are shown to provide an overview of whole-brain morphology. Consecutive coronal sections are shown together with sagittal sections to illustrate the anatomical continuity of the forebrain, midbrain, cerebellum, and brainstem. Scale bars, 10 mm.

Serial coronal and sagittal T2-weighted MRI sections of the Humboldt penguin.

Representative serial T2-weighted MRI sections of the Humboldt penguin brain are shown to provide an overview of whole-brain morphology. Consecutive coronal sections are shown together with sagittal sections to illustrate the anatomical continuity of the forebrain, midbrain, cerebellum, and brainstem. Scale bars, 10 mm.

Serial coronal and sagittal T2-weighted MRI sections of the boat-billed heron.

Representative serial T2-weighted MRI sections of the boat-billed heron brain are shown to provide an overview of whole-brain morphology. Consecutive coronal sections are shown together with sagittal sections to illustrate the anatomical continuity of the forebrain, midbrain, cerebellum, and brainstem. Scale bars, 10 mm.

Serial coronal and sagittal T2-weighted MRI sections of the northern red-billed hornbill.

Representative serial T2-weighted MRI sections of the northern red-billed hornbill brain are shown to provide an overview of whole-brain morphology. Consecutive coronal sections are shown together with sagittal sections to illustrate the anatomical continuity of the forebrain, midbrain, cerebellum, and brainstem. Scale bars, 10 mm.

Serial coronal and sagittal T2-weighted MRI sections of the toco toucan.

Representative serial T2-weighted MRI sections of the toco toucan brain are shown to provide an overview of whole-brain morphology. Consecutive coronal sections are shown together with sagittal sections to illustrate the anatomical continuity of the forebrain, midbrain, cerebellum, and brainstem. Scale bars, 10 mm.

Serial coronal and sagittal T2-weighted MRI sections of the common starling.

Representative serial T2-weighted MRI sections of the common starling brain are shown to provide an overview of whole-brain morphology. Consecutive coronal sections are shown together with sagittal sections to illustrate the anatomical continuity of the forebrain, midbrain, cerebellum, and brainstem. Scale bars, 10 mm.

Serial coronal and sagittal T2-weighted MRI sections of the zebra finch.

Representative serial T2-weighted MRI sections of the zebra finch brain are shown to provide an overview of whole-brain morphology. Consecutive coronal sections are shown together with sagittal sections to illustrate the anatomical continuity of the forebrain, midbrain, cerebellum, and brainstem. Scale bars, 10 mm.

Serial coronal and sagittal T2-weighted MRI sections of the canary.

Representative serial T2-weighted MRI sections of the canary brain are shown to provide an overview of whole-brain morphology. Consecutive coronal sections are shown together with sagittal sections to illustrate the anatomical continuity of the forebrain, midbrain, cerebellum, and brainstem. Scale bars, 10 mm.

Serial coronal and sagittal T2-weighted MRI sections of the large-billed crow.

Representative serial T2-weighted MRI sections of the large-billed crow brain are shown to provide an overview of whole-brain morphology. Consecutive coronal sections are shown together with sagittal sections to illustrate the anatomical continuity of the forebrain, midbrain, cerebellum, and brainstem. Scale bars, 10 mm.

Serial coronal and sagittal T2-weighted MRI sections of the Egyptian rousette.

Representative serial T2-weighted MRI sections of the Egyptian rousette brain are shown to provide an overview of whole-brain morphology. Consecutive coronal sections are shown together with sagittal sections to illustrate the anatomical continuity of the forebrain, midbrain, cerebellum, and brainstem. Scale bars, 10 mm.

Sagittal T2-weighted MRI sections showing the anterior commissure across bird species.

Representative sagittal T2-weighted MRI sections used for the analysis shown in Fig. 2A are shown for the examined bird species. The anterior commissure is indicated by red arrowheads. Species are grouped according to the categories used in the main figure. Scale bars, 10 mm.

T2-weighted MRI, three-dimensional reconstruction, and cerebellar foliation analysis of the Egyptian rousette.

Representative T2-weighted MRI sections and three-dimensional surface reconstructions of the Egyptian rousette brain are shown. The anterior commissure is indicated by red arrowheads in the MRI sections. Cerebellar foliation was assessed using sagittal T2-weighted MRI sections, and representative traces used for branch counting are shown. The number of basal branches and terminal branches is indicated as A and B, respectively. Scale bars, 10 mm.