Quantification of volumetric morphometry and optical property in the cortex of human cerebellum at micrometer resolution

Abstract

The surface of the human cerebellar cortex is much more tightly folded than the cerebral cortex. Volumetric analysis of cerebellar morphometry in magnetic resonance imaging studies suffers from insufficient resolution, and therefore has had limited impact on disease assessment. Automatic serial polarization-sensitive optical coherence tomography (as-PSOCT) is an emerging technique that offers the advantages of microscopic resolution and volumetric reconstruction of large-scale samples. In this study, we reconstructed multiple cubic centimeters of ex vivo human cerebellum tissue using as-PSOCT. The morphometric and optical properties of the cerebellar cortex across five subjects were quantified. While the molecular and granular layers exhibited similar mean thickness in the five subjects, the thickness varied greatly in the granular layer within subjects. Layer-specific optical property remained homogenous within individual subjects but showed higher cross-subject variability than layer thickness. High-resolution volumetric morphometry and optical property maps of human cerebellar cortex revealed by as-PSOCT have great potential to advance our understanding of cerebellar function and diseases.

Keywords: Cell density; Cerebellum; Cross-subject variability; Layer thickness; Myelin; Polarization-sensitive optical coherence tomography.

Fig. 1.

as-PSOCT data acquisition and analysis pipeline.

Fig. 2.

Volumetric reconstruction of human cerebellar tissue (2.8 × 1.8 × 0.75 cm³) from as-PSOCT. The orthogonal views of (A) retardance images and (B) scattering coefficient maps are shown. The locations of the axial (xz) and coronal (yz) planes in A and B are indicated by the dashed lines on xy-plane. Scale bars: 5 mm.

Fig. 3.

Volume rendering of segmented molecular layer (red), granular layer (yellow) and white matter (cyan) for the cerebellar lobules. The whole segmented volume is shown in the composite image of the three structures.

Fig. 4.

Comparison of full manual (top) and semiautomatic (bottom) cerebellum segmentation on a cross-slice viewing plane. Black arrows highlight the inconsistency from manual segmentation and the smoothness from semiautomatic segmentation.

Fig. 5.

Cortical layer thickness in five cerebellar samples. (A) Molecular and granular layer thickness estimation in orthogonal views. The molecular and granular layer thicknesses are indicated by the hot and jet color bars, respectively. White matter is shown in white for anatomical reference. Scale bar: 5 mm. (B) Violin plots of molecular and granular layer thickness in individual subjects. The width and height of the violin plots correspond to the frequency and the value of thickness measurements. The mean and median of the measurements are also shown in the plots as black and blue solid lines. (C) Average cortical layer thickness of the five subjects (p = 0.66, paired-sample t-test). Error bars indicate standard deviations. Mol: molecular layer; Gra: granular layer.

Fig. 6.

Optical scattering coefficient measurements in five cerebellar samples. (A) Violin plots of μ_s measurements in molecular layer, granular layer, and white matter in individual subjects. The width and height of the violin plots correspond with the frequency and the value of μ_s measurements. The mean and median of the measurements are represented as black and blue solid lines. (B) Average μ_s measurements in the molecular layer, granular layer, and white matter across the five subjects (p = 0.0041, one-way ANOVA). Error bars indicate standard deviations. Mol: molecular layer; Gra: granular layer; WM: white matter.

Fig. 7.

Surface mapping of the granular layer. The vertex displacement map on the inflated surface indicates the crown of folium (red) and depth of fissure (blue) (A), surface maps of layer thickness (B), and optical properties (C) of the subject shown in Fig. 2 and Fig. 3. Color bars in (A)-(C) indicate the vertex displacement from the original surface to the inflated one, layer thickness and μs values, respectively, along the surface normal at a depth of 150 μm below the outer surface. Gray regions in (B) and (C) are masked out from the surface map (A).

Fig. 8.

Within-subject variation of morphometric and optical properties in the cerebellar cortex. (A) The variation index (p = 5.2e-5), skewness (p = 6e-5) and kurtosis (p = 0.21) of the molecular and granular layer thickness. (B) The variation index (p = 0.72), skewness (p = 0.24) and kurtosis (p = 0.1) of μ_s in the molecular and granular layers. Error bars indicate standard deviations among the five subjects. Mol: molecular layer; Gra: granular layer. The p-values are obtained from paired-sample t-tests.

Fig. 9.

Contour plots of 2D distribution between thickness and μ_s measurements in the molecular and granular layers of five cerebellar samples. The correlation coefficients (R) between layer thickness and μ_s are shown for each subject in molecular and granular layers. Mol: molecular layer; Gra: granular layer.