Neuroanatomical phenotypes in the reeler mouse

Abstract

The reeler mouse (Reln) has been proposed as a neurodevelopmental model for certain neurological and psychiatric conditions and has been studied by qualitative histochemistry and electron microscopy. Using magnetic resonance microscopy (MRM), we have quantitated for the first time the neuromorphology of Reln mice at a resolution of 21.5 microm. The neuroanatomical phenotypes of heterozygous and homozygous mutant Reln mice were compared to those of wild type (WT) littermates using morphometry and texture analysis. The cortical, hippocampal, and cerebellar phenotypes of the heterozygous and homozygous mutant Reln mice were confirmed, and new features were revealed. The Reln(rl/rl) mice possessed a smaller brain, and both Reln(rl/+) and Reln(rl/rl) mice had increased ventricles compared to WT controls. Shape differences were found between WT and Reln(rl/rl) brains, specifically in cerebellum, olfactory bulbs, dorsomedial frontal and parietal cortex, certain regions of temporal and occipital lobes, as well as in the lateral ventricles and ventral hippocampus. These findings suggest that certain brain regions may be more severely impacted by the Reln mutation than others. Gadolinium-based active staining demonstrated that layers of the hippocampus were disorganized in Reln(rl/rl) mice and differences in thickness of these layers were identified between WT and Reln(rl/rl) mice. The intensity distributions characteristic to the dorsal, middle, and ventral hippocampus were altered in the Reln(rl/rl), especially in the ventral hippocampus. These differences were quantified using skewness and modeling the intensity distributions with a Gaussian mixture. Our results suggest that structural features of Reln(rl/rl) brain most closely phenocopy those of patients with Norman-Roberts lissencephaly.

Figure 1

Mouse brain structures visible by MRM with resolution to 21 μm in Reln mice. Horizontal (Left), coronal (Middle), and sagittal (Right) sections through the brains of a WT control (Top), Reln^rl/+ (Middle), and Reln^rl/rl mouse (Bottom). The stain emphasizes cellular layers of the hippocampus where the pyramidal (Py) and granule cells (GrDG) can be visualized. These cell layers have high contrast and a characteristic shape in WT and Reln^rl/+ mice, but appear disorganized in Reln^rl/rl animals. The ventricles appear enlarged in the Reln^rl/+ and Reln^rl/rl mice; the cerebellum is atrophied in the homozygous mutant animals.

Figure 2

Segmentation of the ventricles and hippocampus in Reln mice. The lateral ventricles (cyan) and hippocampus (yellow) are depicted in WT (Left), Reln^rl/+ (Middle), and Reln^rl/rl (Right) animals in a background of the brain. Enlarged ventricles can be seen for the Reln^rl/+ and Reln^rl/rl mice; the hippocampus is also smaller for the homozygous mutants. The olfactory bulbs appear disorganized in the Reln^rl/rl animals and they seem deformed in Reln^rl/+ mice relative to the WT controls. Layers of the cerebral cortex can be visualized in the WT animal, but are not as easily seen in Reln^rl/rl mice.

Figure 3

Three-dimensional rendering of the whole brain, ventricles, and hippocampus of Reln mice. The mean surfaces are pseudo-colored according to the root mean square of the distance within the WT group (Left). The mean surfaces for the Reln^rl/+ (Middle) and Reln^rl/rl mice (Right) are pseudo-colored according to the distances between the respective mutant group of mice and the WT animals. Histograms of the distance distributions are depicted for whole brain (Top), ventricles (Middle), and hippocampus (Bottom). Most of the values for whole brain in WT and Reln^rl/rl mice are concentrated in the lower range of distances, whereas those of the Reln^rl/rl animals are more evenly distributed across the whole range of values. With regards to the ventricles, distance values for the WT controls are restricted to 0.01–0.8 mm, while those for Reln^rl/+ and Reln^rl/rl mice span a much larger range (0.01–2.65 and 0.01–2.77 mm, respectively). A similar pattern is seen for hippocampus for the three genotypes.

Figure 4

Shape analysis for the surface of the whole brain for Reln^rl/rl mouse. Signed distance map between the mean meshes for the WT and Reln^rl/rl mice indicate inward deformations in the regions of the olfactory bulbs and cerebellum. Temporal cortex (blue) is also deformed. The positive distances are shown in the “warm” color scale, whereas the negative distances are depicted in the “cold” color scale. To the right of each brain are the statistical maps of distances based upon Hotteling tests show that shapes are significantly reduced in the Reln^rl/rl olfactory bulbs, medial frontal and parietal cortex, temporal and occipital lobes, cerebellum, and brainstem.

Figure 5

Caudal view of whole hippocampus of WT and Reln^rl/rl mice. (A) The mean shapes for the WT (green) and Reln^rl/rl (red) hippocampus. Differences can be discerned between genotypes in the dorsal, middle, and ventral hippocampus where this brain area is more compressed in the mutant mouse. (B) Unsigned distance maps quantify differences in shape between WT and Reln^rl/rl hippocampus. The greatest distance between groups is present in the ventral or tail region of the hippocampus. (C) Signed distance map shows a compression in shape of dorsolateral hippocampus in Reln^rl/rl mice.

Figure 6

Grayscale characterization of hippocampus in Reln mice. A slice-by-slice analysis of skewness values is used to evaluate genotype differences along the rostral to caudal axis of the hippocampus. Inspection of the histograms reveals that the skewness values for hippocampus are similar for WT and Reln^rl/+ mice and these values are different from those of the Reln^rl/rl animals. Although differences are observed in the dorsal, middle, and ventral hippocampus, the most robust changes occur in the dorsal (Inset-- Right) and ventral (Inset--Left) hippocampus.