Cytological compartmentalization in the staggerer cerebellum, as revealed by calbindin immunohistochemistry for Purkinje cells

Abstract

The staggerer mouse carries a deletion in a gene encoding the nuclear hormone receptor RORalpha, which leads to severe impairments in phenotypic differentiation of cerebellar Purkinje cells. We previously found parasagittal compartments in the mature staggerer cerebellum, as defined by different transcription levels of Purkinje cell-specific molecules including calbindin. In the present study, we developed a hightiter anti-calbindin antibody to examine morphological features of the staggerer Purkinje cells. Immunohistochemistry for calbindin revealed compartmentalized Purkinje cell populations with different cell sizes, alignments, cell densities, and dendritic arborization, as well as different immunoreactivities, corresponding to the "transcriptional" compartments. Based on these immunohistochemical and cytological characteristics, the rostral cerebellum was clearly subdivided into three to seven parasagittal zones (Zones I-VII). Purkinje cells in Zones I and III were associated with the strongest calbindin immunoreactivities and exhibited morphological features reminiscent of the wild-type cells, i.e., large flask-shaped cell bodies, monolayer alignment, and arborized dendrites. Purkinje cells in Zone V were also labeled strongly, but they were small in cell size, ectopic and possessed long unbranched dendrites. On the other hand, Purkinje cells in Zones II, IV, and VI were very low in calbindin immunoreactivity and marked by small cell size, ectopia, poorly-developed dendrites and low cell density. Considering that this unique cytological compartmentalization emerges as the result of RORalpha gene mutation, it is suggested that normal cytodifferentiation of Purkinje cells is governed by both RORalpha-dependent and -independent mechanisms, and further that the latter mechanism might exert unevenly along the mediolateral cerebellar axis.

Hematopoietic stem cell gene therapy leads to marked visceral organ improvements and a delayed onset of neurological abnormalities in the acid sphingomyelinase deficient mouse model of Niemann–Pick disease

“…Calbindin D28k is a member of the large EF-hand family that is believed to function, in part, as a cytosolic calcium buffer, and encodes a calcium binding protein that is expressed in the cerebellum exclusively in PC. [27][28][29] Our results demonstrated that at 5 months after transplantation, there was an increased number of PC in treated mice as compared with affected, irradiation control mice. These findings were most likely due to the presence of retrovirally transduced, bone marrow-derived microglial cells in the CNS that were capable of secreting ASM in sufficient amounts to delay the disease-specific cell death in PC neurons.…”

P/Q-Type Ca2+Channel α1A Regulates Synaptic Competition on Developing Cerebellar Purkinje Cells

“…We used rabbit calbindin antiserum (1:10,000), guinea pig vesicular glutamate transporter 1 (VGluT1) antibody (1 g/ml), and guinea pig VGluT2 antibody (1 g/ml), the specificities of which have been reported previously (Nakagawa et al, 1998;Miyazaki et al, 2003). For immunofluorescence, FITC-, indocarbocyanine-, and aminomethylcoumarin acetate-labeled species-specific secondary antibodies were used at a dilution of 1:200 (Jackson Immunoresearch, West Grove, PA).…”

Control of Synaptic Connection by Glutamate Receptor δ2 in the Adult Cerebellum

“…), mice were perfused transcardially with 4% paraformaldehyde in 0.1 M sodium phosphate buffer, pH 7.4, and processed for paraffin sections (5 m) with a sliding microtome (SM2000R; Leica Microsystems, Nussloch, Germany). Paraffin sections were stained with hematoxylin or immunostained with rabbit antiGluR␦2 antibody (Araki et al, 1993) and guinea pig anti-calbindin antibody (Nakagawa et al, 1998) followed by incubation with speciesspecific FITC-and Cy3-conjugated secondary antibodies (Jackson ImmunoResearch, West Grove, PA). Photographs were taken by a fluorescence microscope (AX-70; Olympus, Tokyo, Japan) or a confocal laser scanning microscope (Fluoview; Olympus).…”