Immunohistochemical Study of Calpain-Mediated Breakdown Products to α-Spectrin Following Controlled Cortical Impact Injury in the Rat

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Abstract
This study examined the effect of unilateral controlled cortical impact on the appearance of calpain-mediated α-spectrin breakdown products (BDPs) in the rat cortex and hippocampus at various times following injury. Coronal sections were taken from animals at 15 min, 1 h, 3 h, 6 h, and 24 h after injury and immunolabeled with an antibody that recognizes calpain-mediated BDPs to α-spectrin (Roberts-Lewis et al., 1994). Sections from a separate group of rats were also taken at the same times and stained with hematoxylin and eosin. Analyses of early time points (15 min, 1 h, 3 h, and 6 h following injury) revealed α-spectrin BDPs in structurally intact neuronal soma and dendrites in cortex ipsilateral to site of injury that was not present in tissue from sham-injured control rats. By 24 h after injury labeling was not restricted to clearly defined neuronal structures in ipsilateral cortex, although there was an increased extent of diffuse labeling. BDPs to α-spectrin in axons were not detected until 24 h after injury, in contrast to the more rapid accumulation of BDPs observed in neuronal soma and dendrites. The presence of BDPs to α-spectrin in the cortex at the site of impact, and in the rostral and contralateral cortex, coincided with morphopathology detected by hematoxylin and eosin. α-Spectrin BDPs were also observed in the hippocampus ipsilateral to the injury in the absence of overt cell death. This investigation provides further evidence that calpain is activated after controlled cortical impact and could contribute to necrosis at the site of injury. The appearance of calpain-mediated BDPs at sites distal to the contusion site and in the hippocampus also suggests that calpain activation may precede and/or occur in the absence of extensive morphopathological changes.