The morphological changes in the anterior
horn of the L4 and L5 spinal segments were observed following anterior root avulsion in the
adult male Sprague-Dawley rat (300~350 gm) at 5 days, 1 week, 2 weeks and 3 weeks postlesion. The
animals were perfused with 4% paraformaldehyde, 0.15% picric
acid in 0.1 M
phosphate buffer solution and cryostat sections were prepared.
Immunohistochemistry was used to identify changes of the
phenotype in the
anterior horn cells. Primary
antibodies,
goat anti-
choline acetyltransferase (ChaT, 1 500, Chemicon),
mouse antirat ED-1 (1 200, Serotec),
rabbit anti-
glial fibrillary acidic protein (GFAP, 1 200, DAKO) and
rabbit anti-
vascular endothelial growth factor (
VEGF, 1 500, Santa Cruz
Biotechnology) were used.
Avidin-
Biotin complex
method was performed for immunohistochemical reaction and
color reaction was developed with DAB-H2O2. Following results were observed in the anterior
horn of lumbar
spinal cord; 1. The number of ChaT-immunoreactive (ir)
cells were reduced 20% level of control
animals at 3 weeks after avulsion. 2. ED-1-ir
microglia were significantly increased at 1 week and processes of ED-1-ir
microglia surrounded around the axotomized neuronal
cell bodies. 3.
Gliosis defined by extensive GFAP immunoreactivity was observed both ipsilateral and contralateral side of lesion but the
VEGF-ir
cells were significantly increased in the ipsilateral side of lesion. Therefore, this study suggested that the majority of axotomized
motor neurons were degenerated and the
cellular proliferation and
phenotype changes including
glial cell activation were observed in the lumbar
spinal cord after anterior root avulsion of
adult rats.