Upregulation of glial fibrillary acidic protein in satellite glia cells in the chronically

1
Upregulation of glial fibrillary acidic protein
in satellite glia cells in the chronically
compressed dorsal root ganglion Xiaofeng Mei1
Chao Ma1 Fletcher.A. White2 Pu Zhang1 R.H.
LaMotte1 1. Dept. Anesthesiol., Yale Univ. Sch.
Med., New Haven, CT, USA 2. Dept. Cell Biology,
Neurobiology Anatomy, and Anesthesiology,
Loyola Univ Medical Center, Maywood, IL, USA
511.22
GFAP-IR associated with different sized neurons
24 hours after CCD
GFAP-IR appears 6 12 hours after CCD
Introduction
Highest expression of GFAP at 3 days

Satellite glia cells (SGCs) in the dorsal root
ganglion (DRG) play a very important role in
response to ischemia, injury and inflammation.
Very little is known regarding the function and
the regulatory mechanisms of the SGCs in the
peripheral ganglia. Glial fibrillary acidic
protein (GFAP) is a biomarker for glial
activation in the central and peripheral nerve
system. Using immunohistofluorescence techniques,
GFAP was applied as a biomarker to evaluate the
activity of SGCs in a rat model of chronically
compressed dorsal root ganglion (CCD). Our
present study shows that chronical compression of
experimental DRG induces a rapid upregulation of
GFAP in SGCs. These results indicate that
satellite glia cells in DRG are possibly involved
in the early immune/inflammatory response to
compression and may contribute to the neuronal
excitability and pain behavior in CCD animals.
control
6 hours
12 hours
24 hours after CCD surgery, GFAP-IR was around
different size neurons .
As early as 6 hours after CCD surgery , GFAP-IR
was observed in compressed Ipsi-L4 DRG, 12 hours
became more significant.
3 days after CCD surgery, GFAP-IR reached the
highest level in DRG.
Glial activation More rapid and extensive in
compressed than in adjacent non-compressed DRGs
GFAP-IR Disappeared 1 2 weeks after onset of
CCD
Materials and Methods
1 week
24 hours
control
2 weeks
Animals and surgery Female adult Sprague Dawley
rats (160-200 g-wt, Charles River) were
anesthetized and a back midline incision was made
from L3 to L6. The right side paraspinal muscles
were separated from the mammillary process and
transvers process. A stainless steel rod,
L-shaped, 4mm in length and 0.6 mm in diameter,
was implanted into intervertebral foramen at L4
and the other at the L5. The muscle and skin
layers were sutured. Immunofluorescence After
defined survival times, control and injured rats
were terminally anesthetized and perfused through
the ascending aorta with PBS, followed by 4
paraformaldehyde in 0.1M phosphate buffer, pH7.4,
at 4C. After perfusion, the L3, L4, L5, L6,
contralateral L3, L4, L5 and L6 DRGs were removed
and postfixed in the same fixative for 2 hr and
then replaced with 30 sucrose in PBS overnight.
Ten-micron DRG sections were cut using a cryostat
and mounted onto slides. After postfixing in 4
paraformaldehyde-PB, the tissue sections were
blocked with 10 horse serum in 0.2 Triton X-100
for 1 hr at RT and incubated over night at 4C
with rabbit anti-GFAP antibody and Washed twice
in PBS. Sections were then incubated with
Cy3-conjugated donkey anti rabbit antibody for 1
hour. The Bisbenzimide H 33258 was employed as
nuclear staining. The slides were finally washed
and covered with cover slides. The slides were
then examined using a confocal microscope. The
images were analyzed using imageJv1.33 software.
GFAP-IR was always present at 24 hrs. in the
ipsilateral DRG but significantly less so at 1
week and virtually absent at 2 weeks
GFAP-IR is in non-compressed, adjacent DRG but
not in contralateral DRGs
Conclusions
Contralateral L4
Ipsi-L3 after 3 days
Ipsi-L3 after 1 week
Ipsi-L3 after 24 hours

• Satellite glial cells are activated in compressed
  and adjacent, non-compressed DRGs in rat CCD
  model
• Activation was more rapid and intense in the
  compressed DRG CCD surgery
• Activation was present around neurons of all
  sizes
• Time course of GFAP-IR parallels tactile
  allodynia after CCD (J. Neurophysiol. 823347).

GFAP-IR present in ipsilateral non-compressed L3
DRG(left) after CCD surgery at 24 hours, reaching
its highest incidence at 3 days and maintained
for 1 week. No GFAP-IR was found in contralateral
DRG from 6 hours to two weeks.
Supported by NIH grant NS 014624