plasticity after SCI. To test the role of CP-AMPARs on spinal-
mediated behavioral plasticity, we delivered the specific CP-AMPAR
antagonist Naspm or vehicle following INS, and tested for adaptive
plasticity using a spinal instrumental learning assay. Naspm restored
adaptive learning. These findings highlight 1) the vulnerability of the
injured spinal cord, 2) the capacity for nociceptive input to produce
maladaptive plasticity after SCI, 3) the critical involvement of CP-
AMPARs in activity-dependent spinal plasticity and 4) the nociception-
induced loss of adaptive functional recovery after SCI. (Supported by
NIH-NS-038079, NIH-NS-069537, NIH-NS-067092).
Key words
AMPA receptor, nociception, spinal plasticity
T1-14
NEUROPROTECTIVE EFFECTS OF PAM3-CSK4 IN SPINAL
CORD INJURY
Okada, S.L.
1,2
, Stivers, N.
1,2
, Stirling, D.P.
1–3
1
Kentucky Spinal Cord Injury Research Center, University of Louis-
ville, Louisville, USA
2
Department of Neurological Surgery, University of Louisville,
Louisville, USA
3
Department of Microbiology and Immunology, University of Louis-
ville, Louisville, USA
Toll-like receptor 2 (TLR2) plays a protective role following spinal cord
injury (SCI); however, the TLR2-mediated cellular and/or molecular
mechanism(s) remains unclear. One possibility is that TLR2 signaling
modulates activation of macrophage/microglia and their subsequent
pro-inflammatory responses following injury. We hypothesized that
following SCI, the synthetic TLR2-specific agonist Pam3-CSK4 could
alternatively activate a neuroprotective macrophage/microglia phenotype
and improve neurological outcomes. In this study, we assessed the effect
of Pam3-CSK4 on 1) axonal retraction and secondary axonal degener-
ation; 2) macrophage/microglia immune functions; and 3) locomotive
recovery. We first used an
ex vivo
laser-induced SCI (LiSCI) model to
visualize myelinated axons responses after SCI in real-time with Pam3-
CSK4 or control. We then used a T9/10 moderate contusion SCI to
evaluate whether Pam3-CSK4 alters microglia and/or macrophage subset
dynamics and/or their functions (6-color flow cytometry) as well as
neurological recovery outcomes (Basso Mouse Scale). We found Pam3-
CSK4 tended to inhibit axonal retraction/dieback in
ex vivo
spinal cord
isolates after LiSCI. Moreover, Pam3-CSK4 decreased secondary axonal
degeneration. Our flow cytometry results indicated Pam3-CSK4 did not
alter the predominant pro-inflammatory M1 macrophage response which
is typical for contusion SCI; however, it significantly down-regulated
inducible nitric oxide synthase (iNOS) production in these cells. Char-
acterization of microglia responding to contusion SCI using the same
M1- and anti-inflammatory M2-associated markers revealed the major
microglia subset population exhibited a M2-like phenotype that was
functionally heterogeneous than macrophages. Nonetheless, similar to
M1 macrophages, Pam3-CSK4 significantly decreased iNOS production
in these M2-like microglia. Finally, Pam3-CSK4-treated mice showed
greater functional recovery after SCI compared to control-treated mice.
Overall, our data indicates that Pam3-CSK4 may improve neurological
recovery following SCI by down-regulating inflammatory macrophage/
microglia responses and modulating the extent of axonal injuries. This
research is supported by funding from the PVA Research Foundation
(grant 2934) and NIH (P30 GM103507).
Key words
axon, macrophage, microglia, Pam3-CSK4, SCI, TLR2
T1-15
RHOA/RHO KINASE REGULATES CPLA2 ACTIVATION IN
SPINAL CORD NEURONAL TOXICITY INDUCED BY TNF-
a
AND GLUTAMATE
Wu, X.B.
1–3
, Walker, C.L.
1–3
, Lu, Q.B.
1–3
, Xu, X.M.
1–3
1
Spinal Cord and Brain Injury Research Group, Indianapolis, USA
2
Stark Neurosciences Research Institute, Indianapolis, USA
3
Indiana University School of Medicine, Department of Neurological
Surgery, Indianapolis, USA
Activation of RhoA/Rho kinase leads to growth cone collapse and neurite
retraction. Although RhoA/Rho kinase inhibition has been shown to im-
prove axon regeneration, remyelination and functional recovery, its role in
neuronal death remains unclear. To determine whether RhoA/Rho kinase
play a role in neuronal death, we investigated the relationship between
RhoA/Rho kinase and cytosolic phospholipase A
2
(cPLA
2
), a lipase that
mediates inflammation and cell death, using an
in vitro
spinal cord neuronal
death model. We found that co-administration of TNF-
a
and glutamate
induced spinal neuronal death and activation of RhoA, Rho kinase and
cPLA
2
. Inhibitors of RhoA (CT04), Rho kinase (Y27632) and cPLA
2
(ATK) significantly reduced cell death by 33%, 52%and 43%, respectively
(
p
<
0.001). Additionally, RhoA and Rho kinase inhibition signifi-
cantly down-regulated cPLA
2
activation by 46% and 35%, respectively
(
p
<
0.01). RhoA and Rho kinase inhibition also reduced the release of
arachidonic acid (AA), a downstream substrate of cPLA
2
. Co-immuno-
precipitation (Co-IP) assay showed that ROCK1 or ROCK2, two isoforms
of Rho kinase, bound directly with the active phospho-cPLA
2
. Moreover,
application of lysophosphatidic acid (LPA), a small GTPase Rho activator,
activated Rho kinase and cPLA
2
and induced spinal cord neuronal death
in vitro
. Such cell death, however, was markedly reduced in postnatal
cortical neurons isolated from cPLA
2
knockout C57BL/6 mice. Taken
together, our results suggest that RhoA/Rho kinase may play a role in
mediating neuronal death by regulating cPLA
2
activation.
This work was supported in part by National Institutes of Health
NIH 1R01 NS059622.
Key words
cell signaling, cytosolic phospholipase A2, RhoA, Rho kinase
T1-16
18F-FDG PET IMAGING OF RAT SPINAL CORD INJURY
SHOWS DEPRESSED GLUCOSE UPTAKE CORRELATING
WITH LESION VOLUME & FUNCTIONAL RECOVERY
von Leden, R.E.
3
, Jaiswal, S.
2
, Wilson, C.M.
2
, Khayrullina, G.
1
,
Selwyn, R.G.
2
, Byrnes, K.R.
1,3
1
Department of Anatomy, Physiology, and Genetics, Uniformed Ser-
vices University, Bethesda, USA
2
Department of Radiology and Radiological Sciences, Uniformed
Services University, Bethesda, USA
3
Neuroscience Program, Uniformed Services University, Bethesda, USA
The use of non-invasive markers of neurological outcome after spinal
cord injury (SCI) are essential to predicting future outcome for pa-
tients and detecting responses to therapy and treatments. SCI results in
acute reduction in neuronal and glial cell viability near the injury site
and disruption in axonal tract integrity, as well as delayed but pro-
longed increase in glial activity and inflammation, all of which can
influence glucose uptake. Positron emission tomography (PET)-based
measurements of glucose uptake may serve as a novel biomarker for SCI.
This study aimed to determine the glucose uptake pattern in the spinal
cord after moderate SCI and correlate these findings with neuronal
A-7
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