Page 146 - NNS 2014 Program & Abstract Book

D2-03
PRIMARY BLAST DOES NOT INCREASE VULNERABILITY
OF BRAIN TO SUBSEQUENT PRIMARY BLAST OR GLU-
TAMATE EXPOSURE
Effgen, G.B.1, Nammalwar, S.1, Bass, C.R.2 , Meaney, D.F.3, Morrison
III, B.1
1Columbia University, New York, NY,
2Duke University, Durham, NC,
3University of Pennsylvania, Philadelphia, PA
Soldiers who use explosives to breach perimeters can experience 20
explosive detonations during 5 days of practical training and report
symptoms of traumatic brain injury. Non-blast injury increases vul-
nerability of the brain to subsequent mechanical or excitotoxic injury.
It is unclear if primary blast increases the brain’s vulnerability to
subsequent injury, worsening outcome for soldiers exposed to blast.
Organotypic hippocampal slice cultures (OHSC) were generated
from P8-10 rat pups. A shock wave was generated with a shock tube
(424 – 6.4 kPa, 2.3 – 0.3ms, 248 – 3.4 kPa-ms). OHSC were placed in a
fluid-filled receiver at the exit of the tube. Sham cultures were treated
identically except the shock tube was not fired. For repetitive blast
studies, OHSC received 3 blast exposures within 10 minutes. For blast
and glutamate studies, OHSC received blast alone, blast followed by
glutamate (2.5mM, 3 hours), or sham followed by glutamate. Cell death
was quantified as the percentage area of a specific region of interest
(ROI: DG, CA3, or CA1) exhibiting propidium iodide fluorescence
above a threshold. As a positive control for cell death, OHSC were
treated with glutamate (10mM, 3 hours) following experimentation.
Cell death did not increase significantly following repetitive primary
blast (blast: ROI < 1%, n = 18; sham: ROI < 1%, n = 10). There was no
significant difference among samples receiving blast and glutamate
(ROI <1.5%, n = 6), blast alone (ROI < 1.5%, n = 6), or glutamate alone
(ROI <2%, n = 6). Cell death induced by 10mM glutamate (63% <
ROI < 90%, n = 18) indicated OHSC contained viable cells that were
not killed by the other injury paradigms.
These data suggest that primary blast does not increase brain tissue
vulnerability to subsequent glutamate or primary blast exposure. Our
data suggest that poor outcome after repetitive blast exposure may be
attributed to other blast-loading mechanisms such as tertiary blast
injury (i.e. inertia-driven injury), which have known potential to injure
brain.
Key words
blast, cell death, glutamate, hippocampus, shock tube, slice culture
D2-04
CELLULAR MECHANISMS OF PRIMARY BLAST- INDUCED
TRAUMATIC BRAIN INJURY: SHOCK-WAVE NEURO-
TRAUMA
Thorpe, C.N.1 , Ereifej, E.2, Hampton, C.2, Sykes, J.N.3, Rzigalinski,
B.A.1, VandeVord, P.2
1Edward Via College of Osteopathic Medicine, Blacksburg, USA
2Virginia Tech - Wake Forest Center for Injury Biomechanics,
Blacksburg, USA
3Virginia Polytechnic Institute and State University, Blacksburg, USA
Blast-induced traumatic brain injury (bTBI) is one of the most prevalent
injuries of American soldiers. Blast related TBI are conventionally di-
vided into four distinct phases: primary, secondary, tertiary, and qua-
ternary. Primary blast injuries occur as a direct result of blast-wave
induced changes in atmospheric pressure (barotrauma). While primary
blast forces are reported to contribute to brain injury, the exact mecha-
nism(s) by which primary blast damages the brain is poorly understood.
Blast-related pathologies are likely the results of blast loading conditions
such as pressure duration, peak magnitude, and rate of pressure change.
The focus of this study was to gain a better understanding of the primary
blast injury mechanism by observing the molecular response of primary
brain cells exposed to a shock wave. We utilized a novel shock wave
generator (SWG) that uses exploding wire in water to create the surge of
pressure. Primary mixed neuronal - glial cell cultures were blasted in the
SWG at pressures ranging from 5–15 psi for duration of less than one
millisecond. RNA was extracted at several time points post exposure and
Real-time PCR (RT-PCR) was performed to observe the gene expression
of cytoskeletal, astrocyte, and mechanotransduction markers. Results
indicate cells exposed to shock wave overpressure have cytoskeleton and
membrane disruption which lead to an increase of cell apoptotic markers
(Bax/Bcl2) and a decrease in cellular proliferation markers (MAP2k1)
over time. At 24 hours post blast exposures, cells had significantly higher
gene expression levels of Piezo2 (p < 0.003) as compared to the control.
Within 24 hours after exposure, the cells also showed an increase in
relative GFAP gene expression as compared to the sham. This study
indicates that shock wave overpressure leads to membrane damage which
could have led to cell death and a decrease in proliferation.
Key words
blast, traumatic brain injury
D2-05
EFFECTS OF MILD BLAST-INDUCED NEUROTRAUMA ON
BLOOD-BRAIN BARRIER PERMEABILITY
Ruppert, K.A., Parsley, M., Patrikeev, I., Motamedi, M., Sell, S.L.,
Prough, D.S., DeWitt, D.S.
University of Texas Medical Branch, Galveston, USA
The blood-brain barrier (BBB) regulates permeability of molecules
between the CNS and vascular circulation. Although much is known
of BBB integrity in other TBI models, effects of mild blast-induced
neurotrauma (BINT) on BBB remain unknown.
Adult, male rats were exposed to BINT using a device driven by
blank nail gun cartridges. Cerebral blood flow (CBF) and mean ar-
terial blood pressure (MAP) were recorded for one hour post-sham or
BINT injury by laser Doppler flowmetry (LDF) and tail artery, re-
spectively. Cerebral vascular reactivity was determined in isolated
middle cerebral arterial segments (MCA) following sham injury or
BINT. The integrity of the BBB at acute time points following sham
injury or BINT was determined by Evan’s Blue (EB) extravasation
quantified via fluorescent signal detection using an in vivo imaging
system (IVIS). Examining the role of peroxynitrite (ONOO-) in blast-
induced BBB permeability changes, ONOO- scavenger, penicilla-
mine, was injected following sham injury or BINT. Sham and BINT
animals also performed Morris water maze (MWM).
Following BINT, MAP significantly decreased to 39.81% of
baseline values (P < 0.0001; Sham, n = 6. BINT, n = 12). Cerebral
perfusion was significantly reduced to 43.05% of baseline (P = 0.0054;
Sham n = 6, BINT n = 12) following BINT. Dilator responses to re-
duced intravascular pressure in MCA were reduced significantly by
BINT (P < 0.0001; Sham n = 4, BINT n = 12). EB extravasation was
significantly greater 30 mins, 120 mins, 24 hours, and 3 days post-
BINT (P < 0.0001; per time point Sham n = 5, BINT n = 5). Sig-
nificantly less extravasation occurred in penicillamine-treated versus
untreated BINT animals at 30 mins (P < 0.0001). MWM latencies
were significantly longer (P < 0.01) at 24 hrs in BINT animals.
A-114