The complex neurological and neuropsychiatric clinical picture presented by injured soldiers and civilians exposed to IEDs far exceed the physical injuries following exposure to low-level blast waves (Crabtree, 2006
; Mekel et al., 2009
). The spectrum of moderate to severe brain injury is easily detectable, both clinically by focal neurological signs and by neuroimaging (Mintz et al., 2002
) as well as by altered (deficient functional synchronization) EEG activity (Sponheim et al., 2011
). However, the affective, cognitive, and behavioral changes that frequently follow mild, or mild-to moderate brain injury are more problematic, particularly in cases with absent or transient focal neurological signs and neuroimaging studies are show no evidence of abnormalities. In these cases, the mere existence of a clinical entity has been controversial (Brenner et al., 2009
; Pietrzak et al., 2009
). As a consequence, symptom reports can be attributed erroneously to motives for secondary gain (Hoge et al., 2009
). Various laboratory models of blast induced mild TBI have addressed this gap in knowledge, but none of them used “real-world” conditions. In order to mimic the exposure to a mild blast in as-realistically-as-possible, found that mice exposed to very low intensity explosions exhibit both behavioral and MRI results compatible with diffuse mild brain injury.
Study of the pathophysiological mechanisms that underlie blast-induced brain injury is needed both for a better understanding of the clinical entity, and for the further development of possible treatments. This can be done best in an animal model that controls for as many variables as possible as well as resembles, as much as possible, “real life” conditions. We recognize that inferences from an animal-model are limited by factors that include the structural differences between murine and human brains and biophysical factors related to head morphology (including skull thickness). However, the mechanisms of injury and responses can provide insights for rational translational research in humans.
Our mouse-model of blast-induced brain injury fulfills the requirements of replicating key features of clinical blast-related mTBI. Almost all mice exposed to blast (64 mice for a 7m distance/2.5PSI and 62 mice for a 4m distance/5.5PSI) survived and recovered from anesthesia in a manner similar to the control mice. Moreover, upon visual and light microscopic examination, no gross anatomical damage was found acutely in the blast-exposed mice. By seven days after blast exposure, there was only a marginal trend toward alteration of blood-brain barrier integrity. However, the neurological score test, no differences between the blast exposed and the sham group during the first week after blast exposure. These findings suggest that the degree of injury in this model is comparable with human ‘mild’ blast induced brain injury.
The results from mice in our open field model are consistent with findings from other species that show damage after low overpressure exposures in shock tube models of brain blast injury. Pioneering studies by Saljo and co-workers (Saljo et al., 2001
) documented hippocampal and cerebral cortical signs after single relatively high level shock wave exposure (198–202 dB peak overpressure re: 20 μ Pa; 25–35 PSI) to rats in a blast tube. These changes included evidence of apoptotic neurons, persistent changes in phosphorylated neurofilament proteins and transcription factors and a proliferation of microglia and astrocytes for at least 3 weeks after exposure. The mouse model did not exhibit small parenchymal hemorrhages that have reported in the occipital cortex, cerebellar cortex and medulla of 30–40% pigs exposed to impulse noise in the 1.3–6 PSI range (Saljo et al., 2008
). However, the behavioral findings from our mouse model are consistent with the 174 dB SPL (1.5 PSI) over-pressure threshold for abnormal Morris water maze behavior from single blast exposure in rats (Saljo et al. 2009
), Despite potential differences due to species and experimental design and, the mouse, rat and pig data all indicate that low blast exposures have persistent and evolving neurobehavioral consequences.
The combination of the behavioral studies and the MRI findings suggests diffuse but subtle CNS damage. This might be expected to appear following the double impact of the blast wave: a high-pressure phase, followed closely by a low-pressure phase. (Courtney and Courtney, 2009
; Elder and Cristian, 2009
; Moss et al., 2009
). Behavioral and cognitive testing at one week and at one-month post exposure, suggest that there are both persistent and progressive deficits after only a single, low-level blast exposure. The rearing behavior on the staircase test and the discrimination index in the novel object recognition test showed deficits at 7 days that persisted at 30 days after blast exposure, but did not vary in severity between the two low level (2.5 and 5 PSI) blast exposures. However, the discrimination index for Y-maze performance varies with blast exposure. After the lower 2.5 PSI peak exposure a deficit at 7 days resolved partially by 30 days after the blast. A single 5 PSI peak exposure, though, produced a deficit at 7 days that persisted at 30 days.
We suggest the following interpretation of these abnormal tests. A recent evaluation of the novel object recognition test (McTighe et al., 2010
) has indicated that,, rather than a loss of memory for the familiar object or deficit in executive function, there is a “false memory” of the novel object as familiar after perirhinal cortex damage. Furthermore, although no motor impairments were found at the blast group (as seen in the “steps ascended” parameter of the staircase test), the equivalent of a typical clinical picture of restlessness/agitation recognized in people with blast induced mTBI, was evident in the blasted mice from the “rearing” parameter of this test. The fact that the blast-induced changes in these two different tests were not identical might be due to the fact that these are two separate systems. Together, these results suggest a clear and pronounced, combined cognitive and behavioral deficit in the blast-exposed mice that develops during a subacute to chronic post-exposure period.
In the MRI study, significant alterations were found with T1 weighted images showing an increased BBB permeability one month post-blast. BBB rupture is a common consequence of traumatic brain injury and a leading cause for secondary brain damage immediately after injury (Beaumont et al., 2000
; Vajtr et al., 2009
). However, we did not see BBB disruption until one month after these very low intensity blast exposures, and only in the mice exposed to the lower blast level (2.5 PSI/7 m). Because factors such as inflammatory processes can interfere with repair of leakage in some neuroinflammatory conditions (de Vries et al., 1997
), an explanation may come from elucidation the long- term time course of inflammatory mechanisms after similar exposures.
Diffusion Tensor Imaging (DTI) has been suggested as a diagnostic tool for microstructural alteration in brain tissue, especially for axonal and myelin pathologies (Jiang, Q et al., 2006
; Jiang, Y et al., 2010
; Knake et al., 2010
; Mori et al., 2006
; Song et al., 2002
). DTI investigates tissue microstructure of the brain and allows extraction of FA indices (fractional anisotropy), which relates to the level of white matter organization; ADC map (apparent diffusion coefficient), which is related to the isotropic mobility of water; and axial λ1
and radial λ2,
, are related to the diffusivity along the long and short axes of the axons, respectively.
Our data revealed a significant increase in FA values with a concomitant decrease in λ3
values in blasted mice compared with controls, particularly in the hypothalamus,. These results suggest blast-induced microstructural changes. Elevated FA values are correlated with axonal cytotoxicity and edema (Bazarian et al., 2007
; Wilde et al., 2008
). Reduced λ3
values are compatible with myelin abnormalities. Both FA and λ3
values exhibit a more pronounced change over time (30 days>/< 7 days, respectively). This may reflect a time dependent process. These DTI results are in agreement with some of the cognitive and behavioral results, where more deficits are found at 30 days post blast. These findings correlate with Sponheim et al., (2011)
who recently published an elegant study where blast injured patients exhibited diminished EEG phase synchrony of lateral frontal sites with contralateral frontal brain regions, suggesting diminished inter-hemispheric coordination of brain activity following the blast injury. A similar trend in the DTI test was recently found in mTBI patients after blunt force (Chu et al., 2010
) and blast trauma (MacDonald et al., 2011
), suggesting a possible apoptosis of synapses as the underlying mechanism of damage (Coleman and Perry, 2002
). However, a similar study with blast mTBI patients showing no changes in DTI parameters, possibly due to the length of time (29 months) from the injury (Levin et al., 2010
). Reduced λ3
values might reflect an increased affinity of myelin to the axon (due to alteration in ganglioside composition) thus preventing its regeneration following injury. It is noteworthy that an abnormal brain condition is known as “axonal outgrowth- inhibition” suggested previously in other models of TBI (Schnaar 2010
; Vyas et al., 2002
Examination of histological sections of decalcified heads showed normal structure of the brain, spinal cord, vasculature and meninges at survival times up to 72 hours. However, immunohistochemical observations revealed changes suggestive of early inflammatory and oxidative stress responses in the hypothalamic regions that showed later changes in the imaging studies. Neuronal immunoreactivity for the mitochondrial anti-oxidant enzyme, manganese superoxide dismutase 2, was augmented markedly in the ventral aspect of the hypothalamus at 72 hours after the higher blast exposure. Increased immunoreactivity for the CXC chemokine receptor 3 (CXCR3) was also increased in association with blood vessel profiles in the fornix, optic tract and crus cerebri. The apparent upregulation of CXCR3 is of interest because this receptor has been linked to both vascular remodeling and development of autoimmune responses, such as endocrine autoimmunity and multiple sclerosis (Omari et al 2005
; Rotondi et al. 2007
; Lacotte et al. 2009
). Because the primary ligands for CXCR3 (CXCL9, CXCL10 and CXCL11) are induced by interferon-γ, it will be important to explore the interplay between inflammatory and vascular remodeling mechanisms as factors in the chronic behavioral and cognitive consequences of mild blast TBI., The importance of such studies for longer-term neurological disorders, such as dementia and Alzheimer’s disease, has recently been emphasized (DeKosky et al, 2010