Treatment of Swine Closed Head Injury with Perfluorocarbon NVX-428.

Pre-hospital treatment of traumatic brain injury (TBI) with co-existing polytrauma is complicated by requirements for intravenous fluid volume vs. hypotensive resuscitation. A low volume, small particle-size-oxygen-carrier perfluorocarbon emulsion NVX-428 (dodecafluoropentane emulsion; 2% w/v) could improve brain tissue with minimal additional fluid volume. This study examined whether the oxygen-carrier NVX-428 shows safety and efficacy for pre-hospital treatment of TBI. Anesthetized swine underwent fluid percussion injury TBI and received 1 mL/kg IV NVX-428 (TBI-NVX) at 15 min (T15) or normal saline (no-treatment) (TBI-NON). Similarly, uninjured swine received NVX-428 (SHAM-NVX) or normal saline (SHAM-NON). Animals were monitored and measurements were taken for physiological and neurological parameters before euthanasia at the six-hour mark (T360). Histopathological analysis was performed on paraffin embedded tissues. Physiological, biochemical and blood gas parameters were not different, with the exception of a significant but transient increase in mean pulmonary artery pressure observed in the TBI-experimental group immediately after drug administration. There were no initial differences in brain oxygenation at baseline, but over time oxygen decreased ~50% in both TBI groups. Histological brain injury scores were similar between TBI-NVX and TBI-NON, although a number of subcategories (spongiosis-ischemic/dead neurons-hemorrhage-edema) in TBI-NVX had a tendency for lower scores. The cerebellum showed significantly lower spongiosis and ischemic/dead neuron injury scores and a lower number of Fluoro-Jade-B-positive cerebellar-Purkinje-cells after NVX-428 treatment compared to controls. NVX-428 may assist in mitigating secondary cellular brain damage.

Histopathological Evidence of Multiple Organ Damage After Simulated Aeromedical Evacuation in a Swine Acute Lung Injury Model.

INTRODUCTION: Rapid aeromedical evacuation (AE) is standard of care in current conflicts. However, not much is known about possible effects of hypobaric conditions. We investigated possible effects of hypobaria on organ damage in a swine model of acute lung injury. METHODS: Lung injury was induced in anesthetized swine via intravenous oleic acid infusion. After a stabilization phase, animals were subjected to a 4 hour simulated AE at 8000 feet (HYPO). Control animals were kept at normobaria. After euthanasia and necropsy, organ damage was assessed by combined scores for hemorrhage, inflammation, edema, necrosis, and microatelectasis. RESULTS: Hemodynamic, neurological, or hematologic measurements were similar prior to transport. Hemodynamic instability became apparent during the last 2 hours of transport in the HYPO group. Histological injury scores in the HYPO group were higher for all organs (lung, kidney, liver, pancreas, and adrenal glands) except the brain, with the largest difference in the lungs (P < 0.001). CONCLUSIONS: Swine with mild acute lung injury subjected to a 4 hour simulated AE showed more injury to most organs and, in particular, to the lungs compared with ground transport. This may exacerbate otherwise subclinical pathology and, eventually, manifest as abnormalities in gas exchange or possibly end-organ function.

N-acetylcysteine Amide Ameliorates Blast-Induced Changes in Blood-Brain Barrier Integrity in Rats.

Traumatic brain injury resulting from exposure to blast overpressure (BOP) is associated with neuropathology including impairment of the blood-brain barrier (BBB). This study examined the effects of repeated exposure to primary BOP and post-blast treatment with an antioxidant, N-acetylcysteine amide (NACA) on the integrity of BBB. Anesthetized rats were exposed to three 110 kPa BOPs separated by 0.5 h. BBB integrity was examined in vivo via a cranial window allowing imaging of pial microcirculation by intravital microscopy. Tetramethylrhodamine isothiocyanate Dextran (TRITC-Dextran, mw = 40 kDa or 150 kDa) was injected intravenously 2.5 h after the first BOP exposure and the leakage of TRITC-Dextran from pial microvessels into the brain parenchyma was assessed. The animals were randomized into 6 groups (n = 5/group): four groups received 40 kDa TRITC-Dextran (BOP-40, sham-40, BOP-40 NACA, and sham-40 NACA), and two groups received 150 kDa TRITC-Dextran (BOP-150 and sham-150). NACA treated groups were administered NACA 2 h after the first BOP exposure. The rate of TRITC-Dextran leakage was significantly higher in BOP-40 than in sham-40 group. NACA treatment significantly reduced TRITC-Dextran leakage in BOP-40 NACA group and sham-40 NACA group presented the least amount of leakage. The rate of leakage in BOP-150 and sham-150 groups was comparable to sham-40 NACA and thus these groups were not assessed for the effects of NACA. Collectively, these data suggest that BBB integrity is compromised following BOP exposure and that NACA treatment at a single dose may significantly protect against blast-induced BBB breakdown.

Exposure to Blast Overpressure Impairs Cerebral Microvascular Responses and Alters Vascular and Astrocytic Structure.

Exposure to blast overpressure may result in cerebrovascular impairment, including cerebral vasospasm. The mechanisms contributing to this vascular response are unclear. The aim of this study was to evaluate the relationship between blast and functional alterations of the cerebral microcirculation and to investigate potential underlying changes in vascular microstructure. Cerebrovascular responses were assessed in sham- and blast-exposed male rats at multiple time points from 2 h through 28 days after a single 130-kPa (18.9-psi) exposure. Pial microcirculation was assessed through a cranial window created in the parietal bone of anesthetized rats. Pial arteriolar reactivity was evaluated in vivo using hypercapnia, barium chloride, and serotonin. We found that exposure to blast leads to impairment of arteriolar reactivity >24 h after blast exposure, suggesting delayed injury mechanisms that are not simply attributed to direct mechanical deformation. Observed vascular impairment included a reduction in hypercapnia-induced vasodilation, increase in barium-induced constriction, and reversal of the serotonin effect from constriction to dilation. A reduction in vascular smooth muscle contractile proteins consistent with vascular wall proliferation was observed, as well as delayed reduction in nitric oxide synthase and increase in endothelin-1 B receptors, mainly in astrocytes. Collectively, the data show that exposure to blast results in delayed and prolonged alterations in cerebrovascular reactivity that are associated with changes in the microarchitecture of the vessel wall and astrocytes. These changes may contribute to long-term pathologies involving dysfunction of the neurovascular unit, including cerebral vasospasm.

Affective profiling for anxiety-like behavior in a rodent model of mTBI.

Mild traumatic brain injury is a common outcome of blast exposure, and current literature indicates high rates of comorbid posttraumatic stress disorder (PTSD) in military personnel. Blast-exposed rats display PTSD-like behavior, suggesting relationships may exist between PTSD and blast exposure. Other studies demonstrate the roles of stathmin and corticosterone associated with fear- and anxiety-like behaviors in rodent models. Furthermore, studies have observed ranges of responses to both physical and psychological trauma in animal populations (Elder 2012, Ritov 2016). This study exposed rodents to repeated blast overpressure (BOP) and analyzed behavioral responses and molecular variables at 3 weeks and 6 months after exposure. We applied a modified version of a previously reported behavioral profiling approach that separates "affected" and "unaffected" rats based on the presence of anxiety-like behaviors (Ritov, 2016). We report that "affected" 3 week animals showed higher plasma corticosterone and amygdalar stathmin levels, while "affected" 6 month animals had lower prefrontal cortex stathmin. Higher corticosterone also paralleled anxiety behavior in "affected" 3 week animals, which was not observed in 6 month animals, indicating possible negative feedback loop mechanisms. Elevated levels of amygdalar stathmin correlated with anxiety behaviors in "affected" 3 week and 6 month animals, indicating sustained molecular changes. We conclude that this unique analysis may provide more information about response to blast. This type of analysis should also be considered when treating clinical populations, since individual differences may affect behavioral and long-term outcomes. Future studies should elucidate relationships of stress and fear responses in the context of BOP.

Blood transfusions, blood storage, and correlation with elevated pulmonary arterial pressures.

BACKGROUND: The aim of this study was to determine if transfusion with RBCs is associated with a rise in mean pulmonary artery pressure (MPAP) and whether such a rise is influenced by the duration of RBC storage. STUDY DESIGN AND METHODS: A retrospective chart review of intensive care unit patients with pulmonary artery catheters was conducted at two military medical centers. RESULTS: RBC transfusion is associated with a sustained (≥4 hours) statistically significant 2- to 3-mm Hg rise in MPAP relative to both pretransfusion levels (p < 0.05) and compared to asanguinous fluid infusions (p < 0.05). The magnitude of the rise (all infusions, RBCs, and asanguinous) correlates positively with in-hospital mortality (p < 0.01) and hospital length of stay (p < 0.01). The duration of RBC storage was not statistically correlated with the magnitude of rise in the population studied. Mean infusion volume was greater for RBC (vs. asanguinous) infusions, but volume adjustment of MPAP values did not alter the pattern or statistical significance of the results. CONCLUSIONS: Analysis of retrospectively collected data suggests that transfusion of RBC-containing fluids results in a sustained elevation of MPAP. In the patient population studied, the duration of RBC storage did not correlate with the magnitude of MPAP rise. Future prospective studies of transfusion effects should consider including assessment of MPAP and subpopulation analyses.

Low-level blast exposure disrupts gliovascular and neurovascular connections and induces a chronic vascular pathology in rat brain.

Much concern exists over the role of blast-induced traumatic brain injury (TBI) in the chronic cognitive and mental health problems that develop in veterans and active duty military personnel. The brain vasculature is particularly sensitive to blast injury. The aim of this study was to characterize the evolving molecular and histologic alterations in the neurovascular unit induced by three repetitive low-energy blast exposures (3 × 74.5 kPa) in a rat model mimicking human mild TBI or subclinical blast exposure. High-resolution two-dimensional differential gel electrophoresis (2D-DIGE) and matrix-assisted laser desorption/ionization (MALDI) mass spectrometry of purified brain vascular fractions from blast-exposed animals 6 weeks post-exposure showed decreased levels of vascular-associated glial fibrillary acidic protein (GFAP) and several neuronal intermediate filament proteins (α-internexin and the low, middle, and high molecular weight neurofilament subunits). Loss of these proteins suggested that blast exposure disrupts gliovascular and neurovascular interactions. Electron microscopy confirmed blast-induced effects on perivascular astrocytes including swelling and degeneration of astrocytic endfeet in the brain cortical vasculature. Because the astrocyte is a major sensor of neuronal activity and regulator of cerebral blood flow, structural disruption of gliovascular integrity within the neurovascular unit should impair cerebral autoregulation. Disrupted neurovascular connections to pial and parenchymal blood vessels might also affect brain circulation. Blast exposures also induced structural and functional alterations in the arterial smooth muscle layer. Interestingly, by 8 months after blast exposure, GFAP and neuronal intermediate filament expression had recovered to control levels in isolated brain vascular fractions. However, despite this recovery, a widespread vascular pathology was still apparent at 10 months after blast exposure histologically and on micro-computed tomography scanning. Thus, low-level blast exposure disrupts gliovascular and neurovascular connections while inducing a chronic vascular pathology.

Corrigendum: Repeated Low Intensity Blast Exposure Is Associated With Damaged Endothelial Glycocalyx and Downstream Behavioral Deficits.

[This corrects the article DOI: 10.3389/fnbeh.2017.00104.].

Hypobaria during long-range flight resulted in significantly increased histopathological evidence of lung and brain damage in a swine model.

BACKGROUND: Aeromedical evacuation to definitive care is standard in current military conflicts. However, there is minimal knowledge on the effects of hypobaria (HYPO) on either the flight crew or patients. The effects of HYPO were investigated using healthy swine. METHODS: Anesthetized Yorkshire swine underwent a simulated 4 h "transport" to an altitude of 2,441 m (8,000 feet.; HYPO, N = 6) or at normobaric conditions (NORMO, N = 6). Physiologic and biochemical data were collected. Organ damage was assessed for hemorrhage, inflammation, edema, necrosis, and for lungs only, microatelectasis. RESULTS: All parameters were similar prior to and after "transport" with no significant effects of HYPO on hemodynamic, neurologic, or oxygen transport parameters, nor on blood gas, chemistry, or complete blood count data. However, the overall Lung Injury Score was significantly worse in the HYPO than the NORMO group (10.78 ± 1.22 vs. 2.31 ± 0.71, respectively) with more edema/fibrin/hemorrhage in the subpleural, interlobular and alveolar space, more congestion in alveolar septa, and evidence of microatelectasis (vs. no microatelectasis in the NORMO group). There was also increased severity of pulmonary neutrophilic (1.69 ± 0.20 vs. 0.19 ± 0.13) and histiocytic inflammation (1.83 ± 0.23 vs. 0.47 ± 0.17) for HYPO versus NORMO, respectively. On the other hand, there was increased renal inflammation in NORMO compared with HYPO (1.00 ± 0.13 vs. 0.33 ± 0.17, respectively). There were no histopathological differences in brain (whole or individual regions), liver, pancreas, or adrenals. CONCLUSION: Hypobaria, itself, may have an adverse effect on the respiratory system, even in healthy individuals, and this may be superimposed on combat casualties where there may be preexisting lung injury. The additional effects of anesthesia and controlled ventilation on these results are unknown, and further studies are indicated using awake models to better characterize the mechanisms for this pathology and the factors that influence its severity.

Blast-induced "PTSD": Evidence from an animal model.

A striking observation among veterans returning from the recent conflicts in Iraq and Afghanistan has been the co-occurrence of blast-related mild traumatic brain injury (mTBI) and post-traumatic stress disorder (PTSD). PTSD and mTBI might coexist due to additive effects of independent psychological and physical traumas experienced in a war zone. Alternatively blast injury might induce PTSD-related traits or damage brain structures that mediate responses to psychological stressors, increasing the likelihood that PTSD will develop following a subsequent psychological stressor. Rats exposed to repetitive low-level blasts consisting of three 74.5 kPa exposures delivered once daily for three consecutive days develop a variety of anxiety and PTSD-related behavioral traits that are present for at least 9 months after blast exposure. A single predator scent challenge delivered 8 months after the last blast exposure induces additional anxiety-related changes that are still present 45 days later. Because the blast injuries occur under general anesthesia, it appears that blast exposure in the absence of a psychological stressor can induce chronic PTSD-related traits. The reaction to a predator scent challenge delivered many months after blast exposure suggests that blast exposure in addition sensitizes the brain to react abnormally to subsequent psychological stressors. The development of PTSD-related behavioral traits in the absence of a psychological stressor suggests the existence of blast-induced "PTSD". Findings that PTSD-related behavioral traits can be reversed by BCI-838, a group II metabotropic glutamate receptor antagonist offers insight into pathogenesis and possible treatment options for blast-related brain injury. This article is part of the Special Issue entitled "Novel Treatments for Traumatic Brain Injury".

Effect of acute restraint stress in a polytrauma rat model.

INTRODUCTION: A stressful environment may contribute to poor outcomes after TBI. The current study evaluates the impact of acute stress in a polytrauma rat model. METHODS: Rats were stressed by a 45-minute immobilization period before instrumentation under ketamine (t1). Polytrauma was produced by blast overpressure and controlled hemorrhage (t2). Rats were euthanized immediately after a 3 h simulated Medevac-transport time (t3) or after 72 h post-trauma (t4). Corticosterone, ACTH, and ACTH receptor gene expression were measured at these time points. Physiological parameters were monitored throughout the study. RESULTS: HR was higher in stressed compared to unstressed animals at t1. Corticosterone and ACTH levels were similar for all conditions at t1 and t2; ACTH and corticosterone became elevated in all groups at t3 and at t4, respectively. The ACTH receptor gene expression trended towards higher values at t4 for the stressed animals whether being injured or not. Survival after injury was 83% in both unstressed and stressed animals. CONCLUSION: Overall, corticosterone was not significantly affected following acute stress in ketamine-anesthetized rats. Early mortality was primarily due to polytrauma and change in the animal's biochemical parameters appeared at t4 post trauma. The findings indicate that ketamine-anesthesia and/or surgery may have overshadowed the effect of the initial stress.

PTSD-Related Behavioral Traits in a Rat Model of Blast-Induced mTBI Are Reversed by the mGluR2/3 Receptor Antagonist BCI-838.

Battlefield blast exposure related to improvised explosive devices (IEDs) has become the most common cause of traumatic brain injury (TBI) in the recent conflicts in Iraq and Afghanistan. Mental health problems are common after TBI. A striking feature in the most recent veterans has been the frequency with which mild TBI (mTBI) and posttraumatic stress disorder (PTSD) have appeared together, in contrast to the classical situations in which the presence of mTBI has excluded the diagnosis of PTSD. However, treatment of PTSD-related symptoms that follow blast injury has become a significant problem. BCI-838 (MGS0210) is a Group II metabotropic glutamate receptor (mGluR2/3) antagonist prodrug, and its active metabolite BCI-632 (MGS0039) has proneurogenic, procognitive, and antidepressant activities in animal models. In humans, BCI-838 is currently in clinical trials for refractory depression and suicidality. The aim of the current study was to determine whether BCI-838 could modify the anxiety response and reverse PTSD-related behaviors in rats exposed to a series of low-level blast exposures designed to mimic a human mTBI or subclinical blast exposure. BCI-838 treatment reversed PTSD-related behavioral traits improving anxiety and fear-related behaviors as well as long-term recognition memory. Treatment with BCI-838 also increased neurogenesis in the dentate gyrus (DG) of blast-exposed rats. The safety profile of BCI-838 together with the therapeutic activities reported here, make BCI-838 a promising drug for the treatment of former battlefield Warfighters suffering from PTSD-related symptoms following blast-induced mTBI.

Brain oxygenation with a non-vasoactive perfluorocarbon emulsion in a rat model of traumatic brain injury.

OBJECTIVE: The aim of this study was to assess, in two experiments, the safety and efficacy of the PFC emulsion Oxycyte as an oxygen therapeutic for TBI to test the hypothesis that early administration of this oxygen-carrying fluid post-TBI would improve brain tissue oxygenation (Pbt O2 ). METHODS: The first experiment assessed the effects of Oxycyte on cerebral vasoactivity in healthy, uninjured rats using intravital microscopy. The second experiment investigated the effect of Oxycyte on cerebral Pbt O2 using the PQM in TBI model. Animals in the Oxycyte group received a single injection of Oxycyte (6 mL/kg) shortly after TBI, while NON animals received no treatment. RESULTS: Oxycyte did not cause vasoconstriction in small- (<50 µm) or medium- (50-100 µm) sized pial arterioles nor did it cause a significant change in blood pressure. Treatment with Oxycyte while breathing 100% O2 did not improve Pbt O2 . However, in rats ventilated with ~40% O2 , Pbt O2 improved to near pre-TBI values within 105 minutes after Oxycyte injection. CONCLUSIONS: Although Oxycyte did not cause cerebral vasoconstriction, its use at the dose tested while breathing 100% O2 did not improve Pbt O2 following TBI. However, Oxycyte treatment while breathing a lower enriched oxygen concentration may improve Pbt O2 after TBI.

Lack of chronic neuroinflammation in the absence of focal hemorrhage in a rat model of low-energy blast-induced TBI.

Blast-related traumatic brain injury (TBI) has been a common cause of injury in the recent conflicts in Iraq and Afghanistan. Blast waves can damage blood vessels, neurons, and glial cells within the brain. Acutely, depending on the blast energy, blast wave duration, and number of exposures, blast waves disrupt the blood-brain barrier, triggering microglial activation and neuroinflammation. Recently, there has been much interest in the role that ongoing neuroinflammation may play in the chronic effects of TBI. Here, we investigated whether chronic neuroinflammation is present in a rat model of repetitive low-energy blast exposure. Six weeks after three 74.5-kPa blast exposures, and in the absence of hemorrhage, no significant alteration in the level of microglia activation was found. At 6 weeks after blast exposure, plasma levels of fractalkine, interleukin-1ß, lipopolysaccharide-inducible CXC chemokine, macrophage inflammatory protein 1α, and vascular endothelial growth factor were decreased. However, no differences in cytokine levels were detected between blast-exposed and control rats at 40 weeks. In brain, isolated changes were seen in levels of selected cytokines at 6 weeks following blast exposure, but none of these changes was found in both hemispheres or at 40 weeks after blast exposure. Notably, one animal with a focal hemorrhagic tear showed chronic microglial activation around the lesion 16 weeks post-blast exposure. These findings suggest that focal hemorrhage can trigger chronic focal neuroinflammation following blast-induced TBI, but that in the absence of hemorrhage, chronic neuroinflammation is not a general feature of low-level blast injury.

Storage of nitroglycerin (NTG) admixed with HBOC-201 for 30 days in polyolefin plastic bags: a pilot study.

Hemorrhaged animals have benefited from resuscitation with the hemoglobin-based oxygen carrier (HBOC-201). Co-infusion of nitric oxide (NO) via separate intravascular lines is effective in attenuating HBOC-induced elevation of blood pressure. We tested whether nitroglycerin (NTG) and HBOC-201 can be packaged together as a single drug for resuscitation. Since NTG binds easily to plastics such as polyvinylchloride, we assessed the stability of this combination in oxygen barrier double-layer ethylene-vinyl alcohol/polyolefin bags over a 30-day period. Outcome measures indicative of the stability of HBOC/NTG were reported as changes in levels of hemoglobin (Hb), methemoglobin (MetHb), NTG, and nitrite over time. Individual tightly sealed small aliquots of HBOC/NTG were prepared under nitrogen and analyzed in a timely fashion from 0 to 30 days using hematology instruments, HPLC, FPLC, and chemiluminescence. The level of NTG in the HBOC/NTG mixture was reduced significantly over time whereas it was stable in control mixtures of NTG/saline. The level of total Hb in the HBOC/NTG and HBOC/saline mixtures remained stable over time. MetHb formed and increased to 6% up to day 1 and then slowly decreased in the HBOC/NTG mixture whereas it remained unchanged in the HBOC/saline mixture. Nitrite was produced in the HBOC/NTG group upon mixing, was increased at day 1, and then became undetectable. The reaction between HBOC-201 and NTG occurring upon mixing and developing over time in polyolefin bags makes the long-term storage of this mixed combination inappropriate.

Repeated Low Intensity Blast Exposure Is Associated with Damaged Endothelial Glycocalyx and Downstream Behavioral Deficits.

Current clinical research into mild traumatic brain injury (mTBI) has focused on white matter changes as identified by advanced MRI based imaging techniques. However, perivascular tau accumulation in the brains of individuals diagnosed with mTBI suggests that the vasculature plays a key role in the pathology. This study used a rat model to examine whether the endothelial glycocalyx, a layer of the vasculature responsible for sensing luminal shear forces, is damaged by exposure to repeated low intensity blast, and whether this layer is associated with observed behavioral deficits. The blast exposure used consisted of 12, 40 kPa blast exposures conducted with a minimum of 24 h between blasts. We found that repeated blast exposure reduced glycocalyx length and density in various brain regions indicating damage. This blast exposure paradigm was associated with a mild performance decrement in the Morris water maze (MWM) which assesses learning and memory. Administration of hyaluronidase, an enzyme that binds to and degrades hyaluronan (a major structural component of the glycocalyx) prior to blast exposure reduced the observed behavioral deficits and induced a thickening of the glycocalyx layer. Taken together these findings demonstrate that the endothelial glycocalyx degradation following repeated blast is associated with behavioral decrements which can be prevented by treatment with hyaluronidase.

Protective Effect of N-Acetylcysteine Amide on Blast-Induced Increase in Intracranial Pressure in Rats.

Blast-induced traumatic brain injury is associated with acute and possibly chronic elevation of intracranial pressure (ICP). The outcome after TBI is dependent on the progression of complex processes which are mediated by oxidative stress. So far, no effective pharmacological protection against TBI exists. In this study, rats were exposed to a single or repetitive blast overpressure (BOP) at moderate intensities of 72 or 110 kPa in a compressed air-driven shock tube. The degree and duration of the increase in ICP were proportional to the intensity and frequency of the blast exposure(s). In most cases, a single dose of antioxidant N-acetylcysteine amide (NACA) (500 mg/kg) administered intravenously 2 h after exposure to BOP significantly attenuated blast-induced increase in ICP. A single dose of NACA was not effective in improving the outcome in the group of animals that were subjected to repetitive blast exposures at 110 kPa on the same day. In this group, two treatments with NACA at 2 and 4 h post-BOP exposure resulted in significant attenuation of elevated ICP. Treatment with NACA prior to BOP exposure completely prevented the elevation of ICP. The findings indicate that oxidative stress plays an important role in blast-induced elevated ICP as treatment with NACA-ameliorated ICP increase, which is frequently related to poor functional recovery after TBI.

Protection against Blast-Induced Traumatic Brain Injury by Increase in Brain Volume.

Blast-induced traumatic brain injury (bTBI) is a leading cause of injuries in recent military conflicts and it is responsible for an increased number of civilian casualties by terrorist attacks. bTBI includes a variety of neuropathological changes depending on the intensity of blast overpressure (BOP) such as brain edema, neuronal degeneration, diffuse axonal damage, and vascular dysfunction with neurological manifestations of psychological and cognitive abnormalities. Internal jugular vein (IJV) compression is known to reduce intracranial compliance by causing an increase in brain volume and was shown to reduce brain damage during closed impact-induced TBI. We investigated whether IJV compression can attenuate signs of TBI in rats after exposure to BOP. Animals were exposed to three 110 ± 5 kPa BOPs separated by 30 min intervals. Exposure to BOP resulted in a significant decrease of neuronal nuclei (NeuN) together with upregulation of aquaporin-4 (AQP-4), 3-nitrotyrosine (3-NT), and endothelin 1 receptor A (ETRA) expression in frontal cortex and hippocampus one day following exposures. IJV compression attenuated this BOP-induced increase in 3-NT in cortex and ameliorated the upregulation of AQP-4 in hippocampus. These results suggest that elevated intracranial pressure and intracerebral volume have neuroprotective potential in blast-induced TBI.

Determination of methemoglobin and hemoglobin levels in small volume samples.

Background/aims Hemoglobin-based oxygen carriers (HBOCs) have been previously studied as resuscitation fluids. Due to HBOCs specific molecular conformation, hemoglobin (Hb) and methemoglobin (MetHb) determination is not always possible with automated apparatus. A practical technique was designed that allows simultaneous reading of MetHb and Hb in small volume samples. Methods A spectrophotometric method for measuring low levels of MetHb and Hb from limited volume samples was developed using a 96-well-plate by downsizing the Evelyn-Malloy and Drabkin methods. Either blood or buffer samples were spiked with one of five HBOCs (HBOC-201, M101, MP4CO-NP, Sanguinate and Oxyvita C). After treatment with cyanides, the samples were read at 540, 630, and 680 nm, and Hb and MetHb results were compared to certificate-of-analysis. Results Hb levels ranging from 0.2 to 2.8 g/dl were detected accurately with the 96-well-plate method with HBOC-201. Similarly, this method accurately measured Hb from either plasma or buffer samples containing any of the HBOCs. The MetHb plasma samples with HBOC-201 were also in agreement with ABL results (R = 0.99719). MetHb from all HBOCs in buffer measured with this method was comparable to reference but the accuracy was compromised for HBOCs in blood. Conclusions A useful 96-well-plate method of measuring HBOCs' Hb was designed for small-volume plasma samples. It was accurate for measuring MetHb from samples, that contained M101, MP4CO-NP, Sanguinate, and Oxyvita C diluted in buffer. This well-plate method allows reading of batch samples, multiple replicates, and using small volumes to accommodate limited animal blood collection which would not be otherwise detected by automated instrumentation.

Cerebral Vasoactivity and Oxygenation with Oxygen Carrier M101 in Rats.

The severity of traumatic brain injury (TBI) may be reduced if oxygen can be rapidly provided to the injured brain. This study evaluated if the oxygen-carrier M101 causes vasoconstricton of pial vasculature in healthy rats (Experiment 1) and if M101 improves brain tissue oxygen (PbtO2) in rats with controlled cortical impact (CCI)-TBI (Experiment 2). M101 (12.5 mL/kg intravenous [IV] over 2 h) caused a mild (9 mm Hg) increase in the mean arterial blood pressure (MAP) of healthy rats without constriction of cerebral pial arterioles. M101 (12 mL/kg IV over 1 h) caused a modest (27 mm Hg) increase in MAP (peak, 123 ± 5 mm Hg [mean ± standard error of the mean]) of CCI-TBI rats and restored PbtO2 to near pre-injury levels. In both M101 and untreated control (NON) groups, PbtO2 was ∼30 ± 2 mm Hg pre-injury and decreased (p ≤ 0.05) to ∼16 ± 2 mm Hg 15 min after CCI. In NON, PbtO2 remained ∼50% of baseline but M101 administration resulted in a sustained increase in PbtO2 (peak, 25 ± 5 mm Hg), which was not significantly different from pre-injury until the end of the study, when it decreased again below pre-injury (but was still higher than NON). Histopathology showed no differences between groups. In conclusion, M101 increased systemic blood pressures without concurrent cerebral pial vasoconstriction (in healthy rats) and restored PbtO2 to 86% of pre-injury for at least 80 min when given soon after CCI-TBI. M101 should be evaluated in a clinically-relevant large animal model for pre-hospital treatment of TBI.