Journal of Cerebral Blood Flow and Metabolism, 2003
The cellular and molecular pathways initiated by traumatic brain injury (TBI) may compromise the ... more The cellular and molecular pathways initiated by traumatic brain injury (TBI) may compromise the function and structural integrity of mitochondria, thereby contributing to cerebral metabolic dysfunction and cell death. The extent to which TBI affects regional mitochondrial populations with respect to structure, function, and swelling was assessed 3 hours and 24 hours after lateral fluid-percussion brain injury in the rat. Significantly less mitochondrial protein was isolated from the injured compared with uninjured parietotemporal cortex, whereas comparable yields were obtained from the hippocampus. After injury, cortical and hippocampal tissue ATP concentrations declined significantly to 60% and 40% of control, respectively, in the absence of respiratory deficits in isolated mitochondria. Mitochondria with ultrastructural morphologic damage comprised a significantly greater percent of the population isolated from injured than uninjured brain. As determined by photon correlation spectroscopy, the mean mitochondrial radius decreased significantly in injured cortical populations (361 +/- 40 nm at 24 hours) and increased significantly in injured hippocampal populations (442 +/- 36 at 3 hours) compared with uninjured populations (Ctx: 418 +/- 44; Hipp: 393 +/- 24). Calcium-induced deenergized swelling rates of isolated mitochondrial populations were significantly slower in injured compared with uninjured samples, suggesting that injury alters the kinetics of mitochondrial permeability transition (MPT) pore activation. Cyclosporin A (CsA)-insensitive swelling was reduced in the cortex, and CsA-sensitive and CsA-insensitive swelling both were reduced in the hippocampus, demonstrating that regulated MPT pores remain in mitochondria isolated from injured brain. A proposed mitochondrial population model synthesizes these data and suggests that cortical mitochondria may be depleted after TBI, with a physically smaller, MPT-regulated population remaining. Hippocampal mitochondria may sustain damage associated with ballooned membranes and reduced MPT pore calcium sensitivity. The heterogeneous mitochondrial response to TBI may underlie posttraumatic metabolic dysfunction and contribute to the pathophysiology of TBI.
Abstract: The activation of poly(ADP-ribose) polymerase, a DNA base excision repair enzyme, is in... more Abstract: The activation of poly(ADP-ribose) polymerase, a DNA base excision repair enzyme, is indicative of DNA damage. This enzyme also undergoes site-specific proteolysis during apoptosis. Because both DNA fragmentation and apoptosis are known to occur following experimental brain injury, we investigated the effect of lateral fluid percussion brain injury on poly(ADP-ribose) polymerase activity and cleavage. Male Sprague-Dawley rats (n = 52) were anesthetized, subjected to fluid percussion brain injury of moderate severity (2.5-2.8 atm), and killed at 30 min, 2 h, 6 h, 24 h, 3 days, or 7 days postinjury. Genomic DNA from injured cortex at 24 h, but not at 30 min, was both fragmented and able to stimulate exogenous poly(ADP-ribose) polymerase. Endogenous poly(ADP-ribose) polymerase activity, however, was enhanced in the injured cortex at 30 min but subsequently returned to baseline levels. Slight fragmentation of poly(ADP-ribose) polymerase was detected in the injured cortex in the first 3 days following injury, but significant cleavage was detected at 7 days postinjury. Taken together, these data suggest that poly(ADP-ribose) polymerase-mediated DNA repair is initiated in the acute posttraumatic period but that subsequent poly(ADP-ribose) polymerase activation does not occur, possibly owing to delayed apoptosis-associated proteolysis, which may impair the repair of damaged DNA.
Traumatic brain injury (TBI) increases susceptibility to Alzheimer's disease (AD), but it is not ... more Traumatic brain injury (TBI) increases susceptibility to Alzheimer's disease (AD), but it is not known if TBI affects the progression of AD. To address this question, we studied the neuropathological consequences of TBI in transgenic (TG) mice with a mutant human Aβ precursor protein (APP) mini-gene driven by a platelet-derived (PD) growth factor promoter resulting in overexpression of mutant APP (V717F), elevated brain Aβ levels, and AD-like amyloidosis. Since brain Aβ deposits first appear in 6-month-old TG (PDAPP) mice and accumulate with age, 2-year-old PDAPP and wild-type (WT) mice were subjected to controlled cortical impact (CCI) TBI or sham treatment. At 1, 9, and 16 weeks after TBI, neuron loss, gliosis, and atrophy were most prominent near the CCI site in PDAPP and WT mice. However, there also was a remarkable regression in the Aβ amyloid plaque burden in the hippocampus ipsilateral to TBI compared to the contralateral hippocampus of the PDAPP mice by 16 weeks postinjury. Thus, these data suggest that previously accumulated Aβ plaques resulting from progressive amyloidosis in the AD brain also may be reversible.
Lateral (parasagittal) fluid-percussion brain injury of mild (1.0–1.5 atm) and moderate (2.1–2.4 ... more Lateral (parasagittal) fluid-percussion brain injury of mild (1.0–1.5 atm) and moderate (2.1–2.4 atm) severity induced expression of mRNAs for the immediate early genes (IEGs) c-fos, c-jun and junB. At 5 min following mild brain injury, c-fos and junB mRNA were co-induced in the cortex ipsilateral to the impact site. Expression remained elevated up to 2 h after injury and returned to control levels by 6 h. Levels of c-fos mRNA increased in the cells of the hippocampal dentate gyrus as early as 5 min after mild brain injury and additionally in the areas CA1–3 by 30 min. By 2 h, no hippocampal c-fos mRNA was detectable. Induction of junB mRNA in the hippocampus was delayed, occurring at 30 min after injury, and remained elevated up to 2 h post injury. Increased levels of junB mRNA were also observed in the striatum ipsilateral to the injury. Increased expression of c-jun mRNA was restricted to the ipsilateral dentate gyrus and was observed at 5 min after injury and remained elevated up to 6 h. Although the temporal pattern of induction of individual IEGs after brain injury of moderate severity was similar to that observed after mild severity, moderate injury induced IEG mRNA in both injured and contralateral hemispheres. These data suggest that traumatic brain injury invokes a complex acute regional and cellular response which may involve the activation of multiple signal transduction pathways.
Little is known regarding the molecular (genomic) events associated with the pathophysiology of t... more Little is known regarding the molecular (genomic) events associated with the pathophysiology of traumatic brain injury (TBI). This review focusses on the experimental efforts to date elucidating the acute alterations in expression of immediate early genes (IEGs), heat shock proteins (HSPs) and cytokines following experimental brain injury. The immediate early genes, c-fos, c-jun and junB were observed to be bilaterally induced in the cortex and hippocampus as early as 5 min following lateral fluid-percussion (FP) brain injury in the rat. While levels of c-fos and junB mRNA returned to control levels by 2h, c-jun mRNA remained elevated up to 6h post-injury. Increased levels of mRNA for the inducible heat-shock protein (hsp72) were observed up to 12h following injury and were restricted to the cortex ipsilateral to the impact site. Mild induction of the glucose-regulated proteins (grp78 and grp94), which share sequence homology with hsp72, was apparent in the ipsilateral cortex. The cytokines IL-1β and TNFα were induced at 1h following FP brain injury and remained elevated up to 6h post-injury. These data, while indicative of the complex genomic response to TBI, are also suggestive of the trauma-induced activation of multiple signal transduction pathways.
In this investigation, the relationships between stretch and both morphological and electrophysio... more In this investigation, the relationships between stretch and both morphological and electrophysiological signs of axonal injury were examined in the guinea pig optic nerve stretch model. Additionally, the relationship between axonal morphology and electrophysiological impairment was assessed. Axonal injury was produced in vivo by elongating the guinea pig optic nerve between 0 and 8 mm (Ntotal = 70). Morphological damage was detected using neurofilament immunohistochemistry (SMI 32). Electrophysiological impairment was determined using changes in visual evoked potentials (VEPs) measured prior to injury, every 5 min for 40 min following injury, and at sacrifice (72 h). All nerves subjected to ocular displacements greater than 6 mm demonstrated axonal swellings and retraction bulbs, while nerves subjected to displacements below 4 mm did not show any signs of morphological injury. Planned comparisons of latency shifts of the N35 peak in the VEPs showed that ocular displacements greater than 5 mm produced electrophysiological impairment that was significantly different from sham animals. Logit analysis demonstrated that less stretch was required to elicit electrophysiological changes (5.5 mm) than morphological signs of damage (6.8 mm). Moreover, Student t tests indicated that the mean latency shift measured in animals exhibiting morphological injury was significantly greater than that calculated from animals lacking morphological injury (p < 0.01). These data show that distinct mechanical thresholds exist for both morphological and electrophysiological damage to the white matter. In a larger context, the distinct injury thresholds presented in the report will aid in the biomechanical assessment of animate models of head injury, as well as assist in extending these findings to predict the conditions that cause white matter injury in humans.
The neuroprotective effect of magnesium chloride (MgCl2), a compound previously demonstrated to i... more The neuroprotective effect of magnesium chloride (MgCl2), a compound previously demonstrated to improve behavioral and neurochemical outcome in several models of experimental brain injury, was evaluated in the present study. Male Sprague-Dawley rats were anesthetized and subjected to lateral fluid-percussion brain injury of moderate severity (2.5-2.8 atm). A cannula was implanted in the left femoral vein and at 1 h following injury, animals randomly received a 15 min i.v. infusion of either MgCl2 (125 micromol/rat) or saline. A second group of animals received anesthesia, surgery, and either MgCl2 or vehicle to serve as uninjured (sham) controls. Two weeks following brain injury, animals were sacrificed, brains removed, and coronal sections were taken for quantitative analysis of cortical lesion volume and hippocampal CA3 cell counts. Traumatic brain injury resulted in a lesion in the ipsilateral cortex and loss of pyramidal neurons in the CA3 region of the hippocampus in vehicle-treated animals (p < 0.01 vs. uninjured animals). Administration of MgCl2 significantly reduced the injury-induced damage in the cortex (p < 0.01) but did not alter posttraumatic cell loss in the CA3 region of the ipsilateral hippocampus. The present study demonstrates that, in addition to its beneficial effects on behavioral outcome, MgCl2 treatment attenuates cortical histological damage when administered following traumatic brain injury.
Mild, traumatic repetitive head injury (RHI) leads to neurobehavioral impairment and is associate... more Mild, traumatic repetitive head injury (RHI) leads to neurobehavioral impairment and is associated with the early onset of neurodegenerative disease. The authors developed an animal model to investigate the behavioral and pathological changes associated with RHI. Adult male C57BL/6 mice were subjected to a single injury (43 mice), repetitive injury (two injuries 24 hours apart; 49 mice), or no impact (36 mice). Cognitive function was assessed using the Morris water maze test, and neurological motor function was evaluated using a battery of neuroscore, rotarod, and rotating pole tests. The animals were also evaluated for cardiovascular changes, blood-brain barrier (BBB) breakdown, traumatic axonal injury, and neurodegenerative and histopathological changes between 1 day and 56 days after brain trauma. No cognitive dysfunction was detected in any group. The single-impact group showed mild impairment according to the neuroscore test at only 3 days postinjury, whereas RHI caused pronounced deficits at 3 days and 7 days following the second injury. Moreover, RHI led to functional impairment during the rotarod and rotating pole tests that was not observed in any animal after a single impact. Small areas of cortical BBB breakdown and axonal injury. observed after a single brain injury, were profoundly exacerbated after RHI. Immunohistochemical staining for microtubule-associated protein-2 revealed marked regional loss of immunoreactivity only in animals subjected to RHI. No deposits of beta-amyloid or tau were observed in any brain-injured animal. On the basis of their results, the authors suggest that the brain has an increased vulnerability to a second traumatic insult for at least 24 hours following an initial episode of mild brain trauma.
A characteristic feature of severe diffuse axonal injury in man is radiological evidence of the “... more A characteristic feature of severe diffuse axonal injury in man is radiological evidence of the “shearing injury triad” represented by lesions, sometimes haemorrhagic, in the corpus callosum, deep white matter and the rostral brain stem. With the exception of studies carried out on the non-human primate, such lesions have not been replicated to date in the multiple and diverse rodent laboratory models of traumatic brain injury. The present report describes tissue tears in the white matter, particularly in the fimbria of Sprague-Dawley rats killed 12, 24, and 48 h and 7 days after lateral fluid percussion brain injury of moderate severity (2.1–2.4 atm). The lesions were most easily seen at 24 h when they appeared as foci of tissue rarefaction in which there were a few polymorphonuclear leucocytes. At the margins of these lesions, large amounts of accumulated amyloid precursor protein (APP) were found in axonal swellings and bulbs. By 1 week post-injury, there was macrophage infiltration with marked astrocytosis and early scar formation. This lesion is considered to be due to severe deformation of white matter and this is the first time that it has been identified reproducibly in a rodent model of head injury under controlled conditions.
Using the neural stem cell (NSC) clone C17.2, we evaluated the ability of transplanted murine NSC... more Using the neural stem cell (NSC) clone C17.2, we evaluated the ability of transplanted murine NSCs to attenuate cognitive and neurological motor deficits after traumatic brain injury. Nonimmunosuppressed C57BL/6 mice (n = 65) were anesthetized and subjected to lateral controlled cortical impact brain injury (n = 52) or surgery without injury (sham operation group, n = 13). At 3 days postinjury, all brain-injured animals were reanesthetized and randomized to receive stereotactic injection of NSCs or control cells (human embryonic kidney cells) into the cortex-hippocampus interface in either the ipsilateral or the contralateral hemisphere. One group of animals (n = 7) was killed at either 1 or 3 weeks postinjury to assess NSC survival in the acute posttraumatic period. Motor function was evaluated at weekly intervals for 12 weeks in the remaining animals, and cognitive (i.e., learning) deficits were assessed at 3 and 12 weeks after transplantation. Brain-injured animals that received either ipsilateral or contralateral NSC transplants showed significantly improved motor function in selected tests as compared with human embryonic kidney cell-transplanted animals during the 12-week observation period. Cognitive dysfunction was unaffected by transplantation at either 3 or 12 weeks postinjury. Histological analyses showed that NSCs survive for as long as 13 weeks after transplantation and were detected in the hippocampus and/or cortical areas adjacent to the injury cavity. At 13 weeks, the NSCs transplanted ipsilateral to the impact site expressed neuronal (NeuN) or astrocytic (glial fibrillary acidic protein) markers but not markers of oligodendrocytes (2'3'cyclic nucleotide 3'-phosphodiesterase), whereas the contralaterally transplanted NSCs expressed neuronal but not glial markers (double-labeled immunofluorescence and confocal microscopy). These data suggest that transplanted NSCs can survive in the traumatically injured brain, differentiate into neurons and/or glia, and attenuate motor dysfunction after traumatic brain injury.
We evaluated the efficacy of insulin-like growth factor-1 (IGF-1) in attenuating neurobehavioral ... more We evaluated the efficacy of insulin-like growth factor-1 (IGF-1) in attenuating neurobehavioral deficits following lateral fluid percussion (FP) brain injury. Male Sprague–Dawley rats (345–425 g,n = 88) were anesthetized and subjected to FP brain injury of moderate severity (2.4–2.9 atm). In Study 1, IGF-1 (1.0 mg/kg,n = 9) or vehicle (n = 14) was administered by subcutaneous injection at 15 min postinjury and similarly at 12-h intervals for 14 days. In animals evaluated daily for 14 days, IGF-1 treatment attenuated motor dysfunction over the 2-week period (P < 0.02). In Study 2, IGF-1 (4 mg/kg/day,n = 8 uninjured,n = 13 injured) or vehicle (n = 8 uninjured,n = 13 injured) was administered for 2 weeks via a subcutaneous pump implanted 15 min postinjury. IGF-1 administration was associated with increased body weight and mild, transient hypoglycemia which was more pronounced in brain-injured animals. At 2 weeks postinjury (P < 0.05), but not at 48 h or 1 week, brain-injured animals receiving IGF-1 showed improved neuromotor function compared with those receiving vehicle. IGF-1 administration also enhanced learning ability (P < 0.03) and memory retention (P < 0.01) in brain-injured animals at 2 weeks postinjury. Taken together, these data suggest that chronic, posttraumatic administration of the trophic factor IGF-1 may be efficacious in ameliorating neurobehavioral dysfunction associated with traumatic brain injury.
A growing body of evidence suggests that neurons undergo apoptotic cell death following traumatic... more A growing body of evidence suggests that neurons undergo apoptotic cell death following traumatic brain injury (TBI). Since the expression of several tumor suppressor and cell cycle genes have been implicated in neuronal apoptosis, the present study used in situ hybridization (ISH) histochemistry to evaluate the regional and temporal patterns of expression of the mRNAs for the tumor suppressor gene, p53, and the cell cycle gene, cyclin D1, following lateral fluid-percussion (FP) brain injury in the rat. Anesthetized adult male Sprague–Dawley rats (n=16) were subjected to lateral FP brain injury of moderate severity (2.4–2.7 atm), while sham controls (n=6) were surgically prepared but did not receive brain injury. Animals were killed by decapitation at 6 h (n=6 injured and 2 sham), 24 h (n=6 injured and 2 sham), or 3 days (n=4 injured and 2 sham), and their brains processed for ISH. Little to no expression of p53 mRNA was observed in sham brains. At 6 h post-injury, p53 mRNA was induced predominantly in cells that are vulnerable to TBI, such as those in the contused cortex, lateral and medial geniculate nuclei of the thalamus, and the CA3 and hilar neurons of the hippocampus. Increased p53 mRNA was also detected in hippocampal CA1 neurons, cells that are relatively resistant to FP brain injury. Levels of p53 mRNA returned to sham levels in all regions of the injured brain by 24 h. In contrast to p53, cyclin D1 mRNA was detectable in the brains of uninjured animals and was not altered by brain injury. These results suggest that the tumor suppressor gene p53, but not cyclin D1, is upregulated and may participate in molecular response to TBI.
Genetic strategies provide new ways to define the molecular cascades that regulate the responses ... more Genetic strategies provide new ways to define the molecular cascades that regulate the responses of the mammalian nervous system to injury. Genetic interventions also provide opportunities to manipulate and control key molecular steps in these cascades, so as to modify the outcome of CNS injury. Most current genetic strategies involve the use of mice, an animal that has not heretofore been used extensively for neurotrauma research. Therefore, one purpose of the present review is to consider how mice respond to neural trauma, focusing especially on recent information that reveals important differences between mice and rats, and between different inbred strains of mice. The second aim of this review is to provide a brief introduction to the opportunities, caveats, and potential pitfalls of studies that use genetically modified animals for neurotrauma research.
Journal of Cerebral Blood Flow and Metabolism, 2003
The cellular and molecular pathways initiated by traumatic brain injury (TBI) may compromise the ... more The cellular and molecular pathways initiated by traumatic brain injury (TBI) may compromise the function and structural integrity of mitochondria, thereby contributing to cerebral metabolic dysfunction and cell death. The extent to which TBI affects regional mitochondrial populations with respect to structure, function, and swelling was assessed 3 hours and 24 hours after lateral fluid-percussion brain injury in the rat. Significantly less mitochondrial protein was isolated from the injured compared with uninjured parietotemporal cortex, whereas comparable yields were obtained from the hippocampus. After injury, cortical and hippocampal tissue ATP concentrations declined significantly to 60% and 40% of control, respectively, in the absence of respiratory deficits in isolated mitochondria. Mitochondria with ultrastructural morphologic damage comprised a significantly greater percent of the population isolated from injured than uninjured brain. As determined by photon correlation spectroscopy, the mean mitochondrial radius decreased significantly in injured cortical populations (361 +/- 40 nm at 24 hours) and increased significantly in injured hippocampal populations (442 +/- 36 at 3 hours) compared with uninjured populations (Ctx: 418 +/- 44; Hipp: 393 +/- 24). Calcium-induced deenergized swelling rates of isolated mitochondrial populations were significantly slower in injured compared with uninjured samples, suggesting that injury alters the kinetics of mitochondrial permeability transition (MPT) pore activation. Cyclosporin A (CsA)-insensitive swelling was reduced in the cortex, and CsA-sensitive and CsA-insensitive swelling both were reduced in the hippocampus, demonstrating that regulated MPT pores remain in mitochondria isolated from injured brain. A proposed mitochondrial population model synthesizes these data and suggests that cortical mitochondria may be depleted after TBI, with a physically smaller, MPT-regulated population remaining. Hippocampal mitochondria may sustain damage associated with ballooned membranes and reduced MPT pore calcium sensitivity. The heterogeneous mitochondrial response to TBI may underlie posttraumatic metabolic dysfunction and contribute to the pathophysiology of TBI.
Abstract: The activation of poly(ADP-ribose) polymerase, a DNA base excision repair enzyme, is in... more Abstract: The activation of poly(ADP-ribose) polymerase, a DNA base excision repair enzyme, is indicative of DNA damage. This enzyme also undergoes site-specific proteolysis during apoptosis. Because both DNA fragmentation and apoptosis are known to occur following experimental brain injury, we investigated the effect of lateral fluid percussion brain injury on poly(ADP-ribose) polymerase activity and cleavage. Male Sprague-Dawley rats (n = 52) were anesthetized, subjected to fluid percussion brain injury of moderate severity (2.5-2.8 atm), and killed at 30 min, 2 h, 6 h, 24 h, 3 days, or 7 days postinjury. Genomic DNA from injured cortex at 24 h, but not at 30 min, was both fragmented and able to stimulate exogenous poly(ADP-ribose) polymerase. Endogenous poly(ADP-ribose) polymerase activity, however, was enhanced in the injured cortex at 30 min but subsequently returned to baseline levels. Slight fragmentation of poly(ADP-ribose) polymerase was detected in the injured cortex in the first 3 days following injury, but significant cleavage was detected at 7 days postinjury. Taken together, these data suggest that poly(ADP-ribose) polymerase-mediated DNA repair is initiated in the acute posttraumatic period but that subsequent poly(ADP-ribose) polymerase activation does not occur, possibly owing to delayed apoptosis-associated proteolysis, which may impair the repair of damaged DNA.
Traumatic brain injury (TBI) increases susceptibility to Alzheimer's disease (AD), but it is not ... more Traumatic brain injury (TBI) increases susceptibility to Alzheimer's disease (AD), but it is not known if TBI affects the progression of AD. To address this question, we studied the neuropathological consequences of TBI in transgenic (TG) mice with a mutant human Aβ precursor protein (APP) mini-gene driven by a platelet-derived (PD) growth factor promoter resulting in overexpression of mutant APP (V717F), elevated brain Aβ levels, and AD-like amyloidosis. Since brain Aβ deposits first appear in 6-month-old TG (PDAPP) mice and accumulate with age, 2-year-old PDAPP and wild-type (WT) mice were subjected to controlled cortical impact (CCI) TBI or sham treatment. At 1, 9, and 16 weeks after TBI, neuron loss, gliosis, and atrophy were most prominent near the CCI site in PDAPP and WT mice. However, there also was a remarkable regression in the Aβ amyloid plaque burden in the hippocampus ipsilateral to TBI compared to the contralateral hippocampus of the PDAPP mice by 16 weeks postinjury. Thus, these data suggest that previously accumulated Aβ plaques resulting from progressive amyloidosis in the AD brain also may be reversible.
Lateral (parasagittal) fluid-percussion brain injury of mild (1.0–1.5 atm) and moderate (2.1–2.4 ... more Lateral (parasagittal) fluid-percussion brain injury of mild (1.0–1.5 atm) and moderate (2.1–2.4 atm) severity induced expression of mRNAs for the immediate early genes (IEGs) c-fos, c-jun and junB. At 5 min following mild brain injury, c-fos and junB mRNA were co-induced in the cortex ipsilateral to the impact site. Expression remained elevated up to 2 h after injury and returned to control levels by 6 h. Levels of c-fos mRNA increased in the cells of the hippocampal dentate gyrus as early as 5 min after mild brain injury and additionally in the areas CA1–3 by 30 min. By 2 h, no hippocampal c-fos mRNA was detectable. Induction of junB mRNA in the hippocampus was delayed, occurring at 30 min after injury, and remained elevated up to 2 h post injury. Increased levels of junB mRNA were also observed in the striatum ipsilateral to the injury. Increased expression of c-jun mRNA was restricted to the ipsilateral dentate gyrus and was observed at 5 min after injury and remained elevated up to 6 h. Although the temporal pattern of induction of individual IEGs after brain injury of moderate severity was similar to that observed after mild severity, moderate injury induced IEG mRNA in both injured and contralateral hemispheres. These data suggest that traumatic brain injury invokes a complex acute regional and cellular response which may involve the activation of multiple signal transduction pathways.
Little is known regarding the molecular (genomic) events associated with the pathophysiology of t... more Little is known regarding the molecular (genomic) events associated with the pathophysiology of traumatic brain injury (TBI). This review focusses on the experimental efforts to date elucidating the acute alterations in expression of immediate early genes (IEGs), heat shock proteins (HSPs) and cytokines following experimental brain injury. The immediate early genes, c-fos, c-jun and junB were observed to be bilaterally induced in the cortex and hippocampus as early as 5 min following lateral fluid-percussion (FP) brain injury in the rat. While levels of c-fos and junB mRNA returned to control levels by 2h, c-jun mRNA remained elevated up to 6h post-injury. Increased levels of mRNA for the inducible heat-shock protein (hsp72) were observed up to 12h following injury and were restricted to the cortex ipsilateral to the impact site. Mild induction of the glucose-regulated proteins (grp78 and grp94), which share sequence homology with hsp72, was apparent in the ipsilateral cortex. The cytokines IL-1β and TNFα were induced at 1h following FP brain injury and remained elevated up to 6h post-injury. These data, while indicative of the complex genomic response to TBI, are also suggestive of the trauma-induced activation of multiple signal transduction pathways.
In this investigation, the relationships between stretch and both morphological and electrophysio... more In this investigation, the relationships between stretch and both morphological and electrophysiological signs of axonal injury were examined in the guinea pig optic nerve stretch model. Additionally, the relationship between axonal morphology and electrophysiological impairment was assessed. Axonal injury was produced in vivo by elongating the guinea pig optic nerve between 0 and 8 mm (Ntotal = 70). Morphological damage was detected using neurofilament immunohistochemistry (SMI 32). Electrophysiological impairment was determined using changes in visual evoked potentials (VEPs) measured prior to injury, every 5 min for 40 min following injury, and at sacrifice (72 h). All nerves subjected to ocular displacements greater than 6 mm demonstrated axonal swellings and retraction bulbs, while nerves subjected to displacements below 4 mm did not show any signs of morphological injury. Planned comparisons of latency shifts of the N35 peak in the VEPs showed that ocular displacements greater than 5 mm produced electrophysiological impairment that was significantly different from sham animals. Logit analysis demonstrated that less stretch was required to elicit electrophysiological changes (5.5 mm) than morphological signs of damage (6.8 mm). Moreover, Student t tests indicated that the mean latency shift measured in animals exhibiting morphological injury was significantly greater than that calculated from animals lacking morphological injury (p &lt; 0.01). These data show that distinct mechanical thresholds exist for both morphological and electrophysiological damage to the white matter. In a larger context, the distinct injury thresholds presented in the report will aid in the biomechanical assessment of animate models of head injury, as well as assist in extending these findings to predict the conditions that cause white matter injury in humans.
The neuroprotective effect of magnesium chloride (MgCl2), a compound previously demonstrated to i... more The neuroprotective effect of magnesium chloride (MgCl2), a compound previously demonstrated to improve behavioral and neurochemical outcome in several models of experimental brain injury, was evaluated in the present study. Male Sprague-Dawley rats were anesthetized and subjected to lateral fluid-percussion brain injury of moderate severity (2.5-2.8 atm). A cannula was implanted in the left femoral vein and at 1 h following injury, animals randomly received a 15 min i.v. infusion of either MgCl2 (125 micromol/rat) or saline. A second group of animals received anesthesia, surgery, and either MgCl2 or vehicle to serve as uninjured (sham) controls. Two weeks following brain injury, animals were sacrificed, brains removed, and coronal sections were taken for quantitative analysis of cortical lesion volume and hippocampal CA3 cell counts. Traumatic brain injury resulted in a lesion in the ipsilateral cortex and loss of pyramidal neurons in the CA3 region of the hippocampus in vehicle-treated animals (p &lt; 0.01 vs. uninjured animals). Administration of MgCl2 significantly reduced the injury-induced damage in the cortex (p &lt; 0.01) but did not alter posttraumatic cell loss in the CA3 region of the ipsilateral hippocampus. The present study demonstrates that, in addition to its beneficial effects on behavioral outcome, MgCl2 treatment attenuates cortical histological damage when administered following traumatic brain injury.
Mild, traumatic repetitive head injury (RHI) leads to neurobehavioral impairment and is associate... more Mild, traumatic repetitive head injury (RHI) leads to neurobehavioral impairment and is associated with the early onset of neurodegenerative disease. The authors developed an animal model to investigate the behavioral and pathological changes associated with RHI. Adult male C57BL/6 mice were subjected to a single injury (43 mice), repetitive injury (two injuries 24 hours apart; 49 mice), or no impact (36 mice). Cognitive function was assessed using the Morris water maze test, and neurological motor function was evaluated using a battery of neuroscore, rotarod, and rotating pole tests. The animals were also evaluated for cardiovascular changes, blood-brain barrier (BBB) breakdown, traumatic axonal injury, and neurodegenerative and histopathological changes between 1 day and 56 days after brain trauma. No cognitive dysfunction was detected in any group. The single-impact group showed mild impairment according to the neuroscore test at only 3 days postinjury, whereas RHI caused pronounced deficits at 3 days and 7 days following the second injury. Moreover, RHI led to functional impairment during the rotarod and rotating pole tests that was not observed in any animal after a single impact. Small areas of cortical BBB breakdown and axonal injury. observed after a single brain injury, were profoundly exacerbated after RHI. Immunohistochemical staining for microtubule-associated protein-2 revealed marked regional loss of immunoreactivity only in animals subjected to RHI. No deposits of beta-amyloid or tau were observed in any brain-injured animal. On the basis of their results, the authors suggest that the brain has an increased vulnerability to a second traumatic insult for at least 24 hours following an initial episode of mild brain trauma.
A characteristic feature of severe diffuse axonal injury in man is radiological evidence of the “... more A characteristic feature of severe diffuse axonal injury in man is radiological evidence of the “shearing injury triad” represented by lesions, sometimes haemorrhagic, in the corpus callosum, deep white matter and the rostral brain stem. With the exception of studies carried out on the non-human primate, such lesions have not been replicated to date in the multiple and diverse rodent laboratory models of traumatic brain injury. The present report describes tissue tears in the white matter, particularly in the fimbria of Sprague-Dawley rats killed 12, 24, and 48 h and 7 days after lateral fluid percussion brain injury of moderate severity (2.1–2.4 atm). The lesions were most easily seen at 24 h when they appeared as foci of tissue rarefaction in which there were a few polymorphonuclear leucocytes. At the margins of these lesions, large amounts of accumulated amyloid precursor protein (APP) were found in axonal swellings and bulbs. By 1 week post-injury, there was macrophage infiltration with marked astrocytosis and early scar formation. This lesion is considered to be due to severe deformation of white matter and this is the first time that it has been identified reproducibly in a rodent model of head injury under controlled conditions.
Using the neural stem cell (NSC) clone C17.2, we evaluated the ability of transplanted murine NSC... more Using the neural stem cell (NSC) clone C17.2, we evaluated the ability of transplanted murine NSCs to attenuate cognitive and neurological motor deficits after traumatic brain injury. Nonimmunosuppressed C57BL/6 mice (n = 65) were anesthetized and subjected to lateral controlled cortical impact brain injury (n = 52) or surgery without injury (sham operation group, n = 13). At 3 days postinjury, all brain-injured animals were reanesthetized and randomized to receive stereotactic injection of NSCs or control cells (human embryonic kidney cells) into the cortex-hippocampus interface in either the ipsilateral or the contralateral hemisphere. One group of animals (n = 7) was killed at either 1 or 3 weeks postinjury to assess NSC survival in the acute posttraumatic period. Motor function was evaluated at weekly intervals for 12 weeks in the remaining animals, and cognitive (i.e., learning) deficits were assessed at 3 and 12 weeks after transplantation. Brain-injured animals that received either ipsilateral or contralateral NSC transplants showed significantly improved motor function in selected tests as compared with human embryonic kidney cell-transplanted animals during the 12-week observation period. Cognitive dysfunction was unaffected by transplantation at either 3 or 12 weeks postinjury. Histological analyses showed that NSCs survive for as long as 13 weeks after transplantation and were detected in the hippocampus and/or cortical areas adjacent to the injury cavity. At 13 weeks, the NSCs transplanted ipsilateral to the impact site expressed neuronal (NeuN) or astrocytic (glial fibrillary acidic protein) markers but not markers of oligodendrocytes (2&amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;#39;3&amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;#39;cyclic nucleotide 3&amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;#39;-phosphodiesterase), whereas the contralaterally transplanted NSCs expressed neuronal but not glial markers (double-labeled immunofluorescence and confocal microscopy). These data suggest that transplanted NSCs can survive in the traumatically injured brain, differentiate into neurons and/or glia, and attenuate motor dysfunction after traumatic brain injury.
We evaluated the efficacy of insulin-like growth factor-1 (IGF-1) in attenuating neurobehavioral ... more We evaluated the efficacy of insulin-like growth factor-1 (IGF-1) in attenuating neurobehavioral deficits following lateral fluid percussion (FP) brain injury. Male Sprague–Dawley rats (345–425 g,n = 88) were anesthetized and subjected to FP brain injury of moderate severity (2.4–2.9 atm). In Study 1, IGF-1 (1.0 mg/kg,n = 9) or vehicle (n = 14) was administered by subcutaneous injection at 15 min postinjury and similarly at 12-h intervals for 14 days. In animals evaluated daily for 14 days, IGF-1 treatment attenuated motor dysfunction over the 2-week period (P < 0.02). In Study 2, IGF-1 (4 mg/kg/day,n = 8 uninjured,n = 13 injured) or vehicle (n = 8 uninjured,n = 13 injured) was administered for 2 weeks via a subcutaneous pump implanted 15 min postinjury. IGF-1 administration was associated with increased body weight and mild, transient hypoglycemia which was more pronounced in brain-injured animals. At 2 weeks postinjury (P < 0.05), but not at 48 h or 1 week, brain-injured animals receiving IGF-1 showed improved neuromotor function compared with those receiving vehicle. IGF-1 administration also enhanced learning ability (P < 0.03) and memory retention (P < 0.01) in brain-injured animals at 2 weeks postinjury. Taken together, these data suggest that chronic, posttraumatic administration of the trophic factor IGF-1 may be efficacious in ameliorating neurobehavioral dysfunction associated with traumatic brain injury.
A growing body of evidence suggests that neurons undergo apoptotic cell death following traumatic... more A growing body of evidence suggests that neurons undergo apoptotic cell death following traumatic brain injury (TBI). Since the expression of several tumor suppressor and cell cycle genes have been implicated in neuronal apoptosis, the present study used in situ hybridization (ISH) histochemistry to evaluate the regional and temporal patterns of expression of the mRNAs for the tumor suppressor gene, p53, and the cell cycle gene, cyclin D1, following lateral fluid-percussion (FP) brain injury in the rat. Anesthetized adult male Sprague–Dawley rats (n=16) were subjected to lateral FP brain injury of moderate severity (2.4–2.7 atm), while sham controls (n=6) were surgically prepared but did not receive brain injury. Animals were killed by decapitation at 6 h (n=6 injured and 2 sham), 24 h (n=6 injured and 2 sham), or 3 days (n=4 injured and 2 sham), and their brains processed for ISH. Little to no expression of p53 mRNA was observed in sham brains. At 6 h post-injury, p53 mRNA was induced predominantly in cells that are vulnerable to TBI, such as those in the contused cortex, lateral and medial geniculate nuclei of the thalamus, and the CA3 and hilar neurons of the hippocampus. Increased p53 mRNA was also detected in hippocampal CA1 neurons, cells that are relatively resistant to FP brain injury. Levels of p53 mRNA returned to sham levels in all regions of the injured brain by 24 h. In contrast to p53, cyclin D1 mRNA was detectable in the brains of uninjured animals and was not altered by brain injury. These results suggest that the tumor suppressor gene p53, but not cyclin D1, is upregulated and may participate in molecular response to TBI.
Genetic strategies provide new ways to define the molecular cascades that regulate the responses ... more Genetic strategies provide new ways to define the molecular cascades that regulate the responses of the mammalian nervous system to injury. Genetic interventions also provide opportunities to manipulate and control key molecular steps in these cascades, so as to modify the outcome of CNS injury. Most current genetic strategies involve the use of mice, an animal that has not heretofore been used extensively for neurotrauma research. Therefore, one purpose of the present review is to consider how mice respond to neural trauma, focusing especially on recent information that reveals important differences between mice and rats, and between different inbred strains of mice. The second aim of this review is to provide a brief introduction to the opportunities, caveats, and potential pitfalls of studies that use genetically modified animals for neurotrauma research.
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Papers by Ramesh Raghupathi