Shaun W. Carlson

Rate of Neurodegeneration in the Mouse Controlled Cortical Impact Model Is Influenced by Impactor Tip Shape: Implications for Mechanistic and Therapeutic Studies

Controlled cortical impact (CCI), one of the most common models of traumatic brain injury, is being increasingly used with mice for exploration of cell injury mechanisms and pre-clinical evaluation of therapeutic strategies. Although CCI brain injury was originally effected using an impactor with a rounded tip, the majority of studies with mouse CCI use a flat or beveled tip. Recent finite element modeling analyses demonstrate that tip geometry is a significant determinant of predicted cortical tissue strains in rat CCI, and that cell death is proportional to predicted tissue strains. In the current study, a three-dimensional finite element model of a C57BL/6J mouse brain predicted higher maximum principal strains during a simulated 1.0-mm, 3.5-m/s CCI injury with a flat tip when compared to a rounded tip. Consistent with this prediction, experimental CCI with a flat-tip impactor resulted in greater acute cortical hemorrhage and neuron loss in adult male C57BL/6J mice. The amount of neocortical tissue damage was equivalent for the two tip geometries at 9 days following injury, but the rate of neocortical neurodegeneration was markedly slower following CCI with a rounded-tip impactor, with damage reaching a plateau after 24?h as opposed to after 4?h for the flat tip. The flat-tip impactor was associated in general with more regional hippocampal neurodegeneration, especially at early time points such as 4?h. Impactor tip geometry did not have a notable effect on blood?brain barrier breakdown, traumatic axonal injury, or motor and cognitive dysfunction. Execution of CCI injury with a rounded-tip impactor is posited to provide a substantially enhanced temporal window for the study of cellular injury mechanisms and therapeutic intervention while maintaining critical aspects of the pathophysiological response to contusion brain injury.

Conditional overexpression of insulin-like growth factor-1 enhances hippocampal neurogenesis and restores immature neuron dendritic processes after traumatic brain injury.

Traumatic brain injury (TBI) is associated with neuronal damage or neuronal death in the hippocampus, a region critical for cognitive function. Immature neurons within the hippocampal neurogenic niche are particularly susceptible to TBI. Therapeutic strategies that protect immature hippocampal neurons or enhance posttraumatic neurogenesis may be advantageous for promoting functional recovery after TBI. Insulin-like growth factor-1 (IGF-1) promotes neurogenesis in the adult brain, but its effects on neurogenesis after TBI are unknown. We used an astrocyte-specific conditional IGF-1-overexpressing mouse model to supplement IGF-1 in regions of neuronal damage and reactive astrocytosis after controlled cortical impact injury. Although early loss of immature neurons was not significantly attenuated, overexpression of IGF-1 resulted in a marked increase in immature neuron density in the subgranular zone at 10 days after injury. This delayed increase seemed to be driven by enhanced neuron differentiation rather than by increased cellular proliferation. In wild-type mice, dendrites of immature neurons exhibited significant decreases in total length and number of bifurcations at 10 days after injury versus neurons in sham-injured mice. In contrast, the morphology of immature neuron dendrites in brain-injured IGF-1-overexpressing mice was equivalent to that in sham controls. These data provide compelling evidence that IGF-1 promotes neurogenesis after TBI.

Emerging Therapies in Traumatic Brain Injury

Despite decades of basic and clinical research, treatments to improve outcomes after traumatic brain injury (TBI) are limited. However, based on the recent recognition of the prevalence of mild TBI, and its potential link to neurodegenerative disease, many new and exciting secondary injury mechanisms have been identified and several new therapies are being evaluated targeting both classic and novel paradigms. This includes a robust increase in both preclinical and clinical investigations. Using a mechanism-based approach the authors define the targets and emerging therapies for TBI. They address putative new therapies for TBI across both the spectrum of injury severity and the continuum of care, from the field to rehabilitation. They discussTBI therapy using 11 categories, namely, (1) excitotoxicity and neuronal death, (2) brain edema, (3) mitochondria and oxidative stress, (4) axonal injury, (5) inflammation, (6) ischemia and cerebral blood flow dysregulation, (7) cognitive enhancement, (8) augmentation of endogenous neuroprotection, (9) cellular therapies, (10) combination therapy, and (11) TBI resuscitation. The current golden age of TBI research represents a special opportunity for the development of breakthroughs in the field.

Astrocyte-Specific Overexpression of Insulin-Like Growth Factor-1 Protects Hippocampal Neurons and Reduces Behavioral Deficits following Traumatic Brain Injury in Mice

Traumatic brain injury (TBI) survivors often suffer from long-lasting cognitive impairment that stems from hippocampal injury. Systemic administration of insulin-like growth factor-1 (IGF-1), a polypeptide growth factor known to play vital roles in neuronal survival, has been shown to attenuate posttraumatic cognitive and motor dysfunction. However, its neuroprotective effects in TBI have not been examined. To this end, moderate or severe contusion brain injury was induced in mice with conditional (postnatal) overexpression of IGF-1 using the controlled cortical impact (CCI) injury model. CCI brain injury produces robust reactive astrocytosis in regions of neuronal damage such as the hippocampus. We exploited this regional astrocytosis by linking expression of hIGF-1 to the astrocyte-specific glial fibrillary acidic protein (GFAP) promoter, effectively targeting IGF-1 delivery to vulnerable neurons. Following brain injury, IGF-1Tg mice exhibited a progressive increase in hippocampal IGF-1 levels which was coupled with enhanced hippocampal reactive astrocytosis and significantly greater GFAP levels relative to WT mice. IGF-1 overexpression stimulated Akt phosphorylation and reduced acute (1 and 3d) hippocampal neurodegeneration, culminating in greater neuron survival at 10d after CCI injury. Hippocampal neuroprotection achieved by IGF-1 overexpression was accompanied by improved motor and cognitive function in brain-injured mice. These data provide strong support for the therapeutic efficacy of increased brain levels of IGF-1 in the setting of TBI.

Rit GTPase Signaling Promotes Immature Hippocampal Neuronal Survival

The molecular mechanisms governing the spontaneous recovery seen following brain injury remain elusive, but recent studies indicate that injury-induced stimulation of hippocampal neurogenesis contributes to the repair process. The therapeutic potential of endogenous neurogenesis is tempered by the demonstration that traumatic brain injury (TBI) results in the selective death of adult-born immature neurons, compromising the cell population poised to compensate for trauma-induced neuronal loss. Here, we identify the Ras-related GTPase, Rit, as a critical player in the survival of immature hippocampal neurons following brain injury. While Rit knock-out (Rit−/−) did not alter hippocampal development, hippocampal neural cultures derived from Rit−/− mice display increased cell death and blunted MAPK cascade activation in response to oxidative stress, without affecting BDNF-dependent signaling. When compared with wild-type hippocampal cultures, Rit loss rendered immature (Dcx+) neurons susceptible to oxidative damage, without altering the survival of neural progenitor (Nestin+) cells. Oxidative stress is a major contributor to neuronal cell death following brain injury. Consistent with the enhanced vulnerability of cultured Rit−/− immature neurons, Rit−/− mice exhibited a significantly greater loss of adult-born immature neurons within the dentate gyrus after TBI. In addition, post-TBI neuronal remodeling was blunted. Together, these data identify a new and unexpected role for Rit in injury-induced neurogenesis, functioning as a selective survival mechanism for immature hippocampal neurons within the subgranular zone of the dentate gyrus following TBI.

IGF-1 Mediated Neurogenesis Involves a Novel RIT1 /Akt/Sox2 Cascade

Insulin-like growth factor 1 (IGF-1) is known to have diverse effects on brain structure and function, including the promotion of stem cell proliferation and neurogenesis in the adult dentate gyrus. However, the intracellular pathways downstream of the IGF-1 receptor that contribute to these diverse physiological actions remain relatively uncharacterized. Here, we demonstrate that the Ras-related GTPase, RIT1, plays a critical role in IGF-1-dependent neurogenesis. Studies in hippocampal neuronal precursor cells (HNPCs) demonstrate that IGF-1 stimulates a RIT1-dependent increase in Sox2 levels, resulting in pro-neural gene expression and increased cellular proliferation. In this novel cascade, RIT1 stimulates Akt-dependent phosphorylation of Sox2 at T118, leading to its stabilization and transcriptional activation. When compared to wild-type HNPCs, RIT1−/− HNPCs show deficient IGF-1-dependent Akt signaling and neuronal differentiation, and accordingly, Sox2-dependent hippocampal neurogenesis is significantly blunted following IGF-1 infusion in knockout (RIT1−/−) mice. Consistent with a role for RIT1 function in the modulation of activity-dependent plasticity, exercise-mediated potentiation of hippocampal neurogenesis is also diminished in RIT1−/− mice. Taken together, these data identify the previously uncharacterized IGF1-RIT1-Akt-Sox2 signaling pathway as a key component of neurogenic niche sensing, contributing to the regulation of neural stem cell homeostasis.

Lithium increases hippocampal SNARE protein abundance after traumatic brain injury

ABSTRACT Rodent models of traumatic brain injury (TBI) reproduce secondary injury sequela and cognitive impairments observed in patients afflicted by a TBI. Impaired neurotransmission has been reported in the weeks following experimental TBI, and may be a contributor to behavioral dysfunction. The soluble N‐ethylmaleimide‐sensitive factor attachment protein receptor (SNARE) complex, the machinery facilitating vesicular docking and fusion, is a highly‐conserved mechanism important for neurotransmission. Following TBI, there is a reduction in both the formation of the SNARE complex and the abundance of multiple SNARE proteins, including the chaperone protein cysteine string protein &agr; (CSP&agr;). Treatment with lithium in naïve rats reportedly increases the expression of CSP&agr;. In the context of TBI, brain‐injured rats treated with lithium exhibit improved outcome in published reports, but the mechanisms underlying the improvement are poorly understood. The current study evaluated the effect of lithium administration on the abundance of SNARE proteins and SNARE complex formation, hemispheric tissue loss, and neurobehavioral performance following controlled cortical impact (CCI). Sprague Dawley rats were subjected to CCI or sham injury, and treated daily with lithium chloride or vehicle for up to 14 days. Administration of lithium after TBI modestly improved spatial memory at 14 days post‐injury. Semi‐quantitative immunoblot analysis of hippocampal lysates revealed that treatment with lithium attenuated reductions in key SNARE proteins and SNARE complex formation at multiple time points post‐injury. These findings highlight that treatment with lithium increased the abundance of synaptic proteins that facilitate neurotransmission and may contribute to improved cognitive function after TBI. HighlightsSNARE complex formation was reduced in the weeks after TBI.Lithium modestly improved cognitive performance after TBI.Lithium increased SNARE protein and SNARE complex abundance after TBI.

Central Infusion of IGF-1 Increases Hippocampal Neurogenesis and Improves Neurobehavioral Function following Traumatic Brain Injury

Traumatic brain injury (TBI) produces neuronal dysfunction and cellular loss that can culminate in lasting impairments in cognitive and motor abilities. Therapeutic agents that promote repair and replenish neurons post-TBI hold promise in improving recovery of function. Insulin-like growth factor-1 (IGF-1) is a neurotrophic factor capable of mediating neuroprotective and neuroplasticity mechanisms. Targeted overexpression of IGF-1 enhances the generation of hippocampal newborn neurons in brain-injured mice; however, the translational neurogenic potential of exogenously administered IGF-1 post-TBI remains unknown. In a mouse model of controlled cortical impact, continuous intracerebroventricular infusion of recombinant human IGF-1 (hIGF) for 7 days, beginning 15 min post-injury, resulted in a dose-dependent increase in the number of immature neurons in the hippocampus. Infusion of 10 μg/day of IGF-1 produced detectable levels of hIGF-1 in the cortex and hippocampus and a concomitant increase in protein kinase B activation in the hippocampus. Both motor function and cognition were improved over 7 days post-injury in IGF-1-treated cohorts. Vehicle-treated brain-injured mice showed reduced hippocampal immature neuron density relative to sham controls at 7 days post-injury. In contrast, the density of hippocampal immature neurons in brain-injured mice receiving acute onset IGF-1 infusion was significantly higher than in injured mice receiving vehicle and equivalent to that in sham-injured control mice. Importantly, the neurogenic effect of IGF-1 was maintained with as much as a 6-h delay in the initiation of infusion. These data suggest that central infusion of IGF-1 enhances the generation of immature neurons in the hippocampus, with a therapeutic window of at least 6 h post-injury, and promotes neurobehavioral recovery post-TBI.

Lateral Fluid Percussion Injury Impairs Hippocampal Synaptic Soluble N-Ethylmaleimide Sensitive Factor Attachment Protein Receptor Complex Formation

Traumatic brain injury (TBI) and the activation of secondary injury mechanisms have been linked to impaired cognitive function, which, as observed in TBI patients and animal models, can persist for months and years following the initial injury. Impairments in neurotransmission have been well documented in experimental models of TBI, but the mechanisms underlying this dysfunction are poorly understood. Formation of the soluble N-ethylmaleimide-sensitive factor attachment protein receptor (SNARE) complex facilitates vesicular docking and neurotransmitter release in the synaptic cleft. Published studies highlight a direct link between reduced SNARE complex formation and impairments in neurotransmitter release. While alterations in the SNARE complex have been described following severe focal TBI, it is not known if deficits in SNARE complex formation manifest in a model with reduced severity. We hypothesized that lateral fluid percussion injury (lFPI) reduces the abundance of SNARE proteins, impairs SNARE complex formation, and contributes to impaired neurobehavioral function. To this end, rats were subjected to lFPI or sham injury and tested for acute motor performance and cognitive function at 3 weeks post-injury. lFPI resulted in motor impairment between 1 and 5 days post-injury. Spatial acquisition and spatial memory, as assessed by the Morris water maze, were significantly impaired at 3 weeks after lFPI. To examine the effect of lFPI on synaptic SNARE complex formation in the injured hippocampus, a separate cohort of rats was generated and brains processed to evaluate hippocampal synaptosomal-enriched lysates at 1 week post-injury. lFPI resulted in a significant reduction in multiple monomeric SNARE proteins, including VAMP2, and α-synuclein, and SNARE complex abundance. The findings in this study are consistent with our previously published observations suggesting that impairments in hippocampal SNARE complex formation may contribute to neurobehavioral dysfunction associated with TBI.

Novel therapies for combating chronic neuropathological sequelae of TBI

ABSTRACT Traumatic brain injury (TBI) is a risk factor for development of chronic neurodegenerative disorders later in life. This review summarizes the current knowledge and concepts regarding the connection between long‐term consequences of TBI and aging‐associated neurodegenerative disorders including Alzheimers disease (AD), chronic traumatic encephalopathy (CTE), and Parkinsonism, with implications for novel therapy targets. Several aggregation‐prone proteins such as the amyloid‐beta (A&bgr;) peptides, tau proteins, and &agr;‐synuclein protein are involved in secondary pathogenic cascades initiated by a TBI and are also major building blocks of the hallmark pathological lesions in chronic human neurodegenerative diseases with dementia. Impaired metabolism and degradation pathways of aggregation‐prone proteins are discussed as potentially critical links between the long‐term aftermath of TBI and chronic neurodegeneration. Utility and limitations of previous and current preclinical TBI models designed to study the link between TBI and chronic neurodegeneration, and promising intervention pharmacotherapies and non‐pharmacologic strategies to break this link, are also summarized. Complexity of long‐term neuropathological consequences of TBI is discussed, with a goal of guiding future preclinical studies and accelerating implementation of promising therapeutics into clinical trials. This article is part of the Special Issue entitled “Novel Treatments for Traumatic Brain Injury”. HIGHLIGHTSTBI is a risk factor for development of chronic neurodegenerative disorders later in life.Prolonged secondary injury cascades after TBI can result in chronic neurodegeneration.Chronic TBI and aging‐related dementias involve similar changes in aggregation prone molecules.Novel preclinical animal models of TBI are needed to better mimic the human condition.Polypathology of TBI supports the need for development of multifunctional therapies.