MCC950 Attenuates Microglial NLRP3-Mediated Chronic Neuroinflammation and Memory Impairment in a Rat Model of Repeated Low-Level Blast Exposure.

Blast-induced traumatic brain injury is typically regarded as a signature medical concern for military personnel who are exposed to explosive devices in active combat zones. However, soldiers as well as law enforcement personnel may be repeatedly exposed to low-level blasts during training sessions with heavy weaponries as part of combat readiness. Service personnel who sustain neurotrauma from repeated low-level blast (rLLB) exposure do not display overt pathological symptoms immediately but rather develop mild symptoms including cognitive impairments, attention deficits, mood changes, irritability, and sleep disturbances over time. Recently, we developed a rat model of rLLB by applying controlled low-level blast pressures (≤ 70 kPa) repeated five times successively to mimic the pressures experienced by service members. Using this model, we assessed anxiety-like symptoms, motor coordination, and short-term memory as a function of time. We also investigated the role of the NLRP3 inflammasome, a complex involved in chronic microglial activation and pro-inflammatory cytokine interleukin (IL)-1ß release, in rLLB-induced neuroinflammation. NLRP3 and caspase-1 protein expression, microglial activation, and IL-1ß release were examined as factors likely contributing to these neurobehavioral changes. Animals exposed to rLLB displayed acute and chronic short-term memory impairments and chronic anxiety-like symptoms accompanied by increased microglial activation, NLRP3 expression, and IL-1ß release. Treatment with MCC950, an NLRP3 inflammasome complex inhibitor, suppressed microglial activation, reduced NLRP3 expression and IL-1ß release, and improved short-term memory deficits after rLLB exposure. Collectively, this study demonstrates that rLLB induces chronic neurobehavioral and neuropathological changes by increasing NLRP3 inflammasome protein expression followed by cytokine IL-1ß release.

Dorsal root ganglion neurons recapitulate the traumatic axonal injury of CNS neurons in response to a rapid stretch in vitro.

Introduction: In vitro models of traumatic brain injury (TBI) commonly use neurons isolated from the central nervous system. Limitations with primary cortical cultures, however, can pose challenges to replicating some aspects of neuronal injury associated with closed head TBI. The known mechanisms of axonal degeneration from mechanical injury in TBI are in many ways similar to degenerative disease, ischemia, and spinal cord injury. It is therefore possible that the mechanisms that result in axonal degeneration in isolated cortical axons after in vitro stretch injury are shared with injured axons from different neuronal types. Dorsal root ganglia neurons (DRGN) are another neuronal source that may overcome some current limitations including remaining healthy in culture for long periods of time, ability to be isolated from adult sources, and myelinated in vitro. Methods: The current study sought to characterize the differential responses between cortical and DRGN axons to mechanical stretch injury associated with TBI. Using an in vitro model of traumatic axonal stretch injury, cortical and DRGN neurons were injured at a moderate (40% strain) and severe stretch (60% strain) and acute alterations in axonal morphology and calcium homeostasis were measured. Results: DRGN and cortical axons immediately form undulations in response to severe injury, experience similar elongation and recovery within 20 min after the initial injury, and had a similar pattern of degeneration over the first 24 h after injury. Additionally, both types of axons experienced comparable degrees of calcium influx after both moderate and severe injury that was prevented through pre-treatment with tetrodotoxin in cortical neurons and lidocaine in DRGNs. Similar to cortical axons, stretch injury also causes calcium activated proteolysis of sodium channel in DRGN axons that is prevented by treatment with lidocaine or protease inhibitors. Discussion: These findings suggest that DRGN axons share the early response of cortical neurons to a rapid stretch injury and the associated secondary injury mechanisms. The utility of a DRGN in vitro TBI model may allow future studies to explore TBI injury progression in myelinated and adult neurons.

A Method for Evaluating Brain Deformation Under Sagittal Blunt Impacts Using a Half-Skull Human-Scale Surrogate.

Trauma to the brain is a biomechanical problem where the initiating event is a dynamic loading (blunt, inertial, blast) to the head. To understand the relationship between the mechanical parameters of the injury and the spatial and temporal deformation patterns in the brain, there is a need to develop a reusable and adaptable experimental traumatic brain injury (TBI) model that can measure brain motion under varying parameters. In this effort, we aim to directly measure brain deformation (strain and strain rates) in different brain regions in a human head model using a drop tower. METHODS: Physical head models consisting of a half, sagittal plane skull, brain, and neck were constructed and subjected to crown and frontal impacts at two impact speeds. All tests were recorded with a high-speed camera at 1000 frames per second. Motion of visual markers within brain surrogates were used to track deformations and calculate spatial strain histories in 6 brain regions of interest. Principal strains, strain rates and strain impulses were calculated and reported. RESULTS: Higher impact velocities corresponded to higher strain values across all impact scenarios. Crown impacts were characterized by high, long duration strains distributed across the parietal, frontal and hippocampal regions whereas frontal impacts were characterized by sharply rising and falling strains primarily found in the parietal, frontal, hippocampal and occipital regions. High strain rates were associated with short durations and impulses indicating fast but short-lived strains. 2.23 m/s (5 mph) crown impacts resulted in 53% of the brain with shear strains higher than 0.15 verses 32% for frontal impacts. CONCLUSIONS: The results reveal large differences in the spatial and temporal strain responses between crown and forehead impacts. Overall, the results suggest that for the same speed, crown impact leads to higher magnitude strain patterns than a frontal impact. The data provided by this model provides unique insight into the spatial and temporal deformation patterns that have not been provided by alternate surrogate models. The model can be used to investigate how anatomical, material and loading features and parameters can affect deformation patterns in specific regions of interest in the brain.

Learning Environments and Evidence-Based Practices in Bioengineering and Biomedical Engineering.

This paper provides a synopsis of discussions related to the Learning Environments track of the Fourth BME Education Summit held at Case Western Reserve University in Cleveland, Ohio in May 2019. This summit was organized by the Council of Chairs of Bioengineering and Biomedical Engineering, and participants included over 300 faculty members from 100+ accredited undergraduate programs. The Learning Environments track had six interactive workshops that provided facilitated discussion and provide recommendations in the areas of: (1) Authentic project/problem identification in clinical, industrial, and global settings, (2) Experiential problem/project-based learning within courses, (3) Experiential learning in co-curricular learning settings, (4) Team-based learning, (5) Teaching to reach a diverse classroom, and (6) innovative platforms and pedagogy. A summary of the findings, best practices and recommendations from each of the workshops is provided under separate headings below, and a list of resources is provided at the end of this paper.

Animal model of repeated low-level blast traumatic brain injury displays acute and chronic neurobehavioral and neuropathological changes.

Blast-induced neurotrauma (BINT) is not only a signature injury to soldiers in combat field and training facilities but may also a growing concern in civilian population due to recent increases in the use of improvised explosives by insurgent groups. Unlike moderate or severe BINT, repeated low-level blast (rLLB) is different in its etiology as well as pathology. Due to the constant use of heavy weaponry as part of combat readiness, rLLB usually occurs in service members undergoing training as part of combat readiness. rLLB does not display overt pathological symptoms; however, earlier studies report chronic neurocognitive changes such as altered mood, irritability, and aggressive behavior, all of which may be caused by subtle neuropathological manifestations. Current animal models of rLLB for investigation of neurobehavioral and neuropathological alterations have not been adequate and do not sufficiently represent rLLB conditions. Here, we developed a rat model of rLLB by applying controlled low-level blast pressures (<10 psi) repeated successively five times to mimic the pressures experienced by service members. Using this model, we assessed anxiety-like symptoms, motor coordination, and short-term memory as a function of time. We also examined levels of superoxide-producing enzyme NADPH oxidase, microglial activation, and reactive astrocytosis as factors likely contributing to these neurobehavioral changes. Animals exposed to rLLB displayed acute and chronic anxiety-like symptoms, motor and short-term memory impairments. These changes were paralleled by increased microglial activation and reactive astrocytosis. Conversely, animals exposed to a single low-level blast did not display significant changes. Collectively, this study demonstrates that, unlike a single low-level blast, rLLB exerts a cumulative impact on different brain regions and produces chronic neuropathological changes in so doing, may be responsible for neurobehavioral alterations.

Engineering Fiber-Based Nervous Tissue Constructs for Axon Regeneration.

Biomaterial-based scaffolds used in nerve conduits including channels for confining regenerating axons and 3-dimensional (3D) gels as substrates for growth have made improvements in models of nerve repair. Many biomaterial strategies, however, continue to fall short of autologous nerve grafts, which remain the current gold standard in repairing severe nerve lesions (<20 mm). Intraluminal nerve conduit fibers have also shown considerable promise in directing regenerating axons in vitro and in vivo and have gained increasing interest for nerve repair. It is unknown, however, how growing axons respond to a fiber when encountered in a 3D environment. In this study, we considered a construct consisting of a compliant collagen hydrogel matrix and a fiber component to assess contact-guided axon growth. We investigated preferential axon outgrowth on synthetic and natural polymer fibers by utilizing small-diameter microfibers of poly-L-lactic acid and type I collagen representing 2 different fiber stiffnesses. We found that axons growing freely in a 3D hydrogel culture preferentially attach, turn and follow fibers with outgrowth rates and distances that far exceed outgrowth in a hydrogel alone. Wet-spun type I collagen from rat tail tendon performed the best, associated with highly aligned and accelerated outgrowth. This study also evaluated the response of dorsal root ganglion neurons from adult rats to provide data more relevant to axon regenerative potential in nerve repair. We found that ECM treatments on fibers enhanced the regeneration of adult axons indicating that both the physical and biochemical presentation of the fibers are essential for enhancing axon guidance and growth.

Behavioral Deficits in Animal Models of Blast Traumatic Brain Injury.

Blast exposure has been identified to be the most common cause for traumatic brain injury (TBI) in soldiers. Over the years, rodent models to mimic blast exposures and the behavioral outcomes observed in veterans have been developed extensively. However, blast tube design and varying experimental parameters lead to inconsistencies in the behavioral outcomes reported across research laboratories. This review aims to curate the behavioral outcomes reported in rodent models of blast TBI using shockwave tubes or open field detonations between the years 2008-2019 and highlight the important experimental parameters that affect behavioral outcome. Further, we discuss the role of various design parameters of the blast tube that can affect the nature of blast exposure experienced by the rodents. Finally, we assess the most common behavioral tests done to measure cognitive, motor, anxiety, auditory, and fear conditioning deficits in blast TBI (bTBI) and discuss the advantages and disadvantages of these tests.

Outcome measures from experimental traumatic brain injury in male rats vary with the complete temporal biomechanical profile of the injury event.

Millions suffer a traumatic brain injury (TBI) each year wherein the outcomes associated with injury can vary greatly between individuals. This study postulates that variations in each biomechanical parameter of a head trauma lead to differences in histological and behavioral outcome measures that should be considered collectively in assessing injury. While trauma severity typically scales with the magnitude of injury, much less is known about the effects of rate and duration of the mechanical insult. In this study, a newly developed voice-coil fluid percussion injury system was used to investigate the effects of injury rate and fluid percussion impulse on a collection of post-injury outcomes in male rats. Collectively the data suggest a potential shift in the specificity and progression of neuronal injury and function rather than a general scaling of injury severity. While a faster, shorter fluid percussion first presents as a mild TBI, neuronal loss and some behavioral tasks were similar among the slower and faster fluid percussion injuries. This study concludes that the sequelae of neuronal degeneration and behavioral outcomes are related to the complete temporal profile of the fluid percussion and do not scale only with peak pressure.

Blast exposure predisposes the brain to increased neurological deficits in a model of blast plus blunt traumatic brain injury.

Soldiers are often exposed to more than one traumatic brain injury (TBI) over the course of their service. In recent years, more attention has been drawn to the increased risk of neurological deficits caused by the 'blast plus' polytrauma, which typically is a blast trauma combined with other forms of TBI. In this study, we investigated the behavioral and neuronal deficits resulting from a blast plus injury involving a mild-moderate blast followed by a mild blunt trauma using the fluid percussion injury model. We identified that the blast injury predisposed the brain to increased cognitive deficits, chronic ventricular enlargement, increased neurodegeneration at acute time points and chronic neuronal loss. Interestingly, a single blast and single blunt injury differed in their onset and manifestation of cognitive and regional neuronal loss. We also identified the presence of cleaved RIP1 from caspase 8 mediated apoptosis in the blunt injury while the blast injury did not activate immediate apoptosis but led to decreased hilar neuronal survival over time.

Mechanical stretch induces myelin protein loss in oligodendrocytes by activating Erk1/2 in a calcium-dependent manner.

Myelin loss in the brain is a common occurrence in traumatic brain injury (TBI) that results from impact-induced acceleration forces to the head. Fast and abrupt head motions, either resulting from violent blows and/or jolts, cause rapid stretching of the brain tissue, and the long axons within the white matter tracts are especially vulnerable to such mechanical strain. Recent studies have shown that mechanotransduction plays an important role in regulating oligodendrocyte progenitors cell differentiation into oligodendrocytes. However, little is known about the impact of mechanical strain on mature oligodendrocytes and the stability of their associated myelin sheaths. We used an in vitro cellular stretch device to address these questions, as well as characterize a mechanotransduction mechanism that mediates oligodendrocyte responses. Mechanical stretch caused a transient and reversible myelin protein loss in oligodendrocytes. Cell death was not observed. Myelin protein loss was accompanied by an increase in intracellular Ca2+ and Erk1/2 activation. Chelating Ca2+ or inhibiting Erk1/2 activation was sufficient to block the stretch-induced loss of myelin protein. Further biochemical analyses revealed that the stretch-induced myelin protein loss was mediated by the release of Ca2+ from the endoplasmic reticulum (ER) and subsequent Ca2+ -dependent activation of Erk1/2. Altogether, our findings characterize an Erk1/2-dependent mechanotransduction mechanism in mature oligodendrocytes that de-stabilizes the myelination program.

Biomechanical Forces Regulate Gene Transcription During Stretch-Mediated Growth of Mammalian Neurons.

At birth, there are 100 billion neurons in the human brain, with functional neural circuits extending through the spine to the epidermis of the feet and toes. Following birth, limbs and vertebrae continue to grow by several orders of magnitude, forcing established axons to grow by up to 200 cm in length without motile growth cones. The leading regulatory paradigm suggests that biomechanical expansion of mitotic tissue exerts tensile force on integrated nervous tissue, which synchronizes ongoing growth of spanning axons. Here, we identify unique transcriptional changes in embryonic rat DRG and cortical neurons while the corresponding axons undergo physiological levels of controlled mechanical stretch in vitro. Using bioreactors containing cultured neurons, we recapitulated the peak biomechanical increase in embryonic rat crown-rump-length. Biologically paired sham and "stretch-grown" DRG neurons spanned 4.6- and 17.2-mm in length following static or stretch-induced growth conditions, respectively, which was associated with 456 significant changes in gene transcription identified by genome-wide cDNA microarrays. Eight significant genes found in DRG were cross-validated in stretch-grown cortical neurons by qRT-PCR, which included upregulation of Gpat3, Crem, Hmox1, Hpse, Mt1a, Nefm, Sprr1b, and downregulation of Nrep. The results herein establish a link between biomechanics and gene transcription in mammalian neurons, which elucidates the mechanism underlying long-term growth of axons, and provides a basis for new research in therapeutic axon regeneration.

Characterization of a controlled shock wave delivered by a pneumatic table-top gas driven shock tube.

Blast simulators facilitate the creation of shock waves and measurement of pressure morphology in a controlled laboratory setting and are currently a vital model for replicating blast-induced neurotrauma. Due to the maintenance and operation cost of conventional blast simulators, we developed a pneumatic, table-top, gas-driven shock tube to test an alternative method of shock wave generation using a membrane-less driver section. Its unique operational mechanism based on air gun technology does not rely on a plastic membrane rupture for the generation of pressure pulses, allowing the simulator to be quickly reset and thus decreasing the experimental turnaround time. The focus of this study is to demonstrate that this proof-of-concept device can generate shock waves with diverse characteristics based on the selection of driver gas, driver pressurization, and driven section material. Pressure waves were generated using compressed nitrogen or helium at 15 psig and 80 psig and were analyzed based on their velocity and profile shape characteristics. At 15 psig, independent of the type of driver gas, driver pressurization, and driven section material, pressure pulses travelled at sonic velocities. At 80 psig, generation of shock waves was observed in all conditions. The choice of the driver gas affected the velocities of the resulting pressure waves and the shape of pressure waveforms, particularly the peak overpressure and rise time values. Our results demonstrate that depending on the selection of driver gas and magnitude of driver pressurization, the shock wave signatures can be controlled and altered using a piston-based driver section.

Animal Models of Traumatic Brain Injury and Assessment of Injury Severity.

Traumatic brain injury (TBI) contributes a major cause of death, disability, and mental health disorders. Most TBI patients suffer long-term post-traumatic stress disorder, cognitive dysfunction, and disability. The underlying molecular and cellular mechanisms of such neuropathology progression in TBI remain elusive. In part, it is due to non-standardized classification of mild, moderate, and severe injury in various animal models of TBI. Thus, a better diagnosis and treatment requires a better understanding of the injury mechanisms in a well-defined severity of mild, moderate, and severe injury in different models that may potentially reflect the various types of human brain injuries. The purpose of this review article is to highlight the classification of mild, moderate, and severe injury in various animal models of TBI with special focus on mixed injury that represents a translational concussive head injury. We will classify animal models of TBI broadly into focal injury, diffuse injury, and mixed injury. Focal injury, a localized injury, is represented by animal models of controlled cortical impact, penetrating ballistic-like brain injury, and Feeney or Shohami weight drop injury. A global diffuse injury is best represented by shock tube model of primary blast injury, and Marmarou or Maryland weight drop model. A mixed injury consists of focal and diffuse injury which reproduces the concussive clinical syndrome, and it is best studied in animal model of lateral fluid percussion injury.

High Ca2+ Influx During Traumatic Brain Injury Leads to Caspase-1-Dependent Neuroinflammation and Cell Death.

We investigated the hypothesis that high Ca2+ influx during traumatic brain injury induces the activation of the caspase-1 enzyme, which triggers neuroinflammation and cell apoptosis in a cell culture model of neuronal stretch injury and an in vivo model of fluid percussion injury (FPI). We first established that stretch injury causes a rapid increase in the intracellular Ca2+ level, which activates interleukin-converting enzyme caspase-1. The increase in the intracellular Ca2+ level and subsequent caspase-1 activation culminates into neuroinflammation via the maturation of IL-1ß. Further, we analyzed caspase-1-mediated apoptosis by TUNEL staining and PARP western blotting. The voltage-gated sodium channel blocker, tetrodotoxin, mitigated the stretch injury-induced neuroinflammation and subsequent apoptosis by blocking Ca2+ influx during the injury. The effect of tetrodotoxin was similar to the caspase-1 inhibitor, zYVAD-fmk, in neuronal culture. To validate the in vitro results, we demonstrated an increase in caspase-1 activity, neuroinflammation and neurodegeneration in fluid percussion-injured animals. Our data suggest that neuronal injury/traumatic brain injury (TBI) can induce a high influx of Ca2+ to the cells that cause neuroinflammation and cell death by activating caspase-1, IL-1ß, and intrinsic apoptotic pathways. We conclude that excess IL-1ß production and cell death may contribute to neuronal dysfunction and cognitive impairment associated with TBI.

Traumatic brain injury induced matrix metalloproteinase2 cleaves CXCL12α (stromal cell derived factor 1α) and causes neurodegeneration.

Traumatic brain injury (TBI), even at mild levels, can activate matrix metalloproteinases (MMPs) and the induction of neuroinflammation that can result in blood brain barrier breakdown and neurodegeneration. MMP2 has a significant role in neuroinflammation and neurodegeneration by modulating the chemokine CXCL12α (stromal cell derived factor SDF-1α) signaling pathway and the induction of apoptosis. SDF-1α is responsible for cell proliferation and differentiation throughout the nervous system and is also implicated in various neurodegenerative illnesses. We hypothesized that TBI leads to MMP2 activation and cleavage of the N-terminal 4 amino acid residues of CXCL12α with generation of the highly neurotoxic fragment SDF-1(5-67). Using an in vitro stretch-injury model of rat neuronal cultures and the in vivo fluid percussion injury (FPI) model in rats, we found that oxidative stress has a significant role in the activation of MMP2. This is initiated by the induction of free radical generating enzyme NADPH oxidase 1 (NOX1). Induction of NOX1 correlated well with the signatures of oxidative stress marker, 4HNE in the injured neuronal cultures and cerebral cortex of rats. Further, using MMP2 siRNA and pharmacological MMP2 inhibitor, ARP100, we established the neurodegenerative role of MMP2 in cleaving SDF-1α to a neurotoxic fragment SDF-1(5-67). By immunofluorescence, western blotting and TUNEL experiments, we show the cleaved form of SDF leads to apoptotic cell death in neurons. This work identifies a new potential therapeutic target to reduce the complications of brain damage in TBI.

Quantitative optical coherence elastography based on fiber-optic probe for in situ measurement of tissue mechanical properties.

We developed a miniature quantitative optical coherence elastography (qOCE) instrument with an integrated Fabry-Perot force sensor, for in situ elasticity measurement of biological tissue. The technique has great potential for biomechanics modeling and clinical diagnosis. We designed the fiber-optic qOCE probe that was used to exert a compressive force to deform tissue at the tip of the probe. Using the space-division multiplexed optical coherence tomography (OCT) signal detected by a spectral domain OCT engine, we were able to quantify the probe deformation that was proportional to the force applied, and to quantify the tissue deformation corresponding to the external stimulus. Simultaneous measurement of force and displacement allowed us to extract Young's modulus of biological tissue. We experimentally calibrated our qOCE instrument, and validated its effectiveness on tissue mimicking phantoms and biological tissues.

Role of Matrix Metalloproteinases in the Pathogenesis of Traumatic Brain Injury.

Traumatic brain injury (TBI) is a major cause of mortality and morbidity worldwide. Studies revealed that the pathogenesis of TBI involves upregulation of MMPs. MMPs form a large family of closely related zinc-dependent endopeptidases, which are primarily responsible for the dynamic remodulation of the extracellular matrix (ECM). Thus, they are involved in several normal physiological processes like growth, development, and wound healing. During pathophysiological conditions, MMPs proteolytically degrade various components of ECM and tight junction (TJ) proteins of BBB and cause BBB disruption. Impairment of BBB causes leakiness of the blood from circulation to brain parenchyma that leads to microhemorrhage and edema. Further, MMPs dysregulate various normal physiological processes like angiogenesis and neurogenesis, and also they participate in the inflammatory and apoptotic cascades by inducing or regulating the specific mediators and their receptors. In this review, we explore the roles of MMPs in various physiological/pathophysiological processes associated with neurological complications, with special emphasis on TBI.

Developmental axon stretch stimulates neuron growth while maintaining normal electrical activity, intracellular calcium flux, and somatic morphology.

Elongation of nerve fibers intuitively occurs throughout mammalian development, and is synchronized with expansion of the growing body. While most tissue systems enlarge through mitosis and differentiation, elongation of nerve fibers is remarkably unique. The emerging paradigm suggests that axons undergo stretch as contiguous tissues enlarge between the proximal and distal segments of spanning nerve fibers. While stretch is distinct from growth, tension is a known stimulus which regulates the growth of axons. Here, we hypothesized that the axon stretch-growth process may be a natural form of injury, whereby regenerative processes fortify elongating axons in order to prevent disconnection. Harnessing the live imaging capability of our axon stretch-growth bioreactors, we assessed neurons both during and following stretch for biomarkers associated with injury. Utilizing whole-cell patch clamp recording, we found no evidence of changes in spontaneous action potential activity or degradation of elicited action potentials during real-time axon stretch at strains of up to 18% applied over 5 min. Unlike traumatic axonal injury, functional calcium imaging of the soma revealed no shifts in free intracellular calcium during axon stretch. Finally, the cross-sectional areas of nuclei and cytoplasms were normal, with no evidence of chromatolysis following week-long stretch-growth limited to the lower of 25% strain or 3 mm total daily stretch. The neuronal growth cascade coupled to stretch was concluded to be independent of the changes in membrane potential, action potential generation, or calcium flux associated with traumatic injury. While axon stretch-growth is likely to share overlap with regenerative processes, we conclude that developmental stretch is a distinct stimulus from traumatic axon injury.

Effect of acute stretch injury on action potential and network activity of rat neocortical neurons in culture.

The basis for acute seizures following traumatic brain injury (TBI) remains unclear. Animal models of TBI have revealed acute hyperexcitablility in cortical neurons that could underlie seizure activity, but studying initiating events causing hyperexcitability is difficult in these models. In vitro models of stretch injury with cultured cortical neurons, a surrogate for TBI, allow facile investigation of cellular changes after injury but they have only demonstrated post-injury hypoexcitability. The goal of this study was to determine if neuronal hyperexcitability could be triggered by in vitro stretch injury. Controlled uniaxial stretch injury was delivered to a spatially delimited region of a spontaneously active network of cultured rat cortical neurons, yielding a region of stretch-injured neurons and adjacent regions of non-stretched neurons that did not directly experience stretch injury. Spontaneous electrical activity was measured in non-stretched and stretch-injured neurons, and in control neuronal networks not subjected to stretch injury. Non-stretched neurons in stretch-injured cultures displayed a three-fold increase in action potential firing rate and bursting activity 30-60 min post-injury. Stretch-injured neurons, however, displayed dramatically lower rates of action potential firing and bursting. These results demonstrate that acute hyperexcitability can be observed in non-stretched neurons located in regions adjacent to the site of stretch injury, consistent with reports that seizure activity can arise from regions surrounding the site of localized brain injury. Thus, this in vitro procedure for localized neuronal stretch injury may provide a model to study the earliest cellular changes in neuronal function associated with acute post-traumatic seizures.

Fluid percussion injury device for the precise control of injury parameters.

BACKGROUND: Injury to the brain can occur from a variety of physical insults and the degree of disability can greatly vary from person to person. It is likely that injury outcome is related to the biomechanical parameters of the traumatic event such as magnitude, direction and speed of the forces acting on the head. NEW METHOD: To model variations in the biomechanical injury parameters, a voice coil driven fluid percussion injury (FPI) system was designed and built to generate fluid percussion waveforms with adjustable rise times, peak pressures, and durations. Using this system, pathophysiological outcomes in the rat were investigated and compared to animals injured with the same biomechanical parameters using the pendulum based FPI system. RESULTS IN COMPARISON WITH EXISTING METHODS: Immediate post-injury behavior shows similar rates of seizures and mortality in adolescent rats and similar righting times, toe pinch responses and mortality rates in adult rats. Interestingly, post injury mortality in adult rats was sensitive to changes in injury rate. Fluoro-Jade labeling of degenerating neurons in the hilus and CA2-3 hippocampus were consistent between injuries produced with the voice coil and pendulum operated systems. Granule cell population spike amplitude to afferent activation, a measure of dentate network excitability, also showed consistent enhancement 1 week after injury using either system. CONCLUSIONS: Overall our results suggest that this new FPI device produces injury outcomes consistent with the commonly used pendulum FPI system and has the added capability to investigate pathophysiology associated with varying rates and durations of injury.