Impaired Axonal Transport Research Articles

Microtubule dysfunction precedes transport impairment and mitochondria damage in MPP+‐induced neurodegeneration

Dysfunction of the microtubule (MT) system is an emerging theme in the pathogenesis of Parkinson's disease. This study was designed to investigate the putative role of MT dysfunction in dopaminergic neuron death induced by the neurotoxin 1-methyl-4-phenylpiridinium (MPP(+)). In nerve growth factor-differentiated PC12 cells, we have analyzed post-translational modifications of tubulin known to be associated with differently dynamic MTs and show that MPP(+) causes a selective loss of dynamic MTs and a concomitant enrichment of stable MTs. Through a direct live cell imaging approach, we show a significant reduction of MT dynamics following exposure to MPP(+) and a reorientation of MTs. Furthermore, these alterations precede the impairment of intracellular transport as revealed by changes in mitochondria movements along neurites and their accumulation into varicosities. We have also analyzed activation of caspase 3 and mitochondrial injury, well-known alterations induced by MPP(+), and found that they are noticeable only when MT dysfunction is already established. These data provide the first evidence that axonal transport impairment and mitochondrial damage might be a consequence of MT dysfunction in MPP(+) -induced neurodegeneration, lending support to the concept that alterations of MT organization and dynamics could play a pivotal role in neuronal death in Parkinson's disease.

Puerarin attenuates glutamate-induced neurofilament axonal transport impairment

Puerarin (Pur) is a primary component of the most functional extracts of Pueraria lobata used in traditional Chinese medicine for centuries. Since it has been postulated that Pur protects the brain against glutamate (Glu) neurotoxicity, we investigated the effects of Pur on Glu-induced axonal transport impairment in primary hippocampal neurons in this study. Primary hippocampal cultures were prepared from 2-day-old Sprague-Dawley rats. Intracellular calcium concentration [Ca(2+)](i), neurofilament (NF) phosphorylation and protein kinase activity for Cdk5 were measured. Time-lapse imaging technology was used to capture the NF axonal transport in the cultured neurons with transiently transfected fluorescence protein linked to the N-terminus of NF-M (EGFP-NFM). The results showed that Pur significantly diminished the Glu-induced elevation of [Ca(2+)](i) in dose-dependent manner and antagonized the Glu-evoked increases in NF phosphorylation at protein levels. The neurons under the Glu treatment displayed the accumulation of immobile NF clusters in the cell body and the reduced rates of axonal transport of NFs by 72.8% compared to the control neurons. Intriguingly, Pur reversed the slowed rate of the axonal transport by 35.6%. Pur also remarkably attenuated Glu-evoked activation of Cdk5. Pur may play a role in protecting against Glu-induced NF axonal transport impairment in rat primary hippocampal neurons by inhibiting the increased [Ca(2+)](i) and by impeding the activation of Cdk5.

A serial study of retinal changes following optic neuritis with sample size estimates for acute neuroprotection trials

Following an episode of optic neuritis, thinning of the retinal nerve fibre layer, which indicates axonal loss, is observed using optical coherence tomography. The longitudinal course of the retinal changes has not been well characterized. We performed a serial optical coherence tomography study in patients presenting with optic neuritis in order to define the temporal evolution of retinal nerve fibre layer changes and to estimate sample sizes for proof-of-concept trials of neuroprotection using retinal nerve fibre layer loss as the outcome measure. Twenty-three patients (7 male, 16 female, mean age 31 years) with acute clinically isolated unilateral optic neuritis were recruited to undergo optical coherence tomography, visual assessments and visual evoked potentials at presentation (median 16 days from onset of visual loss) and after 3, 6, 12 and 18 months. Compared with the clinically unaffected fellow eye, the retinal nerve fibre layer thickness of the affected eye was significantly increased at presentation and significantly reduced at all later time points. The evolution of retinal nerve fibre layer changes in the affected eye fitted well with an exponential model, with thinning appearing a mean of 1.6 months from symptom onset and the rate of ongoing retinal nerve fibre layer loss decreasing thereafter. At presentation, increased retinal nerve fibre layer thickness was associated with impaired visual acuity and prolonged visual evoked potential latency. Visual function after 12 months was not related to the extent of acute retinal nerve fibre layer swelling but was significantly associated with the extent of concurrent retinal nerve fibre layer loss. Sample size calculations for placebo-controlled trials of acute neuroprotection indicated that the numbers needed after 6 months of follow up are smaller than those after 3 months and similar to those after 12 months of follow-up. Study power was greater when investigating differences between clinically unaffected and affected eyes rather than retinal nerve fibre layer thickness of the affected eye alone. Inflammation in the optic nerve and impaired axonal transport (implied by retinal nerve fibre layer swelling) are associated with visual dysfunction and demyelination (long visual evoked potential latency) during acute optic neuritis. Retinal nerve fibre layer thinning is usually evident within 3 months. Optical coherence tomography-measured retinal nerve fibre layer loss after 6 months is a suitable outcome measure for proof-of-concept trials of acute neuroprotection in optic neuritis.

The Legs at odd angles (Loa) Mutation in Cytoplasmic Dynein Ameliorates Mitochondrial Function in SOD1G93A Mouse Model for Motor Neuron Disease

Amyotrophic lateral sclerosis (ALS) is a debilitating and fatal late-onset neurodegenerative disease. Familial cases of ALS (FALS) constitute ∼10% of all ALS cases, and mutant superoxide dismutase 1 (SOD1) is found in 15–20% of FALS. SOD1 mutations confer a toxic gain of unknown function to the protein that specifically targets the motor neurons in the cortex and the spinal cord. We have previously shown that the autosomal dominant Legs at odd angles (Loa) mutation in cytoplasmic dynein heavy chain (Dync1h1) delays disease onset and extends the life span of transgenic mice harboring human mutant SOD1G93A. In this study we provide evidence that despite the lack of direct interactions between mutant SOD1 and either mutant or wild-type cytoplasmic dynein, the Loa mutation confers significant reductions in the amount of mutant SOD1 protein in the mitochondrial matrix. Moreover, we show that the Loa mutation ameliorates defects in mitochondrial respiration and membrane potential observed in SOD1G93A motor neuron mitochondria. These data suggest that the Loa mutation reduces the vulnerability of mitochondria to the toxic effects of mutant SOD1, leading to improved mitochondrial function in SOD1G93A motor neurons.

Polyglutamine Diseases: Where does Toxicity Come from? What is Toxicity? Where are We Going?

Although the genetic basis of polyglutamine diseases has been recognized for 20 years, their molecular basis is still unclear. We have no therapeutic strategies for these intractable neurodegenerative disorders. To adequately treat patients, we must clarify the molecular basis of polyglutamine diseases. Three main issues address their molecular pathogenesis: whether the specific structure of expanded polyglutamine diseases results in cellular toxicity; what type of dysfunction causes them; and how the toxic structure causes dysfunction, that is, the link between structure and dysfunction. For structures, expanded polyglutamine proteins undergo transformation from monomers to oligomers and inclusions. One can hypothesize that one of these structures might cause the polyglutamine disease. Although the expanded polyglutamine protein is toxic, it does not explain the selective vulnerability of specific neurons in each polyglutamine disease. The normal function of each protein, including protein-protein interaction and modification, might also be crucial for pathogenesis. For dysfunction, various molecular mechanisms have been proposed, including dysregulation of transcription, impairment of the ubiquitin-proteasome system, mitochondrial dysfunction, dysregulation of intracellular Ca(2+) homeostasis, impairment of axonal transport and genotoxic stress. These hypotheses might correlate with each other. In addition, the disease pathogenesis of might not be exclusive to one particular structure or dysfunction. To develop a therapeutic strategy for patients with polyglutamine disease, identifying the most toxic structure and the earliest event in the pathogenesis is important. We review the current understanding of the toxic structure and dysfunction by expanded polyglutamine proteins and suggest directions for future studies of polyglutamine diseases.

Nerve growth factor as a paradigm of neurotrophins related to Alzheimer's disease

Converging lines of evidence on the possible connection between NGF signaling and Alzheimer's diseases (AD) are unraveling new facets which could depict this neurotrophin (NTF) in a more central role. AD animal models have provided evidence that a shortage of NGF supply may induce an AD-like syndrome. In vitro experiments, moreover, are delineating a possible temporal and causal link between APP amiloydogenic processing and altered post-translational tau modifications. After NGF signaling interruption, the pivotal upstream players of the amyloid cascade (APP, beta-secretase, and active form of gamma-secretase) are up-regulated, leading to an increased production of amyloid beta peptide (Abeta) and to its intracellular aggregation in molecular species of different sizes. Contextually, the Abeta released pool generates an autocrine toxic loop in the same healthy neurons. At the same time tau protein undergoes anomalous, GSKbeta-mediated, phosphorylation at specific pathogenetic sites (Ser262 and Thr 231), caspase(s) and calpain- I- mediated truncation, detachment from microtubules with consequent cytoskeleton collapse and axonal transport impairment. All these events are inhibited when the amyloidogenic processing is reduced by beta and gamma secretase inhibitors or anti-Abeta antibodies and appear to be causally correlated to TrkA, p75CTF, Abeta, and PS1 molecular association in an Abeta-mediated fashion. In this scenario, the so-called trophic action exerted by NGF (and possibly also by other neurotrophins) in these targets neurons is actually the result of an anti-amyloidogenic activity.

Increased expression of cdk5/p25 in N2a cells leads to hyperphosphorylation and impaired axonal transport of neurofilament proteins

Aims Alzheimer's disease (AD) is the leading cause of dementia. The increased cdk5 expression and enhanced phosphorylation of tau and NFs have been seen in AD patients. Our study aimed at investigating the effects of increased cdk5 activity on axonal transport of neurofilaments (NFs). Main methods In this study, we used a molecular engineering approach to overexpress cdk5/p25 in neuroblastoma N2a cells and investigated the effects on axonal transport with live cell imaging techniques. Key findings In stably transfected cells, there was a 2.5-fold increase in cdk5 activity compared to non-transfected cells, which in turn led to a dramatic increase in phosphorylation of NFs and tau at several phosphorylation sites. Using time-lapse imaging technology, the transport of NFs was captured in the cells overexpressing cdk5/p25, which were also transiently transfected with fluorescence protein linked to the N-terminus of NF-M (EGFP-NFM). The cdk5/p25 cells displayed significantly slower rates of axonal transport of NFs, with accumulation of immobile NF clusters observed in the cell body. Roscovitine, an inhibitor of cdk5, significantly reversed this defect in axonal transport. Significance These results suggest that increased cdk5 activity found in AD subjects may be crucially related to the pathogenesis of AD via an underlying mechanism by which it promotes accumulation of excessively phosphorylated cytoskeletal NF proteins, leading to the enduring impairment of axonal transport of NFs.

Retinal nerve fiber layer thickness is associated with lesion length in acute optic neuritis

Acute optic neuritis occurs with and without papillitis. The presence of papillitis has previously been thought to imply an anterior location of the neuritis, but imaging studies seeking to test this hypothesis have been inconclusive. This prospective observational cohort study included 41 patients with unilateral optic neuritis and 19 healthy volunteers. All patients were evaluated and examined within 28 days of onset of symptoms. The peripapillary retinal nerve fiber layer thickness (RNFLT), an objective quantitative measure of optic nerve head edema, was measured by optical coherence tomography and the length and location of the inflammatory optic nerve lesion were evaluated using MRI. Ophthalmoscopically, 34% of the patients had papillitis. The retinal nerve fiber layer in affected eyes (mean 123.1 microm) was higher during the acute phase than that of fellow eyes (mean 98.1 microm, p < 0.0001) and higher than that in healthy control eyes (mean 97.1 microm, p < 0.0001). The RNFLT was related to the length of the optic nerve lesion (p = 0.0002), but not to the location of the optic nerve lesions (p = 0.72). In this study of the acute phase of optic neuritis, the degree of optic nerve head edema depended upon the extent of the optic nerve lesion, but not on its location. This suggests that factors other than inflammation, such as compromised venous drainage, vascular leakage, impaired axonal transport, and other mechanisms, are involved in the development of optic nerve head edema in optic neuritis.

A Versatile High Throughput Screening System for the Simultaneous Identification of Anti-Inflammatory and Neuroprotective Compounds

In many chronic neurodegenerative diseases including Frontotemporal Dementia and Alzheimer's disease (AD), microglial activation is suggested to be involved in pathogenesis or disease progression. Activated microglia secrete a variety of cytokines, including interleukin-1beta, interleukin-6, and tumor necrosis factor as well as reactive oxygen and nitrogen species (ROS/RNS). ROS and RNS contribute to alterations in neuronal glucose uptake, inhibition of mitochondrial enzymes, a decrease in mitochondrial membrane potential, impaired axonal transport, and synaptic signaling. In addition, ROS act as signaling molecules in pro-inflammatory redox-active signal transduction pathways. To establish a high throughput screening system for anti-inflammatory and neuroprotective compounds, we have constructed an "Enhanced Green Fluorescent protein" (EGFP) expressing neuronal cell line and set up a murine microglia/neuron co-culture system with these EGFP expressing neuronal cells. We show that microglia activation leads to neuronal cell death, which can be conveniently measured by loss of neuronal EGFP fluorescence. Moreover, we used this system to test selected polyphenolic compounds for their ability to downregulate inflammatory markers and to protect neurons against microglial insult. We suggest that this system might allow accelerated drug discovery for the treatment of inflammation-mediated neurodegenerative diseases.

Lack of Axonal Sprouting of Spared Propriospinal Fibers Caudal to Spinal Contusion Injury Is Attributed to Chronic Axonopathy

We have previously shown that a small percentage of long descending propriospinal tract (LDPT) axons are spared, whereas few short thoracic propriospinal (TPS) fibers survive 2 weeks following severe (50 mm weight drop) low thoracic spinal cord contusion injury (SCI). Here, we extended those findings to a moderate (25 mm weight drop) T9 SCI and assessed the effects of this lesion severity on propriospinal tract fibers at different time periods after injury. We anterogradely labeled fibers with fluororuby (FR) or WGA-HRP to determine their location and number 2, 4, 6, and 16 weeks post-SCI. Findings were compared with non-injured controls. At chronic time points, surviving FR-labeled LDPT fibers rostral to the injury remained as reactive endings or as putative regenerative sprouts. Caudal to the injury, spared LDPT fibers ran along a rim of lateral and ventral white matter, and ended as small abnormal-appearing putative terminal boutons or reactive endings within the intermediate gray matter of lumbosacral cord, with little axonal arborization and no evidence of injury-induced sprouting. One striking difference in the WGA-HRP experimental operates was the increased density of labeling of spared axons within the white matter caudal to the injury compared to controls. This labeling pattern was reminiscent of the labeling found after axotomy in studies by others, and raises a question as to contusion injury-induced impaired axonal transport. We hypothesize that axonal sprouting of axons after partial spinal cord injury seen in previous investigations was not found in the present investigation because of the additional pathological effects of contusion injury, similar to what is observed after traumatic brain injury.

Cerebellar Disorders—At the Crossroad of Molecular Pathways and Diagnosis

Our understanding of the pathogenesis of cerebellar ataxias has started several decades ago and is continuously growing. The numerous mechanisms of cerebellar dysfunction are being discovered by numerous groups of researchers worldwide. Neuronal damage results from a complex interaction of metabolic pathways, which leads to symptoms observed in cerebellar disorders. The main mechanisms at the molecular level are the following: impairment of DNA repair and replication, deregulation of transcription/deficits of processing/transport of RNA, abnormal protein transport and misfolding, aggregates both at the nuclear and cytosolic level, activation of caspases, apoptosis, involvement of autophagic mechanisms, oxidative stress and mitochondrial dysfunction, excitotoxicity, abnormal lipid metabolism, impaired axonal transport and vesicle trafficking, and defects of the neurotransmission. The convergence of the current clinical classification with molecular findings is expected. This integration is a basic substrate for the rationale development of therapies.

Negative Regulation of Active Zone Assembly by a Newly Identified SR Protein Kinase

Author SummaryNeurons communicate with each other through electrochemical impulses transmitted primarily at specialized intercellular junctions termed synapses. At each synapse, the primary site of synaptic vesicle fusion occurs at the active zone, an electron-dense presynaptic membrane with associated fibrillary matrix. Many active zones also possess one or more electron-dense cytosolic projections that are believed to facilitate vesicle mobilization to the active zone membrane and are required for normal synaptic transmission. These electron-dense projections are referred to as T-bars in Drosophila or ribbons in vertebrates. The molecular composition of these structures remains poorly characterized, and very little is known about how these structures are specifically assembled and stabilized at the presynaptic membrane. Here, we identify in Drosophila a neuronally expressed serine-arginine kinase called SRPK79D that localizes to the presynaptic active zone and that through its kinase activity appears to repress T-bar formation within peripheral axons. Our study thus provides evidence for kinase-dependent repression of active zone assembly, with implications for the development and growth of synaptic connections throughout the nervous system.

Impaired Axonal Transport in Motor Neurons Correlates with Clinical Prion Disease

Prion diseases are fatal neurodegenerative disorders causing motor dysfunctions, dementia and neuropathological changes such as spongiosis, astroglyosis and neuronal loss. The chain of events leading to the clinical disease and the role of distinct brain areas are still poorly understood. The role of nervous system integrity and axonal properties in prion pathology are still elusive. There is no evidence of both the functional axonal impairments in vivo and their connection with prion disease. We studied the functional axonal impairments in motor neurons at the onset of clinical prion disease using the combination of tracing as a functional assay for axonal transport with immunohistochemistry experiments. Well-established and novel confocal and ultramicroscopy techniques were used to image and quantify labeled neurons. Despite profound differences in the incubation times, 30% to 45% of neurons in the red nucleus of different mouse lines showed axonal transport impairments at the disease onset bilaterally after intracerebral prion inoculation and unilaterally—after inoculation into the right sciatic nerve. Up to 94% of motor cortex neurons also demonstrated transport defects upon analysis by alternative imaging methods. Our data connect axonal transport impairments with disease symptoms for different prion strains and inoculation routes and establish further insight on the development of prion pathology in vivo. The alterations in localization of the proteins involved in the retrograde axonal transport allow us to propose a mechanism of transport disruption, which involves Rab7-mediated cargo attachment to the dynein-dynactin pathway. These findings suggest novel targets for therapeutic and diagnostic approaches in the early stages of prion disease.

Axonal degeneration in multiple sclerosis: The mitochondrial hypothesis

Multiple sclerosis (MS) is a chronic disease of the central nervous system, affecting more than 2 million people worldwide. Traditionally considered an inflammatory demyelinating disease, recent evidence now points to axonal degeneration as crucial to the development of irreversible disability. Studies show that axonal degeneration occurs throughout the entire course of MS. Although the specific mechanisms causing axonal damage may differ at various stages, mitochondrial failure seems to be a common underlying theme. This review addresses the mitochondrial hypothesis for axonal degeneration in MS, highlighting the mechanisms by which mitochondrial dysfunction leads to axonal disruption in acute inflammatory lesions and the chronic axonopathy in progressive MS. Emphasis is placed on Ca(2+), free radical production, and permeability transition pore opening as key players in mitochondrial failure, axonal transport impairment, and subsequent axonal degeneration. In addition, the role of mitochondria as therapeutic targets for neuroprotection in MS is addressed.

Mechanical membrane injury induces axonal beading through localized activation of calpain

Diffuse axonal injury (DAI), a major component of traumatic brain injury, is characterized by a sequence of neurochemical reactions initiated at the time of trauma and resulting in axonal degeneration and cell death. Calcium influx through mechanically induced axolemmal pores and subsequent activation of calpains are thought to be responsible for the cytoskeletal damage leading to impaired axonal transport. Focal disruption of cytoskeleton accompanied by the accumulation of transported membranous cargo leads to axonal beading which is the characteristic morphology of DAI. By applying fluid shear stress injury on cultured primary neurons, acute calcium (Ca 2+) and calpain responses of axons to mechanical trauma were investigated. Intracellular Ca 2+ concentration ([Ca 2+] i) shows a steady increase following injury that can be blocked by sealing membrane pores with Poloxamer 188 and by chelating intra- or extracellular Ca 2+. Calpain activity increases in response to mechanical injury and this increase depends on Ca 2+ availability and on axolemmal permeability. Both the [Ca 2+] i increase and calpain activity exhibit focal peaks along the axons which co-localize with mitochondria and predict future axonal bead locations. These findings suggest that mechanoporation may be the initiating mechanism resulting in ensuing calcium fluxes and subsequent calpain activity and that post-injury membrane repair may be a valid therapeutic approach for acute intervention in DAI.

Phosphorylated Tau Interacts with c-Jun N-terminal Kinase-interacting Protein 1 (JIP1) in Alzheimer Disease

In Alzheimer disease (AD) and frontotemporal dementia the microtubule-associated protein Tau becomes progressively hyperphosphorylated, eventually forming aggregates. However, how Tau dysfunction is associated with functional impairment is only partly understood, especially at early stages when Tau is mislocalized but has not yet formed aggregates. Impaired axonal transport has been proposed as a potential pathomechanism, based on cellular Tau models and Tau transgenic mice. We recently reported K369I mutant Tau transgenic K3 mice with axonal transport defects that suggested a cargo-selective impairment of kinesin-driven anterograde transport by Tau. Here, we show that kinesin motor complex formation is disturbed in the K3 mice. We show that under pathological conditions hyperphosphorylated Tau interacts with c-Jun N-terminal kinase- interacting protein 1 (JIP1), which is associated with the kinesin motor protein complex. As a result, transport of JIP1 into the axon is impaired, causing JIP1 to accumulate in the cell body. Because we found trapping of JIP1 and a pathological Tau/JIP1 interaction also in AD brain, this may have pathomechanistic implications in diseases with a Tau pathology. This is supported by JIP1 sequestration in the cell body of Tau-transfected primary neuronal cultures. The pathological Tau/JIP1 interaction requires phosphorylation of Tau, and Tau competes with the physiological binding of JIP1 to kinesin light chain. Because JIP1 is involved in regulating cargo binding to kinesin motors, our findings may, at least in part, explain how hyperphosphorylated Tau mediates impaired axonal transport in AD and frontotemporal dementia.

Neuron–glia interactions underlie ALS-like axonal cytoskeletal pathology

Amyotrophic lateral sclerosis (ALS) is a devastating disorder involving loss of movement due to degeneration of motor neurons. Studies suggest that in ALS axonal dysfunction precedes the death of motor neurons. Pathologically, ALS is characterized by neurofilamentous swellings (spheroids) within the axons of motor neurons. However, the causes of this axonopathy and possible resulting axonal dysfunction are not known. Using a novel model of cultured mouse motor neurons, we have determined that these neurons are susceptible to proximal axonopathy, which is related to the glial environment. This axonopathy showed remarkable similarity, both morphologically and neurochemically, to spheroids that develop over months in SOD1(G93A) transgenic mice. Focal ubiquitination, as well as perturbations of neurofilaments and microtubules, occurred in the axonal spheroid-like swellings in vitro, and visualization of mitochondrial dynamics demonstrated that axonopathy resulted in impaired axonal transport. These data provide strong evidence for the involvement of non-neuronal cells in axonal dysfunction in ALS. This cell culture model may be of benefit for the development of therapeutic interventions directed at axonal preservation.

Ultramicroscopy Reveals Axonal Transport Impairments in Cortical Motor Neurons at Prion Disease

The functional imaging of neuronal circuits of the central nervous system is crucial for phenotype screenings or investigations of defects in neurodegenerative disorders. Current techniques yield either low penetration depth, yield poor resolution, or are restricted by the age of the animals. Here, we present a novel ultramicroscopy protocol for fluorescence imaging and three-dimensional reconstruction in the central nervous system of adult mice. In combination with tracing as a functional assay for axonal transport, retrogradely labeled descending motor neurons were visualized with >4 mm penetration depth. The analysis of the motor cortex shortly before the onset of clinical prion disease revealed that >80% neurons have functional impairments in axonal transport. Our study provides evidence that prion disease is associated with severe axonal transport defects in the cortical motor neurons and suggests a novel mechanism for prion-mediated neurodegeneration.

Impaired axonal transport and neurofilament compaction occur in separate populations of injured axons following diffuse brain injury in the immature rat

Diffuse brain injury is a leading cause of mortality in infants and children under 4 years of age and results in cognitive deficits in survivors. The anatomic basis for these behavioral deficits may be traumatic axonal injury (TAI), which manifests as impaired axonal transport (IAT) and neurofilament compaction (NFC), and may occur as a result of glutamate receptor activation. The extent of IAT and NFC was evaluated at 6, 24 and 72 h following non-contusive brain trauma in the 17 day-old rat to examine the causal relationship between these two pathologic entities; in addition, the effect of antagonists to the ionotropic glutamate receptors on TAI was evaluated. At 6 h post-injury, NFC was observed primarily in the cingulum, and appeared as swollen axons and terminal bulbs. By 24 h, swollen axons were additionally present in the corpus callosum and lateral white matter tracts, and appeared to increase in diameter. At 72 h, the extent of axonal swellings exhibiting compacted neurofilaments appeared to decrease, and was accompanied by punctate immunoreactivity within axon tracts suggestive of axonal degeneration. Although NFC was present in the same anatomical locations where axonal accumulation of amyloid precursor protein (APP) has been observed, double-label immunohistochemistry revealed no evidence of colocalization of compacted neurofilament and APP. Pre-injury treatment with either the NMDA receptor antagonist, ifenprodil, or the AMPA receptor antagonist, NBQX, had no significant effect on the extent of TAI, suggesting that excitotoxicity may not be a primary mechanism underlying TAI. Importantly, these data are indicative of the heterogeneity of mechanisms underlying TAI in the traumatically-injured immature brain.

Loss of Modifier of Cell Adhesion Reveals a Pathway Leading to Axonal Degeneration

Axonal dysfunction is the major phenotypic change in many neurodegenerative diseases, but the processes underlying this impairment are not clear. Modifier of cell adhesion (MOCA) is a presenilin binding protein that functions as a guanine nucleotide exchange factor for Rac1. The loss of MOCA in mice leads to axonal degeneration and causes sensorimotor impairments by decreasing cofilin phosphorylation and altering its upstream signaling partners LIM kinase and p21-activated kinase, an enzyme directly downstream of Rac1. The dystrophic axons found in MOCA-deficient mice are associated with abnormal aggregates of neurofilament protein, the disorganization of the axonal cytoskeleton, and the accumulation of autophagic vacuoles and polyubiquitinated proteins. Furthermore, MOCA deficiency causes an alteration in the actin cytoskeleton and the formation of cofilin-containing rod-like structures. The dystrophic axons show functional abnormalities, including impaired axonal transport. These findings demonstrate that MOCA is required for maintaining the functional integrity of axons and define a model for the steps leading to axonal degeneration.