SARM1 can be a potential therapeutic target for spinal cord injury.

Injury to the spinal cord is devastating. Studies have implicated Wallerian degeneration as the main cause of axonal destruction in the wake of spinal cord injury. Therefore, the suppression of Wallerian degeneration could be beneficial for spinal cord injury treatment. Sterile alpha and armadillo motif-containing protein 1 (SARM1) is a key modulator of Wallerian degeneration, and its impediment can improve spinal cord injury to a significant degree. In this report, we analyze the various signaling domains of SARM1, the recent findings on Wallerian degeneration and its relation to axonal insults, as well as its connection to SARM1, the mitogen-activated protein kinase (MAPK) signaling, and the survival factor, nicotinamide mononucleotide adenylyltransferase 2 (NMNAT2). We then elaborate on the possible role of SARM1 in spinal cord injury and explicate how its obstruction could potentially alleviate the injury.

Predictors of motor outcome after childhood arterial ischemic stroke.

AIM: To identify clinical and radiological predictors of long-term motor outcome after childhood-onset arterial ischemic stroke (AIS) in the middle cerebral artery (MCA) territory. METHOD: Medical records of 69 children (36 females, 33 males; median age at index AIS 3y 3mo, range: 1mo-16y) who presented to Great Ormond Street Hospital with first AIS in the MCA territory were reviewed retrospectively. Cases were categorized using the Childhood AIS Standardized Classification and Diagnostic Evaluation (CASCADE). Magnetic resonance imaging (MRI) and angiography were evaluated. An Alberta Stroke Program Early Computed Tomography Score (ASPECTS) was calculated on MRI. The Recurrence and Recovery Questionnaire assessed motor outcome and was dichotomized into good/poor. RESULTS: Eventual motor outcome was good in 49 children and poor in 20. There were no acute radiological predictors of eventual motor outcome. At follow-up, CASCADE 3A (i.e. moyamoya) and Wallerian degeneration were significantly associated with poor motor outcome. In the multivariate analysis, younger age and CASCADE 3A predicted poor motor outcome. INTERPRETATION: In the context of recommendations regarding unproven and potentially high-risk hyperacute therapies for childhood AIS, prediction of outcome could usefully contribute to risk/benefit analysis. Unfortunately, paradigms used in adults, such as ASPECTS, are not useful in children in the acute/early subacute phase of AIS. What this paper adds Adult paradigms, such as the Alberta Stroke Program Early Computed Tomography Score system, are not useful for predicting outcome in children. Younger children tend to have a poorer long-term prognosis than older children. Moyamoya is associated with poor prognosis.

Optimal conditions for graft survival and reinnervation of denervated muscles after embryonic motoneuron transplantation into peripheral nerves undergoing Wallerian degeneration.

Motoneuron transplantation into peripheral nerves undergoing Wallerian degeneration may have applications in treating diseases causing muscle paralysis. We investigated whether functional reinnervation of denervated muscle could be achieved by early or delayed transplantation after denervation. Adult rats were assigned to six groups with increasing denervation periods (0, 1, 4, 8, 12, and 24 weeks) before inoculation with culture medium containing (transplantation group) or lacking (surgical control group) dissociated embryonic motoneurons into the peroneal nerve. Electrophysiological and tissue analyses were performed 3 months after transplantation. Reinnervation of denervated muscles significantly increased relative muscle weight in the transplantation group compared with the surgical control group for denervation periods of 1 week (0.042% ± 0.0031% vs. 0.032% ± 0.0020%, respectively; p = 0.009), 4 weeks (0.044% ± 0.0069% vs. 0.026% ± 0.0045%, respectively; p = 0.0023), and 8 weeks (0.044% ± 0.0029% vs. 0.026% ± 0.0008%, respectively; p = 0.0023). The ratios of reinnervated muscle contractile forces to naïve muscle in the 0, 1, 4, 8, and 12 weeks transplantation groups were 3.79%, 18.99%, 8.05%, 6.30%, and 5.80%, respectively, indicating that these forces were sufficient for walking. The optimal implantation time for transplantation of motoneurons into the peripheral nerve was 1 week after nerve transection. However, the neurons transplanted 24 weeks after denervation survived and regenerated axons. These results indicated that there is time for preparing cells for transplantation in regenerative medicine and suggested that our method may be useful for paralysed muscles that are not expected to recover with current treatment.

Espinogénesis en motoneuronas de la médula espinal tras la lesión farmacológica de la corteza motora de ratas / Spinogenesis in spinal cord motor neurons following pharmacological lesions to the rat motor cortex

INTRODUCCIÓN: Diversas enfermedades neuropatologías asociadas a la degeneración del tracto corticoespinal muestran deterioro de las funciones motoras. Tales alteraciones neurológicas se asocian a diversos fenómenos plásticos subsecuentes, a nivel tanto presináptico como postsináptico. Sin embargo, no existe evidencia que indique la existencia de modificaciones en la transmisión de información del tracto corticoespinal a las motoneuronas espinales. MÉTODOS: Se indujo una lesión por vía estereotáxica en la corteza motora primaria de ratas hembra adultas con ácido kaínico y, 15 días después, se evaluó el desempeño motor mediante la escala BBB y en un dispositivo Rota-Rod. Paralelamente, se cuantificó la densidad numérica y proporcional de las espinas delgadas, en hongo y gordas, en motoneuronas de un segmento torácico-lumbar de la médula espinal. Así mismo, se registró la expresión de las proteínas espinofilina, sinaptofisina β III-tubulina. RESULTADOS: La lesión farmacológica provocó un desempeño motor deficiente. Así mismo, tanto la densidad de espinas como la proporción de espinas delgadas y gordas fue mayor, al igual que la expresión de las 3 proteínas estudiadas. CONCLUSIÓN: La aparición de los síntomas clínicos de daño neurológico provocado por la degeneración walleriana del tracto corticoespinal se acompaña de respuestas plásticas espontáneas de tipo compensador, a nivel sináptico. Lo anterior indica que durante la rehabilitación temprana de este tipo de pacientes, la plasticidad espontánea constituye un factor que se debe considerar para el diseño de estrategias de intervención más eficientes

INTRODUCTION: Motor function is impaired in multiple neurological diseases associated with corticospinal tract degeneration. Motor impairment has been linked to plastic changes at both the presynaptic and postsynaptic levels. However, there is no evidence of changes in information transmission from the cortex to spinal motor neurons. METHODS: We used kainic acid to induce stereotactic lesions to the primary motor cortex of female adult rats. Fifteen days later, we evaluated motor function with the BBB scale and the rotarod and determined the density of thin, stubby, and mushroom spines of motor neurons from a thoracolumbar segment of the spinal cord. Spinophilin, synaptophysin, and β III-tubulin expression was also measured. RESULTS: Pharmacological lesions resulted in poor motor performance. Spine density and the proportion of thin and stubby spines were greater. We also observed increased expression of the 3 proteins analysed. CONCLUSION: The clinical symptoms of neurological damage secondary to Wallerian degeneration of the corticospinal tract are associated with spontaneous, compensatory plastic changes at the synaptic level. Based on these findings, spontaneous plasticity is a factor to consider when designing more efficient strategies in the early phase of rehabilitation

Clinical Features, Risk Factors, and Early Prognosis for Wallerian Degeneration in the Descending Pyramidal Tract after Acute Cerebral Infarction.

BACKGROUND: Wallerian degeneration(WD) occurs in the descending pyramidal tract(DPT) after cerebral infarction commonly, but studies of its degree evaluation, influencing factors and effects on nervous function are still limited. OBJECTIVES: The purpose of this study was to describe these findings and estimate their clinical significance. METHODS: In total, 133 patients confirmed acute cerebral infarction and restricted diffusion in the DPT of the cerebral peduncle by MRI scans. These cases were retrospectively reviewed. We describe their clinical characteristics and analyze influence factors of WD, including the timespan from symptom onset to MRI and TOAST classification. Their NIHSS scores at admission and first 7 days NIHSS improvement rate after admission were also analyzed. RESULTS: These patients were divided into three groups by timespan ≤7 days(n = 45),7-14 days(n = 70) and >14 days(n = 18). The mean WD degree (%)of these three groups was 44.41 ± 22.51,52.35 ± 22.61and 44.31 ± 19.35,respectively(p = 0.122).According to the TOAST classification, the mean WD degree(%) of the cardioembolism group(n = 28, 62.80 ± 25.12) was significantly different from both the large-artery atherosclerosis group(n = 73,45.08 ± 20.03,p = 0.000) and the small-vessel occlusion group(n = 23,39.68 ± 16.95,p = 0.000). The mean NIHSS score upon admission of the WD degree≤50% group(n = 82,8.17 ± 5.87) was different from that of the >50% group(n = 51,11.31 ± 7.00)(p = 0.006). However, the mean 7 days NIHSS improvement rate(%) of the WD degree≤50% group(n = 79,11.83 ± 23.76)and >50% group(n = 50,13.40 ± 27.88) was not significantly different(p = 0.733). CONCLUSIONS: Early WD in ischemic stroke patients has a correlation with serious baseline functional defects. Therefore, we should give close attention to imaging change, especially in those with cardioembolism .

In vivo imaging of injured cortical axons reveals a rapid onset form of Wallerian degeneration.

BACKGROUND: Despite the widespread occurrence of axon and synaptic loss in the injured and diseased nervous system, the cellular and molecular mechanisms of these key degenerative processes remain incompletely understood. Wallerian degeneration (WD) is a tightly regulated form of axon loss after injury, which has been intensively studied in large myelinated fibre tracts of the spinal cord, optic nerve and peripheral nervous system (PNS). Fewer studies, however, have focused on WD in the complex neuronal circuits of the mammalian brain, and these were mainly based on conventional endpoint histological methods. Post-mortem analysis, however, cannot capture the exact sequence of events nor can it evaluate the influence of elaborated arborisation and synaptic architecture on the degeneration process, due to the non-synchronous and variable nature of WD across individual axons. RESULTS: To gain a comprehensive picture of the spatiotemporal dynamics and synaptic mechanisms of WD in the nervous system, we identify the factors that regulate WD within the mouse cerebral cortex. We combined single-axon-resolution multiphoton imaging with laser microsurgery through a cranial window and a fluorescent membrane reporter. Longitudinal imaging of > 150 individually injured excitatory cortical axons revealed a threshold length below which injured axons consistently underwent a rapid-onset form of WD (roWD). roWD started on average 20 times earlier and was executed 3 times slower than WD described in other regions of the nervous system. Cortical axon WD and roWD were dependent on synaptic density, but independent of axon complexity. Finally, pharmacological and genetic manipulations showed that a nicotinamide adenine dinucleotide (NAD+)-dependent pathway could delay cortical roWD independent of transcription in the damaged neurons, demonstrating further conservation of the molecular mechanisms controlling WD in different areas of the mammalian nervous system. CONCLUSIONS: Our data illustrate how in vivo time-lapse imaging can provide new insights into the spatiotemporal dynamics and synaptic mechanisms of axon loss and assess therapeutic interventions in the injured mammalian brain.

Regulation of degenerative spheroids after injury.

Neuronal injury leads to rapid, programmed disintegration of axons distal to the site of lesion. Much like other forms of axon degeneration (e.g. developmental pruning, toxic insult from neurodegenerative disorder), Wallerian degeneration associated with injury is preceded by spheroid formation along axons. The mechanisms by which injury leads to formation of spheroids and whether these spheroids have a functional role in degeneration remain elusive. Here, using neonatal mouse primary sympathetic neurons, we investigate the roles of players previously implicated in the progression of Wallerian degeneration in injury-induced spheroid formation. We find that intra-axonal calcium flux is accompanied by actin-Rho dependent growth of calcium rich axonal spheroids that eventually rupture, releasing material to the extracellular space prior to catastrophic axon degeneration. Importantly, after injury, Sarm1-/- and DR6-/-, but not Wlds (excess NAD+) neurons, are capable of forming spheroids that eventually rupture, releasing their contents to the extracellular space to promote degeneration. Supplementation of exogenous NAD+ or expressing WLDs suppresses Rho-dependent spheroid formation and degeneration in response to injury. Moreover, injured or trophically deprived Sarm1-/- and DR6-/-, but not Wlds neurons, are resistant to degeneration induced by conditioned media collected from wild-type axons after spheroid rupture. Taken together, these findings place Rho-actin and NAD+ upstream of spheroid formation and may suggest that other mediators of degeneration, such as DR6 and SARM1, mediate post-spheroid rupture events that lead to catastrophic axon disassembly.

Augustus Volney Waller: physiologist who described distal degeneration of severed nerve fibres.

Most readers will be familiar with the term 'Wallerian degeneration' - the degenerative changes that take place in nerve fibres distal to the site of their division. Few, I would respectfully suggest, would know very much about the discoverer of this phenomenon, Augustus Waller. This year marks the 200th anniversary of his death.

Axon Degeneration Assays in Superior Cervical Ganglion Explant Cultures.

The ability of peripheral nervous system neurons to extend long, axon-like neurites in vitro makes them ideally suited for studies on mechanisms of axon survival and degeneration. In this chapter, we describe how to prepare explant cultures of sympathetic neurons of the superior cervical ganglion (SCG). We also describe how to induce and assess axon degeneration with an injury or a chemical insult.

Assessing Axonal Degeneration in Embryonic Dorsal Root Ganglion Neurons In Vitro.

The molecular players regulating the axon degeneration pathway have been identified using in vitro experimental models. Here, we describe an in vitro assay to assess the axonal fragmentation induced by mechanical injury to axons in cultured mouse embryonic dorsal root ganglion (DRG) neurons. DRG neurons are pseudounipolar and therefore suitable for an assay of axonal degeneration after injury. In addition, the time course of the axonal fragmentation is stereotyped, enabling the identification of reagents that either expedite or impede the degeneration process. With an image-based quantification method, the in vitro degeneration assay can be utilized as a platform supporting high-throughput screens for pharmacological or genetic reagents delaying axon degeneration.

A Microfluidic Culture Platform to Assess Axon Degeneration.

The field of microfluidics allows for the precise spatial manipulation of small amounts of fluids. Within microstructures, laminar flow of fluids can be exploited to control the diffusion of small molecules, creating desired microenvironments for cells. Cellular neuroscience has benefited greatly from devices designed to fluidically isolate cell bodies and axons. Microfluidic devices specialized for neuron compartmentalization are made of polydimethylsiloxane (PDMS) which is gas permeable, is compatible with fluorescence microscopy, and has low cost. These devices are commonly used to study signals initiated exclusively on axons, somatodendritic compartments, or even single synapses. We have also found that microfluidic devices allow for rapid, reproducible interrogation of axon degeneration. Here, we describe the methodology for assessing axonal degeneration in microfluidic devices. We describe several use cases, including enucleation (removal of cell bodies) and trophic deprivation to investigate axon degeneration in pathological and developmental scenarios, respectively.

A Schwann Cell-Neuron Coculture System to Study Neuron-Glia Interaction During Axon Degeneration.

Autonomous mechanisms of axon degeneration are frequently studied in vitro by mechanical axon injury of isolated sensory neurons. This has led to major advances in understanding the molecular pathways governing axon degeneration. However, this approach does not pay attention to potential glial mechanisms for the regulation of axon death. Here, I describe a straightforward protocol to seed purified rat Schwann cells on neuronal cultures in order to study the interaction between axons and these glia during axon degeneration.

Organotypic Culture Assay for Neuromuscular Synaptic Degeneration and Function.

We describe here an organotypic culture system we have used to investigate mechanisms that maintain structure and function of axon terminals at the neuromuscular junction (NMJ). We developed this by taking advantage of the slow Wallerian degeneration phenotype in mutant Wlds mice, using these to compare preservation of NMJs with degeneration in nerve-muscle preparations from wild-type mice. We take hind limb tibial nerve/flexor digitorum brevis and lumbrical muscles and incubate them in mammalian physiological saline at 32 °C for 24-48 h. Integrity of NMJs can then be compared using a combination of electrophysiological and morphological techniques. We illustrate our method with data showing synaptic preservation ex vivo in nerve-muscle explants from Sarm-1 null-mutant mice. The ex vivo assays of NMJ integrity we describe here may therefore be useful for detailed investigation of synaptic maintenance and degeneration.

In Vivo Analysis of Glial Immune Responses to Axon Degeneration in Drosophila melanogaster.

Axon degeneration elicits a range of immune responses from local glial cells, including striking changes in glial gene expression, morphology, and phagocytic activity. Here, we describe a detailed set of protocols to assess discrete components of the glial reaction to axotomy in the adult nervous system of Drosophila melanogaster. These methods allow one to visualize and quantify transcriptional, morphological, and functional responses of glia to degenerating axons in a model system that is highly amenable to genetic manipulation.

Compound muscle action potential scan and MScanFit motor unit number estimation during Wallerian degeneration after nerve transections.

BACKGROUND: Compound muscle action potential (CMAP) scan and MScanFit have been used to understand the consequences of denervation and reinnervation. This study aimed to monitor these parameters during Wallerian degeneration (WD) after acute nerve transections (ANT). METHODS: Beginning after urgent surgery, CMAP scans were recorded at 1-2 day intervals in 12 patients with ANT of the ulnar or median nerves, by stimulating the distal stump (DS). Stimulus intensities (SI), steps, returners, and MScanFit were calculated. Studies were grouped according to the examination time after ANT. Results were compared with those of 27 controls. RESULTS: CMAP amplitudes and MScanFit progressively declined, revealing a positive correlation with one another. SIs were higher in WD groups than controls. Steps appeared or disappeared in follow-up scans. The late WD group had higher returner% than the early WD and control groups. CONCLUSIONS: MScanFit can monitor neuromuscular dysfunction during WD. SIs revealed excitability changes in DS.

Reduced inflammatory response and accelerated functional recovery following sciatic nerve crush lesion in CXCR3-deficient mice.

Despite the regenerative capacity of the peripheral nerve system (PNS), functional recovery after mechanical nerve trauma is often incomplete, resulting in motor, sensory, and autonomic deficits. The elucidation of key molecules involved in trauma-induced Wallerian degeneration and the ensuing regeneration processes is a prerequisite for the development of disease modifying drugs. The chemokine (C-X-C motif) receptor 3 (CXCR3) has been implicated in the recruitment of macrophages, the major immune cell population during the process of Wallerian degeneration. In this study, we examined whether deletion of CXCR3 affects macrophage recruitment, the expression of the proinflammatory cytokine tumor necrosis factor (TNF)- α and the CXCR3 agonist interferon gamma-induced protein 10 (CXCL10), and functional recovery in the sciatic nerve crush model. CXCR3 mice displayed significantly reduced macrophage counts preceded by diminished expression of CXCL10 and TNF- α. Furthermore, functional recovery of sciatic nerve motor function was significantly accelerated. In summary, these data indicate that the deletion of CXCR3 leads to a diminished inflammatory response and an accelerated functional recovery following sciatic nerve crush injury. Therefore, CXCR3 may be an interesting target for therapeutic interventions after traumatic nerve lesions.

Sleep Regulates Glial Plasticity and Expression of the Engulfment Receptor Draper Following Neural Injury.

Chronic sleep disturbance is associated with numerous health consequences, including neurodegenerative disease and cognitive decline [1]. Neurite damage due to apoptosis, trauma, or genetic factors is a common feature of aging, and clearance of damaged neurons is essential for maintenance of brain function. In the central nervous system, damaged neurites are cleared by Wallerian degeneration, in which activated microglia and macrophages engulf damaged neurons [2]. The fruit fly Drosophila melanogaster provides a powerful model for investigating the relationship between sleep and Wallerian degeneration [3]. Several lines of evidence suggest that glia influence sleep duration, sleep-mediated neuronal homeostasis, and clearance of toxic substances during sleep, raising the possibility that glial engulfment of damaged axons is regulated by sleep [4]. To explore this possibility, we axotomized olfactory receptor neurons and measured the effects of sleep loss or gain on the clearance of damaged neurites. Mechanical and genetic sleep deprivation impaired the clearance of damaged neurites. Conversely, treatment with the sleep-promoting drug gaboxadol accelerated clearance, while genetic induction of sleep promotes Draper expression. In sleep-deprived animals, multiple markers of glial activation were delayed, including activation of the JAK-STAT pathway, upregulation of the cell corpse engulfment receptor Draper, and innervation of the antennal lobe by glial membranes. These markers were all enhanced following genetic and pharmacological sleep induction. Taken together, these findings reveal a critical association between sleep and glial activation following neural injury, providing a platform for further investigations of the molecular mechanisms underlying sleep-dependent modulation of glial function and neurite clearance.

Magnetic resonance imaging evaluation of Wallerian degeneration of bilateral middle cerebellar peduncles after pontine infarction.

We aimed to present a case of symmetrical Wallerian degeneration (WD) in the middle cerebellar peduncles (MCPs) after a unilateral paramedian pontine infarction, which was examined by multimodality magnetic resonance imaging (MRI). In addition, we summarize the small number of reported cases. In our clinic, we observed a case of symmetrical WD of bilateral MCPs that occurred 6 months after the onset of a pontine infarction. We searched the Wanfang (Chinese) and PubMed databases and found 23 reported cases of this condition with characteristic similar to our patient. From the 24 cases, the detection time of WD ranged from 3 to 33 weeks. Symmetrical WD in the bilateral MCPs can occur after unilateral paramedian pontine infarction. Most cases were in Stages 2 and 3 of the disease and showed good clinical prognoses.

Axonal Degeneration Is Mediated by Necroptosis Activation.

Axonal degeneration, which contributes to functional impairment in several disorders of the nervous system, is an important target for neuroprotection. Several individual factors and subcellular events have been implicated in axonal degeneration, but researchers have so far been unable to identify an integrative signaling pathway activating this self-destructive process. Through pharmacological and genetic approaches, we tested whether necroptosis, a regulated cell-death mechanism implicated in the pathogenesis of several neurodegenerative diseases, is involved in axonal degeneration. Pharmacological inhibition of the necroptotic kinase RIPK1 using necrostatin-1 strongly delayed axonal degeneration in the peripheral nervous system and CNS of wild-type mice of either sex and protected in vitro sensory axons from degeneration after mechanical and toxic insults. These effects were also observed after genetic knock-down of RIPK3, a second key regulator of necroptosis, and the downstream effector MLKL (Mixed Lineage Kinase Domain-Like). RIPK1 inhibition prevented mitochondrial fragmentation in vitro and in vivo, a typical feature of necrotic death, and inhibition of mitochondrial fission by Mdivi also resulted in reduced axonal loss in damaged nerves. Furthermore, electrophysiological analysis demonstrated that inhibition of necroptosis delays not only the morphological degeneration of axons, but also the loss of their electrophysiological function after nerve injury. Activation of the necroptotic pathway early during injury-induced axonal degeneration was made evident by increased phosphorylation of the downstream effector MLKL. Our results demonstrate that axonal degeneration proceeds by necroptosis, thus defining a novel mechanistic framework in the axonal degenerative cascade for therapeutic interventions in a wide variety of conditions that lead to neuronal loss and functional impairment.SIGNIFICANCE STATEMENT We show that axonal degeneration triggered by diverse stimuli is mediated by the activation of the necroptotic programmed cell-death program by a cell-autonomous mechanism. This work represents a critical advance for the field since it identifies a defined degenerative pathway involved in axonal degeneration in both the peripheral nervous system and the CNS, a process that has been proposed as an early event in several neurodegenerative conditions and a major contributor to neuronal death. The identification of necroptosis as a key mechanism for axonal degeneration is an important step toward the development of novel therapeutic strategies for nervous-system disorders, particularly those related to chemotherapy-induced peripheral neuropathies or CNS diseases in which axonal degeneration is a common factor.