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All neural processes in the human brain take place in a dense matrix of thin fibers (axons) that release serotonin, an ancient neurotransmitter that supports the plasticity of neural tissue and has profound implications for mental health. The self-organization of the serotonergic matrix is not well understood, despite recent advances in experimental and theoretical approaches. Previous work in our laboratory has demonstrated that individual serotonergic axons produce highly stochastic trajectories, fundamental to the construction of regional fiber densities, but further advances in predictive computer simulations require more accurate experimental information. However, visualizing this dynamic behavior in vivo is currently extremely difficult. Recent technological advances (e.g., digital holotomography, three-dimensional (3D)-cell culture systems, microphysiological systems) have changed the way complex neuronal systems can be studied and offer promising alternatives to in vivo approaches. This research examined single serotonergic axons in culture systems (co-cultures and mono-cultures), by using a set of complementary high-resolution methods: confocal microscopy, holotomography (refractive index-based live imaging), and super-resolution (STED) microscopy. Several 3D-hydrogel systems were additionally tested. This research shows that serotonergic axon walks in neural tissue may strongly reflect the stochastic geometry of the tissue and it also provides new insights into the morphology and branching properties of serotonergic axons. The proposed experimental platform can support next-generation analyses of the serotonergic matrix, including seamless integration with computational approaches.

Irreversible damage to the structure of axons and death of the retinal ganglion cell (RGC) soma in primary open-angle glaucoma (POAG) and Alzheimer’s disease (AD) develop against the background of the already existing clinical manifestation, which is preceded by a slow period of progressive loss of synapses and dendrites of the RGCs. Recent studies have shown that the integrity of the RGC’s dendritic branching can serve as both a target of neuroprotective therapy and a sensitive marker of retinal degeneration in AD and glaucoma. To develop methods of complex neuroprotective therapy, it is necessary to substantiate the targets and tactics of affecting the dendritic tree of the RGCs, the remodeling of which, according to modern concepts, can be closely and antagonistically related to the regeneration of the axon after its damage in trauma and neurodegenerative diseases. RGCs are highly capable of functional modification. Currently, it has been proven that the use of neuroprotective drugs and neurotrophins is promising for maintaining the adaptive plasticity of RGCs and restoring their synaptic contacts at the level of the retina and brain. Understanding the features of the adaptive plasticity of RGCs in AD and glaucoma will make it possible to use technologies to activate the internal potential of neuronal remodeling, including the modification of dendritic branching of RGCs and regeneration of their axons, in the preclinical stages of these diseases. Increasing knowledge about the sequence and mechanisms of early events in the retina’s inner plexiform layer will contribute to the development of targeted neuroprotective therapy and new technologies to detect early POAG, AD, and, possibly, other systemic and local neurodegenerative conditions.

Movement and posture depend on sensory feedback that is regulated by specialized GABAergic neurons (GAD2+) that form axo-axonic contacts onto myelinated proprioceptive sensory axons and are thought to be inhibitory. However, we report here that activating GAD2+ neurons, directly with optogenetics or indirectly by cutaneous stimulation, facilitates sensory feedback to motoneurons in awake rodents and humans. GABAA receptors and GAD2+ innervation at or near nodes of Ranvier of sensory axons cause this facilitation, preventing spike propagation failure at the many axon branch points, which is otherwise common without GABA. In contrast, GABAA receptors are generally lacking from axon terminals (unlike GABAB) and do not presynaptically inhibit transmitter release onto motoneurons. GABAergic innervation near nodes and branch points allows individual branches to function autonomously, with GAD2+ neurons regulating which branches conduct, adding a computational layer to the neuronal networks generating movement and likely generalizing to other CNS axons.

Voltage-gated calcium (Cav) channels are a prerequisite for signal transmission at the first olfactory sensory neuron (OSN) synapse within the glomeruli of the main olfactory bulb (MOB). We showed previously that the N-type Cav channel subunit Cav2.2 is present in the vast majority of glomeruli and plays a central role in presynaptic transmitter release. Here, we identify a distinct subset of glomeruli in the MOB of adult mice that is characterized by expression of the P/Q-type channel subunit Cav2.1. Immunolocalization shows that Cav2.1+ glomeruli reside predominantly in the medial and dorsal MOB, and in the vicinity of the necklace glomerular region close to the accessory olfactory bulb. Few glomeruli are detected on the ventral and lateral MOB. Cav2.1 labeling in glomeruli colocalizes with the presynaptic marker vGlut2 in the axon terminals of OSNs. Electron microscopy shows that Cav2.1+ presynaptic boutons establish characteristic asymmetrical synapses with the dendrites of second-order neurons in the glomerular neuropil. Cav2.1+ glomeruli receive axonal input from OSNs that express molecules of canonical OSNs: olfactory marker protein, the ion channel Cnga2, and the phosphodiesterase Pde4a. In the main olfactory epithelium, Cav2.1 labels a distinct subpopulation of OSNs whose distribution mirrors the topography of the MOB glomeruli, that shows the same molecular signature, and is already present at birth. Together, these experiments identify a unique Cav2.1+ multiglomerular domain in the MOB that may form a previously unrecognized olfactory subsystem distinct from other groups of necklace glomeruli that rely on cGMP signaling mechanisms.

Many oligodendrocyte precursor cells (OPCs) do not differentiate to form myelin, suggesting additional roles of this cell population. The zebrafish optic tectum contains OPCs in regions devoid of myelin. Elimination of these OPCs impaired precise control of retinal ganglion cell axon arbor size during formation and maturation of retinotectal connectivity, and in consequence impaired visual acuity. Therefore, OPCs fine-tune neural circuit structure and function independently of their canonical role to make myelin.Competing Interest StatementThe authors have declared no competing interest.

This study was performed to investigate the effect of platelet-rich fibrin membrane (PRFM), alone and with topical tacrolimus application, on regeneration of the crushed facial nerve (FN). Thirty healthy 7-week-old albino rats were used. The left FN was damaged by crushing in all rats. Three random groups of rats were formed: group 1, untreated; group 2, treated with PRFM; group 3, treated with PRFM plus topical tacrolimus. Functional recovery and histological and immunohistochemical evaluations were performed 4 and 8 weeks later. Anti-S100 was used to detect myelin sheath. At 4 weeks, blinking reflex recovery was more rapid in group 3 than in groups 2 and 1 (4.30 ± 0.48, 3.40 ± 0.52, and 2.20 ± 0.42, respectively); the difference was statistically significant (P = 0.001). Histologically, group 3 showed more apparent normal FN structures than the other groups. Immunohistochemical caspase-3 evaluation of the axon area revealed a significant difference between group 2 (PRFM alone; 8.67 ± 0.029) and group 3 (PRFM plus topical tacrolimus; 4.42 ± 0.028) (P = 0.001). Group 3 showed the greatest positive staining in the myelin sheath. Based on the results of this animal study, clinical studies should be performed to determine whether the combination of PRF and tacrolimus also improves the outcome of nerve regeneration in humans.

Chemical communication controls a wide range of behaviors via conserved signaling networks. Axon regeneration in response to injury is determined by the interaction between the extracellular environment and intrinsic growth potential. In this study, we investigated the role of chemical signaling in axon regeneration inCaenorhabditis elegans. We find that the enzymes involved in ascaroside pheromone biosynthesis, ACOX-1.1, ACOX-1.2, and DAF-22, participate in axon regeneration by producing a dauer-inducing ascaroside, ascr#5. We demonstrate that the chemoreceptor genes,srg-36andsrg-37, which encode G-protein-coupled receptors for ascr#5, are required for adult-specific axon regeneration. Furthermore, the activating mutation inegl-30encoding Gqα suppresses axon regeneration defective phenotype inacox-1.1andsrg-36 srg-37mutants. Therefore, the ascaroside signaling system provides a unique example of a signaling molecule that regulates the regenerative pathway in the nervous system.SIGNIFICANCE STATEMENTInCaenorhabditis elegans, axon regeneration is positively regulated by the EGL-30 Gqα–JNK MAP kinase cascade. However, it remains unclear what signals activate the EGL-30 pathway in axon regeneration. Here, we show that SRG-36 and SRG-37 act as upstream G-protein-coupled receptors (GPCRs) that activate EGL-30.C. eleganssecretes a family of small-molecule pheromones called ascarosides, which serve various functions in chemical signaling. SRG-36 and SRG-37 are GPCRs for the dauer-inducing ascaroside ascr#5. Consistent with this, we found that ascr#5 activates the axon regeneration pathway via SRG-36/SRG-37 and EGL-30. Thus, ascaroside signaling promotes axon regeneration by activating the GPCR–Gqα pathway.

Olfactory ensheathing cells (OECs) from the olfactory bulb (OB) and the olfactory mucosa (OM) have the capacity to repair nerve injury. However, the difference in the therapeutic effect between OB-derived OECs and OM-derived OECs remains unclear. In this study, we extracted OECs from OB and OM and compared the gene and protein expression profiles of the cells using transcriptomics and non-quantitative proteomics techniques. The results revealed that both OB-derived OECs and OM-derived OECs highly expressed genes and proteins that regulate cell growth, proliferation, apoptosis and vascular endothelial cell regeneration. The differentially expressed genes and proteins of OB-derived OECs play a key role in regulation of nerve regeneration and axon regeneration and extension, transmission of nerve impulses and response to axon injury. The differentially expressed genes and proteins of OM-derived OECs mainly participate in the positive regulation of inflammatory response, defense response, cytokine binding, cell migration and wound healing. These findings suggest that differentially expressed genes and proteins may explain why OB-derived OECs and OM-derived OECs exhibit different therapeutic roles. This study was approved by the Animal Ethics Committee of the General Hospital of Ningxia Medical University (approval No. 2017-073) on February 13, 2017.

Mitochondria play a multidimensional role in the function and the vitality of the neurological system. From the generation of neural stem cells to the maintenance of neurons and their ultimate demise, mitochondria play a critical role in regulating our neural pathways’ homeostasis, a task that is critical to our cognitive health and neurological well-being. Mitochondria provide energy via oxidative phosphorylation for the neurotransmission and generation of an action potential along the neuron’s axon. This paper will first review and examine the molecular subtleties of the mitochondria’s role in neurogenesis and neuron vitality, as well as outlining the impact of defective mitochondria in neural aging. The authors will then summarize neurodegenerative diseases related to either neurogenesis or homeostatic dysfunction. Because of the significant detriment neurodegenerative diseases have on the quality of life, it is essential to understand their etiology and ongoing molecular mechanics. The mitochondrial role in neurogenesis and neuron vitality is essential. Dissecting and understanding this organelle’s role in the genesis and homeostasis of neurons should assist in finding pharmaceutical targets for neurodegenerative diseases.

Peripheral nerve injuries commonly occur due to trauma, like a traffic accident. Peripheral nerves get severed, causing motor neuron death and potential muscle atrophy. The current golden standard to treat peripheral nerve lesions, especially lesions with large (≥ 3 cm) nerve gaps, is the use of a nerve autograft or reimplantation in cases where nerve root avulsions occur. If not tended early, degeneration of motor neurons and loss of axon regeneration can occur, leading to loss of function. Although surgical procedures exist, patients often do not fully recover, and quality of life deteriorates. Peripheral nerves have limited regeneration, and it is usually mediated by Schwann cells and neurotrophic factors, like glial cell line-derived neurotrophic factor, as seen in Wallerian degeneration. Glial cell line-derived neurotrophic factor is a neurotrophic factor known to promote motor neuron survival and neurite outgrowth. Glial cell line-derived neurotrophic factor is upregulated in different forms of nerve injuries like axotomy, sciatic nerve crush, and compression, thus creating great interest to explore this protein as a potential treatment for peripheral nerve injuries. Exogenous glial cell line-derived neurotrophic factor has shown positive effects in regeneration and functional recovery when applied in experimental models of peripheral nerve injuries. In this review, we discuss the mechanism of repair provided by Schwann cells and upregulation of glial cell line-derived neurotrophic factor, the latest findings on the effects of glial cell line-derived neurotrophic factor in different types of peripheral nerve injuries, delivery systems, and complementary treatments (electrical muscle stimulation and exercise). Understanding and overcoming the challenges of proper timing and glial cell line-derived neurotrophic factor delivery is paramount to creating novel treatments to tend to peripheral nerve injuries to improve patients' quality of life.

Vocal learning in songbirds is mediated by cortico-basal ganglia circuits that govern diverse functions during different stages of development. We investigated developmental changes in axonal projections to and from motor cortical regions that underlie learned vocal behavior in juvenile zebra finches (Taeniopygia guttata). Neurons in LMAN-core project to RA, a motor cortical region that drives vocal output; these RA-projecting neurons send a transient collateral projection to AId, a region adjacent to RA, during early vocal development. Both RA and AId project to a region of dorsal thalamus (DLM), which forms a feedback pathway to cortico-basal ganglia circuitry. These projections provide pathways conveying efference copy and a means by which information about vocal motor output could be reintegrated into cortico-basal ganglia circuitry, potentially aiding in the refinement of juvenile vocalizations during learning. We used tract-tracing techniques to label the projections of LMAN-core to AId and of RA to DLM in juvenile songbirds. The volume and density of terminal label in the LMAN-core→AId projection declined substantially during early stages of sensorimotor learning. In contrast, the RA→DLM projection showed no developmental change. The retraction of LMAN-core→AId axon collaterals indicates a loss of efference copy to AId and suggests that projections that are present only during early stages of sensorimotor learning mediate unique, temporally restricted processes of goal-directed learning. Conversely, the persistence of the RA→DLM projection may serve to convey motor information forward to the thalamus to facilitate song production during both learning and maintenance of vocalizations. This article is protected by copyright. All rights reserved.

Neuronal remodeling after brain injury is essential for functional recovery. After unilateral cortical lesion, axons from the intact cortex ectopically project to the denervated midbrain, but the molecular mechanisms remain largely unknown. To address this issue, we examined gene expression profiles in denervated and intact mouse midbrains after hemispherectomy at early developmental stages using mice of either sex, when ectopic contralateral projection occurs robustly. The analysis showed that various axon growth-related genes were upregulated in the denervated midbrain, and most of these genes are reportedly expressed by glial cells. To identify the underlying molecules, the receptors for candidate upregulated molecules were knocked out in layer 5 projection neurons in the intact cortex, using the CRISPR/Cas9-mediated method, and axonal projection from the knocked-out cortical neurons was examined after hemispherectomy. We found that the ectopic projection was significantly reduced when integrin subunit β three or neurotrophic receptor tyrosine kinase 2 (also known as TrkB) was knocked out. Overall, the present study suggests that denervated midbrain-derived glial factors contribute to lesion-induced remodeling of the cortico-mesencephalic projection via these receptors.SIGNIFICANCE STATEMENTAfter brain injury, compensatory neural circuits are established that contribute to functional recovery. However, little is known about the intrinsic mechanism that underlies the injury-induced remodeling. We found that after unilateral cortical ablation expression of axon-growth promoting factors is elevated in the denervated midbrain and is involved in the formation of ectopic axonal projection from the intact cortex. Evidence further demonstrated that these factors are expressed by astrocytes and microglia, which are activated in the denervated midbrain. Thus, our present study provides a new insight into the mechanism of lesion-induced axonal remodeling and further therapeutic strategies after brain injury.

Hh signaling has been shown to be activated in intact and injured peripheral nerve. However, the role of Hh signaling in peripheral nerve is not fully understood. In the present study, we observed that Hh signaling responsive cells [Gli1(+) cells] in both the perineurium and endoneurium. In the endoneurium, Gli1(+) cells were classified as blood vessel associated or non-associated. After injury, Gli1(+) cells around blood vessels mainly proliferated to then accumulate into the injury site along with endothelial cells. Hh signaling activity was retained in Gli1(+) cells during nerve regeneration. To understand the role of Hedgehog signaling in Gli1(+) cells during nerve regeneration, we examined mice with Gli1(+) cells-specific inactivation of Hh signaling (Smo cKO). After injury, Smo cKO mice showed significantly reduced numbers of accumulated Gli1(+) cells along with disorganized vascularization at an early stage of nerve regeneration, which subsequently led to an abnormal extension of the axon. Thus, Hh signaling in Gli1(+) cells appears to be involved in nerve regeneration through controlling new blood vessel formation at an early stage.

Neurons in the inferior olive nucleus, the sole origin of cerebellar climbing fibers, project their axons to the cerebellum through the olivocerebellar tract. A single olivocerebellar axon gives rise to multiple climbing fibers (about seven in rat) which typically terminate into longitudinal compartments in the cerebellar cortex. These compartments match with the longitudinal striped arrangement of aldolase C-positive and -negative Purkinje cell subsets. As a result of this topographic arrangement, the olivocerebellar projection relays the synchronous activity of the electrically coupled adjacent inferior olive neurons to complex spike firing of Purkinje cells in a narrow longitudinal stripe. Olivocerebellar axons show a dynamic morphogenetic process. An immature axon has abundant terminal branches that innervate multiple Purkinje cells. Several terminal branches (climbing fibers) grows to eventually establishing a powerful one-to-one synaptic connection between a single climbing fiber terminal and a single target Purkinje cell. Furthermore, these axons are capable of strong compensatory re-innervation after lesion even in adult.

BACKGROUND: The role of neurotrophic factors (brain-derived neurotrophic factors and glial cell line-derived neurotrophic factors) and early electrical stimulation (EES) in the injured nerve has found promising in several studies. However, there is still limited knowledge about the effect of EES in the distal part of the nerve to sustain this level of expression of growth factors. AIM: We aim to evaluate the effects of EES in in neural regeneration by measuring the expression of brain-derived neurotrophic factor (BDNF) and glial cell line-derived neurotrophic factor (GDNF) in animal model. METHODS: The research was conducted starting from April to May 2021 using male Wistar rats. Using general anesthesia, the sciatic nerve was cut. The intervention group was treated with EES in the distal stump, right after nerve resection (20 Hz, 1–2 mA, 2–5 s), while the control group received no treatment after nerve resection. A reoperation on day 3 was performed in both groups to measure BDNF and GDNF expression level of the distal nerve tissue by ELISA as well as histopathological examination of sprouting axons of the injured proximal nerve. RESULTS: A total of 32 samples were included in the study. A statistically significant levels of GDNF is found higher in the EES group (n = 16) than the control group (n = 16) (35. 71 pg/100 mg, confidence interval (CI) 95% 23.93, 47.48, p < 0.05). The number of sprouting axons is found lower in the EES group (p < 0.05). The BDNF level is similar between the two groups, however not significant. After a subgroup analysis, it was found that the greater the level of GDNF, the fewer the axon sprouts in both groups (fewer axon group 58.35 [n = 22, CI 95% 45.14, 71.55] vs. more axon group 47.14 [n = 10, CI 95% 35.33, 58.95]), p < 0.05. CONCLUSION: The EES proves its benefit in accelerating the axonal regeneration by increasing the expression GDNF in the distal nerve stumps in the electrical excited degenerated sciatic nerve in the rat model.

Axon regeneration after spinal cord injury (SCI) is limited by both a decreased intrinsic ability of neurons to grow axons and the growth-hindering effects of extrinsic inhibitory molecules expressed around the lesion. Deletion ofphosphatase and tensin homolog(Pten) augments mechanistic target of rapamycin (mTOR) signaling and enhances the intrinsic regenerative response of injured corticospinal neurons after SCI. Because of the variety of growth-restrictive extrinsic molecules, it remains unclear how inhibition of conserved inhibitory signaling elements would affect axon regeneration and rewiring after SCI. Moreover, it remains unknown how a combinatorial approach to modulate both extrinsic and intrinsic mechanisms can enhance regeneration and rewiring after SCI. In the present study, we deletedRhoAandRhoC, which encode small GTPases that mediate growth inhibition signals of a variety of extrinsic molecules, to remove global extrinsic pathways.RhoA/RhoCdouble deletion in mice suppressed retraction or dieback of corticospinal axons after SCI. In contrast,Ptendeletion increased regrowth of corticospinal axons into the lesion core. Although deletion of bothRhoAandPtendid not promote axon regrowth across the lesion or motor recovery, it additively promoted rewiring of corticospinal circuits connecting the cerebral cortex, spinal cord, and hindlimb muscles. Our genetic findings, therefore, reveal that a combinatorial approach to modulate both intrinsic and extrinsic factors can additively promote neural circuit rewiring after SCI.SIGNIFICANCE STATEMENTSCI often causes severe motor deficits because of damage to the corticospinal tract (CST), the major neural pathway for voluntary movements. Regeneration of CST axons is required to reconstruct motor circuits and restore functions; however, a lower intrinsic ability to grow axons and extrinsic inhibitory molecules severely limit axon regeneration in the CNS. Here, we investigated whether suppression of extrinsic inhibitory cues by genetic deletion ofRhoas well as enhancement of the intrinsic pathway by deletion ofPtencould enable axon regrowth and rewiring of the CST after SCI. We show that simultaneous elimination of extrinsic and intrinsic signaling pathways can additively promote axon sprouting and rewiring of the corticospinal circuits. Our data demonstrate a potential molecular approach to reconstruct motor pathways after SCI.

Many models of motor control emphasize the role of sensorimotor cortex in movement, principally through the projections that corticospinal neurons (CSNs) make to the spinal cord. Additionally, CSNs possess expansive supraspinal axon collaterals, the functional organization of which is largely unknown. Using anatomical and electrophysiological circuit-mapping techniques in the mouse, we reveal dorsolateral striatum as the preeminent target of CSN collateral innervation. We found that this innervation is biased so that CSNs targeting different striatal pathways show biased targeting of spinal cord circuits. Contrary to more conventional perspectives, CSNs encode not only individual movements, but also information related to the onset and offset of motor sequences. Furthermore, similar activity patterns are broadcast by CSN populations targeting different striatal circuits. Our results reveal a logic of coordinated connectivity between forebrain and spinal circuits, where separate CSN modules broadcast similarly complex information to downstream circuits, suggesting that differences in postsynaptic connectivity dictate motor specificity. The authors detail principles underlying the innervation of spinal and striatal circuits by populations of corticospinal neurons, and characterize the behavioral information broadcast through this motor control network.

The corpus callosum (CC) is a major interhemispheric commissure of placental mammals. Early steps of CC formation rely on guidance strategies, such as axonal branching and collateralization. Here we analyze the time-course dynamics of axonal bifurcation during typical cortical development or in a CC dysgenesis mouse model. We use Swiss mice as a typical CC mouse model and find that axonal bifurcation rates rise in the cerebral cortex from embryonic day (E)17 and are reduced by postnatal day (P)9. Since callosal neurons populate deep and superficial cortical layers, we compare the axon bifurcation ratio between those neurons by electroporating ex vivo brains at E13 and E15, using eGFP reporter to label the newborn neurons on organotypic slices. Our results suggest that deep layer neurons bifurcate 32% more than superficial ones. To investigate axonal bifurcation in CC dysgenesis, we use BALB/c mice as a spontaneous CC dysgenesis model. BALB/c mice present a typical layer distribution of SATB2 callosal cells, despite the occurrence of callosal anomalies. However, using anterograde DiI tracing, we find that BALB/c mice display increased rates of axonal bifurcations during early and late cortical development in the medial frontal cortex. Midline guidepost cells adjacent to the medial frontal cortex are significant reduced in the CC dysgenesis mouse model. Altogether these data suggest that callosal collateral axonal exuberance is maintained in the absence of midline guidepost signaling and might facilitate aberrant connections in the CC dysgenesis mouse model.

Much research has focused on the PI3-kinase and PTEN signaling pathway with the aim to stimulate repair of the injured central nervous system. Axons in the central nervous system fail to regenerate, meaning that injuries or diseases that cause loss of axonal connectivity have life-changing consequences. In 2008, genetic deletion of PTEN was identified as a means of stimulating robust regeneration in the optic nerve. PTEN is a phosphatase that opposes the actions of PI3-kinase, a family of enzymes that function to generate the membrane phospholipid PIP3 from PIP2 (phosphatidylinositol (3,4,5)-trisphosphate from phosphatidylinositol (4,5)-bisphosphate). Deletion of PTEN therefore allows elevated signaling downstream of PI3-kinase, and was initially demonstrated to promote axon regeneration by signaling through mTOR. More recently, additional mechanisms have been identified that contribute to the neuron-intrinsic control of regenerative ability. This review describes neuronal signaling pathways downstream of PI3-kinase and PIP3, and considers them in relation to both developmental and regenerative axon growth. We briefly discuss the key neuron-intrinsic mechanisms that govern regenerative ability, and describe how these are affected by signaling through PI3-kinase. We highlight the recent finding of a developmental decline in the generation of PIP3 as a key reason for regenerative failure, and summarize the studies that target an increase in signaling downstream of PI3-kinase to facilitate regeneration in the adult central nervous system. Finally, we discuss obstacles that remain to be overcome in order to generate a robust strategy for repairing the injured central nervous system through manipulation of PI3-kinase signaling.

Many species of lizards are capable of tail regeneration. There has been increased interest in the study of lizard tail regeneration in recent years as it is an amenable regeneration model for amniotes. In this study, Gekko japonicus was used as a model to investigate the initiation of vascularization, innervation and myogenesis during tail regeneration. We found that angiogenesis and axon regeneration occurred almost simultaneously within 4 days post amputation. The results showed that the endothelial cells of the original vasculature proliferated and extended into the blastema as capillary vessels, which inter-connected to form a capillary network. The nerve fibers innervated the regenerated tissue from the original spinal cord and dorsal root ganglia, and the fiber bundles increased during 14 days. Regenerating muscle tissues emerged 2 weeks after amputation. PAX3 and PAX7 expression were detected during myogenesis, with PAX7 showing a continuous increase in expression from day 3 until the day 14, whereas PAX3 reached a peak level on day 10 day post amputation, and then declined quickly to level as normal control on day 14. PCNA and PAX3 double-positive satellite cells were observed in the original rostral tissues, indicating the involvement of satellite cell proliferation during tail regeneration. Taken together, these data suggest that tail regeneration in Gekko japonicus involved rapid angiogenesis from the beginning to the day 10 and followed by capillary remodeling. The innervation of regenerated tail was significant on day 4 and increased gradually during regeneration, while the regenerated muscle tissues was obvious on day 14 after amputation.

Parkinson's disease (PD) is associated with dopamine depletion in the striatum owing to the selective and progressive loss of the nigrostriatal dopaminergic neurons, which results in motor dysfunction and secondary clinical manifestations. The dopamine level in the striatum is preserved because of the innervation of the substantia nigra (SN) dopaminergic neurons into it. Therefore, protection of the SN neurons is crucial for maintaining the dopamine level in the striatum and for ensuring the desired motor coordination. Several strategies have been devised to protect the degenerating dopaminergic neurons or to restore the dopamine levels for treating PD. Most of the methods focus exclusively on preventing cell body death in the neurons. Although advances have been made in understanding the disease, the search for disease-modifying drugs is an ongoing process. The present review describes the evidence from studies involving patients with PD as well as PD models that axon terminals are highly vulnerable to exogenous and endogenous insults and degenerate at the early stage of the disease. Impairment of mitochondrial dynamics, Ca2+ homeostasis, axonal transport, and loss of plasticity of axon terminals appear before the neuronal degeneration in PD. Furthermore, distortion of synaptic morphology and reduction of postsynaptic dendritic spines are the neuropathological hallmarks of early-stage disease. Thus, the review proposes a shift in focus from discerning the mechanism of neuronal cell body loss and targeting it to an entirely different approach of preventing axonal degeneration. The review also suggests appropriate strategies to prevent the loss of synaptic terminals, which could induce regrowth of the axon and its auxiliary fibers and might offer relief from the symptomatic features of PD.

Anesthetics are commonly used in various medical procedures. Accumulating evidence suggests that early-life anesthetics exposure in infants and children affects brain development, causing psychiatric and neurological disorders. However, the underlying mechanisms are poorly understood. Using zebrafish larvae as a model, we found that the proliferation and migration of oligodendrocyte progenitor cells (OPCs) were severely impaired by the exposure of midazolam (MDZ), an anesthetic widely used in pediatric surgery and intensive care medicine, leading to a reduction of oligodendroglial lineage cell in the dorsal spinal cord. This defect was mimicked by the bath application of translocator protein (TSPO) agonists and partially rescued by genetic downregulation of TSPO. Cell transplantation experiments showed that requirement of TSPO for MDZ-induced oligodendroglial lineage cell defects is cell-autonomous. Furthermore, transmission electron microscopy and in vivo electrophysiological recording experiments demonstrated that MDZ exposure caused axon hypomyelination and action potential propagation retardation, resulting in delayed behavior initiation. Thus, our findings reveal that MDZ affects oligodendroglial lineage cell development and myelination in young animals, raising the care about its clinic use in infants and children.

The ischemia and hypoxia microenvironment after spinal cord injury (SCI) makes SCI repair a challenging problem. With various stimulus, chances for neural stem cells (NSCs) to differentiate into neurons, astrocytes, oligodendrocytes are great and is considered as a potential source of the stem cell therapy to SCI. Our research used adeno-associated virus (AAV) to carry the target gene to transfect neural stem cells. Transfected NSCs can express nerve growth factor (NGF) navigated by five hypoxia-responsive elements (5HRE). Therefore, the 5HRE-NGF-NSCs could express NGF specifically in hypoxia sites to promote the tissue repair and function recovery. Based on the regeneration of neurocytes and promotion of the recovery found in SCI models, via locomotor assessment, histochemical staining and molecular examinations, our results demonstrated that 5HRE-NGF-NSCs could improve the motor function, neurons survival and molecules expression of SCI rats. Meanwhile, the downregulated expression of autophagy-related proteins indicated the inhibitive effect of 5HRE-NGF-NSCs on autophagy. Our research showed that 5HRE-NGF-NSCs contribute to SCI repair which might via inhibiting autophagy and improving the survival rate of neuronal cells. The new therapy also hampered the hyperplasia of neural glial scars and induced axon regeneration. These positive functions of 5HRE-NGF-NSCs all indicate a promising SCI treatment.

Wallerian degeneration is a pathological process closely related to peripheral nerve regeneration following injury, and includes the disintegration and phagocytosis of peripheral nervous system cells. Traditionally, morphological changes are observed by performing immunofluorescence staining after sectioning, which results in the loss of some histological information. The purpose of this study was to explore a new, nondestructive, and systematic method for observing axonal histological changes during Wallerian degeneration. Thirty male Thy1-YFP-16 mice (SPF grade, 6 weeks old, 20±5 g) were randomly selected and divided into clear, unobstructed brain imaging cocktails and computational analysis (CUBIC) optical clearing (n=15) and traditional method groups (n=15). Five mice in each group were sacrificed at 1st, 3rd, and 5th day following a crush operation. The histological axon changes were observed by CUBIC light optical clearing treatment, direct tissue section imaging, and HE staining. The results revealed that, compared with traditional imaging methods, there was no physical damage to the samples, which allowed for three-dimensional and deep-seated tissue imaging through CUBIC. Local image information could be nicely obtained by direct fluorescence imaging and HE staining, but it was difficult to obtain image information of the entire sample. At the same time, the image information obtained by fluorescence imaging and HE staining was partially lost. The combining of CUBIC and Thy1-YFP transgenic mice allowed for a clear and comprehensive observation of histological changes of axons in Wallerian degeneration.

During cortical development and throughout adulthood, oligodendrocytes add myelin internodes to glutamatergic projection neurons and GABAergic inhibitory neurons. In addition to directing node of Ranvier formation, to enable saltatory conduction and influence action potential transit time, oligodendrocytes support axon health by communicating with axons via the periaxonal space and providing metabolic support that is particularly critical for healthy ageing. In this review we outline the timing of oligodendrogenesis in the developing mouse and human cortex and describe the important role that oligodendrocytes play in sustaining and modulating neuronal function. We also provide insight into the known and speculative impact that myelination has on cortical axons and their associated circuits during the developmental critical periods and throughout life, particularly highlighting their life-long role in learning and remembering.

Dorsal root ganglion (DRG) neurons are the predominant cell type that innervates the vertebrate skin. They are typically described as pseudounipolar cells that have central and peripheral axons branching from a single root exiting the cell body. The peripheral axon travels within a nerve to the skin, where free sensory endings can emerge and branch into an arbor that receives and integrates information. In some immature vertebrates, DRG neurons are preceded by Rohon-Beard (RB) neurons. While the sensory endings of RB and DRG neurons function like dendrites, we use live imaging in zebrafish to show that they have axonal plus-end-out microtubule polarity at all stages of maturity. Moreover, we show both cell types have central and peripheral axons with plus-end-out polarity. Surprisingly, in DRG neurons these emerge separately from the cell body, and most cells never acquire the signature pseudounipolar morphology. Like another recently characterized cell type that has multiple plus-end-out neurites, ganglion cells in Nematostella, RB and DRG neurons maintain a somatic microtubule organizing center even when mature. In summary, we characterize key cellular and subcellular features of vertebrate sensory neurons as a foundation for understanding their function and maintenance.

Emerging roles for microglia in modifying normal brain development continue to provide new perspectives on the functions of this resident immune cell in the brain. While the molecular underpinnings driving microglia's position in regulating developmental programs remain largely an unchartered territory, innate immune signaling lies at the forefront. At least three innate immune receptors expressed on microglia-fractalkine, complement, and triggering receptor expressed on microglia (TREM2)-modulate developmental synaptic pruning to refine brain circuitry. Our laboratory recently published that microglia with a unique amoeboid morphology invade the corpus callosum and engulf oligodendrocyte progenitor cells (OPCs) during early postnatal development before myelination in a fractalkine receptor (CX3CR1)-dependent manner to modulate ensheathment of axons. Amoeboid microglia are observed in the corpus callosum but not cerebral cortex, and lose their amoeboid shape at the commencement of myelination assuming a resting phenotype. Furthermore, OPCs contacted or engulfed by microglia do not express markers of cell death suggesting a novel homeostatic mechanism facilitating an appropriate OPC:axon ratio for proper myelin ensheathment. The unique morphology of microglia and the restricted window for phagocytic engulfment of OPCs suggest a critical period for OPC engulfment important for action potential propagation during development when activity-dependent mechanisms regulate synaptic pruning. In this review, we summarize the role of activity-dependent mechanisms in sculpting brain circuitry, how myelin ensheathment influences action potential propagation, the spatial and temporal relationship of microglia-dependent elimination of OPCs and synapses, and implications for the synergistic role of microglial phagocytosis in shaping the architecture for neuronal function.

Axons in the adult mammalian central nervous system fail to regenerate after injury. By contrast, spontaneous axon regeneration occurs in the peripheral nervous system (PNS) due to a supportive PNS environment and an increase in the intrinsic growth potential induced by injury via cooperative activation of multifaceted biological pathways. This study compared axon regeneration and injury responses in C57BL/6 male and female mice after sciatic nerve crush (SNC) injury. The extent of axon regeneration in vivo was indistinguishable in male and female mice when observed at 3 days after SNC injury, and primary dorsal root ganglion (DRG) neurons from injured, male and female mice extended axons to a similar length. Moreover, the induction of selected regeneration-associated genes (RAGs), such as Atf3, Sprr1a, Gap43, Sox11, Jun, Gadd45a, and Smad1 were comparable in male and female DRGs when assessed by quantitative real-time reverse transcription polymerase chain reaction. Furthermore, the RNA-seq analysis of male and female DRGs revealed that differentially expressed genes (DEGs) in SNC groups compared to sham-operated groups included many common genes associated with neurite outgrowth. However, we also found that a large number of genes in the DEGs were sex dependent, implicating the involvement of distinct gene regulatory network in the two sexes following peripheral nerve injury. In conclusion, we found that male and female mice mounted a comparable axon regeneration response and many RAGs were commonly induced in response to SNC. However, given that many DEGs were sex-dependently expressed, future studies are needed to investigate whether they contribute to peripheral axon regeneration, and if so, to what extent.

In all animals the optic tectum (OT) (or superior colliculi in mammals) provides guidance for external attention; this is not the only function of the tectum but is critically important for the development of behavioral visual reactions. In fish, the OT is the main primary visual center. It receives signals from most ganglion cells (GC) of different (known) types in the retina. Knowledge of the properties – both structural and physiological – of the neurons in the OT is important for understanding the mechanisms organizing behavior. We recorded extracellular electrical activity in the OT in living adult fish (Carassius auratus gibelio). Simultaneous recordings were made of the responses of retinal GC (from their axon terminals) and tectal neurons (TN), probably from cell bodies. Four types of TN are described with directional selectivity (DS) (henceforth these neurons are termed DS TN) at different (defined) depths of the OT. In addition to these, rare sporadic spikes lacking DS and arising on stimulation at any locus in a large area were consistently recorded simultaneously (superficially) with the responses of caudorostral DS GC with the electrode in one position. These are presumptively the responses of superficial tectal neurons (superficial inhibitory neurons, SIN). Various different types of stimulation were applied with the aim of obtaining clear SIN responses. Comparison of the results of our electrophysiological studies with published data (most studies in this direction have used calcium imaging in transparent Danio rerio fry) showed that DS TN were identical to glutamatergic periventricular interneurons in the OT, while SIN were identical to GABAergic inhibitory interneurons (SIN). These latter presumptively mediate detection of the main object (pop-out) in the field of vision.

Summary In this issue of Neuron, Izadifar et al. (2021) have identified a conserved molecule Wnk as a key regulator in both developmental axon branching and long-term survival of neurons, characterizing its interplay with axon destruction genes including Sarm. The discovery of Wnk will be important to our understanding of neurodevelopmental and neurodegenerative diseases.

Doublecortin-like kinase 1 (DCLK1) is a Doublecortin family kinase involved in a range of brain development processes including cell migration, axon/dendrite growth, and synapse development. The Dclk1 gene potentially generates multiple splicing isoforms, but the detailed expression patterns in the brain as well as in vivo functions of each isoform are still incompletely understood. Here we assessed expression patterns of DCLK1 isoforms using multiple platforms including in silico, in situ, and in vitro datasets in the developing mouse brain, and show quantitative evidence that among the four DCLK1 isoforms, DCLK1-L and DCL are mainly expressed in the embryonic cortex whereas DCLK1-L and CPG16 become dominant compared to DCL and CARP in the postnatal cortex. We also provide compelling evidence that DCLK1 isoforms are distributed in the partially distinct brain regions in the embryonic and the postnatal stages. We further show that overexpression of DCLK1-L, but not the other isoforms, in neural progenitors causes severe migration defects in the cortex, and that the migration defects are dependent on the kinase activity of DCLK1-L. Our data thus uncover partially segregated localization of DCLK1 isoforms in the developing mouse brain and suggest different roles for distinct DCLK1 isoforms in the brain development and function.

Our lab has shown that brief electrical nerve stimulation (ES) has a dramatic impact on remyelination of lysophosphatidyl choline (LPC)-induced focally demyelinated rat peripheral nerves, while also inducing an axon-protective phenotype and shifting macrophages from a predominantly pro-inflammatory toward a pro-repair phenotype. Whether this same potential exists in the central nervous system is not known. Thus, for proof of principle studies, the peripheral nerve demyelination and ES model was adapted to the central nervous system, whereby a unilateral focal LPC-induced demyelination of the dorsal column at the lumbar enlargement where the sciatic nerve afferents enter was created, so that subsequent ipsilateral sciatic nerve ES results in increased neural activity in the demyelinated axons. Data reveal a robust focal demyelination at 7 days post-LPC injection. Delivery of 1-hour ES at 7 days post-LPC polarizes macrophages/microglia toward a pro-repair phenotype when examined at 14 days post-LPC; results in smaller LPC-associated regions of inflammation compared to non-stimulated controls; results in significantly more cells of the oligodendroglial lineage in the demyelinated region; elevates myelin basic protein levels; and shifts the paranodal protein Caspr along demyelinated axons to a more restricted distribution, consistent with reformation of the paranodes of the nodes of Ranvier. ES also significantly enhanced levels of phosphorylated neurofilaments detected in the zones of demyelination, which has been shown to confer axon protection. Collectively these findings support that strategies that increase neural activity, such as brief electrical stimulation, can be beneficial for promoting intrinsic repair following focal demyelinating insults in demyelinating diseases such as multiple sclerosis. All animal procedures performed were approved by the University of Saskatchewan’s Animal Research Ethics Board (protocol# 20090087; last approval date: November 5, 2020).

The axon initial segment (AIS) is essential for maintaining neuronal polarity, modulating protein transport into the axon, and action potential generation. These functions are supported by a distinctive actin and microtubule cytoskeleton that controls axonal trafficking and maintains a high density of voltage-gated ion channels linked by scaffold proteins to the AIS cytoskeleton. However, our knowledge of the mechanisms and proteins involved in AIS cytoskeleton regulation to maintain or modulate AIS structure is limited. In this context, formins play a significant role in the modulation of actin and microtubules. We show that pharmacological inhibition of formins modifies AIS actin and microtubule characteristics in cultured hippocampal neurons, reducing F-actin density and decreasing microtubule acetylation. Moreover, formin inhibition diminishes sodium channels, ankyrinG and ßIV-spectrin AIS density, and AIS length, in cultured neurons and brain slices, accompanied by decreased neuronal excitability. We show that genetic downregulation of the mDia1 formin by interference RNAs also decreases AIS protein density and shortens AIS length. The ankyrinG decrease and AIS shortening observed in pharmacologically inhibited neurons and neuron-expressing mDia1 shRNAs were impaired by HDAC6 downregulation or EB1-GFP expression, known to increase microtubule acetylation or stability. However, actin stabilization only partially prevented AIS shortening without affecting AIS protein density loss. These results suggest that mDia1 maintain AIS composition and length contributing to the stability of AIS microtubules., Open Access funding provided thanks to the CRUE-CSIC agreement with Springer Nature. The work was supported by agrant from Ministerio de Ciencia y Universidades (RTI2018-095156-B-100) to JJG and INSERM funding to DD. Wei Zhang was supported by a fellowship from China Scholarship Council (No.201506300085) and Beatriz Achon by a Master fellowship from Universidad Autónoma de Madrid.

Objective The pathoanatomical correlate of temporal lobe epilepsy is hippocampal sclerosis, characterized by selective neuronal death of mossy cells in the hilus and of pyramidal cells in cornu ammonis 1. Although granule cells survive, they lose mossy cells as a target and redirect their axons (mossy fibers) backward into the molecular cell layer. It has been assumed that this process results in excitatory circuits. We therefore examined whether sprouted mossy fibers form synaptic connection not only with excitatory granule cells but also with inhibitory interneurons, such as basket cells. Methods Resected hippocampal specimens of patients with hippocampal sclerosis were compared to controls of patients with extrahippocampal lesions with only mild sclerosis. Mossy fibers were traced with Neurobiotin or labeled against synaptoporin; inhibitory interneurons were labeled against parvalbumin. Synapses were examined with electron microscopy, labeled with γ-aminobutyric acid immunogold. Results Sprouted mossy fibers of epileptic hippocampi innervate not only excitatory granule cells but also inhibitory parvalbuminergic interneurons. Despite neuronal death in hippocampal sclerosis, the axonal plexus of inhibitory parvalbuminergic interneurons surrounding the granule cells is preserved. Connections of sprouted mossy fibers and inhibitory axon terminals were quantified, showing that the number of inhibitory axon terminals significantly exceeds the number of sprouted excitatory mossy fiber terminals (.03 boutons/µm vs. .11 boutons/µm; p Significance Although no definite conclusions regarding the function of our findings may be derived from this anatomical study, the observed aberrant connectivity might lead to an increased inhibition and synchronization of granule cells, because the preserved inhibitory interneurons show an additional innervation through sprouted mossy fibers. This might result in the instability of a previously balanced network.

Axon regenerative failure in the mature CNS contributes to functional deficits following many traumatic injuries, ischemic injuries, and neurodegenerative diseases. The complement cascade of the innate immune system responds to pathogen threat through inflammatory cell activation, pathogen opsonization, and pathogen lysis, and complement is also involved in CNS development, neuroplasticity, injury, and disease. Here, we investigated the involvement of the classical complement cascade and microglia/monocytes in CNS repair using the mouse optic nerve injury (ONI) model, in which axons arising from retinal ganglion cells (RGCs) are disrupted. We report that central complement C3 protein and mRNA, classical complement C1q protein and mRNA, and microglia/monocyte phagocytic complement receptor CR3 all increase in response to ONI, especially within the optic nerve itself. Importantly, genetic deletion ofC1q,C3, orCR3attenuates RGC axon regeneration induced by several distinct methods, with minimal effects on RGC survival. Local injections of C1q function-blocking antibody revealed that complement acts primarily within the optic nerve, not retina, to support regeneration. Moreover, C1q opsonizes and CR3+microglia/monocytes phagocytose growth-inhibitory myelin debris after ONI, a likely mechanism through which complement and myeloid cells support axon regeneration. Collectively, these results indicate that local optic nerve complement-myeloid phagocytic signaling is required for CNS axon regrowth, emphasizing the axonal compartment and highlighting a beneficial neuroimmune role for complement and microglia/monocytes in CNS repair.SIGNIFICANCE STATEMENTDespite the importance of achieving axon regeneration after CNS injury and the inevitability of inflammation after such injury, the contributions of complement and microglia to CNS axon regeneration are largely unknown. Whereas inflammation is commonly thought to exacerbate the effects of CNS injury, we find that complement proteins C1q and C3 and microglia/monocyte phagocytic complement receptor CR3 are each required for retinal ganglion cell axon regeneration through the injured mouse optic nerve. Also, whereas studies of optic nerve regeneration generally focus on the retina, we show that the regeneration-relevant role of complement and microglia/monocytes likely involves myelin phagocytosis within the optic nerve. Thus, our results point to the importance of the innate immune response for CNS repair.

Chronic sleep restriction (CSR) is common in modern society, adversely affecting cognitive performance and health. Yet how it impacts neurons regulating sleep remains unclear. Several studies using mice reported substantial losses of wake-active orexin/hypocretin and locus coeruleus (LC) noradrenergic neurons, but not rapid eye movement sleep-active melanin-concentrating hormone (MCH) neurons, following CSR. Here, we used immunohistochemistry and stereology to examine orexin, MCH and LC noradrenergic neurons in a rat model of CSR that uses programmed wheel rotation (3 h on/1 h off; '3/1' protocol). Adult male Wistar rats underwent one or four cycles of the 4-day 3/1 CSR protocol, with 2-day recovery between cycles in home cages. Time-matched control rats were housed in locked wheels/home cages. We found no significant differences in the numbers of orexin, MCH and LC noradrenergic neurons following either one- or four-cycle CSR protocol compared to respective controls. Similarly, the four-cycle CSR protocol had no effect on the densities of orexin axon terminals in the LC, noradrenergic dendrites in the LC and noradrenergic axon terminals in the frontal cortex. Body weights, however, decreased after one cycle of CSR and then increased with diminishing slope over the next three cycles. Thus, we found no evidence for loss of orexin or LC noradrenergic neurons following one and four cycles of the 4-day 3/1 CSR protocol in rats. Differences in CSR protocols and/or possible species differences in neuronal vulnerability to sleep loss may account for the discrepancy between the current results in rats and previous findings in mice.

Neurons can change their classical neurotransmitters during ontogeny, sometimes going through stages of dual release. Here, we explored the development of the neurotransmitter identity of neurons of the avian nucleus isthmi parvocellularis (Ipc), whose axon terminals are retinotopically arranged in the optic tectum (TeO) and exert a focal gating effect upon the ascending transmission of retinal inputs. Although cholinergic and glutamatergic markers are both found in Ipc neurons and terminals of adult pigeons and chicks, the mRNA expression of the vesicular acetylcholine transporter, VAChT, is weak or absent. To explore how the Ipc neurotransmitter identity is established during ontogeny, we analyzed the expression of mRNAs coding for cholinergic (ChAT, VAChT, and CHT) and glutamatergic (VGluT2 and VGluT3) markers in chick embryos at different developmental stages. We found that between E12 and E18, Ipc neurons expressed all cholinergic mRNAs and also VGluT2 mRNA; however, from E16 through posthatch stages, VAChT mRNA expression was specifically diminished. Our ex vivo deposits of tracer crystals and intracellular filling experiments revealed that Ipc axons exhibit a mature paintbrush morphology late in development, experiencing marked morphological transformations during the period of presumptive dual vesicular transmitter release. Additionally, although ChAT protein immunoassays increasingly label the growing Ipc axon, this labeling was consistently restricted to sparse portions of the terminal branches. Combined, these results suggest that the synthesis of glutamate and acetylcholine, and their vesicular release, is complexly linked to the developmental processes of branching, growing and remodeling of these unique axons.

Among the spinocerebellar projections vital for sensorimotor coordination of limbs and the trunk, the morphology of spinocerebellar axons originating from the lumbar cord has not been well characterized compared to those from thoracic and sacral cords. We reconstructed 26 single spinocerebellar axons labeled by biotinylated dextran injections into the gray matter of the lumbar spinal cord in mice. Axon terminals were mapped with the zebrin pattern of the cerebellar cortex. Reconstructed axons were primarily classified into ipsilaterally and contralaterally ascending axons, arising mainly from the dorsal and ventral horns, respectively. The majority of ipsilateral and contralateral axons took the dorsal-medullary and ventral-pontine pathways, respectively. The axons of both groups terminated mainly in the vermal and medial paravermal areas of lobules II-V and VIII-IXa, often bilaterally but predominantly ipsilateral to the axonal origin, with a weak preference to particular portions of zebrin stripes. The ipsilateral axons originating from the medial dorsal horn in the upper lumbar cord (n = 3) had abundant (43-147) mossy fiber terminals and no medullary collaterals. The ipsilateral axons originating from the lateral dorsal horn in the lower lumbar cord (n = 9) and the contralateral axons (n = 14) showed remarkable morphology variations. The number of their mossy fiber terminals varied from 2 to 172. Their collaterals, observed in 17 axons out of 23, terminated mainly in the medial cerebellar nucleus, nucleus X, and lateral reticular nucleus in various degrees. The results indicated that the lumbar spinocerebellar projection contains highly heterogeneous axonal populations regarding their pathway, branching, and termination patterns.

BACKGROUND Proper guidance of neuronal axons to their targets is required to assemble neural circuits during the development of the nervous system. However, the mechanism by which the guidance of axonal growth cones is regulated by specific intermediaries activated by receptor signaling pathways to mediate cytoskeleton dynamics is unclear. Vav protein members have been proposed to mediate this process, prompting us to investigate their role in the limb selection of the axon trajectory of spinal lateral motor column (LMC) neurons. RESULTS We found Vav2 and Vav3 expression in LMC neurons when motor axons grew into the limb. Vav2, but not Vav3, loss-of-function perturbed LMC pathfinding, while Vav2 gain-of-function exhibited the opposite effects, demonstrating that Vav2 plays an important role in motor axon growth. Vav2 knockdown also attenuated the redirectional phenotype of LMC axons induced by Dcc, but not EphA4, in vivo and lateral LMC neurite growth preference to Netrin-1 in vitro. This study showed that Vav2 knockdown and ectopic nonphosphorylable Vav2 mutant expression abolished the Src-induced stronger growth preference of lateral LMC neurites to Netrin-1, suggesting that Vav2 is downstream of Src in this context. CONCLUSIONS Vav2 is essential for Netrin-1-regulated LMC motor axon pathfinding through Src interaction.

Glaucoma is a prevalent neurodegenerative disease that is characterized by progressive visual field loss. It is the leading cause of irreversible blindness in the world. The main risk factor for glaucoma is elevated intraocular pressure that results in the damage and death of retinal ganglion cells (RGCs) and their axons. The death of RGCs has been shown to be apoptotic. We tested the hypothesis that blocking the activation of apoptosis may be an effective strategy to prevent RGC death and preserve functional vision in glaucoma. In the magnetic microbead mouse model of induced ocular hypertension, inhibition of RGC apoptosis was targeted through viral-mediated ocular delivery of the X-linked inhibitor of apoptosis (XIAP) gene, a potent caspase inhibitor. Pattern electroretinograms revealed that XIAP therapy resulted in significant protection of both somal and axonal RGC function in glaucomatous eyes. Histology confirmed that the treated optic nerves showed preservation of axon counts and reduced glial cell infiltration. These results show that XIAP is able to provide both functional and structural protection of RGCs in the microbead model of glaucoma and provide important proof-of-principle for XIAP's efficacy as a neuroprotective treatment for glaucoma.

The primary function of the urinary bladder is to store urine (continence) until a suitable time for voiding (micturition). These distinct processes are determined by the coordinated activation of sensory and motor components of the nervous system, which matures to enable voluntary control at the time of weaning. Our aim was to define the development and maturation of the nerve-organ interface of the mouse urinary bladder by mapping the organ and tissue distribution of major classes of autonomic (motor) and sensory axons. Innervation of the bladder was evident from E13 and progressed dorsoventrally. Increasing defasciculation of axon bundles to single axons within the muscle occurred through the prenatal period, and in several classes of axons underwent further maturation until P7. Urothelial innervation occurred more slowly than muscle innervation and showed a clear regional difference, from E18 the bladder neck having the highest density of urothelial nerves. These features of innervation were similar in males and females but varied in timing and tissue density between different axon classes. We also analysed the pelvic ganglion, the major source of motor axons that innervate the lower urinary tract and other pelvic organs. Cholinergic, nitrergic (subset of cholinergic) and noradrenergic neuronal cell bodies were present prior to visualization of these axon classes within the bladder. Examination of cholinergic structures within the pelvic ganglion indicated that connections from spinal preganglionic neurons to pelvic ganglion neurons were already present by E12, a time at which these autonomic ganglion neurons had not yet innervated the bladder. These putative preganglionic inputs increased in density prior to birth as axon terminal fields continued to expand within the bladder tissues. Our studies also revealed in numerous pelvic ganglion neurons an unexpected transient expression of calcitonin gene-related peptide, a peptide commonly used to visualise the peptidergic class of visceral sensory axons. Together, our outcomes enhance our understanding of neural regulatory elements in the lower urinary tract during development and provide a foundation for studies of plasticity and regenerative capacity in the adult system.

Dendrite and axon arbors form scaffolds that connect a neuron to its partners; they are patterned to support the specific connectivity and computational requirements of each neuron subtype. Transcription factor networks control the specification of neuron subtypes, and the consequent diversification of their stereotyped arbor patterns during differentiation. We outline how the differentiation trajectories of stereotyped arbors are shaped by hierarchical deployment of precursor cell and postmitotic transcription factors. These transcription factors exert modular control over the dendrite and axon features of a single neuron, create spatial and functional compartmentalization of an arbor, instruct implementation of developmental patterning rules, and exert operational control over the cell biological processes that construct an arbor.

Sensory disturbance in the orofacial region owing to trigeminal nerve injury is caused by dental treatment or accident. Commercially available therapeutics are ineffective for the treatment of sensory disturbance. Additionally, the therapeutic effects of rapamycin, an allosteric inhibitor of mammalian target of rapamycin (mTOR), which negatively regulates autophagy, on the sensory disturbance are not fully investigated. Thus, we investigated the therapeutic effects of rapamycin on the sensory disturbance in the mandibular region caused by inferior alveolar nerve (IAN) transection (IANX) in rats. The expression levels of the phosphorylated p70S6K, a downstream molecule of mTOR, in the proximal and distal stumps of the transected IAN were significantly reduced by rapamycin administration to the injured site. Conversely, the increments of both Beclin 1 and microtubule-associated protein-1 light chain 3-II protein levels in the proximal and distal stumps of the transected IAN was induced by rapamycin administration. Immunohistochemical analyses revealed that Beclin 1 was located in Schwann cells in the proximal stump of the IAN. Accumulation of myelin protein zero and myelin basic protein in the proximal and distal stumps of the IAN was significantly reduced by rapamycin administration. Rapamycin administration facilitated axon regeneration after IANX and increased the number of brain-derived neurotrophic factor positive neurons in the trigeminal ganglion. Thus, recovery from sensory disturbance in the lower lip caused by IANX was markedly facilitated by rapamycin. These findings suggest that rapamycin administration is a promising treatment for the sensory disturbance caused by IANX.

Axon degeneration is a prominent feature of the injured nervous system, occurs across neurological diseases, and drives functional loss in neural circuits. We have seen a paradigm shift in the last decade with the realization that injured axons are capable of actively driving their own destruction through the sterile-alpha and TIR motif containing 1 (SARM1) protein. Early studies of Wallerian degeneration highlighted a central role for NAD+ metabolites in axon survival, and this association has grown even stronger in recent years with a deeper understanding of SARM1 biology. Here, we review our current knowledge of SARM1 function in vivo and our evolving understanding of its complex architecture and regulation by injury-dependent changes in the local metabolic environment. The field is converging on a model whereby SARM1 acts as a sensor for metabolic changes that occur after injury and then drives catastrophic NAD+ loss to promote degeneration. However, a number of observations suggest that SARM1 biology is more complicated, and there remains much to learn about how SARM1 governs nervous system responses to injury or disease.

Oligodendroglial progenitor cells (OPC) are highly proliferative and migratory bipolar cells, which differentiate into complex myelin forming and axon ensheathing mature oligodendrocytes during myelination. Recent studies indicate that the oligodendroglial cell population is heterogeneous on transcriptional and functional level depending on the location in the CNS. Here, we compared intrinsic properties of OPC from spinal cord and brain on functional and transcriptional level. Spinal cord OPC demonstrated increased migration as well as differentiation capacity. Moreover, transcriptome analysis revealed differential expression of several genes between both OPC populations. In spinal cord OPC we confirmed upregulation of SKAP2, a cytoplasmatic adaptor protein known for its implication in cytoskeletal remodelling and migration in other cell types. Recent findings suggest that actin dynamics determine not only oligodendroglial migration, but also differentiation: Whereas actin polymerization is important for process extension, actin destabilization and depolymerization is required for myelin sheath formation. Downregulation or complete lack of SKAP2 in OPC resulted in reduced migration and impaired morphological maturation in oligodendrocytes. In contrast, overexpression of SKAP2 as well as constitutively active SKAP2 increased OPC migration suggesting that SKAP2 function is dependent on activation by phosphorylation. Furthermore, lack of SKAP2 enhanced the positive effect on OPC migration after integrin activation suggesting that SKAP2 acts as modulator of integrin dependent migration. In summary, we demonstrate the presence of intrinsic differences between spinal cord and brain OPC and identified SKAP2 as a new regulator of oligodendroglial migration and sheath formation.Significance statementOPC play an important role in many still incurable diseases, such as multiple sclerosis, leukodystrophies or neurodegenerative diseases. Their heterogeneity in different CNS regions has recently been identified. Here, we observed increased migration and differentiation capabilities of OPC isolated from the spinal cord compared to brain OPC and confirmed differences in the transcriptome between these two cell populations. Furthermore, we identified SKAP2 as potential modulator of actin dynamics in oligodendrocytes. Whereas knockdown or lack of SKAP2 impairs migration and myelin sheath formation in mouse and human oligodendrocytes, overexpression of wildtype or constitutively active SKAP2 enhances the migratory capacity of murine oligodendrocytes. In summary, we present SKAP2 as modulator of cytoskeletal dynamics regulating OPC migration, differentiation and myelin sheath formation.

Cytoarchitectonical parcellation of the visual cortex into the striate and extrastriate cortex requires complex histogenetic events within a precise spatio-temporal frame to attain the specification of areal domains and associated thalamocortical connections during the fetal brain development. We analyzed a deep subplate cellular monolayer (subplate “corridor” cells) present during a restricted period of 13–15 postconceptional weeks, showing the 3D caudo-ventro-medial position in the human fetal occipital lobe, corresponding to the segregation point of pulvinocortical and geniculocortical fibers at the prospective area 17/18 border. Immunofluorescence stainings revealed subplate “corridor” cells as the specific class of the deepest subplate neurons (NeuN+, Tbr1+, Cplx3+) expressing axon guidance molecules (Sema-3A+, EphA6+), presumably for the attraction of pulvinocortical axons and the repulsion of geniculocortical axons growing at that time (SNAP25+, Syn+, FN+). Furthermore, quantitative analysis of the subplate “corridor” region of interest, considering cell number, immunofluorescence signal intensity per cell and per region, revealed significant differences to other regions across the tangential circumference of the developing cerebral wall. Thus, our study sheds new light on the deepest subplate sublayer, strategically aligned along the growing axon systems in the prospective visual system, suggesting the establishment of the area 17/18 border by differential thalamocortical input during the fetal brain development.

Glial cells play essential roles in the nervous system. Although glial populations are tightly regulated, the mechanisms regulating the population size remain poorly understood. Since Drosophila glial cells are similar to the human counterparts in their functions and shapes, rendering them an excellent model system to understand the human glia biology. Lipid phosphate phosphatases (LPP) are important for regulating bioactive lipids. In Drosophila, there are three known LPP-encoding genes; wunen, wunen-2 and lazaro. The wunens are important for the germ cell migration and survival, and septate junction formation during tracheal development. Lazaro is involved in phototransduction. In the present study, we characterized a novel Drosophila LPP-encoding gene, CG11426. Suppression of CG11426 increased glial cell number in the eye imaginal disc during larval development, while ectopic CG11426 expression decreased it. Both types of mutation also caused defects in axon projection to the optic lobe in larval eye-brain complexes. Moreover, CG11426 promoted apoptosis via inhibiting ERK signaling in the eye imaginal disc. Taken together, these findings demonstrated that CG11426 gene product negatively regulates ERK signaling to promote apoptosis for proper maintenance of glial population in the developing eye disc. This article is protected by copyright. All rights reserved.

Axonal plasticity allows neurons to control their output, which critically determines the flow of information in the brain. Axon diameter can be regulated by activity, yet how morphological changes in an axon impact its function remains poorly understood. Axonal swellings have been found on Purkinje cell axons in the cerebellum both in healthy development and in neurodegenerative diseases, and computational models predicts that axonal swellings impair axonal function. Here we report that in young Purkinje cells, axons with swellings propagated action potentials with higher fidelity than those without, and that axonal swellings form when axonal failures are high. Furthermore, we observed that healthy young adult mice with more axonal swellings learn better on cerebellar-related tasks than mice with fewer swellings. Our findings suggest that axonal swellings underlie a form of axonal plasticity that optimizes the fidelity of action potential propagation in axons, resulting in enhanced learning., Axonal swellings have been found on Purkinje cell axons in the cerebellum both during development and disease. The authors show that axons with swellings propagate action potentials with higher fidelity than those without and that mice with more axonal swellings learn cerebellar-related tasks better.