Axonal and neuromuscular synaptic phenotypes in WldS,SOD1G93A and ostes mutant mice identified by fiber-optic confocal microendoscopy

We used live imaging by fiber-optic confocal microendoscopy (CME) of yellow fluorescent protein (YFP) expression in motor neurons to observe and monitor axonal and neuromuscular synaptic phenotypes in mutant mice. First, we visualized slow degeneration of axons and motor nerve terminals at neuromuscular junctions following sciatic nerve injury in Wld^S mice with slow Wallerian degeneration. Protection of axotomized motor nerve terminals was much weaker in Wld^S heterozygotes than in homozygotes. We then induced covert modifiers of axonal and synaptic degeneration in heterozygous Wld^S mice, by N-ethyl-N-nitrosourea (ENU) mutagenesis, and used CME to identify candidate mutants that either enhanced or suppressed axonal or synaptic degeneration. From 219 of the F1 progeny of ENU-mutagenized BALB/c mice and thy1.2-YFP16/Wld^S mice, CME revealed six phenodeviants with suppression of synaptic degeneration. Inheritance of synaptic protection was confirmed in three of these founders, with evidence of Mendelian inheritance of a dominant mutation in one of them (designated CEMOP_S5). We next applied CME repeatedly to living Wld^S mice and to SOD1^G93A mice, an animal model of motor neuron disease, and observed degeneration of identified neuromuscular synapses over a 1–4 day period in both of these mutant lines. Finally, we used CME to observe slow axonal regeneration in the ENU-mutant ostes mouse strain. The data show that CME can be used to monitor covert axonal and neuromuscular synaptic pathology and, when combined with mutagenesis, to identify genetic modifiers of its progression in vivo.     

Disruption of the midkine gene (Mdk) delays degeneration and regeneration in injured peripheral nerve

Midkine (MK) is a growth factor implicated in the development and repair of various tissues, especially neural tissues. MK acts as a reparative neurotrophic factor in damaged peripheral nerves. A postulated role of MK in the degeneration and regeneration of sciatic nerves was explored by comparing wild‐type (Mdk^+/+) mice with MK‐deficient (Mdk^?/?) mice after freezing injury. In the Mdk^?/? mice, a regenerative delay was observed, preceded by a decelerated Wallerian degeneration (WD). The relative wet weight of the soleus muscle slowly declined, and recovery was delayed compared with that in the Mdk^+/+ mice. In the regenerating nerve, unmyelinated axons were unevenly distributed, and some axons contained myelin‐like, concentrically lamellated bodies. In the endplates of soleus muscles, nerve terminals containing synaptic vesicles disappeared in both mice. In Mdk^?/? mice, the appearance of nerve terminals was delayed in synaptic vesicles of terminal buttons after injury. The recovery of evoked electromyogram was delayed in Mdk^?/? mice compared with Mdk^+/+ mice. Our results suggested a delay in axonal degeneration and regeneration in Mdk^?/? mice compared with Mdk^+/+ mice, and the delayed regeneration was associated with a delayed recovery of motor function. These findings show that a lack of MK following peripheral nerve injury is a critical factor in degeneration and regeneration, and manipulation of the supply of MK may offer interesting therapeutic options for the treatment of peripheral nerve damage. © 2009 Wiley‐Liss, Inc.     

VCP binding influences intracellular distribution of the slow Wallerian degeneration protein, Wld(S)

Wallerian degeneration slow (Wld^S) mice express a chimeric protein that delays axonal degeneration. The N-terminal domain (N70), which is essential for axonal protection in vivo, binds valosin-containing protein (VCP) and targets both Wld^S and VCP to discrete nuclear foci. We characterized the formation, composition and localization of these potentially important foci. Missense mutations show that the N-terminal sixteen residues (N16) of Wld^S are essential for both VCP binding and targeting Wld^S to nuclear foci. Removing N16 abolishes foci, and VCP binding sequences from ataxin-3 or HrdI restore them. In vitro, these puncta co-localize with proteasome subunits. In vivo, Wld^S assumes a range of nuclear distribution patterns, including puncta, and its neuronal expression and intranuclear distribution is region-specific and varies between spontaneous and transgenic Wld^S models. We conclude that VCP influences Wld^S intracellular distribution, and thus potentially its function, by binding within the N70 domain required for axon protection.     

Baculoviral inhibitor of apoptosis protein repeat-containing protein 3 delays early Wallerian degeneration after sciatic nerve injury

Wallerian degeneration is a complex biological process that occurs after nerve injury,and involves nerve degeneration and regeneration.Schwann cells play a crucial role in the cellular and molecular events of Wallerian degeneration of the peripheral nervous system.However,Wallerian degeneration regulating nerve injury and repair remains largely unknown,especially the early response.We have previously reported some key regulators of Wallerian degeneration after sciatic nerve injury.Baculoviral inhibitor of apoptosis protein repeat-containing protein 3(BIRC3)is an important factor that regulates apoptosis-inhibiting protein.In this study,we established rat models of right sciatic nerve injury.In vitro Schwann cell models were also established and subjected to gene transfection to inhibit and overexpress BIRC3.The data indicated that BIRC3 expression was significantly up-regulated after sciatic nerve injury.Both BIRC3 upregulation and downregulation affected the migration,proliferation and apoptosis of Schwan cells and affected the expression of related factors through activating c-fos and ERK signal pathway.Inhibition of BIRC3 delayed early Wallerian degeneration through inhibiting the apoptosis of Schwann cells after sciatic nerve injury.These findings suggest that BIRC3 plays an important role in peripheral nerve injury repair and regeneration.The study was approved by the Institutional Animal Care and Use Committee of Nantong University,China(approval No.2019-nsfc004)on March 1,2019.     

The effects of claudin 14 during early Wallerian degeneration after sciatic nerve injury

Claudin 14 has been shown to promote nerve repair and regeneration in the early stages of Wallerian degeneration(0–4 days) in rats with sciatic nerve injury, but the mechanism underlying this process remains poorly understood. This study reported the effects of claudin 14 on nerve degeneration and regeneration during early Wallerian degeneration. Claudin 14 expression was up-regulated in sciatic nerve 4 days after Wallerian degeneration. The altered expression of claudin 14 in Schwann cells resulted in expression changes of cytokines in vitro. Expression of claudin 14 affected c-Jun, but not Akt and ERK1/2 pathways. Further studies revealed that enhanced expression of claudin 14 could promote Schwann cell proliferation and migration. Silencing of claudin 14 expression resulted in Schwann cell apoptosis and reduction in Schwann cell proliferation. Our data revealed the role of claudin 14 in early Wallerian degeneration, which may provide new insights into the molecular mechanisms of Wallerian degeneration.     

Lack of galectin‐3 speeds Wallerian degeneration by altering TLR and pro‐inflammatory cytokine expressions in injured sciatic nerve

Wallerian degeneration (WD) comprises a series of events that includes activation of non‐neuronal cells and recruitment of immune cells, creating an inflammatory milieu that leads to extensive nerve fragmentation and subsequent clearance of the myelin debris, both of which are necessary prerequisites for effective nerve regeneration. Previously, we documented accelerated axon regeneration in animals lacking galectin‐3 (Gal‐3), a molecule associated with myelin clearance. To clarify the mechanisms underlying this enhanced regeneration, we focus here on the early steps of WD following sciatic nerve crush in Gal‐3^?/? mice. Using an in vivo model of nerve degeneration, we observed that removal of myelin debris is more efficient in Gal‐3^?/? than in wild‐type (WT) mice; we next used an in vitro phagocytosis assay to document that the phagocytic potential of macrophages and Schwann cells was enhanced in the Gal‐3^?/? mice. Moreover, both RNA and protein levels for the pro‐inflammatory cytokines IL‐1β and TNF‐α, as well as for Toll‐like receptor (TLR)‐2 and ‐4, show robust increases in injured nerves from Gal‐3^?/?mice compared to those from WT mice. Collectively, these data indicate that the lack of Gal‐3 results in an augmented inflammatory profile that involves the TLR–cytokine pathway, and increases the phagocytic capacity of Schwann cells and macrophages, which ultimately contributes to speeding the course of WD.     

Genetic influences on cellular reactions to spinal cord injury: A wound-healing response present in normal mice is impaired in mice carrying a mutation (WldS) that causes delayed Wallerian degeneration

Progressive tissue necrosis is a process unique to the injured mammalian spinal cord which often leads to gradually increasing cavitation and enlargement of the lesion. To evaluate the role of neuronal degeneration in initiating this response, histopathological changes were compared in C57BL and Wld^S (delayed Wallerian degeneration mutation) mice. The spinal cord was crushed at T8, producing a primary lesion at the site of the trauma and a secondary lesion extending rostrocaudally in the dorsal columns (where long ascending and descending fiber tracts undergo Wallerian degeneration). Cavitation was relatively mild at both sites and developed mainly at the margins of the lesions. In striking contrast to spinal cord injury in rats, progressive necrosis did not occur in mice; instead, the primary and secondary lesion sites became filled in by macrophages and fibroblasts embedded in a well-vascularized collagenous stroma. Quantitative image analysis revealed that the primary lesion decreased dramatically in size and cavitation between 2 and 3 weeks in C57BL, whereas in Wld^S the reduction in size and cavitation began later (at 4 weeks) and was less complete. The initial development of the secondary lesion began later and its healing was less complete in Wld^S than C57BL. These results are consistent with the hypothesis that neuronal damage, including Wallerian degeneration, triggers inflammatory responses leading to tissue repair. For this reason, any delay in neuronal degeneration, as in the Wld^S mutation, results in deficient tissue repair as reflected in the larger size of both primary and dorsal column lesions. © 1996 Wiley-Liss, Inc.     

Differences in peripheral nerve degeneration/regeneration between wild-type and neuronal nitric oxide synthase knockout mice

Nitric oxide (NO), a unique biological messenger molecule, is synthesized by three isoforms of the enzyme NO synthase (NOS) and diffuses from the site of production across cellular membranes. A postulated role for NO in degeneration and regeneration of peripheral nerves has been explored in a sciatic nerve model comparing wild-type mice and mice lacking neuronal NOS after transection and microsurgical repair. In NOS knockout mice, regenerative delay was observed, preceded by a decelerated Wallerian degeneration (WD). In the regenerated nerve, pruning of uncontrolled sprouts was disturbed, leading to an enhanced number of axons, whereas remyelination seemed to be less affected. Delayed regeneration was associated with a delayed recovery of sensor and motor function. In such a context, possible NO targets are neurofilaments and myelin sheaths of the interrupted axon, filopodia of the growth cone, newly formed neuromuscular endplates, and Schwann cells in the distal nerve stump. The results presented suggest that 1) local release of NO following peripheral nerve injury is a crucial factor in degeneration/regeneration, 2) success of fiber regeneration in the peripheral nervous system depends on a regular WD, and 3) manipulation of NO supply may offer interesting therapeutic options for treatment of peripheral nerve lesions.     

Heat shock protein is a key therapeutic target for nerve repair in autoimmune peripheral neuropathy and severe peripheral nerve injury

Guillain-Barré syndrome (GBS) is an autoimmune peripheral neuropathy and a common cause of neuromuscular paralysis. Preceding infection induces the production of anti-ganglioside (GD) antibodies attacking its own peripheral nerves. In severe proximal peripheral nerve injuries that require long-distance axon regeneration, motor functional recovery is virtually nonexistent. Damaged axons fail to regrow and reinnervate target muscles. In mice, regenerating axons must reach the target muscle within 35 days (critical period) to reform functional neuromuscular junctions and regain motor function. Successful functional recovery depends on the rate of axon regeneration and debris removal (Wallerian degeneration) after nerve injury. The innate-immune response of the peripheral nervous system to nerve injury such as timing and magnitude of cytokine production is crucial for Wallerian degeneration. In the current study, forced expression of human heat shock protein (hHsp) 27 completely reversed anti-GD-induced inhibitory effects on nerve repair assessed by animal behavioral assays, electrophysiology and histology studies, and the beneficial effect was validated in a second mouse line of hHsp27. The protective effect of hHsp27 on prolonged muscle denervation was examined by performing repeated sciatic nerve crushes to delay regenerating axons from reaching distal muscle from 37 days up to 55 days. Strikingly, hHsp27 was able to extend the critical period of motor functional recovery for up to 55 days and preserve the integrity of axons and mitochondria in distal nerves. Cytokine array analysis demonstrated that a number of key cytokines which are heavily involved in the early phase of innate-immune response of Wallerian degeneration, were found to be upregulated in the sciatic nerve lysates of hHsp27 Tg mice at 1 day postinjury. However, persistent hyperinflammatory mediator changes were found after chronic denervation in sciatic nerves of littermate mice, but remained unchanged in hHsp27 Tg mice. Taken together, the current study provides insight into the development of therapeutic strategies to enhance muscle receptiveness (reinnervation) by accelerating axon regeneration and Wallerian degeneration.     

Chronic nerve compression alters Schwann cell myelin architecture in a murine model

Introduction: Myelinating Schwann cells compartmentalize their outermost layer to form actin‐rich channels known as Cajal bands. Herein we investigate changes in Schwann cell architecture and cytoplasmic morphology in a novel mouse model of carpal tunnel syndrome. Methods: Chronic nerve compression (CNC) injury was created in wild‐type and slow‐Wallerian degeneration (Wld^S) mice. Over 12 weeks, nerves were electrodiagnostically assessed, and Schwann cell morphology was thoroughly evaluated. Results: A decline in nerve conduction velocity and increase in g‐ratio is observed without early axonal damage. Schwann cells display shortened internodal lengths and severely disrupted Cajal bands. Quite surprisingly, the latter is reconstituted without improvements to nerve conduction velocity. Conclusions: Chronic entrapment injuries like carpal tunnel syndrome are primarily mediated by the Schwann cell response, where decreases in internodal length and myelin thickness disrupt the efficiency of impulse propagation. Restitution of Cajal bands is not sufficient for remyelination after CNC injury. Muscle Nerve, 2012     

Role of microtubule dynamics in Wallerian degeneration and nerve regeneration after peripheral nerve injury

Wallerian degeneration,the progressive disintegration of distal axons and myelin that occurs after peripheral nerve injury,is essential for creating a permissive microenvironment for nerve regeneration,and involves cytoskeletal reconstruction.However,it is unclear whether microtubule dynamics play a role in this process.To address this,we treated cultured sciatic nerve explants,an in vitro model of Wallerian degeneration,with the microtubule-targeting agents paclitaxel and nocodazole.We found that paclitaxel-induced microtubule stabilization promoted axon and myelin degeneration and Schwann cell dedifferentiation,whereas nocodazole-induced microtubule destabilization inhibited these processes.Evaluation of an in vivo model of peripheral nerve injury showed that treatment with paclitaxel or nocodazole accelerated or attenuated axonal regeneration,as well as functional recovery of nerve conduction and target muscle and motor behavior,respectively.These results suggest that microtubule dynamics participate in peripheral nerve regeneration after injury by affecting Wallerian degeneration.This study was approved by the Animal Care and Use Committee of Southern Medical University,China(approval No.SMUL2015081) on October 15,2015.     

Axonal autophagy during regeneration of the rat sciatic nerve**★

BACKGROUND: The removal of degenerated axonal debris during Wallerian degeneration is very important for nerve regeneration. However, the mechanism by which debris is removed is not been completely understood. Considerable controversy remains as to the clearance pathway and cells that are involved. OBJECTIVE: To investigate axonal autophagy during removal of degenerated axonal debris by transecting the sciatic nerve in a rat Wallerian degeneration model.DESIGN, TIME AND SETTING: Experimental neuropathological analysis. The experiment was conducted at the Laboratory Animal Service Center of the Southern Medical University between January and June 2005. MATERIALS: Fifty-four adult, Wistar rats of either sex, weighing 180-250 g, were obtained from the Laboratory Animal Service Center of the Southern Medical University. Animals were randomly divided into nine groups of six rats. METHODS: Wallerian degeneration was induced by transecting the rat sciatic nerve, and tissue samples from the distal stump were obtained 0.2, 0.4, 1, 2, 3, 4, 7, 10, and 15 days post-transection. Ultrathin sections were prepared for electron microscopy to study ultrastructure and enzyme cytochemistry staining. MAIN OUTCOME MEASURES: Ultrastructure (axon body, autophagic body, and cystoskeleton) of axons and myelin sheaths observed with electron microscopy; acidic phosphatase activity detected by Gomori staining using electron microscopy. RESULTS: The major changes of degenerating axons after transection were axoplasm swelling and separation of axons from their myelin sheath between five hours and two days post-transection. At four days post-transection, the axoplasm condensed and axons were completely separated from the myelin sheath, forming dissociative axon bodies. Vacuoles of different sizes formed in axons during the early phase after lesion. Larger dissociative axon bodies were formed when the axons were completely separated from the myelin sheath during a late phase. The axolemma surrounding the axon body was derived from the neuronal cell membrane; the condensed axoplasm contained many autophagic vacuoles at all levels. A large number of neurofilaments, microtubules, and microfilaments were arranged in a criss-cross pattern. The autophagic vacuoles exhibited acidic phosphatase activity. Axonal bodies were absorbed after degradation from day 7 onwards, and macrophages were observed rarely in the formative cavity. CONCLUSION: The degenerating axons were cleared mainly by axonal autophagy and Schwann cell phagocytosis during regeneration of the rat sciatic nerve, and macrophages exhibited only an assisting function.     

Differences in Sympathetic Innervation of Mouse DRG Following Proximal or Distal Nerve Lesions

Nerve injury leads to novel sympathetic innervation of the dorsal root ganglion (DRG). We have hypothesized previously that the degenerating nerve increases the sympathetic sprouting in the DRG and pain after chronic sciatic constriction injury (CCI) by virtue of its influence on sensory and sympathetic axons spared by the injury. However, L5 spinal nerve ligation and transection (SNL) results in the complete isolation of the L5 DRG from the degenerating stump, yet sympathetic axons invade the ganglion, and sympathetically dependent pain develops. We investigated the role of Wallerian degeneration in both sympathetic sprouting and neuropathic pain in these two models of painful peripheral neuropathy by comparing responses of normal C57B1/6J and C57B1/Wld^smice in which degeneration is impaired. After CCI, Wld^smice, unlike 6J mice, did not develop thermal or mechanoallodynia or sympathetic innervation of the L5 DRG. After SNL, both strains developed mechanoallodynia and sympathetic sprouts in L5, but only 6J mice developed thermal allodynia. Observation of the origins of the invading sympathetic axons revealed that after CCI, sympathetics innervating blood vessels and dura (probably intact) sprouted into the ganglion, but after SNL sympathetics (probably axotomized) invaded from the injured spinal nerve. Based on these findings, we hypothesize that there are two mechanisms for sympathetic sprouting into DRG, differentially dependent on Wallerian degeneration. Analysis of pain behavior in these animals reveals that (i) mechanoallodynia and sympathetic innervation of the DRG tend to coincide and (ii) thermal allodynia and Wallerian degeneration, but not sympathetic innervation of the DRG tend to coincide.     

Gene expression profiling of the rat sciatic nerve in early Wallerian degeneration after injury

Wallerian degeneration is an important area of research in modern neuroscience.A large number of genes are differentially regulated in the various stages of Wallerian degeneration,especially during the early response.In this study,we analyzed gene expression in early Wallerian degeneration of the distal nerve stump at 0,0.5,1,6,12 and 24 hours after rat sciatic nerve injury using gene chip microarrays.We screened for differentially-expressed genes and gene expression patterns.We examined the data for Gene Ontology,and explored the Kyoto Encyclopedia of Genes and Genomes Pathway.This allowed us to identify key regulatory factors and recurrent network motifs.We identified 1 546 differentially-expressed genes and 21 distinct patterns of gene expression in early Wallerian degeneration,and an enrichment of genes associated with the immune response,acute inflammation,apoptosis,cell adhesion,ion transport and the extracellular matrix.Kyoto Encyclopedia of Genes and Genomes pathway analysis revealed components involved in the Jak-STAT,ErbB,transforming growth factor-β,T cell receptor and calcium signaling pathways.Key factors included interleukin-6,interleukin-1,integrin,c-sarcoma,carcinoembryonic antigen-related cell adhesion molecules,chemokine(C-C motif) ligand,matrix metalloproteinase,BH3 interacting domain death agonist,baculoviral IAP repeat-containing 3 and Rac.The data were validated with real-time quantitative PCR.This study provides a global view of gene expression profiles in early Wallerian degeneration of the rat sciatic nerve.Our findings provide insight into the molecular mechanisms underlying early Wallerian degeneration,and the regulation of nerve degeneration and regeneration.     

An 8.5-kb segment of the PMP22 promoter responds to loss of axon signals during Wallerian degeneration, but does not respond to specific axonal signals during nerve regeneration

Altered expression of the PMP22 gene causes Charcot-Marie-Tooth disease type 1A (CMT1A) and hereditary neuropathy with liability to pressure palsies (HNPP). We have examined the promoter activity of 8.5 kb upstream of the first coding exon of the rat peripheral myelin protein-22 (rPmp22) gene in transgenic mice. We found that the -8.5 kb rPmp22/chloramphenicol acetyl transferase (CAT)/beta-galactosidase (lacZ) construct directs reporter gene expression in a weakly developmental and tissue-specific pattern, consistent with the expression pattern of the endogenous Pmp22 gene. The -8.5 kb rPmp22/CAT/lacZ transgene responds to loss of axonal signals during Wallerian degeneration but unlike the endogenous Pmp22 gene, the transgene fails to respond to axonal signals during nerve regeneration after a sciatic nerve crush injury. In conclusion, the function of the -8.5 kb rPmp22/CAT/lacZ transgene suggests that there are separable regulatory elements in the rPmp22 gene that respond differently to axonal signals received by Schwann cells during nerve development, and during remyelination.     

Delayed peripheral nerve degeneration, regeneration, and pain in mice lacking inducible nitric oxide synthase

Inducible nitric oxide synthase (iNOS) may be a critical factor in the repair of injured tissues. In mice lacking iNOS we observed abnormalities in how the peripheral nerve responds to each of 3 fundamental types of injury: chronic constriction partial nerve injury (a model of neuropathic pain), nerve crush, and nerve transection. In each type of injury, mice lacking iNOS had evidence of a regenerative delay, preceded by slowing of myelinated fiber Wallerian degeneration (WD). In wild-type mice, iNOS immunoreactivity and the presence and upregulation of its mRNA were demonstrated distal to injury, but neither was observed in the knockout mice. Slowed WD was suggested by the abnormal persistence of apparent myelinated fiber profiles distal to the injury zones in mice lacking iNOS compared to wild-type controls. In mice lacking iNOS there were fewer regenerating myelinated fibers, smaller caliber regenerating fibers, and slowed reinnervation of muscle endplates distal to the injury zone. Slowed degeneration was also associated with normal initiation but delayed expression of neuropathic pain. Our findings highlight important relationships among nitric oxide, WD, neuropathic pain, and axon regeneration.     

Evidence that the Rate of Wallerian Degeneration is Controlled by a Single Autosomal Dominant Gene

In a substrain of C57BL mice, C57BL/Ola, Wallerian degeneration in the distal segment of the severed sciatic nerve is extremely slow when compared to other mice. Despite this very slow degeneration in the distal segment regeneration of the motor nerves is not impaired. From suitable genetic outcrosses and backcrosses, the authors provide evidence that the rate of Wallerian degeneration in this strain is controlled by a single autosomal gene product. The authors have also shown that the rate of degeneration, in C57BL/Ola mice, is influenced by the environment in which the animals were bred and housed. Wallerian degeneration in the sciatic nerves of mice raised in isolators is slower than in those raised in a conventional animal house. This strain of mouse may prove to be of value in the understanding of nerve degeneration and regeneration.     

Reversible endoneurial changes after nerve injury

Summary Endoneurial changes in the rat sciatic nerve were studied during Wallerian degeneration and subsequent regeneration. After total axotomy two different experimental models were used. In the first the cut ends of the sciatic nerves were left free to allow reinnervation. In the second model the distal end of the transected nerve was sutured to the adjoining muscle to prevent regeneration. Within 2 weeks after the axomoty, a Wallerian type of degeneration was seen with axonal destruction and phagocytosis of myelin sheaths. After 4 weeks endoneurial fibroblastic cells formed circular structures around the Schwann cell columns, i.e., the bands of Buengner in both groups. These fascicle-like structures became more pronounced in the non-regenerating nerves up to 8 weeks, while during reinnervation the cellular reaction in the endoneurium nearly disappeared within this time. Ultrastructurally, the endoneurial fibroblast-like cells showed marked phagocytotic activity and also fragments of basement membrane on their surface. The appearance of thin (25–30 nm in diameter) collagen fibrils closely related to the basement membrane was noted around the bands of Buengner, as well as the appearance of an amorphous extracellular gap between the newly synthetized thin collagen fibrils and normal endoneurial collagen (50–60 nm). The reversible endoneurial compartmentation seems to be important for maintaining the nerve structure, serving as a support for axonal regeneration in addition to the bands of Buengner.Supported in part by grants from the Emil Aaltonen Foundation and from the Research and Science Foundation of Farmos (to V. S.) and institutional grants (to Dept. Med. Chem.) from the Sigrid Jusélius Foundation and the Turku University Foundation