Notochord is essential for oligodendrocyte development in Xenopus spinal cord

Oligodendrocyte precursors originate in the ventral ventricular zone of the developing spinal cord. To examine whether the notochord is essential for the development of oligodendrocytes in Xenopus spinal cord the notochord was prevented from forming, ablated, or transplanted during early stages of development. Differentiated oligodendrocytes did not appear in spinal cord regions lacking a notochord in animals in which notochord failed to develop after UV irradiation at the one-cell stage. Similarly, differentiated oligodendrocytes were not detected in the spinal cord adjacent to the site of segmental notochord ablation at embryonic or larval stages. Transplantation of an additional notochord dorsal to the spinal cord induced the premature appearance of differentiated oligodendrocytes in adjacent lateral and dorsal spinal cord white matter. These results indicate that the development of Xenopus spinal cord oligodendrocytes is dependent on local influences from the notochord and suggest that the notochord is essential for oligodendrocyte development in Xenopus spinal cord.     

Failure of spinal cord oligodendrocyte development in mice lacking neuregulin

Oligodendrocytes develop from a subpopulation of precursor cells within the ventral ventricular zone of the spinal cord. The molecular cues that direct this spatially and temporally restricted event seem to originate in part from structures ventral to and within the spinal cord. Here, we present evidence that the family of ligands termed neuregulins are necessary for the normal generation of mouse spinal cord oligodendrocytes. Oligodendrocytes mature in spinal cord explants from wild-type mice and mice heterozygotic for a null mutation in the neuregulin gene (NRG +/-) in a temporal sequence of developmental events that replicates that observed in vivo. However, in spinal cord explants derived from mice lacking neuregulin (NRG -/-), oligodendrocytes fail to develop. Addition of recombinant neuregulin to spinal cord explants from NRG -/- mice rescues oligodendrocyte development. In wild-type spinal cord explants, inhibitors of neuregulin mimic the inhibition of oligodendrocyte development that occurs in NRG -/- explants. In embryonic mouse spinal cord, neuregulins are present in motor neurons and the ventral ventricular zone where they likely exert their influence on early oligodendrocyte precursor cells.     

The chemokine growth-regulated oncogene-alpha promotes spinal cord oligodendrocyte precursor proliferation

Chemokines, (chemotactic cytokines) are a family of regulatory molecules involved in modulating inflammatory responses. Here we demonstrate that the chemokine growth-regulated oncogene-alpha (GRO-alpha) is a potent promoter of oligodendrocyte precursor proliferation. The proliferative response of immature spinal cord oligodendrocyte precursors to their major mitogen, platelet derived growth factor (PDGF), is dramatically enhanced by GRO-alpha present in spinal cord conditioned medium. One source of GRO-alpha is a subset of spinal cord astrocytes. Cultures of astrocytes contain GRO-alpha mRNA and protein and secrete biologically active concentrations of GRO-alpha. In postnatal spinal cord white matter the location of GRO-alpha-immunoreactive cells is developmentally regulated: GRO-alpha+ cells first appear in ventral and later in dorsal spinal cord white matter. These results suggest that localized proliferation of oligodendrocytes is mediated by synergy between PDGF and GRO-alpha.     

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The reaction of oligodendrocytes in response to traumatic injury of the CNS are poorly understood. In the present report we studied changes in the expression of a major constituent of CNS myelin, myelin basic protein (MBP), by immunohistochemistry and in situ hybridization from 6 h up to 2 weeks following partial transection of the spinal cord in adult rats. MBP immunohistochemistry showed degeneration of myelin at the lesion center and signs of myelin breakdown in necrotic foci in the dorsal and ventral funiculi proximal and distal to the lesion. In situ hybridization revealed that mRNA for MBP was downregulated at the local lesion site within the first day following injury, probably reflecting oligodendrocytes to undergo cell death. From 2 days on, however, MBP mRNA was conspicuously upregulated at the border of the lesion area. This "reactive" response of surviving oligodendrocytes, as indicated by increased levels of MBP mRNA, peaked around 8 days. At this time, oligodendrocytes displaying strong MBP in situ signal formed stripe-like structures which were oriented radially toward the lesion center and arranged in parallel to neurofilament-positive axons. At around 2 weeks post-injury, MBP mRNA at the border of the lesion area was again downregulated to levels comparable to uninjured controls. These results show that traumatic injury of the spinal cord induces a "reactive" response of surviving oligodendrocytes adjacent to lesion sites. This response might represent an important component of local repair mechanisms.     

Effects of neurotransplants and BDNF on the survival and regeneration of injured adult spinal motoneurons

We compared the effects of peripheral nerve grafts, embryonic spinal cord transplants and brain-derived neurotrophic factor (BDNF) on the survival and axon regeneration of adult rat spinal motor neurons undergoing retrograde degeneration after ventral root avulsion. Following implantation into the dorsolateral funiculus of the injured spinal cord segment, neither a peripheral nerve graft nor a combination of peripheral nerve graft with embryonic spinal cord transplant could prevent the retrograde motor neuron degeneration induced by ventral root avulsion. However, intrathecal infusion of BDNF promoted long-term survival of the lesioned motor neurons and induced abundant motor axon regeneration from the avulsion zone along the spinal cord surface towards the BDNF source. A combination of ventral root reconstitution and BDNF treatment might therefore be a promising means for the support of both motor neuron survival and guided motor axon regeneration after ventral root lesions.     

Tissues exhibiting inhibitory [correction of inhibiory] and repulsive activities during the initial stages of neurite outgrowth from the dorsal root ganglion in the chick embryo

To elucidate the mechanisms underlying the projection of dorsal root ganglion (DRG) axons into the dorsal root entry zone in the dorsolateral region of the spinal cord, we examined tissue interactions which affect neurite outgrowth from DRG. We cultured explants or dissociated cells of DRG from embryonic day 4 (E4) chick embryos in combination with E3 spinal cord, notochord, and dermomyotome in three-dimensional collagen gels. The ventral spinal cord, notochord, and dermomyotome, which are located close to the initial projection pathway of DRG but do not receive direct innervation, strongly inhibited DRG neurite outgrowth and repelled DRG neurites. These inhibitory/repulsive cues appear diffusible in nature, because this activity was observed in the absence of direct contacts between tissue explants and DRG neurites. Furthermore, in heterochronic cultures, E9 DRG lost its responsiveness to inhibitory/repulsive factors from E3 ventral spinal cord, while retaining responsiveness to E3 notochord and dermomyotome, suggesting that the E3 ventral spinal cord may secrete a different inhibitory/repulsive signal than notochord and dermomyotome. Putative inhibitory/repulsive signals secreted from tissues along the axonal pathway may serve to guide growing DRG axons to the dorsal root entry zone.     

Sensory axons are guided by local cues in the developing dorsal spinal cord

During development, different classes of sensory neurons establish distinctive central projections within the spinal cord. Muscle spindle afferents (Ia fibers) grow ventrally through the dorsal horn to the ventral cord, whereas cutaneous sensory collaterals remain confined to the dorsal horn. We have studied the nature of the cues used by Ia fibers in establishing their characteristic projections within the dorsal horn. An organotypic culture preparation of embryonic chicken spinal cord and sensory ganglia was used to test the influence of ventral spinal cord and local cues within the dorsal spinal cord on the growing Ia afferents. When the ventral half of the spinal cord was replaced with an inverted duplicate dorsal half, Ia fibers entering through the dorsal columns still grew ventrally within the host dorsal horn. After the fibers entered the duplicate dorsal half, they continued growing in the same direction. With respect to the duplicate dorsal tissue, this was in an opposite, ventral-to-dorsal, direction. In both cases, however, Ia collaterals remained confined to the medial dorsal laminae. Restriction to these laminae was maintained even when the fibers had to change their direction of growth to stay within them. These results show that cues from the ventral cord are not required for the development of correct Ia projections within the dorsal horn. Local, rather than long-range directional, cues appear to determine the pattern of these projections. When the ventral half of the spinal cord was left intact but sensory axons were forced to enter the dorsal gray matter growing rostrally or caudally, their collateral axons grew in random directions, further showing the absence of directional cues even when the ventral cord was present. Taken together, these observations suggest that Ia fibers are guided by local positional cues that keep them confined to the medial gray matter within the dorsal horn, but their direction of growth is determined primarily by their orientation and position as they enter the dorsal gray matter.     

Apoptosis of microglia and oligodendrocytes after spinal cord contusion in rats

Following spinal cord contusion in the rat, apoptosis has been observed in the white matter for long distances remote from the center of the lesion and is primarily associated with degenerating fiber tracts. We have previously reported that many of the apoptotic cells are oligodendrocytes. Here we show that the oligodendrocyte death is maximal at 8 days postinjury and suggest that loss of oligodendrocytes may result in demyelination of axons that have survived the initial trauma. There are two mechanisms that may account for the observed oligodendrocyte apoptosis. The apoptotic cell death may result from the loss of trophic support after axonal degeneration or it may be the consequence of microglial activation. The hypothesis that oligodendrocyte apoptosis is secondary to microglial activation is supported by our observations of microglia with an activated morphology in the same regions as apoptosis and apparent contact between some of the apoptotic oligodendrocytes and microglial processes. In addition to oligodendrocyte apoptosis, a subpopulation of microglia appears to be susceptible to apoptotic cell death as well, as evidenced by the presence of apoptotic bodies in OX42 immunopositive profiles. Thus, the population of apoptotic cells following spinal cord contusion is comprised of oligodendrocytes and putative phagocytic microglia or macrophages. Given the delayed time course of oligodendrocyte death, the apoptotic death of oligodendrocytes may be amenable to pharmacological intervention with subsequent improvement in functional recovery.     

Improving the nutrition of refugees and displaced people in Africa: workshop in Machakos, Kenya, 5-7 December 1994

We report an unusual post-traumatic spinal cord herniation, which became symptomatic 38 years after the trauma. A 44-year-old man presented with a 2-year history of increasing impotence, neuropathic bladder dysfunction and dissociated sensory loss below the level of T6. At the age of 6 years he had a severe blunt spinal injury with transient paraparesis. MRI revealed right lateral and ventral displacement of the spinal cord at the T5/6 level. The spinal cord was surgically exposed and found to herniate through a ventral defect of the arachnoid membrane and the dura mater. As there were no other events that could have precipitated spinal cord herniation the reported blunt trauma in childhood is the most likely cause for the spinal cord herniation in this patient.     

Pharmacology of descending noradrenergic systems in relation to motor function

The function of descending noradrenergic systems in the spinal ventral horn has not been fully elucidated. We have reviewed our own findings and those of others relating to motor function of these noradrenergic systems. We studied the effects of adrenergic drugs on spinal reflexes, decerebrate rigidity, and noradrenaline release from the spinal cord in rats, and motoneuron activity in spinal cord slices isolated from adult rats. It was shown that the descending noradrenergic systems were facilitatory to the motor system, and that alpha 1-antagonistic action at the spinal cord and alpha 2-agonistic action at the brainstem inhibited spinal motor activity by blocking spinal alpha 1-receptors and by reducing the release of noradrenaline in the spinal cord, respectively.     

Local uncoupling of the cerebrovascular and metabolic responses to somatosensory stimulation after neuronal nitric oxide synthase inhibition

In order to investigate the remyelinating potential of mature oligodendrocytes in vivo, we have developed a model of demyelination in the adult rat spinal cord in which some oligodendrocytes survive demyelination. A single intraspinal injection of complement proteins plus antibodies to galactocerebroside (the major myelin sphingolipid) resulted in demyelination followed by oligodendrocyte remyelination. Remyelination was absent when the spinal cord was exposed to 40 Grays of x-irradiation prior to demyelination, a procedure that kills dividing cells. Quantitative Rip immunohistochemical analysis revealed a similar density of surviving oligodendrocytes in x-irradiated and nonirradiated lesions 3 days after demyelination. Rip and bromodeoxyuridine double immunohistochemical analysis of demyelinated lesions indicated that Rip+ oligodendrocytes did not divide as an acute response to demyelination. Oligodendrocytes were also identified by Rip immunostaining and electron microscopy at late time points (3 weeks) within x-irradiated areas of demyelination. These oligodendrocytes extended processes that engaged axons, and on occasion formed myelin membranes, but did not lay down new myelin sheaths. These studies demonstrate that (a) oligodendrocytes that survive within a region of demyelination are not induced to divide in the presence of demyelinated axons, and (b) fully-differentiated oligodendrocytes are therefore postmitotic and do not contribute to remyelination in the adult CNS.     

Hindshaker, a novel myelin mutant showing hypomyelination preferentially affecting the spinal cord

Animals with spontaneous mutations affecting myelin formation have provided useful information about the genetic and cellular mechanisms regulating normal and abnormal myelination. In this paper we describe a novel murine mutation termed hindshaker (hsh), which is inherited in an autosomal recessive manner. Affected mice are characterised by a variable tremor of the hind end which commences at about 2 weeks of age and largely disappears in animals older than 6 weeks. There is hypomyelination affecting predominantly the spinal cord, although the optic nerves and brain are involved to a much lesser degree. The defect of thinly myelinated and naked axons is maximal at 20 days of age and largely resolves with time so that in the adult most axons are myelinated. The myelin structure appears normal and immunostains for the major proteins. Although the distribution of oligodendrocytes in the spinal cord is similar to normal during the period of hypomyelination, there are fewer mature cells. The hsh mutation appears to delay the maturation of oligodendrocytes, particularly in the spinal cord. Additionally, there is a considerable variation in phenotypic expression and in penetrance when the mutation is expressed on different genetic backgrounds, suggesting the hsh locus is subject to the influence of modifying gene(s). Identification of the hsh gene should identify a factor important in the development of oligodendrocytes, particularly those in the spinal cord.     

Regeneration of lesioned corticospinal tract fibers in the adult rat spinal cord under experimental conditions

The absence of fiber regrowth in the injured spinal cord and brain is influenced by several different factors and mechanisms. Among these are factors which inhibit neurite growth which are found on the surface of oligodendrocytes and central myelin. Their neutralization by a specific antibody allowed regeneration of transected corticospinal tract fibers in the adult rat spinal cord. Using a recently introduced novel neuroanatomical tracer, biotin-dextran-amine, we demonstrate the extensive regenerative sprouting of lesioned corticospinal fibers in the lesioned adult spinal cord. In the presence of the antibody against the myelin-associated neurite growth inhibitors, some of these fibers grew over remaining tissue bridges into the caudal spinal cord. They branched extensively in the lumbar spinal cord segments. These branches were decorated with synapse-like boutons. This neuroanatomical configuration probably contributes importantly to the functional recovery observed earlier in these antibody-treated animals.     

Vitamin D metabolism is altered in Asian Indians in the southern United States: a clinical research center study

In order to define precisely the relation between descending monoaminergic systems and the motor system, we measured in the ventral horn of spinal cord of adult rats the variations of extracellular concentrations of 5-HT, 5-HIAA, DA and MHPG. Measurements were performed during rest, endurance running on a treadmill, and a post-exercise period, with microdialysis probes implanted permanently for 45 days. We found a slight decrease in both 5-HT and 5-HIAA during locomotion with a more marked decrease during the post-exercise period compared to the mean of rest values. In contrast, the concentration of DA and MHPG increased slightly during the exercise and decreased thereafter. These results, when compared with those of a previous study, which measured monoamines in the spinal cord white matter [C. Gerin, D. Bécquet, A. Privat, Direct evidence for the link between monoaminergic descending pathways and motor activity: I. A study with microdialysis probes implanted in the ventral funiculus of the spinal cord, Brain Res. 704 (1995) 191-201], highlight the complex regulation of the release of monoamines that occurs in the ventral horn.     

Three-dimensional analysis of the vascular system in the rat spinal cord with scanning electron microscopy of vascular corrosion casts. Part 1: Normal spinal cord

Vascular corrosion casts of polyester resin in the normal spinal cord at C4-C6 and C7-T1 were inspected three-dimensionally by scanning electron microscopy in 13 rats. Arteries and veins were easily differentiated by the impression pattern of endothelial nuclei on the casts. The centrifugal arterial system from the sulcal arteries supplied most of the gray and white matter in the ventral and lateral spinal cord. Each sulcal artery supplied only one side of the cord. The average number of sulcal arteries was 2.6 per mm. The centripetal arterial system from the posterior spinal arteries fed the posterior gray and white matter. In contrast with classical concepts, there was no pial arterial plexus on the ventral and ventrolateral surface except for infrequent transverse branches from the anterior spinal artery. In the posterior columns, two types of large veins were identified: the posterior medial septal veins and the posterior oblique veins that drained the posterior columns, medial posterior gray matter, and posterior gray commissure. The remainder of the gray and white matter was drained by the sulcal veins and the radial veins. This method clearly demonstrates the three-dimensional structure of both the arterial and venous system in the rat spinal cord.     

Effects of hypoxia on the ventral root motor-evoked potential in the in vitro spinal cord preparation

STUDY DESIGN: This study investigated the effects of hypoxia and glucose on motor functions of spinal cord, monitoring ventral root motor-evoked potential in the in vitro cervical cord preparations. OBJECTIVE: To study ischemia-induced changes in ventral root motor-evoked potential of spinal cord. SUMMARY OF BACKGROUND DATA: Previous studies demonstrated ischemic changes caused by local circulatory impairment might be a major pathophysiologic basis of neuron damage in cord compression. METHODS: Ventral root motor-evoked potential elicited by stimulation of ventrolateral funiculus was recorded from the ventral root in the isolated spinal cord preparations obtained from a newborn rat. The preparations were exposed to artificial cerebrospinal fluid equilibrated with severe or mild hypoxia for 90 minutes. Inhibitory and excitatory neurotransmitter antagonists were added to artificial cerebrospinal fluid to investigate synaptic transmission. The artificial cerebrospinal fluids containing various concentrations of glucose were used to study the glucose's effects. RESULTS: Ventral root motor-evoked potential consisted of the early and late components, which were excitatory transsynaptic potentials. The amplitudes were increased in the early phase of severe hypoxia and declined in the prolonged exposure. In mild hypoxia, there was a sustained increase of the amplitudes. The application of inhibitory neurotransmitter antagonists abolished the augmentation of the amplitudes in the early phase of severe hypoxia. Hypoxia without glucose accelerated hypoxic change. CONCLUSION: Inhibitory synaptic transmission was depressed preferentially in the early phase of severe hypoxia or in mild hypoxia. Excitatory and inhibitory transmissions were suppressed in prolonged severe hypoxia. Glucose deficiency aggravated hypoxic inhibition of synaptic transmissions.     

Neurofilament phosphorylation is increased in ventral horn neurons of neonatal rat spinal cord exposed to cerebrospinal fluid from patients with amyotrophic lateral sclerosis

Aberrant neurofilament (NF) phosphorylation in the soma of the ventral horn neurons of neonatal rat spinal cord is observed following exposure to cerebrospinal fluid (CSF) of patients suffering from Amyotrophic Lateral Sclerosis (ALS). CSF samples from ALS and non-ALS neurological patients were injected into the spinal subarachnoid space of 3 day old rat pups. After 48 h, sections of spinal cords were stained for the presence of phosphorylated NF epitopes with SMI-31 antibody. The number of neuronal soma staining with this antibody in the ventral and dorsal horns sides of the spinal cord was counted. There was a significant 3-fold increase in the number of soma stained with SMI-31 antibody in the ventral horns of rat spinal cords exposed to CSF of patients with ALS compared to cords from rats exposed to CSF of non-ALS patients and those which were not exposed to any CSF samples. Such an increase in staining of neuronal soma was not observed in the dorsal horns. Hyperphosphorylation of neuronal soma suggests an initial stage of degenerative changes occurring in the motor (ventral horn) neurons following exposure to circulating factor(s) in the CSF of patients with ALS.     

Density-dependent feedback inhibition of oligodendrocyte precursor expansion

The myelin sheath in the vertebrate CNS is formed by oligodendrocytes. The number of oligodendrocytes in a mature axon tract must be sufficient to myelinate all appropriate axons. How the number of oligodendrocytes is matched to axonal requirements and whether such matching involves axon-oligodendrocyte signaling or intrinsic oligodendrocyte self-regulation are not clear. Using a combination of in vitro analyses, we demonstrate that oligodendrocyte precursors closely regulate their numbers through interactions between adjacent precursors. In low-density rat spinal cord cultures, the number of oligodendrocyte lineage cells increases rapidly. The addition of large numbers of oligodendrocyte precursors substantially reduces precursor expansion and results in a normalization of oligodendrocyte lineage cell numbers in the cultures over time. Thus, the number of oligodendrocyte lineage cells that develop appears dependent on the density of oligodendrocyte lineage cells. This normalization of cell number is reflected in assays of clonal potential and proliferation. For example, precursors gave rise to fewer progeny and proliferated less at high density. Reduced precursor expansion at high density was not attributable to the depletion of growth factors. Cocultures of high and low densities did not inhibit precursor expansion in low-density cultures, suggesting the requirement for local cell-cell interactions. The inhibition of precursor expansion was cell-type-specific and dependent on the presence of oligodendrocyte lineage cells. We propose that this density-dependent feedback inhibition of oligodendrocyte precursor expansion may play a primary role in regulating the number of oligodendrocytes in the developing spinal cord.