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M Schachner

Publications and source records attributed to M Schachner.

At least 199 records · Page 11Linked to original sources
PubMed1995

Detection of tenascin-C in the nervous system of the tenascin-C mutant mouse.

We have investigated the expression of tenascin-C (TN-C) in the somatosensory cortex of early postnatal mutant mice in which lacZ was reported to be expressed in place of tenascin (Saga et al.: Genes Dev 6:1821-1831, 1992). At both the mRNA and protein levels, TN-C was detected at levels lower in the mutant than in wild type animals by in situ hybridization and by immunocytochemistry using several poly- and monoclonal antibodies directed against mouse TN-C. The distribution of TN-C immunoreactivity in coronal sections was abnormal in that the barrel field boundaries in layer 4 of the somatosensory cortex could not be detected intracellularly in most cell bodies, including possibly also neurons. Western blot analysis of homogenates of brain tissue from early postnatal animals showed an abnormal pattern of protein bands immunoreactive for TN-C in mutant animals while beta-galactosidase migrated at its expected molecular weight without incorporation into fusion proteins with TN-C, suggesting disturbed splicing mechanisms. No gross disturbances in the patterning of barrel fields could be detected in the mutant mice as shown by Nissl staining. Our observations show that the mutant mouse designed to be nully disrupted for TN-C expression shows detectable and abnormal TN-C expression.

Animals↗
PubMed1995

Identification of the border between fibronectin type III homologous repeats 2 and 3 of the neural cell adhesion molecule L1 as a neurite outgrowth promoting and signal transducing domain.

To determine the domains of the neural cell adhesion molecule L1 involved in neurite outgrowth, we have generated monoclonal antibodies against L1 and investigated their effects on neurite outgrowth of small cerebellar neurons in culture. When the 10 antibodies were coated as substrate, only antibody 557.B6, which recognizes an epitope represented by a synthetic peptide comprising amino acids 818 to 832 at the border between the fibronectin type III homologous repeats 2 and 3, was as efficacious as L1 in promoting neurite outgrowth, increasing intracellular levels of Ca2+, and stimulating the turnover of inositol phosphates. These findings suggest that neurite outgrowth and changes in these second messengers are correlated. Such a correlation was confirmed by the ability of Ca2+ channel antagonists and pertussis toxin to inhibit neurite outgrowth on L1 and antibody 557.B6. These observations indicate for the first time a distinct site on cell surface-bound L1 as a prominent signal-transducing domain through which the recognition events appear to be funneled to trigger neurite outgrowth, increase turnover of inositol phosphates, and elevate intracellular levels of Ca2+.

Amino Acid Sequence↗
PubMed1995

Tenascin-C mRNA is expressed in cranial neural crest cells, in some placodal derivatives, and in discrete domains of the embryonic zebrafish brain.

A partial zebrafish tenascin-C cDNA clone was isolated from an embryonic zebrafish cDNA library on the basis of homology to mouse tenascin-C. The expression pattern in the head of embryonic zebrafish was analyzed by in situ hybridization. Tenascin-C mRNA was detected in neural crest cells during the period of their migration and differentiation. Expression also occurred in differentiating placodal tissues and in mesodermal cells. In the developing brain, tenascin-C mRNA was expressed in specific domains. In the hindbrain the pattern of the domains was dynamic. At 18 to 22 h postfertilization, expression was widespread in rhombomeres 3, 5, and 6, confined to periventricular cells in rhombomere 2, and not detectable in rhombomere 4. At 32 h postfertilization, tenascin-C was expressed at the rhombomere boundaries. In contrast to the hindbrain, the pattern in the forebrain and midbrain did not show any major changes between 22 and 32 h postfertilization. Domains expressing tenascin-C alternated with regions devoid of it. The most anterior domain of expression was observed at the telencephalic-diencephalic border, surrounding the optic recess. A second domain, at the border between the diencephalon and the midbrain, and a third domain, in the caudal midbrain tegmentum, appeared restricted to the basal plate. Additionally, expression of tenascin-C mRNA was detected in the hypothalamus and in the developing epiphysis. These expression patterns suggest that tenascin-C may play a role in neural crest cell migration and during the differentiation of neural crest, placodal, and mesodermal derivatives. In the developing brain, tenascin-C may be involved in the consolidation of different regional identities.

Amino Acid Sequence↗
PubMed1995

Tenascin-C expression by neurons and glial cells in the rat spinal cord: changes during postnatal development and after dorsal root or sciatic nerve injury.

We have used in situ hybridization with a digoxigenin-labelled probe for tenascin-C mRNA and immunocytochemistry with antibodies against tenascin-C, glial fibrillary acidic protein, OX-42 and the 200 kDa neurofilament protein to study the expression, distribution and cellular relationships of tenascin-C mRNA and protein in the developing (postnatal) and adult spinal cord of rat, and the effects thereon of dorsal root, ventral root and sciatic nerve injuries. The most interesting finding was that on postnatal day 7 (P7), P14 and in the adult, but not on P0 or P3, a group of neurons in the lumbar ventral horn expressed the tenascin-C mRNA gene. They represented about 5% of ventral horn neurons in the adult and were among the smaller such neurons. Since 40-60% of such cells were lost at P13 following sciatic nerve crush on P0, some were almost certainly motor neurons. In addition, we found that at P0 and P3, mRNA-containing glial cells were widespread in grey and white matter but sparse in the developing dorsal columns; tenascin-C immunofluorescence showed a similar distribution. By P7 there were fewer mRNA-containing cells in the ventral horns and in the area of the dorsal columns containing the developing corticospinal tract where immunofluorescence was also weak. At P14 there were no glial-like mRNA-containing cells in the grey matter; such cells were confined to the periphery of the lateral and ventral white columns but were present throughout the dorsal columns where tenascin-C immunofluorescence was also strong. No glial-like mRNA-containing cells were present in the adult lumbar spinal cord and tenascin-C immunofluorescence was confirmed to irregular patches in the ventral horn, especially around immunonegative cell bodies of small neurons, a zone around the central canal, and a thin zone adjacent to the glia limitans. Thus the expression of tenascin-C is differentially developmentally regulated in the grey matter and in different parts of the white matter. Three days after injury of dorsal roots L4-6, many cells containing tenascin-C mRNA, some identified as glial fibrillary acidic protein-positive astrocytes, were present in the ipsilateral dorsal column, but were rare after longer survivals. Immunoreactivity, however, was elevated in the ipsilateral dorsal column at 3 days, remained high for several months and disappeared at 6.5 months. Dorsal root injury had no effect on tenascin-C mRNA or protein in the grey matter. Sciatic nerve or ventral root injury had no effect on these molecules in any part of the spinal cord.

Animals↗
PubMed1995

Tenascin-C expression during wallerian degeneration in C57BL/Wlds mice: possible implications for axonal regeneration.

Schwann cells in the distal stumps of lesioned peripheral nerves strongly express the extracellular matrix glycoprotein tenascin-C. To gain insights into the relationship between Wallerian degeneration, lesion induced tenascin-C upregulation and regrowth of axons we have investigated C57BL/Wlds (C57BL/Ola) mice, a mutant in which Wallerian degeneration is considerably delayed. Since we found a distinct difference in the speed of Wallerian degeneration between muscle nerves and cutaneous nerves in 16-week-old C57BL/Wlds mice, as opposed to 6-week-old animals in which Wallerian degeneration is delayed in both, we chose the older animals for closer investigation. Five days post lesion tenascin-C was upregulated in the muscle branch (quadriceps) but not in the cutaneous branch (saphenous) of the femoral nerve in 16-week-old animals. In addition myelomonocytic cells displaying endogenous peroxidase activity invaded the muscle branch readily whereas they were absent from the cutaneous branch at this time. We could further show that it is only a subpopulation of axon-Schwann cell units (mainly muscle efferents) in the muscle branch which undergo Wallerian degeneration and upregulate tenascin-C at normal speed and that the remaining axon-Schwann cell units (mainly afferents) displayed delayed Wallerian degeneration and no tenascin-C expression. Regrowing axons could only be found in the tenascin-C-positive muscle branch where they always grew in association with axon-Schwann cell units undergoing Wallerian degeneration. These observations indicate a tight relationship between Wallerian degeneration, upregulation of tenascin-C expression and regrowth of axons, suggesting an involvement of tenascin-C in peripheral nerve regeneration.

Animals↗
PubMed1995

Glycans and the modulation of neural-recognition molecule function.

Neural-recognition molecules are carbohydrate-bearing glycoproteins, glycolipids or proteoglycans that are found at the cell surface or in the extracellular matrix that regulate cell interactions during development, modification of synaptic activity and regeneration of nerve connections after damage in the adult. The expression of the carbohydrates appears to be fine tuned to these functions. Among the identified carbohydrates are polysialic acid, a 3'-sulfated glucuronic acid, and oligomannosidic residues. They act not only between apposing partner cell surfaces (trans-interaction) but also between recognition molecules within the surface membrane of one cell (cis-interaction), thereby forming complexes that influence transduction of signals to the cell interior.

Animals↗
PubMed1995

Transient upregulation of NCAM mRNA in astrocytes in response to entorhinal cortex lesions and ischemia.

Axonal sprouting and synaptic reorganization play an important role in the adaptation of the CNS to injury. However, the molecular mechanisms underlying this neuronal plasticity are poorly understood. In the present study we used in situ hybridization to examine the expression of NCAM mRNA in normal hippocampus, and in response to entorhinal cortex (EC) lesions and transient global ischemia. Both neurons and astrocytes were labeled by digoxygenin-tagged cRNA probes which recognize all three major NCAM isoforms of the adult CNS. In contrast, NCAM180-specific probes labeled only neurons in the hippocampus. After unilateral EC lesion, a transient and anatomically restricted upregulation of NCAM120/140 mRNA in reactive astrocytes in the denervated molecular layer of the dentate gyrus was observed. This increase was only present 2-4 days after the lesion whereas the GFAP mRNA increase was present up to 30 days postlesion. Following global ischemia a similar, transient increase of NCAM120/140 mRNA labeling of reactive astrocytes was observed; this increase was anatomically restricted to CA1, where neuronal loss occurred. Results suggest that the transient upregulation of NCAM120/140 mRNA in reactive astrocytes shortly after injury might be an important molecular mechanism in the cascade of events underlying neuronal plasticity in the adult CNS.

Animals↗
PubMed1995

Lack of evidence that myelin-associated glycoprotein is a major inhibitor of axonal regeneration in the CNS.

The MAG-deficient mouse was used to test whether MAG acts as a significant inhibitor of axonal regeneration in the adult mammalian CNS, as suggested by cell culture experiments. Cell spreading, neurite elongation, or growth cone collapse of different cell types in vitro was not significantly different when myelin preparations or optic nerve cryosections from either MAG-deficient or wild-type mice were used as a substrate. More importantly, the extent of axonal regrowth in lesioned optic nerve and corticospinal tract in vivo was similarly poor in MAG-deficient and wild-type mice. However, axonal regrowth increased significantly and to a similar extent in both genotypes after application of the IN-1 antibody directed against the neurite growth inhibitors NI-35 and NI-250. These observations do not support the view that MAG is a significant inhibitor of axonal regeneration in the adult CNS.

3T3 Cells↗
PubMed1995

Protein zero (P0)-deficient mice show myelin degeneration in peripheral nerves characteristic of inherited human neuropathies.

Mutations in the human gene for the myelin recognition molecule protein zero (P0) give rise to severe and progressive forms of dominantly inherited peripheral neuropathies. We have previously reported that mice homozygous for a null mutation in P0 have severely hypomyelinated nerves ten weeks after birth. Here we show hypomyelination already exists at day four with subsequent demyelination and impaired nerve conduction. Furthermore, heterozygous mutants show normal myelination, but develop progressive demyelination after four months of age. Thus, the pathology of homo- and heterozygous P0 mutants resembles that of the severely affected Déjérine-Sottas and the more mildly affected Charcot-Marie-Tooth type 1B patients, respectively.

Animals↗
PubMed1995

Galectin-3, a beta-galactoside-binding animal lectin, binds to neural recognition molecules.

In this study, we have investigated the ability of galectin-3, a beta-galactoside-binding animal lectin, to interact in vitro with different neural tissue-derived glycoproteins involved in cell-cell and cell-substrate adhesion. Galectin-3 interacted to varying degrees with the cell recognition molecules L1, the myelin-associated glycoprotein, and the neural cell adhesion molecule and the extracellular matrix molecules tenascin-C and tenascin-R but not with collagen type I. Binding of galectin-3 to the different glycoproteins tested was carbohydrate dependent and could be specifically inhibited by the addition of lactose and, to a lesser extent, galactose.

Animals↗
PubMed1995

L1 and N-CAM antibodies trigger protein phosphatase activity in growth cone-enriched membranes.

Triggering of the cell adhesion molecules L1 or N-CAM in a nerve growth cone membrane fraction from fetal rat brain with purified L1 or N-CAM or specific antibodies decreases the steady-state levels of protein tyrosine phosphorylation in the membranes. Here we report that triggering of L1 and N-CAM in the growth cone-enriched membrane fraction with a subset of antibodies directed against the extracellular region of L1 and N-CAM elicited dephosphorylation of endogenous protein substrates, indicating the presence of a cell adhesion molecule-activated phosphatase. The most prominent substrates were a membrane-associated 200-kDa protein and tubulin, both of which were dephosphorylated on tyrosine and serine/threonine residues in response to L1 or N-CAM triggering. The antibody-induced phosphatase was inhibited by agents that blocked tyrosine and serine/threonine phosphatases, including sodium orthovanadate, vanadyl sulfate, zinc cations, heparin, and sodium pyrophosphate. Purified L1 and N-CAM fragments and other antibodies reacting with the extracellular region of these adhesion molecules did not activate the phosphatase but did inhibit tyrosine phosphorylation. These properties suggested that triggering of L1 and N-CAM can lead to either phosphatase activation or tyrosine kinase inhibition in growth cone membranes. These findings implicate protein phosphatases in addition to tyrosine kinases as components of L1 and N-CAM intracellular signaling pathways in growth cones.

Amino Acids↗
PubMed1995

The F3 neuronal glycosylphosphatidylinositol-linked molecule is localized to glycolipid-enriched membrane subdomains and interacts with L1 and fyn kinase in cerebellum.

The F3 molecule is a member of the immunoglobulin superfamily anchored to plasma membranes by a glycosylphosphatidylinositol group. In adult mouse cerebellum, F3 is predominantly expressed on a subset of axons, the parallel fibers, and at their synapses. In vitro studies established that it is a plurifunctional molecule that, depending on the cellular context and the ligand with which it interacts, either mediates repulsive interactions or promotes neurite outgrowth. In the present study, we report the isolation of two fractions of F3-containing microdomains from adult cerebellum on the basis of their resistance to solubilization by Triton X-100 at 4 degrees C. Both fractions were composed of vesicles, ranging from 100 to 200 nm in diameter. Lipid composition analysis indicated that the lighter fraction was enriched in cerebrosides and sulfatides. F3 sensitivity to phosphatidylinositol phospholipase C differed between the two fractions, possibly reflecting structural differences in the lipid anchor of the F3 molecule. Both fractions were highly enriched in other glycosylphosphatidylinositol-anchored proteins such as NCAM 120 and Thy-1. It is interesting that these vesicles were devoid of the transmembrane forms (NCAM 180 and NCAM 140), which were recovered in Triton X-100-soluble fractions, but contained the L1 transmembrane adhesion molecule that is coexpressed with F3 on parallel fibers and the fyn tyrosine kinase. Immunoprecipitation experiments indicated that F3, but not NCAM 120 or Thy-1, was physically associated in a complex with both L1 and fyn tyrosine kinase. This strongly suggests that the interaction between L1 and F3, already described to occur with isolated molecules, is present in neural tissue. More important is that our study provides information on the molecular machinery likely to be involved in F3 signaling.

Animals↗
PubMed1995

Characterization of a 21 amino acid peptide sequence of the laminin G2 domain that is involved in HNK-1 carbohydrate binding and cell adhesion.

The N-linked HNK-1 carbohydrate expressed by several recognition molecules mediates the adhesion of early post-natal cerebellar neurons to the G2 domain of the terminal globular domain of the laminin alpha 1 chain (H. Hall et al., submitted). To define this binding site more precisely, G2-derived synthetic peptides were used for binding and competition studies. Peptide 5-G2, comprising the amino acid residues 3431-3451 of G2, inhibited the interaction between the HNK-1-carrying glycolipid and laminin in a concentration-dependent and saturable manner. Peptides which overlap only partially with this sequence interfered less. Peptides comprising other amino acid sequences from G2, and peptides derived from G1 and G3 or a scrambled version of peptide 5-G2, did not show significant effects. Direct binding of peptide 5-G2 to the HNK-1 glycolipid was also demonstrated. Furthermore, peptide 5-G2 interfered in a concentration-dependent and saturable manner with the adhesion of early postnatal cerebellar neurons to laminin. These observations indicate that amino acid residues 3431-3451 of the laminin G2 domain are involved in HNK-1 carbohydrate-mediated cell adhesion.

Amino Acid Sequence↗
PubMed1995

Distinct sites on tenascin-C mediate repellent or adhesive interactions with different neuronal cell types.

In this study we have determined the binding specificities of four different neuronal cell types to tenascin-C (TN-C) and laminin using a cell adhesion assay. TN-C was repulsive for small cerebellar neurons and PC12 phaeochromocytoma cells, since after short-term adhesion to the substrate-bound molecule with a maximum of cell binding at 45 min, the cells detached from the substrate and after 22 h only about 25% of the originally adherent cells were still bound. For N2A neuroblastoma cells and retinal cells TN-C was an adhesive substrate, since the number of adherent cells did not decrease after the initial attachment period. All four cell types adhered well to laminin at all time points studied. For short-term adhesion of small cerebellar neurons and PC12 cells two binding sites were identified on TN-C, one being localized within the epidermal growth factor-like repeats three to five and the second within fibronectin type III-like repeats three and four. One binding site for N2A and retinal cells was localized within fibronectin type III-like repeat seven. Binding of small cerebellar neurons to TN-C was dependent on Ca2+, but not on Mg2+ and was inhibitable by polyclonal antibodies to beta 1 integrin. Short-term adhesion of small cerebellar neurons was also inhibitable with a mixture of recombinant fragments of TN-C encompassing the whole molecule, although the specific inhibitory activity of this mixture was ten-fold lower on a molar basis when compared to the native molecule. Our observations indicate that different neuronal cell types use distinct binding sites on TN-C for repellent or adhesive interactions and that beta 1 integrin is involved in the recognition event leading to repulsion of small cerebellar neurons.

Animals↗
PubMed1995

Focal brain injury and upregulation of a developmentally regulated extracellular matrix protein.

Tenascin is an extracellular matrix glycoprotein expressed during both normal development and neoplastic growth in both neural and nonneural tissues. During development of the central nervous system (CNS), tenascin is synthesized by glial cells, in particular by immature astrocytes, and is concentrated in transient boundaries around emerging groups of functionally distinct neurons. In the mature CNS, only low levels of the glycoprotein can be detected. The present study demonstrates that following trauma to the adult human cerebral cortex, discrete populations of reactive astrocytes upregulate their expression of tenascin and dramatically increase their transcription of the tenascin gene. The enhanced expression of tenascin may be involved in CNS wound healing, and may also affect neurite growth within and around a brain lesion.

Adult↗
PubMed1995

Mice doubly deficient in the genes for P0 and myelin basic protein show that both proteins contribute to the formation of the major dense line in peripheral nerve myelin.

In search for the molecular mechanisms underlying the formation of the major dense line in peripheral nerve myelin we investigated mice deficient in the myelin proteins P0 and MBP. In mice lacking both molecules axons were enwrapped by myelin-like processes devoid of the major dense line, while mice deficient in either protein showed, respectively, partial and normal compaction. Mice heterozygous for P0 but devoid of MBP showed myelin of reduced thickness around axons of normal caliber. Both molecules thus contribute to the formation of the major dense line and to the determination of myelin thickness. Furthermore, our observations modify the view that axon caliber is dependent on normal myelin.

Animals↗
PubMed1994

Training in chicks alters PSA-N-CAM distribution in forebrain cell membranes.

The intermediate and medial hyperstriatum ventrale (IMHV) in the chick is involved in memory formation following one-trial passive avoidance training. Possible links between neural cell adhesion molecule (N-CAM) distribution and memory consolidation were examined in an immunoelectron microscope study of IMHV 6 h after training. An antibody against alpha 2,8 polysialic acid (PSA), characteristic for embryonic N-CAM, and one against the protein backbones of N-CAM were labelled (post-embedding), separately, with 15 nm immunogold, and binding to their epitopes was analysed using the statistics of point processes. No difference in labelling levels between control and trained chicks, for either antibodies, was found, both groups of birds showing that cell membrane regions are 10-30 times more enriched in N-CAM isoforms, and have a two-fold greater proportion of PSA-N-CAM, than tissue as a whole. However, 375-400 nm regular arrays of labelled PSA-N-CAM were revealed statistically in cell membranes of control, but not trained, chicks, which may be related to the possible involvement of such membrane PSA domains in memory-related neuroplasticity.

Animals↗