Biomedical subjects

M Schachner

Publications and source records attributed to M Schachner.

At least 343 records · Page 19Linked to original sources

PubMed1989

Binding properties of the neural cell adhesion molecule to different components of the extracellular matrix.

A soluble form of the neural cell adhesion molecule (N-CAM) was obtained from 100,000-g supernatants of crude brain membrane fractions by incubation for 2 h at 37 degrees C. The isolated N-CAM, consisting of one polypeptide chain with a molecular mass of 110 kilodaltons (N-CAM 110), was studied for its binding specificity to different components of the extracellular matrix (ECM). N-CAM 110 bound to different types of collagen (collagen types I-VI and IX). The binding efficiency was dependent on salt concentration and could be called specific according to the following criteria: (a) Binding showed substrate specificity (binding to collagens, but not to other ECM components, such as laminin or fibronectin). (b) Binding of N-CAM 110 to heat-denatured collagens was absent or substantially reduced. (c) Binding was saturable (Scatchard plot analyses were linear with KD values in the range of 9.3-2.0 X 10(-9) M, depending on the collagen type and buffer conditions). Binding of N-CAM 110 to collagens could be prevented in a concentration-dependent manner by the glycosaminoglycans heparin and chondroitin sulfate. N-CAM 110 also interacted with immobilized heparin, and this interaction could be prevented by heparin and chondroitin sulfate. Thus, in addition to its role in cell-cell adhesion, N-CAM is a binding partner for different ECM components, an observation suggesting that it also serves as a substrate adhesion molecule in vivo.

Animals↗

PubMed1989

J1/tenascin-related molecules are not responsible for the segmented pattern of neural crest cells or motor axons in the chick embryo.

It has been suggested that substrate adhesion molecules of the tenascin family may be responsible for the segmented outgrowth of motor axons and neural crest cells during formation of the peripheral nervous system. We have used two monoclonal antibodies (M1B4 and 578) and an antiserum [KAF9(1)] to study the expression of J1/tenascin-related molecules within the somites of the chick embryo. Neural crest cells were identified with monoclonal antibodies HNK-1 and 20B4. Young somites are surrounded by J1/tenascin immunoreactive material, while old sclerotomes are immunoreactive predominantly in their rostral halves, as described by other authors (Tan et al. 1987--Proc. natn. Acad. Sci. U.S.A. 84, 7977; Mackie et al. 1988--Development 102, 237). At intermediate stages of development, however, immunoreactivity is found mainly in the caudal half of each sclerotome. After ablation of the neural crest, the pattern of immunoreactivity is no longer localised to the rostral halves of the older, neural-crest-free sclerotomes. SDS-polyacrylamide gel electrophoresis of affinity-purified somite tissue, extracted using M1B4 antibody, shows a characteristic set of bands, including one of about 230 x 10(3), as described for cytotactin, J1-200/220 and the monomeric form of tenascin. Affinity-purified somite material obtained from neural-crest-ablated somites reveals some of the bands seen in older control embryos, but the high molecular weight components (120-230 x 10(3] are missing. Young epithelial somites also lack the higher molecular mass components. The neural crest may therefore participate in the expression of J1/tenascin-related molecules in the chick embryo. These results suggest that these molecules are not directly responsible for the segmented outgrowth of precursors of the peripheral nervous system.

Animals↗

PubMed1989

Expression of v-src arrests murine glial cell differentiation.

A replication-defective retroviral vector carrying the v-src oncogene and the gene for neomycin resistance was used to infect neurone- and fibroblast-depleted embryonic mouse brain cells in vitro. Cells resistant to the antibiotic G418 were obtained and continually passaged. Several cell lines were isolated which express high levels of v-src mRNA and v-src tyrosine kinase activity. The antigenic marker profile of either the pooled cells from an individual infection or sublines isolated from individual foci showed the cells to be immature glia: most cells expressed vimentin, A2B5 antigen and/or J1/tenascin glycoproteins, but not fibronectin. Sublines expressed different antigen profiles suggesting that the immortalised cells were derived from glial cells of different phenotypes. The cell lines expressed the 120 and 140 but not the 180 kd components of N-CAM as well as voltage-activated potassium channels, typical for glial cells. 01 antigen-positive oligodendrocytes were never observed in the lines or sublines after long term passage (over 1 year), but some cells expressed glial fibrillary acidic protein, a marker for mature astrocytes. Thus, expression of v-src in murine glial cells appears to arrest their development and prevent their differentiation.

Animals↗

PubMed1988

N-linked oligosaccharides are not required for neuron-neuron interactions mediated by neural cell adhesion molecule.

Recent studies have described the role of various regions of the neural cell adhesion molecule (NCAM) in cell-cell interactions. Monoclonal antibodies (L2/HNK-1) directed against a sulfated, glucuronic acid-containing, N-linked carbohydrate epitope have also been shown to inhibit NCAM-mediated neural cell adhesion. In the present study we show that dissociated retinal neurons in an in vitro model system can bind as well to normal NCAM as to NCAM lacking the L2/HNK-1 epitope or to glycopeptidase F-treated NCAM. These data suggest that N-linked oligosaccharide chains do not confer upon NCAM the adhesional properties associated with its role in neuron-neuron interactions.

Amidohydrolases↗

PubMed1988

Adhesion of neural cells to extracellular matrix constituents. Involvement of glycosaminoglycans and cell adhesion molecules.

Single cell suspensions of early postnatal mouse cerebellum adhere to substrate-bound culture supernatants of the teratocarcinoma cell line PF-HR9 and can be inhibited to adhere by antibodies to the neural cell adhesion molecules L1 and N-CAM. Adhesion can also be inhibited by the glycosaminoglycans heparin and heparan sulfate, and less by chondroitin sulfate or hyaluronic acid. Heparinase treatment of cells, but not of HR9 substrate, reduces adhesion. Adhesion does not appear to be mediated by laminin, a constituent of HR9 extracellular matrix, since L1 and N-CAM antibodies do not interfere with cell adhesion on EHS sarcoma laminin as substrate and since antibodies to EHS sarcoma laminin partially inhibit adhesion to HR9 extracellular matrix which contains laminin. Of the other extracellular matrix constituents analysed in HR9 culture supernatants (collagen type IV, a heparan sulfate proteoglycan and fibronectin) none could be shown to promote adhesion, when coated as substrate, suggesting that yet unidentified compounds are responsible for L1- or N-CAM-mediated cell adhesion. These experiments show for the first time that extracellular matrix constituents can act as binding partners for the neural cell adhesion molecules L1 and N-CAM.

Animals↗

PubMed1988

Characterization of Ca2+-dependent and -independent aggregation mechanisms among mouse cerebellar cells.

To gain insight into the cellular and molecular mechanisms underlying cell interactions in the early postnatal mouse cerebellum, Ca2+-dependent and -independent aggregation mechanisms were characterized using single cell suspensions under conditions that allow discrimination between the two mechanisms. When cerebellar cells were derived from newborn to 10-day-old mouse cerebellum, both mechanisms were active and showed no major change in activity during this time period. Mg2+ could not replace Ca2+ in the Ca2+-dependent mechanism. In contrast to the Ca2+-independent mechanisms, the Ca2+-dependent mechanism was inactive at low temperatures, suggesting a necessity for molecular rearrangement within the surface membrane during aggregation. Neuraminidase, chondroitinase, heparinase or hyaluronidase treatment of cells did not influence the aggregation of cells under Ca2+-dependent and -independent conditions. Chondroitin sulfate inhibited and hyaluronic acid stimulated the Ca2+-dependent mechanism, whereas chondroitin sulfate only slightly and hyaluronic acid strongly inhibited the Ca2+-independent one. Dextran sulfate slightly inhibited both mechanisms, whereas heparin and fucoidan, a complex sulfated carbohydrate, did not influence cell aggregation, while they strongly inhibited attachment of cells to laminin. The polycation poly-L-lysine slightly stimulated the Ca2+-independent mechanism, but inhibited the Ca2+-dependent one. Interestingly, chondroitin sulfate and hyaluronic acid strongly stimulated cell aggregation under conditions where both mechanisms were almost destroyed or inactive. Dextran sulfate showed only a small effect under these conditions. These observations indicate that different molecular mechanisms are active in cell-cell versus cell-extracellular matrix interactions and suggest a hitherto unknown complexity in molecular mechanisms during early postnatal cerebellar development.

Aging↗

PubMed1988

Neural adhesion molecule L1 as a member of the immunoglobulin superfamily with binding domains similar to fibronectin.

Diverse glycoproteins of cell surfaces and extracellular matrices operationally termed 'adhesion molecules' are important in the specification of cell interactions during development, maintenance and regeneration of the nervous system. These adhesion molecules have distinct functions involving different cells at different developmental stages, but may cooperate when expressed together. Families of adhesion molecules which share common carbohydrate domains do exist, despite the structural and functional diversity of these glycoproteins. These include the Ca2+-independent neural adhesion molecules: N-CAM, myelin associated glycoprotein (MAG) and L1. L1 is involved in neuron-neuron adhesion, neurite fasciculation, outgrowth of neurites, cerebellar granule cell migration, neurite outgrowth on Schwann cells and interactions among epithelial cells of intestinal crypts. We show here that in addition to sharing carbohydrate epitopes with N-CAM and MAG, L1 is also a member of the immunoglobulin superfamily. It contains six C2 domains and also shares three type III domains with the extracellular matrix adhesion molecule fibronectin.

Amino Acid Sequence↗

PubMed1988

Immunohistological localization of cell adhesion molecules L1, J1, N-CAM and their common carbohydrate L2 in the embryonic cortex of normal and reeler mice.

The expression of the cell adhesion molecules L1, J1 and N-CAM and their shared carbohydrate L2 was studied in the embryonic cerebral cortex of normal and reeler mutant mice using light and electron microscopic immunocytochemistry. Apart from a general delay in their appearance in the reeler cortex, the 4 antigens were present with a cellular distribution in both genotypes reflecting the anatomical characteristics of normal and mutant phenotypes. The cell surface glycoprotein L1 was exclusively expressed by neurons, particularly axons, but was never detected at sites of neuron-glia contact. L1 was accumulated in the marginal zone and subplate of the normal cortex and in the homologous layers of the reeler cortex. The secreted glycoprotein J1 was found on glia and neurons. Although initially present in regions of fiber outgrowth, J1 became characteristically excluded from the large fiber tracts at later stages. J1 mapped in the marginal zone and subcortical plate of the normal cortex and in the corresponding layers of the mutant cortex. N-CAM had a more ubiquitous distribution and was present in ventricular zones, particularly at early stages, as well as on glia and neurons and large fiber tracts at later developmental stages. The distribution of the L2 epitope was quite similar to that of the J1 molecule but remained present on large fiber tracts, like N-CAM and L1, also at later developmental stages. These comparative observations in normal and reeler mutant mice lend support to previous suggestions that L1, together with N-CAM, may play a role in the aggregation of neuronal cell bodies after migration and in the fasciculation of developing fiber bundles. They also point to a possible function of the extracellular matrix component J1 in the guidance or support of fiber outgrowth in large fiber tracts.

Animals↗

PubMed1988

Ultrastructural features of cultured oligodendrocytes expressing stage-specific cell-surface antigens.

This study was designed to correlate cytological features that had previously been established for oligodendrocytes at different developmental stages in vivo and cytological criteria in vitro with the expression of stage-specific cell-surface antigens of cultured oligodendrocytes. Cells obtained from the corpus callosum of 10-day-old C57BL/6J mice were maintained in monolayer cultures and stained with monoclonal antibodies 01 through 012 by indirect immunofluorescence or immunoperoxidase methods. 0 antigen-positive cells were classified according to two criteria: (a) cell shape (type I-III); and (b) cytoplasmic features at the ultrastructural level (class 1-3). Approximately 95% of all 0 antigen-positive cells could be identified as oligodendrocytes by established cytological criteria, thus supporting previous evidence of their glial character from electrophysiological and cell type-specific marker studies. After 12 days in vitro approximately 90% of all morphologically identified oligodendrocytes expressed antigens 03, 04, 05 or 06, which are the first to appear during development in vivo, whereas only 30-40% expressed antigens 011 or 012 which are the last to appear during development in vivo. 01 through 010 antigen-positive oligodendrocytes belong to 3 morphologically distinct cell types: (1) with small (approximately 10 micron in diameter) round cell bodies and few slender processes; (2) with 'hairy eyeball' morphology with a network of processes; and (3) with large, sometimes bipolar cell bodies (up to 30 micron in diameter) surrounded by high amounts of membranous material devoid of cytoplasm. By cytoplasmic criteria at least 90% of all 0 antigen-positive cells fit the description by Mori and Leblond of 'light' to 'medium' oligodendrocytes in vivo, although a clear-cut correlation with expression of early or late appearing 0 antigens was not observed. Typically 'dark' oligodendrocytes were rarely seen in our cultures. 011 and 012 antigen-positive cells are restricted to the group of large oligodendrocytes with high amounts of membranous material, often organized in more or less compact structures (type III). In contrast to the more uniform localization of antigens 01 through 010 over the whole cell surface, antigens 011 and 012 are less strongly detectable on cell bodies than on processes and membranous whirls.

Animals↗

PubMed1988

Stage-specific cell-surface antigens of oligodendrocytes in the peripheral nervous system. Expression during development and regeneration and in myelin-deficient mutants.

Monoclonal antibodies to stage-specific cell surface antigens of oligodendrocytes have been used to investigate the expression of antigens 05 through 011 in the peripheral nervous system of the mouse by immunohistology. In the adult sciatic nerve antigens 05 through 09 and 011 were diffusely positive. 010 antigen was not detectable in the peripheral nervous system at any age tested. During development antigens 05, 06 and 07 were first detectable at birth in tracts at the proximal part of the sciatic nerve. At day 2 the whole diameter of the nerve was positive for 05 antigen, while antigens 06 and 07 were detectable only in part of the nerve and antigens 08 and 09 were just about to appear. At day 4 antigen 011 was the last to appear. At day 7 all antigens were strongly detectable throughout the nerve. After transection of adult sciatic nerve expression of antigens 05 through 09 and 011 was studied at the proximal and distal ends of the cut. Three days after transection all antigens were fully detectable in the degenerating myelin and its debris. After 15 days residual debris was still distinctly positive, while Schwann cells in the bands of Bünger were antigen-negative. At approximately two weeks a connecting bridge between proximal and distal ends of the cut nerve had developed, but the 0 antigens were not detectable in this bridge until day 21. At day 42 all antigens were again fully detectable in the regenerating nerve. In hypomyelinating mouse mutants no difference to the normal control littermates was seen in staining pattern and intensity for jimpy and shiverer, while quaking showed an increase in staining intensity for 05 through 08 antigens. In trembler antigens 05, 06 and 07, but not 08, 09 and 011 appeared associated with non-myelin-forming Schwann cells, while the few recognizable myelin-forming Schwann cells expressed all antigens. These observations show that we have characterized 4 new monoclonal antibodies as further reagents to look at developmentally distinct steps in myelination of the peripheral nervous system.

Animals↗

PubMed1988

Effect of benzodiazepines and pentobarbital on the GABA-induced depolarization in cultured astrocytes.

We have previously shown that cultured astrocytes from neonatal rat cerebral cortex are depolarized by GABA. The underlying ionic mechanism, activation of a Cl- conductance and responses to an agonist and antagonists were found to be similar to those of the neuronal GABAA receptor (Kettenmann et al.: Brain Research 404:1-9, 1987; Kettenmann and Schachner: Journal of Neuroscience 5:3295-3301, 1985). To characterize further the pharmacological properties of the GABA receptor we have tested the influence of pentobarbital and benzodiazepines on the GABA response. Pentobarbital potentiated and prolonged the GABA-induced depolarization and enhanced the velocity of the depolarization. Agonists of the neuronal benzodiazepine receptor, flunitrazepam, diazepam, and midazolam, increased the GABA-induced depolarization. As in neurons, an antagonist of the benzodiazepine receptor, Ro 15-1788, blocked the flunitrazepam-induced enhancement of the GABA response. In contrast to their effects on neurons, the inverse agonists Ro 22-7497 and DMCM increased the GABA-induced depolarization. The ligand of the putative peripheral benzodiazepine binding site, Ro 5-4864, did not show consistent effects on the GABA response. These studies confirm that cultured astrocytes express GABAA receptors. This receptor is similar to the neuronal GABAA receptor with regard to Cl- conductance and its pharmacological responses to muscimol, bicuculline, picrotoxin, pentobarbital, and benzodiazepine agonists and an antagonist, but it is different in its responses to inverse agonists of the benzodiazepine site. The physiological role of the glial GABAA receptor is at present unknown.

Animals↗

PubMed1988

Glutamate opens Na+/K+ channels in cultured astrocytes.

Glial cells from different brain regions and species are depolarized by the neurotransmitter glutamate. The depolarization or, if voltage-clamped at the resting membrane potential, the inward current induced by glutamate could be due either to activation of receptor-coupled ion channels or electrogenic uptake of the transmitter. In the present study we applied the patch-clamp technique in the whole-cell recording mode to analyze glutamate-induced currents in cultured astrocytes from rat cerebral hemispheres. At the resting membrane potential, glutamate induced an inward current ranging from 40 to 300 pA. This current decreased in size with depolarization and reversed at about 0 mV. The resulting current-to-voltage curve was linear and depended strongly on the transmembrane Na+ but not on the Ca++ or Cl- gradient. In the presence of glutamate, current noise increased at potentials positive or negative from the reversal potential indicating that ionic channels are activated by glutamate. Both kainate and quisqualate mimicked the effect of glutamate. We conclude that glutamate opens a Na+/K+ channel in cultured astrocytes because of activation of a receptor which shares many properties with the neuronal kainate/quisqualate receptor.

Animals↗

PubMed1988

Expression and localization of the fibronectin receptor in the mouse nervous system.

Cell surface receptors for extracellular matrix components have recently been characterized as integral membrane complexes with common features in their structural and functional properties. We have investigated the expression of the mammalian fibronectin receptor in the mouse nervous system using immunocytological and immunochemical methods. The fibronectin receptor was detectable on immature oligodendrocytes and immature and mature astrocytes in culture, while central nervous system neurons did not reveal detectable levels of fibronectin receptor at the developmental stages studied. In the peripheral nervous system both glia and neurons were found to express the fibronectin receptor. The receptor complex in both peripheral and central nervous system has an apparent molecular weight of approximately 140 kD under reducing conditions and resolves into two or three distinct protein bands under nonreducing conditions. The fibronectin receptor expresses the L2/HNK-1 epitope that is characteristic of several adhesion molecules, including L1, N-CAM, the myelin-associated glycoprotein, and J1 and thus is another member of the L2/HNK-1 family of adhesion molecules. The L2/HNK-1 carbohydrate epitope is expressed differently and independently of the fibronectin receptor protein backbone in that it is detectable in neonatal brain but not in adult brain. Our observations attribute a functional role to the fibronectin receptor and its L2/HNK-1 carbohydrate epitope during development and maintenance of cell interactions in the central and peripheral nervous systems.

Animals↗

PubMed1988

Immunocytological localization of the major peripheral nervous system glycoprotein P0 and the L2/HNK-1 and L3 carbohydrate structures in developing and adult mouse sciatic nerve.

Immunocytological localization of the major glycoprotein of peripheral myelin P0 and its associated carbohydrate structures L2/HNK-1 and L3 was performed at the light- and electron-microscopic levels in mouse sciatic nerves at several developmental stages and in adulthood. P0 was first expressed on Schwann cells at the time that Schwann cells associated with axons on a 1:1 basis. P0 remains expressed at all times of myelin formation and in compact myelin. After cessation of myelination P0 is no longer detectable in the uncompacted parts of myelin, i.e., Schmidt-Lanterman incisures, paranodal loops, and outer and inner mesaxons. P0 is not detectable on basement membranes, interstitial collagens, and non-myelin-forming Schwann cells. The associated carbohydrate epitope L2 does not follow the expression of P0 at any developmental or adult stage. Until 21 days the L2 epitope is confined to nonmyelinated fibers. In sciatic nerves of mice older than 8 weeks, however, only a few nonmyelinated fibers remain L2-positive. L2 immunoreactivity is clearly seen in a subpopulation of compact myelin figures largely associated with motor fibers. The L3 epitope is never detectable on nonmyelinated fibers and becomes first visible when compact myelin is discerned. Unlike the L2 epitope L3 is present in most, if not all, compact myelin figures. These observations suggest that P0 may be involved in ensheathment of axons by Schwann cells at the decisive stages of initiation of myelination and later on, possibly in conjunction with the L3 carbohydrate structure, in maintenance of compact myelin. The appearance of the L2 carbohydrate epitopes in compact myelin of largely motor and fewer sensory nerve fibers at times when morphogenesis of myelin has ceased remains to be elucidated in functional terms.

Animals↗

PubMed1988

Expression of cell adhesion molecules in the olfactory system of the adult mouse: presence of the embryonic form of N-CAM.

The expression of the neural cell adhesion molecules N-CAM and L1 was investigated in the olfactory system of the mouse using immunocytochemical and immunochemical techniques. In the olfactory epithelium, globose basal cells and olfactory neurons were stained by the polyclonal N-CAM antibody reacting with all three components of N-CAM (N-CAM total) in their adult and embryonic states. Dark basal cells and supporting cells were not found positive for N-CAM total. The embryonic form of N-CAM (E-N-CAM) was only observed on the majority of globose basal cells, the precursor cells of olfactory neurons, and some neuronal elements, probably immature neurons, since they were localized adjacent to the basal cell layer. Differentiated neurons in the olfactory epithelium did not express E-N-CAM. In contrast to N-CAM total, the 180-kDa component of N-CAM (N-CAM180) and E-N-CAM, L1 was not detectable on cell bodies in the olfactory epithelium. L1 and N-CAM180 were strongly expressed on axons leaving the olfactory epithelium. Olfactory axons were also labeled by antibodies to N-CAM180 and L1 in the lamina propria and the nerve fiber and glomerular layers of the olfactory bulb, but only some axons showed a positive immunoreaction for E-N-CAM. Ensheathing cells in the olfactory nerve were observed to bear some labeling for N-CAM total, L1, and N-CAM180, but not E-N-CAM. In the olfactory bulb, L1 was not present on glial cells. In contrast, N-CAM180 was detectable on some glia and N-CAM total on virtually all glia. Glia in the nerve fiber layer were labeled by E-N-CAM antibody only at the external glial limiting membrane. In the glomerular layer, E-N-CAM expression was particularly pronounced at contacts between olfactory axons and target cells. The presence of E-N-CAM in the adult olfactory epithelium and bulb was confirmed by Western blot analysis. The continued presence of E-N-CAM in adulthood on neuronal precursor cells, a subpopulation of olfactory axons, glial cells at the glia limitans, and contacts between olfactory axons and their target cells indicates the retention of embryonic features in the mammalian olfactory system, which may underlie its remarkable regenerative capacity.

Animals↗

PubMed1988

Growth of embryonic retinal neurites elicited by contact with Schwann cell surfaces is blocked by antibodies to L1.

Explants from embryonic rat retina plated on Schwann cell monolayers were used to examine the mechanisms by which these central neurons interact with Schwann cell surfaces. Embryonic retinal explants extend neurites reliably on Schwann cell surfaces (Kleitman et al., 1988, J. Neurosci. 8: 653). Antibodies to molecules thought to be present on Schwann cell surfaces (laminin and the 217C antigen), on retinal neurite surfaces (Thy-1.1), or on both surfaces (L1) were tested for their ability to influence this neurite growth. Of these, only antibodies to L1 were effective in blocking retinal neurite extension on Schwann cells. Inhibition of neurite growth by anti-L1 was shown to be specific to growth on Schwann cell surfaces because neurite growth on air-dried collagen (a substratum known to support retinal neurite outgrowth) was not affected. This blockage was dose-dependent. At a low titer of anti-L1 Fab fragments defasciculation of neurites was prominent; at high titers 95% of neurite outgrowth was inhibited. This virtual elimination of the ability of Schwann cell surfaces to support embryonic retinal neurite growth in the presence of antibodies to L1 indicates that binding of the L1 molecule is a critical component of the mechanism by which Schwann cells foster the growth of these neurites. The present experiments concur with the growing body of evidence that L1 plays an important role in supporting neurite growth on cell surfaces and raise the possibility that L1 may also mediate the striking ability of adult retinal axons to regenerate in a peripheral nerve environment.

Animals↗

PubMed1988

The L2/HNK-1 carbohydrate of neural cell adhesion molecules is involved in cell interactions.

We investigated whether the L2/HNK-1 carbohydrate epitope, expressed by two unusual glycolipids and several neural adhesion molecules, including L1, neural cell adhesion molecule, J1, and the myelin-associated glycoprotein, is involved in adhesion. Monoclonal L2 antibodies, the L2/HNK-1-reactive, sulfate-3-glucuronyl residue carrying glycolipids (L2 glycolipid) and a tetrasaccharide derived from the L2 glycolipid (L2 tetrasaccharide) were added to microexplant cultures of early postnatal mouse cerebellum, and cell migration and process extension were monitored. On the substrate poly-D-lysine, Fab fragments of L2 antibodies, L2 glycolipid, and L2 tetrasaccharide inhibited outgrowth of astrocytic processes and migration of cell bodies, but only L2 glycolipid and L2 tetrasaccharide reduced neurite outgrowth. On laminin, L2 antibodies, L2 glycolipid, and L2 tetrasaccharide inhibited outgrowth of astrocytic processes. Additionally, L2 glycolipid and L2 tetrasaccharide inhibited cell migration and neurite outgrowth. Several negatively charged glycolipids, lipids, and saccharides were tested for control and found to have no effect on outgrowth patterns, except for sulfatide and heparin, which modified outgrowth patterns in a similar fashion as L2 glycolipid and L2 tetrasaccharide. On astrocytes none of the tested compounds interfered with explant outgrowth. In short-term adhesion assays L2 glycolipid, sulfatide, and heparin inhibited adhesion of neural cells to laminin. L2 glycolipid and sulfatide interfered with neuron to astrocyte and astrocyte to astrocyte adhesion, but not with neuron-neuron adhesion. The most straightforward interpretation of these observations is that the L2/HNK-1 carbohydrate and the sulfated carbohydrates, sulfatide and heparin, act as ligands in cell adhesion.

Animals↗

PubMed1988

Immunoelectron microscopic localization of neural cell adhesion molecules (L1, N-CAM, and myelin-associated glycoprotein) in regenerating adult mouse sciatic nerve.

The localization of the neural cell adhesion molecules L1, N-CAM, and the myelin-associated glycoprotein was studied by pre- and postembedding staining procedures at the light and electron microscopic levels in transected and crushed adult mouse sciatic nerve. During the first 2-6 d after transection, myelinated and nonmyelinated axons degenerated in the distal part of the proximal stump close to the transection site and over the entire length of the distal part of the transected nerve. During this time, regrowing axons were seen only in the proximal, but not in the distal nerve stump. In most cases L1 and N-CAM remained detectable at cell contacts between nonmyelinating Schwann cells and degenerating axons as long as these were still morphologically intact. Similarly, myelin-associated glycoprotein remained detectable in the periaxonal area of the degenerating myelinated axons. During and after degeneration of axons, nonmyelinating Schwann cells formed slender processes which were L1 and N-CAM positive. They resembled small-diameter axons but could be unequivocally identified as Schwann cells by chronical denervation. Unlike the nonmyelinating Schwann cells, only few myelinating ones expressed L1 and N-CAM. At the cut ends of the nerve stumps a cap developed (more at the proximal than at the distal stump) that contained S-100-negative and fibronectin-positive fibroblast-like cells. Most of these cells were N-CAM positive but always L1 negative. Growth cones and regrowing axons expressed N-CAM and L1 at contact sites with these cells. Regrowing axons of small diameter were L1 and N-CAM positive where they made contact with each other or with Schwann cells, while large-diameter axons were only poorly antigen positive or completely negative. 14 d after transection, when regrowing axons were seen in the distal part of the transected nerve, regrowing axons made L1- and N-CAM-positive contacts with Schwann cells. When contacting basement membrane, axons were rarely found to express L1 and N-CAM. Most, if not all, Schwann cells associated with degenerating myelin expressed L1 and N-CAM. In crushed nerves, the immunostaining pattern was essentially the same as in the cut nerve. During formation of myelin, the sequence of adhesion molecule expression was the same as during development: L1 disappeared and N-CAM was reduced on myelinating Schwann cells and axons after the Schwann cell process had turned approximately 1.5 loops around the axon. Myelin-associated glycoprotein then appeared both periaxonally and on the turning loops of Schwann cells in the uncompacted myelin.(ABSTRACT TRUNCATED AT 400 WORDS)

Animals↗