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M Schachner

Publications and source records attributed to M Schachner.

At least 433 records · Page 24Linked to original sources
PubMed1983

Coupling among identified cells in mammalian nervous system cultures.

Cell coupling can be demonstrated among groups of oligodendrocytes, astrocytes, and fibroblasts or fibroblast-like cells, but not for dorsal root ganglion neurons, Schwann cells, or macrophages in mouse or rat cell cultures. Neurons were identified by their ability to generate action potentials. Non-neuronal cells were recognized immunologically by their ability to react with a variety of rhodamine-labeled cell type-specific antibodies. Intracellular injection of the gap junction permeable fluorescent dye Lucifer Yellow and ionic current were used to establish the presence of coupling among these identified cells. Coupling under the culture conditions used in this study does not represent a random membrane interaction between closely apposed cells but rather a form of communication among restricted populations.

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PubMed1983

Exclusive potassium dependence of the membrane potential in cultured mouse oligodendrocytes.

Membrane potential, conductance, and intracellular potassium concentration were measured in oligodendrocytes in 3- to 10-week-old cultures of embryonic mouse spinal cord. After intracellular recording the cells were first injected with Lucifer Yellow and then stained by immunofluorescence using rhodamine-labeled monoclonal antibody 01 specific for oligodendrocyte cell surfaces. The membrane potential of these identified oligodendrocytes was in mV -66 +/- 4.3 SD; it could be reversibly reduced almost to zero by the addition of ouabain. Changes in external K+ but not Na+, Ca++, or Cl- changed the membrane potential. A 10-fold increase in extracellular potassium concentration ([K+]0) depolarized the cell by about 52 mV. This is less than the 61 mV predicted by the Nernst equation for a K+ electrode assuming a constant intracellular potassium concentration ([K+]i). However, when [K+]i was measured with an ion-selective electrode during the increase in [K+]0 it was found to rise. The Nernst equation for K+ accurately predicts the oligodendrocyte membrane potential when the increase in [K+]i is taken into account. Oligodendrocytes may be described as accurate K+ electrodes with a variable reference solution.

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PubMed1982

Immunocytochemical cell identification in nervous system culture combined with intracellular injection of a blue fluorescing dye (SITS).

SITS, a stilbene isothiocyanate derivative, was injected by iontophoresis into neurons and oligodendrocytes in cultures of mouse spinal cord. The dye readily stains the entire cell. When excited with ultraviolet light it emits a blue fluorescence. Thus, SITS-injected cells may be distinguished from those injected with Lucifer yellow CH. Furthermore, the blue cells can be identified in culture after the cells have been labeled with two different antibodies using rhodamine (TRITC) or fluorescein (FITC) as fluorochromes by the use of appropriate filter combinations.

4-Acetamido-4'-isothiocyanatostilbene-2,2'-disulfo↗
PubMed1982

Single potassium channel currents in cultured mouse oligodendrocytes.

Single channel currents were recorded from isolated membrane patches of cultured mouse oligodendrocytes using the giga-seal technique. The observed conductance of the channel was 71 +/- 34 pS. Isolated patches contained 1 to 4 channels with similar conductances and kinetics. Closed times of the channel varied from less than 1 msec to many minutes. The open state was always interrupted by flickering to the closed state. The kinetics of opening and closing appeared insensitive to voltage steps of up to +/- 75 mV from the resting level of the membrane potential, but could be affected by very large voltage steps. The observed changes in channel current in response to changes of potassium concentration on either side of the membrane indicate a high selectivity for potassium. The results show a membrane with constant macroscopic permeability that contains channels which open and close.

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PubMed1982

Cell type-specificity of epidermal growth factor (EGF) binding in primary cultures of early postnatal mouse cerebellum.

Binding of 125I-labeled epidermal growth factor (EGF) to cells of cultured early postnatal mouse cerebellar cells was investigated by autoradiography in conjunction with cell type-specific immunolabeling of neurons, astrocytes and oligodendrocytes. By use of tetanus toxin for recognition of neurons and glial fibrillary acidic (GFA) protein and 04 antigen as markers for astrocytes and oligodendrocytes, respectively, it could be shown that oligodendrocytes do not express receptors for EGF within the limits of sensitivity of the autoradiographic method, and that less than 1% of all small neurons (mostly granule cells) and 50-90% of all GFA protein-sensitive astrocytes show detectable levels of EGF binding.

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PubMed1982

Cultivation of immature astrocytes of mouse cerebellum in a serum-free, hormonally defined medium. Appearance of the mature astrocyte phenotype after addition of serum.

A hormonally defined culture medium is described which supports survival and proliferation of astroglia from primary cultures of early postnatal mouse cerebellum. This medium consists of bovine serum albumin, insulin, transferrin, selenium, hyaluronic acid, protease inhibitor aprotinin, and epidermal growth factor. Trypsin-dissociated single cerebellar cell suspensions are plated in this medium on poly-l-coated glass coverslips and maintained for two weeks before subcultivation. After subcultivation into defined medium more than 99% of all cells are vimentin-positive and fibronectin- and almost completely glial fibrillary acid (GFA) protein-negative, indicating that these cells are less mature astrocytes. After replacement of defined medium by culture medium containing 10% horse serum, expression of GFA protein is detectable in addition to vimentin by indirect immunofluorescence.

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PubMed1982

Cell that are O4 antigen-positive and O1 antigen-negative differentiate into O1 antigen-positive oligodendrocytes.

In freshly dissociated viable cells of early postnatal mouse cerebellum, O1 antigen-positive oligodendrocytes have been eliminated by complement dependent immunocytolysis. Before seeding residual cells in culture, O4 antigen-positive cells were immunolabeled by O4 antibody which had been directly conjugated with rhodamine. After various periods of time in culture, cells were treated with fluorescein conjugated O1 antibody, in order to assay for the appearance of O1-positive oligodendrocytes. After 6.5 h in vitro, the first cells carrying both the previously applied rhodamine label and the freshly applied fluorescein label were seen. The simultaneous appearance of both labels indicates tha O4-positive and O1-negative cells are direct precursors of O1 antigen-positive oligodendrocytes.

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PubMed1982

Immunoelectron microscopic characterization of galactocerebroside and nervous system antigen-1 (NS-1) positive oligodendrocytes in culture.

Immunoelectron microscopy was performed on live, cultured early postnatal mouse cerebellar cells to identify and ultrastructurally characterize those cells which express nervous system antigen-1 (NS-1) and beta-D-galactocerebroside (GalCer) on their surfaces. These two antigens are expressed on oligodendroglia with similar ultrastructural features. These cells are mainly of the immature type, possessing numerous polysomes, mitochondria and microtubules. Filaments are not observed. These oligodendroglia are sometimes surrounded by NS-1 and GalCer-positive membranous material, Few (less than 5%) mature oligodendrocytes are present in the dissociated cerebellar cultures.

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PubMed1982

Immunoselection of oligodendrocytes by magnetic beads. I. Determination of antibody coupling parameters and cell binding conditions.

Oligodendrocytes from early postnatal mouse cerebellum were isolated using polyacrylamide-coated magnetic beads carrying monoclonal antibody to 04 cell surface antigen. Oligodendrocytes were enriched to a purity of 91 +/- 4% starting from a mixed cell population containing approximately 1.5% antigen-positive oligodendrocytes. Viability of 04 antigen-positive oligodendrocytes was approximately 90% as judged by exclusion of trypan blue. Oligodendrocytes were recovered after detachment from the beads with a yield of 19 +/- 6% and after collection by centrifugation onto glass coverslips with yields of approximately 6% of all 04 antigen-positive cells. The final cell yield of oligodendrocytes is approximately 8 x 10(5) cells/gram fresh cerebellar tissue.

Acrylamides↗
PubMed1982

Immunoselection of oligodendrocytes by magnetic beads. II. In vitro maintenance of immunoselected oligodendrocytes.

Oligodendrocytes isolated by immunoaffinity on magnetic beads using a monoclonal antibody to oligodendrocyte cell surfaces were obtained with a purity of 91 +/- 4% [see companion paper, Meier et al, 1982]. Contaminating cells consisted mostly of tetanus toxin-positive neurons with small cell bodies. Very few glial fibrillary acidic protein-positive astrocytes and fibronectin-positive fibroblasts or fibroblast-like cells could be detected by indirect immunofluorescence. Macrophages, as identified by their characteristic morphology, were rarely detectable in the cultures. With increasing time in culture, the degree of homogeneity of the cultured oligodendrocytes increased slightly, while the cell density of oligodendrocytes decreased drastically within the first week of culture. Cultures of immunoselected oligodendrocytes with a lower degree of purity showed a better survival of oligodendrocytes, but a considerable reduction in purity after three weeks in culture, probably due to overgrowth of nonoligodendroglial elements.

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PubMed1982

Development and expression of cytoplasmic antigens in Purkinje cells recognized by monoclonal antibodies. Studies in neurologically mutant mice.

Five monoclonal antibodies reacting with intracellular constituents of Purkinje cells were investigated by means of indirect immunofluorescence on fresh-frozen sections of the cerebellum and retina from developing and adult normal and mutant mice. Antibodies PC1, PC2 and PC3, which recognize Purkinje cells, but no other cerebellar neuron type, label these cells from day 4 onward. PC4 antigen is expressed in addition to Purkinje cells also in granule cells and neurons of deep cerebellar nuclei and appears in Purkinje cells at day 4. M1 antigen (Lagenaur et al. 1980) is first detectable in Purkinje cell bodies by day 5; it is also detectable in deep cerebellar neurons. In the adult retina, only PC4 antigen is detectably expressed and is localized in the inner segments of photoreceptor cells. The neurological mutants weaver, reeler, jimpy and wobbler show detectable levels of these antigens in Purkinje cells. However, the mutants staggerer and Purkinje cell degeneration are abnormal in expression PC1, PC2, PC3, and M1 antigens. Staggerer never starts to express the antigens during development, whereas Purkinje cell degeneration first expresses the antigens, but then loses antigen expression after day 23. PC4 antigen is detectable in the remaining Purkinje cells in staggerer and Purkinje cell degeneration mice at all ages tested in this study. Deep cerebellar neurons are positive for both antigens, PC4 and M1, in all mutants and at all ages studied. In retinas of staggerer and Purkinje cell degeneration mutants. PC4 antigen is normally detectable in the inner segments of photoreceptor cells, even when these have started to degenerate in the case of Purkinje cell degeneration.

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PubMed1982

Cell type specificity of a neural cell surface antigen recognized by the monoclonal antibody A2B5.

The monoclonal antibody A2B5 reacts with the surface membrane of most neurons in monolayer cultures of cerebellum, retina, spinal cord, and dorsal root ganglion of embryonic and early postnatal C57Bl/6 J mice maintained in vitro for culture periods of 2 to 10 days. A small percentage of astroglial cells also expresses A2B5 antigen in murine, chicken and rabbit cerebellum, in chicken retina, and in murine spinal cord and dorsal root ganglion. Less mature astroglial cells are strained for A2B5 antigen to a greater extent than the more mature astrocytes. Astrocytes from rat cerebellum and mouse retina were not found to express A2B5 antigen under the present culture conditons. Some of the less mature oligodendrocytes recognized by 04 antibodies express A2B5 antigen, while the more mature 01 antigen- and galactocerebroside-positive oligodendrocytes were not found to be A2B5 antigen-positive. Fibroblasts or fibroblast-like cells do not express detectable levels of A2B5 antigen. After fixation of the cells with paraformaldehyde and ethanol, all cell types present in culture are labeled by the A2B5 antibody intracellularly.

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PubMed1982

Electron-microscopic localization of A2B5 cell surface antigen in monolayer cultures of murine cerebellum and retina.

Immuno-electron microscopy was performed on live, cultured, early postnatal cerebellar and retinal cells of the mouse to identify A2B5 antigen-bearing elements. In cerebellar cultures, granule cells, some immature oligodendroglia, and astroblasts express A2B5 antigen on their cell surfaces. The typical features of astroblasts include large cisternae of the endoplasmic reticulum and a mixed population of intermediate-sized filaments and microtubules. Immature oligodendroglia cells express the antigen on their cell bodies and on processes filled with cytoplasm. Cytoplasm-free membranous whorls, however, are devoid of A2B5 antigen, but not of 0 or NS-1 antigens. In retinal cultures, A2B5 antigen is observed on differentiating neurons with the exception of photoreceptor cells as identified by ribbon synapses.

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