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Journal of Brachial Plexus and... 2016Our previous experiments demonstrated modulation of the amplitude of the axonal compound action potential (CAP) by electrical stimulation. To verify assumption that...
Our previous experiments demonstrated modulation of the amplitude of the axonal compound action potential (CAP) by electrical stimulation. To verify assumption that glutamate released from axons could be involved in this phenomenon, the modification of the axonal CAP induced by glutamate was investigated. The major objective of this research is to verify the hypothesis that axonal activity would trigger the release of glutamate, which in turn would interact with specific axonal receptors modifying the amplitude of the action potential. Segments of the sciatic nerve were exposed to exogenous glutamate in vitro, and CAP was recorded before and after glutamate application. In some experiments, the release of radioactive glutamate analog from the sciatic nerve exposed to exogenous glutamate was also evaluated. The glutamate-induced increase in CAP was blocked by different glutamate receptor antagonists. The effect of glutamate was not observed in Ca-free medium, and was blocked by antagonists of calcium channels. Exogenous glutamate, applied to the segments of sciatic nerve, induced the release of radioactive glutamate analog, demonstrating glutamate-induced glutamate release. Immunohistochemical examination revealed that axolemma contains components necessary for glutamatergic neurotransmission. The proteins of the axonal membrane can under the influence of electrical stimulation or exogenous glutamate change membrane permeability and ionic conductance, leading to a change in the amplitude of CAP. We suggest that increased axonal activity leads to the release of glutamate that results in changes in the amplitude of CAPs.
PubMed: 28077958
DOI: 10.1055/s-0036-1593441 -
PloS One 2011Neurons are characterized by extremely long axons. This exceptional cell shape is likely to depend on multiple factors including interactions between the cytoskeleton...
Neurons are characterized by extremely long axons. This exceptional cell shape is likely to depend on multiple factors including interactions between the cytoskeleton and membrane proteins. In many cell types, members of the protein 4.1 family play an important role in tethering the cortical actin-spectrin cytoskeleton to the plasma membrane. Protein 4.1B is localized in myelinated axons, enriched in paranodal and juxtaparanodal regions, and also all along the internodes, but not at nodes of Ranvier where are localized the voltage-dependent sodium channels responsible for action potential propagation. To shed light on the role of protein 4.1B in the general organization of myelinated peripheral axons, we studied 4.1B knockout mice. These mice displayed a mildly impaired gait and motility. Whereas nodes were unaffected, the distribution of Caspr/paranodin, which anchors 4.1B to the membrane, was disorganized in paranodal regions and its levels were decreased. In juxtaparanodes, the enrichment of Caspr2, which also interacts with 4.1B, and of the associated TAG-1 and Kv1.1, was absent in mutant mice, whereas their levels were unaltered. Ultrastructural abnormalities were observed both at paranodes and juxtaparanodes. Axon calibers were slightly diminished in phrenic nerves and preterminal motor axons were dysmorphic in skeletal muscle. βII spectrin enrichment was decreased along the axolemma. Electrophysiological recordings at 3 post-natal weeks showed the occurrence of spontaneous and evoked repetitive activity indicating neuronal hyperexcitability, without change in conduction velocity. Thus, our results show that in myelinated axons 4.1B contributes to the stabilization of membrane proteins at paranodes, to the clustering of juxtaparanodal proteins, and to the regulation of the internodal axon caliber.
Topics: Alternative Splicing; Animals; Axons; Electrophysiology; Erythrocytes; Female; Male; Mice; Mice, Knockout; Microfilament Proteins; Microscopy, Fluorescence; Models, Biological; Mutation; Myelin Sheath; Neurons; Protein Isoforms; Rats; Sciatic Nerve; Temperature
PubMed: 21966409
DOI: 10.1371/journal.pone.0025043 -
International Journal of... 2009There is evidence that in the acute axonal motor neuropathy (AMAN) subtype of Guillain-Barré syndrome antibodies to gangliosides, produced through molecular mimicry by...
There is evidence that in the acute axonal motor neuropathy (AMAN) subtype of Guillain-Barré syndrome antibodies to gangliosides, produced through molecular mimicry by antecedent Campylobacter jejuni (C. jejuni) infection, attack gangliosides expressed in human peripheral nerve axolemma, inducing a primary axonal damage. The aim of this study is to investigate whether the T cell response has a role in AMAN pathogenesis. We isolated monocytes from 4 healthy subjects and 5 AMAN patients with antecedent C. jejuni infection and antibodies to GM1 and/or GD1a gangliosides. Immature dendritic cells expressing CD1 molecules cultured with autologous T cells were stimulated with 2 lipopolysaccharides (LPSs) extracted from C. jejuni strains containing GM1 and GD1a-like structures and with GM1 and GD1a. The T cell response to LPSs and to gangliosides was determined by measuring the release of IFN-gamma and TNF-alpha. We observed a T cell response to both LPSs in controls and AMAN patients, whereas only AMAN patients showed T cell reactivity to gangliosides GM1 and GD1a with a tight correlation between T cell reactivity to the ganglioside and individual antibody responses to the same ganglioside. T cells responding to gangliosides were CD1c-restricted CD8 positive and CD27 negative. These findings indicate a contribution of cellular immunity in the pathogenesis of AMAN. A possible role for ganglioside-reactive T cells might be to facilitate the production of antibodies against gangliosides.
Topics: Acute Disease; Adult; Aged; Antibodies; Antigens, CD1; Axons; CD8-Positive T-Lymphocytes; Campylobacter Infections; Campylobacter jejuni; Case-Control Studies; Cells, Cultured; Coculture Techniques; Cytotoxicity, Immunologic; Dendritic Cells; Female; G(M1) Ganglioside; Gangliosides; Glycoproteins; Guillain-Barre Syndrome; Humans; Immunity, Cellular; Immunophenotyping; Interferon-gamma; Lipopolysaccharides; Male; Middle Aged; Motor Neuron Disease; Motor Neurons; Tumor Necrosis Factor Receptor Superfamily, Member 7; Tumor Necrosis Factor-alpha; Young Adult
PubMed: 20074468
DOI: 10.1177/039463200902200420 -
The Journal of Cell Biology Feb 1980Using freeze-fracture techniques, we have analyzed the glial-axonal junction (GAJ) between Schwann cells and axons in the peripheral nervous system, and between... (Comparative Study)
Comparative Study
Rows of dimeric-particles within the axolemma and juxtaposed particles within glia, incorporated into a new model for the paranodal glial-axonal junction at the node of Ranvier.
Using freeze-fracture techniques, we have analyzed the glial-axonal junction (GAJ) between Schwann cells and axons in the peripheral nervous system, and between oligodendrocytes and axons in the central nervous system of the rat. We have identified a new set of dimeric-particles arranged in circumferential rows within the protoplasmic fracture faces (P-faces) of the paranodal axolemma in the region of glial-axonal juxtaposition. These particles, 260 A in length, composed of two 115-A subunits, are observed in both aldehyde-fixed and nonfixed preparations. The rows of dimeric-particles within the axonal P-face are associated with complementary rows of pits within the external fracture face (E-face) of the paranodal axolemma. These axonal particles are positioned between rows of 160-A particles that occur in both fracture faces of the glial loops in the same region. We observed, in addition to these previously described 160-A particles, a new set of 75-A glial particles within the glial P-faces of the GAJ. These 75-A particles form rows that are centered between the rows of 160-A particles and are therefore superimposed over the rows of dimeric-particles within the paranodal axolemma. Our new findings are interpreted with respect to methods of specimen preparation as well as to a potential role for the paranodal organ in saltatory conduction. We conclude that this particle-rich junction between axon and glia could potentially provide an intricate mechanism for ion exchange between these two cell types.
Topics: Animals; Axons; Cell Membrane; Fixatives; Freeze Etching; Freeze Fracturing; Glycerol; Models, Neurological; Neuroglia; Ranvier's Nodes; Rats
PubMed: 7380883
DOI: 10.1083/jcb.84.2.261 -
Frontiers in Cellular Neuroscience 2021In the central nervous system, myelin is attached to the axon in the paranodal region by a trimolecular complex of Neurofascin155 (NF155) in the myelin membrane,...
In the central nervous system, myelin is attached to the axon in the paranodal region by a trimolecular complex of Neurofascin155 (NF155) in the myelin membrane, interacting with Caspr1 and Contactin1 on the axolemma. Alternative splicing of a single Neurofascin transcript generates several different Neurofascins expressed by several cell types, but NF155, which is expressed by oligodendrocytes, contains a domain in the third fibronectinIII-like region of the molecule that is unique. The immunoglobulin 5-6 domain of NF155 is essential for binding to Contactin1, but less is known about the functions of the NF155-unique third fibronectinIII-like domain. Mutations and autoantibodies to this region are associated with several neurodevelopmental and demyelinating nervous system disorders. Here we used Crispr-Cas9 gene editing to delete a 9 bp sequence of NF155 in this unique domain, which has recently been identified as a thrombin binding site and implicated in plasticity of the myelin sheath. This small deletion results in dysmyelination, eversion of paranodal loops of myelin, substantial enlargement of the nodal gap, a complete loss of paranodal septate junctions, and mislocalization of Caspr1 and nodal sodium channels. The animals exhibit tremor and ataxia, and biochemical and mass spectrometric analysis indicates that while NF155 is transcribed and spliced normally, the NF155 protein is subsequently degraded, resulting in loss of the full length 155 kDa native protein. These findings reveal that this 9 bp region of NF155 in its unique third fibronectinIII-like domain is essential for stability of the protein.
PubMed: 33815060
DOI: 10.3389/fncel.2021.576609 -
The Journal of Neuroscience : the... May 1993Nerve injury frequently triggers hyperexcitability and the ectopic initiation of impulses in primary afferent axons. An important consequence is neuropathic paresthesias...
Nerve injury frequently triggers hyperexcitability and the ectopic initiation of impulses in primary afferent axons. An important consequence is neuropathic paresthesias and pain. Electrogenesis in normal afferents depends on appropriate Na+ channel concentrations. Therefore, we have asked whether injury might trigger changes in axolemmal Na+ channel distribution that could account for neuropathic hyperexcitability. We used an Na+ channel-specific antibody, 7493, to immunolocalize Na+ channels ultrastructurally in membranes of normal rat axons, and to assess remodeling following nerve section and neuroma formation. Selective labeling of nodal axolemma and, more weakly, of Schwann cell membrane, confirmed the efficacy of our immunolabeling protocol. In neuromas at postoperative times associated with peak ectopic activity, we found clear evidence of Na+ channel accumulation. Specifically, soon after myelin was stripped from large-diameter axons, the exposed, formerly internodal axolemma became immunopositive. Small-diameter unmyelinated axons and axon sprouts in the neuroma were also marked with 7493 IgG. Activated phagocytic macrophages and endothelial cells were 7493 negative. Both large- and small-diameter axons in neuromas end in swollen, organelle-packed "end bulbs." Most, but not all, of these acquired Na+ channel immunolabeling. We propose that remodeling results from a modification of the normal process of Na+ channel turnover in neural membranes. Na+ channel protein accumulates in preterminal axolemma and neuroma end bulbs due to a combination of permissive factors (especially myelin removal) and promotional factors (removal of normal downstream targets). This accumulation is a likely precursor of afferent hyperexcitability in injured nerve.
Topics: Animals; Axons; Epitopes; Immunologic Techniques; Male; Myelin Sheath; Neuroma; Neuronal Plasticity; Peripheral Nerve Injuries; Peripheral Nerves; Peripheral Nervous System Neoplasms; Ranvier's Nodes; Rats; Rats, Wistar; Sodium Channels; Wounds, Penetrating
PubMed: 7683047
DOI: 10.1523/JNEUROSCI.13-05-01976.1993 -
The Journal of Biophysical and... Aug 1961The extrinsic eye muscles of the killifish (F. heteroclitus) were fixed in OSO(4) (pH 7.6) and subsequently dehydrated, embedded, and sectioned for electron microscopy....
The extrinsic eye muscles of the killifish (F. heteroclitus) were fixed in OSO(4) (pH 7.6) and subsequently dehydrated, embedded, and sectioned for electron microscopy. The fine structures of neuromuscular junctions and of sarcoplasmic reticulum were then observed. The neuromuscular junction consists of the apposition of axolemma (60 to 70 A) and sarcolemma (90 to 100 A), with an intervening cleft space of 200 to 300 A, forming a synaptolemma 400 to 500 A thick. The terminal axons contain synaptic vesicles, mitochondria, and agranular reticulum. The subsynaptic sarcolemma lacks the infolding arrangement characteristic of neuromuscular junctions from other vertebrate skeletal muscle, making them more nearly like that of insect neuromuscular junctions. A comparison between the folded and non-folded subsynaptic membrane types is made and discussed in terms of comparative rates of acetylcholine diffusion from the synaptic cleft and resistances of the clefts and subsynaptic membranes. The sarcoplasmic reticulum consists of segmentally arranged, membrane-limited vesicles and tubular and cisternal elements which surround individual myofibrils in a sleeve-like arrangement. Triadic differentiation occurs at or near the A-I junction. Unit sleeves span the A and I bands alternately and consist of closed terminal cisternae interconnected across the A and I bands by tubular cisternae. The thickness of the sarcoplasmic membranes increases from 30 to 40 A in intertriadic regions to 50 to 70 A at the triads. The location of the triads is compared with previously described striated muscle from Ambystoma larval myotomes, cardiac and sartorius muscles of the albino rat, mouse limb muscle, chameleon lizard muscle, and insect muscle, with reference to their possible role in intracellular impulse conduction.
Topics: Animals; Cell Differentiation; Cytoskeleton; Fishes; Fundulidae; Histological Techniques; Insecta; Larva; Microscopy, Electron; Muscle, Skeletal; Muscles; Myofibrils; Neuromuscular Junction; Oculomotor Muscles; Sarcolemma; Sarcoplasmic Reticulum; Synaptic Vesicles
PubMed: 13740363
DOI: 10.1083/jcb.10.4.111 -
The Journal of Cell Biology Feb 2012Myelinating Schwann cells regulate the localization of ion channels on the surface of the axons they ensheath. This function depends on adhesion complexes that are...
Myelinating Schwann cells regulate the localization of ion channels on the surface of the axons they ensheath. This function depends on adhesion complexes that are positioned at specific membrane domains along the myelin unit. Here we show that the precise localization of internodal proteins depends on the expression of the cytoskeletal adapter protein 4.1G in Schwann cells. Deletion of 4.1G in mice resulted in aberrant distribution of both glial adhesion molecules and axonal proteins that were present along the internodes. In wild-type nerves, juxtaparanodal proteins (i.e., Kv1 channels, Caspr2, and TAG-1) were concentrated throughout the internodes in a double strand that flanked paranodal junction components (i.e., Caspr, contactin, and NF155), and apposes the inner mesaxon of the myelin sheath. In contrast, in 4.1G(-/-) mice, these proteins "piled up" at the juxtaparanodal region or aggregated along the internodes. These findings suggest that protein 4.1G contributes to the organization of the internodal axolemma by targeting and/or maintaining glial transmembrane proteins along the axoglial interface.
Topics: Animals; Axons; COS Cells; Cell Adhesion Molecules, Neuronal; Chlorocebus aethiops; Mice; Mice, Knockout; Microfilament Proteins; Myelin Sheath; Nerve Fibers, Myelinated; Peripheral Nerves
PubMed: 22291039
DOI: 10.1083/jcb.201111127 -
The Journal of Biological Chemistry Oct 1982The binding of a 3H-labeled ethylenediamine derivative of tetrodotoxin ([3H]EN-TTX) and 125I-labeled polypeptide neurotoxins, purified from the sea anemone Anenomia...
The binding of a 3H-labeled ethylenediamine derivative of tetrodotoxin ([3H]EN-TTX) and 125I-labeled polypeptide neurotoxins, purified from the sea anemone Anenomia sulcata (ATXII) and the scorpion Androctonus australis Hector (AaHII), was studied using axolemma-enriched membrane fractions. The membrane fractions were derived from a purified preparation of myelinated axons fractionated via a linear sucrose gradient in a zonal rotor. The specific activity of Na+K+ ATPase in the axolemma-enriched preparation, found in the 28-32% sucrose region of the density gradient, was 59.1 mumol of ATP hydrolyzed/mg of protein/h. As estimated by 3H-specific ouabain binding, this fraction contained 183.6 pmol of Na+K+ ATPase/mg of protein. The 28-32% region of the density gradient was most enriched in the binding capacity for all neurotoxins, while the stoichiometry of the binding activities varied throughout the density gradient. The maximal binding (Bmax) of [3H]EN-TTX was 1 pmol/mg; the dissociation constant (KD) of the neurotoxin for its receptor was 2 X 10(-10) M. The comparable values for ATXII were 3.2 pmol/mg and 1.5 X 10(-7) M, respectively, while AaHII had a Bmax of 0.08 pmol/mg and a KD of 3.3 X 10(-9) M. The relationship of the binding of these neurotoxins to that observed in other axonal plasma membrane preparations is discussed.
Topics: Animals; Axons; Brain Stem; Kinetics; Myelin Sheath; Neurotoxins; Rats; Species Specificity; Synaptosomes; Tetrodotoxin
PubMed: 7118904
DOI: No ID Found -
The Journal of Neuroscience : the... Apr 2008Peroxisomal metabolism is essential for normal brain development both in men and in mice. Using conditional knock-out mice, we recently showed that peroxisome deficiency...
Peroxisomal metabolism is essential for normal brain development both in men and in mice. Using conditional knock-out mice, we recently showed that peroxisome deficiency in liver has a severe and persistent impact on the formation of cortex and cerebellum, whereas absence of functional peroxisomes from the CNS only causes developmental delays without obvious alteration of brain architecture. We now report that a substantial fraction of the latter Nes-Pex5 knock-out mice survive into adulthood but develop progressive motoric and coordination problems, impaired exploration, and a deficit in cognition and die before the age of 6 months. Histopathologically, both the white and gray matter of the CNS displayed a region-specific accumulation of neutral lipids, astrogliosis and microgliosis, upregulation of catalase, and scattered cell death. Nes-Pex5 knock-out mice featured a dramatic reduction of myelin staining in corpus callosum, whereas cerebellum and other white matter tracts were less affected or unchanged. This was accompanied by a depletion of alkenylphospholipids in myelin and differentially reduced immunoreactivity of myelin proteins. EM analysis revealed that myelin wrappings around axons did still form, but they showed a reduction in thickness relative to axon diameters. Remarkably, multifocal axonal damage occurred in the corpus callosum. Thereby, debris accumulated between axolemma and inner myelin surface and axons collapsed, although myelin sheaths remained present. These anomalies of myelinated axons were already present in juvenile mice but aggravated in adulthood. Together, loss of CNS peroxisomal metabolism both affects myelin sheaths and axonal integrity possibly via independent pathways.
Topics: Animals; Apoptosis; Ataxia; Axons; Behavior, Animal; Brain; Catalase; Central Nervous System; Central Nervous System Diseases; Demyelinating Diseases; Dyskinesias; Exploratory Behavior; Gliosis; Intermediate Filament Proteins; Lipid Metabolism; Mice; Mice, Knockout; Myelin Sheath; Nerve Degeneration; Nerve Tissue Proteins; Nestin; Peroxisome-Targeting Signal 1 Receptor; Peroxisomes; Phenotype; Receptors, Cytoplasmic and Nuclear; Severity of Illness Index; Spinal Cord; Up-Regulation
PubMed: 18400901
DOI: 10.1523/JNEUROSCI.4968-07.2008