Molecular domains of myelinated axons

Myelinated axons are organized into specific domains as the result of interactions with glial cells. Recently, distinct protein complexes of cell adhesion molecules, Na(+) channels and ankyrin G at the nodes, Caspr and contactin in the paranodes, and K(+) channels and Caspr2 in the juxtaparanodal region have been identified, and new insights into the role of the paranodal junctions in the organization of these domains have emerged.     

Polarized domains of myelinated axons

The entire length of myelinated axons is organized into a series of polarized domains that center around nodes of Ranvier. These domains, which are crucial for normal saltatory conduction, consist of distinct multiprotein complexes of cell adhesion molecules, ion channels, and scaffolding molecules; they also differ in their diameter, organelle content, and rates of axonal transport. Juxtacrine signals from myelinating glia direct their sequential assembly. The composition, mechanisms of assembly, and function of these molecular domains will be reviewed. I also discuss similarities of this domain organization to that of polarized epithelia and present emerging evidence that disorders of domain organization and function contribute to the axonopathies of myelin and other neurologic disorders.     

Coated microvesicles in central nervous system myelinated axons

The distribution of coated and uncoated 50 nm microvesicles was determined for five CNS regions of myelinated axons of adult rats comparing two methods of fixation. On the average, we found 1.3 microvesicular profiles per axon of which 9.4% were bristle coated. The proportion of microvesicles with coated profiles in myelinated axons was 1.2 to 2 times that observed among microvesicles of nerve terminals depending on fixation method. Relative abundance of coated vs. uncoated vesicles among various brain regions was not significantly different. The results demonstrate for the first time the presence of coated vesicles in axons other than at their terminals. Not only were vesicles present, but the relative proportions in terminal and somal portions were similar.     

Latency distributions in homogeneous bundles of myelinated axons

Using a very simple hypothesis concerning the length of the depolarized area during propagation of action potentials, distributions of latencies in bundles of myelinated axons have been derived. The internodal length, the number of nodes of Ranvier, the depolarized area and the variation in internodal length are the important parameters.To demonstrate the applicability of the derivation proposed here some examples taken from neurophysiological experiments are given.     

Ribosomes in myelinated axons of dorsal root ganglia

Summary In the dorsal root ganglia of the rat, ribosomes were found not only in the initial segment, but they were also observed in the axoplasm of intraganglionar myelinated fibres and in the sensory portion of spinal nerves. Axons of seven-days-old rats contained more ribosomes than those of adult animals. The amount of particles decreased gradually from the initial segment trough intraganglionar internodes to the axons of spinal nerves. No ribosomes were found in axons of dorsal roots. In intraganglionar fibres, ribosomal particles were usually observed near the nodes of Ranvier, in the vicinity of Schmidt-Lantermann clefts and in axons near the Schwann cell nuclei. They were arranged in tetrads, pentads or in larger polysomes, and they were often observed adjacent to a group of mitochondria.The particles had invariably a stable size, their average diameters measuring 234 ± 2 × 197 ± 3 Å, which is practically equal to the diameters of 232 ± 2 × 203 ± 3 Å of ribosomes in the Schwann cell cytoplasm. These values fall within the range of diameters of ribosomes isolated from various cells of eukaryotic organisms as given in the literature. Since no other granular component of the cytoplasm has similarly stable dimensions, the measurements are considered to prove that the axonal particles described here are ribosomes.The author wishes to thank Dr. K. Smetana for his valuable suggestions and Mrs. M. Sobotková, Ing. M. Doubek and Mr. H. Kunz for their skillful technical assistance. The investigation was in part supported by a grant-in-aid from the Muscular Dystrophy Associations of America, Inc.     

Tolperisone--a novel modulator of ionic currents in myelinated axons

The actions of tolperisone on single intact Ranvier nodes of the toad Xenopus were investigated by means of the Hodgkin-Huxley formalism. Adding tolperisone to the bathing medium (100 micromol/l) caused the following fully reversible effects: 1. The sodium permeability P'Na was decreased by about 50% in a nearly potential-independent manner while the so-called sodium inactivation curve was shifted in the negative direction by about 3 mV. 2. The remaining parameters of the sodium system, i.e. m, taum and tauh, did not change. 3. The potassium permeability P'K decreased at strong depolarizing potentials (V > 60 mV); hence the permeability constant P(K) decreased by about 8%. However, weak depolarizations (V < 60 mV) caused P'K to increase by about 7%. 4. The potassium activation curve was shifted in the positive direction by about 9 mV and the exponent of n, b, was reduced from about 3.5 to about 1.5. Concentration-response relations for reduction of the sodium permeability constant PNa and of the potassium permeability constant P(K) yielded apparent dissociation constants of about 0.06 mmol/l and 0.32 mmol/l, respectively. The increase of P'K at V = 40 mV, however, was largely concentration-independent. Our findings show that, in contrast to the prevailing view, tolperisone cannot be said to have a so-called lidocaine-like activity, because its effect on potassium permeability in the threshold region is fundamentally different from that of other known local anaesthetics. We infer that this effect, in combination with the decrease in sodium permeability, is responsible for the tendency of tolperisone to reduce excitability and hence for the antispastic action of tolperisone documented by clinical observations.     

Voltage clamp calculations for myelinated and demyelinated axons

Exact cable theory is used to calculate voltage distributions along fully myelinated axons and those with various patterns of demyelination. The model employed uses an R-C circuit for the soma, an equivalent cable for the dendrites, a myelinated axon with n internodes and a cable representing telodendria. For the case of a voltage clamp at the soma, a system of 2n + 1 equations must be solved to obtain the potential distribution and this is done for arbitrary n. An explicit calculation is performed for one internode whereas computer-generated solutions are obtained for several internodes. The relative importance of the position of a single demyelinated internode is determined. An approximate expression is given for the critical internodal length necessary for action potential generation.     

Microtubules in myelinated and unmyelinated axons of rat sciatic nerve

Summary Tannic acid in glutaraldehyde was used to stain microtubules in myelinated and unmyelinated axons of rat sciatic nerve. In the majority of areas the tannic acid failed to penetrate the unmyelinated axons whilst penetrating neighbouring myelinated axons, suggesting a difference in the ability of the two types of nerves to exclude tannic acid. Where tannic acid had penetrated the unmyelinated axons the 13 protofilament substructure and size of the microtubules appeared identical to those seen in the myelinated axons.     

Spectrin and ankyrin-based cytoskeletons at polarized domains in myelinated axons

In myelinated nerve fibers, action potential initiation and propagation requires that voltage-gated ion channels be clustered at high density in the axon initial segments and nodes of Ranvier. The molecular organization of these subdomains depends on specialized cytoskeletal and scaffolding proteins such as spectrins, ankyrins, and 4.1 proteins. These cytoskeletal proteins are considered to be important for 1) formation, localization, and maintenance of specific integral membrane protein complexes, 2) a barrier restricting the diffusion of both cytoplasmic and membrane proteins to distinct regions or compartments of the cell, and 3) stabilization of axonal membrane integrity. Increased insights into the role of the cytoskeleton could provide important clues about the pathophysiology of various neurological disorders.     

Internodal specializations of myelinated axons in the central nervous system

We have examined the localization of contactin-associated protein (Caspr), the Shaker-type potassium channels, Kv1.1 and Kv1.2, their associated beta subunit, Kvbeta2, and Caspr2 in the myelinated fibers of the CNS. Caspr is localized to the paranodal axonal membrane, and Kv1.1, Kv1.2, Kvbeta2 and Caspr2 to the juxtaparanodal membrane. In addition to the paranodal staining, an internodal strand of Caspr staining apposes the inner mesaxon of the myelin sheath. Unlike myelinated axons in the peripheral nervous system, there was no internodal strand of Kv1.1, Kv1.2, Kvbeta2, or Caspr2. Thus, the organization of the nodal, paranodal, and juxtaparanodal axonal membrane is similar in the central and peripheral nervous systems, but the lack of Kv1.1/Kv1.2/Kvbeta2/Caspr2 internodal strands indicates that the oligodendrocyte myelin sheaths lack a trans molecular interaction with axons, an interaction that is present in Schwann cell myelin sheaths.     

nsf is essential for organization of myelinated axons in zebrafish

BACKGROUND: Myelinated axons are essential for rapid conduction of action potentials in the vertebrate nervous system. Of particular importance are the nodes of Ranvier, sites of voltage-gated sodium channel clustering that allow action potentials to be propagated along myelinated axons by saltatory conduction. Despite their critical role in the function of myelinated axons, little is known about the mechanisms that organize the nodes of Ranvier. RESULTS: Starting with a forward genetic screen in zebrafish, we have identified an essential requirement for nsf (N-ethylmaleimide sensitive factor) in the organization of myelinated axons. Previous work has shown that NSF is essential for membrane fusion in eukaryotes and has a critical role in vesicle fusion at chemical synapses. Zebrafish nsf mutants are paralyzed and have impaired response to light, reflecting disrupted nsf function in synaptic transmission and neural activity. In addition, nsf mutants exhibit defects in Myelin basic protein expression and in localization of sodium channel proteins at nodes of Ranvier. Analysis of chimeric larvae indicates that nsf functions autonomously in neurons, such that sodium channel clusters are evident in wild-type neurons transplanted into the nsf mutant hosts. Through pharmacological analyses, we show that neural activity and function of chemical synapses are not required for sodium channel clustering and myelination in the larval nervous system. CONCLUSIONS: Zebrafish nsf mutants provide a novel vertebrate system to investigate Nsf function in vivo. Our results reveal a previously unknown role for nsf, independent of its function in synaptic vesicle fusion, in the formation of the nodes of Ranvier in the vertebrate nervous system.     

The axoplasmic reticulum within myelinated axons is not transported rapidly

Brain Cell Biology - The axoplasmic reticulum in myelinated axons is an extensive system of branched smooth membranous tubules which is found throughout the length of large axons. To investigate...     

Composition of axolemma-enriched fractions isolated from bovine CNS myelinated axons

Axolemma-enriched fractions were isolated from the white matter of bovine corpus callosum via a purified preparation of myelinated axons which were osmotically shocked and fractionated on a discontinuous density gradient. Two membrane fractions of differing density were obtained; both were somewhat enriched over white matter whole homogenate in specific activity of acetylcholinesterase and 5-nucleotidase and maximal binding capacity for saxitoxin. Both membrane fractions contained appreciable amounts of 2, 3-cyclic nucleotide 3-phospho-hydrolase; the specific activity of antimycin-sensitive NAPH-cytochromec reductase and cytochromec oxidase indicated low levels of contamination by microsomal and mitochondrial membrane. The myelin which is concomitantly isolated with the axolemma-enriched fractions has a lipid and protein composition comparable to that of myelin isolated by other procedures. Both axolemma-enriched fractions contain about one half of their dry weight as lipid comprised of approximately 25% cholesterol, 25% galactolipid (cerebrosides and sulfatides in a molar ratio of about 4:1) and 50% phospholipid, mostly choline phosphatides and ethanolamine phospholes in an equimolar ratio. The axolemma fractions are also enriched in ganglioside content relative to the myelin fraction. The polypeptides of the axolemma-enriched fractions range from 20,000 to over 200,000 in molecular weight; the predominant proteins are in the range from 50,000 to 69,000. The most dense axolemma-enriched fraction is over fourfold enriched in glyco-protein content compared with myelin, with at least 10 different molecular-weight classes of glycoproteins as identified by Schiff stain of polyacrylamide gel protein profiles. The differences and similarities in the molecular composition of axolemma-enriched preparations which have been characterized to date are discussed.     

Simulation of high-frequency sinusoidal electrical block of mammalian myelinated axons

High frequency alternating current (HFAC) sinusoidal waveforms can block conduction in mammalian peripheral nerves. A mammalian axon model was used to simulate the response of nerves to HFAC conduction block. Sinusoidal waveforms from 1 to 40 kHz were delivered to eight simulated axon diameters ranging from 7.3 to 16 μm. Conduction block was obtained between 3 to 40 kHz. The minimum peak to peak current at which block was obtained, defined as the block threshold, increased with increasing frequency. Block threshold varied inversely with axon diameter. Upon initiation, the HFAC waveform produced one or more action potentials. These simulation results closely parallel previous experimental results of high frequency motor block of the rat sciatic and cat pudendal nerve. During HFAC block, the axons showed a dynamic steady state depolarization of multiple nodes, strongly suggesting a depolarization mechanism for HFAC conduction block. Action Editor: Karen Sigvardt     

The local differentiation of myelinated axons at nodes of Ranvier

Efficient and rapid propagation of action potentials in myelinated axons depends on the molecular specialization of the nodes of Ranvier. The nodal region is organized into several distinct domains, each of which contains a unique set of ion channels, cell-adhesion molecules and cytoplasmic adaptor proteins. Voltage-gated Na+ channels - which are concentrated at the nodes - are separated from K+ channels - which are clustered at the juxtaparanodal region - by a specialized axoglial contact that is formed between the axon and the myelinating cell at the paranodes. This local differentiation of myelinated axons is tightly regulated by oligodendrocytes and myelinating Schwann cells, and is achieved through complex mechanisms that are used by another specialized cell-cell contact - the synapse.     

Clustering and activity tuning of Kv1 channels in myelinated hippocampal axons

Precise localization of axonal ion channels is crucial for proper electrical and chemical functions of axons. In myelinated axons, Kv1 (Shaker) voltage-gated potassium (Kv) channels are clustered in the juxtaparanodal regions flanking the node of Ranvier. The clustering can be disrupted by deletion of various proteins in mice, including contactin-associated protein-like 2 (Caspr2) and transient axonal glycoprotein-1 (TAG-1), a glycosylphosphatidylinositol-anchored cell adhesion molecule. However, the mechanism and function of Kv1 juxtaparanodal clustering remain unclear. Here, using a new myelin coculture of hippocampal neurons and oligodendrocytes, we report that tyrosine phosphorylation plays a critical role in TAG-1-mediated clustering of axonal Kv1.2 channels. In the coculture, myelin specifically ensheathed axons but not dendrites of hippocampal neurons and clustered endogenous axonal Kv1.2 into internodes. The trans-homophilic interaction of TAG-1 was sufficient to position Kv1.2 clusters on axonal membranes in a neuron/HEK293 coculture. Mutating a tyrosine residue (Tyr⁴⁵⁸) in the Kv1.2 C terminus or blocking tyrosine phosphorylation disrupted myelin- and TAG-1-mediated clustering of axonal Kv1.2. Furthermore, Kv1.2 voltage dependence and activation threshold were reduced by TAG-1 coexpression. This effect was eliminated by the Tyr⁴⁵⁸ mutation or by cholesterol depletion. Taken together, our studies suggest that myelin regulates both trafficking and activity of Kv1 channels along hippocampal axons through TAG-1.