Distribution Of Myelin Basic Protein Research Articles

Behavioral and Molecular Consequences of Chronic Sleep Restriction During Development in Fragile X Mice

Sleep is critical for brain development and synaptic plasticity. In male wild-type mice, chronic sleep restriction during development results in long-lasting impairments in behavior including hypoactivity, decreased sociability, and increased repetitive behavior. Disordered sleep is characteristic of many neurodevelopmental disorders. Moreover, the severity of behavioral symptoms is correlated with the degree of disordered sleep. We hypothesized that chronic developmental sleep restriction in a mouse model of fragile X syndrome (FXS) would exacerbate behavioral phenotypes. To test our hypothesis, we sleep-restricted Fmr1 knockout (KO) mice for 3 h per day from P5 to P52 and subjected mice to behavioral tests beginning on P42. Contrary to our expectations, sleep restriction improved the hyperactivity and lack of preference for social novelty phenotypes in Fmr1 KO mice but had no measurable effect on repetitive activity. Sleep restriction also resulted in changes in regional distribution of myelin basic protein, suggesting effects on myelination. These findings have implications for the role of disrupted sleep in the severity of symptoms in FXS.

Abnormal activation of Yap/Taz contributes to the pathogenesis of tuberous sclerosis complex.

The multi-systemic genetic disorder tuberous sclerosis complex (TSC) impacts multiple neurodevelopmental processes including neuronal morphogenesis, neuronal migration, myelination and gliogenesis. These alterations contribute to the development of cerebral cortex abnormalities and malformations. Although TSC is caused by mTORC1 hyperactivation, cognitive and behavioral impairments are not improved through mTORC1 targeting, making the study of the downstream effectors of this complex important for understanding the mechanisms underlying TSC. As mTORC1 has been shown to promote the activity of the transcriptional co-activator Yap, we hypothesized that altered Yap/Taz signaling contributes to the pathogenesis of TSC. We first observed that the levels of Yap/Taz are increased in human cortical tuber samples and in embryonic cortices of Tsc2 conditional knockout (cKO) mice. Next, to determine how abnormal upregulation of Yap/Taz impacts the neuropathology of TSC, we deleted Yap/Taz in Tsc2 cKO mice. Importantly, Yap/Taz/Tsc2 triple conditional knockout (tcKO) animals show reduced cortical thickness and cortical neuron cell size, despite the persistence of high mTORC1 activity, suggesting that Yap/Taz play a downstream role in cytomegaly. Furthermore, Yap/Taz/Tsc2 tcKO significantly restored cortical and hippocampal lamination defects and reduced hippocampal heterotopia formation. Finally, the loss of Yap/Taz increased the distribution of myelin basic protein in Tsc2 cKO animals, consistent with an improvement in myelination. Overall, our results indicate that targeting Yap/Taz lessens the severity of neuropathology in a TSC animal model. This study is the first to implicate Yap/Taz as contributors to cortical pathogenesis in TSC and therefore as potential novel targets in the treatment of this disorder.

Distribution and orientation of nerve fibers and myelin assembly in a brain section retrieved by small-angle neutron scattering

The structural connectivity of the brain has been addressed by various imaging techniques such as diffusion weighted magnetic resonance imaging (DWMRI) or specific microscopic approaches based on histological staining or label-free using polarized light (e.g., three-dimensional Polarized Light Imaging (3D-PLI), Optical Coherence Tomography (OCT)). These methods are sensitive to different properties of the fiber enwrapping myelin sheaths i.e. the distribution of myelin basic protein (histology), the apparent diffusion coefficient of water molecules restricted in their movements by the myelin sheath (DWMRI), and the birefringence of the oriented myelin lipid bilayers (3D-PLI, OCT). We show that the orientation and distribution of nerve fibers as well as myelin in thin brain sections can be determined using scanning small angle neutron scattering (sSANS). Neutrons are scattered from the fiber assembly causing anisotropic diffuse small-angle scattering and Bragg peaks related to the highly ordered periodic myelin multilayer structure. The scattering anisotropy, intensity, and angular position of the Bragg peaks can be mapped across the entire brain section. This enables mapping of the fiber and myelin distribution and their orientation in a thin brain section, which was validated by 3D-PLI. The experiments became possible by optimizing the neutron beam collimation to highest flux and enhancing the myelin contrast by deuteration. This method is very sensitive to small microstructures of biological tissue and can directly extract information on the average fiber orientation and even myelin membrane thickness. The present results pave the way toward bio-imaging for detecting structural aberrations causing neurological diseases in future.

Cyfip1 haploinsufficient rats show white matter changes, myelin thinning, abnormal oligodendrocytes and behavioural inflexibility

The biological basis of the increased risk for psychiatric disorders seen in 15q11.2 copy number deletion is unknown. Previous work has shown disturbances in white matter tracts in human carriers of the deletion. Here, in a novel rat model, we recapitulated low dosage of the candidate risk gene CYFIP1 present within the 15q11.2 interval. Using diffusion tensor imaging, we first showed extensive white matter changes in Cyfip1 mutant rats, which were most pronounced in the corpus callosum and external capsule. Transmission electron microscopy showed that these changes were associated with thinning of the myelin sheath in the corpus callosum. Myelin thinning was independent of changes in axon number or diameter but was associated with effects on mature oligodendrocytes, including aberrant intracellular distribution of myelin basic protein. Finally, we demonstrated effects on cognitive phenotypes sensitive to both disruptions in myelin and callosal circuitry.

Aging-Related Peculiarities of the Distribution of Myelin Basic Protein in Cerebral Structures of Gerbils

Using an indirect immuno-enzyme technique, we measured the contents of myelin basic protein (MBP) in different cerebral structures of young adult and old (6- and 24-month-old, respectively) mongolian gerbils. The mean MBP content in the cerebellum of old gerbils was about 47% of the corresponding value in younger individuals (1.04 ± 0.10 and 2.20 ± 0.50 μg per 100 mg tissue, respectively). At the same time, the mean MBP contents in the thalamus and hippocampus of old gerbils demonstrated no decrease and even somewhat exceeded the corresponding values in young animals (the differences were, however, insignificant). Thus, aging-related changes in the MBP content in different brain structures of gerbils demonstrate clearly expressed specificities; there are reasons to believe that age-related processes of demyelination develop, first of all, in the cerebellum.

Interaction of myelin basic protein and 2',3'-cyclic nucleotide phosphodiesterase with mitochondria.

The content and distribution of myelin basic protein (MBP) isoforms (17 and 21.5 kDa) as well as 2',3'-cyclic nucleotide-3'-phosphodiesterase (CNPase) were determined in mitochondrial fractions (myelin fraction, synaptic and nonsynaptic mitochondria) obtained after separation of brain mitochondria by Percoll density gradient. All the fractions could accumulate calcium, maintain membrane potential, and initiate the opening of the permeability transition pore (mPTP) in response to calcium overloading. Native mitochondria and structural contacts between membranes of myelin and mitochondria were found in the myelin fraction associated with brain mitochondria. Using Western blot, it was shown that addition of myelin fraction associated with brain mitochondria to the suspension of liver mitochondria can lead to binding of CNPase and MBP, present in the fraction with liver mitochondria under the conditions of both closed and opened mPTP. However, induction of mPTP opening in liver mitochondria was prevented in the presence of myelin fraction associated with brain mitochondria (Ca2+ release rate was decreased 1.5-fold, calcium retention time was doubled, and swelling amplitude was 2.8-fold reduced). These results indicate possible protective properties of MBP and CNPase.

Treatment with afobazole at delayed time points following ischemic stroke improves long-term functional and histological outcomes

There is currently a significant lack of therapeutic options for acute ischemic stroke, and no drug has been approved for treating patients at delayed time points (≥6h post-stroke). Afobazole, an anxiolytic currently used clinically in Russia, has been shown to reduce neuronal and glial cell injury in vitro following ischemia. Experiments using the permanent middle cerebral artery occlusion (MCAO) rat model were carried out to determine if afobazole can reduce ischemic stroke damage in vivo and expand the therapeutic window for stroke treatment. Post-stroke (24h) application of afobazole (0.3–3mg/kg) significantly decreased infarct volume at 96h post-surgery, as determined by Fluoro-Jade and NeuN staining of brain sections. Moreover, afobazole helped preserve both the levels and normal histological distribution of myelin basic protein, indicating a reduction in white matter injury. A time-dependence study showed that either pre-treatment or treatment started 6 to 48h post-stroke with the drug yields improved outcomes at 96h. The decrease in infarct volume produced by afobazole was blocked by the application of either a σ-1 (BD 1063, 30mg/kg) or a σ-2 (SM-21, 1mg/kg) antagonist, indicating that both receptor subtypes are involved in the effects of afobazole. Treatment with afobazole starting at 24h post-stroke resulted in enhanced survival one month following surgery. Behavioral testing of animals 28–32days post-surgery using the elevated body swing and forelimb grip-strength tests revealed that treatment with afobazole starting 24h post-stroke significantly reduces behavioral deficits caused by ischemic stroke. The increase in survival and improved functional outcomes are accompanied by a reduction in infarct volume, as determined by thionin staining of brain sections. Taken together, our data support the use of afobazole as a post-stroke pharmacological agent to expand the current therapeutic window.

Expression of myelin basic protein in the human auditory nerve—An immunohistochemical and comparative study

The aim of this study is to analyse the expression and distribution of myelin basic protein (MBP or Myelin A1 protein) in the human spiral ganglion and auditory nerve. Cryostat sections were made from freshly fixed human cochlear specimens removed at surgery in patients with life-threatening petro-clival meningiomas compressing the brain stem. The sections were subjected to immunohistochemistry using antibodies against MBP, S-100 and Tubulin. The immunoreaction was documented using laser confocal microscopy. Type I spiral ganglion nerve somata (SGN) were surrounded by so-called "satellite glial cells" (SGCs) that lacked expression of MBP consistent with earlier light and electron microscopic findings indicating that these cells are non-myelinating. S-100 labeling showed that the SGCs form a continuous network in the apical region. The pattern of myelination in human spiral ganglion is different from that in other species' spiral ganglion. The striking differences in myelin outline should be investigated further in combination with its influence on signal coding and preservation properties in man.

Presence of α-globin mRNA and migration of bone marrow cells after sciatic nerve injury suggests their participation in the degeneration/regeneration process

We have previously reported that in the distal stump of ligated sciatic nerves, there is a change in the distribution of myelin basic protein (MBP) and P 0 protein immunoreactivities. These results agreed with the studies of myelin isolated from the distal stump of animals submitted to ligation of the sciatic nerve, showing a gradual increase in a 14 kDa band with an electrophoretic mobility similar to that of an MBP isoform, among other changes. This band, which was resolved into two bands of 14 and 15 kDa using a 16% gel, was found to contain a mixture of MBP fragments and peptides with great homology with α- and β-globins. In agreement with these results, we have demonstrated that the mRNA of α-globin is present in the proximal and distal stumps of the ligated nerve. It is also detected at very low levels in Schwann cells isolated from normal nerves. These results could be due to the presence of α- and/or β-globin arising from immature cells of the erythroid series. Also, they could be present in macrophages, which spontaneously migrate to the injured nerve to promote the degradation of myelin proteins. Cells isolated from normal adult rat bone marrow which were injected intraortically were found to migrate to the injured area. These cells could contribute to the remyelination of the damaged area participating in the removal of myelin debris, through their transdifferentiation into Schwann cells or through their fusion with preexisting Schwann cells in the distal stump of the injured sciatic nerve.

A periaxonal net in the zebrafish central nervous system

We produced a monoclonal antibody, named A20, which specifically recognizes a 35 kDa protein and stains myelinated axons in zebrafish brain. The A20 antigen is located at the outside of the myelin layer of large axons, and comprises a fine meshwork composed of thin unit fibers about 1–2 μm in length and about 100–200 nm in thickness. The unit fibers form pentagonal and hexagonal structures, which further polymerize into an envelope structure on the axons. The A20 monoclonal antibody did not stain neuronal cell bodies nor synapses. Instead, the distribution of the A20 antigen was along axons, practically coincident with the distribution of myelin basic protein. The monoclonal antibody stained only axons in the central nervous system (CNS), and not the extracellular matrix surrounding Schwann cells. These results suggest that this antigenic meshwork (which we call the periaxonal net) is synthesized by oligodendrocytes. During the development of the zebrafish brain, the periaxonal net appeared after the formation of myelin on the axons. The periaxonal net developed first at the brain stem, then gradually appeared at the caudal end of the spinal cord. The thickness of the periaxonal net around the Mauthner axon changed during development. Although the thickness of the Mauthner axon continues to grow throughout life, the thickness of periaxonal net stopped growing at 6 months after fertilization.

PPAR ? agonists stimulate oligodendrocyte differentiation in tissue culture

Peroxisome proliferator-activated receptors (PPARs) are ligand-activated transcription factors of the nuclear hormone receptor superfamily that have been described as master genes that switch cells from an undifferentiated phenotype to a differentiated phenotype. In the present investigation, we examined the possibility that ligands for PPARs are potent activators of oligodendrocyte (OL) differentiation and/or proliferation. Primary glial cultures and enriched OL cultures of neonatal mouse cerebra were treated with three different PPAR agonists: a PPAR gamma-selective agonist, a PPAR delta-selective agonist, and a pan agonist selective for both PPAR gamma and delta. Treatment with PPAR gamma agonist does not have an effect on the differentiation of OLs; however, PPAR delta agonist and the pan agonist treatment accelerates the differentiation of OLs within 24 h of application in mixed glial cultures. The number of OLs with processes and huge membrane sheets increases two- to threefold in both groups. The increase in the size of the sheets is also mirrored by changes in the intensity and distribution of myelin basic protein (MBP) and proteolipid protein (PLP) mRNAs. As compared to controls, the PPAR delta agonist-treated groups contain more OLs that have MBP and PLP mRNA extending into distal processes. These results indicate that PPAR delta plays a significant role in the maturation of OLs and regulates the size of OL sheets. BrdU immunostaining reveals that these agonists do not significantly stimulate proliferation of OLs expressing glycolipids. The studies in enriched OL cultures reproduce the effects of the PPAR agonists seen in the mixed glial cultures, indicating that the effect of the PPAR agonists is directly on the OLs and not via astrocytes. In the enriched cultures, the total number of OLs increases significantly in the PPAR delta agonist-treated groups, but BrdU immunostaining does not show an increased proliferation of cells. These findings suggest that PPAR delta increases the survival of cells and/or prevents cell death in enriched cultures. Although PPAR delta is expressed in various cell types, its role as a factor in the transcriptional regulation of OL differentiation has not been explored. We show for the first time that a ligand that serves as an agonist for PPAR delta activates the program of OL differentiation in primary and enriched OL cultures.

Distribution of myelin basic protein messages relative to the cytoskeleton

In situ hybridization is performed on oligodendrocytes using oligonucleotide probes to determine the subcellular distribution of myelin basic protein (MBP) encodulg. messages which, are associated with the cytoskeleton, relative to sub-cellular distribution of all MBP-encodlng messages In the cells. [Neural Res 1997; 19: 451-453]

Distribution of Myelin Basic Protein and P2 mRNAs in Rabbit Spinal Cord Oligodendrocytes

Myelin basic protein (MBP) and P2 protein are small positively charged proteins found in oligodendrocytes of rabbit spinal cord. Both proteins become incorporated into compact myelin. We have begun investigations into the mechanisms by which MBP and P2 become incorporated into the myelin membrane. We find that P2, like the MBPs, is synthesized on free polysomes in rabbit spinal cord. Cell fractionation experiments reveal that rabbit MBP mRNAs are preferentially segregated to the peripheral myelinating regions whereas P2 mRNAs are predominantly localized within the perikaryon of the cell. In vitro synthesized rabbit MBP readily associates with membranes added to translation mixtures, whereas P2 protein does not. It is possible that P2 requires a "receptor" molecule, perhaps a membrane-anchored protein, for association with the cytoplasmic face of the myelin membrane.

Differential Ultrastructural Localization of Myelin Basic Protein, Myelin/Oligodendroglial Glycoprotein, and 2′,3′‐Cyclic Nucleotide 3′‐Phosphodiesterase in the CNS of Adult Rats

In a light and electron microscopic immunocytochemical study we have examined the distribution of myelin basic protein (MBP), 2',3'-cyclic nucleotide 3'-phosphodiesterase (CNP), and myelin/oligodendroglial glycoprotein (MOG) within CNS myelin sheaths and oligodendrocytes of adult Sprague-Dawley rats. Ultrastructural immunocytochemistry allowed quantitative analysis of antigen density in different myelin and oligodendrocyte zones: MBP was detectable in high density over the whole myelin sheath, but not in regions of loops, somata, or the oligodendrocyte plasma membrane. CNP reactivity was highest at the myelin/axon interface, and found in lower concentration over the outer lamellae of myelin sheaths, at the cytoplasmic face of oligodendrocyte membranes, and throughout the compact myelin. MOG was preferentially detected at the extracellular surface of myelin sheaths and oligodendrocytes and in only low amounts in the lamellae of compacted myelin and the myelin/axon border zone. Our studies, thus, indicate further the presence of different molecular domains in compact myelin, which may be functionally relevant for the integrity and maintenance of the myelin sheath.

Immunocytochemical localization of MAG, MBP and P 0 protein in acute and relapsing demyelinating lesions of Theiler's virus infection

Acute demyelinating and relapsing demyelinating lesions from spinal cords of mice infected with the WW strain of Theiler's encephalomyelitis virus (TMEV) were studied immunocytochemically with antisera to various myelin constituents. Acute lesions were studied for differences in the distribution of myelin basic protein (MBP) and myelin associated glycoprotein. (MAG). Relapsing lesions, characterized by demyelination of areas previously remyelinated by Schwann cells, were studied for differences in the distriution of P 0 and MAG. In both instances the earliest lesions were characterized by preferential disappearance of MBP and P 0 respectively when compared to MAG. In well-developed lesions, MAG, MBP and P 0 were absent in essentially equal proportion. These observations are in agreement with previous findings suggesting a primary loss of myelin rather than a direct attack on oligodendrocytes as the main pathogenetic mechanism of demyelination in this viral model.

Myelin basic protein in the optic nerve of the developing albino rat: an immunoperoxidase study of paraffin embedded tissue.

The distribution of myelin basic protein (MBP) was determined by application of the unlabeled peroxidase-anti-peroxidase method to sections of paraffin embedded optic nerve taken from the developing albino rat. MBP was not detected in optic nerves from animals younger than 90 hours postnatum. MBP staining presented first as faint, diffuse deposits of chromagen which were found in approximately one-third of the 90 hour subjects. The number of MBP positive regions and the intensity of staining of these regions increased rapidly with age. There was no obvious radial gradient in the distribution of MBP in cross sections of developing optic nerves, nor were smooth central/peripheral gradients apparent in longitudinal sections.

Myelin basic protein and myelin-associated glycoprotein in chronic, relapsing experimental allergic encephalomyelitis

One-μm plastic sections of spinal cords from SJL/J mice with chronic relapsing experimental allergic encephalomyelitis (EAE) were reacted immunocytochemically with antiserum to myelin basic protein and myelin-associated glycoprotein. The distribution of myelin basic protein and myelin-associated glycoprotein in myelin sheaths was compared in acute and chronic areas of demyelination. No difference in the size of the lesion was seen with the two antisera. Myelin-associated glycoprotein was seen periaxonally in both normal myelin sheaths and sheaths which showed extensive splitting and ballooning as seen with toluidine blue stain and myelin basic protein antiserum. At least at the level of the light microscope, myelin basic protein antiserum gave intense staining of myelin while antiserum to myelin-associated glycoprotein showed little or no affinity to stain the myelin sheath itself, in contrast to other recent electron microscope observations. A few myelin basic protein or myelin-associated glycoprotein-containing oligodendrocytes were seen in lesion areas and remyelination by oligodendrocytes was rare. These observations are in agreement with findings from other models of EAE and multiple sclerosis where a primary loss of myelin has been implicated.

Ultracytochemical distribution of myelin basic protein after injection into the cerebrospinal fluid: Evidence for transport through the blood-brain barrier and binding to the luminal surface of cerebral veins

Distribution of myelin basic protein (MBP) in the central nervous system (CNS) following injection into the cerebrospinal fluid (CSF) was studied by different qualitative and quantitative immunelectron-microscopic techniques. Endogenous MBP was present in myelin sheaths in injected as well as in control animals. After injection of exogenous MBP into CSF this protein was present in the subarachnoid space, on the surface of meningeal cells, on the surface of collagen fibers, in the basement membrane of the glia limitans, in vessel walls, and in the extracellular space of spinal roots. In meningeal veins, endothelial vesicles filled with peroxidase reaction product were found on the abluminal side of endothelial cells, in the endothelial cytoplasm and sometimes opening into the vascular lumen. In addition patchy staining of the luminal surface of endothelial cells was noted, indicating binding of antigen at this location. Quantitative immunelectron microscopy (an indirect technique with rabbit anti-MBP serum as primary layer and gold-labeled anti-rabbit IgG as secondary layer) revealed highly significant MBP binding on the luminal surface of endothelial cells after injection of this antigen into the CSF. The present results indicate that MBP, when liberated in CNS is transported through the blood-brain barrier and presented on the luminal surface of endothelial cells of the cerebral and meningeal veins. This observation may be important in interpretation of pathogenesis of initial inflammatory infiltrates in experimental allergic encephalitis (EAE).

An electronmicroscopic study of the distribution of myelin basic protein in multilayer dispersions of diacylphosphatidylserine

Although dispersions containing lipid and protein are widely used as model systems to explore the properties of biomembranes, the extent of mixing of the two components has generally not been determined. Here, the distribution of bovine myelin basic protein in dispersions with bovine brain L-α-diacylphosphatidylserine (PS) has been examined electronmicroscopically. Dispersions of PS were prepared by hydrating a known amount of dried lipid with buffer or with buffer containing an equal weight of myelin basic protein or lysozyme. The lipid-protein complexes were separated from unbound protein by centrifugation in 0–60% sucrose density gradients. In both systems only a few percent of the protein was unbound and the resultant recombinants, which gave single bands on the gradients, contained about 50% protein by weight. After removal of the sucrose by dialysis the dispersions were fixed in 2.5% glutaraldehyde and 1% osmium tetroxide, dehydrated and embedded in epoxy resin. Thin sections cut from these blocks were incubated, after removal of osmium tetroxide, with antiserum raised in rabbits against human myelin basic protein. Excess antiserum was removed and the antigen-antibody complexes on the thin sections were labelled with 13 nm diameter colloidal gold particles stabilized with protein A. The distribution of these gold particles were examined under an electronmicroscope. Comparison of the labelling patterns for PS, PS-lysozyme and PS-basic protein demonstrated specific labelling in the last, and showed the gold partciles to be uniformly dispersed. It was concluded that in these dispersions the protein and lipid werre intimately mixed at the molecular level.

Regional distribution of myelin basic protein in the central nervous system of quaking, jimpy, and normal mice during development and aging.

Myelin basic protein (MBP) was quantified using a RIA technique in the spinal cord, cerebellum, diencephalon plus brainstem region and cerebral hemispheres of two dysmyelinating murine mutants, quaking (qk) and jimpy (jp) mice. Comparison was made with normal control values. The whole life-span has been investigated: ie, ages ranging from 0 to 26 days for the jp, O to one year for the qk, and prenatal stage to three years for the control animals. Assays in the mutants at early ages were rendered feasible by the use of marker genes, which has allowed the diagnosis of the mutation at birth, 12 days before the expression of their typical tremor phenotype. Special care was given to the period of early myelinogenesis in order to clarify the dysynchrony between the various parts of the central nervous system. In normal mice, MBP was already detected in the brain of 19-day-old embryos. During development, rapid accumulation of MBP first occurred in the spinal cord then in the diencephalon, the brainstem, the cerebellum, and finally in the cerebral hemispheres. In the 25-day-old jimpy mutant, levels of MBP were found dramatically decreased, never exceeding 6% of the normal controls in any of the areas investigated. The situation for the quaking mouse was quite different. This mutant could be investigated up to one year old. At that age, a high discrepancy was observed between the values found in the brain and in the spinal cord (respectively, 10% and 35%) compared to normal controls. In both mutants, not only were the levels of MBP decreased, but also its appearance during development was delayed. Nevertheless, in both mutants the caudo-rostral timing of myelination as assayed by MBP levels was maintained. Furthermore, the later myelination occurred, the stronger weas the deficit in MBP. Interestingly, in the quaking mutant, the specific plasticity of the spinal cord was exemplified by its ability to reduce constantly, even at an advanced age, its initial deficit of MBP.