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Mouse Anti-Neurofilament 200 Monoclonal Antibody, Unconjugated, Clone NE14

RRID:AB_260781

Antibody ID

AB_260781

Target Antigen

Neurofilament 200 human, porcine, human, pig

Proper Citation

(Sigma-Aldrich Cat# N5389, RRID:AB_260781)

Clonality

monoclonal antibody

Comments

Vendor recommendations: Immunohistochemistry; Western Blot; Immunoblotting, Immunohistochemistry (formalin-fixed, paraffin-embedded), Immunohistochemistry (frozen)

Clone ID

Clone NE14

Host Organism

mouse

Vendor

Sigma-Aldrich

Cat Num

N5389

R-Ras1 and R-Ras2 Are Essential for Oligodendrocyte Differentiation and Survival for Correct Myelination in the Central Nervous System.

  • Sanz-Rodriguez M
  • J. Neurosci.
  • 2018 May 30

Literature context:


Abstract:

Rapid and effective neural transmission of information requires correct axonal myelination. Modifications in myelination alter axonal capacity to transmit electric impulses and enable pathological conditions. In the CNS, oligodendrocytes (OLs) myelinate axons, a complex process involving various cellular interactions. However, we know little about the mechanisms that orchestrate correct myelination. Here, we demonstrate that OLs express R-Ras1 and R-Ras2. Using female and male mutant mice to delete these proteins, we found that activation of the PI3K/Akt and Erk1/2-MAPK pathways was weaker in mice lacking one or both of these GTPases, suggesting that both proteins coordinate the activity of these two pathways. Loss of R-Ras1 and/or R-Ras2 diminishes the number of OLs in major myelinated CNS tracts and increases the proportion of immature OLs. In R-Ras1-/- and R-Ras2-/--null mice, OLs show aberrant morphologies and fail to differentiate correctly into myelin-forming phenotypes. The smaller OL population and abnormal OL maturation induce severe hypomyelination, with shorter nodes of Ranvier in R-Ras1-/- and/or R-Ras2-/- mice. These defects explain the slower conduction velocity of myelinated axons that we observed in the absence of R-Ras1 and R-Ras2. Together, these results suggest that R-Ras1 and R-Ras2 are upstream elements that regulate the survival and differentiation of progenitors into OLs through the PI3K/Akt and Erk1/2-MAPK pathways for proper myelination.SIGNIFICANCE STATEMENT In this study, we show that R-Ras1 and R-Ras2 play essential roles in regulating myelination in vivo and control fundamental aspects of oligodendrocyte (OL) survival and differentiation through synergistic activation of PI3K/Akt and Erk1/2-MAPK signaling. Mice lacking R-Ras1 and/or R-Ras2 show a diminished OL population with a higher proportion of immature OLs, explaining the observed hypomyelination in main CNS tracts. In vivo electrophysiology recordings demonstrate a slower conduction velocity of nerve impulses in the absence of R-Ras1 and R-Ras2. Therefore, R-Ras1 and R-Ras2 are essential for proper axonal myelination and accurate neural transmission.

Funding information:
  • Intramural NIH HHS - ZIA BC011010-06(United States)

Crosstalk control and limits of physiological c-Jun N-terminal kinase activity for cell viability and neurite stability in differentiated PC12 cells.

  • Waetzig V
  • Mol. Cell. Neurosci.
  • 2018 Apr 24

Literature context:


Abstract:

The c-Jun N-terminal kinases (JNKs) are important mediators of cell viability and structural integrity in postmitotic neurons, which is required for maintaining synaptic connections and neural plasticity. In the present study, we chose differentiated PC12 cells as a well-characterised neuronal model system to selectively examine the regulation of basal JNK activity by extracellular signal-regulated kinase 1/2 (ERK1/2) and Akt. We detected a complex interaction between the kinases to prevent cell death and neurite loss. Especially the appropriate level of JNK activation determined cellular survival. Basal activity of ERK1/2 attenuated the potentiation of JNK phosphorylation and thereby the induction of apoptosis. Importantly, when JNK activity was too low, cell viability and the number of neurite-bearing cells also decreased, even though the activation of ERK1/2 was enhanced. In this case, the JNK-mediated survival signals via activating transcription factor-3 (ATF3) were inhibited. Furthermore, the phosphorylation of ERK1/2 induced by the JNK inhibitor SP600125 inhibited the basal activity of Akt, which normally supported cell viability. Thus, controlling JNK activity is crucial to promote survival and neurite stability of differentiated neuronal cells.

The signaling lipid sphingosine 1-phosphate regulates mechanical pain.

  • Hill RZ
  • Elife
  • 2018 Mar 21

Literature context:


Abstract:

Somatosensory neurons mediate responses to diverse mechanical stimuli, from innocuous touch to noxious pain. While recent studies have identified distinct populations of A mechanonociceptors (AMs) that are required for mechanical pain, the molecular underpinnings of mechanonociception remain unknown. Here, we show that the bioactive lipid sphingosine 1-phosphate (S1P) and S1P Receptor 3 (S1PR3) are critical regulators of acute mechanonociception. Genetic or pharmacological ablation of S1PR3, or blockade of S1P production, significantly impaired the behavioral response to noxious mechanical stimuli, with no effect on responses to innocuous touch or thermal stimuli. These effects are mediated by fast-conducting A mechanonociceptors, which displayed a significant decrease in mechanosensitivity in S1PR3 mutant mice. We show that S1PR3 signaling tunes mechanonociceptor excitability via modulation of KCNQ2/3 channels. Our findings define a new role for S1PR3 in regulating neuronal excitability and establish the importance of S1P/S1PR3 signaling in the setting of mechanical pain thresholds.

Funding information:
  • Howard Hughes Medical Institute - Faculty Scholar Award()
  • Medical Research Council - G0800297(United Kingdom)
  • National Institute of Arthritis and Musculoskeletal and Skin Diseases - AR051219()
  • National Institute of Arthritis and Musculoskeletal and Skin Diseases - AR059385()
  • National Institute of General Medical Sciences - GM007367()
  • National Institute of Neurological Disorders and Stroke - NS077224()
  • National Institute of Neurological Disorders and Stroke - NS098097()
  • National Institute of Neurological Disorders and Stroke - NS105449()

Nimodipine confers clinical improvement in two models of experimental autoimmune encephalomyelitis.

  • Ingwersen J
  • J. Neurochem.
  • 2018 Feb 23

Literature context:


Abstract:

Multiple sclerosis is characterised by inflammatory neurodegeneration, with axonal injury and neuronal cell death occurring in parallel to demyelination. Regarding the molecular mechanisms responsible for demyelination and axonopathy, energy failure, aberrant expression of ion channels and excitotoxicity have been suggested to lead to Ca2+ overload and subsequent activation of calcium-dependent damage pathways. Thus, the inhibition of Ca2+ influx by pharmacological modulation of Ca2+ channels may represent a novel neuroprotective strategy in the treatment of secondary axonopathy. We therefore investigated the effects of the L-type voltage-gated calcium channel blocker nimodipine in two different models of mouse experimental autoimmune encephalomyelitis (EAE), an established experimental paradigm for multiple sclerosis. We show that preventive application of nimodipine (10 mg/kg per day) starting on the day of induction had ameliorating effects on EAE in SJL/J mice immunised with encephalitic myelin peptide PLP139-151 , specifically in late-stage disease. Furthermore, supporting these data, administration of nimodipine to MOG35-55 -immunised C57BL/6 mice starting at the peak of pre-established disease, also led to a significant decrease in disease score, indicating a protective effect on secondary CNS damage. Histological analysis confirmed that nimodipine attenuated demyelination, axonal loss and pathological axonal β-amyloid precursor protein accumulation in the cerebellum and spinal cord in the chronic phase of disease. Of note, we observed no effects of nimodipine on the peripheral immune response in EAE mice with regard to distribution, antigen-specific proliferation or activation patterns of lymphocytes. Taken together, our data suggest a CNS-specific effect of L-type voltage-gated calcium channel blockade to inflammation-induced neurodegeneration.

Funding information:
  • NIAID NIH HHS - R01AI59159(United States)

Acrylamide Retards the Slow Axonal Transport of Neurofilaments in Rat Cultured Dorsal Root Ganglia Neurons and the Corresponding Mechanisms.

  • An L
  • Neurochem. Res.
  • 2016 May 18

Literature context:


Abstract:

Chronic acrylamide (ACR) exposure induces peripheral-central axonopathy in occupational workers and laboratory animals, but the underlying mechanisms remain unclear. In this study, we first investigated the effects of ACR on slow axonal transport of neurofilaments in cultured rat dorsal root ganglia (DRG) neurons through live-cell imaging approach. Then for the underlying mechanisms exploration, the protein level of neurofilament subunits, motor proteins kinesin and dynein, and dynamitin subunit of dynactin in DRG neurons were assessed by western blotting and the concentrations of ATP was detected using ATP Assay Kit. The results showed that ACR treatment results in a dose-dependent decrease of slow axonal transport of neurofilaments. Furthermore, ACR intoxication significantly increases the protein levels of the three neurofilament subunits (NF-L, NF-M, NF-H), kinesin, dynein, and dynamitin subunit of dynactin in DRG neurons. In addition, ATP level decreased significantly in ACR-treated DRG neurons. Our findings indicate that ACR exposure retards slow axonal transport of NF-M, and suggest that the increase of neurofilament cargoes, motor proteins, dynamitin of dynactin, and the inadequate ATP supply contribute to the ACR-induced retardation of slow axonal transport.

Distribution of the auxiliary GABAB receptor subunits KCTD8, 12, 12b, and 16 in the mouse brain.

  • Metz M
  • J. Comp. Neurol.
  • 2011 Jun 1

Literature context:


Abstract:

GABA(B) receptors are the G-protein-coupled receptors for γ-aminobutyric acid (GABA). KCTD8, 12, 12b, and 16 were recently identified as auxiliary GABA(B) receptor subunits and distinctly influence biophysical and pharmacological properties of the receptor response. Here we examined the expression patterns of the KCTDs in the mouse brain. Using in situ hybridization analysis, we found that most neurons express KCTD transcripts, supporting biochemical data showing that most GABA(B) receptors in the brain incorporate KCTD proteins. In the adult brain, KCTD12 and 16 have a widespread and KCTD8 and 12b a restricted expression pattern. Individual neurons can coexpress multiple KCTDs, as shown for granule cells and CA1/CA3 pyramidal cells in the hippocampus that coexpress KCTD12 and 16. In contrast, granule, Purkinje, and Golgi cells in the cerebellum selectively express one KCTD at a time. The expression levels of individual KCTD transcripts vary during postnatal brain development. Immunohistochemistry reveals that individual KCTD proteins can exhibit distinct axonal or dendritic localizations in neuronal populations. KCTDs are also detectable in nonneuronal tissues not expected to express GABA(B) receptors, suggesting that the role of KCTD proteins extends beyond GABA(B) receptors. In summary, our findings support that most brain GABA(B) receptors associate with KCTD proteins, but that the repertoire and abundance of KCTDs varies during development, among brain areas, neuronal populations, and at subcellular sites. We propose that the distinct spatial and temporal KCTD distribution patterns underlie functional differences in native GABA(B) responses.

Funding information:
  • Canadian Institutes of Health Research - MH-71313(Canada)
  • NIGMS NIH HHS - R01 GM079719-02(United States)

Developmental expression of the actin depolymerizing factor ADF in the mouse inner ear and spiral ganglia.

  • Herde MK
  • J. Comp. Neurol.
  • 2010 May 15

Literature context:


Abstract:

Hair cells, the inner ear's sensory cells, are characterized by tens to hundreds of actin-rich stereocilia that form the hair bundle apparatus necessary for mechanoelectrical transduction. Both the number and length of actin filaments are precisely regulated in stereocilia. Proper cochlear and vestibular function also depends on actin filaments in nonsensory supporting cells. The formation of actin filaments is a dynamic, treadmill-like process in which actin-binding proteins play crucial roles. However, little is known about the presence and function of actin binding molecules in the inner ear, which set up, and maintain, actin-rich structures and regulate actin turnover. Here we examined the expression and subcellular location of the actin filament depolymerizing factor (ADF) in the cochlea and vestibular organs. By means of immunocytochemistry and confocal microscopy, we analyzed whole-mount preparations and cross-sections in fetal and postnatal mice (E15-P26). We found a transient ADF expression in immature hair cells of the organ of Corti, the utricle, and the saccule. Interestingly, the stereocilia were not labeled. By P26, ADF expression was restricted to supporting cells. In addition, we localized ADF in presynaptic terminals of medio-olivocochlear projections after hearing onset. A small population of spiral ganglion neurons strongly expressed ADF. Based on their relative number, peripheral location within the ganglion, smaller soma size, and coexpression of neurofilament 200, we identified these cells as Type II spiral ganglion neurons. The developmentally regulated ADF expression suggests a temporally restricted function in the stereocilia and, thus, a hitherto undescribed role of ADF.

Synaptic activity-related classical protein kinase C isoform localization in the adult rat neuromuscular synapse.

  • Besalduch N
  • J. Comp. Neurol.
  • 2010 Jan 10

Literature context:


Abstract:

Protein kinase C (PKC) is essential for signal transduction in a variety of cells, including neurons and myocytes, and is involved in both acetylcholine release and muscle fiber contraction. Here, we demonstrate that the increases in synaptic activity by nerve stimulation couple PKC to transmitter release in the rat neuromuscular junction and increase the level of alpha, betaI, and betaII isoforms in the membrane when muscle contraction follows the stimulation. The phosphorylation activity of these classical PKCs also increases. It seems that the muscle has to contract in order to maintain or increase classical PKCs in the membrane. We use immunohistochemistry to show that PKCalpha and PKCbetaI were located in the nerve terminals, whereas PKCalpha and PKCbetaII were located in the postsynaptic and the Schwann cells. Stimulation and contraction do not change these cellular distributions, but our results show that the localization of classical PKC isoforms in the membrane is affected by synaptic activity.

Funding information:
  • NIAMS NIH HHS - R37 AR038648-21(United States)