Publications by authors named "Wiebke Möbius"

105 Publications

DEAD-box RNA helicase Dbp4/DDX10 is an enhancer of α-synuclein toxicity and oligomerization.

PLoS Genet 2021 Mar 3;17(3):e1009407. Epub 2021 Mar 3.

Department of Molecular Microbiology and Genetics, Institute for Microbiology and Genetics, University of Goettingen, Göttingen, Germany.

Parkinson's disease is a neurodegenerative disorder associated with misfolding and aggregation of α-synuclein as a hallmark protein. Two yeast strain collections comprising conditional alleles of essential genes were screened for the ability of each allele to reduce or improve yeast growth upon α-synuclein expression. The resulting 98 novel modulators of α-synuclein toxicity clustered in several major categories including transcription, rRNA processing and ribosome biogenesis, RNA metabolism and protein degradation. Furthermore, expression of α-synuclein caused alterations in pre-rRNA transcript levels in yeast and in human cells. We identified the nucleolar DEAD-box helicase Dbp4 as a prominent modulator of α-synuclein toxicity. Downregulation of DBP4 rescued cells from α-synuclein toxicity, whereas overexpression led to a synthetic lethal phenotype. We discovered that α-synuclein interacts with Dbp4 or its human ortholog DDX10, sequesters the protein outside the nucleolus in yeast and in human cells, and stabilizes a fraction of α-synuclein oligomeric species. These findings provide a novel link between nucleolar processes and α-synuclein mediated toxicity with DDX10 emerging as a promising drug target.
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http://dx.doi.org/10.1371/journal.pgen.1009407DOI Listing
March 2021

Blood-brain barrier resealing in neuromyelitis optica occurs independently of astrocyte regeneration.

J Clin Invest 2021 Mar;131(5)

Institute of Neuropathology and.

Approximately 80% of neuromyelitis optica spectrum disorder (NMOSD) patients harbor serum anti-aquaporin-4 autoantibodies targeting astrocytes in the CNS. Crucial for NMOSD lesion initiation is disruption of the blood-brain barrier (BBB), which allows the entrance of Abs and serum complement into the CNS and which is a target for new NMOSD therapies. Astrocytes have important functions in BBB maintenance; however, the influence of their loss and the role of immune cell infiltration on BBB permeability in NMOSD have not yet been investigated. Using an experimental model of targeted NMOSD lesions in rats, we demonstrate that astrocyte destruction coincides with a transient disruption of the BBB and a selective loss of occludin from tight junctions. It is noteworthy that BBB integrity is reestablished before astrocytes repopulate. Rather than persistent astrocyte loss, polymorphonuclear leukocytes (PMNs) are the main mediators of BBB disruption, and their depletion preserves BBB integrity and prevents astrocyte loss. Inhibition of PMN chemoattraction, activation, and proteolytic function reduces lesion size. In summary, our data support a crucial role for PMNs in BBB disruption and NMOSD lesion development, rendering their recruitment and activation promising therapeutic targets.
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http://dx.doi.org/10.1172/JCI141694DOI Listing
March 2021

Lack of astrocytes hinders parenchymal oligodendrocyte precursor cells from reaching a myelinating state in osmolyte-induced demyelination.

Acta Neuropathol Commun 2020 12 24;8(1):224. Epub 2020 Dec 24.

Institute of Neuropathology, University Medical Center Göttingen, 37075, Göttingen, Germany.

Demyelinated lesions in human pons observed after osmotic shifts in serum have been referred to as central pontine myelinolysis (CPM). Astrocytic damage, which is prominent in neuroinflammatory diseases like neuromyelitis optica (NMO) and multiple sclerosis (MS), is considered the primary event during formation of CPM lesions. Although more data on the effects of astrocyte-derived factors on oligodendrocyte precursor cells (OPCs) and remyelination are emerging, still little is known about remyelination of lesions with primary astrocytic loss. In autopsy tissue from patients with CPM as well as in an experimental model, we were able to characterize OPC activation and differentiation. Injections of the thymidine-analogue BrdU traced the maturation of OPCs activated in early astrocyte-depleted lesions. We observed rapid activation of the parenchymal NG2 OPC reservoir in experimental astrocyte-depleted demyelinated lesions, leading to extensive OPC proliferation. One week after lesion initiation, most parenchyma-derived OPCs expressed breast carcinoma amplified sequence-1 (BCAS1), indicating the transition into a pre-myelinating state. Cells derived from this early parenchymal response often presented a dysfunctional morphology with condensed cytoplasm and few extending processes, and were only sparsely detected among myelin-producing or mature oligodendrocytes. Correspondingly, early stages of human CPM lesions also showed reduced astrocyte numbers and non-myelinating BCAS1 oligodendrocytes with dysfunctional morphology. In the rat model, neural stem cells (NSCs) located in the subventricular zone (SVZ) were activated while the lesion was already partially repopulated with OPCs, giving rise to nestin progenitors that generated oligodendroglial lineage cells in the lesion, which was successively repopulated with astrocytes and remyelinated. These nestin stem cell-derived progenitors were absent in human CPM cases, which may have contributed to the inefficient lesion repair. The present study points to the importance of astrocyte-oligodendrocyte interactions for remyelination, highlighting the necessity to further determine the impact of astrocyte dysfunction on remyelination inefficiency in demyelinating disorders including MS.
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http://dx.doi.org/10.1186/s40478-020-01105-2DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7761156PMC
December 2020

Oligodendrocytes support axonal transport and maintenance via exosome secretion.

PLoS Biol 2020 12 22;18(12):e3000621. Epub 2020 Dec 22.

Institute of Developmental Biology and Neurobiology, Johannes Gutenberg University of Mainz, Mainz, Germany.

Neurons extend long axons that require maintenance and are susceptible to degeneration. Long-term integrity of axons depends on intrinsic mechanisms including axonal transport and extrinsic support from adjacent glial cells. The mechanisms of support provided by myelinating oligodendrocytes to underlying axons are only partly understood. Oligodendrocytes release extracellular vesicles (EVs) with properties of exosomes, which upon delivery to neurons improve neuronal viability in vitro. Here, we show that oligodendroglial exosome secretion is impaired in 2 mouse mutants exhibiting secondary axonal degeneration due to oligodendrocyte-specific gene defects. Wild-type oligodendroglial exosomes support neurons by improving the metabolic state and promoting axonal transport in nutrient-deprived neurons. Mutant oligodendrocytes release fewer exosomes, which share a common signature of underrepresented proteins. Notably, mutant exosomes lack the ability to support nutrient-deprived neurons and to promote axonal transport. Together, these findings indicate that glia-to-neuron exosome transfer promotes neuronal long-term maintenance by facilitating axonal transport, providing a novel mechanistic link between myelin diseases and secondary loss of axonal integrity.
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http://dx.doi.org/10.1371/journal.pbio.3000621DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7787684PMC
December 2020

Microglia facilitate repair of demyelinated lesions via post-squalene sterol synthesis.

Nat Neurosci 2021 01 21;24(1):47-60. Epub 2020 Dec 21.

Department of Neurogenetics, Max Planck Institute of Experimental Medicine, Göttingen, Germany.

The repair of inflamed, demyelinated lesions as in multiple sclerosis (MS) necessitates the clearance of cholesterol-rich myelin debris by microglia/macrophages and the switch from a pro-inflammatory to an anti-inflammatory lesion environment. Subsequently, oligodendrocytes increase cholesterol levels as a prerequisite for synthesizing new myelin membranes. We hypothesized that lesion resolution is regulated by the fate of cholesterol from damaged myelin and oligodendroglial sterol synthesis. By integrating gene expression profiling, genetics and comprehensive phenotyping, we found that, paradoxically, sterol synthesis in myelin-phagocytosing microglia/macrophages determines the repair of acutely demyelinated lesions. Rather than producing cholesterol, microglia/macrophages synthesized desmosterol, the immediate cholesterol precursor. Desmosterol activated liver X receptor (LXR) signaling to resolve inflammation, creating a permissive environment for oligodendrocyte differentiation. Moreover, LXR target gene products facilitated the efflux of lipid and cholesterol from lipid-laden microglia/macrophages to support remyelination by oligodendrocytes. Consequently, pharmacological stimulation of sterol synthesis boosted the repair of demyelinated lesions, suggesting novel therapeutic strategies for myelin repair in MS.
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http://dx.doi.org/10.1038/s41593-020-00757-6DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7116742PMC
January 2021

Mannan-MOG35-55 Reverses Experimental Autoimmune Encephalomyelitis, Inducing a Peripheral Type 2 Myeloid Response, Reducing CNS Inflammation, and Preserving Axons in Spinal Cord Lesions.

Front Immunol 2020 19;11:575451. Epub 2020 Nov 19.

Laboratory of Molecular Genetics, Department of Immunology, Hellenic Pasteur Institute, Athens, Greece.

CNS autoantigens conjugated to oxidized mannan (OM) induce antigen-specific T cell tolerance and protect mice against autoimmune encephalomyelitis (EAE). To investigate whether OM-peptides treat EAE initiated by human MHC class II molecules, we administered OM-conjugated murine myelin oligodendrocyte glycoprotein peptide 35-55 (OM-MOG) to humanized HLA-DR2b transgenic mice (DR2b.Ab°), which are susceptible to MOG-EAE. OM-MOG protected DR2b.Ab° mice against MOG-EAE by both prophylactic and therapeutic applications. OM-MOG reversed clinical symptoms, reduced spinal cord inflammation, demyelination, and neuronal damage in DR2b.Ab° mice, while preserving axons within lesions and inducing the expression of genes associated with myelin () and neuron () recovery in B6 mice. OM-MOG-induced tolerance was peptide-specific, not affecting PLP178-191-induced EAE or polyclonal T cell proliferation responses. OM-MOG-induced immune tolerance involved rapid induction of PD-L1- and IL-10-producing myeloid cells, increased expression of (Ym1) in secondary lymphoid organs and characteristics of anergy in MOG-specific CD4 T cells. The results show that OM-MOG treats MOG-EAE in a peptide-specific manner, across mouse/human MHC class II barriers, through induction of a peripheral type 2 myeloid cell response and T cell anergy, and suggest that OM-peptides might be useful for suppressing antigen-specific CD4 T cell responses in the context of human autoimmune CNS demyelination.
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http://dx.doi.org/10.3389/fimmu.2020.575451DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7711156PMC
November 2020

Structural myelin defects are associated with low axonal ATP levels but rapid recovery from energy deprivation in a mouse model of spastic paraplegia.

PLoS Biol 2020 11 16;18(11):e3000943. Epub 2020 Nov 16.

Department of Neurogenetics, Max Planck Institute of Experimental Medicine, Göttingen, Germany.

In several neurodegenerative disorders, axonal pathology may originate from impaired oligodendrocyte-to-axon support of energy substrates. We previously established transgenic mice that allow measuring axonal ATP levels in electrically active optic nerves. Here, we utilize this technique to explore axonal ATP dynamics in the Plpnull/y mouse model of spastic paraplegia. Optic nerves from Plpnull/y mice exhibited lower and more variable basal axonal ATP levels and reduced compound action potential (CAP) amplitudes, providing a missing link between axonal pathology and a role of oligodendrocytes in brain energy metabolism. Surprisingly, when Plpnull/y optic nerves are challenged with transient glucose deprivation, both ATP levels and CAP decline slower, but recover faster upon reperfusion of glucose. Structurally, myelin sheaths display an increased frequency of cytosolic channels comprising glucose and monocarboxylate transporters, possibly facilitating accessibility of energy substrates to the axon. These data imply that complex metabolic alterations of the axon-myelin unit contribute to the phenotype of Plpnull/y mice.
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http://dx.doi.org/10.1371/journal.pbio.3000943DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7704050PMC
November 2020

A role of oligodendrocytes in information processing.

Nat Commun 2020 10 30;11(1):5497. Epub 2020 Oct 30.

Department of Neurogenetics, Max Planck Institute of Experimental Medicine, Göttingen, Germany.

Myelinating oligodendrocytes enable fast propagation of action potentials along the ensheathed axons. In addition, oligodendrocytes play diverse non-canonical roles including axonal metabolic support and activity-dependent myelination. An open question remains whether myelination also contributes to information processing in addition to speeding up conduction velocity. Here, we analyze the role of myelin in auditory information processing using paradigms that are also good predictors of speech understanding in humans. We compare mice with different degrees of dysmyelination using acute multiunit recordings in the auditory cortex, in combination with behavioral readouts. We find complex alterations of neuronal responses that reflect fatigue and temporal acuity deficits. We observe partially discriminable but similar deficits in well myelinated mice in which glial cells cannot fully support axons metabolically. We suggest a model in which myelination contributes to sustained stimulus perception in temporally complex paradigms, with a role of metabolically active oligodendrocytes in cortical information processing.
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http://dx.doi.org/10.1038/s41467-020-19152-7DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7599337PMC
October 2020

Intranasal mesenchymal stem cell therapy to boost myelination after encephalopathy of prematurity.

Glia 2021 Mar 12;69(3):655-680. Epub 2020 Oct 12.

Department for Developmental Origins of Disease, University Medical Center Utrecht, Utrecht University, Utrecht, The Netherlands.

Encephalopathy of prematurity (EoP) is a common cause of long-term neurodevelopmental morbidity in extreme preterm infants. Diffuse white matter injury (dWMI) is currently the most commonly observed form of EoP. Impaired maturation of oligodendrocytes (OLs) is the main underlying pathophysiological mechanism. No therapies are currently available to combat dWMI. Intranasal application of mesenchymal stem cells (MSCs) is a promising therapeutic option to boost neuroregeneration after injury. Here, we developed a double-hit dWMI mouse model and investigated the therapeutic potential of intranasal MSC therapy. Postnatal systemic inflammation and hypoxia-ischemia led to transient deficits in cortical myelination and OL maturation, functional deficits and neuroinflammation. Intranasal MSCs migrated dispersedly into the injured brain and potently improved myelination and functional outcome, dampened cerebral inflammationand rescued OL maturation after dWMI. Cocultures of MSCs with primary microglia or OLs show that MSCs secrete factors that directly promote OL maturation and dampen neuroinflammation. We show that MSCs adapt their secretome after ex vivo exposure to dWMI milieu and identified several factors including IGF1, EGF, LIF, and IL11 that potently boost OL maturation. Additionally, we showed that MSC-treated dWMI brains express different levels of these beneficial secreted factors. In conclusion, the combination of postnatal systemic inflammation and hypoxia-ischemia leads to a pattern of developmental brain abnormalities that mimics the clinical situation. Intranasal delivery of MSCs, that secrete several beneficial factors in situ, is a promising strategy to restore myelination after dWMI and subsequently improve the neurodevelopmental outcome of extreme preterm infants in the future.
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http://dx.doi.org/10.1002/glia.23919DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7821154PMC
March 2021

CMTM6 expressed on the adaxonal Schwann cell surface restricts axonal diameters in peripheral nerves.

Nat Commun 2020 09 9;11(1):4514. Epub 2020 Sep 9.

Department of Neurogenetics, Max Planck Institute of Experimental Medicine, 37075, Göttingen, Germany.

The velocity of nerve conduction is moderately enhanced by larger axonal diameters and potently sped up by myelination of axons. Myelination thus allows rapid impulse propagation with reduced axonal diameters; however, no myelin-dependent mechanism has been reported that restricts radial growth of axons. By label-free proteomics, STED-microscopy and cryo-immuno electron-microscopy we here identify CMTM6 (chemokine-like factor-like MARVEL-transmembrane domain-containing family member-6) as a myelin protein specifically localized to the Schwann cell membrane exposed to the axon. We find that disruption of Cmtm6-expression in Schwann cells causes a substantial increase of axonal diameters but does not impair myelin biogenesis, radial sorting or integrity of axons. Increased axonal diameters correlate with accelerated sensory nerve conduction and sensory responses and perturbed motor performance. These data show that Schwann cells utilize CMTM6 to restrict the radial growth of axons, which optimizes nerve function.
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http://dx.doi.org/10.1038/s41467-020-18172-7DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7481192PMC
September 2020

Macromolecular and electrical coupling between inner hair cells in the rodent cochlea.

Nat Commun 2020 06 25;11(1):3208. Epub 2020 Jun 25.

Institute for Auditory Neuroscience and InnerEarLab, University Medical Center Göttingen, Göttingen, Germany.

Inner hair cells (IHCs) are the primary receptors for hearing. They are housed in the cochlea and convey sound information to the brain via synapses with the auditory nerve. IHCs have been thought to be electrically and metabolically independent from each other. We report that, upon developmental maturation, in mice 30% of the IHCs are electrochemically coupled in 'mini-syncytia'. This coupling permits transfer of fluorescently-labeled metabolites and macromolecular tracers. The membrane capacitance, Ca-current, and resting current increase with the number of dye-coupled IHCs. Dual voltage-clamp experiments substantiate low resistance electrical coupling. Pharmacology and tracer permeability rule out coupling by gap junctions and purinoceptors. 3D electron microscopy indicates instead that IHCs are coupled by membrane fusion sites. Consequently, depolarization of one IHC triggers presynaptic Ca-influx at active zones in the entire mini-syncytium. Based on our findings and modeling, we propose that IHC-mini-syncytia enhance sensitivity and reliability of cochlear sound encoding.
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http://dx.doi.org/10.1038/s41467-020-17003-zDOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7316811PMC
June 2020

MICOS assembly controls mitochondrial inner membrane remodeling and crista junction redistribution to mediate cristae formation.

EMBO J 2020 Jul 22;39(14):e104105. Epub 2020 Jun 22.

Department of NanoBiophotonics, Max Planck Institute for Biophysical Chemistry, Göttingen, Germany.

Mitochondrial function is critically dependent on the folding of the mitochondrial inner membrane into cristae; indeed, numerous human diseases are associated with aberrant crista morphologies. With the MICOS complex, OPA1 and the F F -ATP synthase, key players of cristae biogenesis have been identified, yet their interplay is poorly understood. Harnessing super-resolution light and 3D electron microscopy, we dissect the roles of these proteins in the formation of cristae in human mitochondria. We individually disrupted the genes of all seven MICOS subunits in human cells and re-expressed Mic10 or Mic60 in the respective knockout cell line. We demonstrate that assembly of the MICOS complex triggers remodeling of pre-existing unstructured cristae and de novo formation of crista junctions (CJs) on existing cristae. We show that the Mic60-subcomplex is sufficient for CJ formation, whereas the Mic10-subcomplex controls lamellar cristae biogenesis. OPA1 stabilizes tubular CJs and, along with the F F -ATP synthase, fine-tunes the positioning of the MICOS complex and CJs. We propose a new model of cristae formation, involving the coordinated remodeling of an unstructured crista precursor into multiple lamellar cristae.
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http://dx.doi.org/10.15252/embj.2019104105DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7361284PMC
July 2020

Neuronal activity disrupts myelinated axon integrity in the absence of NKCC1b.

J Cell Biol 2020 Jul;219(7)

Centre for Discovery Brain Sciences, University of Edinburgh, Edinburgh, UK.

Through a genetic screen in zebrafish, we identified a mutant with disruption to myelin in both the CNS and PNS caused by a mutation in a previously uncharacterized gene, slc12a2b, predicted to encode a Na+, K+, and Cl- (NKCC) cotransporter, NKCC1b. slc12a2b/NKCC1b mutants exhibited a severe and progressive pathology in the PNS, characterized by dysmyelination and swelling of the periaxonal space at the axon-myelin interface. Cell-type-specific loss of slc12a2b/NKCC1b in either neurons or myelinating Schwann cells recapitulated these pathologies. Given that NKCC1 is critical for ion homeostasis, we asked whether the disruption to myelinated axons in slc12a2b/NKCC1b mutants is affected by neuronal activity. Strikingly, we found that blocking neuronal activity completely prevented and could even rescue the pathology in slc12a2b/NKCC1b mutants. Together, our data indicate that NKCC1b is required to maintain neuronal activity-related solute homeostasis at the axon-myelin interface, and the integrity of myelinated axons.
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http://dx.doi.org/10.1083/jcb.201909022DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7337504PMC
July 2020

In Situ Investigation of the Formation Kinematics of Plasma-Generated Silver Nanoparticles.

Nanomaterials (Basel) 2020 Mar 19;10(3). Epub 2020 Mar 19.

Faculty of Engineering and Health, HAWK University of Applied Sciences and Arts, Von-Ossietzky-Str. 99/100, 37085 Göttingen, Germany.

In this publication, it is shown how to synthesize silver nanoparticles from silver cations out of aqueous solutions by the use of an atmospheric pressure plasma source. The use of an atmospheric pressure plasma leads to a very fast reduction of silver ions in extensive solvent volumes. In order to investigate the nanoparticle synthesis process, ultraviolet/visible (UV/VIS) absorption spectra were recorded in situ. By using transmission electron microscopy and by the analysis of UV/VIS spectra, the kinetics of silver nanoparticle formation by plasma influence can be seen in more detail. For example, there are two different sections visible in the synthesis during the plasma exposure process. The first section of the synthesis is characterized by a linear formation of small spherical particles of nearly constant size. The second section is predominated by saturation effects. Here, particle faults are increasingly formed, induced by changes in the particle shape and the fusion of those particles. The plasma exposure time, therefore, determines the shape and size distribution of the nanoparticles.
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http://dx.doi.org/10.3390/nano10030555DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7153378PMC
March 2020

Pathology of myelinated axons in the PLP-deficient mouse model of spastic paraplegia type 2 revealed by volume imaging using focused ion beam-scanning electron microscopy.

J Struct Biol 2020 05 8;210(2):107492. Epub 2020 Mar 8.

Department of Neurogenetics, Max Planck Institute of Experimental Medicine, 37075 Göttingen, Germany; Electron Microscopy Core Unit, Max Planck Institute of Experimental Medicine, 37075 Göttingen, Germany; Center Nanoscale Microscopy and Molecular Physiology of the Brain (CNMPB), Göttingen, Germany. Electronic address:

Advances in electron microscopy including improved imaging techniques and state-of-the-art detectors facilitate imaging of larger tissue volumes with electron microscopic resolution. In combination with genetic tools for the generation of mouse mutants this allows assessing the three-dimensional (3D) characteristics of pathological features in disease models. Here we revisited the axonal pathology in the central nervous system of a mouse model of spastic paraplegia type 2, the Plp mouse. Although PLP is a bona fide myelin protein, the major hallmark of the disease in both SPG2 patients and mouse models are axonal swellings comprising accumulations of numerous organelles including mitochondria, gradually leading to irreversible axonal loss. To assess the number and morphology of axonal mitochondria and the overall myelin preservation we evaluated two sample preparation techniques, chemical fixation or high-pressure freezing and freeze substitution, with respect to the objective of 3D visualization. Both methods allowed visualizing distribution and morphological details of axonal mitochondria. In Plp mice the number of mitochondria is 2-fold increased along the entire axonal length. Mitochondria are also found in the excessive organelle accumulations within axonal swellings. In addition, organelle accumulations were detected within the myelin sheath and the inner tongue. We find that 3D electron microscopy is required for a comprehensive understanding of the size, content and frequency of axonal swellings, the hallmarks of axonal pathology.
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http://dx.doi.org/10.1016/j.jsb.2020.107492DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7196930PMC
May 2020

Saltatory Conduction along Myelinated Axons Involves a Periaxonal Nanocircuit.

Cell 2020 01 26;180(2):311-322.e15. Epub 2019 Dec 26.

Department of Axonal Signalling, Netherlands Institute for Neuroscience, Royal Netherlands Academy for Arts and Sciences, Meibergdreef 47, 1105 BA Amsterdam, the Netherlands; Cell Biology, Neurobiology and Biophysics, Department of Biology, Faculty of Science, Utrecht University, Padualaan 8, 3584 CH Utrecht, the Netherlands. Electronic address:

The propagation of electrical impulses along axons is highly accelerated by the myelin sheath and produces saltating or "jumping" action potentials across internodes, from one node of Ranvier to the next. The underlying electrical circuit, as well as the existence and role of submyelin conduction in saltatory conduction remain, however, elusive. Here, we made patch-clamp and high-speed voltage-calibrated optical recordings of potentials across the nodal and internodal axolemma of myelinated neocortical pyramidal axons combined with electron microscopy and experimentally constrained cable modeling. Our results reveal a nanoscale yet conductive periaxonal space, incompletely sealed at the paranodes, which separates the potentials across the low-capacitance myelin sheath and internodal axolemma. The emerging double-cable model reproduces the recorded evolution of voltage waveforms across nodes and internodes, including rapid nodal potentials traveling in advance of attenuated waves in the internodal axolemma, revealing a mechanism for saltation across time and space.
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http://dx.doi.org/10.1016/j.cell.2019.11.039DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC6978798PMC
January 2020

Two adhesive systems cooperatively regulate axon ensheathment and myelin growth in the CNS.

Nat Commun 2019 10 22;10(1):4794. Epub 2019 Oct 22.

Institute of Neuronal Cell Biology, Technical University Munich, Munich, Germany.

Central nervous system myelin is a multilayered membrane produced by oligodendrocytes to increase neural processing speed and efficiency, but the molecular mechanisms underlying axonal selection and myelin wrapping are unknown. Here, using combined morphological and molecular analyses in mice and zebrafish, we show that adhesion molecules of the paranodal and the internodal segment work synergistically using overlapping functions to regulate axonal interaction and myelin wrapping. In the absence of these adhesive systems, axonal recognition by myelin is impaired with myelin growing on top of previously myelinated fibers, around neuronal cell bodies and above nodes of Ranvier. In addition, myelin wrapping is disturbed with the leading edge moving away from the axon and in between previously formed layers. These data show how two adhesive systems function together to guide axonal ensheathment and myelin wrapping, and provide a mechanistic understanding of how the spatial organization of myelin is achieved.
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http://dx.doi.org/10.1038/s41467-019-12789-zDOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC6805957PMC
October 2019

Correction to: Ketogenic diet ameliorates axonal defects and promotes myelination in Pelizaeus-Merzbacher disease.

Acta Neuropathol 2019 Oct 3;138(4):673-674. Epub 2019 Sep 3.

Department of Neurogenetics, Max-Planck-Institute of Experimental Medicine, Hermann-Rein-Str. 3, 37075, Göttingen, Germany.

The original article was published.
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http://dx.doi.org/10.1007/s00401-019-02064-2DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC6778063PMC
October 2019

FIB-SEM of mouse nervous tissue: Fast and slow sample preparation.

Methods Cell Biol 2019 7;152:1-21. Epub 2019 May 7.

Department of Neurogenetics, Electron Microscopy Core Unit, Max Planck Institute of Experimental Medicine, Göttingen, Germany; Center Nanoscale Microscopy and Molecular Physiology of the Brain (CNMPB), Göttingen, Germany. Electronic address:

Focused ion beam-scanning electron microscopy (FIB-SEM) has become a widely used technique in life sciences. To achieve the best data quality, sample preparation is important and has to be adapted to the specimen and the specific application. Here we illustrate three preparation procedures for mouse nervous tissue: First, the use of high-pressure freezing followed by direct imaging of vitrified tissue without any staining in the FIB-SEM under cryo-conditions as direct and fast procedure. Second, a slow procedure involving freeze substitution of frozen samples combined with additional staining for enhanced contrast and plastic embedding. Third, a fast preparation applying microwave-assisted chemical fixation and processing for resin embedding. All three methods of sample preparation are suitable for obtaining data stacks by FIB-SEM acquisition and 3D reconstruction.
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http://dx.doi.org/10.1016/bs.mcb.2019.03.009DOI Listing
April 2020

A liquid-like spindle domain promotes acentrosomal spindle assembly in mammalian oocytes.

Science 2019 06;364(6447)

Department of Meiosis, Max Planck Institute for Biophysical Chemistry, 37077 Göttingen, Germany.

Mammalian oocytes segregate chromosomes with a microtubule spindle that lacks centrosomes, but the mechanisms by which acentrosomal spindles are organized and function are largely unclear. In this study, we identify a conserved subcellular structure in mammalian oocytes that forms by phase separation. This structure, which we term the liquid-like meiotic spindle domain (LISD), permeates the spindle poles and forms dynamic protrusions that extend well beyond the spindle. The LISD selectively concentrates multiple microtubule regulatory factors and allows them to diffuse rapidly within the spindle volume. Disruption of the LISD via different means disperses these factors and leads to severe spindle assembly defects. Our data suggest a model whereby the LISD promotes meiotic spindle assembly by serving as a reservoir that sequesters and mobilizes microtubule regulatory factors in proximity to spindle microtubules.
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http://dx.doi.org/10.1126/science.aat9557DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC6629549PMC
June 2019

Platelets, endothelial cells and leukocytes contribute to the exercise-triggered release of extracellular vesicles into the circulation.

J Extracell Vesicles 2019 28;8(1):1615820. Epub 2019 May 28.

Institute of Developmental Biology and Neurobiology, Biology of Extracellular Vesicles, University of Mainz, Mainz, Germany.

Physical activity initiates a wide range of multi-systemic adaptations that promote mental and physical health. Recent work demonstrated that exercise triggers the release of extracellular vesicles (EVs) into the circulation, possibly contributing to exercise-associated adaptive systemic signalling. Circulating EVs comprise a heterogeneous collection of different EV-subclasses released from various cell types. So far, a comprehensive picture of the parental and target cell types, EV-subpopulation diversity and functional properties of EVs released during exercise (ExerVs) is lacking. Here, we performed a detailed EV-phenotyping analysis to explore the cellular origin and potential subtypes of ExerVs. Healthy male athletes were subjected to an incremental cycling test until exhaustion and blood was drawn before, during, and immediately after the test. Analysis of total blood plasma by EV Array suggested endothelial and leukocyte characteristics of ExerVs. We further purified ExerVs from plasma by size exclusion chromatography as well as CD9-, CD63- or CD81-immunobead isolation to examine ExerV-subclass dynamics. EV-marker analysis demonstrated increasing EV-levels during cycling exercise, with highest levels at peak exercise in all EV-subclasses analysed. Phenotyping of ExerVs using a multiplexed flow-cytometry platform revealed a pattern of cell surface markers associated with ExerVs and identified lymphocytes (CD4, CD8), monocytes (CD14), platelets (CD41, CD42, CD62P), endothelial cells (CD105, CD146) and antigen presenting cells (MHC-II) as ExerV-parental cells. We conclude that multiple cell types associated with the circulatory system contribute to a pool of heterogeneous ExerVs, which may be involved in exercise-related signalling mechanisms and tissue crosstalk.
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http://dx.doi.org/10.1080/20013078.2019.1615820DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC6542154PMC
May 2019

Myelinating Glia-Specific Deletion of Fbxo7 in Mice Triggers Axonal Degeneration in the Central Nervous System Together with Peripheral Neuropathy.

J Neurosci 2019 07 13;39(28):5606-5626. Epub 2019 May 13.

Department of Neurology, RWTH University Hospital, 52074 Aachen, Germany,

Myelination of axons facilitates the rapid propagation of electrical signals and the long-term integrity of axons. The ubiquitin-proteasome system is essential for proper protein homeostasis, which is particularly crucial for interactions of postmitotic cells. In our study, we examined how the E3 ubiquitin ligase FBXO7-SCF (SKP1, Cul1, F-box protein) expressed in myelinating cells affects the axon-myelin unit. Deletion of in oligodendrocytes and Schwann cells in mice using the Cnp1- driver line led to motor impairment due to hindlimb paresis. It did not result in apoptosis of myelinating cells, nor did it affect the proper myelination of axons or lead to demyelination. It however triggered axonal degeneration in the CNS and resulted in the severe degeneration of axons in the PNS, inducing a full-blown neuropathy. Both the CNS and PNS displayed inflammation, while the PNS was also characterized by fibrosis, massive infiltration of macrophages, and edema. Tamoxifen-induced deletion of , after myelination using the Plp1- line, led to a small number of degenerated axons and hence a very mild peripheral neuropathy. Interestingly, loss of also resulted in reduced proteasome activity in Schwann cells but not in cerebellar granule neurons, indicating a specific sensitivity of the former cell type. Together, our results demonstrate an essential role for FBXO7 in myelinating cells to support associated axons, which is fundamental to the proper developmental establishment and the long-term integrity of the axon-myelin unit. The myelination of axons facilitates the fast propagation of electrical signals and the trophic support of the myelin-axon unit. Here, we report that deletion of in myelinating cells in mice triggered motor impairment but had no effect on myelin biogenesis. Loss of Fbxo7 in myelinating glia, however, led to axonal degeneration in the CNS and peripheral neuropathy of the axonal type. In addition, we found that Schwann cells were particularly sensitive to Fbxo7 deficiency reflected by reduced proteasome activity. Based on these findings, we conclude that Fbxo7 is essential for the support of the axon-myelin unit and long-term axonal health.
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http://dx.doi.org/10.1523/JNEUROSCI.3094-18.2019DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC6616290PMC
July 2019

Publisher Correction: NRG1 type I dependent autoparacrine stimulation of Schwann cells in onion bulbs of peripheral neuropathies.

Nat Commun 2019 Apr 16;10(1):1840. Epub 2019 Apr 16.

Department of Neurogenetics, Max-Planck-Institute of Experimental Medicine, Hermann-Rein-Str. 3, 37075, Göttingen, Germany.

Michael W. Sereda was incorrectly associated with the Department of Cellular Neurophysiology, Hanover Medical School, Carl-Neuberg-Str. 1, 30625 Hanover, Germany. The correct affiliations for Michael W. Sereda are Department of Neurogenetics, Max-Planck-Institute of Experimental Medicine, Hermann-Rein-Str. 3, 37075 Göttingen, Germany and Department of Clinical Neurophysiology, University Medical Center Göttingen, Robert-Koch-Str. 40, 37075 Göttingen, Germany.
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http://dx.doi.org/10.1038/s41467-019-09886-4DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC6467885PMC
April 2019

Biological Sample Preparation by High-pressure Freezing, Microwave-assisted Contrast Enhancement, and Minimal Resin Embedding for Volume Imaging.

J Vis Exp 2019 03 19(145). Epub 2019 Mar 19.

Electron Microscopy Core Unit, Max Planck Institute of Experimental Medicine; Center Nanoscale Microscopy and Molecular Physiology of the Brain (CNMPB);

The described sample preparation technique is designed to combine the best quality of ultrastructural preservation with the most suitable contrast for the imaging modality in a focused ion beam scanning electron microscope (FIB-SEM), which is used to obtain stacks of sequential images for 3D reconstruction and modelling. High-pressure freezing (HPF) allows close to native structural preservation, but the subsequent freeze substitution often does not provide sufficient contrast, especially for a bigger specimen, which is needed for high-quality imaging in the SEM required for 3D reconstruction. Therefore, in this protocol, after the freeze substitution, additional contrasting steps are carried out at room temperature. Although these steps are performed in a microwave, it is also possible to follow traditional bench processing, which requires longer incubation times.The subsequent embedding in minimal amounts of resin allows for faster and more precise targeting and preparation inside the FIB-SEM. This protocol is especially useful for samples that require preparation by high-pressure freezing for a reliable ultrastructural preservation but do not gain enough contrast during the freeze substitution for volume imaging using FIB-SEM. In combination with the minimal resin embedding, this protocol provides an efficient workflow for the acquisition of high-quality volume data.
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http://dx.doi.org/10.3791/59156DOI Listing
March 2019

NRG1 type I dependent autoparacrine stimulation of Schwann cells in onion bulbs of peripheral neuropathies.

Nat Commun 2019 04 1;10(1):1467. Epub 2019 Apr 1.

Department of Neurogenetics, Max-Planck-Institute of Experimental Medicine, Hermann-Rein-Str. 3, 37075, Göttingen, Germany.

In contrast to acute peripheral nerve injury, the molecular response of Schwann cells in chronic neuropathies remains poorly understood. Onion bulb structures are a pathological hallmark of demyelinating neuropathies, but the nature of these formations is unknown. Here, we show that Schwann cells induce the expression of Neuregulin-1 type I (NRG1-I), a paracrine growth factor, in various chronic demyelinating diseases. Genetic disruption of Schwann cell-derived NRG1 signalling in a mouse model of Charcot-Marie-Tooth Disease 1A (CMT1A), suppresses hypermyelination and the formation of onion bulbs. Transgenic overexpression of NRG1-I in Schwann cells on a wildtype background is sufficient to mediate an interaction between Schwann cells via an ErbB2 receptor-MEK/ERK signaling axis, which causes onion bulb formations and results in a peripheral neuropathy reminiscent of CMT1A. We suggest that diseased Schwann cells mount a regeneration program that is beneficial in acute nerve injury, but that overstimulation of Schwann cells in chronic neuropathies is detrimental.
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http://dx.doi.org/10.1038/s41467-019-09385-6DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC6443727PMC
April 2019

Ketogenic diet ameliorates axonal defects and promotes myelination in Pelizaeus-Merzbacher disease.

Acta Neuropathol 2019 07 27;138(1):147-161. Epub 2019 Mar 27.

Department of Neurogenetics, Max-Planck-Institute of Experimental Medicine, Hermann-Rein-Str. 3, 37075, Göttingen, Germany.

Pelizaeus-Merzbacher disease (PMD) is an untreatable and fatal leukodystrophy. In a model of PMD with perturbed blood-brain barrier integrity, cholesterol supplementation promotes myelin membrane growth. Here, we show that in contrast to the mouse model, dietary cholesterol in two PMD patients did not lead to a major advancement of hypomyelination, potentially because the intact blood-brain barrier precludes its entry into the CNS. We therefore turned to a PMD mouse model with preserved blood-brain barrier integrity and show that a high-fat/low-carbohydrate ketogenic diet restored oligodendrocyte integrity and increased CNS myelination. This dietary intervention also ameliorated axonal degeneration and normalized motor functions. Moreover, in a paradigm of adult remyelination, ketogenic diet facilitated repair and attenuated axon damage. We suggest that a therapy with lipids such as ketone bodies, that readily enter the brain, can circumvent the requirement of a disrupted blood-brain barrier in the treatment of myelin disease.
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http://dx.doi.org/10.1007/s00401-019-01985-2DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC6570703PMC
July 2019

Mapping developmental maturation of inner hair cell ribbon synapses in the apical mouse cochlea.

Proc Natl Acad Sci U S A 2019 03 13;116(13):6415-6424. Epub 2019 Mar 13.

Molecular Architecture of Synapses Group, Institute for Auditory Neuroscience, InnerEarLab and Center for Biostructural Imaging of Neurodegeneration, University Medical Center Göttingen, 37075 Göttingen, Germany;

Ribbon synapses of cochlear inner hair cells (IHCs) undergo molecular assembly and extensive functional and structural maturation before hearing onset. Here, we characterized the nanostructure of IHC synapses from late prenatal mouse embryo stages (embryonic days 14-18) into adulthood [postnatal day (P)48] using electron microscopy and tomography as well as optical nanoscopy of apical turn organs of Corti. We find that synaptic ribbon precursors arrive at presynaptic active zones (AZs) after afferent contacts have been established. These ribbon precursors contain the proteins RIBEYE and piccolino, tether synaptic vesicles and their delivery likely involves active, microtubule-based transport pathways. Synaptic contacts undergo a maturational transformation from multiple small to one single, large AZ. This maturation is characterized by the fusion of ribbon precursors with membrane-anchored ribbons that also appear to fuse with each other. Such fusion events are most frequently encountered around P12 and hence, coincide with hearing onset in mice. Thus, these events likely underlie the morphological and functional maturation of the AZ. Moreover, the postsynaptic densities appear to undergo a similar refinement alongside presynaptic maturation. Blockwise addition of ribbon material by fusion as found during AZ maturation might represent a general mechanism for modulating ribbon size.
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http://dx.doi.org/10.1073/pnas.1812029116DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC6442603PMC
March 2019

Transmission Electron Microscopy of Oligodendrocytes and Myelin.

Methods Mol Biol 2019 ;1936:343-375

Max Planck Institute of Experimental Medicine, Department of Neurogenetics, Electron Microscopy Core Unit, Göttingen, Germany.

In this chapter, we describe protocols to study different aspects of oligodendrocytes and myelin using electron microscopy. First, we describe in detail how to prepare central nervous system tissue routinely by perfusion fixation of the animal and conventional embedding in Epon resin. Then, we explain how, with some modifications, chemically fixed tissue can be used for immunoelectron microscopy on cryosections. Chemical fixation and Epon embedding can also be applied to purified myelin to assess the quality of the preparation. Furthermore, we describe how cryopreparation by high-pressure freezing can be used to study the fine structure of myelin in nerve, brain, and spinal cord tissue. The differences in the structural appearance of oligodendrocytes and myelin between cryopreserved and conventionally processed samples are compared using representative images. Since primary cultured oligodendrocytes are used to study structure and function in vitro, we provide protocols for chemical fixation and Epon embedding of these cultures. Finally, we explain how the cytoskeleton of cultured oligodendrocytes can be visualized by using transmission electron microscopy on platinum-carbon replicas. In this chapter, we provide a wide range of protocols that can be applied to shed light on the different biological aspects of myelin and oligodendrocytes.
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http://dx.doi.org/10.1007/978-1-4939-9072-6_20DOI Listing
July 2019

Seeding variability of different alpha synuclein strains in synucleinopathies.

Ann Neurol 2019 05 27;85(5):691-703. Epub 2019 Mar 27.

Department of Neurology, University Medicine Goettingen and the German Center for Neurodegenerative Diseases (DZNE), Göttingen, Germany.

Objectives: Currently, the exact reasons why different α-synucleinopathies exhibit variable pathologies and phenotypes are still unknown. A potential explanation may be the existence of distinctive α-synuclein conformers or strains. Here, we intend to analyze the seeding activity of dementia with Lewy bodies (DLB) and Parkinson's disease (PD) brain-derived α-synuclein seeds by real-time quaking-induced conversion (RT-QuIC) and to investigate the structure and morphology of the α-synuclein aggregates generated by RT-QuIC.

Methods: A misfolded α-synuclein-enriched brain fraction from frontal cortex and substantia nigra pars compacta tissue, isolated by several filtration and centrifugation steps, was subjected to α-synuclein/RT-QuIC analysis. Our study included neuropathologically well-characterized cases with DLB, PD, and controls (Ctrl). Biochemical and morphological analyses of RT-QuIC products were conducted by western blot, dot blot analysis, Raman spectroscopy, atomic force microscopy, and transmission electron microscopy.

Results: Independently from the brain region, we observed different seeding kinetics of α-synuclein in the RT-QuIC in patients with DLB compared to PD and Ctrl. Biochemical characterization of the RT-QuIC product indicated the generation of a proteinase K-resistant and fibrillary α-synuclein species in DLB-seeded reactions, whereas PD and control seeds failed in the conversion of wild-type α-synuclein substrate.

Interpretation: Structural variances of α-synuclein seeding kinetics and products in DLB and PD indicated, for the first time, the existence of different α-synuclein strains in these groups. Therefore, our study contributes to a better understanding of the clinical heterogeneity among α-synucleinopathies, offers an opportunity for a specific diagnosis, and opens new avenues for the future development of strain-specific therapies. Ann Neurol 2019;85:691-703.
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http://dx.doi.org/10.1002/ana.25446DOI Listing
May 2019

Anillin facilitates septin assembly to prevent pathological outfoldings of central nervous system myelin.

Elife 2019 01 23;8. Epub 2019 Jan 23.

Department of Neurogenetics, Max Planck Institute of Experimental Medicine, Göttingen, Germany.

Myelin serves as an axonal insulator that facilitates rapid nerve conduction along axons. By transmission electron microscopy, a healthy myelin sheath comprises compacted membrane layers spiraling around the cross-sectioned axon. Previously we identified the assembly of septin filaments in the innermost non-compacted myelin layer as one of the latest steps of myelin maturation in the central nervous system (CNS) (Patzig et al., 2016). Here we show that loss of the cytoskeletal adaptor protein anillin (ANLN) from oligodendrocytes disrupts myelin septin assembly, thereby causing the emergence of pathological myelin outfoldings. Since myelin outfoldings are a poorly understood hallmark of myelin disease and brain aging we assessed axon/myelin-units in -mutant mice by focused ion beam-scanning electron microscopy (FIB-SEM); myelin outfoldings were three-dimensionally reconstructed as large sheets of multiple compact membrane layers. We suggest that anillin-dependent assembly of septin filaments scaffolds mature myelin sheaths, facilitating rapid nerve conduction in the healthy CNS.
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http://dx.doi.org/10.7554/eLife.43888DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC6344079PMC
January 2019