Investigating mechanisms of myelin sheath length regulation and plasticity

Myelination of axons is important for proper functioning of the nervous system and breakdown of myelin can cause severe disabilities. By regulating nerve conduction, myelination is also critical for learning and memory processes. Myelination greatly influences conduction properties and remodeling of myelin has been proposed as a potential mechanism to adjust and modulated nervous system function. However, to date it is still unclear if existing myelin is able to remodel and therefore participate in brain plasticity. I used existing zebrafish transgenic lines and generated new constructs to visualize myelinated axons in the CNS and to perform in vivo life imaging. Furthermore, I developed a single cell ablation method with high spatial and temporal precision to selectively demyelinate axon stretches and assess remyelination and remodeling dynamics. Using these tools, I was able to describe the growth dynamics of single myelin sheaths and show that they are independent of neighboring sheaths and time of initiation. Myelin sheath growth can be divided into three different growth phases, an oligodendrocyte intrinsic, highly uniformly growth phase that lasts for about 8 hours, followed by a second variable growth phase, likely regulated by axon intrinsic mechanisms, in which sheath length differences are established. The last growth phase compensates for body growth, and is highly predictable by the length increase of the animal. By demyelination of short axon stretches I was able to show that myelin segments are able to deviate from their otherwise very stereotypic growth dynamics. Ablation of a myelin sheath resulted in reinitiation of fast sheath growth in the neighboring sheath to remyelinate the gap. A new sheath was formed in the gap and grew which often led to a pushing back of the neighboring sheaths that had invaded the demyelinated territory. Thereby, often re-establishing the pre-ablation pattern, indicating a homeostatic regulation of myelin sheath length along an axon. Similarly, partially myelinated axons regularly restored their pre-ablation pattern after demyelination. Together, these results indicated axonal control of myelin sheath length and node of Ranvier positioning to guide the restoration of pre-ablation patterns. Furthermore, I observed a high number of asymmetrically grown sheath that could not be explained by physical barriers like neighboring sheaths or axon collaterals, indicating the existence of a molecular growth barrier on the axon. To collect further evidence, I investigated the dynamics of the nodal marker Neurofascin and found that it forms clusters along unmyelinated axons which are predictive for node of Ranvier positions. In order to test if the formation of clusters and the positioning of nodes is axonal activity dependent I established an optogenetic setup for long-term stimulation of freely swimming fish. By manipulating axonal activity by optogenetics I was able to induce myelin sheath remodeling supporting the hypothesis of axonal regulation of node or Ranvier positioning, however, similar effects were observed in control animals. Together, I was able to describe the dynamics of myelin sheath growth and could show that existing myelin segments can remodel and are therefore able to participate in brain plasticity. Additionally, I collected evidence that node or Ranvier positioning and therefore also myelin sheath length are regulated by axonal mechanisms

Investigating mechanisms of myelin sheath length regulation and plasticity

Myelination of axons is important for proper functioning of the nervous system and breakdown of myelin can cause severe disabilities. By regulating nerve conduction, myelination is also critical for learning and memory processes. Myelination greatly influences conduction properties and remodeling of myelin has been proposed as a potential mechanism to adjust and modulated nervous system function. However, to date it is still unclear if existing myelin is able to remodel and therefore participate in brain plasticity. I used existing zebrafish transgenic lines and generated new constructs to visualize myelinated axons in the CNS and to perform in vivo life imaging. Furthermore, I developed a single cell ablation method with high spatial and temporal precision to selectively demyelinate axon stretches and assess remyelination and remodeling dynamics. Using these tools, I was able to describe the growth dynamics of single myelin sheaths and show that they are independent of neighboring sheaths and time of initiation. Myelin sheath growth can be divided into three different growth phases, an oligodendrocyte intrinsic, highly uniformly growth phase that lasts for about 8 hours, followed by a second variable growth phase, likely regulated by axon intrinsic mechanisms, in which sheath length differences are established. The last growth phase compensates for body growth, and is highly predictable by the length increase of the animal. By demyelination of short axon stretches I was able to show that myelin segments are able to deviate from their otherwise very stereotypic growth dynamics. Ablation of a myelin sheath resulted in reinitiation of fast sheath growth in the neighboring sheath to remyelinate the gap. A new sheath was formed in the gap and grew which often led to a pushing back of the neighboring sheaths that had invaded the demyelinated territory. Thereby, often re-establishing the pre-ablation pattern, indicating a homeostatic regulation of myelin sheath length along an axon. Similarly, partially myelinated axons regularly restored their pre-ablation pattern after demyelination. Together, these results indicated axonal control of myelin sheath length and node of Ranvier positioning to guide the restoration of pre-ablation patterns. Furthermore, I observed a high number of asymmetrically grown sheath that could not be explained by physical barriers like neighboring sheaths or axon collaterals, indicating the existence of a molecular growth barrier on the axon. To collect further evidence, I investigated the dynamics of the nodal marker Neurofascin and found that it forms clusters along unmyelinated axons which are predictive for node of Ranvier positions. In order to test if the formation of clusters and the positioning of nodes is axonal activity dependent I established an optogenetic setup for long-term stimulation of freely swimming fish. By manipulating axonal activity by optogenetics I was able to induce myelin sheath remodeling supporting the hypothesis of axonal regulation of node or Ranvier positioning, however, similar effects were observed in control animals. Together, I was able to describe the dynamics of myelin sheath growth and could show that existing myelin segments can remodel and are therefore able to participate in brain plasticity. Additionally, I collected evidence that node or Ranvier positioning and therefore also myelin sheath length are regulated by axonal mechanisms

Evidence for Myelin Sheath Remodeling in the CNS Revealed by In Vivo Imaging

The length of myelin sheaths affects conduction speed along axons and information propagation. It has recently become clear that myelin may be adaptively modified to modulate circuit function, implying that length remodeling of myelin sheaths should occur. However, direct evidence for such events is lacking. We have investigated how myelination patterns are formed, maintained, and remodeled using long-term imaging and myelin ablation in zebrafish. We demonstrate that length differences between myelin sheaths are established by rapid and variable growth within 3 days after their formation, independently of their time of formation, and even along discontinuously myelinated axons. Afterward, sheaths continue extending at similar rates to compensate for overall animal growth. In consequence, once axon myelination patterns are established, they are maintained over long periods of time. We tested whether mature myelin sheaths can remodel by removing individual sheaths from single axons by targeted ablation. Remarkably, extensive changes in sheath length and number occurred, which frequently restored the original myelination pattern. Our results show that axons can control myelin growth and remodeling, and we provide evidence for a homeostatic control of axon myelination patterns by maintenance and remodeling of myelin sheath length, with implications for circuit development, function, and repair

Clusters of neuronal neurofascin prefigure the position of a subset of nodes of Ranvier along individual central nervous system axons in vivo

The spacing of nodes of Ranvier crucially affects conduction properties along myelinated axons. It is assumed that node position is primarily driven by growing myelin sheaths. Here, we reveal an additional mechanism of node positioning that is driven by the axon. Through longitudinal live imaging of node formation dynamics in the zebrafish central nervous system, we show that stable clusters of the cell adhesion molecule neurofascin a can accumulate at specific sites along axons prior to myelination. While some of these clusters are pushed into future node position by extending myelin sheaths, others are not and thus prefigure the position of where a mature node forms. Animals that lack full-length neurofascin a show increased internodal distances and less regular nodal spacing along single axons. Together, our data reveal the existence of an axonal mechanism to position nodes of Ranvier that does not depend on regulation by myelin sheath growth

Impact of Disease-Modifying Treatments on the Longitudinal Evolution of Anti-JCV Antibody Index in Multiple Sclerosis

Background: Risk of natalizumab-related progressive multifocal leukoencephalopathy is associated with the presence of anti-JC-virus (JCV) antibodies.Objective: To investigate the impact of disease-modifying treatments (DMT) on the longitudinal evolution of anti-JCV antibody index.Methods: Patients with multiple sclerosis who had serum sampling at intervals of 6 ± 3 months over up to 6 years and who either started DMT (interferon-β, glatiramer acetate or natalizumab) during the observation period with at least one serum sample available before and after treatment initiation or received no DMT during the observation period were included. Anti-JCV antibody serological status and index were determined by 2-step second-generation anti-JCV antibody assay.Results: A total of 89 patients were followed for a median time of 55.2 months. Of those, 62 (69.7%) started DMT and 27 (30.3%) were without therapy during the observation period. Variation of longitudinal anti-JCV antibody index ranged from 9 to 15% and was similar in patients with and without DMT. Applying a mixed model considering the combined effects of treatment and time as well as individual heterogeneity did not show a significant change of anti-JCV antibody index by the start of treatment with interferon-β, glatiramer acetate, or natalizumab.Conclusion: Evaluated DMTs do not impact longitudinal anti-JCV antibody index evolution

Immune profiling in multiple sclerosis: a single-center study of 65 cytokines, chemokines, and related molecules in cerebrospinal fluid and serum

IntroductionThe understanding of the pathophysiology of multiple sclerosis (MS) has evolved alongside the characterization of cytokines and chemokines in cerebrospinal fluid (CSF) and serum. However, the complex interplay of pro- and anti-inflammatory cytokines and chemokines in different body fluids in people with MS (pwMS) and their association with disease progression is still not well understood and needs further investigation. Therefore, the aim of this study was to profile a total of 65 cytokines, chemokines, and related molecules in paired serum and CSF samples of pwMS at disease onset.MethodsMultiplex bead-based assays were performed and baseline routine laboratory diagnostics, magnetic resonance imaging (MRI), and clinical characteristics were assessed. Of 44 participants included, 40 had a relapsing–remitting disease course and four a primary progressive MS.ResultsThere were 29 cytokines and chemokines that were significantly higher in CSF and 15 in serum. Statistically significant associations with moderate effect sizes were found for 34 of 65 analytes with sex, age, CSF, and MRI parameters and disease progression.DiscussionIn conclusion, this study provides data on the distribution of 65 different cytokines, chemokines, and related molecules in CSF and serum in newly diagnosed pwMS

Combining Computational Prediction of Cis-Regulatory Elements with a New Enhancer Assay to Efficiently Label Neuronal Structures in the Medaka Fish

The developing vertebrate nervous system contains a remarkable array of neural cells organized into complex, evolutionarily conserved structures. The labeling of living cells in these structures is key for the understanding of brain development and function, yet the generation of stable lines expressing reporter genes in specific spatio-temporal patterns remains a limiting step. In this study we present a fast and reliable pipeline to efficiently generate a set of stable lines expressing a reporter gene in multiple neuronal structures in the developing nervous system in medaka. The pipeline combines both the accurate computational genome-wide prediction of neuronal specific cis-regulatory modules (CRMs) and a newly developed experimental setup to rapidly obtain transgenic lines in a cost-effective and highly reproducible manner. 95% of the CRMs tested in our experimental setup show enhancer activity in various and numerous neuronal structures belonging to all major brain subdivisions. This pipeline represents a significant step towards the dissection of embryonic neuronal development in vertebrates

Catalyst development for the dehydrogenation of perhydro-dibenzyltoluene

In dieser Arbeit wurde die katalytische Dehydrierung von Perhydro-Dibenzyltoluol bei Temperaturen um 300 °C untersucht. Die Reaktion stellt einen wichtigen Teilschritt bei der Wasserstoffspeicherung mittels Liquid Organic Hydrogen Carrier (LOHC) dar. Ein Katalysatorsystem zur Freisetzung von Wasserstoff, das bereits technisch eingesetzt wird, wurde in einer vorhergehenden Arbeit entwickelt. In dieser Arbeit wurde dieses Katalysatorsystem hinsichtlich einer reduzierten Nebenproduktbildung weiterentwickelt. In diesem Zuge wurde der Einfluss verschiedener Katalysatoreigenschaften auf die Bildung von Nebenprodukten untersucht und ein Verständnis für die Vorgänge auf molekularer Ebene erhalten. Ziel war es, wesentliche Charakteristika eines aktiven und selektiven Katalysators für die Dehydrierung von Perhydro-Dibenzyltoluol herauszuarbeiten.In this thesis the catalytic dehydrogenation of perhydro-dibenzyltoluene at temperatures around 300 °C was investigated. The reaction is an important step in the technology of hydrogen storage by Liquid Organic Hydrogen Carrier (LOHC). A catalyst system for the release of hydrogen, which is already technically used, was developed in a previous work. The object of this work was to further develop this catalyst system with regard to reduced by-product formation. In this context, the influence of different catalyst properties on the formation of by-products was investigated and a deeper understanding of the processes at a molecular level was obtained. The aim was to identify essential characteristics of an active and at the same time selective catalyst for the dehydrogenation of perhydro-dibenzyltoluene