• Fig. 1 Visualization of the dynamic microtubule cytoskeleton through the +TIP protein EB3

    Still image of movie depicting microtubule dynamics. Overlay of 30 images (1 image/s). EB3 marks microtubule plus ends (image 30). Red: EB3. Green: Trace of microtubule growth as a function of EB3 mobility over 29 s (images 1-29).

    Microtubules (MTs) are dynamic structures of alpha- and beta-tubulin. The dynamics of MTs can be visualized by the +TIP protein EB3. MTs mediate various functions in cells and act as tracks for cargo transport by molecular motor proteins. MTs are highly modified through posttranslational modifications (PTMs), such as acetylation, polyglutamylation or de-tyrosination. PTM patterns are thought to mediate an identity of MTs that encode cellular functions, such as cargo entry into specific subcellular compartments or cargo directionality (tubulin code hypothesis).

    The FOR 2419 project P1 uses tubulin polyglutamylation-deficient mutant mice for experiments that aim at i.) identifying whether and how neuronal activity and behavioral preconditioning alter PTMs at MT, and ii.) understanding whether and how conditions that alter the tubulin code, affect the synaptic delivery of plasticity-related proteins and ultimately learning and memory.

    Project P2 investigates the interplay between different motor proteins (Kevenaar, J.T. et al., Curr Biol 2016, 26: 849-61) with regard to i.) whether organelle positioning in dendrites is determined by a specialization of the local organization of the MT and actin cytoskeleton, and ii.) how local synapto-dendritic Ca2+ signaling affects the activity of different motors attached to the same cargo.

    Figure adapated from Janke, C., and Kneussel, M., Trends Neurosci 2010, 33:362-72.

    Fig. 2 F-actin is enriched in spines of live Purkinje neurons in dissociated cultures

    Filamentous actin (F-actin) is markedly enriched within spines as demonstrated in this figure with dissociated cerebellar cultures of live Purkinje neurons using a laser scanning confocal microscope (Z-stack projection of a Purkinje cell nucleofected with cell volume and F-actin makers).

    The postsynaptic actin cytoskeleton and its dynamic turnover are required for the remodeling of postsynaptic specializations during synaptic plasticity, for the regulation of synaptic AMPA receptor numbers, and for the formation and maintenance of spines. Project P3 of FOR 2419 examines the question of how myosin motors cooperate with the spine actin cytoskeleton to promote synaptic plasticity.

    Figure adapted from Wagner, W. et al.,. Nat Cell Biol 2011, 13:40-U101.

    Fig. 3 TAO2 accumulates on spines of mature cultured neurons and co-localizes with postsynaptic marker SynGAP

    By immunofluorescence staining of cultured cortical neurons and confocal imaging, the thousand-and-one-amino acid 2 kinase (TAO2), a member of the mitogen-activated protein Kinase Kinase Kinase (MAPKKK) family, was shown to accumulate on spines and to co-localize with a postsynaptic marker. Several findings suggest that it plays an important role in establishing brain connectivity (de Anda et al., Nat Neurosci 2012, 15:1022-31).

    Using primary neurons and the developing mouse brain as model systems, the project P5 investigates i.) whether TAO2 modulates spine formation via actin cytoskeleton / microtubules remodeling, and ii.) whether TAO2 affects synapse function mediating the transport of endoplasmatic reticulum (ER) into dendritic spines.

    Project P4 investigates the question whether the transient invasion of ER into individual spine synapses mediates a kind of metaplasticity (Holbro, N. et al., Proc Natl Acad Sci USA 2009, 106: 15055-60.).

    Fig. 4 Electron microscopy analysis of the fine structure of hippocampal region CA3 after high-pressure freezing

    A: Low-power electron micrograph illustrating all major tissue components of CA3 such as the cell bodies of pyramidal neurons (PC), proximal pyramidal cell dendrites (D), that give rise to large complex spines (S) in contact with mossy fiber boutons (MFB), and bundles of unmyelinated preterminal mossy fibers (MF). B: High-power electron micrograph of an MFB forming synaptic contacts (arrows) with a large complex spine (S). Arrowheads label non-synaptic puncta adhaerentia with a dendritic shapft (D). Scale bars = 1 µm.

    The FOR 2419 project P6 applies high-pressure freezing and electron microscopy (EM) immunogold labeling to study structural modifications at MF synapses in the hippocampus after induction of chemical long-term potentiation. These EM studies at defined time points are supplemented with 2-photon-microscopy studies allowing for time course analyses of structural changes using different fluorescent dyes to label presynaptic and postsynaptic elements of identified MF synapses.

    Figure adapted from Zhao, S. et al., J Comp Neurol 2012, 520: 2340-51.

    Fig. 5 Optical long-term plasticity experiments at single synapses in hippocampal neurons

    CA3 neurons expressing ChR2 (spectrally shifted channelrhodopsins) and a red fluorescent marker for synaptic vesicle clusters are stimulated with blue light to evoke transmission at CA1-CA3 Schaffer collateral synapses in organotypic hippocampal slice cultures. Spine volume and Ca2+ influx through NMDA receptors in post-synaptic spines are monitored by two-photon imaging of mCerulean (excitation: 810 nm) and GCaMP6s (980 nm). EPSC (excitatory postsynaptic current) is recorded in a reporter neuron.

    The working hypothesis of FOR 2419 project P7 is that the time of synapse removal is determined by the temporal correlation of pre- and postsynaptic spiking in a time window of several days (Wiegert, J.S., and Oertner, T.G., Proc Natl Acad Sci USA 2013, 110: E4510-19).

Logo FOR 2419
Funding Period 2016 - 2018

Plasticity versus Stability:

Molecular Mechanisms of Synaptic Strength

Coordinator: Prof. Dr. Matthias Kneussel

Research Unit FOR 2419 funded by the DFG

Contact: Dr. Eva-Maria Suciu, Tel.: +49 (0) 40 7410-55081

eva-maria.suciu@zmnh.uni-hamburg.de

  • P1 Matthias Kneussel, ZMNH Institute for Molecular Neurogenetics

    Delivery of Plasticity-Related Proteins (PRPs) in Synaptic Consolidation

    We will investigate the role of the “tubulin code” in regulating synapse delivery via microtubules, using three newly generated tubulin knock-in mouse lines that carry mutations to abolish polyglutamylation (poly-Glu). Synaptic delivery of AMPA- and NMDA receptors will be assessed by FRAP and live cell imaging in neurons. Optogenetic protocols will help to specifically induce neuronal activity in hippocampal slices. Poly-Glu tubulin will be visualized by EM. We will ask whether altered poly-Glu tubulin affects synaptic plasticity and learning and vice versa whether behavioral training alters poly-Glu patterns along the cytoskeleton.

    P2 Marina Mikhaylova, ZMNH Research Group Neuronal Protein Transport

    Functional Interplay of Microtubule and Actin Motors in Dendritic Compartmentalization

    Dendritic secretory organelles are local supply stations for plasticity-related products. In this project we would like to understand how the interplay between motor proteins allows for controlled cargo delivery, retention or release in response to synaptic activity in dendritic branch compartments. We hypothesize that i) the organelle positioning in dendrites is determined by local organization of the microtubule and actin cytoskeleton and ii) by local synapto-dendritic Ca2+ signals, which can regulate the activity of different motors on the same cargo allowing for controlled coordination of motility. We aim to gain a better understanding of basic molecular mechanisms involved in regulation of synaptic and dendritic transport mediated by different microtubule- and actinassociated motor proteins and to translate these findings to the models of synaptic plasticity.

    P3 Wolfgang Wagner, Institute for Molecular Neurogenetics

    Mechanisms of Actomyosin-Dependent Regulation of Postsynaptic Function and Plasticity in Purkinje Cells

    Members of the myosin family of actin-based cytoskeletal motors play crucial roles for synaptic plasticity at excitatory synapses. Two neuronal myosins that remain to be characterized in this respect are myosin XVI and myosin Id. Strikingly, these myosins physically interact with synaptic plasticity key regulators and appear to be genetically associated with psychiatric disorders. We hypothesize that myosin XVI and myosin Id regulate AMPA receptor trafficking and/or the actin cytoskeleton at the postsynaptic side. Our aim is to test this hypothesis and to uncover the function of these myosins in vitro and in vivo using cerebellar Purkinje cells as a model system. We expect to shed new light on the mechanisms of myosin-dependent synaptic plasticity regulation.

    P4 Thomas Oertner, ZMNH Institute for Synaptic Physiology

    Impact of Spine Endoplasmic Reticulum on Synaptic Function and Plasticity

    A subset of spines on hippocampal pyramidal cells contains endoplasmic reticulum (ER), either as a single thin tube protruding into the spine or in the form of a differentiated ‘spine apparatus‘. We demonstrated that synapses with ER reduce their strength after activation of mGluRs, but synapses on spines that lack ER do not support this form of long-term depression (Holbro et al., PNAS 2009). We would like to investigate which signals and molecular mechanisms direct ER tubules into specific spines and how much time it takes to assemble a fully fledged spine apparatus. Furthermore, we want to find out how the presence of ER affects synaptic plasticity and structure and long-term stability of the spine

    P5 Froylan Calderon de Anda, ZMNH Research Group Neuronal Development

    The Role of TAO2 in Synapse Formation and Plasticity

    TAO2 is a member of the MAP Kinase Kinase Kinase (MAPKKK) family. In humans, the gene encoding TAO2 is located on chromosome 16p11.2, a region that carries substantial susceptibility to autism spectrum disorders (ASD) and schizophrenia. We demonstrated a novel role for TAO2 in the development of axons and dendrites (Calderon de Anda F. et al., Nat. Neurosci. 2012). In the proposed project, we will test whether TAO2 is a key player of synapse formation and function. To directly test this, we will elucidate: a) whether TAO2 modulates spine formation via actin cytoskeleton / microtubules remodeling, and b) whether TAO2 depletion affects synapse function mediating endoplasmic reticulum (ER) transport into dendritic spines.

    P6 Michael Frotscher, ZMNH Institute for Structural Neurobiology

    Structural Plasticity of Hippocampal Mossy Fiber Synapses

    In this project we aim to characterize the molecular and structural changes associated with functional plasticity of identified hippocampal mossy fiber (MF) synapses. We will use high pressure freezing (HPF) for our electron microscopic studies to minimize tissue alteration such as protein denaturation and tissue shrinkage. We will use 2-photon microscopy to monitor the time course of activity-induced structural changes at identified MF synapses. 2-photon microscopy will also be used to record calcium transients in spines postsynaptic to MF boutons as a read-out of synaptic strength.

    P7 Christine E. Gee and J. Simon Wiegert, ZMNH Institute for Synaptic Physiology

    Dynamic Rewiring of Hippocampal Circuits Following Synaptic Plasticity

    We propose to study how the lifetime of excitatory synapses is regulated by activity. In organotypic slice cultures of rat hippocampus, we will combine optogenetic induction of long-term plasticity (LTP, LTD) at identified synapses with optogenetic modulation of synaptic activity during the following days. Controlling the activity of identified neurons allows us to investigate the long-term consequences of synaptic plasticity rules on the network level. Do new synapses form at random or do they preferentially form between best synchronized pre- and postsynaptic neuronal populations? The ultimate goal is to understand the connection between functional and structural synaptic plasticity and to decipher the rules of activity-dependent brain wiring.

    Summary Research Foci of FOR 2419

    Methodological Approches of FOR 2419
    Methodological Approches of FOR 2419

    Neuronal networks operate in intricate circuits to regulate cognitive processes such as learning and memory. Individual neurons are highly plastic by forming and retracting synapses in a neuronal activity-dependent manner. At the molecular level, our understanding of synaptic plasticity is still like a peek through and keyhole. Basic research is therefore required to unravel the mechanisms underlying the structural and functional modifiability of synapses in a given network and, in the long-term to fight synaptopathies.

    The DFG Research Unit FOR 2419 combines molecular biology and mouse genetics with network physiology and optogenetic approaches to address the conflict of “plasticity” versus “stability” at neuronal synapses. Since the majority of molecular components at synapses are highly dynamic and undergo rapid turnover, we ask how does a dynamic system of this kind encode stability in neuronal connectivity and ultimately behavior?

    A central question will be to investigate the molecular mechanisms that stabilize or consolidate synaptic structure and function in order for plastic changes to become persistent. To address these questions we combine experts in studying cytoskeleton transport and local synaptic trafficking with experts in neurophysiology, calcium imaging, optogenetics and synapse structure.

    A main goal is to understand the crosstalk between activity- and calcium-dependent processes with the delivery and removal of synaptic components. Combined investigations of synaptic trafficking with optogenetics and physiology creates a powerful interdisciplinary approach that is currently unique across Germany. In the long-term, we aim to take advantage of optogenetic approaches in order to bridge the molecular level of synaptic research with our understanding of temporal network coordination and cognitive performance in intact animals.

    More Details and References (pdf)

  • December 15, 2016 at 2 pm

    Prof. Dr. Victor Tarabykin , Institute of Cell Biology and Neurobiology, Charite Berlin

    Molecular control of the neocortex development

    Venue: Center for Molecular Neurobiology Hamburg (ZMNH), seminar room E.82

    January 19, 2017 at 2 pm

    Dr. Benjamin Rost , German Center for Neurodegenerative Diseases e.V. (DZNE), Berlin

    Synaptic Wiring and Information Processing

    Venue: Center for Molecular Neurobiology Hamburg (ZMNH), seminar room E.82

    February 16, 2017 at 2 pm

    Prof. Dr. Imre Vida , Institute of Integrative Neuroanatomy and NeuroCure Cluster, Berlin

    Venue: Center for Molecular Neurobiology Hamburg (ZMNH), seminar room E.82

    March 16, 2017 at 2 pm

    Dr. Frédéric Gambino , Interdisciplinary Institute for Neuroscience, University Bordeaux

    Dendritic mechanisms for associative learning in behaving animals

    Venue: Center for Molecular Neurobiology Hamburg (ZMNH), seminar room E.82

    April 20, 2017 at 2 pm

    Prof. Dr. Anthony Holtmaat , FG Structural synaptic plasticity, Geneva Neuroscience Center

    Venue: Center for Molecular Neurobiology Hamburg (ZMNH), seminar room E.82

    June 22,2017 at 2 pm

    Prof. Dr. Siegrid Löwel , Bernstein Focus for Neurotechnology, University of Göttingen

    Venue: Center for Molecular Neurobiology Hamburg (ZMNH), seminar room E.82

    July 13, 2017 at 2 pm

    Prof. Dr. Wieland Huttner , MPI of Molecular Biology and Genetics, Dresden

    Neural stem and progenitor cells and neocortex expansion in development and evolution

    Venue: Center for Molecular Neurobiology Hamburg (ZMNH), seminar room E.82

    September 14, 2017 at 2 pm

    Dr. Andrea Barberis , Istituto Italiano di Tecnologia, Genova, Italy

    Venue: Center for Molecular Neurobiology Hamburg (ZMNH), seminar room E.82

    October 12, 2017 at 2 pm

    Prof. Dr. Claudia Bagni , University of Lausanne

    Venue: Center for Molecular Neurobiology Hamburg (ZMNH), seminar room E.82

    December 14, 2017 at 2 pm

    Dr. Sabine Lévi , Institut du Fer à Moulin, Paris

    Venue: Center for Molecular Neurobiology Hamburg (ZMNH), seminar room E.82

    Past FOR 2419 - Seminars

  • The 37th Blankenese Conference entitled Synaptic Plasticity versus Stability - Information Update, Processing and Coding will cover the focus of FOR 2419 complemented by aspects of the plasticity of cellular circuits in the olfactory and taste systems to illuminate the role of synapses in information processing in the central and peripheral nervous system.

    Confirmed speakers (pdf)

    Registration is limited to 100 participants (including invited speakers) on a first-come first-serve basis. Please apply via the Blankenese Conference website . The deadline for registration is March 1st, 2017.

  • Each doctoral student involved in a project of the FOR 2419 is tutored by a principal investigator of the FOR 2419. He or she concludes a supervision agreement with this main supervisor and two mentors; one of the mentors is another principal investigator of the FOR 2419. The supervision agreement specifies the rights and duties of the persons involved such as a yearly thesis committee meeting in which the doctoral student presents a progress report on the dissertation to the main supervisor and the mentors. Successfully completed doctoral studies will lead to the award of a doctoral degree in conformity with the regulations of the Faculty of Mathematics, Informatics and Natural Sciences (MIN Faculty) or the Faculty of Medicine of Universität Hamburg.

    This supervision concept is part of the ZMNH PhD Program which aims at ensuring the best possible supervision and support of doctoral students as well as a high quality of interdisciplinary academic education. Furthermore, two ombudspersons and two PhD student representatives, annually elected by the ZMNH Scientists’ Conference, support doctoral students’ activities at the ZMNH. For example, the ZMNH Doctoral Students’ Journal Club meets regularly and there are biweekly Internal PhD-Seminars for discussing own research findings.

    In addition to the support and supervision of doctoral students’ thesis research, the attendance of ZMNH-Seminars, seminars of the Hamburg Center of NeuroScience and of the lectures and research methods courses of the ZMNH-based Graduate Program in Molecular Biology (ASMB) is offered. Moreover, the doctoral students may join interdisciplinary, research methods and academic key skill courses offered by the Faculty of Medicine, the MIN Faculty as well as the Career Center of Universität Hamburg.

  • Peer-reviewed Publications

    Anda FC, Madabhushi R, Rei D, Meng J, Graff J, Durak O, Meletis K, Richter M, Schwanke B, Mungenast A, Tsai LH (2016) Cortical neurons gradually attain a post-mitotic state. Cell Res 26:1033-1047.

    Bär J, Kobler O, van Bommel B, Mikhaylova M (2016) Periodic F-actin structures shape the neck of dendritic spines. Sci Rep. 2016 Nov 14;6:37136. doi: 10.1038/srep37136.

    Brill MS, Kleele T, Ruschkies L, Wang M, Marahori NA, Reuter MS, Hausrat TJ, Weigand E, Fisher M, Ahles A, Engelhardt S, Bishop DL, Kneussel M, Misgeld T (2016) Branch-Specific Microtubule Destabilization Mediates Axon Branch Loss during Neuromuscular Synapse Elimination. Neuron, http://dx.doi.org/10.1016/j.neuron.2016.09.049

    Kneussel, M., and Hausrat, T.J. (2016). Postsynaptic Neurotransmitter Receptor Reserve Pools for Synaptic Potentiation. Trends Neurosci 39, 170-182. Review

    Muhia, M., Thies, E., Labonte, D., Ghiretti, A.E., Gromova, K.V., Xompero, F., Lappe-Siefke, C., Hermans-Borgmeyer, I., Kuhl, D., Schweizer, M., Ohana, O., Schwarz, J.R., Holzbaur, E.L., and Kneussel, M. (2016). The Kinesin KIF21B Regulates Microtubule Dynamics and Is Essential for Neuronal Morphology, Synapse Function, and Learning and Memory. Cell Rep 15, 968-977.

    van Bommel B, Mikhaylova M (2016) Talking to the neighbours: The molecular and physiological mechanisms of clustered synaptic plasticity. Neurosci Biobehav Rev 71:352-361. Review

    Other Publications

    Kneussel, M (2016) DFG Forschergruppe FOR 2419 „Plastizität versus Stabilität: Molekulare Mechanismen der Synapsenstärke“. Neuroforum 22, 60-61.