1. Physics of Living Systems
  2. Structural Biology and Molecular Biophysics

Nanoresolution real-time 3D orbital tracking for studying mitochondrial trafficking in vertebrate axons in vivo

  1. Fabian Wehnekamp
  2. Gabriela Plucińska
  3. Rachel Thong
  4. Thomas Misgeld  Is a corresponding author
  5. Don C Lamb  Is a corresponding author
  1. Ludwig Maximilian University of Munich, Germany
  2. Technische Universität München, Germany
https://doi.org/10.7554/eLife.46059
Share this article
Cite this article
  1. Fabian Wehnekamp
  2. Gabriela Plucińska
  3. Rachel Thong
  4. Thomas Misgeld
  5. Don C Lamb
(2019)
Nanoresolution real-time 3D orbital tracking for studying mitochondrial trafficking in vertebrate axons in vivo
eLife 8:e46059.
https://doi.org/10.7554/eLife.46059
  2. Download BibTeX
  3. Download .RIS

Abstract

We present the development and in vivo application of a feedback-based tracking microscope to follow individual mitochondria in sensory neurons of zebrafish larvae with nanometer precision and millisecond temporal resolution. By combining various technical improvements, we tracked individual mitochondria with unprecedented spatiotemporal resolution over distances of >100µm. Using these nanoscopic trajectory data, we discriminated five motional states: a fast and a slow directional motion state in both the anterograde and retrograde directions and a stationary state. The transition pattern revealed that mitochondria predominantly persist in the original direction of travel after a short pause, while transient changes of direction often exhibited longer pauses. Moreover, mitochondria in the vicinity of a second, stationary mitochondria displayed an increased probability to pause. The capability of following and optically manipulating a single organelle with high spatiotemporal resolution in a living organism offers a new approach to elucidating their function in its complete physiological context.

Data availability

The analysis software program is available on Gitlab and the wide-field images and trajectories are available on Zenodo. Source data files have been provided for all the figures.

The following data sets were generated

Article and author information

Author details

  1. Fabian Wehnekamp

    Physical Chemistry, Department for Chemistry and Center for Nanoscience, Ludwig Maximilian University of Munich, Munich, Germany
    Competing interests
    The authors declare that no competing interests exist.

  2. Gabriela Plucińska

    Institute of Neuronal Cell Biology, Technische Universität München, Munich, Germany
    Competing interests
    The authors declare that no competing interests exist.

  3. Rachel Thong

    Institute of Neuronal Cell Biology, Technische Universität München, Munich, Germany
    Competing interests
    The authors declare that no competing interests exist.

  4. Thomas Misgeld

    Institute of Neuronal Cell Biology, Technische Universität München, Munich, Germany
    For correspondence
    thomas.misgeld@tum.de
    Competing interests
    The authors declare that no competing interests exist.

  5. Don C Lamb

    Physical Chemistry, Department for Chemistry and Center for Nanoscience, Ludwig Maximilian University of Munich, Munich, Germany
    For correspondence
    d.lamb@lmu.de
    Competing interests
    The authors declare that no competing interests exist.
    ORCID icon "This ORCID iD identifies the author of this article:" 0000-0002-0232-1903

Funding

Deutsche Forschungsgemeinschaft (SFB1032 (Project B3))

  • Thomas Misgeld
  • Don C Lamb

Fakultät für Chemie und Pharmazie, Ludwig-Maximilians-Universität München (Center for NanoScience (CeNS) and the BioImaging Network (BIN))

  • Don C Lamb

H2020 European Research Council (ERC Grant Agreement n. 616791)

  • Thomas Misgeld

German Center for Neurodegenerative Diseases

  • Thomas Misgeld

Deutsche Forschungsgemeinschaft (research grants Mi 694/7)

  • Thomas Misgeld
  • Don C Lamb

Deutsche Forschungsgemeinschaft (Priority Program SPP1710)

  • Thomas Misgeld
  • Don C Lamb

Deutsche Forschungsgemeinschaft (SFB870 15 (Project A11))

  • Thomas Misgeld
  • Don C Lamb

The funders had no role in study design, data collection and interpretation, or the decision to submit the work for publication.

Copyright

© 2019, Wehnekamp et al.

This article is distributed under the terms of the Creative Commons Attribution License permitting unrestricted use and redistribution provided that the original author and source are credited.

Download links

Share this article

https://doi.org/10.7554/eLife.46059

Categories and tags

Research organism

Further reading

    1. Neuroscience
    2. Physics of Living Systems

    Annihilation of action potentials induces electrical coupling between neurons

    Moritz Schloetter, Georg U Maret, Christoph J Kleineidam
    Research Article

    Neurons generate and propagate electrical pulses called action potentials which annihilate on arrival at the axon terminal. We measure the extracellular electric field generated by propagating and annihilating action potentials and find that on annihilation, action potentials expel a local discharge. The discharge at the axon terminal generates an inhomogeneous electric field that immediately influences target neurons and thus provokes ephaptic coupling. Our measurements are quantitatively verified by a powerful analytical model which reveals excitation and inhibition in target neurons, depending on position and morphology of the source-target arrangement. Our model is in full agreement with experimental findings on ephaptic coupling at the well-studied Basket cell-Purkinje cell synapse. It is able to predict ephaptic coupling for any other synaptic geometry as illustrated by a few examples.

    1. Physics of Living Systems

    Modularity of the segmentation clock and morphogenesis

    James E Hammond, Ruth E Baker, Berta Verd
    Research Article

    Vertebrates have evolved great diversity in the number of segments dividing the trunk body, however, the developmental origin of the evolvability of this trait is poorly understood. The number of segments is thought to be determined in embryogenesis as a product of morphogenesis of the pre-somitic mesoderm (PSM) and the periodicity of a molecular oscillator active within the PSM known as the segmentation clock. Here, we explore whether the clock and PSM morphogenesis exhibit developmental modularity, as independent evolution of these two processes may explain the high evolvability of segment number. Using a computational model of the clock and PSM parameterised for zebrafish, we find that the clock is broadly robust to variation in morphogenetic processes such as cell ingression, motility, compaction, and cell division. We show that this robustness is in part determined by the length of the PSM and the strength of phase coupling in the clock. As previous studies report no changes to morphogenesis upon perturbing the clock, we suggest that the clock and morphogenesis of the PSM exhibit developmental modularity.

    1. Physics of Living Systems

    Emergence of ion-channel-mediated electrical oscillations in Escherichia coli biofilms

    Emmanuel Akabuogu, Victor Carneiro da Cunha Martorelli ... Thomas A Waigh
    Research Article

    Bacterial biofilms are communities of bacteria usually attached to solid strata and often differentiated into complex structures. Communication across biofilms has been shown to involve chemical signaling and, more recently, electrical signaling in Gram-positive biofilms. We report for the first time, community-level synchronized membrane potential dynamics in three-dimensional Escherichia coli biofilms. Two hyperpolarization events are observed in response to light stress. The first requires mechanically sensitive ion channels (MscK, MscL, and MscS) and the second needs the Kch-potassium channel. The channels mediated both local spiking of single E. coli biofilms and long-range coordinated electrical signaling in E. coli biofilms. The electrical phenomena are explained using Hodgkin-Huxley and 3D fire-diffuse-fire agent-based models. These data demonstrate that electrical wavefronts based on potassium ions are a mechanism by which signaling occurs in Gram-negative biofilms and as such may represent a conserved mechanism for communication across biofilms.