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

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.

Metrics

  • 2,416
    views
  • 345
    downloads
  • 35
    citations

Views, downloads and citations are aggregated across all versions of this paper published by eLife.

Download links

A two-part list of links to download the article, or parts of the article, in various formats.

Downloads (link to download the article as PDF)

Open citations (links to open the citations from this article in various online reference manager services)

Cite this article (links to download the citations from this article in formats compatible with various reference manager tools)

  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

Share this article

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

Further reading

    1. Cell Biology
    2. Physics of Living Systems
    Deb Sankar Banerjee, Shiladitya Banerjee
    Research Article

    Accurate regulation of centrosome size is essential for ensuring error-free cell division, and dysregulation of centrosome size has been linked to various pathologies, including developmental defects and cancer. While a universally accepted model for centrosome size regulation is lacking, prior theoretical and experimental works suggest a centrosome growth model involving autocatalytic assembly of the pericentriolar material. Here, we show that the autocatalytic assembly model fails to explain the attainment of equal centrosome sizes, which is crucial for error-free cell division. Incorporating latest experimental findings into the molecular mechanisms governing centrosome assembly, we introduce a new quantitative theory for centrosome growth involving catalytic assembly within a shared pool of enzymes. Our model successfully achieves robust size equality between maturing centrosome pairs, mirroring cooperative growth dynamics observed in experiments. To validate our theoretical predictions, we compare them with available experimental data and demonstrate the broad applicability of the catalytic growth model across different organisms, which exhibit distinct growth dynamics and size scaling characteristics.

    1. Cell Biology
    2. Physics of Living Systems
    Marta Urbanska, Yan Ge ... Jochen Guck
    Research Article

    Cell mechanical properties determine many physiological functions, such as cell fate specification, migration, or circulation through vasculature. Identifying factors that govern the mechanical properties is therefore a subject of great interest. Here, we present a mechanomics approach for establishing links between single-cell mechanical phenotype changes and the genes involved in driving them. We combine mechanical characterization of cells across a variety of mouse and human systems with machine learning-based discriminative network analysis of associated transcriptomic profiles to infer a conserved network module of five genes with putative roles in cell mechanics regulation. We validate in silico that the identified gene markers are universal, trustworthy, and specific to the mechanical phenotype across the studied mouse and human systems, and demonstrate experimentally that a selected target, CAV1, changes the mechanical phenotype of cells accordingly when silenced or overexpressed. Our data-driven approach paves the way toward engineering cell mechanical properties on demand to explore their impact on physiological and pathological cell functions.