Optical estimation of absolute membrane potential using fluorescence lifetime imaging

  1. Julia Rose Lazzari-Dean
  2. Anneliese M M Gest
  3. Evan W Miller  Is a corresponding author
  1. University of California, Berkeley, United States

Abstract

All cells maintain ionic gradients across their plasma membranes, producing transmembrane potentials (Vmem). Mounting evidence suggests a relationship between resting Vmem and the physiology of non-excitable cells with implications in diverse areas, including cancer, cellular differentiation, and body patterning. A lack of non-invasive methods to record absolute Vmem limits our understanding of this fundamental signal. To address this need, we developed a fluorescence lifetime-based approach (VF-FLIM) to visualize and optically quantify Vmem with single-cell resolution in mammalian cell culture. Using VF-FLIM, we report Vmem distributions over thousands of cells, a 100-fold improvement relative to electrophysiological approaches. In human carcinoma cells, we visualize the voltage response to growth factor stimulation, stably recording a 10-15 mV hyperpolarization over minutes. Using pharmacological inhibitors, we identify the source of the hyperpolarization as the Ca2+-activated K+ channel KCa3.1. The ability to optically quantify absolute Vmem with cellular resolution will allow a re-examination of its signaling roles.

Data availability

All data presented in the manuscript is available in the supporting / supplementary information.

Article and author information

Author details

  1. Julia Rose Lazzari-Dean

    Department of Chemistry, University of California, Berkeley, Berkeley, United States
    Competing interests
    No competing interests declared.
    ORCID icon "This ORCID iD identifies the author of this article:" 0000-0003-2971-5379
  2. Anneliese M M Gest

    Department of Chemistry, University of California, Berkeley, Berkeley, United States
    Competing interests
    No competing interests declared.
  3. Evan W Miller

    Department of Chemistry, University of California, Berkeley, Berkeley, United States
    For correspondence
    evanwmiller@berkeley.edu
    Competing interests
    Evan W Miller, is listed as an inventor on a patent describing voltage-sensitive fluorophores. This patent (US20170315059) is owned by the Regents of the University of California.
    ORCID icon "This ORCID iD identifies the author of this article:" 0000-0002-6556-7679

Funding

National Science Foundation (GRFP)

  • Julia Rose Lazzari-Dean

National Institutes of Health (R35GM119855)

  • Evan W Miller

Alfred P. Sloan Foundation (FG-2016-6359)

  • Evan W Miller

March of Dimes Foundation (5-FY-16-65)

  • Evan W Miller

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

Reviewing Editor

  1. Lawrence Cohen, Yale, United States

Version history

  1. Received: February 14, 2019
  2. Accepted: September 16, 2019
  3. Accepted Manuscript published: September 23, 2019 (version 1)
  4. Version of Record published: October 25, 2019 (version 2)

Copyright

© 2019, Lazzari-Dean 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.

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  1. Julia Rose Lazzari-Dean
  2. Anneliese M M Gest
  3. Evan W Miller
(2019)
Optical estimation of absolute membrane potential using fluorescence lifetime imaging
eLife 8:e44522.
https://doi.org/10.7554/eLife.44522

Share this article

https://doi.org/10.7554/eLife.44522

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    Funding:

    This work was supported by the Weizmann Krenter Foundation and the Weizmann – Ichilov (Tel Aviv Sourasky Medical Center) Collaborative Grant in Biomedical Research, by the Minerva Foundation, by the ISF KillCorona grant 3777/19.