Cited by 197Mitochondrial Function and PathologyAdvanced Fluorescence Microscopy TechniquesMolecular Sensors and Ion Detection

Imaging Mitochondrial Flux in Single Cells with a FRET Sensor for Pyruvate

Open access
San Martín, Alejandro, Ceballo, Sebastián, Baeza-Lehnert, Felipe, Lerchundi, Rodrigo, Valdebenito, Rocío, Contreras-Baeza, Yasna, Alegría, Karin, Barros, L. Felipe·2014·PLoS ONE

Abstract

Mitochondrial flux is currently accessible at low resolution. Here we introduce a genetically-encoded FRET sensor for pyruvate, and methods for quantitative measurement of pyruvate transport, pyruvate production and mitochondrial pyruvate consumption in intact individual cells at high temporal resolution. In HEK293 cells, neurons and astrocytes, mitochondrial pyruvate uptake was saturated at physiological levels, showing that the metabolic rate is determined by intrinsic properties of the organelle and not by substrate availability. The potential of the sensor was further demonstrated in neurons, where mitochondrial flux was found to rise by 300% within seconds of a calcium transient triggered by a short theta burst, while glucose levels remained unaltered. In contrast, astrocytic mitochondria were insensitive to a similar calcium transient elicited by extracellular ATP. We expect the improved resolution provided by the pyruvate sensor will be of practical interest for basic and applied researchers interested in mitochondrial function.

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  • For PdhR-based FRET sensors, increasing pyruvate concentrations result in enhanced CFP emission and reduced YFP emission.

    “Similar to other previously reported PdhR-based FRET sensors74,75, increasing pyruvate concentrations result in enhanced CFP emission and reduced YFP emission.”

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Authors
San Martín, A., Ceballo, S., Baeza-Lehnert, F., Lerchundi, R., Valdebenito, R., Contreras-Baeza, Y., Alegría, K., & Barros, L. F.
Affiliations
  • Centro de Estudios Científicos (CL)
  • Austral University of Chile (CL)
Corresponding author
Barros, L. Felipe
Venue
PLoS ONE
Published
2014-01-21
Volume
9
Issue
1
Pages
e85780
ISSN
1932-6203 (ISSN-L)
OpenAlex
W1977850703
Cited by
197
References
37
Citations by year
2026: 9 · 2025: 18 · 2024: 18 · 2023: 15 · 2022: 17 · 2021: 19 · 2020: 21 · 2019: 20 · 2018: 17 · 2017: 16 · 2016: 10 · 2015: 10 · 2014: 7
FWCI
5.87
Type
journal article
Language
en
Open access
Locations
  • PLoS ONE (publishedVersion, cc-by)
  • Europe PMC (PubMed Central) (publishedVersion, cc-by)
  • PubMed Central (publishedVersion, cc-by)
  • PubMed (publishedVersion)
  • Americanae (AECID Library) (submittedVersion, other-oa)
  • DOAJ (DOAJ: Directory of Open Access Journals) (submittedVersion, cc-by-sa)
  • Figshare (submittedVersion, cc-by)
  • LA Referencia (Red Federada de Repositorios Institucionales de Publicaciones Científicas) (submittedVersion, other-oa)
Funders
  • Deutsche Forschungsgemeinschaft
  • Comisión Nacional de Investigación Científica y Tecnológica
Topics
  • Life Sciences › Biochemistry, Genetics and Molecular Biology › Molecular Biology › Mitochondrial Function and Pathology
  • Life Sciences › Biochemistry, Genetics and Molecular Biology › Biophysics › Advanced Fluorescence Microscopy Techniques
  • Physical Sciences › Chemistry › Spectroscopy › Molecular Sensors and Ion Detection
Keywords
pyruvate, FRET biosensor, genetically encoded biosensors, pyruvate transport, single-cell imaging, calcium transients, theta burst stimulation, extracellular ATP, neurons, astrocytes, HEK293 cells, mitochondrial metabolism
MeSH terms
Animals, Bacterial Proteins, Brain, Cytosol, Glycolysis, Humans, Luminescent Proteins, Male, Mammals, Mice, Inbred C57BL, Mitochondria, Repressor Proteins, Transcription, Genetic, Biosensing Techniques, Pyruvic Acid, Escherichia coli Proteins, Fluorescence Resonance Energy Transfer, Green Fluorescent Proteins, Mice, Molecular Imaging, HEK293 Cells, Single-Cell Analysis

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