Difference between revisions of "Serna 2022 MitoFit"

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Serna 2022 MitoFit

Serna JDC, Ramos VM, Cabral-Costa JV, Vilas-Boas EA, Amaral AG, Ohya G, da Silva CCC, Kowaltowski AJ (2022) Measuring mitochondrial Ca²⁺ efflux in isolated mitochondria and permeabilized cells. MitoFit Preprints 2022.21. https://doi.org/10.26124/mitofit:2022-0021

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Measuring mitochondrial Ca²⁺ efflux in isolated mitochondria and permeabilized cells

Serna Julian DC, Ramos Vitor M, Cabral-Costa Joao V, Vilas-Boas Eloisa A, Amaral Andressa G, Ohya Georgia, da Silva Camille CC, Kowaltowski Alicia J (2022-05-24) MitoFit Prep

Abstract:

Mitochondrial Ca²⁺ efflux is essential for mitochondrial and cell Ca²⁺ homeostasis. Mitochondrial inner membrane Ca²⁺/H⁺ and Na⁺/Li⁺/Ca²⁺ (NCLX) exchangers are known today to be plastic transporters, with important roles in physiological responses and pathological states. Until now, however, no consensus protocols were available to measure mitochondrial Ca²⁺ efflux, and we find that some published protocols may induce mitochondrial permeability transition, underestimating the effects of these exchangers. In this work we describe a method to measure Na⁺-sensitive and insensitive mitochondrial Ca²⁺ efflux activity in isolated mitochondria and permeabilized cells using the Ca²⁺ Green indicator and a fluorimeter. A checklist is provided to avoid artefacts as well as pinpoint adaptations necessary in specific experimental models. • Keywords: mitochondria, Ca²⁺ efflux, NCLX, mPTP, liver • Bioblast editor: Cecatto C

ORCID: Serna Julian DC, Ramos Vitor M, Cabral-Costa Joao V, Vilas-Boas Eloisa A, Amaral Andressa G, Ohya Georgia, da Silva Camille CC, Kowaltowski Alicia J

Labels: MiParea: Exercise physiology;nutrition;life style, Pharmacology;toxicology

Bioblast 2022

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 |year=2022-05-24
 |journal=MitoFit Prep
-|abstract=[[File:.png|right|250px|Graphical abstract]]
+|abstract=[[File:Serna 2022 MitoFit graphical abstract.png|right|250px|Graphical abstract]]
 Mitochondrial Ca<sup>2+</sup> efflux is essential for mitochondrial and cell Ca<sup>2+</sup> homeostasis. Mitochondrial inner membrane Ca<sup>2+</sup>/H<sup>+</sup> and Na<sup>+</sup>/Li<sup>+</sup>/Ca<sup>2+</sup> (NCLX) exchangers are known today to be plastic transporters, with important roles in physiological responses and pathological states. Until now, however, no consensus protocols were available to measure mitochondrial Ca<sup>2+</sup> efflux, and we find that some published protocols may induce mitochondrial permeability transition, underestimating the effects of these exchangers. In this work we describe a method to measure Na<sup>+</sup>-sensitive and insensitive mitochondrial Ca<sup>2+</sup> efflux activity in isolated mitochondria and permeabilized cells using the Ca<sup>2+</sup> Green indicator and a fluorimeter. A checklist is provided to avoid artefacts as well as pinpoint adaptations necessary in specific experimental models.