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. https://doi.org/10.26124/mitofit:2022-0021 — 2022-07-28 published in Bioenerg Commun 2022.7.

» MitoFit Preprints 2022.21.

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) MitoFit Prep

Abstract:

Serna 2022 Abstract Bioblast: 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 • O2k-Network Lab: BR Sao Paulo Kowaltowski AJ

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: Instruments;methods

Stress:Permeability transition Organism: Human, Mouse Tissue;cell: Liver, Other cell lines Preparation: Permeabilized cells, Isolated mitochondria

Regulation: Calcium, Ion;substrate transport Coupling state: LEAK, OXPHOS Pathway: S, ROX HRR: Oxygraph-2k

Bioblast 2022

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 {{MitoFit page name}}
 {{Publication
-|title=Serna JDC, Ramos VM, Cabral-Costa JV, Vilas-Boas EA, Amaral AG, Ohya G, da Silva CCC, Kowaltowski AJ (2022) Measuring mitochondrial Ca<sup>2+</sup> efflux in isolated mitochondria and permeabilized cells. MitoFit Preprints 2022.21. https://doi.org/10.26124/mitofit:2022-0021
+|title=Serna JDC, Ramos VM, Cabral-Costa JV, Vilas-Boas EA, Amaral AG, Ohya G, da Silva CCC, Kowaltowski AJ (2022) Measuring mitochondrial Ca<sup>2+</sup> efflux in isolated mitochondria and permeabilized cells. https://doi.org/10.26124/mitofit:2022-0021 — ''2022-07-28 published in [https://www.bioenergetics-communications.org/index.php/bec/article/view/serna_2022 '''Bioenerg Commun 2022.7.''']''
-|info=[[File:MitoFit Preprints pdf.png|left|160px|link=https://wiki.oroboros.at/images/d/d2/Serna_2022_Mitofit.pdf|MitoFit pdf]] [https://wiki.oroboros.at/images/d/d2/Serna_2022_Mitofit.pdf Measuring mitochondrial Ca<sup>2+</sup> efflux in isolated mitochondria and permeabilized cells]<br/>
+|info=MitoFit Preprints 2022.21. [[File:MitoFit Preprints pdf.png|left|160px|link=https://wiki.oroboros.at/images/5/5a/Serna_2022_Mitofit.pdf|MitoFit pdf]] [https://wiki.oroboros.at/images/5/5a/Serna_2022_Mitofit.pdf Measuring mitochondrial Ca<sup>2+</sup> efflux in isolated mitochondria and permeabilized cells]<br/>
 |authors=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
-|year=2022-05-24
+|year=2022
 |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.
+[[Serna 2022 Abstract Bioblast]]: 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.
 |keywords=mitochondria, Ca<sup>2+</sup> efflux, NCLX, mPTP, liver
 |editor=[[Cecatto C]]
-|mipnetlab=
+|mipnetlab=BR Sao Paulo Kowaltowski AJ
 }}
-ORC'''ID''':[[File:ORCID.png|20px|link=https://orcid.org/]] Serna Julian DC, [[File:ORCID.png|20px|link=https://orcid.org/]] Ramos Vitor M, [[File:ORCID.png|20px|link=https://orcid.org/]] Cabral-Costa Joao V, [[File:ORCID.png|20px|link=https://orcid.org/]] Vilas-Boas Eloisa A, [[File:ORCID.png|20px|link=https://orcid.org/]] Amaral Andressa G, [[File:ORCID.png|20px|link=https://orcid.org/]] Ohya Georgia, [[File:ORCID.png|20px|link=https://orcid.org/]] da Silva Camille CC, [[File:ORCID.png|20px|link=https://orcid.org/]] Kowaltowski Alicia J
+ORC'''ID''':[[File:ORCID.png|20px|link=https://orcid.org/0000-0002-2285-5769]] Serna Julian DC, [[File:ORCID.png|20px|link=https://orcid.org/0000-0001-7004-0823]] Ramos Vitor M, [[File:ORCID.png|20px|link=https://orcid.org/0000-0001-5441-1807]] Cabral-Costa Joao V, [[File:ORCID.png|20px|link=https://orcid.org/0000-0003-1565-1618]] Vilas-Boas Eloisa A, [[File:ORCID.png|20px|link=https://orcid.org/0000-0002-1261-2982]] Amaral Andressa G, [[File:ORCID.png|20px|link=https://orcid.org/0000-0001-6608-7282]] Ohya Georgia, [[File:ORCID.png|20px|link=https://orcid.org/0000-0001-5158-071X]] da Silva Camille CC, [[File:ORCID.png|20px|link=https://orcid.org/0000-0002-3807-2419]] Kowaltowski Alicia J
 {{Labeling
-|area=Exercise physiology;nutrition;life style, Pharmacology;toxicology
+|area=Instruments;methods
-|preparations=
+|injuries=Permeability transition
-|enzymes=
+|organism=Human, Mouse
-|topics=
+|tissues=Liver, Other cell lines
-|couplingstates=
+|preparations=Permeabilized cells, Isolated mitochondria
-|instruments=
+|topics=Calcium, Ion;substrate transport
-|additional=Bioblast 2022,
+|couplingstates=LEAK, OXPHOS
+|pathways=S, ROX
+|instruments=Oxygraph-2k
+|additional=Bioblast 2022
 }}