Difference between revisions of "Serna 2022 MitoFit"
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{{Publication | {{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. MitoFit Preprints 2022.21. https://doi.org/10.26124/mitofit:2022-0021 | ||
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|journal=MitoFit Prep | |journal=MitoFit Prep | ||
|abstract=[[File:Serna 2022 MitoFit graphical abstract.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. Β | 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. | ||
Β | |||
|keywords=mitochondria, Ca<sup>2+</sup> efflux, NCLX, mPTP, liver | |keywords=mitochondria, Ca<sup>2+</sup> efflux, NCLX, mPTP, liver | ||
|editor=[[Cecatto C]] | |editor=[[Cecatto C]] | ||
}} | }} | ||
{{Labeling | {{Labeling | ||
|area= | |area=Instruments;methods | ||
| | |injuries=Permeability transition | ||
| | |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 | |||
}} | }} | ||
[[File:Bioblast2022 banner.jpg|link=Bioblast_2022]] | |||
{{MitoFit page name}} | |||
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 |
Revision as of 14:33, 24 May 2022
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 |
Β»
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. β’ Keywords: mitochondria, Ca2+ efflux, NCLX, mPTP, liver β’ Bioblast editor: Cecatto C
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
Serna 2022 MitoFit
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