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Chicco 2022 MitoFit

Chicco AJ, Zilhaver PT, Whitcomb LA, Fresa KJ, Izon CS, Gonzalez-Franquesa A, Izon CS, Dometita C, Irving BA, Garcia-Roves PM (2022) Resolving the Rotenone Paradox: elucidating the complexity of multi-substrate respirometry protocols. MitoFit Preprints 2022.17. https://doi.org/10.26124/mitofit:2022-0017

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Resolving the Rotenone Paradox: elucidating the complexity of multi-substrate respirometry protocols

Chicco Adam J, Zilhaver Philip T, Whitcomb Luke A, Fresa Kyle J, Izon Cheyanne S, Gonzalez-Franquesa Alba, Dometita Crystal, Irving Brian A, Garcia-Roves Pablo Miguel (2022-05-02) MitoFit Prep

Abstract: Multi-substrate respirometry protocols are frequently used to resolve the relative contributions of NADH-producing (N-pathway or CI-linked) substrates and succinate (S-pathway or CII-linked substrate) to mitochondrial oxygen consumption rate (J_O₂). Similarly, rotenone (a selective CI inhibitor) is utilized in the presence of N+S substrates to deduce the contribution of N-pathway flux to the total (NS-pathway) J_O₂. However, under S- and some NS-pathway states, rotenone elicits a paradoxical increase in J_O₂, revealing a complex interaction of N- and S-pathway substrate oxidation on J_O₂ in vitro. Herein, we demonstrate inhibitory effects of >1 mM malate or malonate (a CII inhibitor) on J_O₂ supported by pyruvate and/or glutamate, suggesting that endogenous succinate oxidation interacts with malate concentration to potently regulate J_O₂ supported by N-pathway substrates in a tissue-specific manner. Potential mechanisms are discussed to stimulate further experimentation aimed at elucidating the biological bases for variations in NS-pathway flux in multi-substrate respirometry protocols. • Keywords: Mitochondrial respiration, electron transport chain, succinate, glutamate, oxidative phosphorylation, high-resolution respirometry • Bioblast editor: Tindle-Solomon L

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Bioblast 2022

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 |title=Chicco AJ, Zilhaver PT, Whitcomb LA, Fresa KJ, Izon CS, Gonzalez-Franquesa A, Izon CS, Dometita C, Irving BA, Garcia-Roves PM (2022) Resolving the Rotenone Paradox: elucidating the complexity of multi-substrate respirometry protocols. MitoFit Preprints 2022.17. https://doi.org/10.26124/mitofit:2022-0017
 |info=[[File:MitoFit Preprints pdf.png|left|160px|link=https://wiki.oroboros.at/images/3/33/Chicco_2022_MitoFit.pdf|MitoFit pdf]] [https://wiki.oroboros.at/images/3/33/Chicco_2022_MitoFit.pdf Resolving the Rotenone Paradox: elucidating the complexity of multi-substrate respirometry protocols]<br/>
 |authors=Chicco Adam J, Zilhaver Philip T, Whitcomb Luke A, Fresa Kyle J, Izon Cheyanne S, Gonzalez-Franquesa Alba, Dometita Crystal, Irving Brian A, Garcia-Roves Pablo Miguel
 |year=2022-05-02
 |journal=MitoFit Prep
-|abstract=            Multi-substrate respirometry protocols are frequently used to resolve the relative contributions of NADH-producing (N-pathway or CI-linked) substrates and succinate (S-pathway or CII-linked substrate) to mitochondrial oxygen consumption rate (''J<sub>O2</sub>''). Similarly, rotenone (a selective CI inhibitor) is utilized in the presence of N+S substrates to deduce the contribution of N-pathway flux to the total (N+S-pathway) ''J<sub>O2</sub>''.  However, under S- and some N+S pathway states, rotenone elicits a paradoxical increase in ''J<sub>O2</sub>'', revealing a complex interaction of N- and S-pathway substrate oxidation on ''J<sub>O2</sub>'' ''in vitro''.  Herein, we demonstrate inhibitory effects of >1 mM malate or malonate (a CII inhibitor) on JO2 supported by pyruvate and/or glutamate, suggesting that endogenous succinate oxidation interacts with malate concentration to potently regulate ''J<sub>O2</sub>'' supported by N-pathway substrates in a tissue-specific manner.  Potential mechanisms are discussed to stimulate further experimentation aimed at elucidating the biological bases for variations in NS-pathway flux in multi-substrate respirometry protocols. <br><br>
+|abstract=Multi-substrate respirometry protocols are frequently used to resolve the relative contributions of NADH-producing (N-pathway or CI-linked) substrates and succinate (S-pathway or CII-linked substrate) to mitochondrial oxygen consumption rate (''J''<sub>O<sub>2</sub></sub>). Similarly, rotenone (a selective CI inhibitor) is utilized in the presence of N+S substrates to deduce the contribution of N-pathway flux to the total (NS-pathway) ''J''<sub>O<sub>2</sub></sub>.  However, under S- and some NS-pathway states, rotenone elicits a paradoxical increase in ''J''<sub>O<sub>2</sub></sub>, revealing a complex interaction of N- and S-pathway substrate oxidation on ''J''<sub>O<sub>2</sub></sub> ''in vitro''. Herein, we demonstrate inhibitory effects of >1 mM malate or malonate (a CII inhibitor) on ''J''<sub>O<sub>2</sub></sub> supported by pyruvate and/or glutamate, suggesting that endogenous succinate oxidation interacts with malate concentration to potently regulate ''J''<sub>O<sub>2</sub></sub> supported by N-pathway substrates in a tissue-specific manner. Potential mechanisms are discussed to stimulate further experimentation aimed at elucidating the biological bases for variations in NS-pathway flux in multi-substrate respirometry protocols.
 |keywords=Mitochondrial respiration, electron transport chain, succinate, glutamate, oxidative phosphorylation, high-resolution respirometry
 |editor=Tindle-Solomon L
 }}
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