|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. https://doi.org/10.26124/mitofit:2022-0017|
» MitoFit Preprints 2022.17.
Abstract: Chicco 2022 Abstract Bioblast: 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 (JO2). 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) JO2. However, under S- and some NS-pathway states, rotenone elicits a paradoxical increase in JO2, revealing a complex interaction of N- and S-pathway substrate oxidation on JO2 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 JO2 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 • O2k-Network Lab: US CO Fort Collins Chicco AJ, US LA Baton Rouge Irving BA, ES Barcelona Garcia-Roves PM
Labels: MiParea: Respiration
Preparation: Isolated mitochondria
Pathway: N, S, NS HRR: Oxygraph-2k