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Assessing Mitochondrial Function in Sciatic Nerve by High-Resolution Respirometry

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Mitochondrial Sensitivity to Submaximal [ADP] Following Bed Rest: A Novel Two-Phase Approach Associated With Fibre

Lucrezia Zuccarelli¹, Maria De Martino¹, Antonio Filippi¹

¹Department of Medicine, University of Udine, Udine, Italy.

Journal of Cachexia, Sarcopenia and Muscle

|April 26, 2025

Summary

A new mathematical model reveals two distinct phases of mitochondrial respiration linked to muscle fiber type. This finding helps understand oxidative metabolism changes during inactivity and their relation to specific myosin heavy chain isoforms.

Keywords:

ADP bed rest mitochondrial sensitivity myosin heavy chains skeletal muscle mitochondria

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Area of Science:

Exercise Physiology
Mitochondrial Biology
Skeletal Muscle Research

Background:

Inactivity/simulated microgravity impairs oxidative metabolism upstream of mitochondrial function.
Previous studies evaluated maximal ADP-stimulated mitochondrial respiration (JO2max) ex vivo.
This study aimed to assess mitochondrial sensitivity to submaximal ADP concentrations.

Purpose of the Study:

To evaluate mitochondrial sensitivity to submaximal ADP concentrations using an alternative approach.
To identify responses associated with skeletal muscle fibre type composition.
To apply a novel mathematical model to respiration data.

Main Methods:

High-resolution respirometry on isolated permeabilized vastus lateralis fibres from young males before and after 10-day bed rest.
Submaximal ADP titrations (12.5–10,000 μM) to assess complex I+II-linked ADP sensitivity.
Analysis using traditional Michaelis-Menten kinetics and two sequential hyperbolic equations; myosin heavy chain (MyHC) isoform expression analysis.

Main Results:

A novel two-phase hyperbolic model provided an alternative fit to JO2 versus [ADP] data.
Phase 1: Low Vmax1 (28±10 pmol s⁻¹ mg⁻¹) and apparent Km1 (62±54 μM).
Phase 2: Higher Vmax2 (61±16 pmol s⁻¹ mg⁻¹) and apparent Km2 (1784±833 μM); both phases correlated with MyHC isoform percentages.

Conclusions:

A novel mathematical approach using two sequential hyperbolic functions offers an alternative to Michaelis-Menten kinetics for fitting respiration data.
This model identified two distinct ADP response phases in skeletal muscle fibres, linked to fibre type composition.
The low apparent Km/Vmax phase relates to less oxidative (Type 2A+2X) MyHC isoforms, while the high apparent Km/Vmax phase relates to more oxidative (Type 1) MyHC isoforms.