Dynamic label-free imaging of lipid droplets and their link to fatty acid and pyruvate oxidation in mouse eggs

Abstract

Mammalian eggs generate most of their ATP by mitochondrial oxidation of pyruvate from the surrounding medium or from fatty acids that are stored as triacylglycerols within lipid droplets. The balance between pyruvate and fatty acid oxidation in generating ATP is not established. We have combined coherent anti-Stokes Raman scattering (CARS) imaging with deuterium labelling of oleic acid to monitor turnover of fatty acids within lipid droplets of living mouse eggs. We found that loss of labelled oleic acid is promoted by pyruvate removal but minimised when β-oxidation is inhibited. Pyruvate removal also causes a significant dispersion of lipid droplets, while inhibition of β-oxidation causes droplet clustering. Live imaging of luciferase or FAD autofluorescence from mitochondria, suggest that inhibition of β-oxidation in mouse eggs only leads to a transient decrease in ATP because there is compensatory uptake of pyruvate into mitochondria. Inhibition of pyruvate uptake followed by β-oxidation caused a similar and successive decline in ATP. Our data suggest that β-oxidation and pyruvate oxidation contribute almost equally to resting ATP production in resting mouse eggs and that reorganisation of lipid droplets occurs in response to metabolic demand.