Time-lapse confocal imaging-induced calcium ion discharge from the cumulus–oocyte complex at the time of cattle oocyte activation
Hanna J. McLennan
A B C E , Melanie L. Sutton-McDowall A B C , Sabrina Heng A C D , Andrew D. Abell A C D and Jeremy G. Thompson A B C
+ Author Affiliations
- Author Affiliations
A Australian Research Council Centre of Excellence for Nanoscale BioPhotonics, Australia.
B Robinson Research Institute, Adelaide Medical School, University of Adelaide, SA 5005, Australia.
C Institute for Photonics and Advanced Sensing, School of Physical Sciences, University of Adelaide, SA 5005, Australia.
D Department of Chemistry, School of Physical Sciences, University of Adelaide, SA 5005, Australia.
E Corresponding author. Email: hanna.mclennan@adelaide.edu.au
Reproduction, Fertility and Development 32(14) 1223-1238 https://doi.org/10.1071/RD20143
Submitted: 2 June 2020 Accepted: 15 September 2020 Published: 8 October 2020
Abstract
Oocyte activation, the dynamic transformation of an oocyte into an embryo, is largely driven by Ca2+ oscillations that vary in duration and amplitude across species. Previous studies have analysed intraoocyte Ca2+ oscillations in the absence of the oocyte’s supporting cumulus cells. Therefore, it is unknown whether cumulus cells also produce an ionic signal that reflects fertilisation success. Time-lapse confocal microscopy and image analysis on abattoir-derived cattle cumulus–oocyte complexes coincubated with spermatozoa revealed a distinct discharge of fluorescence from the cumulus vestment. This study demonstrated that this Ca2+ fluorescence discharge was an artefact induced by the imaging procedure independently of oocyte activation success. The fluorescence discharge was a direct result of cumulus cell membrane integrity loss, and future studies should consider the long-term effect of fluorescent labels on cells in time-lapse imaging. However, this study also demonstrated that the distinctive pattern of a coordinated fluorescence discharge was associated with both the presence of spermatozoa and subsequent embryo development to the morula stage, which was affected by Ca2+ chelation and a reduction in the active efflux of the fluorophore. This indicates that the cumulus vestment may have a relationship with oocyte activation at and beyond fertilisation that requires further investigation.
Keywords: calcium oscillations, confocal imaging, embryo development, fertilisation, fluorescent probes, oocyte activation.
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