Cell death in isolated Xenopus laevis (African clawed frog) oocytes is a temperature-dependent process.

Abstract

After isolation of Xenopus laevis oocytes from adult frogs, the cells deteriorate over a few days in vitro under incubation conditions proposed by most protocols. As we have shown in our previous study, changes in storage conditions significantly increase viability, mainly due to cooling to 5-8 °C. Aim of this study was to elucidate the influence of apoptosis on deterioration during aging. Batches of defolliculated stage V-VI oocytes were stored in salt solutions (MBS-based) with or without apoptosis and maturation inhibitors (metamizole, theophylline, ibuprofen, fenbufen) and an additional energy source (glucose-6-phosphate) either cooled (5 or 8 °C) or at 18 °C. Throughout the storage period of up to 4 weeks, oocytes from each condition were injected at different days post-isolation with RNA for expressing GABA receptors (Drosophila RDL) or potassium channels (human KCNQ1). After 2‑3 days expression at 18°C, oocytes were optically checked for deterioration and current responses recorded by the two-electrode voltage clamp technique. No significant influence was found for solutions containing inhibitors, survival times being 16.1 ± 1.2 days on average. Viability differed between batches in cooled oocytes, from 7-25 days of incubation, with a declining number of viable oocytes during a run. Caspase-3 activity was increased significantly in uncooled oocytes over a few days, but the active protein remains to be detected via Western blotting. Our results indicate a temperature-dependent mechanism involved in aging-related deterioration that may differ from apoptosis.