Retention of Structure and Function of the Cat Germinal Vesicle after Air-Drying and Storage at Suprazero Temperature

Abstract

The study explored a novel approach for preserving the maternal genome without the entire oocyte by air-drying the cat germinal vesicle (GV) in the presence of the disaccharide trehalose. Specifically, we examined GV structure and function after desiccation, storage at 4 °C (up to 32 wk), and rehydration including the ability to resume meiosis after injection into a fresh, conspecific cytoplast. In experiment 1, DNA integrity was similar to fresh controls after 1 and 4 wk storage in the presence of trehalose, but was more fragmented at later time points (especially after 32 wk). Nuclear envelope integrity was sustained in >90% of oocytes stored for 0, 4, or 16 wk regardless of protective treatment. In experiment 2, compacted, air-dried GVs were stored for 2 or 4 wk, rehydrated, and injected into fresh cytoplasts. After culture for 24 h in vitro, up to 73% of oocytes reconstructed with desiccated GVs preserved in trehalose resumed meiosis compared to 30% of those dried in the absence of the disaccharide. At each storage time point, trehalose presence during air-drying was advantageous for resumption of meiosis, with >20% of oocytes completing nuclear maturation to metaphase II. This demonstrates a potential for preserving the female genome using the GV alone and for multiple weeks after desiccation. Trehalose enhanced the process by retaining the ability of a dried and rehydrated GV to resume communication with the surrounding cytoplasm of the recipient oocyte to permit reaching metaphase II and likely sustain subsequent embryo development.