Meiotic arrest, resumption, and TZP retraction in bovine COCs undergoing pre-IVM: lessons from a refined GV stage classification.

Abstract

The nuclear, cytoplasmic, and molecular maturation of the mammalian oocyte is a finely orchestrated sequence of events that relies on proper cumulus-oocyte communication. Bovine oocytes enter the in vitro maturation systems at the germinal vesicle stage (GV) exhibiting four different chromatin configurations (GV0-GV3). Herein, we associate the oocyte chromatin and nuclear lamina configurations to propose a refined GV classification (GV0, GV1.1-GV1.3, GV2.1-GV2.3, and GV3.1-GV3.3). This refined GV classification system was correlated with oocyte meiosis resumption and transzonal projections (TZPs) density of COCs submitted to three IVM systems (control IVM; and a modified IVM preceded or not by a pre-IVM step). Pre-IVM resulted in lower polar body extrusion rates at 19 h IVM, albeit ~24% of the oocytes extruded their first polar body at 9 h IVM. Pre-IVM sustained 80% of oocytes meiotically arrested but altered GV distribution, reducing GV2 and increasing GV1.3 and GV3.3 categories. Pre-IVM reduced TZP densities predominantly in pre-matured GV3 and GVBD COCs. At 9 h of IVM, both groups matured in modified IVM showed lower TZP densities compared to immature and IVM control. Gene expression supports the TZP density differences, with ERK2 and PRKACA upregulation in pre-matured cumulus and in modified IVM groups at 9 h of IVM. GDF9 and BMP15 levels were similar between treated and control groups at all time points. Our findings indicate that despite the IVM system, the initial oocyte GV stage influences pre-IVM and IVM outcomes. The refined GV classification system is a useful tool to oocyte biologists.