Cytochalasin D treatment induces meiotic resumption in follicular sheep oocytes

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Abstract
Ovine cumulus‐enclosed oocytes collected from antral follicles (3–5 mm in diameter) were cultured in vitro with 2 ± 106 granulosa cell/ml in the presence or absence of gonadotropins or in the presence of cytochalasin D (CD). The maturation rate was assessed after 24 h of culture. In the control group, in the presence of gonadotropins (follicle‐stimulating hormone‐luteinizing hormone (FSH‐LH; 10 μg/ml) 100% of the oocytes reached metaphase II. Whereas intercellular junctions were no longer present after 6–7 h of culture, germinal vesicle breakdown (GVBD) occurred by the same time. In contrast, in the absence of gonadotropin, the majority of the oocytes (59%) remained blocked in GV stage. The inhibition exerted by the granulosa cells on meiotic resumption was overcome when the cumulus‐oocyte complexes (COCs) were incubated in CD (5 μg/ml) for 6 h at the beginning of the culture. Under these conditions, 85% of the oocytes matured with extrusion of the first polar body. Cytological analysis by cytofluorescence (NBD phallacidin) and electron microscopy showed that, after 6 h of treatment, CD provoked a redistribution of the microfilaments, mainly in the cumulus cells and to a lesser extent in the oocyte cortex. Intercellular junctions disappeared concomitantly with a significant decrease of the intercellular transport of tritiated uridine. The initiation of GVBD occurred at the same time. These results indicate that the resumption of meiosis was correlated with a loss of both junctional complexes (intermediate and gap junctions) between the cumulus cells and the oocyte.