RAT OOCYTE MATURATION - EFFECTS OF PROTEIN-SYNTHESIS INHIBITORS

Puromycin evidently blocks the meiosis of cultured mouse oocytes before extrusion of the 1st polar body, but allows meiosis resumption as revealed by germinal vesicle breakdown (GVB). GVB evidently occurs independently of protein synthesis. Rat oocytes were treated with puromycin during 2 successive culture periods. During the 1st period meiosis resumption was prevented either by the concomitant presence of dibutyryl-cyclic AMP (dbcAMP) or by oocyte incubation within their intact preovulatory follicles. Isolated oocytes, with or without cumulus cells, were then transferred to plain medium or medium with puromycin alone for a 2nd culture period. Cumulus-surrounded oocytes matured spontaneously in culture. DbcAMP prevented GVB, if added within 1 h from the start of culture, whereas puromycin did not prevent GVB but blocked polar body formation, confirming earlier observations. When oocytes were first cultured for 0.5-4 h in the presence of both puromycin and dbcAMP and then cultured for 4 h with puromycin alone, GVB was prevented in a time-dependent way: the longer the 1st culture period the lower the rate of GVB. GVB inhibition was fully reversed when no drugs were present during the 2nd culture. The block of GVB produced by puromycin was sustained even when the 2nd culture was continued for 18 h. Cycloheximide, but not puromycin aminonucleoside, produced the same effects as puromycin. Similar results were obtained when denuded oocytes were used, suggesting that the effects of puromycin were directly on the oocyte and not mediated via the cumulus cells. When follicle-enclosed oocytes were incubated for 3 h with puromycin and the oocytes then isolated and cultured with puromycin, GVB was also inhibited. The rise in O2 consumption seen in maturing oocytes could be prevented in oocytes maturing in medium containing puromycin. This effect of puromycin was reversible, indicating that puromycin did not inflict permanent oocyte damage. Resumption of meiosis is evidently dependent on some proteins with high turnover rates existing in the oocyte before the start of meiosis. Polar body formation might be explained in a similar way.