Transient exposure of rhesus macaque oocytes to calyculin-a and okadaic acid stimulates germinal vesicle breakdown permitting subsequent development and fertilization

Abstract

Exposure of mammalian oocytes to the protein phosphatase (PP)-1 (PP1) and PP2A inhibitor okadaic acid (OA) stimulates oocyte meiosis. However, treated oocytes do not develop beyond metaphase I (MI), and they display morphological aberrations. Experiments were conducted to define inhibitor treatment conditions for macaque oocytes that would result in germinal vesicle breakdown (GVB) stimulation and completion of meiosis without significant cytoplasmic abnormalities. As described above for OA, continual exposure of macaque oocytes to 50 nM calyculin-a (CL-A) significantly enhanced GVB at 24 h compared to that in controls, and the majority of the treated oocytes displayed cytoplasmic abnormalities. However, transient exposure (10 min) of rhesus macaque oocytes to either 50 nM CL-A or 1.0 μ OA enhanced GVB rates compared to that in controls and did not increase the incidence of cytoplasmic abnormalities. Meiotic maturation from germinal vesicle-intact oocytes to MII was enhanced following transient treatment with CL-A or OA compared to that in controls; however, development from MI to MII occurred at a similar frequency. In vitro-matured oocytes transiently exposed to OA and CL-A were capable of fertilization. In addition, ovarian immunohistochemical analysis revealed that both PP1 and PP2A were present in macaque oocytes. PP1 was localized throughout the cytoplasm with a predominance in the nucleus, whereas PP2A was evenly distributed throughout the cytoplasm with a reduction in the nuclear area. These results taken together - differential developmental responses to inhibitor treatment and intracellular enzyme localizations - may be indicative of multiple regulatory role. of PP1 and/or PP2A during meiosis.