Roles of gap junctional communication of cumulus cells in cytoplasmic maturation of porcine oocytes cultured in vitro.
Cumulus cells of the oocyte play important roles in in vitro maturation and subsequent development. One of the routes by which the factors are transmitted from cumulus cells to the oocyte is gap junctional communication (GJC). The function of cumulus cells in in vitro maturation of porcine oocytes was investigated by using a gap junction inhibitor, heptanol. Cumulus-oocyte complexes (COCs) were collected from the ovaries of slaughtered gilts by aspiration. After selection of COCs with intact cumulus cell layers and uniform cytoplasm, they were cultured in a medium with 0, 1, 5, or 10 mM of heptanol for 48 h. After culture in vitro, one group of oocytes was assessed for nuclear maturation and glutathione (GSH) content, and another group was assigned to in vitro fertilization and assessed for the penetrability of oocytes and the degree of progression to male pronuclei (MPN) of penetrated spermatozoa. At the end of in vitro maturation, the oocytes reached metaphase II at a high rate (about 80%) regardless of the presence of heptanol at various concentrations. Cumulus cell expansion and the morphology of oocytes cultured in the medium with heptanol were similar to those of control COCs matured without heptanol. The amount of GSH in cultured oocytes tended to decrease as the concentration of heptanol in the medium was increased. Although there was no difference in the rates of penetrated oocytes cultured in media with different concentrations of heptanol, the proportion of oocytes forming MPN after insemination decreased significantly (P < 0.01) at all concentrations tested. A higher rate of sperm (P < 0.01) failed to degrade their nuclear envelopes after penetration into the oocytes that were treated with heptanol. GJC between the oocyte and cumulus cells might play an important role in regulating the cytoplasmic factor(s) responsible for the removal of sperm nuclear envelopes as well as GSH inflow from cumulus cells.
Mori
T
T
Animal Breeding and Reproduction, Division of Bioresources and Product Science, Graduate School of Agriculture, Hokkaido University, Sapporo 060-8589, Japan. tamori@anim.agr.hokudai.ac.jp
Amano
T
T
Shimizu
H
H
D000818:Animals; D002450:Cell Communication / Q000502:physiology*; D002467:Cell Nucleus / Q000502:physiology; D002478:Cells, Cultured; D003470:Culture Media; D003593:Cytoplasm / Q000502:physiology; D005260:Female; D005307:Fertilization in Vitro; D017629:Gap Junctions / Q000502:physiology*; D005978:Glutathione / Q000378:metabolism; D019850:Heptanol / Q000494:pharmacology; D008297:Male; D009865:Oocytes / Q000166:cytology / Q000502:physiology*; D013084:Sperm-Ovum Interactions / Q000502:physiology; D013552:Swine
D003470:Culture Media; D019850:Heptanol; D005978:Glutathione