Parthenogenetic activation of pig eggs by exposure to protein kinase inhibitors

Abstract

The objective of the present study was to assess the effect of low concentrations of protein kinase inhibitors on activation. Pig eggs were electrostimulated or cultured with the following: 10 microM 1-[5-isoquinolinylsulfonyl]-2-methylpiperazine, HCl H7 for 24 h; 100 microM H7 for 24 h; 10 nM staurosporine for 24 h; or with 20 microM staurosporine for 20 min followed by Whitten's medium for 24 h. Rates of pronuclear formation in eggs (n = 1240) subjected to these treatments were: untreated, 6.2%; electrostimulated, 77.1%; 10 microM H7, 10.0%; 100 microM H7, 65%; 10 nM staurosporine, 24.2%; and 20 microM staurosporine, 67.3% (significance at P < or = 0.05: 10 microM H7 vs untreated, not significant; 20 microM staurosporine vs 100 microM H7, not significant). Percentages of eggs (n = 125) expressing a 22-kDa band after treatment were: untreated, 37.5%; electrostimulated, 100%; 10 microM H7, 72%; 100 microM H7, 66.7%; 10 nM staurosporine, 40.0%; and 20 microM staurosporine, 77.3% (significance at P < or = 0.10: 100 microM H7, 10 nM staurosporine and 20 microM staurosporine vs 10 microM H7, not significant; 100 microM H7 and 10 nM staurosporine vs untreated, not significant). Transmission electron microscopy of ultrathin sections of treated eggs revealed that cortical granules were present in over half the untreated eggs, as well as over half of the eggs treated with 100 microM H7 or 10 nM staurosporine; in contrast, all cortical granules were absent from electrically-activated eggs. The results indicate that long-term exposure of eggs to low concentrations of broad-spectrum protein kinase inhibitors induces some of the events commonly associated with fertilization.