Activation by Stimuli of Sperm Penetration in Bovine Oocytes Treated with Higher Concentration of BAPTA/AM.

Abstract

We attempted to determine whether [Ca2+]i elevation in penetrated bovine oocytes is essential for the activation events, including inactivation of MPF and MAP kinase, pronucleus formation, and second polar body extrusion. After maturing for 20 h, oocytes were pretreated with various concentrations of BAPTA/AM for 1 h, coincubated with sperm for 7 h and cultured for 17 h. Percentages of polyspermy increased with BAPTA/AM concentration, and nearly all of the penetrated oocytes that had been pretreated with more than 25 μM BAPTA/AM were penetrated by two or more spermatozoa. In oocytes pretreated with higher concentrations of BAPTA/AM (25 and 50 μM), most of the penetrated oocytes formed one or more pronuclei and an extruded second polar body. In oocytes with or without BAPTA/AM treatments, cdc2 kinase activity decreased to the basal level at 8 h after onset of developmental culture. The MAP kinase activities were significantly reduced at 8 h after onset of developmental culture compared to that before in vitro fertilization. Therefore, we propose that the activation of bovine oocytes at fertilization may be directly induced by as yet unknown sperm-derived activator(s) that is(are) different from the factor inducing oscillation of calcium in spermatozoa.