Parthenogenetic Activation of Pig Oocytes with Calcium lonophore and the Block to Sperm Penetration after Activation'

Calcium ionophore A23187 can parthenogenetically activate oocytes in many animals. The present study was designed to an-alyze functionally the mechanism of A23187 activation of pig oocytes matured in vitro. In experiment 1, effects of the con-centration of A23187 on intracellular calcium transients, corti-cal granule (CG) exocytosis, nuclear activation, and zona reac-tion, which was determined by zona hardening and sperm pen-etrability, were examined. Cumulus-free oocytes were exposed to 0-100 FiM A23187 for 5 min. It was found that the amplitude of the intracellular calcium transients, percentage of CG exo-cytosis, and percentage of pronuclear formation were increased in a concentration-dependent manner. The time for dissolution of zona pellucida (ZP) was increased in the oocytes treated with 25-100 ILM A23187. Penetration of ZP-intact oocytes by sper-matozoa was decreased and only 3-4 % of oocytes were pene-trated by spermatozoa after 50-100 PiM A23187 treatment. In experiment 2, oocytes were treated with 100 PiM A23187 for 5 min and then cultured for 10 min or 3.5 h before insemination. No difference in penetration rates was observed between the two groups of oocytes (12.0 % vs. 12.2%), but the penetration rates were significantly lower than those in controls (85.2 % vs. 82.4%). In experiment 3, treatment of oocytes with 100 FiM A23187 for 5 min was followed by removal of the ZP from a portion of the oocytes. ZP-intact and ZP-free oocytes were then inseminated for examination of sperm penetration. One of 65 (2%) oocytes with ZP and 48 of 52 (92%) oocytes without ZP were penetrated by spermatozoa. These results indicate that ac-tivation of pig oocytes by A23187 is the result of A23187-in-duced intracellular calcium increase and that A23187-induced cortical reaction can prevent sperm penetration of ZP-intact oo-cytes, but not ZP-free oocytes