The present study examined the mechanism of A23187-induced activation in pi
g oocytes, with special reference to the effects of extracellular calcium o
n oocyte activation. The following endpoints were evaluated: intracellular
free calcium concentration ([Ca2+](i)), intracellular pH ([pH](i)), cortica
l granule (CG) exocytosis, pronuclear formation, and blastocyst development
. In experiment one, when oocytes were exposed to 50 mu M A23187 for 5 min
in a medium with, or without, calcium, a significant (P < 0.004) increase i
n the [Ca2+](i) was observed in medium with calcium but not in medium witho
ut calcium. An increased [pH](i) (0.08 unit in medium with calcium and 0.13
unit in medium without calcium), cortical granule exocytosis and pronuclea
r formation were observed in oocytes treated with A23187 irrespective of th
e presence or absence of calcium in the medium. In experiment two, the effe
cts of treatment time (0, 0.5, i, 2, and 5 min) on nuclear activation of oo
cytes with A23187 were further examined in medium with, or without, calcium
. It was found that a 2 min treatment activated more (71-74%) oocytes than
the other treatments. Treatment for 5 min in medium without calcium resulte
d in chromatin condensation in some oocytes. Microtubules were not found in
these oocytes. In experiment three, developmental ability was examined of
the oocytes treated with A23187 in medium with, or without, calcium. In vit
ro fertilized oocytes were used as a positive control. It was found that 16
%, 6% and 38% of the oocytes treated with A23187 in medium with calcium, in
medium without calcium, and in vitro fertilized oocytes developed to blast
ocysts after culture for 7 days, respectively. These results indicate that
A23187 can induce pig oocyte activation in calcium-free medium without a ty
pical increase in the [Ca2+](i) and that A23187-induced pig oocyte activati
on is accompanied by an increase in [pH](i). Oocytes activated with A23187
can develop to blastocysts regardless of activation in medium with, or with
out, calcium. (C) 1999 Wiley-Liss, Inc.