The role of
calcium in cortical granule exocytosis and activation of the cell cycle at fertilization was examined in the mouse egg using the
calcium chelator BAPTA (1,2-bis(o-aminophenoxy)ethane-N,N,N',N'-tetraacetic acid) and the fluorescent
calcium indicator fluo-3.
BAPTA and
fluo-3 were introduced into
zona-free mouse eggs by a 30-min incubation with 0.01-50 microM
BAPTA acetoxymethyl
ester (AM) and/or 1-20 microM
fluo-3 AM prior to in vitro fertilization. Incubation of eggs in greater than or equal to 5.0 microM
BAPTA AM inhibited cortical granule exocytosis in all cases. Introduction of the
calcium chelator into the egg blocked second polar body formation at greater than or equal to 1.0 microM
BAPTA AM. Sperm entry occurred in all eggs regardless of the
BAPTA AM concentration. Sperm induce a large transient increase in
calcium lasting 2.3 +/- 0.6 min, followed by repetitive transients lasting 0.5 +/- 0.1 min and occurring at 3.4 +/- 1.4-min intervals. Incubation with greater than or equal to 5.0 microM
BAPTA AM inhibited all
calcium transients. Introduction of
BAPTA also inhibited
calcium transients, exocytosis, and the resumption of meiosis following application of the
calcium ionophore A23187 or SrCl2, which activate eggs. These results demonstrate that the
calcium increase at fertilization is required for cortical granule exocytosis and resumption of the cell cycle in a mammalian egg.