The passage of Ca2+ and fluorescent markers between the sperm and egg after fusion in the mouse.
Mouse sperm-egg fusion was examined using two-photon and confocal microscopy. A delay of several minutes occurred between the first observable event of fusion (which was the diffusion of Ca2+-sensitive dyes from egg into sperm) and any change in egg cytoplasmic Ca2+. When indo-1 dextran was used to obtain ratiometric two-photon images, there was no detectable local increase in egg cytoplasmic Ca2+ near the site of sperm fusion. However, the sperm underwent a Ca2+ transient which appeared to be coincident with the egg cytoplasm Ca2+ transient, which suggested that there was a high permeability pathway for Ca2+ between egg and sperm. To exclude this pathway from providing trigger Ca2+ for the egg transient, we reduced bathing [Ca2+] to approx. 18 microM and 13nM (with EGTA). In these conditions the first egg Ca2+ transient was not prevented, which makes an obligatory role for extracellular Ca2+ in the initiation of the egg Ca2+ transient unlikely. Both FITC-albumin (70 kDa) and 10 kDa dextran-linked Ca2+ indicators were able to diffuse into the sperm from the egg. In addition, phycoerythrin (240 kDa) rapidly diffused into the sperm shortly after fusion (but before any changes in Ca2+ occurred). This suggests that the 'pore(s)' that form during sperm-egg fusion must be at least 8 nm in diameter. These data are compatible with the idea that a diffusible sperm protein could trigger the observed changes in intracellular Ca2+ in the egg, but do not exclude the possibility that other second messengers are generated during sperm-egg fusion.
Jones
K T
KT
Department of Anatomy and Developmental Biology, University College, London WC1E 6BT, UK.
Soeller
C
C
Cannell
M B
MB
D000818:Animals; D002118:Calcium / Q000378:metabolism*; D003911:Dextrans; D004058:Diffusion; D005260:Female; D005306:Fertilization / Q000502:physiology*; D005456:Fluorescent Dyes; D008297:Male; D051379:Mice; D008810:Mice, Inbred C57BL; D008808:Mice, Inbred CBA; D018613:Microscopy, Confocal; D010063:Ovum / Q000166:cytology / Q000502:physiology*; D010799:Phycoerythrin / Q000493:pharmacokinetics; D013084:Sperm-Ovum Interactions / Q000502:physiology*; D013094:Spermatozoa / Q000166:cytology / Q000502:physiology*
D003911:Dextrans; D005456:Fluorescent Dyes; D010799:Phycoerythrin; D002118:Calcium