Spatiotemporal Dynamics of the [Ca2+]iRise Induced by Microinjection of Sperm Extract into Mouse Eggs: Preferential Induction of a Ca2+Wave from the Cortex Mediated by the Inositol 1,4,5-Trisphosphate Receptor

Abstract

Hamster sperm extract (SE) possessing Ca2+ oscillation-inducing activity was microinjected into the peripheral or central region of mouse eggs, and the first increase in intracellular Ca2+ concentration ([Ca2+]i), together with the spread of fluorescence-labeled SE in the ooplasm, was investigated by imaging with confocal microscopy. Injection into the periphery always induced a Ca2+ wave that started from the injection site after a delay of 5 to 30 s depending on the concentration of SE. The diluted SE caused a wave of two-step [Ca2+]i rises, which was always observed at fertilization. Injection into the center could induce a radial Ca2+ wave with relatively high dose of SE, but lower dose of SE caused a [Ca2+]i rise after a longer delay which was initiated synchronously over the ooplasm or was preceded in a peripheral area. Injection of diluted SE remarkably prolonged the delay time and reduced the rate of [Ca2+]i rise. The critical concentration of SE needed to induce [Ca2+]i rise was significantly lower in the periphery. These results indicate that the sensitivity to SE is higher in the cortex. SE-induced [Ca2+]i rises were blocked by an antibody against the type 1 inositol 1,4,5-trisphosphate receptor (InsP3R). The cortex was substantially more sensitive to injected InsP3 induction of Ca2+ release than the center. It is suggested that the cortex of mouse eggs may involve a functionally specialized organization of InsP3Rs and Ca2+ pools in which a cytosolic sperm factor(s) could act upon sperm-egg fusion to cause Ca2+ release, leading to the Ca2+ wave at fertilization.

The Role of EF-hand Domains and C2 Domain in Regulation of Enzymatic Activity of Phospholipase Cζ

“…Preparation of Eggs and [Ca 2ϩ ] i Measurement-Mature mouse eggs were obtained from the oviducts of B6D2F1 female mice superovulated by intraperitoneal injection of gonadotropins (8). They were loaded with the Ca 2ϩ -sensitive dye fura-2 (5 M; dextran, Molecular Probes Inc.) in M2 medium for 8 min at 37°C and then transferred to a plastic dish mounted on an inverted fluorescence microscope and heated at 30 -32°C.…”

“…Injection of recombinant PLC into mouse eggs induces Ca 2ϩ oscillations as well (7). The Ca 2ϩ oscillation-inducing activity of sperm extract (4,8) is lost when the extract is pretreated with an antibody against PLC (1). Thus, PLC is a strong candidate for the sperm-derived Ca 2ϩ oscillation-inducing protein.…”

Mammalian Fertilization: From Sperm Factor to Phospholipase Cζ

“…Interestingly, the ability of mammalian sperm extracts to trigger [Ca 2+ ] i oscillations extends to eggs across species (Figs. 1C and D) and even across phyla (Wilding et al, 1997;Wu et al, 1997;Oda et al, 1999;Parrington, et al, 1999;Dong et al, 2000;Jones et al, 2000;Stricker et al, 2000;Li et al, 2001;Coward et al, 2003). Moreover, these extracts are also able to trigger [Ca 2+ ] i oscillations in somatic cells (Berrie et al, 1996).…”

Calcium wave pacemakers in eggs

“…The appearance of cortical ER-rich domains during maturation correlates with an increase in Journal of Cell Science 115 (18) sensitivity to Ins(1,4,5)P3 and to sperm-induced Ca 2+ release (Chiba et al, 1990;Shiraishi et al, 1995;Mehlmann and Kline, 1994;Terasaki et al, 2001) (reviewed in Sardet et al, 2002). Local injections of Ins(1,4,5)P3 and sperm extracts in mouse eggs have revealed that the egg cortex is a region of higher sensitivity to Ins(1,4,5)P3 and to sperm extracts (Oda et al, 1999). Indeed, although the abundance of ER in the egg cortex renders this region more sensitive to Ins(1,4,5)P3, it is also exposed to the highest concentrations of Ins(1,4,5)P3 as it is closest to the source of PtdIns(4,5)P2 in the plasma membrane (Halet et al, 2002;Sardet et al, 2002).…”