Modulation of spermatozoa and zona pellucida properties by the soluble acrosome reaction‐inducing factor of the ovulated egg‐cumulus complex

Abstract

Three sources of hamster periovulatory fluids (+/- heat inactivation at 56 degrees C), with bovine serum albumin (BSA) as control, were tested for effects on penetration of three classes of eggs by hamster sperm precapacitated in BSA. These fluids were a soluble extract of cumulus oophorus fluid (COF) from the ovulated hamster egg-cumulus complex, serum, and follicular fluid. Egg types were ovulated, salt-stored (ovulated), and follicular. In both COF and serum, there were significant differences among egg types in mean penetration, and significant effects of fluid addition. In contrast, there was no effect of follicular fluid and no differences between follicular and stored eggs. For the follicular eggs (combined data, normalized, ranked), patterns of response to the three factors (+/- heating) were different: only unheated COF and heated serum increased penetration significantly above BSA control levels (average rank 20.2, 41.4, 38, for BSA, COF (unheated), serum (heated), respectively). This indicated that the active component in COF was heat labile, not present in either serum or follicular fluid, and, therefore, of oviductal origin. Oviduct and/or COF exposure of eggs and sperm was tested for effects as an acrosome reaction inducing factor (ARIF) for acrosome reactions (AR; zona-bound and free-swimming sperm) and on sperm:zona binding and penetration. The COF ARIF for free-swimming sperm AR was heat stable. Penetration of follicular eggs increased after incubation in COF prior to sperm addition, but a greater response occurred when COF was added to eggs with sperm. In kinetic experiments, 25 min following sperm attachment, follicular eggs had lost 41% of initially bound sperm, vs. 23% for ovulated eggs, and had only 16 AR sperm/egg, vs. 26 for ovulated. Follicular eggs incubated in COF (then washed three times) had the same number of bound AR sperm as ovulated eggs. Acid solubilized zona pellucida (ASZP) from ovulated eggs was more effective as an ARIF per zona than ASZP from follicular eggs. Zonae of follicular eggs, as evidenced by dissolution times in beta-mercaptoethanol (beta-MEOH), were not "harder" than those of ovulated eggs. There were differences in lectin binding antigens on zonae of both fresh and stored, follicular and ovulated, eggs. We conclude that multiple biological factors orchestrate sperm:egg interactions in the ampulla. Our data are consistent with the presence of at least three effective components: 1) the oviductal lectin-binding antigen (ZPO or oviductin), 2) an additional heat-labile component, and 3) the heat-stable ARIF for free-swimming sperm.

Identification of a Sperm Penetration Factor in the Oviduct of the Golden Hamster1

“…There is considerable interest in identifying molecules present at the site of in vivo fertilization that act as cofactors for penetration and fertilization, potentially acting as acrosome reaction-inducing factor(s) (ARIF; [13][14][15][16]). Previously, we reported that the ovulated hamster egg cumulus complex contained a soluble, non-albumin ARIF that stimulated hamster sperm acrosome reactions and enhanced penetration through the zona pellucida [13,16,17]. The egg penetration-enhancing substance for follicular oocytes appeared to be of oviductal origin and functioned partly to increase the ARIF and spermbinding activities of the zona pellucida [17].…”

“…Previously, we reported that the ovulated hamster egg cumulus complex contained a soluble, non-albumin ARIF that stimulated hamster sperm acrosome reactions and enhanced penetration through the zona pellucida [13,16,17]. The egg penetration-enhancing substance for follicular oocytes appeared to be of oviductal origin and functioned partly to increase the ARIF and spermbinding activities of the zona pellucida [17]. The objectives of the current study were to identify the oviductal component that enhances sperm penetration into follicular eggs and to determine whether or not its mechanism of action is to stimulate acrosome reactions in capacitated sperm.…”

Secreted Proteins of the Oviduct

“…Second, the oviduct is seen as a biologically active, dynamic tissue synthesizing and releasing macromolecules into the lumen throughout the estrous or menstrual cycle. Recent studies suggest a variety of oviduct-gamete [Buhi et al, 1993;Boatman et al, 1994] and oviduct-embryo [Gandolfi, 1995] interactions which indicate some regulation of these events. In this review, discussion will focus on the biosynthetic activity of the oviduct, identification and characterization of secretory protein molecules and current literature on the oviductal microenvironment.…”

Epidermal growth factor stimulates hydrolysis of phosphatidylinositol4,5‐bisphosphate, generation of diacylglycerol and exocytosis in mouse spermatozoa

“…The mass of DAG generated after EGF stimulation exceeds that potentially arising via PtdIns (4,5) Fig. 3.…”

“…It is generally believed that zona pellucida (ZP), the acellular oocyte coat, is the inducer of acrosomal exocytosis [1], with one of its glycoproteins (ZP3 in the mouse) being responsible for its exocytosis-inducing activity [2]. However, recent work has suggested that a series of molecules present in the cumulus-oocyte complex or in the oviductal ampulla (the site of fertilization) could also induce exocytosis or may act as co-factors during the initiation or completion of this process [3][4][5].…”