The Experts below are selected from a list of 255 Experts worldwide ranked by ideXlab platform
Janice P Evans - One of the best experts on this subject based on the ideXlab platform.
-
Preventing Polyspermy in mammalian eggs—Contributions of the membrane block and other mechanisms
Molecular Reproduction and Development, 2020Co-Authors: Janice P EvansAbstract:: The egg's blocks to Polyspermy (fertilization of an egg by more than one sperm) were originally identified in marine and aquatic species with external fertilization, but Polyspermy matters in mammalian reproduction too. Embryonic triploidy is a noteworthy event associated with pregnancy complications and loss. Polyspermy is a major cause of triploidy with up to 80% of triploid conceptuses being the result of dispermic fertilization. The mammalian female reproductive tract regulates the number of sperm that reach the site of fertilization, but mammals also utilize egg-based blocks to Polyspermy. The egg-based blocks occur on the mammalian egg coat (the zona pellucida) and the egg plasma membrane, with apparent variation between different mammalian species regarding the extent to which one or both are used. The zona pellucida block to Polyspermy has some similarities to the slow block in water-dwelling species, but the mammalian membrane block to Polyspermy differs substantially from the fast electrical block that has been characterized in marine and aquatic species. This review discusses what is known about the incidence of Polyspermy in mammals and about the mammalian membrane block to Polyspermy, as well as notes some lesser-characterized potential mechanisms contributing to Polyspermy prevention in mammals.
-
preventing Polyspermy in mammalian eggs contributions of the membrane block and other mechanisms
Molecular Reproduction and Development, 2020Co-Authors: Janice P EvansAbstract:: The egg's blocks to Polyspermy (fertilization of an egg by more than one sperm) were originally identified in marine and aquatic species with external fertilization, but Polyspermy matters in mammalian reproduction too. Embryonic triploidy is a noteworthy event associated with pregnancy complications and loss. Polyspermy is a major cause of triploidy with up to 80% of triploid conceptuses being the result of dispermic fertilization. The mammalian female reproductive tract regulates the number of sperm that reach the site of fertilization, but mammals also utilize egg-based blocks to Polyspermy. The egg-based blocks occur on the mammalian egg coat (the zona pellucida) and the egg plasma membrane, with apparent variation between different mammalian species regarding the extent to which one or both are used. The zona pellucida block to Polyspermy has some similarities to the slow block in water-dwelling species, but the mammalian membrane block to Polyspermy differs substantially from the fast electrical block that has been characterized in marine and aquatic species. This review discusses what is known about the incidence of Polyspermy in mammals and about the mammalian membrane block to Polyspermy, as well as notes some lesser-characterized potential mechanisms contributing to Polyspermy prevention in mammals.
-
camkii can participate in but is not sufficient for the establishment of the membrane block to Polyspermy in mouse eggs
Journal of Cellular Physiology, 2007Co-Authors: Allison J Gardner, Jason G Knott, Keith T Jones, Janice P EvansAbstract:Fertilization triggers initiation of development and establishment of blocks on the egg coat and plasma membrane to prevent fertilization by multiple sperm (Polyspermy). The mechanism(s) by which mammalian eggs establish the membrane block to Polyspermy is largely unknown. Ca2+/calmodulin-dependent protein kinase II (CaMKII) appears to be the key regulator of several egg activation events (completion of meiosis, progression to embryonic interphase, recruitment of maternal mRNAs). Since sperm-induced increases in cytosolic Ca2+ play a role in establishment of the membrane block to Polyspermy in mouse eggs, we hypothesized that CaMKII was a Ca2+-dependent effector leading to this change in egg membrane function. To test this hypothesis, we modulated CaMKII activity in two ways: activating eggs parthenogenetically by introducing constitutively active CaMKIIα (CA-CaMKII) into unfertilized eggs, and inhibiting endogenous CaMKII in fertilized eggs with myristoylated autocamtide 2-related inhibitory peptide (myrAIP). We find that eggs treated with myrAIP establish a less effective membrane block to Polyspermy than do control eggs, but that CA-CaMKII is not sufficient for membrane block establishment, despite the fact that CA-CaMKII-activated eggs undergo other egg activation events. This suggests that: (1) CaMKII activity contributes to the membrane block, but this not faithfully mimicked by CA-CaMKII and furthermore, other pathways, in addition to those activated by Ca2+ and CaMKII, also participate in membrane block establishment; (2) CA-CaMKII has a range of effects as a parthenogenetic trigger of egg activation (high levels of cell cycle resumption, modest levels of cortical granule exocytosis, and no membrane block establishment). J. Cell. Physiol. 212: 275–280, 2007. © 2007 Wiley-Liss, Inc.
-
mammalian membrane block to Polyspermy new insights into how mammalian eggs prevent fertilisation by multiple sperm
Reproduction Fertility and Development, 2006Co-Authors: Allison J Gardner, Janice P EvansAbstract:To inhibit fertilisation by more than one sperm (a condition known as Polyspermy), eggs have developed preventative mechanisms known as blocks to Polyspermy. The block at the level of the egg extracellular coat (the zona pellucida in mammals, the vitelline envelope in non-mammals) has been well characterised in many different animal species and the block at the level of the egg plasma membrane is understood in some non-mammalian species. However, virtually nothing is known about the membrane block to Polyspermy in mammalian eggs, despite data dating back 50–90 years that provide evidence for its existence. In the present review, we will discuss the background on blocks to Polyspermy used by animal eggs and then focus on the membrane block to Polyspermy in mammalian eggs. This will include a summary of classical studies that provide evidence for this block in mammalian eggs, assays used to study the mammalian membrane block and what has been elucidated from recent experimental studies about the cellular signalling events that lead to membrane block establishment and the mechanism of how the membrane block may prevent additional fertilisation.
-
involvement of calcium signaling and the actin cytoskeleton in the membrane block to Polyspermy in mouse eggs
Biology of Reproduction, 2002Co-Authors: Beth A Mcavey, Genevieve B Wortzman, Carmen J Williams, Janice P EvansAbstract:This study examines the effects of actin microfilament-disrupting drugs on events of fertilization, with emphasis on gamete membrane interactions. Mouse eggs, freed of their zonae pellucidae, were treated with drugs that perturb the actin cytoskeleton by different mechanisms (cytochalasin B, cytochalasin D, jasplakinolide, latrunculin B) and then inseminated. Cytochalasin B, jasplakinolide, and latrunculin B treatments resulted in a decrease in the percentage of eggs fertilized and the average number of sperm fused per egg. However, cytochalasin D treatment resulted in an increase in the average number of sperm fused per egg and the percentage of polyspermic eggs. This increase in Polyspermy occurred despite the observation that cytochalasin D treatment caused a decrease in sperm-egg binding and did not affect spontaneous acrosome reactions or sperm motility. This suggested that cytochalasin D-treated eggs had an impaired ability to establish a block to Polyspermy at the level of the plasma membrane. The effect of cytochalasin D on the block to Polyspermy was not due to a general disruption of egg activation because sperm-induced calcium oscillations and cortical granule exocytosis were similar in cytochalasin Dtreated and control eggs. However, buffering of intracellular calcium levels with the calcium chelator BAPTA-AM resulted in an increase in Polyspermy. Together, these data suggest that a postfertilization decrease in egg membrane receptivity to sperm requires functions of the egg actin cytoskeleton that are disrupted by cytochalasin D. Furthermore, egg activation-associated increased intracellular calcium levels are necessary but not sufficient to affect postfertilization membrane dynamics that contribute to a membrane block to Polyspermy. calcium, fertilization, gamete biology, ovum, sperm
Anne E Carlson - One of the best experts on this subject based on the ideXlab platform.
-
inhibiting actin polymerization does not prevent the fast block to Polyspermy in the african clawed frog xenopus laevis
bioRxiv, 2020Co-Authors: Maiwase Tembo, Monica L Sauer, Bennett W Wisner, David O Beleny, Marc A Napolitano, Anne E CarlsonAbstract:Fertilization of an egg by more than one sperm presents one of the earliest and most prevalent obstacles to successful reproduction. As such, eggs employ multiple mechanisms to prevent sperm entry into the nascent zygote. The fast block to Polyspermy is a depolarization of the egg membrane initiated by sperm entry and is employed by diverse external fertilizers including frogs and sea urchins. For some external fertilizers, sperm entry is associated with actin polymerization during the initiation of the fast block. We therefore sought to determine whether the fast block to Polyspermy in the African clawed frog, Xenopus laevis, requires actin polymerization. Although actin polymerization is required for sperm entry into eggs from diverse external fertilizers, including sea urchins and zebrafish, here we demonstrate that actin polymerization is not required for the fast block to Polyspermy in X. laevis.
-
ion channels and signaling pathways used in the fast Polyspermy block
Molecular Reproduction and Development, 2020Co-Authors: Katherine L Wozniak, Anne E CarlsonAbstract:: Fertilization of an egg by multiple sperms, Polyspermy, is lethal to most sexually reproducing species. To combat the entry of additional sperm into already fertilized eggs, organisms have developed various Polyspermy blocks. One such barrier, the fast Polyspermy block, uses a fertilization-activated depolarization of the egg membrane to electrically inhibit supernumerary sperm from entering the egg. The fast block is commonly used by eggs of oviparous animals with external fertilization. In this review, we discuss the history of the fast block discovery, as well as general features shared by all organisms that use this Polyspermy block. Given the diversity of habitats of external fertilizers, the fine details of the fast block-signaling pathways differ drastically between species, including the identity of the depolarizing ions. We highlight the known molecular mediators of these signaling pathways in amphibians and echinoderms, with a fine focus on ion channels that signal these fertilization-evoked depolarizations. We also discuss the investigation for a fast Polyspermy block in mammals and teleost fish, and we outline potential fast block triggers. Since the first electrical recordings made on eggs in the 1950s, the fields of developmental biology and electrophysiology have substantially matured, and yet we are only now beginning to discern the intricate molecular mechanisms regulating the fast block to Polyspermy.
-
plc and ip3 evoked ca2 release initiate the fast block to Polyspermy in xenopus laevis eggs
The Journal of General Physiology, 2018Co-Authors: Katherine L Wozniak, Maiwase Tembo, Wesley A Phelps, Anne E CarlsonAbstract:The prevention of Polyspermy is essential for the successful progression of normal embryonic development in most sexually reproducing species. In external fertilizers, the process of fertilization induces a depolarization of the egg’s membrane within seconds, which inhibits supernumerary sperm from entering an already-fertilized egg. This fast block requires an increase of intracellular Ca2+ in the African clawed frog, Xenopus laevis, which in turn activates an efflux of Cl− that depolarizes the cell. Here we seek to identify the source of this intracellular Ca2+. Using electrophysiology, pharmacology, bioinformatics, and developmental biology, we explore the requirement for both Ca2+ entry into the egg from the extracellular milieu and Ca2+ release from an internal store, to mediate fertilization-induced depolarization. We report that although eggs express Ca2+-permeant ion channels, blockade of these channels does not alter the fast block. In contrast, insemination of eggs in the presence of Xestospongin C—a potent inhibitor of inositol 1,4,5-trisphosphate (IP3)-induced Ca2+ release from the endoplasmic reticulum (ER)—completely inhibits fertilization-evoked depolarization and increases the incidence of Polyspermy. Inhibition of the IP3-generating enzyme phospholipase C (PLC) with U73122 similarly prevents fertilization-induced depolarization and increases Polyspermy. Together, these results demonstrate that fast Polyspermy block after fertilization in X. laevis eggs is mediated by activation of PLC, which increases IP3 and evokes Ca2+ release from the ER. This ER-derived Ca2+ then activates a Cl− channel to induce the fast Polyspermy block. The PLC-induced cascade of events represents one of the earliest known signaling pathways initiated by fertilization.
-
calcium signaling required for the fast Polyspermy block in xenopus laevis
Biophysical Journal, 2016Co-Authors: Katherine L Wozniak, Anne E CarlsonAbstract:Preventing Polyspermy is essential for the normal embryonic development of most animals. Polyspermy, or the fertilization of an egg by more than one sperm, causes severe chromosomal defects and embryonic mortality. The eggs of nearly all externally fertilizing species employ a fast block to Polyspermy in the form of a fertilization signaled depolarization. In the African clawed frog, Xenopus laevis, the fast block requires an increase in intracellular calcium that activates a Ca2+-activated Cl- channel; an efflux of chloride from the egg then depolarizes the membrane. Although increased Ca2+ is crucial for the fast block, the source of this Ca2+ remains unknown. Here we tested the hypothesis that Ca2+ enters the egg to signal the fast block in X. laevis using whole cell recordings on eggs inseminated in the nominal absence of Ca2+or in the presence of 10 μM Gd3+. We recorded normal depolarizations, as well as normal embryonic development, from eggs fertilized in both treatments thereby suggesting that Ca2+ is not entering from the extracellular environment. To test the importance Ca2+ released from the endoplasmic reticulum (ER) in the fast block, the membrane potential was recorded from eggs that had inhibited inositol trisphophate (IP3) synthesis or IP3 receptor binding. Our data show that inseminated eggs under both of these conditions failed to depolarize. Taken together, these data suggest that the primary source of calcium needed for the fast block to Polyspermy is the ER.
Yasuhiro Iwao - One of the best experts on this subject based on the ideXlab platform.
-
the electrical block to Polyspermy induced by an intracellular ca2 increase at fertilization of the clawed frogs xenopus laevis and xenopus tropicalis
Molecular Reproduction and Development, 2019Co-Authors: Mami Watabe, Kenta Izaki, Shohei Fujino, Mei Maruyama, Chiho Kojima, Azusa Hiraiwa, Shuichi Ueno, Yasuhiro IwaoAbstract:: Polyspermy blocking, to ensure monospermic fertilization, is necessary for normal diploid development in most animals. We have demonstrated here that monospermy in the clawed frog, Xenopus tropicalis, as well as in X. laevis, is ensured by a fast, electrical block to Polyspermy on the egg plasma membrane after the entry of the first sperm, which is mediated by the positive-going fertilization potential. An intracellular Ca2+ concentration ([Ca2+ ]i ) at the sperm entry site was propagated as a Ca2+ wave over the whole egg cytoplasm. In the X. tropicalis eggs fertilized in 10% Steinberg's solution, the positive-going fertilization potential of +27 mV was generated by opening of Ca2+ -activated Cl- -channels (CaCCs). The fertilization was completely inhibited when the egg's membrane potential was clamped at +10 mV and 0 mV in X. tropicalis and X. laevis, respectively. In X. tropicalis, a small number of eggs were fertilized at 0 mV. In the eggs whose membrane potential was clamped below -10 mV, a large increase in inward current, the fertilization current, was recorded and allowed Polyspermy to occur. A small initial step-like current (IS current) was observed at the beginning of the increase in the fertilization current. As the IS current was elicited soon after a small increase in [Ca2+ ]i , this is probably mediated by the opening of CaCCs. This study not only characterized the fast and electrical Polyspermy in X. tropicalis, but also explained that the initial phase of [Ca2+ ]i increase causes IS current during the early phase of egg activation of Xenopus fertilization.
-
universality and diversity of a fast electrical block to Polyspermy during fertilization in animals
2018Co-Authors: Yasuhiro Iwao, Kenta IzakiAbstract:In the sexual reproduction of animals, fertilization is indispensable for the initiation of diploid embryonic development. Most animals exhibit monospermy, in which only one sperm enters an egg during normal fertilization. In monospermic species, a fast, electrical block on the egg membrane is one of the most important blocks to Polyspermy. A fertilizing primary sperm usually causes a positive-going fertilization potential to prevent the subsequent entry of excess sperm. An increase in intracellular Ca2+ ([Ca2+]i) in the egg cytoplasm induced by the fertilizing sperm is necessary for egg activation and blocks Polyspermy. The mechanism of voltage-dependent fertilization in monospermic amphibians is presented as a model system of vertebrate fertilization. The electrical Polyspermy blocks in various animals are reviewed and their universality and diversity across the animal kingdom are discussed. Relationships between the fast, electrical block and [Ca2+]i increases in egg cytoplasm are discussed, as well as their changes throughout the course of animal evolution.
-
Egg Activation in Polyspermy: Its Molecular Mechanisms and Evolution in Vertebrates
Sexual Reproduction in Animals and Plants, 2014Co-Authors: Yasuhiro IwaoAbstract:In amphibians, most urodeles (newts) exhibit Polyspermy physiologically, but primitive urodeles (Hynobius) and anurans (frogs) exhibit monospermy. Several fertilizing sperm induce multiple small Ca2+ waves in the polyspermic egg, but a single large Ca2+ wave occurs in the monospermic egg. The Ca2+ waves in newt eggs are caused by a sperm-specific citrate synthase localized outside the mitochondria. The single Ca2+ wave at monospermy is necessary for eliciting a fast block to Polyspermy, whereas the small multiple Ca2+ waves provide slower egg activation to permit the entry of several sperm at Polyspermy. Physiological Polyspermy seems to be evolved in association with the increase in size of eggs in urodeles, reptiles, and birds laying larger yolky eggs. The sperm factor (citrate synthase) operating in slower egg activation in polyspermic eggs is already prepared in the monospermic urodele Hynobius. We have focused on comparative studies in fertilization among amphibians to understand the role of egg activation in establishment of Polyspermy with discussion of the evolution in vertebrates.
-
egg activation in physiological Polyspermy
Reproduction, 2012Co-Authors: Yasuhiro IwaoAbstract:: Fertilization is indispensable not only for restoring diploid genomes but also for the initiation of early embryonic cell cycles in sexual reproduction. While most animals exhibit monospermy, which is ensured by Polyspermy blocks to prevent the entry of extra sperm into the egg at fertilization, several animals exhibit physiological Polyspermy, in which the entry of several sperm is permitted but only one sperm nucleus participates in the formation of a zygote nucleus. Polyspermy requires that the sperm transmit the egg activation signal more slowly, thus allowing the egg to accept several sperm. An increase in intracellular Ca(2+) concentration induced by the fertilizing sperm is both necessary and sufficient for egg activation in Polyspermy. Multiple small Ca(2+) waves induced by several fertilizing sperm result in a long-lasting Ca(2+) rise, which is a characteristic of polyspermic amphibian eggs. We introduced a novel soluble sperm factor for egg activation, sperm-specific citrate synthase, into polyspermic newt eggs to cause Ca(2+) waves. Citrate synthase may perform dual functions: as an enzyme in mitochondria and as a Ca(2+)-inducing factor in egg cytoplasm. We also discuss the close relationship between the mode of fertilization and the Ca(2+) rise at egg activation and consider changes in this process through evolution in vertebrates.
Manuel Aviles - One of the best experts on this subject based on the ideXlab platform.
-
what controls Polyspermy in mammals the oviduct or the oocyte
Biological Reviews, 2009Co-Authors: Manuel AvilesAbstract:: A block to Polyspermy is required for successful fertilisation and embryo survival in mammals. A higher incidence of Polyspermy is observed during in vitro fertilisation (IVF) compared with the in vivo situation in several species. Two groups of mechanisms have traditionally been proposed as contributing to the block to Polyspermy in mammals: oviduct-based mechanisms, avoiding a massive arrival of spermatozoa in the proximity of the oocyte, and egg-based mechanisms, including changes in the membrane and zona pellucida (ZP) in reaction to the fertilising sperm. Additionally, a mechanism has been described recently which involves modifications of the ZP in the oviduct before the oocyte interacts with spermatozoa, termed "pre-fertilisation zona pellucida hardening". This mechanism is mediated by the oviductal-specific glycoprotein (OVGP1) secreted by the oviductal epithelial cells around the time of ovulation, and is reinforced by heparin-like glycosaminoglycans (S-GAGs) present in oviductal fluid. Identification of the molecules contributing to the ZP modifications in the oviduct will improve our knowledge of the mechanisms of sperm-egg interaction and could help to increase the success of IVF systems in domestic animals and humans.
-
oviduct specific glycoprotein and heparin modulate sperm zona pellucida interaction during fertilization and contribute to the control of Polyspermy
Proceedings of the National Academy of Sciences of the United States of America, 2008Co-Authors: Sebastian Canovas, Irene Mondejar, Maria Dolores Saavedra, Raquel Romar, Luis Alberto Grullon, C Matas, Manuel AvilesAbstract:Polyspermy is an important anomaly of fertilization in placental mammals, causing premature death of the embryo. It is especially frequent under in vitro conditions, complicating the successful generation of viable embryos. A block to Polyspermy develops as a result of changes after sperm entry (i.e., cortical granule exocytosis). However, additional factors may play an important role in regulating Polyspermy by acting on gametes before sperm–oocyte interaction. Most studies have used rodents as models, but ungulates may differ in mechanisms preventing Polyspermy. We hypothesize that zona pellucida (ZP) changes during transit of the oocyte along the oviductal ampulla modulate the interaction with spermatozoa, contributing to the regulation of Polyspermy. We report here that periovulatory oviductal fluid (OF) from sows and heifers increases (both, con- and heterospecifically) ZP resistance to digestion with pronase (a parameter commonly used to measure the block to Polyspermy), changing from digestion times of ≈1 min (pig) or 2 min (cattle) to 45 min (pig) or several hours (cattle). Exposure of oocytes to OF increases monospermy after in vitro fertilization in both species, and in pigs, sperm–ZP binding decreases. The resistance of OF-exposed oocytes to pronase was abolished by exposure to heparin-depleted medium; in a medium with heparin it was not altered. Proteomic analysis of the content released in the heparin-depleted medium after removal of OF-exposed oocytes allowed the isolation and identification of oviduct-specific glycoprotein. Thus, an oviduct-specific glycoprotein–heparin protein complex seems to be responsible for ZP changes in the oviduct before fertilization, affecting sperm binding and contributing to the regulation of Polyspermy.
Katherine L Wozniak - One of the best experts on this subject based on the ideXlab platform.
-
ion channels and signaling pathways used in the fast Polyspermy block
Molecular Reproduction and Development, 2020Co-Authors: Katherine L Wozniak, Anne E CarlsonAbstract:: Fertilization of an egg by multiple sperms, Polyspermy, is lethal to most sexually reproducing species. To combat the entry of additional sperm into already fertilized eggs, organisms have developed various Polyspermy blocks. One such barrier, the fast Polyspermy block, uses a fertilization-activated depolarization of the egg membrane to electrically inhibit supernumerary sperm from entering the egg. The fast block is commonly used by eggs of oviparous animals with external fertilization. In this review, we discuss the history of the fast block discovery, as well as general features shared by all organisms that use this Polyspermy block. Given the diversity of habitats of external fertilizers, the fine details of the fast block-signaling pathways differ drastically between species, including the identity of the depolarizing ions. We highlight the known molecular mediators of these signaling pathways in amphibians and echinoderms, with a fine focus on ion channels that signal these fertilization-evoked depolarizations. We also discuss the investigation for a fast Polyspermy block in mammals and teleost fish, and we outline potential fast block triggers. Since the first electrical recordings made on eggs in the 1950s, the fields of developmental biology and electrophysiology have substantially matured, and yet we are only now beginning to discern the intricate molecular mechanisms regulating the fast block to Polyspermy.
-
plc and ip3 evoked ca2 release initiate the fast block to Polyspermy in xenopus laevis eggs
The Journal of General Physiology, 2018Co-Authors: Katherine L Wozniak, Maiwase Tembo, Wesley A Phelps, Anne E CarlsonAbstract:The prevention of Polyspermy is essential for the successful progression of normal embryonic development in most sexually reproducing species. In external fertilizers, the process of fertilization induces a depolarization of the egg’s membrane within seconds, which inhibits supernumerary sperm from entering an already-fertilized egg. This fast block requires an increase of intracellular Ca2+ in the African clawed frog, Xenopus laevis, which in turn activates an efflux of Cl− that depolarizes the cell. Here we seek to identify the source of this intracellular Ca2+. Using electrophysiology, pharmacology, bioinformatics, and developmental biology, we explore the requirement for both Ca2+ entry into the egg from the extracellular milieu and Ca2+ release from an internal store, to mediate fertilization-induced depolarization. We report that although eggs express Ca2+-permeant ion channels, blockade of these channels does not alter the fast block. In contrast, insemination of eggs in the presence of Xestospongin C—a potent inhibitor of inositol 1,4,5-trisphosphate (IP3)-induced Ca2+ release from the endoplasmic reticulum (ER)—completely inhibits fertilization-evoked depolarization and increases the incidence of Polyspermy. Inhibition of the IP3-generating enzyme phospholipase C (PLC) with U73122 similarly prevents fertilization-induced depolarization and increases Polyspermy. Together, these results demonstrate that fast Polyspermy block after fertilization in X. laevis eggs is mediated by activation of PLC, which increases IP3 and evokes Ca2+ release from the ER. This ER-derived Ca2+ then activates a Cl− channel to induce the fast Polyspermy block. The PLC-induced cascade of events represents one of the earliest known signaling pathways initiated by fertilization.
-
calcium signaling required for the fast Polyspermy block in xenopus laevis
Biophysical Journal, 2016Co-Authors: Katherine L Wozniak, Anne E CarlsonAbstract:Preventing Polyspermy is essential for the normal embryonic development of most animals. Polyspermy, or the fertilization of an egg by more than one sperm, causes severe chromosomal defects and embryonic mortality. The eggs of nearly all externally fertilizing species employ a fast block to Polyspermy in the form of a fertilization signaled depolarization. In the African clawed frog, Xenopus laevis, the fast block requires an increase in intracellular calcium that activates a Ca2+-activated Cl- channel; an efflux of chloride from the egg then depolarizes the membrane. Although increased Ca2+ is crucial for the fast block, the source of this Ca2+ remains unknown. Here we tested the hypothesis that Ca2+ enters the egg to signal the fast block in X. laevis using whole cell recordings on eggs inseminated in the nominal absence of Ca2+or in the presence of 10 μM Gd3+. We recorded normal depolarizations, as well as normal embryonic development, from eggs fertilized in both treatments thereby suggesting that Ca2+ is not entering from the extracellular environment. To test the importance Ca2+ released from the endoplasmic reticulum (ER) in the fast block, the membrane potential was recorded from eggs that had inhibited inositol trisphophate (IP3) synthesis or IP3 receptor binding. Our data show that inseminated eggs under both of these conditions failed to depolarize. Taken together, these data suggest that the primary source of calcium needed for the fast block to Polyspermy is the ER.
