Sequential forms of ATPase activity correlated with changes in cation binding and membrane potential from meiosis to first clevage in R. pipiens

Changes in cortical membrane potential, cation binding, subcellular aggregation, ATPase activity, phosphoprotein phosphatase activity and inorganic phosphate levels have been measured in the R. pipiens egg from meiotic prophase to first cleavage.

1.

1. Between meiotic prophase (oocyte) and metaphase of the second maturation division (ovulated egg), the egg cortex depolarizes and the egg cytoplasm becomes positive relative to the external environment. Fertilization is accompanied by a transient positive hyperpolarization and the potential returns to the level of the ovulated egg by the completion of meiosis (15 to 20 min). The egg cytoplasm remains positive until first cleavage when the cortex slowly repolarizes and the cytoplasm again becomes negative relative to the external medium.     

A direct effect of some rifamycin derivatives on the morphology of mammalian mitochondria

Protein synthesis has been investigated in cell-free preparations from mature ovarian oocytes, unfertilized and fertilized eggs, and early embryos of Drosophila melanogaster. Preparations from unfertilized eggs have a specific activity that is 5- to 6-fold higher than the activity of fractions from ovarian oocytes. There is an additional small increase in activity of preparations from fertilized eggs. The specific activity that is rapidly attained in the fertilized egg remains essentially constant for 2 to 2.5 h after fertilization, decreases sharply during blastoderm formation, and again increases during gastrulation. The activities of unfertilized eggs decline slightly during the first 2 h after oviposition, and then decrease more sharply. About 35 % of the ribosomes in preparations from both unfertilized and fertilized eggs sediment in the polyribosome region of sucrose density gradients, whereas no polyribosomes could be detected in preparations from ovarian oocytes. In both ovarian oocytes and fertilized eggs, less than 1 % of the ribosome populations were present as subunits. Additional ribonucleoprotein material of buoyant densities different from those of ribosomal subunits or ribosomes was found throughout the sucrose gradients. About 3.5 % of the ribosomes were found to be membrane-bound in preparations from both unfertilized and fertilized eggs.     

Cytokeratin intermediate filament organisation and dynamics in the vegetal cortex of living Xenopus laevis oocytes and eggs

Cytokeratin intermediate filaments are prominent constituents of developing Xenopus oocytes and eggs, forming radial and cortical networks. In order to investigate the dynamics of the cortical cytokeratin network, we expressed EGFP-tagged Xenopus cytokeratin 1(8) in oocytes and eggs. The EGFP-cytokeratin co-assembled with endogenous partner cytokeratin proteins to form fluorescent filaments. Using time-lapse confocal microscopy, cytokeratin filament assembly was monitored in live Xenopus oocytes at different stages of oogenesis, and in the artificially-activated mature egg during the first cell cycle. In stage III to V oocytes, cytokeratin proteins formed a loose cortical geodesic network, which became more tightly bundled in stage VI oocytes. Maturation of oocytes into metaphase II-arrested eggs induced disassembly of the EGFP-cytokeratin network. Imaging live eggs after artificial activation allowed us to observe the reassembly of cytokeratin filaments in the vegetal cortex. The earliest observable structures were loose foci, which then extended into curly filament bundles. The position and orientation of these bundles altered with time, suggesting that forces were acting upon them. During cortical rotation, the cytokeratin network realigned into a parallel array that translocated in a directed manner at 5 microm/minute, relative to stationary cortex. The cytokeratin filaments are, therefore, moving in association with the bulk cytoplasm of the egg, suggesting that they may provide a structural role at the moving interface between cortex and cytoplasm.     

The secretions of the cumulus-oocyte complex in relation to fertilization and early mouse embryonic development: a histochemical study

Summary A study has been made of the histochemical composition of the murine cumulus—oocyte complex and zona pellucida following treatment of immature females with exogenous gonadotrophins. Selected developmental stages were studied in detail, namely (i) the ovulated and unfertilized egg, (ii) the fertilized oocyte and (iii) the preimplantation embryo. In addition, the histochemical features observed in normal fertilized embryos have been compared with those of haploid and diploid parthenogenetic embryos at comparable stages following activation. Shortly after fertilization, glycosaminoglycans, which form a major component of the extracellular matrix surrounding the cumulus cells, become incorporated into the zona pellucida of the fertilized egg. In oocytes with few or no attendant cumulus cells, there appeared to be a diminished uptake of glycosaminoglycans and a reduced intensity of the zona staining reaction to Alcian Blue. In these oocytes, uptake of glycosaminoglycans appeared to be from the secretions lining the oviduct. There was little incorporation of the glycosaminoglycans from the extra-cellular matrix of the surrounding cumulus cells into the zona pellucida in unfertilized or parthenogenetic eggs despite the activation stimulus. After fertilization or activation, the zona pellucida became increasingly PAS-positive. Enzymic studies clearly indicate that the composition of the zona pellucida of the early embryo is histochemically different from the zona that surrounds the oocyte in the preovulatory follicle. These findings are discussed in relation to the decreased viability of embryos from oocytes which have been ovulated.The death of Mrs Carol Grainge is sadly recorded.     

Inhibition of the onset of vitellogenesis by abnormal retention of mature eggs in the stick insect,Carausius morosus

Mature eggs stored in the lateral oviducts are unusual in Carausius morosus. When egg laying is experimentally prevented, ovarian production is inhibited by the accumulation of mature eggs within the lateral oviducts. Such storage affects only the very small terminal oocytes and not the vitellogenic follicles, which attain to term and are ovulated. The fact that unilateral retention of eggs affects only the corresponding ovary strongly suggests that the mature eggs themselves inhibit ovarian production in the stick insect without the involvement of an endocrine organ. It is likely that this mode of action, different from that suggested for the housefly or Rhodnius prolixus, is correlated with the independence of ovarian development from the corpus allatum hormone in the stick insect.     

Effects of cytochalasin B on the cortex of the unfertilized sea urchin egg

The peripheral cytoplasm of the unfertilized sea urchin egg contains approximately 18,000 cortical granules. These granules remain monolayered within the normal boundaries of the cortex when the egg is centrifuged at forces sufficient to stratify other intracellular inclusions. Exposure of unfertilized eggs to the microfilament disrupting agent, cytochalasin B (CB) causes the granules to rearrange into several layers and occasionally to undergo exocytosis or break down in situ. When these eggs are centrifuged, the cortical granules are dislodged from the cortex and migrate centrifugally among the densest intracellular components. In addition, cytoplasmic inclusions, which normally are excluded from the cortex, impinge directly upon the egg plasma membrane in CB-treated, centrifuged eggs. These results are consistent with the existence of a microfilamentous network which confines the cortical granules within and excludes other intracellular inclusions from the cortex of the unfertilized egg.     

Changes in elemental composition of fertilized and unfertilized northern pike (Esox lucius) eggs incubated in buffer with and without dimethyl sulfoxide: An X-ray microanalysis study

Fertilized and unfertilized eggs from the northern pike (Esox lucius) were incubated 2 hr in buffer with 0 and 10% (v/v) dimethyl sulfoxide and then quickly frozen in the wells of aluminum blocks submerged in liquid nitrogen. Control eggs and ovarian fluid were similarly frozen immediately after collection. The frozen eggs were sectioned, freeze dried, mounted on stubs, and carbon coated. X-ray microanalysis was used to determine changes in element levels and dimethyl sulfoxide (Me₂SO) penetration in the zona radiata, cytoplasm, cortical alveoli, and egg yolk. Unfertilized eggs incubated without Me₂SO showed decreased levels of Na, Cl, and K in the zona radiata; fertilized eggs, incubated without Me₂SO showed decreased levels of Na, P, and Cl in the zona radiata and increased levels of K in the cytoplasm; unfertilized eggs, incubated with 10% Me₂SO showed decreased Na and Cl in the zona radiata, decreased K in the cytoplasm and increased K in the cortical alveoli; fertilized eggs incubated in buffer with 10% Me₂SO showed decreased levels of Na, P, Cl, and K (zona radiata), P, Cl, and K (cytoplasm), Na (yolk), and increased Cl in the yolk (all P<.01). Me₂SO (v/v) levels reached 1.5-3.1% in the zona radiata, 0-3.2% in cytoplasm, 2.3-8.7% in cortical alveoli, and 0-1.6% in the yolk. Unfertilized eggs showed more Me₂SO penetration than fertilized eggs.     

Accumulation of fluorescently labeled actin in the cortical layer in sea urchin eggs after fertilization

Actin from sea urchin eggs was fluorescently labeled with fluorescein isothiocyanate (FITC), N-(7-dimethylamino-4-methylcoumarinyl)-maleimide (DACM), or 5-iodoacetamidofluorescein (IAF) and microinjected into sea urchin eggs and oocytes. It distributed evenly in the cytoplasm of unfertilized eggs. Upon fertilization, actin accumulated first around the sperm binding site and, soon afterwards, in the fertilization cone. The accumulation propagated all over the cortex after a latent period of 10-20 sec. In the case of Clypeaster japonicus eggs, propagation of the accumulation coincided with a shape change in the egg, suggesting that the accumulated actin in the cortex generates forces. FITC-actin was incorporated into microvilli and retained in the cortex after cleavage. On the other hand, DACM- or IAF-actin was not incorporated into microvilli and was dispersed from the cortex by cleavage. These differences may be attributable to differences in the properties of the actins labeled at different sites. After photobleaching by laser light irradiation, FITC- or IAF-actin redistributed in the cortex of fertilized egg as quickly as it did before fertilization. When an unfertilized egg was injected with both actin and a calcium buffer (intracellular free Ca2+ concentration 9 microM), the actin accumulation was similar to that during fertilization but without the latent period. This suggests that the accumulation depended on the increase in the intracellular free Ca2+ concentration. When the unfertilized egg was injected with 0.2 M EGTA after injection of labeled actin and then inseminated, it accumulated only in the protrusion of cytoplasm where the sperm had entered, and fertilization was not completed. In immature oocytes, the accumulation was observed in the cortical region, including the huge protrusion of the cytoplasm where the sperm had entered. These results suggest that actin accumulation in the sperm binding site plays an important role in the sperm reception mechanism of the egg.     

Identification of a novel C1q family member in color crucian carp (Carassius auratus) ovary

Potential roles of C1q/tumor necrosis factor (TNF) superfamily proteins have been observed in vertebrate oogenesis and oocyte maturation, but no ovary-specific member has been identified so far. In this study, we have cloned and identified a novel member of C1q family with a C1q domain in the C-terminal from fully grown oocyte cDNA library of color crucian carp and demonstrated that the gene might be specifically expressed in ovary and therefore designated as Carassius auratus ovary-specific C1q-like factor, CaOC1q-like factor. It encodes a 213 amino acid protein with a 17 amino acid signal peptide. There is only one protein band of about 24.5 kDa in the extracts from phase I to phase IV oocytes, but two positive protein bands are detected in the extracts of mature eggs and fertilized eggs. Furthermore, the mobility shift of the smaller target protein band cannot be eliminated by phosphatase treatment, but the larger protein band increases its mobility on the gel after phosphatase treatment, suggesting that the larger protein might be a phosphorylated form. Immunofluorescence localization indicates that the CaOC1q-like proteins localize in cytoplasm, cytoplasm membrane and egg envelope of the oocytes at cortical granule stage and vitellogenesis stage, whereas they were compressed to cytoplasm margin in ovulated mature eggs and discharged into perivitelline space between cytoplasm membrane and egg envelope after egg fertilization. Further studies on distribution and translocation mechanism of the CaOC1q-like factor will be benefit to elucidate the unique function in oogenesis, oocyte maturation and egg fertilization.     

Changes in membrane potential during mouse egg development

The electrical membrane potential (E_m) was measured in the developing mouse egg with intracellular microelectrodes. The oocyte had a low negative E_m of ?8.3 ± 0.8 mV (mean ± SE) when immature, which decreased and reversed polarity to a small positive value (+1.9 ± 0.2 mV) in the mature ovulated oocyte. After fertilization E_m returned to a negative value (?9.2 ± 0.5 mV) similar in magnitude to that observed in immature oocytes and then increased significantly (P < 0.01) at both the two-cell (?10.7 ± 0.3 mV) and morula stage (?12.8 ± 0.7 mV) and leveled out at the blastocyst stage (?12.9 ± 0.7 mV). Average potential difference recorded across the blastocoele wall of not fully expanded blastocysts was ?5.0 ± 0.5 mV. These data represent the first report on membrane potentials of the mammalian egg during development. A striking similarity is seen in the relative changes in E_m throughout development of the mouse egg in comparison to those seen in other invertebrate and vertebrate eggs.     

A novel procedure for obtaining denuded sea urchin eggs and observations on the role of the vitelline layer in sperm reception and egg activation

A procedure is described for the complete removal of the vitelline layer of the eggs of the sea urchin, Strongylocentrotus purpuratus. The method involves treatment of unfertilized eggs with an S. purpuratus cortical granule protease preparation followed by incubation in an alkaline dithiothreitol seawater solution. Eggs denuded of their vitelline layers react metabolically to parthenogenetic agents and sperm like unfertilized eggs, whereas the fertilizability of denuded eggs and receptivity to sperm is much less than controls. The present method is superior to previous methods using mercaptans in that all of the vitelline layer is removed and to procedures using other proteolytic enzymes in that no ¹²⁵I-labelled plasma membrane proteins are extensively modified. Thus the cortical granule protease dithiothreitol procedure is ideal for studies of the plasma membrane of the unfertilized egg and for studies on the role of the vitelline layer in normal fertilization and development.     

Stages of oocyte development in the zebrafish,Brachydanio rerio

Oocyte development has been divided into five stages in the zebrafish Brachydanio rerio, based on morphological criteria and on physiological and biochemical events. In stage I (primary growth stage), oocytes reside in nests with other oocytes (Stage IA) and then within a definitive follicle (Stage IB), where they greatly increase in size. In stage II (cortical alveolus stage), oocytes are distinguished by the appearance of variably sized cortical alveoli and the vitelline envelope becomes prominent. In stage III (vitellogenesis), yolk proteins appear in oocytes and yolk bodies with crystalline yolk accrue during this major growth stage. Ooctes develop the capacity to respond in vitro to the steroid 17α, 20β-dihydroxy-4-pregnen-3-one (DHP) by undergoing oocyte maturation. In stage IV (oocyte maturation), oocytes increase slightly in size, become translucent, and their yolk becomes non-crystalline as they undergo final meiotic maturation in vivo (and in response to DHP in vitro). In stage V (mature egg), eggs (approx. 0.75 mm) are ovulated into the ovarian lumen and are capable of fertilization. This staging series lays the foundation for future studies on the cellular processes occurring during oocyte development in zebrafish and should be useful for experimentation that requires an understanding of stage-specific events. © 1993 Wiley-Liss, Inc.     

Morphological changes in the oocyte and its surrounding vestments during in vivo fertilization in the dasyurid marsupial Sminthopsis crassicaudata

This light and transmission electron microscopical study shows that the first polar body is given off before ovulation and that part of its cell membrane and that of the surrounding oocyte have long microvilli at the time of its ejection. Several layers of cumulus cells initially surround the secondary oocyte and first polar body, but the ovulated oocytes in the oviducts in the process of being fertilized do not have cumulus cells around them. Partly expelled second polar bodies occur in the oviduct; they are elongated structures that lack organelles and have electron-dense nuclei. A small fertilization cone appears to form around the sperm tail at the time of sperm entry into the egg and an incorporation cone develops around the sperm head in the egg cytoplasm. In three fertilized eggs a small hole was seen in the zona, which was presumably formed by the spermatozoon during penetration. Cortical granules, present in ovarian oocytes, are not seen in fertilized tubal or uterine eggs; release of their contents probably reduces the chances of polyspermy, although at least one polyspermic fertilized egg was seen and several other fertilized eggs had spermatozoa within the zona pellucida. In the zygote the pronuclei come to lie close together, but there was no evidence of fusion. A "yolk mass," which becomes eccentric before ovulation, is extruded by the time the two-cell embryos are formed, but many vacuoles remain in the non-yolky pole of the egg. A shell membrane of variable thickness is present around all uterine eggs but its origin remains undetermined.     

Histochemical observations on the cortical granules in the eggs of Indian lizards

Summary The development, distribution and histochemical nature of cortical granules have been investigated in growing eggs of two species of Indian lizards (Hemidactylus flaviviridis Rüppel and Uromastix hardwickii). Numerous cortical granules develop in the peripheral ooplasm of growing oocytes and are finally arranged in the cortical cytoplasm of the egg. They consist of a carbohydrate-protein complex; most of the carbohydrate component is an acid mucopolysaccharide. The cortical granules in the eggs of lizards have been compared and contrasted to those in Amphioxus, fishes, amphibians, and mammals.     

Morphologic comparison of ovulated and in vitro–matured porcine oocytes,with particular reference to polyspermy after in vitro fertilization

This study was conducted to evaluate morphologic differences in pig oocytes matured in vivo and in vitro, with particular reference to the potential relationship between oocyte morphology and the occurrence of polyspermy after in vitro fertilization (IVF). In vivo–matured oocytes were surgically recovered from the oviducts of gilts with ovulated follicles on day 2 of estrus, and in vitro–matured oocytes were obtained by culturing follicular oocytes in a oocyte maturation system that has resulted previously in production of live offspring following IVF. Comparisons were made of the cytoplasm density, the diameter of oocytes with or without zona pellucida (ZP), the thickness of the ZP, the size of the perivitelline space (PVS), ZP dissolution time, and cortical granule (CG) distribution before IVF, and CG exocytosis and polyspermic penetration after IVF. Oviductal oocytes have clear areas in the cytoplasm cortex, while in vitro–matured oocytes have very dense cortex. The diameter of ovulated oocytes with ZPs was significantly (P < 0.001) greater than that of in vitro–matured oocytes. However, no difference was observed in the diameter of the oocyte proper. Significantly (P < 0.001) thicker ZPs and wider PVSs were observed in the ovulated oocytes. The ZPs of ovulated oocytes were not dissolved by exposure to 0.1% pronase within 2 hr, but the ZPs of in vitro–matured oocytes were dissolved within 131.7 ± 7.6 sec. The ZPs of ovulated oocytes, but not of in vitro–matured oocytes, were strongly labeled by a lectin from archis hypogaea that is specific for β-D-Gal(1–3)-D-GalNAc. Polyspermy rate was significantly (P < 0.01) higher for in vitro–matured oocytes (65%) than for ovulated oocytes (28%). CGs of oviductal oocytes appeared more aggregated than those of in vitro–matured oocytes. Most of CGs were released from both groups of oocytes 6 hr after IVF regardless of whether they were polyspermic or monospermic oocytes. These results indicate that in vitro–matured and in vivo–matured pig oocytes possess equal ability to release CGs on sperm penetration. Unknown changes in the extracellular matrix and/or cytoplasm of the oocytes while in the oviduct may play an important role(s) in the establishment of a functional block to polyspermy in pig oocytes. Mol. Reprod. Dev. 49:308–316, 1998. © 1998 Wiley-Liss, Inc.     

Actin in triton-treated cortical preparations of unfertilized and fertilized sea urchin eggs

Triton-treated cortical fragments of unfertilized and fertilized sea urchin eggs prepared in the presence of greater than or equal to 5 mM EGTA contain 15-30% of the total egg actin. However, actin filaments are not readily apparent by electron microscopy on the cortical fragments of unfertilized eggs but are numerous on those of fertilized eggs. The majority of the actin associated with cortical fragments of unfertilized eggs is solubilized by dialysis against a low ionic strength buffer at pH 7.5. This soluble actin preparation (less than 50% pure actin) does not form proper filaments in 0.1 M KCl and 3 mM MgCl2, whereas actin purified from this preparation does, as judged by electron microscopy. Optical diffraction analysis reveals that these purified actin filaments have helical parameters very similar to those of muscle actin. Furthermore, the properties of the purified actin with regard to activation of myosin ATPase are similar to those of actin from other cell types. The possibility that actin is maintained in a nonfilamentous form on the inner surface of the unfertilized egg plasma membrane and is induced to assemble upon fertilization is discussed.     

Action of trypsin on the eggs of Dendraster excentricus

Crystalline trypsin in 3 × 10^?8 M concentration and higher, elicits fertilization membranes in the unfertilized eggs of Dendraster excentricus. These membranes are adequate in artificial parthenogenesis. If the action of trypsin on these eggs is continued for two or three hours the result is first, digestion of the membranes, followed later by reduction of the egg to amoeboid form. When fertilized, some of the partially digested eggs segment and form irregular cell masses, thus demonstrating that, in response to trypsin, there is first the cortical reaction giving rise to the fertilization membrane, and second, the progressive digestion and disintegration of the cytoplasm.Chymotrypsin causes rounding of the unfertilized eggs and, in rare instances, a few membranes, but the enzyme is not an adequate parthenogenetic agent.Fertilization of the egg renders the cytoplasm resistant to trypsin. The facts lead to the suggestion that fertilization liberates trypsin inhibitors in the cytoplasm.     

Changes in microtubule structures during the first cell cycle of physiologically polyspermic newt eggs

The unfertilized egg of the newt, Cynops pyrrhogaster, has a second meiotic spindle at the animal pole and numerous cortical cytasters. After physiologically polyspermic fertilization, all sperm nuclei incorporated into the egg develop sperm asters, and the cortical cytasters change into bundles of cortical microtubules. The size of the sperm asters in the animal hemisphere is ∼5.6-fold larger than that in the vegetal hemisphere. Only one sperm nucleus moves toward the center of the animal hemisphere to form a zygote nucleus with the egg nucleus. This movement is inhibited by nocodazole, but not by cytochalasin B. The centrosome in the zygote nucleus divides into two parts to form a bipolar spindle for the first cleavage synchronously with the nuclear cycle, but centrosomes of accessory sperm nuclei in the vegetal hemisphere remained to form monopolar interphase asters and subsequently degenerate around the first cleavage stage. The size of sperm asters in monospermically fertilized Xenopus eggs was ∼37-fold larger than those in Cynops eggs. Since sperm asters that formed in polyspermically fertilized Xenopus eggs exclude each other, the formation of a zygote nucleus is inhibited. Cynops sperm nuclei form larger asters in Xenopus eggs, whereas Xenopus sperm nuclei form smaller asters in Cynops eggs compared with those in homologous eggs. Since there was no significant difference in the concentration of monomeric tubulin between those eggs, the size of sperm asters is probably regulated by a component(s) in egg cytoplasm. Smaller asters in physiologically polyspermic newt eggs might be useful for selecting only one sperm nucleus to move toward the egg nucleus. Mol. Reprod. Dev. 47:210–221, 1997. © 1997 Wiley-Liss, Inc.     

A long-lasting electrically mediated block, due to the egg membrane hyperpolarization at fertilization, ensures physiological monospermy in eggs of the crab Maia squinado

In response to fertilization, the membrane potential (Em) of the crab egg hyperpolarizes from about -50 mV to about -80 mV in 400 msec. To establish whether this fast hyperpolarization is correlated with physiological polyspermy or conversely mediates an electrical block to polyspermy, we examined the morphological and electrophysiological characteristics of eggs from the crab Maia squinado. Fertilized naturally spawned eggs were found to be physiologically monospermic and their average Em was constant at -77 +/- 0.5 mV. To examine a possible electrical block ensuring this monospermy, unfertilized eggs were voltage clamped at various Em values ranging from +20 to -90 mV, inseminated, and examined morphologically. All eggs clamped at +20 to -65 mV responded by developing a fertilization current, If. It consisted of an outwardly directed K+ current in one or several steps, each caused by a single spermatozoon interacting with the egg membrane. The percentage of eggs clamped at values more negative than -65 mV, which responded at insemination by developing an If, decreased and dropped to 0 at -80 mV. This indicated that the membrane processes occurring during the contact between gametes and eliciting an electrical response by the egg membrane are voltage dependent. Further, the spermatozoon never penetrated into eggs voltage clamped at a Em between +20 and -60 mV and at voltages more negative than -75 mV. Em values between -65 and -75 mV were required for spermatozoon incorporation into the egg, indicating that sperm entry is also voltage dependent. It is proposed that the hyperpolarization of the egg membrane in response to fertilization constitutes a long-lasting electrical block to polyspermy in crab eggs.     

Intracellular pH change does not accompany egg activation in the mouse

In the sea urchin, some other marine invertebrates, and the frog, Xenopus, egg activation at fertilization is accompanied by an increase in intracellular pH (pH_i). We measured pH_i, in germinal vesicle (GV)-intact mouse oocytes, ovulated eggs, and in vivo fertilized zygotes using the pH indicator dye, SNARF-1. The mean pH_i was 6.96 ± 0.004 (± SEM) in GV-intact oocytes, 7.00 ± 0.01 in ovulated, unfertilized eggs, and 7.02 ± 0.01 in fertilized zygotes, indicating no sustained changes in pH_i after germinal vesicle breakdown (GVBD) or fertilization. To examine whether transient changes in pH_i occur shortly after egg activation, mouse eggs were parthenogenetically activated by 7% ethanol in phosphate buffered saline (PBS); no significant change in pH_i followed ethanol activation. Since increased Na⁺/H⁺ antiporter activity is responsible for pH_i increase in the sea urchin, pH_i was measured in the absence of added bicarbonate or CO₂ la condition under which the antiporter would be the only major pH_i regulatory mechanism able to operate, since the others were bicarbonate- dependent) in GV-intact oocytes, ovulated eggs, and in vivo fertilized zygotes to determine whether a Na⁺/H⁺ antiporter was activated. There was no physiologically significant difference in pH_i after GVBD or fertilization, when pH_i was measured in bicarbonate-free medium, nor any change upon parthenogenetic activation. Thus, a change in pH_i is not a feature of egg activation in the mouse. © 1996 Wiley-Liss, Inc.