Egg Periphery Research Articles

Arabinogalactan proteins and arabinan pectins abound in the specialized matrices surrounding female gametes of the fern Ceratopteris richardii.

Both male and female gametes of archegoniates are highly specialized cells surrounded by an extraprotoplasmic matrix rich in AGPs, which are speculated to facilitate development and gamete fusion through Ca 2+) oscillations. An additional layer, the egg envelope, forms around the egg periphery, except at the fertilization pore, and contains arabinose-rich polymers that presumably impart flexibility for the rapidly growing zygote and embryo. The abundant AGPs and arabinan pectins associated with the eggs of C. richardii not only are integral to development, fertilization, and early embryogenesis, but also may be involved in desiccation tolerance important to the survival of the reproductive gametophyte. A defining feature of gametogenesis in archegoniates is the deposition of a special matrix outside of the plasmalemma of both egg and sperm cells that displaces the primary cell wall away from the protoplasm. It is within this matrix that gamete differentiation occurs. In leptosporangiate ferns, maturation of the egg cell involves the deposition of a second specialized wall, the so-called egg envelope that surrounds the cell except at the fertilization pore, a narrow site where gamete fusion takes place. We provide the first conclusive evidence of the macromolecular constituents in the unique structures surrounding fern egg cells before and after fertilization. To test the hypotheses that the egg extracellular matrix contains arabinogalactan proteins (AGPs) as does the sperm cell matrix, and that cell wall polysaccharides, especially pectins, are components of the egg envelope, we examined the expression patterns of AGPs and cell wall constituents during oogenesis in Ceratopteris richardii. Utilizing histochemical stains for callose, cellulose and AGPs coupled with immunogold localizations employing a suite of monoclonal antibodies to cell wall components (JIM13, JIM8, LM2, LM5, LM6, LM19, LM20 and anticallose), we demonstrate that AGPs, but not pectins, are abundant in the matrix around egg cells and degrading neck canal and ventral canal cells during archegonial development. A striking finding is that both AGPs and (1,5)-α-L-arabinan pectin epitopes are principle components of the egg envelope before and after fertilization, suggesting that they are important in both egg maturation and gamete fusion.

A study of embryonic development in eriophyoid mites (Acariformes, Eriophyoidea) with the use of the fluorochrome DAPI and confocal microscopy.

The embryonic development of four eriophyoid mite species, Cecidophyopsis ribis, Phytoptus avellanae, Oziella liroi and Loboquintus subsquamatus, has been studied with the use of fluorochrome DAPI and confocal microscopy. The first three nuclear divisions occur on the egg periphery (the groups of 2, 4, and 6 nuclei have been recorded), while the biggest part of yolk remains undivided. After four or five nuclear divisions all nuclei are situated only in one sector of the embryo, while other sectors contain only yolk suggesting possible meroblastic cleavage. Later, the formation of superficial blastoderm takes place. A few large yolk cells are situated inside the embryo. Germ band formation initiates as funnel-like cell invagination and leads to formation of a typical stage with four paired prosomal buds (chelicerae, palps, legs I and II). Each palp contains two lobes (anterior and posterior), the adult subcapitulum is presumably a fusion product of the anterior pair of the lobes. Neither rudiments of legs III and IV, traces of opisthosomal segments nor remnants of the prelarval exuvium under the egg shell were detected. Overall, the pattern of embryonic development in eriophyoids re-emphasizes the peculiarity of this ancient group of miniaturized phytoparasitic animals, and invites researches to pursue a deeper investigation of various fundamental aspects of this aberrant group of Acari. Further studies using various fluorescent dyes and transmission electron microscopy are needed to visualize plasma membranes and clarify the pattern of early cleavage of eriophyoids.

The embryonic development of the centipede Strigamia maritima

The geophilomorph centipede Strigamia maritima is an emerging model for studies of development and evolution among the myriapods. A draft genome sequence has recently been completed, making it also an important reference for comparative genomics, and for studies of myriapod physiology more generally. Here we present the first detailed description of myriapod development using modern techniques. We describe a timeline for embryonic development, with a detailed staging system based on photographs of live eggs and fixed embryos. We show that the early, cleavage and nuclear migration, stages of development are remarkably prolonged, accounting for nearly half of the total developmental period (approx 22 of 48days at 13°C). Towards the end of this period, cleavage cells migrate to the egg periphery to generate a uniform blastoderm. Asymmetry quickly becomes apparent as cells in the anterior half of the egg condense ventrally to form the presumptive head. Five anterior segments, the mandibular to the first leg-bearing segment (1st LBS) become clearly visible through the chorion almost simultaneously. Then, after a short pause, the next 35 leg-bearing segments appear at a uniform rate of 1 segment every 3.2h (at 13°C). Segment addition then slows to a halt with 40–45 LBS, shortly before the dramatic movements of germ band flexure, when the left and right halves of the embryo separate and the embryo folds deeply into the yolk. After flexure, segment morphogenesis and organogenesis proceed for a further 10days, before the egg hatches. The last few leg-bearing segments are added during this period, much more slowly, at a rate of 1–2segments/day. The last leg-bearing segment is fully defined only after apolysis of the embryonic cuticle, so that at hatching the embryo displays the final adult number of leg-bearing segments (typically 47–49 in our population).

Early embryonic development in the spider Achaearanea tepidariorum: Microinjection verifies that cellularization is complete before the blastoderm stage

The spider Achaearanea tepidariorum is emerging as a non-insect model for studying developmental biology. However, the availability of microinjection into early embryos of this spider has not been reported. We defined the early embryonic stages in A. tepidariorum and applied microinjection to its embryos. During the preblastoderm 16- and 32-nucleus stages, the energids were moving toward the egg periphery. When fluorochrome-conjugated dextran was microinjected into the peripheral region of 16-nucleus stage embryos, it was often incorporated into a single energid and inherited in the progeny without leaking out to surrounding energids. This suggested that 16-nucleus stage embryos consisted of compartments, each containing a single energid. These compartments were considered to be separate cells. Fluorochrome-conjugated dextran could be introduced into single cells of 16- to 128-nucleus stage embryos, allowing us to track cell fate and movement. Injection with mRNA encoding a nuclear localization signal/green fluorescent protein fusion construct demonstrated exogenous expression of the protein in live spider embryos. We propose that use of microinjection will facilitate studies of spider development. Furthermore, these data imply that in contrast to the Drosophila syncytial blastoderm embryo, the cell-based structure of the Achaearanea blastoderm embryo restricts diffusion of cytoplasmic gene products.

Histochemical Characterization of the Embryonic Stages in <I>Diatraea saccharalis</I> (Lepidoptera: Crambidae)

Embryonic development in Diatraea saccharalis F. (Lepidoptera: Crambidae) has been studied by means of whole mounts, histochemical techniques, and light microscopy. Three embryonic stages were identified up to larval eclosion. In the first stage, 0–10 h, intense multiplication of energids occurred, and significant changes in RNA levels were detected in distinct zones before the start of blastulation. During the second stage, 10–24 h, new protein and RNA sites were detected in the egg’s periphery. This was probably because of migration of energids and formation of the blastula. Gastrulation occurs during this stage and is characterized by intense synthetic and mitotic processes. The third stage, 25–168 h, showed ortochromatic and metachromatic regions at the posterior pole and in the yolk, egg periphery, and body of the embryo. Organogenesis closes approximately on the sixth day when the embryo starts feeding on the remaining yolk. Its development is complete by the end of the seventh day on which the larva ecloses.

Determination of first cleavage plane: the relationships between the orientation of the mitotic apparatus for first cleavage and the position of meiotic division-related structures in starfish eggs

In order to understand when the orientation of the first cleavage plane is fixed along the animal–vegetal axis in starfish eggs, the behavior of the sperm aster was examined by indirect immunofluorescence staining. After duplication, the sperm aster organizes the mitotic apparatus for first cleavage perpendicular to the cleavage plane. The sperm aster located in the egg periphery just after fertilization and moved to the site close to the animal pole rather than the egg center by meiosis II. At early metaphase II, duplication of the sperm aster was detected but the axis of the resultant sperm diaster randomly pointed. Subsequently, its axis had already turned perpendicular to the animal–vegetal axis before pronucleus fusion. These results indicate that the orientation processes of the sperm diaster consist of positioning before its duplication and successive determining its azimuth. Furthermore, the azimuth and position of the mitotic apparatus for first cleavage did not change by shifting or eliminating the meiotic division-related structures such as the germinal vesicle, meiotic spindle, and female pronucleus by micromanipulation. These results show that none of them determines the first cleavage plane. Therefore, we discuss the pointing mechanism of the first cleavage plane without the influence of these meiotic division-related structures.

The secretion of phenolic compounds following fertilization in Acrocarpia paniculata (Fucales, Phaeophyta)

We have used light and electron microscopy to study the secretion of phenolic substances by zygotes of the brown alga Acrocarpia paniculata (Turner) Areschoug. Phenolic compounds, packaged within vesicles known as physodes, are a major cytoplasmic component of eggs and zygotes of Acrocarpia paniculata. Physodes accumulate at the egg periphery, and were identified using several different histochemical stains. Immediately after fertilization, zygotes secrete large numbers of phenolic bodies. The magnitude of phenolic release is so great that the zygote membrane is distorted during this process. However, we have found no cytologic evidence for endocytosis. The secreted phenolics surround the zygote and appear to impair sperm motility by causing sperm to slow down and become motionless. As the zygote wall is secreted, the phenolics form part of the developing wall, becoming oxidized and brown in color. We conclude that phenolics play an important role in the early developmental processes of A. paniculata, and we discuss their possible functional significance.

Long-lasting exocytosis and massive structural reorganisation in the egg periphery during cortical reaction inPlatynereis dumerilii(Annelida, Polychaeta)

The course of the cortical reaction in the Platynereis dumerilii egg is described from live observation and from sectioned fixed material and is found to differ in several aspects from the course of cortical reactions in better-known systems. Cortical granules are unusually numerous. They are discharged by exocytosis during a period of about 25 min following fertilisation (18 degrees C). Most of the surplus membrane material brought to the egg surface by exocytosis is set free into the perivitelline space. Swelling of egg jelly precursor secreted by cortical granule exocytosis may be causal for the detachment of the vitelline envelope from the egg cell surface which, however, remains attached punctately to the vitelline envelope by about 30,000 microvilli. Under the strain of the distending vitelline envelope, the bases of the microvilli move and line up, pulling the cell surface into a network of ridges. The grooves in between the ridges are the sites of exocytoses. Cytochalasin B, generally destabilising actin filaments, induces rupture of the microvilli and exaggerated distension of the vitelline envelope during the cortical reaction. In a final phase of the cortical reaction the vitelline envelope wrinkles and falls back onto the egg cell surface, the microvilli shorten and the egg cell transiently becomes deformed by local contractions. The cortical reaction in the nereid egg is discussed as a process of distortion and reorganisation of the egg cortex and plasmalemma. The abundance of cortical granules accommodating egg jelly precursor in the Platynereis oocyte is attributed to the mode of so-called diffuse oogenesis characteristic of nereids, i.e. of differentiation of oocytes freely suspended in the coelomic fluid. In nereids, egg jelly therefore forms after fertilisation as opposed to ovulation.

Structure of yolk granules in oocytes and eggs of Blattella germanica and their interaction with vitellophages and endosymbiotic bacteria during granule degradation

Oocytes and embryos of the cockroach Blattella germanica were examined by optical and electron microscopy to study yolk granule degradation during embryo development. During vitellogenesis, progressively larger yolk granules are formed in the ooplasm and by chorionogenesis, the mature granules are packed so tightly that their shape is highly distorted. Throughout ovarian development, endosymbiotic bacteria lie at the follicle cell/oocyte interface. Just prior to chorionogenesis the endosymbionts transit the oocyte plasma membrane and cluster at the periphery. Bacteria become more numerous over the ventral region of the egg by day 1 postovulation and begin to invade the interior of the yolk mass from the ventral periphery. At that time, lysis of the nearby yolk granules occurs while those in the central ooplasm remain intact and free of bacteria up to day 4. Vitellophages become evident by day 2 postovulation. These cells are also distributed over the egg's periphery but are most numerous in the ventral region. Vitellophages, in association with the endosymbionts, protrude toward the yolk granules and extend filo- and lamellipodia over the granule surface. Portions of the yolk granules are then engulfed and sequestered as large vacuoles in the vitellophage's cytoplasm. The vacuoles then become vesiculated. As embryo development proceeds, the vesiculated portions partition into smaller multivesicular bodies. This study describes the dynamics of yolk granule-vitellophage interaction in embryos of B. germanica and suggests that yolk utilization entails the cooperative efforts of both vitellophages and endosymbiont bacteria.

The early embryonic development of the jumping bristletail Pedetontus unimaculatus machida (Hexapoda: Microcoryphia, Machilidae).

The early development of Pedetontus unimaculatus from maturation to germ rudiment formation has been described by light and electron microscopy. Newly laid eggs of P. unimaculatus are in the metaphase of the first maturation division, and two successive maturation divisions produce two polar bodies. Nuclear divisions up to the eighth or ninth are accompanied by cytoplasmic divisions, and are holoblastic. Each resulting blastomere contains a single nucleus. Most cleavages are radial, but a few are tangential resulting in the formation of primary yolk nuclei. After the eighth or ninth nuclear division, cytoplasmic divisions are restricted to the egg periphery, and these later cleavages are superficial. Boundaries of blastomeres gradually disappear. Nuclei, which settle in the peripheral cytoplasm, proliferate and differentiate into blastoderm cells, and they also give rise to secondary yolk nuclei. A posterior circular region of blastoderm thickens and concentrates to form a germ rudiment 50-100 μm in diameter. During the formation of a germ rudiment the serosal cuticle begins to form. A similar pattern of cleavage was observed in other species of the Machilidae belonging to four genera in two subfamilies (Machilinae, genus Haslundichilis; Petrobiinae, genera Pedetontus, Pedetontinus, Petrobiellus). The cleavage pattern of the machilids closely resembles that found in the myriapod groups, the Symphyla, Diplopoda, ana Pauropoda, as well as in the apterygote Collembola, but it differs from the purely superficial cleavage pattern characteristic of the apterygote Thysanura sensu stricto (Zygentoma) and the Pterygota. It is concluded 1) that the pattern of early total cleavage changing later to superficial cleavage is a plesiomorphic character for the Antennata, and 2) that the purely superficial pattern is an apomorphic character within the Hexapoda.

The cortical reaction in the egg of Discoglossus pictus: A study of the changes in the endoplasmic reticulum at activation

In Discoglossus pictus previous ultrastructural observations have shown that at the animal dimple, where sperm fuse with the egg, cortical granules (CG), vacuoles, and tightly packed clusters of small cisternae are present. At fertilization the clusters open (i.e., become loose) and give rise to longer cisternae arranged in whorls and chains which migrate toward the plasma membrane. The vacuoles fuse to form cisternae and exocytose along with the CG. In the rest of the egg periphery, while exocytosis occurs, the clusters do not open as a result of activation ( C. Campanella, R. Talevi, U. Atripaldi, and L. Quaglia (1986). In “Molecular and Cellular Biology of Fertilization” (J. L. Hedrick, Ed.). Plenum, New York ). We have recently conducted electrophysiological studies which have detected inward currents at the dimple center, outward current at the rest of the egg surface, and an eightfold increase in [Ca 2+] i which propagates from the site of activation throughout the egg ( R. Nuccitelli, D. Kline, W. Busa, R. Talevi, and C. Campanella (1988). Dev. Biol. 130, 120–132 ). In this paper we have asked whether the anionic current and the Ca 2+ increase could be causally related to the changes of the smooth endoplasmic reticulum (SER) at activation. The results obtained by activating the eggs in ion-substituted Ringers indicate that (1) the migration of cisternae is not dependent on the polarity of the activation current crossing the dimple, but is strongly impaired, together with CG exocytosis, by 5× Cl − Ringer; (2) TMB-8, a drug which partially blocks calcium release ( C. Y. Choiu and M. J. Malagodi (1975). Brit. J. Pharmacol. 53, 279–288 ), partially inhibits opening of cisternae clusters and the formation of an SER network in the dimple. This suggests a causal relationship between the Ca 2+ rise and the cluster transformation at activation.

Subcortical rotation in Xenopus eggs: An early step in embryonic axis specification

The amphibian egg undergoes a rotation of its subcortical cytoplasm relative to its surface during the first cell cycle. Nile blue spots applied to the egg periphery move with the subcortical cytoplasm and make rotation directly observable ( J.-P. Vincent, G. F. Oster, and J. C. Gerhart (1986). Dev. Biol. 113, 484 ). We have previously shown that the direction of rotation accurately predicts the orientation of the embryonic axis developed by the egg. This suggests an important role for subcortical rotation in axis specification. In this report, we provide two kinds of experimental evidence for the essential role of rotation, and against a role for other concurrent cytoplasmic movements such as the convergence of subcortical cytoplasm toward the sperm entry point in the animal hemisphere. First, dispermic eggs develop only one embryonic axis, which is oriented accurately in line with the direction of the single rotation movement and not with the two convergence foci that form in the animal hemisphere. Rotation probably modifies the vegetal, not animal, hemisphere since axial development is normal in dispermic eggs despite highly altered animal subcortical movement. Second, we show that the amount of rotation correlates with the extent of dorsal development. UV irradiation of the vegetal hemisphere, or cold shock of the egg, inhibits rotation effectively. When there is no rotation, there is no dorsal development. On average within the egg population, increasing amounts of rotation correlate with the increasingly anterior limit of the dorsal structures of the embryonic body axis. However, individual partially inhibited eggs vary greatly in the amount of axis formed following a given amount of movement. Furthermore, the egg normally rotates more than is necessary for the development of a complete axis. These findings suggest that rotation, although essential, does not directly pattern the antero-posterior dimension of the body axis, but triggers a response system which varies from egg to egg in its sensitivity to rotation. This system is artificially sensitized by exposure of the egg to D 2O shortly before rotation. We show that D 2O-treated eggs produce extensive axes despite very limited rotation, often developing into hyperdorsal embryos. However, like normal eggs, they depend on rotation and cannot form dorsal structures if it is eliminated.

Subcortical rotation in Xenopus eggs: a preliminary study of its mechanochemical basis.

The amphibian egg undergoes a 30 degree rotation of its subcortical contents relative to its surface during the first cell cycle, a displacement of 350 micron in 50 min. This is directly visualized by following the movement of an array of Nile blue (a subcortical stain) spots applied to the egg periphery (Vincent, Oster, and Gerhart: Dev Bio 113:484-500, '86). We have investigated the mechanochemical basis of this unusual cell motility. Subcortical rotation depends on microtubule integrity during its entire course and is insensitive to inhibitors of microfilament assembly. It does not depend on newly synthesized proteins for its operation or timing, and it does not involve calcium-dependent processes. Finally, we show that vegetal fragments of the egg can complete rotation on their own, indicating that mechanochemical components can operate locally in this hemisphere.

Cleavage and blastoderm formation in normal and experimentally deformed naked eggs of a dipteran insect

The eggs of the gall midgeHeteropeza pygmaea develop parthenogenetically inside of the mother larva. They lack a chorion and remain enveloped by the follicular epithelium. After experimental elimination of the follicular epithelium “naked” eggs are formed, which reach the blastoderm stage but remain spherical instead of assuming an elongated shape. To analyze this peculiar egg development and the roles of egg shape and envelope during development, the ultrastructure of cleaving normal and naked eggs was investigated. It was shown that the number of elements of Golgi apparatus and endoplasmic reticulum strongly increases during early cleavage. Their association with cleavage furrows and nuclei suggests that these organelles play a dominant role in membrane production. Egg yolk consists of lipids and glycogen, wheareas no proteins are found. Cleaving eggs contain numerous vesicles with lysosomal characteristics, indicating intense autophagic processes. Cleavage furrow formation occurs independently from the positioning of cleavage nuclei. The numerous microtubules, which are associated with cleavage furrows and nuclei and located in the egg periphery, the intercellular bridges, and in the central part of the egg, suggest that the cytoskeleton has an important role in cleavage furrow formation, blastoderm layer establishment, and yolk localization. Since these processes are accurately accomplished in naked spherical eggs, they can be considered as independent of normal egg shape and the follicular epithelium.

Ooplasmic segregation in theTubifex egg: Mode of pole plasm accumulation and possible involvement of microfilaments.

Ooplasmic segregation, i.e. the accumulation of pole plasm in theTubifex egg, consists of two steps: (1) Cytoplasm devoid of yolk granules and lipid droplets migrates toward the egg periphery and forms a continuous subcortical layer around the whole egg; (2) the subcortical cytoplasm moves along the surface toward the animal pole in the animal hemisphere and toward the vegetal pole in the vegetal hemisphere, and finally accumulates at both poles of the egg to form the animal and vegetal pole plasms. Whereas the subcortical layer increases in volume during the first step, it decreases during the second step. This is ascribed to the compact rearrangement in the subcortical layer of membraneous organelles such as endoplasmic reticulum and mitochondria. The number of membraneous organelles associated with the cortical layer increases during the second step. Electron microscopy reveals the presence of microfilaments not only in the cortical layer but also in the subcortical layer. Subcortical microfilaments link membraneous organelles to form networks; some are associated with bundles of cortical microfilaments. The thickness of the cortical layer differs regionally. The pattern of this difference does not change during the second step. On the other hand, the subcortical cytoplasm moves ahead of the 'stationary' cortical layer. The accumulation of pole plasm is blocked by cytochalasin B but not by colchicine. The first step of this process is less sensitive to cytochalasin B than the second step, suggesting that these two steps are controlled by differnt mechanisms. The mechanical aspects of ooplasmic segregation in theTubifex egg are discussed in the light of the present observations.

The embryonic development of the primitive moth, Neomicropteryx nipponensis Issiki (lepidoptera, micropterygidae): Morphogenesis of the embryo by external observation.

The micropterygid moth Neomicropteryx nipponensis belongs to the most primitive suborder Zeugloptera of the Lepidoptera. During embryogenesis the small circular germ disk formed on the ventral egg surface invaginates deeply into the yolk. It finally separates from the egg periphery or rudimentary serosa, and becomes a sac-shaped germ rudiment. Its anterior part later develops into the germ band, while its posterior part is the future amnion. Just before revolution of the embryo, the embryo assumes a completely superficial position beneath the yolk. Neither amnion nor serosa rupture during revolution; by completion of dorsal closure they have been incorporated into the yolk to form the secondary dorsal organ. The formation of the germ rudiment and embryonic membranes in N. nipponensis resembles those of swift moths, Endoclyta (suborder Monotrysia) and of the caddisflies, Stenopsyche (Trichoptera), but differs from those of ditrysian Lepidoptera. The secondary dorsal organ has never been found in any other lepidopteran embryos; however, it is formed in N. nipponensis and in the Trichoptera. The results of the present study strongly support the general phylogenetic views that the Zeugloptera have a close affinity to the Trichoptera.

ELECTRON MICROSCOPIC OBSERVATIONS ON THE EXTRA-GRANULAR ZONE OF THE EMBRYO OF THE SEA URCHIN, HEMICENTROTUS PULCHERRIMUS.

"Extra-granular zone" proposed by Dan and Dan (3) was studied with the electron microscope. Two stages are noticed during the process of its formation; at first, a relatively yolk granule-free zone constitutes the egg periphery which should correspond to that observed by Dan and Dan, and later, this zone is occupied by a layer of the vacuoles (about 1.3 μm in mean diameter) with highly electron-transparent inclusions which migrated to the egg surface from the inner cytoplasm after fertilization. These vacuoles are scarcely found in the micromeres of 16-cell stage embryo.

DNA and RNA Synthesis by Fertilised, Cleavage Arrested Sea Urchin Eggs

Eggs of Lytechinus variegatus which have been fertilised and maintained at 10 degrees C do not undergo segmentation, but do replicate DNA. It has been found that RNA synthesis is initiated in these eggs following a period of DNA synthesis resulting in amounts of DNA/egg comparable to those amounts found in late blastulae (1000 genome equivalents). The RNA synthesised has been studied by autoradiographic and sucrose density gradient centrifugation. RNAse-sensitive grains appear in autoradiographs of cleavage-arrested eggs exposed to tritiated uridine beginning two days after fertilisation at 10 degrees C. Incorporation of labelled uridine into the 18 and 25 s region of sucrose gradients parallels the appearance of acid-insoluble radioactivity in the uridine autoradiographs. Localisation of the RNA in the autoradiographs was shown to be in close association with Feulgen-positive areas of the egg periphery. Since differentiation is generally thought to involve changing patterns of transcription of the zygote genome, transcription patterns in the absence of cleavage and normal morphogenesis is of considerable interest. The study of molecular events occurring in a time span during which there is no cell division may have bearing on the role of cell division in differentiation.

An electron microscope study of cytoplasmic organelle transformations in developing mouse oocytes.

The development of the mouse oocyte during the primordial, primary and secondary follicular growth stages was studied by means of the electron microscope.During the early stages of oocyte maturation, mitochondrial multiplication takes place along with an apparent temporary transition from round to oval shape. The internal structure of many of the mitochondria is altered by separation of membranes of a crista to form a vacuole. This enlarges to pear-shaped configurations and gradually it forms so large a structure as to result in compression of adjacent cristae, thereby altering the entire appearance of the organelle.Dense round bodies encapsulated by a single membrane are found in the cytoplasm of oocytes of primary follicles near the periphery. The Golgi complex appears in primary follicle oocytes as an aggregation of vesicles. Gradually the number of lamellae in the complexes increase and these organelles become more peripherally located. The "Balbiani yolk nuclei" apparently is represented by a conglomeration of Golgi complexes and are present only in primordial and young primary follicle oocytes.The endoplasmic reticulum appears in the early stages only as rough-surfaced vesicles. At later stages individual cisternae become prominent. Apparently, a modified form of E. R. appears during maturation of the secondary follicle oocyte.Multivesicular complexes, each consisting of two components, small vesicles and larger vesicles enclosing microvesicles (multivesicular bodies), were commonly found during all stages of oocyte growth. The secondary follicle oocytes frequently contain multilamellar bodies. These are commonly found in juxtaposition to the multivesicular complexes and also near the egg periphery and occasionally near the nuclear envelope.

Cytochemical Studies on the Second Gradient Axis of Fertilized Oyster Eggs

1. The activity of glucose-6-phosphatase, glucose-6-phosphate dehydrogenase and malate dehydrogenase in the oyster eggs and the relationship between the egg nucleolus and egg periphery were studied.2. It is the peripheral zone of eggs where some enzymes blocked in unfertilized eggs are released from the blockage and increases their activity remarkably after the fertilization.3. Upon the insemination, the egg periphery evokes the activity of the creative area and the nucleolus.4. Changing the direction of the first gradient axis parallel to the egg axis (the second gradient axis), KP axis regains its stability of position which has been lost by the insemination.5. These changes of metabolism in oyster eggs are the primary event of fertilization.