Previtellogenic oocytes of South African largemouth bass Micropterus salmoides Lacépède 1802 (Actinopterygii,Perciformes) - the Balbiani body,cortical alveoli and developing eggshell

The ovaries of the largemouth bass Micropterus salmoides, an alien and invasive species in South Africa, contain a germinal epithelium which consists of germline and somatic cells, as well as previtellogenic and late vitellogenic ovarian follicles. The ovarian follicle consists of an oocyte surrounded by follicular cells and a basal lamina; thecal cells adjacent to this lamina are covered by an extracellular matrix. In this article, we describe the Balbiani body and the polarization and ultrastructure of the cytoplasm (ooplasm) in previtellogenic oocytes. The nucleoplasm in all examined oocytes contains lampbrush chromosomes, nuclear bodies and several nucleoli near the nuclear envelope. The ultrastructure of the nucleoli is described. Numerous nuage aggregations are present in the perinuclear cytoplasm in germline cells as well as in the ooplasm. Possible roles of these aggregations are discussed. The ooplasm contains the Balbiani body, which defines the future vegetal region in early previtellogenic oocytes. It is comprised of nuage aggregations, rough endoplasmic reticulum, Golgi apparatus, mitochondria, complexes of mitochondria with nuage-like material, and lysosome-like organelles. In mid-previtellogenic oocytes, the Balbiani body surrounds the nucleus and later disperses in the ooplasm. The lysosome-like organelles fuse and transform into vesicles containing material which is highly electron dense. As a result of the fusion of the vesicles of Golgi and rough endoplasmic reticulum, the cortical alveoli arise and distribute uniformly throughout the ooplasm of late previtellogenic oocytes. During this stage, the deposition of the eggshell (zona radiata) begins. The eggshell is penetrated by canals containing microvilli and consists of the following: the internal and the external egg envelope. In the external envelope three sublayers can be distinguished.     

Fine structure of the human ovum in the pronuclear stage

A penetrated ovum was recovered from the oviduct of a 33 year old surgical patient who had had sexual intercourse 26 hr before the operation. The ovum was in the pronuclear stage. The ooplasmic organelles were mainly represented by mitochondria, endoplasmic reticulum components, and Golgi elements. Small vesicles were found in the space between the two sheets of the pronuclear envelope. These vesicles appeared to be morphologically similar to the ER vesicles in the ooplasm and were considered to be involved in pronuclear development. Numerous annulate lamellae were seen in the ooplasm as well as in the pronuclei. Ooplasmic crystalloids were also observed. These were thought to represent cytoplasmic yolk. Remnants of the penetrating spermatozoon were found in close relation to one of the pronuclei. The fine structure of the first and second polar body is also described. The nuclear complement of the first polar body consisted of isolated chromosomes, whereas the second polar body contained a membrane-bounded nucleus. In consideration of the possibility that polar body fertilization may take place, these differences in nuclear organization could be of importance. Other recognizable differences between the two polar bodies were presence of dense cortical granules and microvilli in the first polar body, and absence of these structures in the second. These dissimilarities were considered to be related to the organization of the egg cytoplasm at the time of polar body separation.     

Cytochemical studies of oocyte development in Sarcophaga lineatocollis Macq. (Muscidae: Diptera)

The ovary of Sarcophaga lineatocollis is a typical polytrophic ovary. Each of its 25-30 ovarioles is composed of a small terminal filament, a small germarium and a vitellarium consisting of the egg follicle. The tunica propria is a noncellular, PAS-positive membrane. The ovarian follicle contains fifteen trophocytes and one oocyte. RNA is synthesized with the aid of the nuclei in the trophocyte cytoplasm, which are RNA- and PAS-positive. Protein is deposited intensively in the early stages of the trophocytes. The trophocytes of Sarcophaga lineatocollis synthesize RNA and protein more actively than the oocyte. In this fly, protein yolk precursor (PYP) bodies are supplied by the trophocyte cytoplasm to the ooplasm at an advanced stage of development. Nucleolar budding and vacuolation are observed in the trophocytes. RNA, DNA, protein and PYP bodies appear to be transported to the ooplasm from the trophocytes. Pyknotic trophocyte nuclei can be seen entering the ooplasm. The perinuclear Golgi bodies of the trophocytes help in the production and maturation of PYP bodies in the trophocytes before they are organized and passed on to the oocytes. Some RNA is contributed to the oocyte by the follicular epithelium. All these processes leading to maturation and development of the oocyte are discussed and interpreted.     

The human pronuclear ovum: Fine strcture of monospermic and polyspermic fertilization in vitro

The fine structure of pronuclear ova (monospermy and polyspermy) and one-cell embryos has been investigated in our IVF programme. Sixteen oocytes were collected at laparoscopy after appropriate hormonal stimulation and were matured and fertilized in vitro by methods that have given rise to normal pregnancies. Pronuclear ova showing monospermic fertilization had two vesicular pronuclei surrounded by aggregations of cellular organelles. The male pronucleus was closely associated with a sperm axoneme, while the female pronucleus was dismantling its envelope and condensing its chromatin ahead of its counterpart in late pronuclear ova. Each pronucleus had dispersed chromatin, dense compact nucleoli, and intranuclear annulate lamellae. Smooth endoplasmic reticulum, annulate lamellae, Golgi complexes, and mitochondria formed a conspicuous part of the perinuclear ooplasm. The one-cell embryos were either in syngamy or in the process of undergoing first cleavage. Positive evidence of cortical granule release and second polar bodies were detected in the perivitelline space. A block to polyspermy seemed to operate at the level of the inner zona. Dispermic and polyspermic ova had 3–16 pronuclei resembling those of monospermic ova and had sperm tails in the ooplasm. Sperm were also seen penetrating the inner zona and were occasionally found in the perivitelline space. Incomplete cortical granule release and early signs of cytoplasmic fragmentation were noted in polyspermic ova. Both normal and abnormal features of these ova are reported and compared with pronuclear structure in vivo and in vitro.     

Fine structure of spermatogonia and primary spermatocytes in rabbits

Summary The fine structure of rabbit Spermatogonia and primary spermatocytes in meiotic prophase has been studied with different methods of preparation, including a technique for acid phosphatase activity. The spermatogonial cytoplasm is rich in free ribosomes and containes moderate amounts of vesicular, smooth-surfaced endoplasmic reticulum and mitochondria, a simple Golgi-apparatus, some micropinocytotic vesicles, and occasional multivesicular bodies, vacuoles and dense bodies with acid phosphatase activity. The large type A Spermatogonia have a prominent nucleolus and their mitochondria sometimes form clusters with a dense intermitochondrial substance, similar to that in spermatocytes.The nucleus and cytoplasm of primary spermatocytes increase markedly in volume and density during meiotic prophase. The Golgi apparatus enlarges and becomes more differentiated and finally forms small proacrosome granules. The endoplasmic reticulum produces numerous small, mainly smooth vesicles and might also be the source of a new organelle: numerous piles of narrow cisternae with opaque contents. These piles disintegrate late in prophase. The mitochondria become aggregated in clusters with dense intermitochondrial substance and their internal structure is characterized by highly dilated cristae and small particles, interpreted as mitochondrial ribosomes, in the matrix. The role of these structures in the formation of new mitochondria is discussed. The clusters of mitochondria finally disperse and their cores of dense intermitochondrial substance, possibly containing ribonucleoprotein, coalesce into a large chromatoid body similar to that in spermatids. Micropinocytosis and a few lysosomes occur in most spermatocytes. The pachytene nuclei show prominent nucleoli and a distinct sex vesicle without any synaptinemal complex.The importance for spermatid differentiation of some events taking place in the cytoplasm of primary spermatocytes is emphasized.Financial support for this study was received from the Swedish Medical Research Council.     

Ultrastructure of the ovarian follicles in the placentotrophic Andean lizard of the genus Mabuya (Squamata: Scincidae)

We studied the ultrastructural organization of the ovarian follicles in a placentotrophic Andean lizard of the genus Mabuya. The oocyte of the primary follicle is surrounded by a single layer of follicle cells. During the previtellogenic stages, these cells become stratified and differentiated in three cell types: small, intermediate, and large globoid, non pyriform cells. Fluid‐filled spaces arise among follicular cells in late previtellogenic follicles and provide evidence of cell lysis. In vitellogenic follicles, the follicular cells constitute a monolayered granulosa with large lacunar spaces; the content of their cytoplasm is released to the perivitelline space where the zona pellucida is formed. The oolemma of younger oocytes presents incipient short projections; as the oocyte grows, these projections become organized in a microvillar surface. During vitellogenesis, cannaliculi develop from the base of the microvilli and internalize materials by endocytosis. In the juxtanuclear ooplasm of early previtellogenic follicles, the Balbiani's vitelline body is found as an aggregate of organelles and lipid droplets; this complex of organelles disperses in the ooplasm during oocyte growth. In late previtellogenesis, membranous organelles are especially abundant in the peripheral ooplasm, whereas abundant vesicles and granular material occur in the medullar ooplasm. The ooplasm of vitellogenic follicles shows a peripheral band constituted by abundant membranous organelles and numerous vesicular bodies, some of them with a small lipoprotein core. No organized yolk platelets, like in lecithotrophic reptiles, were observed. Toward the medullary ooplasm, electron‐lucent vesicles become larger in size containing remains of cytoplasmic material in dissolution. The results of this study demonstrate structural similarities between the follicles of this species and other Squamata; however, the ooplasm of the mature oocyte of Mabuya is morphologically similar to the ooplasm of mature oocytes of marsupials, suggesting an interesting evolutionary convergence related to the evolution of placentotrophy and of microlecithal eggs. J. Morphol., 2010. © 2010 Wiley‐Liss, Inc.     

A comparative histochemical study of fish (Channa maruleus) and amphibian (Bufo stomaticus) oogenesis

Summary Comparative histochemical studies on the fish (Channa maruleus) and amphibian (Bufo stomaticus) oogenesis demonstrate a great similarity in the growth and differentiation of their egg follicle. The ooplasm, germinal vesicle and egg-membranes show distinct morphological and cytochemical changes during previtellogenesis and vitellogenesis.During previtellogenesis the various components of the follicle are engaged in the synthesis of protoplasm as shown by the proliferation of yolk nucleus substance, mitochondria and some lipid bodies in the ooplasm and of nucleoli in the germinal vesicle. The substance of the yolk nucleus consisting of proteins, lipoproteins and RNA first appears adjacent to the nuclear membrane. Numerous mitochondria of lipoprotein composition, and some lipid bodies consisting of unsaturated phospholipids lie in association with the yolk nucleus which forms substratum for the former. The lipid bodies, present inside the germinal vesicle, follicular epithelium, and adjacent to the plasma membrane in association with some pinocytotic vacuoles, have been considered to play a significant role in the active transport of some substances from the environment into the ooplasm and from the latter into the germinal vesicle. The follicular epithelium itself is very poorly developed, negating its appreciable role in the contribution of specific substances into the oocyte, which seem to be contributed by the germinal vesicle showing a considerable development of nuclear sap, basophilic granules and nucleoli consisting of RNA and proteins; many large nucleoli bodily pass into the cytoplasm during the previtellogenesis of Channa, where their substance is gradually dissolved. The intense, diffuse, basophilic substance of the cytoplasm is believed due to free ribosomes described in many previous ultrastructural studies.During vitellogenesis, the various deutoplasmic inclusions, namely carbohydrate yolk, proteid yolk and fatty yolk, are deposited in the ooplasm. The carbohydrate yolk bodies rich in carbohydrates originate in association with the plasma membrane and correspond to vesicles and cortical granules of previous studies. The proteid yolk consisting of proteins and some lipoproteins, and fatty yolk containing first phospholipids and some triglycerides and then triglycerides only are deposited under the influence of yolk nucleus substance, mitochondria and cytoplasm. The mitochondria and yolk nucleus substance foreshadow in some way the pattern of these two deutoplasmic inclusions and persist at the animal pole of mature egg while the other inclusions of previtellogenesis disappear from view. The pigment granules, which also show a gradient from the animal to vegetal pole in Bufo, are also formed in association with yolk nucleus substance and mitochondria. Some glycogen also appears in both the species. The nuclear membrane becomes irregular due to the formation of lobes. The lipid bodies of the germinal vesicle come to lie outside the nuclear membrane, suggesting active transport of some substances into the ooplasm; many nucleoli bodily pass into the ooplasm of Bufo, where they are gradually absorbed. The amount of basophilic granules is considerably increased in the germinal vesicle during vitellogenesis. Various egg-membranes such as outer epithelium, thin theca, single-layered follicular epithelium, zona pellucida or vitelline membrane surround the vitellogenic oocytes. The zona pellucida formed between the oocyte and follicle cells consists of a carbohydrate-protein complex. The follicle cells show lipid droplets, mitochondria and basophilic substance in their cytoplasm. The various changes that occur in the components of the follicle during vitellogenesis seem to be initiated by gonadrotrophins formed under the influence of specific environmental conditions.The author wishes to express sincere appreciation and gratitude to Dr. Gilbert S. Greenwald, who has made the completion of this investigation possible.Ph. D. Population Council Post-doctoral Fellow.     

Dirofilaria immitis: ultrastructural aspects of oocyte development and zygote formation.

Ultrastructural observations of the ovary and uterus of Dirofilaria immitis reveal some characteristics of oogonia, oocytes, and uterine sperm. Oogonia are confined to the distal portion of the ovary including a blind tip, where a morphologically distinct terminal cap cell was not observed. These cells contain a nucleus with a nucleolus, numerous dense bodies, scanty ribosomes, lipid droplets, and an occasional mitochondrion. Endoplasmic reticulum is lacking and Golgi complexes were observed only in fully grown oogonia. Primary oocytes located in the middle portion of the ovary are large, elongate, and have a complete set of organelles including many small mitochondria, fragmentary endoplasmic reticulum, ribosomes, Golgi complexes, and very few dense bodies. These cells are arranged into many rosettes about central cytoplasmic masses, the rachises, to which they maintain cytoplasmic continuity by pseudopodlike processes. The rachises contain no organelle except a few dense granules and are bound by winding membranes. Oocytes from the proximal portion differ from those of the middle portion of the ovary in their larger size, round shape, absence of many organelles, presence of small dense granules, and lacking a rachis. Dense bodies are specific to the oogonia and exhibit DNase susceptibility and a positive reaction for a mitochondrial enzyme. These findings together with their decreased number and a concomitant increase of mitochondria in the oocytes suggest a relationship between these bodies and mitochondria.Uterine sperm of D. immitis are of the amoeboid type and contain several chromatin masses without a nuclear envelope, many mitochondria, and specialized membranous organelles referred to as mesosomelike vesicles. The vesicles are probably originated from the sperm plasma membrane. Upon fertilization, the entire spermatozoon penetrates the oocyte and its contents are gradually dissolved in the ooplasm with a simultaneous appearance of large numbers of ribosomes at the site of dissolution. Ribosomes were later found in the nucleus. A pronucleus was not observed. These findings are basically in agreement with those described for Ascaris but differ in the morphologic features and number of rachises, presence of dense bodies, absence of refringent granules in the oocytes and the absence of a refringent body and presence of several chromatin masses in the sperm.     

Morphology of the epithelium of the extratesticular rete testis, ductuli efferentes and ductus epididymidis of the adult male rabbit

The fine structure of the epithelium lining the extratesticular rete testis, ductuli efferentes and ductus epididymidis of the rabbit has been investigated. In the ductuli efferentes the epithelium is composed of two cell types, principal cells and ciliated cells. The latter type is distinguished from principal cells by the presence of cilia projecting into the lumen and the position of the nucleus in the apical half of the cell. Principal cells in this segment are characterized by micropinocytotic vesicles on the surface plasma membrane and a variety of small dense bodies scattered throughout the cytoplasm. In the ductus epididymidis basal cells replace ciliated cells as the second cell type, but differences between various segments of the epididymis are related to the fine structure of the principal cells. In the proximal caput epididymidis (Nicander's region 1) the principal cells are tall with long microvilli. They typically contain a small Golgi apparatus and a cluster of dense bodies adjacent to the nucleus. In the distal caput epididymidis (Nicander's regions 2-5) the apical cytoplasm of principal cells is filled with numerous micropinocytotic vesicles and large multivesicular bodies; these features are interpreted as signs of absorptive activity. The multivesicular bodies are absent from the cytoplasm of principal cells in the corpus epididymidis (Nicander's region 6) and, instead, numerous elements of smooth endoplasmic reticulum, a large Golgi apparatus, lipid droplets and dense bodies characterize principal cells in this segment. Towards the proximal cauda epididymidis (Nicander's region 7), the number of dense bodies (lysosomes) in the cytoplasm increases considerably. In the globose cauda (Nicander's region 8), the principal cells are reduced in height, and in addition to the features described in region 7, are characterized by a concentric array of rough endoplasmic reticulum in the basal cytoplasm. These observations are discussed in relation to the role of the epididymis in promoting the maturation and survival of spermatozoa.     

Cytodifferentiation and vitellogenesis during oogenesis in arachnida: Cytological studies on developing oocytes of a harvestman

Light and electron microscope studies were made on harvestman oocytes during the course of their origin, differentiation, and vitellogenesis. The germ cells appear to originate from the ovarian epithelium. They subsequently migrate to the outer surface of the epithelium, where they remain attached often by means of stalk cells which suspend them in the hemocoel during oogenesis. The “Balbiani bodies,” “yolk nuclei,” or “nuage” constitute a prominent feature of young, previtellogenic oocytes, and take the form of large, but variable sizes of electron-dense cytoplasmic aggregates with small fibrogranular components. The cytoplasmic aggregates fragment and disperse, and cannot be detected in vitellogenic oocytes. The young oocytes become surrounded by a vitelline envelope that appears to represent a secretory product of the oocyte. The previtellogenic oocytes are impermeable to horseradish peroxidase under both in vivo and in vitro conditions. In addition to mitochondria, dictyosomes, and abundant ribosomes, the ooplasm of the previtellogenic oocyte acquires both vesicular and lamellar forms of the rough-surfaced endoplasmic reticulum. In many areas, a dense homogeneous product appears within the cisternae of the endoplasmic reticulum and represents nascent yolk protein synthesized by the oocyte during early stages of vitellogenesis. Later in vitellogenesis, the oocyte becomes permeable to horseradish peroxidase under both in vivo and in vitro conditions. This change is associated with a massive process of micropinocytosis which is reflected in the presence of large numbers of vesicles of variable form and structure in the cortical ooplasm. Both spherical and tubular vesicles are present, as are coated and uncoated vesicles. Stages in the fusion of the vesicles with each other and with developing yolk platelets are illustrated. In the harvester oocytes, vitellogenesis is a process that involves both autosynthetic and heterosynthetic mechanisms.     

COTTON EMBRYOGENESIS: THE EARLY DEVELOPMENT OF THE FREE NUCLEAR ENDOSPERM

An electron microscope study was made of the central cell and the development of the free nuclear endosperm surrounding the zygote and synergids during the first three days after pollination. The cytoplasm of the central cell, concentrated around the partially-fused polar nuclei, contains many ribosomes, mitochondria and large, dense, starch-containing plastids, some dictyosomes and lipid bodies, and long, single cisternae of rough endoplasmic reticulum (RER) that frequently terminate in whorls. Dense, core-containing microbodies are closely associated with the RER. After fertilization the cytoplasm of the 2-and 4-nucleate endosperm shows an increase in number of dictyosomes, and in amount of RER which becomes stacked in arrays of parallel cisternae. Cup-shaped plastids are associated with many long, helical polysomes. Perinuclear aggregates of dense, granular material also appear after fertilization. Granular aggregates and helical polysomes disappear after the first few divisions of the primary endosperm nucleus. During the second and third days of development there is an increase in dictyosome number and RER proliferation, and endosperm nuclei become deeply lobed. Concurrently, there is a sharp decline in the starch and lipid reserves of the central cell and elaborate transfer walls are formed at the micropylar end of the embryo sac and on the outer surface of the degenerating synergid. The transfer walls contain groups of small, membrane-bound vesicles, and are associated with large numbers of mitochondria and with the smooth endoplasmic reticulum.     

Previtellogenic and vitellogenic oocytes in ovarian follicles of cultured siberian sturgeon Acipenser baerii (Chondrostei,Acipenseriformes)

Previtellogenic and vitellogenic oocytes in ovarian follicles from cultured Siberian sturgeon Acipenser baerii were examined. In previtellogenic oocytes, granular and homogeneous zones in the cytoplasm (the ooplasm) are distinguished. Material of nuclear origin, rough endoplasmic reticulum, Golgi complexes, complexes of mitochondria with cement and round bodies are numerous in the granular ooplasm. In vitellogenic oocytes, the ooplasm comprises three zones: perinuclear area, endoplasm and periplasm. The endoplasm contains yolk platelets, lipid droplets, and aggregations of mitochondria and granules immersed in amorphous material. In the nucleoplasm, lampbrush chromosomes, nucleoli, and two types of nuclear bodies are present. The first type of nuclear bodies is initially composed of fibrillar threads only. Their ultrastructure subsequently changes and they contain threads and medium electron dense material. The second type of nuclear bodies is only composed of electron dense particles. All nuclear bodies impregnate with silver, stain with propidium iodide, and are DAPI‐negative. Their possible role is discussed. All oocytes are surrounded by follicular cells and a basal lamina which is covered by thecal cells. Egg envelopes are not present in previtellogenic oocytes. In vitellogenic oocytes, the plasma membrane (the oolemma) is covered by three envelopes: vitelline envelope, chorion, and extrachorion. Vitelline envelope comprises four sublayers: filamentous layer, trabecular layer 2 (t2), homogeneous layer, and trabecular layer 1 (t1). In the chorion, porous layer 1 and porous layer 2 are distinguished in most voluminous examined oocytes. Three micropylar cells that are necessary for the formation of micropyles are present between follicular cells at the animal hemisphere. J. Morphol. 278:50–61, 2017. ©© 2016 Wiley Periodicals,Inc.     

Ultrastructure of the Developing and Fertilized Embryo Sac of Amaranthus hypochondriacus L.

Amaranthus hypochondriacus embryo sac development was investigatedbefore and after fertilization. During the early stages of development,the young embryo sac displays three antipodal cells at the chalazalpole that degenerate very early in the maturation process, beforethe synergids and egg cell are completely differentiated. Themature embryo sac is composed only of the female germ unit.The synergid cells organize a filiform apparatus accompaniedby the presence of mitochondria and dictyosomes with numerousvesicles. The involvement of the synergids in transport andsecretory functions related to pollen tube attraction and guidance,are discussed. The egg cell is located at the micropylar polenear the synergids and displays exposed plasma membranes atthe chalazal pole. The fertilized egg cell does not exhibitmarked changes after fertilization except for the closure ofthe cell wall. The central cell is the largest cell of thisvery long embryo sac. The fused nucleus is close to the eggapparatus before fertilization and displays a remarkable chalazalmigration after gamete delivery. The ultrastructure of the centralcell cytoplasm and the numerous wall ingrowths around this cellsuggest an important role in nutrient transportation. Aftergamete delivery, the embryo sac displays electron dense bodiesthat aggregate within the intercellular space between the synergids,egg cell and central cell. These bodies, that appear in theembryo sac of several plants, are probably involved in gametedelivery for double fertilization. The possibility of biparentalinheritance of mitochondria in this plant is also discussed.Copyright 1999 Annals of Botany Company Amaranthus hypochondriacus, grain amaranth, embryo sac, fertilization.     

Cytochemical localization of GalNAc and GalNAcβ1,4Galβ1,4 disaccharide in mouse zona pellucida

Carbohydrate residues contained in the zona pellucida play a key role in the process of sperm-egg interaction. In vitro fertilization experiments have shown that a specific monoclonal antibody against GalNAcş,4Galş,4 disaccharide inhibits fertilization in mice. In the present study, the ultrastructural cytochemical localization of GalNAc residues and the GalNAcş,4Galş,4 disaccharide was carried out in ovarian and postovulatory oocytes by using lectin-gold cytochemistry and immunocytochemistry. Plant lectins SBA and DBA showed an affinity for the entire zona pellucida matrix of ovarian oocytes throughout the follicular maturation; however, immunoreactivity for GalNAcş,4Galş,4 disaccharide was not detected in ovarian oocytes at the earliest stages of follicular development but was found to be associated with the inner region of the zona matrix at the trilaminar primary follicle stage. The Golgi apparatus, vesicular aggregates, and cortical granules of the oocyte were intensely labeled by SBA and DBA throughout follicular development. Immunoreactivity to GalNAcş,4Galş,4 disaccharide was first observed in the Golgi apparatus and vesicular aggregates in trilaminar primary follicles. No immunoreactivity was observed in the cortical granules. In postovulatory oocytes, results were similar to those observed in ovarian oocytes. Our results thus suggest that (1) GalNAcş,4Galş,4 disaccharide residues are present only in the inner region of the zona pellucida and, therefore, might be involved in sperm penetration through the zona pellucida, (2) the inner and outer regions of the zona pellucida contain different oligosaccharide chains, (3) the vesicular aggregates detected in the oocyte could represent an intermediate step in the secretory pathway of zona pellucida glycoproteins and might be involved in the formation of cortical granules.     

ULTRASTRUCTURAL ZONATION OF ADRENOCORTEX IN THE RAT

The fine structure of the different zones in the adrenal cortex of the adult rat has been studied under the electron microscope. Four regions mainly differentiated by the mitochondrial morphology, the lipid droplets, and the structure of the ground cytoplasm were recognized. In the glomerular zone mitochondria are thin and elongated with an abundant matrix. The inner structure is characterized by the presence of tubules of 300 A that are straight or bend at an angle and which may be grouped in parallel array giving a pseudocrystalline pattern. The wall of each tubule is a finger-like projection of the inner membrane and its cavity corresponds to the outer chamber of the mitochondrion. In the intermediary zone mitochondria are larger and irregular. The matrix is filled with convoluted tubules and vesicular elements. The lipid droplets are larger and irregular in the glomerulosa and and small in the intermedia. The ground substance is dense and contains free ribosomes in the glomerulosa and starts to be vacuolated in the intermedia. In the fasciculata mitochondria are round or oval and are filled with vesicular elements with a mean size of 450 A. Larger vesicles and more clear elements (vacuoles) are seen near the edge as if their content was diluted. Some of these vacuoles protrude on the surface. In the reticular zone mitochondria are also vesicular but frequently show signs of alteration and disruption. Dense elements recognized as microbodies are observed in the fasciculata but they increase in number in the reticularis. These results are discussed on the light of the so called zonal theory of the adrenal cortex. Two stages in the differentiation of the mitochondria are postulated. The tubular structure of the glomerulosa undergoes a process of disorientation and dilatation of the tubules to form the tubulo-vesicular elements of the intermediary zone. In a second stage of differentiation, by fragmentation of the tubules, the vesicular structure of fasciculata is formed. These findings are discussed from the viewpoint of the relationship between mitochondria and synthesis of steroid hormones. A secretory process that starts within mitochondria by the formation of vesicles and proceeds into the ground cytoplasm, as extruded and more clear vacuoles, is postulated.     

Electron microscopic observations on sperm entry and pronuclear formation in naked eggs of the rose bitterling in polyspermic fertilization

Unfertilized eggs of the rose bitterling (Rhodeus ocellatus ocellatus) were squeezed out of females that had an elongated ovipositor and were dechorionated mechanically with fine forceps in physiological saline. The dechorionated eggs were transferred into fresh water then inseminated at once by spermatozoa of the same species. A large number of spermatozoa was found on the surface of eggs that had not yet had cortical reaction following insemination. The surface of the naked eggs responded by formation of many small cytoplasmic protrusions (viz., fertilization cones) at sperm attachment sites. The formed fertilization cones were rosettelike structures formed by the aggregation of some bleblike swellings devoid of microvilli and microplicae. About 10 min after insemination, the fertilization cones retracted, but marks of their presence characterized by less microvilli and microplicae remained in the eggs 15 min after insemination. Many spermatozoa penetrated into the cytoplasm of each naked egg. The sperm nuclear envelope disappeared by means of vesiculation resulting from fusion of the inner and outer membranes. The sperm nucleus decondensed and developed into a larger male pronucleus. Smooth-surfaced vesicles surrounded the decondensing sperm nucleus and formed the new male pronuclear envelope. Sperm mitochondria and flagella were found in the egg 15 min after insemination. The response of the egg surface to sperm entry and pronucleus formation are discussed.     

Oogenesis in the leech Glossiphonia heteroclita (Annelida, Hirudinea, Glossiphoniidae) II. Vitellogenesis, follicle cell structure and egg shell formation

By the end of previtellogenesis, the oocytes of Glossiphonia heteroclita gradually protrude into the ovary cavity. As a result they lose contact with the ovary cord (which begins to degenerate) and float freely within the hemocoelomic fluid. The oocyte's ooplasm is rich in numerous well-developed Golgi complexes showing high secretory activity, normal and transforming mitochondria, cisternae of rER and vast amounts of ribosomes. The transforming mitochondria become small lipid droplets as vitellogenesis progresses. The oolemma forms microvilli, numerous coated pits and vesicles occur at the base of the microvilli, and the first yolk spheres appear in the peripheral ooplasm. A mixed mechanism of vitellogenesis is suggested. The eggs are covered by a thin vitelline envelope with microvilli projecting through it. The envelope is formed by the oocyte. The vitelline envelope is produced by exocytosis of vesicles containing two kinds of material, one of which is electron-dense and seems not to participate in envelope formation. The cortical ooplasm of fully grown oocytes contains many cytoskeletal elements (F-actin) and numerous membrane-bound vesicles filled with stratified content. Those vesicles probably are cortical granules. The follicle cells surrounding growing oocytes have the following features: (1) they do not lie on a basal lamina; (2) their plasma membrane folds deeply, forming invaginations which eventually seem to form channels throughout their cytoplasm; (3) the plasma membrane facing the ovary lumen is lined with a layer of dense material; and (4) the plasma membrane facing the oocyte forms thin projections which intermingle with the oocyte microvilli. In late oogenesis, the follicle cells detach from the oocytes and degenerate in the ovary lumen.     

Cotton embryogenesis: The pollen cytoplasm

Summary The ultrastructure and composition of cotton (Gossypium hirsutum) pollen, exclusive of the wall, was examined immediately before and after germination. The pollen grain before germination consists of two parts: the outer layer and a central core. The outer layer contains large numbers of mitochondria and dictyosomes as well as endoplasmic reticulum (ER). The core contains units made of spherical pockets of ER which are lined with lipid droplets and filled with small vesicles; the ER is rich in protein and may contain carbohydrate while the vesicles are filled with carbohydrate. Starch-containing plastids are also present in the core as are small vacuoles. The cytoplasm of the pore regions contains many 0.5 spherical bodies containing carbohydrate. After germination the ER pockets open and the lipid droplets and small vesicles mix with the other portions of the cytoplasm. With germination the pore region becomes filled with mitochondria and small vesicles. The vegetative nucleus is large, extremely dense and contains invaginations filled with coils of ER. A greatly reduced nucleolus is present in the generative cell which is surrounded by a carbohydrate wall. The cytoplasm of the generative cell is dense and contains many ribosomes, a few dictyosomes and mitochondria, many vesicles of several sizes, and some ER. No plastids were identified. The generative nucleus is also dense with masses of DNA clumped near the nuclear membrane. An unusual tubular structure of unknown origin or function was observed in the generative cell.     

Oogenesis in the annelid Enchytraeus albidus with special reference to the origin and cytochemistry of yolk

In the annelid Enchytraeus albidus the ovary is composed of packets containing eight synchronously developing oocytes. Each oocyte in the packet is connected, via a bridge, to a common cytoplasmic mass. Developmental synchrony of oocytes within individual packets is probably related to the ooplasmic continuity. The young previtellogenic oocyte contains many polysomes, a few cisternae of smooth and rough endoplasmic reticulum, small Golgi complexes, and mitochondria. Many of the mitochondria are dumbbell-shaped and may thus represent division stages. Vitellogenesis is marked by the appearance of peripherally located lipid yolk and small, densely staining granules scattered throughout the ooplasm. There is an increase of smooth endoplasmic reticulum, mitochondria, and enlarged Golgi elements. Small multivesicular-like bodies, the early stages of developing yolk, are derived from the Golgi complex. The mature yolk sphere is bipartite and consists of (a) a variable number of dense spheres, the core bodies, which are produced in the ooplasm by the Golgi complex and which become embedded in (b) a dense matrix. The electron opaque tracer, horseradish peroxidase is incorporated into the oocyte and deposited in the matrix suggesting that this component of the yolk sphere is obtained by micropinocytosis. Enzyme digestions and various cytochemical techniques suggest that the core bodies are rich in carbohydrate, probably as glyco- or mucoproteins, and that the matrix is rich in lipid.     

The Maturation of Oocyte Follicular Epithelium of Podarcis s. sicula Is Promoted by d-Aspartic Acid

We investigated whether the maturation of oocyte follicular epithelium of lizard is affected by d-aspartic acid (d-Asp). Our results demonstrated that d-Asp is endogenously present in the oocytes, and its distribution varies during the reproductive cycle and following intraperitoneal administration. At previtellogenesis, it is observed in the cytoplasm and nucleus of pyriform cells, in intermediate cells, in some small cells of the granulosa, in the ooplasm, and in some thecal elements. At vitellogenesis, d-Asp is localized in the proximity of the zona pellucida, in the theca, and in the ooplasm. Injected d-Asp is mainly captured by pyriform cells and ooplasm of previtellogenic oocytes, but a moderate accumulation is evident in the cytoplasm of some small granulosa cells and in the theca. d-Asp also increases the ovarian and plasmatic levels of 17β-estradiol and decreases those of testosterone. As a direct and/or indirect consequence of d-Asp, previtellogenic oocytes grow up and mature, resulting in a higher accumulation of carbohydrates in the granulosa, zona pellucida, and ooplasm, but also a reduction in the thickness of the granulosa layer and an increase of the theca stratum. Taken together, our results show that d-Asp may be related to the synchrony of reproduction, either enhancing the growth and maturation of follicular epithelium or influencing its endocrine functions. (J Histochem Cytochem 58:157–171, 2010)