Freeze-fracture analysis of structural reorganization during meiotic maturation in oocytes of Xenopus laevis

Summary During meiotic maturation, the cortex of oocytes of Xenopus laevis undergoes structural reorganization, visualized in this study by freeze-fracture electron microscopy. In the full-grown but immature oocyte, annulate lamellae are dispersed throughout the subcortex of the egg, 5 to 20 m from the plasma membrane. The annulate lamellae consist of well-organized stacks of membrane with visible pores. Stimulation of meiotic maturation by progesterone leads to disruption of the annulate lamellae and formation of an elaborate cortical endoplasmic reticulum which surrounds the cortical granules and intertwines throughout the cortex of the mature egg. Pore-like structures similar to those previously observed in the subcortical annulate lamellae are observed in the mature cortical endoplasmic reticulum. The cortical endoplasmic reticulum is often in close apposition with the plasma membrane and with membranes of cortical granules, but no junctions are visualized. This study provides further evidence that the cortical endoplasmic reticulum develops during progesterone-stimulated meiotic maturation in vitro, and that the annulate lamellae are precursors to the cortical endoplasmic reticulum.     

Ultrastructural changes during nuclear maturation in Bufo arenarum oocytes.

During progesterone-induced nuclear maturation the oocytes of Bufo arenarum undergo a series of nuclear and cytoplasmic changes. The breakdown of heterocellular communications between the follicular cell projections and the oocyte microvilli, and the consequent enlargement of the perivitelline space, were observed at the animal pole. The more evident cytoplasmic feature during nuclear maturation comprised the gathering of glycogen granules in clusters, some phagocytosed by empty vesicles. With respect to the location of these vesicles, some were observed in close proximity to the oolemma and others were freely suspended in the perivitelline space, extruded from the oocyte. Other visible events were the disruption of the annulate lamellae, the formation of an elaborate cortical endoplasmic reticulum and the rearrangement of the cortical granules in a monolayer immediately beneath the oolemma together with aggregates of endoplasmic reticulum cisternae. Our results show that during nuclear maturation the nuclear oocyte changes include a flattening of the spherical oocyte nucleus, its migration towards the surface of the animal pole, the disappearance of the nucleoli and the dissolution of the nuclear envelope.     

An ultrastructural analysis of mitosis and cytokinesis in the zygote of the sea urchin, Arbacia punctulata

Post-fertilization events leading to the cleavage of the zygote of the sea-urchin, Arbacia punctulata were examined with the light and electron microscopes. Prior to prophase of the first cleavage division, endoplasmic reticulum and annulate lamellae become organized around the zygotic nucleus to produce a crescent-shaped structure which is defined as the streak (Harvey, '56). With the advent of prophase the streak undergoes morphogenic events which lead to the formation of the mitotic asters. During this transition there is a loss of annulate lamellae and a concomitant increase in endoplasmic reticulum. Annulate lamellae are not found as a part of the mitotic apparatus and are not again observed within the embryo until the two cell stage. During telophase, karyomeres are formed which consist of chromosomes delimited by a porous bilaminar envelope. Blastomere nuclei are produced following the fusion of the outer laminae, and subsequently by the fusion of the inner laminae of the envelopes encompassing the karyomeres.     

Self-organization of endoplasmic reticulum aggregates in cells with overexpression of nucleoporin POM121

The nuclear pore complexes are complex protein structures located in the nuclear envelope, where they control the nuclear-cytoplasmic transport, and inside the stacks of endoplasmic reticulum cisternae, annulate lamellae. After overexpression of some nucleoporins, numerous granules are visible in the cytoplasm. According to the published data, these granules are the annulate lamellae. In the current paper, the structural organization of POM121-containing granules was analyzed using correlative light and electron microscopy. The ultrastructural study demonstrates that POM121-containing granules are not annulate lamellae but aggregates of endoplasmic reticulum membranes. Thus, overexpressed POM121 is not able to induce the annulate lamella formation. The mechanisms of self-organization of non-functional structures (such as the aggregates of endoplasmic reticulum membranes described here) and possible involvement of these mechanisms in the formation of cellular structures are discussed.     

Ultrastructure of in vivo fertilization in the goat

In vivo fertilization of goat eggs has been studied by electron microscopy. Eggs were recovered from superovulated or natural cyclic goats, 32 to 52 hours after the onset of oestrus; only eggs recovered between 46 and 52 hours were fertilized. Spermatozoa penetrated the zona pellucida tangentially leaving vesiculated products of the acrosome reaction at the zona surface. As sperm penetrated into the ooplasm, the second meiotic division completed and cortical granule exocytosis occurred. However a few unreacted cortical granules usually remained in the cortex of the two fertilized eggs, adjacent to the plasma membrane. After swelling the two pronuclei presented similar ultrastructural morphology: they contained small, compact, agranular nucleoli and unevenly distributed chromatin. The cytoplasm in close vicinity to the apposed pronuclei contained large stacks of annulate lamellae, smooth endoplasmic reticulum, prominent Golgi complexes, as well as dense areas of unidentified material. The abundance of cytoplasmic organelles near the pronuclei might be the expression of intensive metabolic activity. Conversely, in the cortex of fertilized ova several large organelles-free cytoplasmic areas were randomly distributed.     

Structural organization and possible functional role of annulate lamellae containing cytoplasmic pores

This review is devoted to annulate lamellae, a specific compartment of endoplasmic reticulum that occurs, presumably, in actively growing and rapidly dividing cells (oocytes, embryonic and tumor cells). We summarized both earlier and recent data on the dustribution of annulate lamellae in various cell types, on their morphology, and the distribution of interaction with intracellular structures at various treatments. As the annulate lamellae contain cytoplasmic pore complexes, a special attention was paid to their relation with nuclear pores. Possible functions of the annulate lamellae in intracellular processes and, particularly, in nuclear envelope assembly, are discussed.     

Unusual intracytoplasmic lamellar bodies in a malignant gonadal stromal tumor

Characteristic intracytoplasmic lamellar bodies were found in a malignant gonadal stromal tumor. These bodies consisted of the stacks of up to 200 tubular cisternae arranged in parallel. Each cisterna had a circular section in tangential view and a diameter of about 85 nm. The cisternae on the outermost side of these lamellar bodies tended to be dilated and adorned with ribosomes. The ends of cisternae were often contiguous with rough-surfaced endoplasmic reticulum. The latter feature is also seen in annulate lamellae, but periodically spaced annuli or discontinuities characteristic of annulate lamellae were never observed. Furthermore, fine ribosomal granules resembling a rosary were recognizable along the whole circumference of the outer surface of each cisterna. The unique structure we describe is a cytoplasmic organelle which, like annulate lamellae, is closely associated with the endoplasmic reticulum and is presumed to be related to the genesis of rough-surfaced endoplasmic reticulum in tumor cells.     

Annulate lamellae and crystalline inclusions in granular endoplasmic reticulum of the islet organ and associated tissues of a cyclostome,Myxine glutinosa

Cytoplasmic annylate lamellae were found in the islet organ of a cyclostome, the hagfish (Myxine glutinosa), predominantly in cells interpreted as young proliferating beta-cells, and also in endocrine cells and enterocytes of the bile duct and gut and in the endothelial cells of small blood vessels. A close association was observed annulate lamellae and granular endoplasmic reticulum. Both in cells with and in those without annulate lamellae, crystalline inclusions of proteinaceous nature were seen in granular endoplasmic reticulum. These inclusions were occasionally closely associated to annulate lamellae, and a direct continuity could be seen between granular endoplasmic reticulum and the outer nuclear membrane surrounding an inclusion partially situated in the perinuclear cisterna. Rod-shaped structures and rounded electron dense bodies were seen in the nuclei of some islet parenchymal cells. The presence of annulate lamellae in the islet organ and associated tissues of Myxine glutinosa is believed to be related to the very high phylogenetic age of this species. The close association observed between annulate lamellae, granular endoplasmic reticulum, crystalline inclusions, and sometimes also nuclear membranes, may be of functional significance.     

Annulate lamellae in plant cells: Formation during microsporogenesis and pollen development in Canna generalis Bailey

Summary The occurrence of stacked annulate lamellae is documented for a plant cell system, namely for pollen mother cells and developing pollen grains of Canna generalis. Their structural subarchitecture and relationship to endoplasmic reticulum (ER) and nuclear envelope cisternae is described in detail. The results demonstrate structural homology between plant and animal annulate lamellae and are compatible with, though do not prove, the view that annulate lamellar cisternae may originate as a degenerative form of endoplasmic reticulum.     

Intranuclear and cytoplasmic annulate lamellae in the polyploid giant cells of the trophoblast

Intranuclear and cytoplasmic annulate lamellae in polyploid giant cells of the trophoblast have been studied in rat placenta on days 12--17 of development. The annulate lamellae are present in the cytoplasm within a limited time, being visible on day 12 only. These are arranged in bundles near the nucleus to be moving then to the cytoplasm. The end parts of annulate lamellae are broadened to make cisterns of rough endoplasmic reticulum. Unlike the cytoplasmic annulate lamellae, those found within the nucleus are seen in part of the nuclei investigated throughout the whole period examined to look as single structures (not gathered in bundles), they can be branching, separating closed spaces within the nucleus (making local swellings in the loci of branching; the latter having electron dense or transparent vesicles). Association with nuclear chromatin in some regions is a peculiar feature of the intranuclear annulate lamellae. This association is especially obvious at endoprophase in the cycle ofthe polytene nucleus during the somatic conjugation--chromonemes unite in a bundle and condense. Ultrastructural changes of the annulate lamellae is noted throughout the polytene nucleus cycle and during the cell differentiation. It is supposed that in the case of temporary labile chromosome polyteny in the nuclear cycle, which is characteristic of mammalian trophoblasts, annulate lamellae can well compare, in their function, with the synaptonemal complex--these prevent from too tight associations of homologues in the course of somatic conjugation of chromosomes.     

INTRANUCLEAR AND CYTOPLASMIC ANNULATE LAMELLAE IN TUNICATE OOCYTES

Electron microscope studies were made on various tunicate oocytes at different stages of growth and development. Both the inner and outer lamellae of the perforated nuclear envelope demonstrate considerable blebbing activity. The blebs of the inner lamella detach into the nucleoplasm where they undergo a special type of fusion process resulting in the formation of numerous, usually single, differentiated annulate lamellae of various lengths. The blebbing of the outer layer of the nuclear envelope contributes to the vesicular and granular endoplasmic reticulum characteristically present in the ooplasm and perhaps to the differentiation of cytoplasmic annulate lamellae as well. Cytoplasmic stacks of annulate lamellae frequently have ribosomes associated with them. In addition, granular accumulations are sometimes observed around or between the annuli. The morphological evidence suggests that, at least in many cases, the annuli in the annulate lamellae are patent.     

Are annulate lamellae in the Drosophila embryo the result of overproduction of nuclear pore components?

Annulate lamellae are cytoplasmic organelles composed of stacked sheets of membrane containing pores that are structurally indistinguishable from nuclear pores. The functions of annulate lamellae are not well understood. Although they may be found in virtually any eucaryotic cell, they occur most commonly in transformed and embryonic tissues. In Drosophila, annulate lamellae are found in the syncytial blastoderm embryo as it is cleaved to form the cellular blastoderm. The cytological events of the cellularization process are well documented, and may be used as temporal landmarks when studying changes in annulate lamellae. By using morphometric techniques to analyze electron micrographs of embryos, we are able to calculate the number of pores per nucleus in nuclear envelopes and annulate lamellae during progressive stages of cellularization. We find that annulate lamellae pores remain at a low level while nuclear envelopes are expanding and acquiring pores in early interphase. Once nuclear envelopes are saturated with pores, however, the number of annulate lamellae pores increases more than 10-fold in 9 min. Over the next 30 min it gradually declines to the initial low level. On the basis of these results, we propose (a) that pore synthesis and assembly continues after nuclear envelopes have been saturated with pores; (b) that these supernumerary pores accumulate transiently in cytoplasmic annulate lamellae; and (c) that because these pores are not needed by the embryo they are subsequently degraded.     

A FINE STRUCTURAL ANALYSIS OF CLEAVAGE INDUCTION AND FURROWING IN THE EGGS OF ARBACIA PUNCTULATA

A fine structural study has been carried out on the various formed elements present before, during, and after the first cleavage division, not only in normally developing Arbacia eggs, but also in eggs which have been induced to cleave prematurely by high-pressure centrifugation. The aim has been to ascertain whether or not any of the morphologically identifiable components may be involved in initiating the furrowing process. Also, attention has been given to the fine structure of the cytoplasmic cortex, particulary in the walls of the furrow, in the hope of reaching a better understanding of the mechanics of cleavage. The annulate lamellae and the membranous envelope of the nucleus are the only formed elements which disappear shortly before cleavage, not only in eggs undergoing normal division, but also in eggs which have been induced to cleave ahead of schedule by high-pressure, high-force centrifugation. Therefore, it is suggested as a tentative hypothesis that materials liberated upon disintegration of the nuclear membrane and the annulate lamellae play an essential role in initiating and effecting the furrowing reaction, especially since the stratification of these elements in experimentally induced eggs corresponds to the position of the developing furrow. Another of the membranous elements in the egg, the Golgi complex, shows considerable modification as a result of high-pressure centrifugation, but these structures do not undergo disintegration. Rather, they become curled into rounded bodies. The vacuole population is not greatly affected by inducing treatments. During cleavage, both naturally occurring and experimentally induced, a considerable number of 50 A filaments appear in the denser cytoplasmic cortex, but only in the walls of the furrow. These filaments are similar to those which have been demonstrated in a number of contractile cells. Accordingly, it is suggested that this fibrillar system may be actively involved in the development of the cleavage force.     

The Origin and Fate of Annulate Lamellae in Maturing Sand Dollar Eggs

Electron micrograph evidence is presented that the nuclear envelope of the mature ovum of Dendraster excentricus is implicated in a proliferation of what appear as nuclear envelope replicas in the cytoplasm. The proliferation is associated with intranuclear vesicles which apparently coalesce to form comparatively simple replicas of the nuclear envelope closely applied to the inside of the nuclear envelope. The envelope itself may become disorganized at the time when fully formed annulate lamellae appear on the cytoplasmic side and parallel with it. The concept of interconvertibility of general cytoplasmic vesicles with most of the membrane systems of the cytoplasm is presented. The structure of the annuli in the annulate lamellae is shown to include small spheres or vesicles of variable size embedded in a dense matrix. Dense particles which are about 150 A in diameter are often found closely associated with annulate lamellae in the cytoplasm. Similar structures in other echinoderm eggs are basophilic. In this species, unlike other published examples, the association apparently takes place in the cytoplasm only after the lamellae have separated from the nucleus. If 150 A particles are synthesized by annulate lamellae, as their close physical relationship suggests, then in this species at least the necessary synthetic mechanisms and specificity must reside in the structure of annulate lamellae.     

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.     

The origin and fate of annulate lamellae in maturing sand dollar eggs

Electron micrograph evidence is presented that the nuclear envelope of the mature ovum of Dendraster excentricus is implicated in a proliferation of what appear as nuclear envelope replicas in the cytoplasm. The proliferation is associated with intranuclear vesicles which apparently coalesce to form comparatively simple replicas of the nuclear envelope closely applied to the inside of the nuclear envelope. The envelope itself may become disorganized at the time when fully formed annulate lamellae appear on the cytoplasmic side and parallel with it. The concept of interconvertibility of general cytoplasmic vesicles with most of the membrane systems of the cytoplasm is presented. The structure of the annuli in the annulate lamellae is shown to include small spheres or vesicles of variable size embedded in a dense matrix. Dense particles which are about 150 A in diameter are often found closely associated with annulate lamellae in the cytoplasm. Similar structures in other echinoderm eggs are basophilic. In this species, unlike other published examples, the association apparently takes place in the cytoplasm only after the lamellae have separated from the nucleus. If 150 A particles are synthesized by annulate lamellae, as their close physical relationship suggests, then in this species at least the necessary synthetic mechanisms and specificity must reside in the structure of annulate lamellae.     

Spontaneous assembly of pore complex-containing membranes ("annulate lamellae") in Xenopus egg extract in the absence of chromatin.

Extract prepared from activated Xenopus eggs is capable of reconstituting nuclei from added DNA or chromatin. We have incubated such extract in the absence of DNA and found that numerous flattened membrane cisternae containing densely spaced pore complexes (annulate lamellae) formed de novo. By electron and immunofluorescence microscopy employing a pore complex-specific antibody we followed their appearance in the extract. Annulate lamellae were first detectable at a 30-min incubation in the form of short cisternae which already contained a high pore density. At 90-120 min they were abundantly present and formed large multilamellar stacks. The kinetics of annulate lamellae assembly were identical to that of nuclear envelope formation after addition of DNA to the extract. However, in the presence of DNA or chromatin, i.e., under conditions promoting the assembly of nuclear envelopes, annulate lamellae formation was considerably reduced and, at sufficiently high chromatin concentrations, completely inhibited. Incubation of the extract with antibodies to lamin LIII did not interfere with annulate lamellae assembly, whereas in the presence of DNA formation of nuclear envelopes around chromatin was inhibited. Our data show that nuclear membrane vesicles are able to fuse spontaneously into membrane cisternae and to assemble pore complexes independently of interactions with chromatin and a lamina. We propose that nuclear envelope precursor material will assemble into a nuclear envelope when chromatin is available for binding the membrane vesicles, and into annulate lamellae when chromatin is absent or its binding sites are saturated.     

Annulate lamellae in frog adenohypophysis under normal and experimental conditions

Cytoplasmic annulate lamellae have been observed in frog (Rana ridibunda) adenohypophysis pars distalis from normal spring animals and from others which were submitted to experimental conditions inducing selective activation of different cell types. Cell activation, because of either the normal active period in the frog cycle or the experimental treatments, seems to be correlated with the occurrence annulate lamellae. These annulate lamellae consist of a succession of two relatively parallel membranes interrupted periodically by discontinuities similar to nuclear pores. Sometimes they have been observed connected to endoplasmic reticulum.     

Annulate lamellae and "yolk nuclei" in oocytes of the dragonfly, Libellula pulchella

Annulated membranes in the form of single and short lamellae are present adjacent to and parallel to the nuclear envelope in oogonia and early oocyte (synaptene) stages of the dragonfly, Libellula pulchella. These solitary and short annulate lamellae are usually continuous with long, part rough- and part smooth-surfaced cisternae which extend into more distal areas of the oogonial ooplasm. These particular annulate lamellae then either disappear or decrease in number to be replaced by a much more extensive system of annulate lamellae in the cortical ooplasm of previtellogenic oocytes. The differentiation of extensive stacks of annulate lamellae is consistently observed to be restricted to large cytoplasmic areas of considerable electron density. These cytoplasmic regions consist of material which stains basophilic and contains RNA but differs structurally from the large number of ribosomes which surround the dense masses. The cytoplasmic dense masses, in terms of their formation and staining reactions, are comparable to the "yolk nuclei" or "Balbiani bodies" described in insect oocytes in earlier studies. The results of the present study thus provide evidence that the appearance of cortical ooplasmic stacks of annulate lamellae in the dragonfly oocyte is specifically limited to cytoplasmic areas of high electron density which contain RNA but which do not have a ribosomal morphology.     

Annulate lamellae in adenomas of human pituitary glands.

Twelve nontumorous adenohypophyses and 36 various pituitary adenomas, removed by surgery, have been investigated by electron microscopy in order to shed some light on annulate lamellae, primarily on their ultrastructural features, incidence, origin, fate and functional significance. No annulate lamellae were found in the nontumorous adenohypophyses and in 33 pituitary adenomas. They were, however, detected in two adenomas consisting of undifferentiated cells and one adenoma composed of sparsely granulated prolactin cells indicating that these unique membrane configurations cannot be regarded as an exceedingly rare finding and, furthermore, that they may be disclosed not only in undifferentiated but occasionally in highly differentiated cells. Annulate lamellae may arise from endoplasmic reticulum and/or nuclear envelope and consist of arrays of smooth walled double membrane sheets exhibiting regularly spaced interruptions as well as continuities with the endoplasmic reticulum. No relationship was established between annulate lamellae and adenohypophysial secretory activity. Our findings seem to be consistent with the view that annulate lamellae are present in those cells which have the tendency to proliferate.