bluuebunny/crossref_metadata_2025 huggingface dataset:
Asynchronization of Cell Division is Concurrently Related with Ciliogenesis in Sea Urchin Blastulae
Authors: MASUDA MICHITAKA , SATO HIDEMI
Published: January 2005

During the early development of the sea urchins, Temnopleurus toreumaticus, Temnopleurus hardwickii and Hemicentrotus pulcherrimus, the division synchrony in all blastomeres lasted only until the 4th cleavage and a regional synchrony or a graded activity of cell division appeared. In the midblastula stage prior to hatching, the regional synchrony vanished simultaneously with the formation of cilia, then the division proceeded asynchronously. The analysis of cell pedigrees confirmed that a variable extension of intercleavage times occurred after the ciliogenesis. In blastomeres derived from mesomeres of T. toreumaticus embryos, the mean intercleavage time extended from 48 min of the 8th cycle (pre‐ciliated) to 115 min of the 9th cycle (ciliated), and the coefficient of variation increased from 15% to 39%. We attempted a kinetic analysis of cell proliferation on the basis of the transition probability model of cell cycle control. We concluded that the minimum time required for the completion of the cell cycle was the decisive factor in the cell cycle succession of pre‐ciliated blastomeres, and that a sudden and sharp decrease in the transition probability of the ciliated blastomeres probably interpreted the abrupt slowing and asynchronization of the cleavage cycle at the time of ciliogenesis.
Somite Chondrogenesis in vitro: Differential Induction by Modified Matrix‐ a Biochemical and Morphological Study*
Authors: VASAN N. S. , MILLER E.
Published: January 2005

The stimulation of somite chondrogenesis by extracellular matrix components was studied by monitering the synthesis of cartilage‐specific large proteoglycan aggregates. Chick embryonic sternal proteoglycans were separated into various components: monomers, hyaluronic acid, link protein and glycosaminoglycan side chains. The effects of these components, either individually or in various combinations, on somite chondrogenesis was examined. Proteoglycan monomers, alone or in a mixture with other components, induced chondrogenesis. The other components did not have any stimulating effect of their own. The results of these induction studies were also observed on a Sepharose CL–2B column and correlated using electron microscopy. Stimulation of somites resulted in an increase in the amount of proteoglycan aggregation (material excluded from the column) and was in agreement with the morphological appearance of the matrix in that there was increased accumulation of large proteoglycan granules. A matrix mixture of collagen and proteoglycans showed significant stimulation. When the matrix environment of the somites was altered to be unfavorable to the explants (medium containing hyaluronic acid) there was altered synthesis of cartilage‐specific molecules. The results presented in this report strongly suggest that the composition of the extracellular matrix material is critical for somite chondrogenesis.
Inductive Capacities for the Dorsal Mesoderm of the Dorsal Marginal Zone and Pharyngeal Endoderm in the Very Early Gastrula of the Newt, and Presumptive Pharyngeal Endoderm as an Initiator of the Organization Center*
Authors: HAMA TADAO , TSUJIMURA HIDENOBU , KANÉDA TERUO , TAKATA KENZO , OHARA AKITO
Published: January 2005

The organization center of Cynops pyrrhogaster was divided into Parts 1, 2 and 3 of equal size (0.3×0.4 mm2) with presumptive fates as pharyngeal, pharyngeal+prechordal+trunk notochord, and trunk‐tail notochord, respectively. Movements and changes in size and shape of each part were followed through gastrulation. Differentiation tendencies of each part were examined under three conditions: I, isolated; II, sandwiched with presumptive ectoderm; 111, sandwiched with presumptive ectoderm after preculture in isolation for various times. In I, Parts 2 and 3 differentiated into dorsal mesoderm. In II, each part induced dorsal mesoderm and neural tissues, the frequency being highest in Part 2 and lowest in Part 3. In III, Parts 1 and 2 realized their presumptive fates, through changes in inductive capacities from trunk‐tail to head. This change progressed rapidly in Part 1, and slowly in Part 2. Part 3 required induction by neighbouring Part 2 to realize its presumptive fate. Changes of inductive capacity of Parts 1 and 2 respectively, were chronologically similar in normal development and in preculture experiments. Lastly, the primary presumptive pharyngeal zone at blastula was proposed to act as an initiator of the organization center, its programmed information being transmitted to Part 2, and then to Part 3.
Release of a Gonad‐Stimulating Substance from the Radial Nerves of the Cushion Star, Asterina pectinifera, in response to Excess Potassium*
Authors: TAKAHASHI NOBUAKI
Published: January 2005

The release of a gonad‐stimulating substance (GSS) from radial nerves stimulated by a K+‐rich solution was examined in the cushion star, Asterina pectinifera. The threshold concentration of K+necessary for the release of a sufficient amount of GSS to induce shedding of mature oocytes from ovarian fragments was 50 to 75 mM. The minimal amount of fresh nerve required for this release was 3 mg per ml of medium. The time of K+stimulation necessary to induce maximal GSS release was about 20 min. Repeated stimulations of radial nerves with 100 mM K+caused repeated release of GSS.
DNA Replication in Permeabilized Cells of Sea Urchin Embryos
Authors: MORIOKA MIZUE , SHIMADA HIRAKU
Published: January 2005

For study of the regulation of DNA replication in sea urchin embryos during the early stages of development, an embryonic cell system that was permeable to exogenously supplied nucleotides was established. Embryos were permeabilized by incubating them in hypotonic buffer containing 0.3 M glucose. The permeabilized embryonic cells maintained their morphological integrity, and synthesized DNA when supplied with exogenous dNTPs.

DNA synthesis in these permeabilized embryonic cells required the presence of ATP and three other deoxyribonucleoside triphosphates in addition to labeled dTTP. DNA synthesis was almost completely inhibited by N‐ethylmaleimide, and proceeded in a discontinuous fashion. Only cells permeabilized during the S phase could incorporate nucleoside triphosphates into DNA: cells permeabilized during other phases did not synthesize DNA. During a 60 min‐incubation period, over 10% of the genomic DNA was replicated under the experimental conditions used.
Isolation of Acrosomal Vesicles and Their Surrounding Membranes from Starfish Sperm*
Authors: CHRISTEN RICHARD
Published: January 2005

Intact acrosomal vesicles and their surrounding membranes were isolated from starfish sperm. The identification of the acrosomal vesicle and confirmation of its purification away from other sperm organelles was made by electron microscopy and SDS‐polyacrylamide gel electrophoresis.
Acceleration of the Cleavage in Sea Urchin Eggs by Treatments with Local Anesthetics (II) Treatments with Some Other Local Anesthetics than Procaine*
Authors: KOJIMA MANABU K.
Published: January 2005

Unfertilized sea urchin eggs were exposed to sea water solutions of local anesthetics, such as caffeine, tetracaine and ethyl urethane, and the herbicide, isopropyl N‐phenyl carbamate (IPC) for 10min and returned to normal sea water. Then they were inseminated 5min later. When eggs were pre‐treated with 1–2 mM caffeine, 0.02–0.05 mM tetracaine, 50–100 mM ethyl urethane and 2% saturated sea water of IPC, respectively, they could cleave and hatch earlier than the control eggs. However, when fertilized eggs were continuously post‐treated with solutions of the agents except IPC at the same concentrations as those in the case of the pre‐treatments, the fertilized eggs could not cleave or were retarded in development. The possible mechanisms of the cleavage acceleration by pre‐treatments with local anesthetics were discussed.
Cell‐type Conversion at Low Temperature in Dictyostelium discoideum
Authors: MAEDA MINEKO
Published: January 2005

The effects of low temperature (5°C) on cell‐type conversion in whole slugs of Dictyostelium discoideum and their anterior prestalk‐ and posterior prespore‐isolates were examined immunohistochemically and electronmicroscopically. When slugs were incubated for nine days at 5°C, the proportion of cells containing spore‐antigens increased from about 75% to 85%. More important, the proportion of prespore and spore cells increased from about 3% to 40% in anterior prestalk isolates incubated at 5°C for 12 days, but no cell‐type conversion from prespore to prestalk cells occurred in posterior prespore isolates. Therefore, the mechanism regulating the proportions of cells that operates at 21 °C does not operate at low temperature. The cells with full competence for stalk differentiation could change into stalk cells even at 5°C, because a short stalk was always formed when early culminants were transferred to low temperature. The effects of low temperature on several sequential steps of cell differentiation are discussed on the basis of these findings. The ultrastructural characteristics during the process of cell‐type conversion are also described.
Prestalk/prespore Differentiation of Dictyostelium Cells under the Conditions Favoring Stalk or Spore Cell Formation*
Authors: NAKAHARA YASUO , NOCE TOSHIAKI , TAKEUCHI IKUO
Published: January 2005

The differentiation processes of Dictyostelium discoideum cells under the conditions which favored either stalk or spore cell formation were examined by the use of prestalk‐ and prespore‐specific antibodies. In stalk cell‐forming conditions, cells reactive with prestalk‐specific monoclonal antibody (C1) increased rapidly early in development and later differentiated into stalk cells. No or only a few cells became reactive with prespore‐specific monoclonal (B6) and polyclonal (antispore) antibodies. Despite the fact that most cells terminally became spores under spore cell‐forming conditions, cells were first stained with the C1 antibody before becoming reactive with the B6 antibody. Unlike the case of normal development where cells coincidentally become reactive with the B6 and antispore antibodies, the appearance of the cells reactive with the latter was either delayed or suppressed. In conclusion, under either spore or stalk cell‐forming conditions, the appearance of the prestalk antigen preceded that of the prespore one, which is consistent with normal development.
Isolation and Characterization of Mesenchymal Cells from the Tail of Bullfrog Tadpoles*
Authors: YOSHIZATO KATSUTOSHI , NISHIKAWA AKIO
Published: January 2005

Mesenchymal cells were separated with the aid of EDTA, dispase, collagenase and hyaluronidase from tail fins of Rana catesbeiana tadpoles and were cultured to examine their morphology and response to thyroid hormone, which controls metamorphosis. The mensenchymal tissue consisted of two main types of cells, macrophage‐like cells (M cells) and fibroblastic cells (F cells), distinguishable by their morphological and biochemical characteristics. M cells were preferentially located near the boundary between the mesenchyme and the epidermis. They were more adhesive to the plastic culture dishes than F cells. The M cells developed a bizarre rod‐like branching process during one week of culture, which appeared to be identical to the process formed by mammalian macrophages undergoing clasmatosis. M cells cultured for more than one week tended to fuse together forming multinuclear giant cells. F cells were found uniformly in the tissue and were active in collagen biosynthesis. Thyroid hormone at a physiological concentration greatly reduced the life span of F cells, but did not reduce that of M cells.
SEM observation of the External Yolk Syncytial Layer in Blastopore Closure of the Medaka, Oryzias latipes
Authors: KAGEYAMA TETSUO
Published: January 2005

The embryonic surface of the teleost, Oryzias latipes, was observed by scanning electron microscopy (SEM) to examine the last phase of epiboly or blastopore closure. The surface of the external yolk syncytial layer (E–YSL), a surface cytoplasmic layer encompassing the yolk sphere situated beyond the blastoderm, was highly undulated with surface folds of random orientation throughout most of epiboly (st. 14–20). Scattered microvilli were observed on the surface of the margin of the yolk plug in st. 18–20. The microvilli, 1 to 6 μm in length, were projected in a bunch at the end of blastopore closure (st. 20–21). The appearance of these microvilli in the last phase of epiboly is discussed with respect to the mechanism of epiboly.
The Effects of Cyclophosphamide on Alkaline Phosphatase Activity and on in vitro Post‐Implantation Murine Blastocyst Development†
Authors: KOLA ISMAIL , FOLB PETER I.
Published: January 2005

Cyclophosphamide, administered in doses of 20 and 40 mg/kg body weight to pregnant inbred CBA mice on the third day of gestation (60 h after the estimated time of ovulation) reduced the activity of non‐specific alkaline phosphatase (E.C. 3.1.3.1.) in 84 h‐old blastocysts in a dose‐related fashion compared with controls (p < 0.02 in each case; comparison of groups by Kruskal Wallis analysis of variance). This effect was not demonstrated with 4 mg/kg body weight (p < 0.1). Forty mg/kg body weight, but not the lower doses of cyclophosphamide, significantly retarded the hatching of embryos from the zona pellucida, attachment to the culture dish, and the formation of trophoblast outgrowths when the blastocysts were subsequently cultured in vitro for 120 h. The growth of an expanded inner cell mass was impaired in the 20 and 40 mg/kg groups. The differentiation of the inner cell mass into endoderm and ectoderm was significantly affected in the 4 mg, 20 mg and 40 mg/kg groups (p < 0.05, p < 0.01 and p < 0.001 respectively). The possible relationships of these various findings are discussed in the text. These results suggest that alkaline phosphatase may be a useful indicator of impaired growth and differentiation of the inner cell mass after exposure of preimplantation embryos to teratogens.
Presence of an Allocyclic Early Replicating X Chromosome in Female Embryos of the Rat, Chinese Hamster and Rabbit
Authors: SUGAWARA OSAMU , TAKAGI NOBUO
Published: January 2005

Previous studies on early female mouse embryos revealed the presence of two kinds of inactive X chromosomes, one replicating late and the other early in the DNA synthetic period. The X chromosome that replicates early is of special interest because of its paternal origin, preferential occurrence in trophectoderm and primitive endoderm derivatives, and programmed shift to the late replicator. This study by BrdU labeling and acridine orange fluorescence staining was undertaken to examine whether the inactive X chromosome behaves in a similar manner in other laboratory mammals. In rat embryos the paternal X chromosome was found to show the same behavior in extraembryonic tissues. Early replicating chromosomes were also found in the extraembryonic regions of Chinese hamster and rabbit embryos, although their parental origin could not be determined due to the absent of X chromosome polymorphism in these species. Probably the early replicating X chromosome occurs commonly in mammals. Its functional significance is unknown.
The Role of Cell Adhesion in the Differentiation of Mesendodermal Tissues in the Starfish, Asterina pectinifera
Authors: KOMINAMI TETSUYA
Published: January 2005

The role of cell‐to‐cell adhesion in the early embryogenesis of the starfish Asterina pectinifera was studied by using concanvalin A (ConA), an agent known to weaken cellular contact by binding to glycosides at the cell surface. The major change in morphology was a diminution in the volume of the endodermal tissues (the digestive tract) of the treated larvae. It was found by pulse treatment that this effect of ConA was stage‐specific, and that the effective period corresponded to the stage when blastomeres become more cohesive. The number of cells in the mesodermal tissues, however, was relatively constant while the volume of the endodermal tissue varied considerably. It was suggested that cell‐to‐cell adhesion during this stage is involved in the allocation of endodermal tissues. In contrast, mechanisms other than cell adhesion were considered to be important for the differentiation of the mesodermal tissues.
Hormonal Control of Gametogenesis in the Terrestrial Snail, Euhadra peliomphala
Authors: TAKAYANAGI HIROSHI , TAKEDA NAOKUNI
Published: January 2005

Effects of the cerebral ganglion (brain) and the optic gland on oogenesis and spermatogenesis were studied in the terrestrial snail, Euhadra peliomphula. Removal of the optic tentacles inhibited both oogenesis and spermatogenesis. These effects were abolished by the injection of the optic tentacle extract. In the tissue of the optic tentacle, only the optic gland showed the recovery action. Furthermore, in vitro the optic gland extract stimulated spermatogenesis but had no effect on oogenesis. On the contrary, the brain extract promoted oogenesis not only in vitro and but also in viva The hermaphroditic gland extract reduced the nuclear volume of the optic gland cells, but the brain extract did not. In addition, among sex steroid hormones secreted from the hermaphroditic gland, only testosterone also reduced the nuclear volume of optic gland cells. On the basis of these results, hormonal mechanisms of gametogenesis in Euhadra peliomphala are discussed.
Induction and Inhibition of Meiotic Maturation in Follicle‐enclosed Mouse Oocytes by Forskolin
Authors: HASHIMOTO NAOHIRO , KISHIMOTO TAKEO , NAGAHAMA YOSHITAKA
Published: January 2005

A continuous exposure of follicle‐enclosed mouse oocytes to ovine luteinizing hormone (LH, 10 μg/ml) in vitro resulted in a 3‐fold elevation of CAMP levels in the follicle cells, but not the oocytes, with subsequent oocyte maturation. When follicle‐enclosed oocytes were exposed to forskolin (0.01–10 μM) for 2 hr and then incubated in forskolin‐free medium (transient exposure group), oocytes underwent germinal vesicle breakdown in a dose‐dependent manner. In contrast, a continuous exposure of the follicles to forskolin (10 μM) for up to 10 hr failed to induce resumption of meiosis. Follicle cell cAMP levels increased within 2 hr after the initial exposure to forskolin, and thereafter decreased rapidly regardless of whether forskolin treatment was transient or continuous. A similar transient increase in oocyte cAMP levels was observed after transient or continuous treatment with forskolin. It was evident, however, that at any time examined oocyte cAMP levels were consistently higher in the continuous exposure group than in the transient exposure group. Furthermore, a continuous exposure to forskolin also blocked LH‐induced meiotic maturation. These findings suggest that elevated levels of cAMP in the oocyte block meiotic maturation in mouse oocytes. The present results further suggest that an increase in follicle cell cAMP levels is essential to the LH‐induced meiotic maturation.
Changes during Development in the Activities and Intracellular Locations of Glycerol Kinase and Hexokinase in Rat Brain and Liver
Authors: KANEKO MUNEKIYO , ISHIBASHI SADAHIKO
Published: January 2005

The changes in activities and intracellular locations of glycerol kinase in rat brain and liver during development were compared. Glycerol kinase activity was consistently much higher in the liver than in the brain from just before the birth to the adult stage. Most of the activity was bound to mitochondria in the brain, but the intracellular distribution of the activity in the liver changed during development, the amount of activity bound to mitochondria being high just after birth and then decreasing gradually. These changes of glycerol kinase during development were compared with those of hexokinase, and the significance of the changes in the two enzymes is discussed in relation to dietary changes during development.
Inhibitory Effects of Diltiazem and Verapamil, Calcium Antagonists, on Fertilization‐Induced Increase in the Phosphorylase Reaction in Echiuroid Eggs
Authors: TAZAWA EIGORO , FUJIWARA AKIKO , KOMURAKI MASAYUKI , FUJINO YUKIO , YASUMASU IKUO
Published: January 2005

The calcium antagonists diltiazem and verapamil at 100 μM caused considerable inhibition of the glycolysis system in recently fertilized eggs of the echiuroid, Urechis unicinctus. The levels of glycolytic intermediates in eggs were found to be higher 5 min after insemination than before fertilization while the levels of adenine nucleotides and inorganic phosphate were almost the same before and after fertilization. Addition of diltiazem or verapamil 30 sec after insemination did not inhibit fertilization, but resulted in maintenance of as low levels of glycolytic intermediates as in unfertilized eggs. The apparent mass action ratio in the phosphorylase step, calculated from the levles of glucose‐1‐phosphate and inorganic phosphate was normally higher in fertilized eggs than in unfertilized eggs, but was maintained at as low a level as in unfertilized eggs by adding these compounds 30 sec after insemination. Phosphorylase a activity also normally increased after insemination, but was maintained at a low level in fertilized eggs by adding these compounds. These compounds also inhibited the increased 45Ca2+ uptake normally observed after fertilization. These results suggest that after fertilization, the Ca2+ level increases associated with fertilization‐induced Ca2+ influx and that this stimulates Ca2+ dependent protein kinase to phosphorylate phosphorylase b, resulting in an increased rate of the phosphorylase reaction.
Distribution of Intramembrane Particles and Its Changes during the Acrosome Reaction in Spermatozoa of the Japanese Abalone
Authors: SAKAI YOSHI T. , SUZUKI FUMIE , SHIROYA YOKO
Published: January 2005

The distribution of intramembrane particles in the plasma and acrosomal membranes of sperm of the Japanese abalone, Haliotis discus, and its changes during the acrosome reaction were studied by the freeze‐fracture replica technique. The P face of the plasma membrane covering the acrosome has sparse membrane particles except in the apical region, which includes the trigger and ‘truncated cone’ regions. Large particles with an average diameter of 10 nm are located in this apical region. The E face of the plasma membrane has only a few particles. On the outer acrosomal membrane, many particles are randomly distributed throughout the P face, but only a small number of particles are found on the E face. Numerous particles on the P face of the inner acrosomal membrane show a regular arrangement as a dense lattice or with a concentric circular pattern. The initial change in the acrosome reaction is clearance of membrane particles from both the P and E faces of the plasma and outer acrosomal membranes around the apical region, where fusion of the two membranes occurs. As the acrosomal process elongates, the dense arrangement of particles on the inner acrosomal membrane changes via a loose lattice arrangement to a patchy distribution with particle‐free areas. Then the arrangement is further disorganized becoming a sparse, random distribution.
Developmental Changes of Cell Process Meshwork and Extracellular Space in Subepidermal Mesenchyme in Mouse Forelimb Buds
Authors: DATU ABDUL RAZAK
Published: January 2005

This study was designed to reveal morphological changes in epithelial‐mesenchymal interface during limb development. An electron microscopic and morphometric analysis was done on the cell process meshwork (CPM) and subepidermal extracellular space (SEECS) in mesenchyme in mouse forelimb buds at embryonic ages from day 9.5 to 12.5 (vaginal plug=day 0). At days 9.5 and 10.0, when the apical ectodermal ridge (AER) had not yet appeared, no spatial differences were noted in the CPM density and SEECS width. However, at days 10.5, 11.0 and 11.5, when the AER was distinct, the SEECS was wide and the number of mesenchymal cell processes was small beneath the AER as compared with those at days 9.5 and 10.0; they differed significantly from those beneath the dorsal or ventral non‐ridge epidermis. Between days 10.0 and 10.5, the SEECS width decreased and the number of cell processes increased in the dorsal region. At days 12.0 and 12.5, when the AER was regressing, the spatial differences in the CPM density and the SEECS width became less obvious. These findings indicate that spatial and temporal differences of the SEECS width and CPM density exist in mouse limb buds, and these differences are closely associated with the AER development.
Transdifferentiation in Animal Cells: Fact or Artifact?
Authors: OKADA T. S.
Published: January 2005

Some specialized animal cells can overtly change from one differentiated phenotype to another through the process of transdifferentiation. In this review article, representative and convincing examples of transdifferentiation in cells of various animals ranging from mammals to coelenterates are presented. Several basic nature of the cellular properties that underly the transdifferentiation process is discussed. Studies are in progress to understand the mechanism of transdifferentiation in terms of gene expression.
Presence of a Ciliary Patch in Preoral Epithelium of Sea Urchin Plutei
Authors: NAKAJIMA YOKO
Published: January 2005

Removal of the hyaline layer from sea urchin embryos at the pluteus stage discloses a densely ciliated region in the preoral area of the ectodermal epithelium. In four‐armed plutei, this ciliary path is located between the anterolateral arms and in eight‐armed plutei it becomes surrounded by preoral and anterolateral arms. The area of the patch and the number of cilia increase with age. This patch is covered by cilia of unusual morphology and orientation. There are more than two cilia per cell which are coiled together several times around a small cone at the apical end of the cell. These coiled cilia run parallel to the surface of the cell but do not extend beyond the hyaline layer. The ciliary axoneme consists of a “9+2” microtubular structure, but no outer or inner dynein arms are observed. Although the cells with coiled cilia are present in a cluster constituting a part of the epithelium, they have axons that project from their basal (inner) ends. The structural characteistics of the ciliary patch suggest that it possesses a sensory function.
Cell to Cell Adhesion Systems in Xenopus laevis, the South African Clawed Frog I. Detection of Ca2+ Dependent and Independent Adhesion Systems in Adult and Embryonic Cells
Authors: NOMURA K. , UCHIDA M. , KAGEURA H. , SHIOKAWA K. , YAMANA K.
Published: January 2005

The reaggregation kinetics of frog (Xenopus laevis) cells prepared using several different dissociation procedures were monitored. Two distinct modes of cell adhesion were revealed, one mediated by Ca2+ dependent cell‐cell adhesion system (CDS) and the other mediated by Ca2+ independent one (CIDS).

CDS was detected in frog embryonic (two‐cell stage to neurula) cells and in cells of epithelial cell lines (A6 and A8), while CIDS was detected only in A6 cells. Frog CDS was resistant to 0.01% tryptic digestion in the presence of 1 mM Ca2+ and CIDS was resistant to dilute (0.0001%) tryptic digestion. Cells with both mechanisms were prepared by dissociation with 1 mM EDTA and both mechanisms were absent in cells dissociated with 0.01% trypsin and 1 mM EDTA. Frog CDS and CIDS functioned in a temperature dependent and independent manner, respectively. Properties of frog CDS and CIDS revealed in this study were the same as those of mammalian or avian CDS and CIDS, respectively. Frog cells with CDS cross‐adhered to mammalian epithelial (F9) cells but not to fibroblastic (V79) cells Ca2+ dependently. Monoclonal antibody ECCD‐1 raised against mouse epithelial CDS blocked aggregation of frog A8 cells.

These results suggest the similarity between frog CDS and mammalian epithelial type CDS.
Somatic Embryogenesis in Cultured Carrot Cells
Authors: NOMURA KOJI , KOMAMINE ATSUSHI
Published: January 2005

Somatic embryogenesis in cultured plant cells is an ideal system for investigating the whole process of differentiation and development from single cells to whole plants, and especially the molecular mechanism of expression of totipotency. This review reports recent progress the studies on somatic embryogenesis.
Prospective Neural Areas and Their Morphogenetic Movements during Neural Plate Formation of Xenopus Embryos. I. Development of Vegetal Half Embryos and Chimera Embryos
Authors: SUZUKI AKIO S. , HARADA KEIICHI
Published: January 2005

Development of animal cap‐less Xenopus gastrulae was examined. In vegetal halves from which the animal cap was removed 0.6 mm above the blastopore, an apparently normal array of craniocaudal structures developed. Histological examination showed differentiation of central nervous system (CNS) structures in the cap‐less embryos, but differentiation of sensory organs, such as a lens and ear vesicle in only a few embryos. Only the dorsal midline of the embryos was covered with epidermis, and its lateral‐ventral areas consisted of bare endoderm and mesoderm. The development of animal cap was also investigated by exchanging the animal cap of X. laevis embryos with that of X. borealis embryos, which can be distinguished by quinacrine fluorescence staining. The central nervous system of chimera embryos consisted mainly of X. laevis cells stained homogeneously with quinacrine but a small number of punctately‐stained X. borealis cells was in the anterior tip of the forebrain. Cells of the lens and ear vesicle were punctately stained. More than two‐thirds of the epidermal area consisted of punctately‐stained cells and only the dorsal midline of the posterior head‐ and trunk‐epidermis consisted of homogeneously‐stained cells.

Areas of the prospective central nervous system and their movement during embryogenesis of Xenopus are discussed.
Modification of Dorsal‐Ventral Polarity in Xenopus laevis Embryos Following Withdrawal of Egg Contents before First Cleavage
Authors: WAKAHARA MASAMI
Published: January 2005

When fertilized Xenopus laevis eggs were pricked just beneath the marginal zone with a thick glass needle prior to the first cleavage, a small amount of cytoplasm escaped into the exudate. Those eggs were placed in a poly L‐lysine‐coated plastic dish filled with 10% Ficoll solution. The location of the sperm entrance site (SES) of each egg was marked by scratching the surface of the plastic dish. The pricked embryos were anchored to the dish through poly L‐lysine, and developed, therefore, without changing their original position. Consequently, development of the dorsalventral polarity was conveniently monitored with respect to the location of the SES. Embryos which developed from eggs pricked on the side opposite the SES showed modification of the dorsal‐ventral polarity: Semi‐quantitative studies showed that an exudation approximately 1.5–12.5% of the whole egg contents from the presumptive dorsal side caused a reversal of the dorsal‐ventral polarity. That is, the dorsal lip of the blastopore formed on the same side of the SES, whereas the dorsal lip formed on the side opposite the SES in the normal control and sham‐operated embryos. Half of the embryos which had larger cytoplasmic exudates more than 12.5% of the whole egg contents failed to form the dorsal lip by the time all controls and the embryos with smaller exudates showed normal dorsal lip formation. When eggs were pricked on the SES side, the normal topographic relationship between the SES and future dorsal lip side was reinforced.
Shift in Steroidogenesis in the Ovarian Follicles of the Goldfish (Carassius auratus) during Gonadotropin‐Induced Oocyte Maturation
Authors: NAGAHAMA YOSHITAKA , GOETZ FREDERICK W. , TAN JOSEFA D.
Published: January 2005

Both partially purified chum salmon gonadotropin and 17α‐hydroxyprogesterone stimulated in vitro production of testosterone by postvitellogenic follicles of goldfish (Carassius auratus). Chum salmon gonadotropin further enhanced the conversion of exogenously supplied 17α‐hydroxyprogesterone to 17α, 20β‐dihydroxy‐4‐pregnen‐3‐one. The increased medium concentrations of 17α, 20β‐dihydroxy‐4‐pregnen‐3‐one were associated with the induction of final oocyte maturation.

The capacity of postvitellogenic follicles to produce steroids in response to exogenous 17α‐hydroxyprogesterone was examined in females at various stages of final oocyte maturation following the administration of human chorionic gonadotropin in vivo combined with elevation of holding temperature. The maximum production of testosterone in response to 17α‐hydroxyprogesterone was obtained in follicles from initial controls. In contrast, 17α 20β‐diOHprog production was very low in initial controls and markedly increased during oocyte maturation (3–6 hr following injection), followed by a significant decrease in follicles collected at 15 hr. Estradiol‐17β production by the follicles was very low at any stages of gonadotropin‐induced oocyte maturation. These results suggest that gonadotropin‐induced shift in the biosynthetic pathway in the follicle from the secretion of predominantly testosterone to 17α, 20β‐dihydroxy‐4‐pregnen‐3‐one secretion is a prerequisite step for the induction of oocyte maturation in goldfish.
Direct Evidence for the Presence of Germ Cell Determinant in Vegetal Pole Cytoplasm of Xenopus laevis and in a Subcellular Fraction of It
Authors: IKENISHI KOHJI , NAKAZATO SAKIKO , OKUDA TAKASHI
Published: January 2005

To test for the presence of germ cell determinant in Xenopus embryos, vegetal pole cytoplasm containing the “germ plasm”, or a subcellular fraction of it, was microinjected into single somatic blastomeres isolated from 32‐cell embryos. Injected or non‐injected (control) blastomeres were cultured in 3H‐thymidine until normal control embryos reached the neurula stage. The labeled explants were then implanted into unlabeled host neurulae, which were allowed to develop to the tadpole stage. Labeled PGCs of explant origin in the genital ridges of the experimental tadpoles were examined by autoradiography.

Isolated blastomeres were injected with vegetal pole cytoplasm of 32‐cell embryos or with a 20,000 g pellet made from vegetal pole cytoplasm of 2‐cell embryos. Labeled PGCs were found in 7.6% and 2.3% of the experimental tadpoles, respectively. No labeled PGCs were found in the control tadpoles, except for one tadpole in the first experiment. These results strongly suggest that the vegetal pole cytoplasm and its subcellular fractions act as germ cell determinant.
Elongated Microvilli on Vegetal Pole Cells in Sea Urchin Embryos
Authors: AMEMIYA SHONAN
Published: January 2005

The ultrastructure of cells in the vegetal pole region of sea urchin embryos during early development to the mesenchyme blastula stage was examined by scanning electron microscopy. Vegetal pole cells in the ectoderm with longer microvilli than those of neighboring cells were first detectable at the early blastula stage just before hatching. These cells with elongated microvilli remained in the central region of the vegetal plate when most vegetal plate cells ingressed into the blastocoel to form primary mesenchyme. When first detectable in the sea urchin, Anthocidaris crassispina, four vegetal pole cells had elongated microvilli, but at the time of primary mesenchyme cell ingression, the number of cells with elongated microvilli had increased to eight, apparently by cell division. These vegetal pole cells were wedge‐shaped with a broad surface adhering to the hyaline layer at the time of primary mesenchyme cell ingression. SEM observation of the outer surface of embryos showed that the microvilli extended into the hyaline layer. The reinforced attachment of vegetal pole cells to the hyaline layer through their elongated microvilli may explain why these cells could remain at the vegetal pole when the surrounding cells ingressed into the blastocoel as primary mesenchyme cells.
Development of Fertilized Starfish Eggs in Which Cytokinesis is Prevented by Iturin A–2
Authors: TSUCHIMORI NOBORU , MIYASHIRO SHIGEYOSHI , TSUJI TOMOKO , KIDA TAKAO , SHIBAI HIROSHIRO , IKEGAMI SUSUMU
Published: January 2005

Iturin A–2, a cyclic peptide obtained from cultures of Bacillus subtilis inhibited cell division but not nuclear divisions of fertilized eggs of the starfish Asterina pectinifera, resulting in the formation of non‐cleaving eggs containing all the embryonic nuclei in a common cytoplasm. Fertilized eggs in which cleavage was prevented by iturin A–2 (50μg/ml) synthesized DNA, RNA and protein at comparable rates to those of normal embryos up to the onset of blastulation. In single‐cell embryos, however, elevation of the level of transcription, which is a characteristic marker of blastulation, did not take place and the chromatin masses eventually dispersed in the cytoplasm.

Treatment of oocytes with iturin A–2 (50μg/ml) resulted in loss of their response to the maturation‐inducing substance 1‐methyladenine, whose receptors are located in the cell membrane or other components of the oocyte cortex. Furthermore, iturin A–2 bound to oocyte cortices quantitatively, suggesting that its site of action is the cortices of oocytes and eggs, and that its arrest of cleavage of fertilized eggs is due to loss of the ability of the cell membrane to form a cleavage furrow.
Heat Shock May Relieve a Posttranscriptional Block on σ Crystallin Synthesis in Cultures of Chick Embryo Neuroretinal Cells
Authors: DE POMERAI D. I. , CARR A.
Published: January 2005

The lens protein δ crystallin is expressed at high levels in lentoids fromed by the transdifferentiation of chick embryo neuroretinal cells in long‐term culture under permissive conditions. Previous work has shown that one non‐permissive medium (F199) allows transcription of δ crystallin RNA in late neural retinal cultures, but that these δ transcripts remain confined to the nuclei and are neither translated nor indeed detectable in the cytoplasm. We show here that F199 cultures maintained at 43°c throughout transdifferentiate extensively into lentoids, in contrast to similar cultures at 37°c. Following transfer of late (30 day) F199 cultures from 37°c to 43°c, δ crystallin synthesis becomes increasingly prominent over a period of at least 5 days, suggesting that recovery from heat shock (rather than heat shock per se) may be important in this phenomenon. A similar response can be elicited by chemical agents (sodium arsenite, azetidine 2‐carboxylic acid) which induce heat‐shock (stress) proteins at 37°c. Treatment of late F199 cultures with actinomycin D does not prevent this increase in δ crystallin synthesis, provided the drug is not added until after the initiation of a heat‐shock response (6 hr after temperature shift). This suggests that the observed increase in δ synthesis following heat shock involves primarily the export and/or stabilisation of δ mRNAs already present in the nuclei in late F199 cultures. Possible wider implications of this posttranscriptional level of control over δ crystallin synthesis are discussed.
Probable Role of Allylisothiocyanate‐Sensitive H+, K+‐ATPase in Spicule Calcification in Embryos of the Sea Urchin, Hemicentrotus pulcherrimus
Authors: MITSUNAGA KEIKO , FUJINO YUKIO , YASUMASU IKUO
Published: January 2005

In embryos of the sea urchin, Hemicentrotus pulcherrimus, as well as in cultured cells derived from isolated micromeres, spicule formation was inhibited by allylisothiocyanate, an inhibitor of H+, K+‐ATPase, at above 0.5 μM and was almost completely blocked at above 10 μM. Amiloride, an inhibitor of Na+, H+ antiporter, at above 100 μM exerted only slight inhibitory effect, if any, on spicule formation. Intravesicular acidification, determined using [dimethylamine‐14C]‐aminopyrine as a pH probe, was observed in the presence of ATP and 200 mM KCl in microsome fraction obtained from embryos at the post gastrula stage, at which embryos underwent spicule calcification. Intravesicular acidification and K+‐dependent ATPase activity were almost completely inhibited by allylisothiocyanate at 10 μM. Allylisothiocyanate‐sensitive ATPase activity was found mainly in the mesenchyme cells with spicules isolated from prisms. H+, K+‐ATPase, an H+ pump, probably mediates H+ release to accelerate CaCO3 deposition from Ca2+, CO2 and H2O in the primary mesenchyme cells. Intravesicular acidification was stimulated by valinomycin at the late gastrula and the prism stages but not at the pluteus stage. K+ permeability probably increases after the prism stage to activate H+ release.
Changes in the Activity of the Maturation‐Promoting Factor Are Correlated with Those of a Major Cyclic AMP and Calcium‐Independent Protein Kinase During the First Mitotic Cell Cycles in the Early Starfish Embryo
Authors: PICARD ANDRÉ , LABBÉ JEAN‐CLAUDE , PEAUCELLIER GÉRARD , LE BOUFFANT FRANÇOISE , LE PEUCH CHRISTIAN , DOREE MARCEL
Published: January 2005

Many studies suggest that MPF activation depends on protein phosphorylation or that MPF is itself a protein kinase. In the present report, cyclic variations of MPF activity have been correlated in vivo with changes in the extent of protein phosphorylation or in vitro with changes of a major protein kinase during the first cell cycles of fertilized starfish eggs. This cycling protein kinase neither requires cAMP nor Ca2+. Neither colchicine nor aphidicoline, which inhibits cleavage and chromosome replication respectively, was found to suppress the synchronous and cyclic variations of both MPF and protein kinase activities. Protein synthesis was found to be required for both MPF and protein kinase activities to reappear after their simultaneous drop at the time of mitotic or meiotic cleavages. Production of either MPF or protein kinase activities is not the immediate result of protein synthesis since there is a delay at each cell cycle between the time when protein synthesis is required and the time when both MPF and protein kinase activities are produced. This suggests that both MPF and protein kinase activities might involve some post‐translational modification of a precursor protein synthesized during the preceeding cell cycle.
Changes in the Level of Histone H4 mRNA in Xenopus leavies Embryogenesis.
Authors: SHIOKAWA KOICHIRO , ATSUCHI YASUO , WOODLAND HUGH R.
Published: January 2005

In Xenopus laevis, the change in the amount of histone H4 mRNA per embryo measured by Northern blotting methods follows a unique change during early embryogenesis: It starts to increase first at the blastula stage, doubles by the gastrula stage then decreases considerably at the neurula stage, and then increases again from the tailbud stage on. The present paper establishes these developmental changes, and furthermore, provides evidence that the synthesis of H4 mRNA starts or at least increases to a detectable level at the midblastula stage as shown by S–1 protection analysis of the expression of paternal histone H4 genes in X. borealis (♀) and X. laevis (♂) hybrid embryos.
Inhibitory Effect of Omeprazole, a Specific Inhibitor of H+, K+‐ATPase, on Spicule Formation in Sea Urchin Embryos and in Cultured Micromere‐Derived Cells.
Authors: FUJINO YUKIO , MITSUNAGA KEIKO , YASUMASU IKUO
Published: January 2005

Embryos kept with omeprazole, a specific H+, K+‐ATPase inhibitor, in a period of development between the mesenchyme blastula and the pluteus corresponding stage became abnormal plutei having quite small spicules, somewhat poor pluteus arms and apparently normal archenterons. In micro‐mere‐derived cells, kept with omeprazole at pH 8.2 in a period between 15 and 40 hr of culture at 20°C, omeprazole strongly inhibited spicule formation but did not block the outgrowth of pseudopodial cables, in which spicule rods were to be formed. These indicate that omeprazole probably exerts no obvious inhibitory effects other than spicule rods formation. Omeprazole‐sensitive H+, K+‐ATPase, an H+pump, seems to be indispensable for CaCO3 deposition (formation of spicule rod) in these spicule forming cells. H+, produced in overall reaction for CaCO3 formation: Ca2++ CO2+H2O°CaCO3+2H+, is probably released from the cells by this H+pump and hence, this reaction tends to go to CaCO3 production to form spicule rods. Omeprazole, known to become effective following its conversion to a specific inhibitor of H+, K+‐ATPase at acidic pH, is able to inhibit formation of spicule rod at alkaline pH in sea water. This is probably due to an acidification of sea water near the cell surface by H+ejection in H+, K+‐ATPase reaction.
Development of Sea Urchin Embryos in Artificial Sea Water Containing Br−in Place of Cl−
Authors: FUJINO YUKIO , MITSUNAGA KEIKO , YASUMASU IKUO
Published: January 2005

In artificial sea water in which the Cl−concentration was reduced to less than 10% of that in normal sea water by its replacement with Br−, sea urchin eggs were fertilized and developed into abnormal plutei following almost the same time schedule as in natural sea water. These embryos had poorly developed spicules, short pluteus arms, somewhat jagged embryo‐walls and quasi‐normal archenterons. Similar embryos were obtained in another artificial sea water in which 90% of the Cl−concentration in normal sea water was reduced by Br−and 10% by acetate. In artificial sea water, in which either 90% of the Cl−was replaced by Br−or 10% was replaced by acetate, embryos developed into plutei with quasi‐normal spicules, pluteus arms and archenterons. These findings indicate that deficiency of Cl−results in somewhat abnormal sea urchin embryos. When cells derived from isolated micromeres, were cultured in these Cl−‐deficient artificial sea waters, containing Br−in place of more than 70% of the normal Cl−concentration in sea water, spicule formation was strongly inhibited, but pseudopodial cables were well developed. Thus, external Cl−seems to be necessary for at least normal formation of spicule rods.
In Vitro Induction of Starfish Oocyte Maturation by Cysteine Alkylesters
Authors: MITA MASATOSHI , UETA NOBUO , NAGAHAMA YOSHITAKA
Published: January 2005

The effect of various disulfide‐reducing agents including cysteine and its alkylesters on the induction of germinal vesicle breakdown (GVBD) in starfish (Asterina pectinifera) oocytes was investigated in vitro. Although cysteine did not induce GVBD, its alkylesters were effective. Cysteine alkylesters significantly mimicked the effect of 1‐methyladenine (1‐MeAde), the naturally occurring maturation‐inducing hormone of starfish, on oocyte maturation. However, the effective concentrations and pH optimum for stimulation of oocyte maturation varied between 1‐MeAde and the cysteine alkylesters. By comparing pKa values of the disulfide‐reducing agents to pH of the medium, it is suggested that the redox potential of a disulfide‐reducing agent is an important indicator its ability to induce oocyte maturation.

With the use of fluorescent probes for thiol groups, it was shown that the fluorescence in oocyte cortices increased within 5 min after administration of 1‐MeAde. The fluorescence intensity in the cortices also increased after treatment with cysteine and its alkylesters, although the intensity was much stronger with the latter. Furthermore, both 1‐MeAde and the disulfide‐reducing agents were suggested to cause reduction of thiol groups within the plasma membrane as opposed to those on the external and internal surfaces. Thus, it is suggested that disulfide‐reducing agents and 1‐MeAde induce starfish oocyte maturation by changing the redox state of the thiol groups located within the oocyte plasma membrane.