Maturation Division Research Articles

Ultrastructure of germ cells, the Leydig cells, and Sertoli cells during spermatogenesis in Boleophthalmus pectinirostris (Teleostei, Perciformes, Gobiidae)

The ultrastructures of germ cells, Leydig cells, and Sertoli cells during spermatogenesis in male Boleophthalmus pectinirostris were investigated by electron microscopic observations. During the period of maturation divisions, well-developed Leydig cells have three major morphological characteristics: a vesicular nucleus, mitochondria with tubular cristae, and a number of smooth endoplasmic reticulum. Based on cytoplasmic features, it appears that Leydig cells are responsible for the synthesis of male sex steroids. Although no clear evidence of steroidogenesis was found in the Sertoli cells, they were found to perform a phagocytic function in the seminiferous lobules. Most Sertoli cells contain granules thought to represent deposited glycogen or lipid but there is no indication of a transfer of nutrients to the spermatids. During the period of germ cell degeneration, several characteristics of phagocytosis appear in the cytoplasm of the Sertoli cells. In particular, it is assumed that the Sertoli cells are involved in the degeneration and resorption of undischarged spermatids after spermiation. No acrosome of the sperm is formed. The structure of the spermatozoon in B. pectinirostris is very similar and closely resembles to those of suborder Gobioidei (perciform type teleosts). The flagellum or sperm tail shows the typical 9 + 2 array of microtubules.

New data on the cytology of parthenogenetic weevils (Coleoptera, Curculionidae)

Parthenogenesis and, in particular, polyploidy are rare in animals. A number of cases, known among weevils, represent apomictic parthenogenesis--a reproductive mode in which eggs undergo one maturation division, the chromosomes divide equationally, and no reduction takes place. Among parthenogenetic weevils there are two diploids, 48 triploids, 18 tetraploids, six pentaploids, three hexaploids and one decaploid. Eight examined parthenogenetic species are triploids with 33 chromosomes of different morphology, confirming that triploidy is the most common level of ploidy in weevils. The karyotypes are heterogeneous with the presence of meta-, submeta-, subtelo- and acrocentric chromosomes. The C-banding method showed that only two species possess a large amount of heterochromatin visible as a band around the centromere during mitotic metaphase. This agrees with observations that weevils are characterized by a small amount of heterochromatin, undetectable in metaphase plates after C-banding. In three species an atypical course of apomictic oogenesis occurs with stages similar to meiosis, in which chromosomes form bivalents and multivalent clusters. This association of chromosomes probably represents the remnants of meiosis, although these events have nothing to do with recombination. The results support the hypothesis that the evolution of apomictic parthenogenesis in weevils has proceeded through a stage of automixis.

Experimental induction of gonadal maturation and spawning in the giant jellyfish Nemopilema nomurai (Scyphozoa: Rhizostomeae)

We studied the processes of gonadal maturation, spawning, fertilization and embryonic development of the giant jellyfish Nemopilema nomurai, which has been blooming in recent years in the Sea of Japan. Healthy medusae actively swimming offshore had immature gonads, but damaged and ashore-drifted animals had relatively mature gonads. The animals maintained in a small net on the nearshore to mimic the drifted condition showed induced gonadal maturation by the increase in oocyte diameter and darkness in the matrix. A similar maturation process also occurred in isolated pieces of ovary incubated at the same temperature. Fully grown oocytes that probably stopped at the prophase of the first meiosis reinitiated maturation divisions in response to light exposure, and massive spawnings occurred 80–100 min after the exposure. The spawned oocytes were first released within the subgenital sinus, then transported peripherally and finally shed into the gastrovascular cavity. Maturation and spawning in male gonads were similar to those of females except that the male spawning occurred within 30 min after the light exposure and always preceded the female spawning. Hence, it was suggested that fertilization might take place in the female gastrovascular cavity by the sperm that came in from surrounding seawater by animals’ pumping activities after dawn. Having mechanical damages might promote fertilization success of this species in the Sea of Japan.

Induction of mictic females in the rotiferBrachionus: oocytes of amictic females respond individually to population‐density signal only during oogenesis shortly before oviposition

Summary1. One at a time during the reproductive period of amictic females, oocytes fill with yolk and undergo a mitotic maturation division (oogenesis), are oviposited as single cells, and then develop parthenogenetically into females. Sexual reproduction inBrachionusand several other genera is initiated when amictic females are crowded and oviposit some eggs induced to differentiate into mictic females. Mictic females produce haploid eggs that can develop parthenogentically into males or be fertilised and develop into diapausing embryos called resting eggs.2. This study examines the time when oocytes in amictic females respond to maternal population density. Is the fate of all oocytes in the germarium irreversibly determined during the early postnatal life of the mother, or is each oocyte labile until just before oviposition? In the former case, the probability of an amictic female producing a mictic daughter at any time throughout her reproductive period would reflect the population density she experienced while young and not that at the time she oviposited an egg.3. Amictic females of two clones of a Florida strain ofB. calycifloruswere cultured singly from birth at a low or high density (in a large or small volume) until about halfway through their reproductive period and then switched (experimental treatment), or not (control treatment), to the other density condition. The results indicate that the female fate of an oocyte is determined by maternal population density during oogenesis. Eggs oviposited soon after transfer from low to high density had the same, or a higher, probability of becoming mictic females compared with those produced by control females kept at the high density; eggs oviposited after transfer from the high to the low density had the same low probability of becoming mictic females as those produced by control females kept at the low density.4. Control females kept at the high density were less likely to produce mictic daughters as they aged. This decline is not because of a decreased propensity of older females to respond to crowding, as older females responded maximally when transferred from a low to a high population density.5. As oocytes in amictic females respond to maternal population density only during oogenesis, there is a negligible lag between the population‐density signal in the environment and the commitment to sexual reproduction. This minimises the obligatory two‐generation lag between this signal and production of resting eggs, and thus reduces the possibility that crowding will lead to food limitation before production of these eggs.

Germ-plasm specification and germline development in the parthenogenetic pea aphid Acyrthosiphon pisum: Vasa and Nanos as markers

The germarium, oocytes and embryos of the parthenogenetic viviparous pea aphid Acyrthosiphon pisum are contained within a single ovariole. This species provides an excellent model for studying how maternally-inherited germ plasm is specified and how it is transferred to primordial germ cells. Previous studies have shown that germ cells are first segregated at the embryonic posterior after formation of the blastoderm. We used two cross-reacting antibodies against the conserved germline markers Vasa and Nanos, which specifically identified these presumptive germ cells, to investigate whether germ cells were determined during early development. We observed randomly-distributed weak expression of Vasa signals in the developing oocyte but no localization in the oocyte segregated from the germarium. Localized Vasa was not apparent until it was detected at the posterior in the embryo undergoing the second nuclear division. Nanos, on the other hand, was localized to a nuage-like structure surrounding the nucleus in the developing and segregated oocytes. At the beginning of the oocyte maturation division, Nanos localization shifted to the posterior and could be identified in successive stages until it was incorporated into the germ cells. Taken together, our results suggest that germ plasm is specified in the developing oocyte and that Nanos is an earlier germline marker than Vasa. Germ cells stained for Vasa remained at a dorsal location in the egg during mid-development and then were guided into abdominal segments A1 to A6 during germ-band retraction. We infer that germ cells coalesce with segmented gonadal mesoderm during this period.

Deciphering reproductive polyphenism in aphids

Summary Polyphenism, which allows one given genotype to produce several discrete phenotypes, is an extreme case of phenotypic plasticity and is mainly found in arthropods. Social insects are the canonical example of polyphenism with the development of castes in the colonies. However, aphids display one of the largest range of polyphenisms, notably by producing winged or wingless, as well as asexual or sexual forms, depending on environmental conditions. During spring and summer, aphids reproduce by viviparous parthenogenesis, whereas in autumn they enter sexual reproduction. This switch in reproductive mode is triggered by changes in photoperiod and temperature. Here, the data accumulated since the 1960s on the identification of photoperiodic clocks, counter and putative neural photoreceptors that participate in this reproductive shift are reviewed. After perception, the photoperiodic signal is transduced through the secretion of hormones (juvenile hormones may well be involved) which, in turn, may act on the target cells, namely the oocytes. In short-day conditions, oocytes enter meiosis and produce haploid eggs which develop a 2n embryo after fertilisation. By contrast, in long-day conditions, a single maturation division produces 2n oocytes which immediately enter parthenogenetic embryogenesis. A physiological model of the determination of sexual vs. asexual reproduction in aphids is proposed and viewed from the perspective of newly initiated molecular studies.

Cytogenetics of three parthenogenetic Psocid species (Psocoptera, Psocomorpha)

A novel method was applied to study chromosomes and their behavior in female meiosis in three parthenogenetic psocid species (Psocoptera). Two of these species Valenzuela flavidus (Steph.), Caeciliusidae and Peripsocus subfasciatus (Rambur), Peripsocidae displayed a triploid chromosome number 3x=27 while the third species Trichadenotecnum majus (Kolbe) was diploid (2n=18). In all species meiosis consisted of only a single, mitosis-like maturation division, i.e. meiosis was of apomictic type. Models for the formation of polyploid parthenogenetic forms are discussed.

Polyaxial cleavage in sponges (Porifera): a new pattern of metazoan cleavage.

Cleavage is an important stage of animal embryo-genesis marked by the emergence of multicellularity ofan organism and by ooplasmic segregation of morpho-genetic determinants. In addition, cleavage is a highlycoordinated process of diagnostic significance. Cleav-age patterns are often used to investigate the phyloge-netic relationships in various animal groups [1].Analysis of early development of sponges( Porifera ), including cleavage, is of particular interestbecause of their key position at the base of the phyloge-netic tree for Metazoa. However, studies speciallyaddressing the stage of egg cleavage in sponges werevery few until the past two decades [2–5]. It is now wellknown that, in all sponge species, the whole egg dividescompletely (holoblastic cleavage). The process is usu-ally uniform, but the cleavage patterns may signifi-cantly vary from species to species. In most species,cleavage division planes are randomly oriented (chaoticcleav age). A unique type of cleavage called tablepalyntomy occurs in some species of the subclassCalcaronea [4]. In the glass sponge Oopsacas minuta (Hexactinellida), cleavage is pseudospiral [5]. In manyoviparous species of the class Demospongiae, cleavagefollows a nearly radial pattern [6].Of particular interest is analysis of cleavage insponges of the order Halisarcida, which are the mostprimitive of Demospongiae [7, 8]. The goal of thisstudy was to reexamine the cleavage period of Hal-isarca dujardini development using light and electron(scanning and transmission) microscopy. Sponges ofthis species are dioecious and viviparous. Theirembryos develop within temporary follicles. Each fol-licle consists of pinacocyte-like follicular cells and anouter collagen layer. Before maturation division, theoocyte is oval, with a round nucleus at the center. Yolkgranules are uniformly distributed throughout theoocyte.The first cleavage division in H. dujardini is uniformand proceeds in the meridional plane. The cleavage fur-rows are of the bipolar type. The subsequent cleavageproceeds asynchronously. During the second cleavagedivision, the orientation of mitotic spindles relative toeach other varies from parallel to perpendicular(Fig. 1a). Thereafter, cleavage goes on asynchronously.From the third cleavage division onward, mitotic spin-dles are always formed parallel to the surface; there-fore, the cleavage furrows run perpendicular to the sur-face (Fig. 1b). The eight-cell embryo is round orslightly oval in the cross-section and never appears as alamina-like structure described earlier [7].Cleavage leads to the formation of an apolar coelo-blastula with a small cavity inside. The cavity isbounded with long wedge-shaped cells (Fig. 1c). At thestage of approximately 100 cells, blastomeres exhibit adistinct apicobasal polarity.In our previous studies [2, 8], some primitive traitswere described in H. dujardini cleavage. These were,first of all, the absence of any determinate spatial rela-tionships among blastomeres within the embryo. Con-ceivably, the origin of these primitive traits should besought in the absence of mature egg patterning (prepat-terning) [9], which is usually regarded to be a commonprimitive trait of egg organization in sponges [10].Throughout the H. dujardini egg, its structure remainsuniform. Holoblastic uniform asynchronous cleavage istypical of all the Halisarcida species studied, such as H. dujardini, H. metschnicovi , and H. nahantensis [2, 7, 8, 11]. A distinctive feature of the cleavage stageof H. dujardini development is the long retention of theblastula-like appearance by the embryo (from the eight-cell to the prelarval stage), which is due to the early for-mation of cleavage division planes of similar type run-ning parallel to the embryo surface [2, 8]. The samecleavage pattern was described for H. nahantensis [11].This cleavage pattern is unique among the viviparousDemospongiae but quite usual in Calcinea (class Cal-carea), which form a monolayer flagellated blastula [12].At present, eight major patterns of holoblastic cleav-age are distinguished in animals [1]. Chaotic cleavageproduces no patterning in blastomere distribution. Pas-sage of all cleavage furrows along the animal–vegetal

Reproductive Maternal Centrosomes Are Cast off into Polar Bodies during Maturation Division in Starfish Oocytes

Animal egg inherits a maternal centrosome from the meiosis-II spindle and sperm can introduce another centrosome at fertilization. It has been believed that in most animals only the sperm centrosome provides the division poles for mitosis in zygotes. This uniparental (paternal) inheritance of the centrosome must depend on the loss of the maternal centrosome. In starfish, suppression of polar body (PB) extrusion is a prerequisite for induction of parthenogenesis (Washitani-Nemoto et al. (1994) Dev. Biol. 163, 293–301), suggesting that the centrosomes cast off into PBs have reproducing capacity. Due to the absence of centriole duplication in meiosis II of starfish oocytes, each centrosome of a meiosis-II spindle has only one single centriole, whereas in meiosis I each has a pair of centrioles (Sluder et al. (1989) Dev. Biol. 131, 567–579; Kato et al. (1990) Dev. Growth Differ. 32, 41–49). Hence, the first PB (PB1) has two centrioles, whereas the second PB (PB2) and the mature egg have only one centriole, respectively. The present study examined the reproductive capacity of PB centrosomes by transplanting them into artificially activated eggs, and then the recipient egg nucleus with the surrounding cytoplasm was removed. A transplanted PB2 centrosome with a single centriole formed a monopolar spindle at the first mitosis, followed by formation of a bipolar spindle in the next mitosis, leading to actual cleavage and subsequent development. This proves the reproducing capacity of the single centriole in the PB2 centrosome. The behavior of the transplanted PB1 centrosome was exactly the same as in the PB2 centrosome, in spite of the difference in the number of centrioles. These results clearly show that (1) four maternal centrioles are heterogeneous in duplicating capacity, (2) during meiosis only one centriole in each of the centrosomes of a meiosis-I spindle pole retains duplicating capacity, (3) the reproductive centrioles are successively cast off into PBs, and finally a mature egg inheriting a nonreproductive centriole alone is formed, and (4) the presence of a single reproductive centriole is sufficient condition for embryonic development in starfish.

Centriole behavior during meiosis in oocytes of the sea urchin Hemicentrotus pulcherrimus.

Ultrastructural changes in the maturing oocyte of the sea urchin Hemicentrotus pulcherrimus were observed, with special reference to the behavior of centrioles and chromosomes, using oocytes that had spontaneously started the maturation division process in vitro after dissection from ovaries. The proportion of oocytes entering the maturation process differed from batch to batch. In those eggs that accomplished the maturation division, it took ~4.5-5 h from the beginning of germinal vesicle breakdown to the formation of a second polar body. Serial sections revealed that a young oocyte before germinal vesicle breakdown had a pair of centrioles with procentrioles, located between the presumed animal pole and the germinal vesicle and accompanied by amorphous aggregates of moderately dense material and dense granules (granular aggregate). Just before germinal vesicle breakdown, a pair of fully grown centrioles located in the granular aggregate, which is present until this stage and then disappears, had already separated from another pair of centrioles. In meiosis I, each division pole had two centrioles, whereas in meiosis II each had only one. The two centrioles in the secondary oocyte separated into single units and formed the mitotic figure of meiosis II. The first polar body had two centrioles and the second had only one. The two centrioles in the first polar body did not form the mitotic figure nor did they separate at the time of meiosis II. These results indicate that, in sea urchins, duplication of the centrioles does not occur during the two successive meiotic divisions and the egg inherits only one centriole from the primary oocyte, confirming the results previously reported for starfish oocytes.

Cell Proliferation and CD11b Expression Are Controlled Independently during HL60 Cell Differentiation Initiated by 1,25α-Dihydroxyvitamin D3 or All-trans-Retinoic Acid

When 1α,25-dihydroxyvitamin D3 (D3) induces HL60 cells to differentiate to monocytes, a burst of approximately three shortened cell cycles (“maturation divisions”) precedes exit from cell cycle and completion of maturation. Here we show that similar maturation divisions occur during neutrophil differentiation induced by all-trans-retinoic acid (ATRA), but without shortening of the cell cycle. Both ATRA and D3 initiate these maturation divisions as cells pass through a “window of sensitivity” during early G1. We also investigated whether the initiation of maturation divisions and of the expression of CD11b, an early-expressed maturation marker, are linked. Cells treated with D3 or ATRA start to express CD11b after 9–14 h, before completing the first maturation division. Elutriation was used to isolate small HL60 cells (almost all in G1) and larger cells (in G1 and S phases) from unsynchronized populations. When these were cultured with D3 or ATRA, most reentered cycle synchronously, multiplied, and differentiated. Following D3 treatment, the G1-enriched small cells expressed CD11b slightly faster than unsynchronized cultures or fractions dominated by late G1 cells and/or S phase cells. D3-induced CD11b expression occurred at a similar rate even in G1 cells that were held at the G1/S boundary by thymidine. In conclusion, changes in the control of the cell cycle that characterize the onset of monocytic and neutrophil differentiation are only triggered in early G1, but CD11b expression can be initiated from most points in the cell cycle. Differentiating agents must therefore regulate the proliferation and the maturation of differentiating myeloid cells by mechanisms that are at least partly independent.

GENESIS OF THE APYRENE PARASPERM IN THE APPLE SNAIL POMACEA CANALICULATA (GASTROPODA: AMPULLARIIDAE): AN ULTRASTRUCTURAL STUDY

The apyrene paraspermatogenesis in the freshwater gastropod Pomacea canaliculata has been studied with electron microscopy. Mature apyrene parasperm result from a cytodifferentiation process without maturation division. The atypical condition is recognized early. Paraspermatogonia are characterized by voluminous nuclei with irregular clusters of heterochromatin, numerous dilated cisternae of the rough endoplasmic reticulum and small electron-dense granules spread through the cytoplasm. As this process advances, the nuclei become lobed and chromatin degenerates. The remnant chromatin condenses to form dense bodies which are finally excreted from the cells. At the cytoplasmic level a centriolar multiplication with the consequent flagellogenesis takes place. The axonemal microtubules run along the entire length of the cell and emerge from the posterior end forming a tuft of three or more free flagella. The rough endoplasmic reticulum and the Golgi complex are involved in the production of secretory granules, some of which are later released by exocytosis. At the end of paraspermatogenesis mature apyrene parasperm are fusiform-shaped anuclear ciliated cells.

Nuclear phenomena during conjugation of Heliophrya erhardi (Ciliata, Suctoria). II. The orientation of postmeiotic and metagamic division spindles is organized by the cytoskeleton

Summary Whereas the spindles of the meiotic divisions appear in a random orientation within the co-conjugants of Heliophrya erhardi, the spindles of the postmeiotic and metagamic nuclei show a distinct fixed position within the conjugational bridge and in the cytoplasm of each partner’s cell body. In postmeiotic divisions, the spindle orientation results in a different functional determination of the division products. For their later fate, mostly a concentration gradient of “cytoplasmic factors” was postulated [29]. But above all, the question arises, what is the mechanistic basis for the spindle orientation? We followed the sequential steps of conjugation by time lapse microcinematography and by EM-observation. The result of the maturation divisions are 12 to 24 haploid nuclei [11] from which only one nucleus reaching the conjugational bridge survives and starts the postmeiotic division. The position of this gonal nucleus is fixed on the left marginal side of the bridge by a dense microfilament network surrounding the nucleus in a cage-like manner and anchoring it to the epiplasmic septum separating the co-conjugants. During the following division, the spindle extends parallel to the septum fixed by cytoskeletal elements during anaphasic spindle elongation. Additionally, at metaphase, a basal body appearing at the spindle pole proximal to the pellicle nucleates microtubules (MTs) which penetrate the microfilamentous cage and contact also the nuclear envelope. The fixation of the spindle axis parallel to the separating septum by cytoskeletal elements guarantees the positioning of one pronucleus as the future stationary nucleus on the left side, and the other — the presumptive migratory nucleus — on the right side of the conjugational bridge. After exchange of the migratory nuclei and subsequent fusion with formation of the diploid synkaryon, one metagamic division follows immediately in each partner cell. From meta- to telophase the spindles exhibit in most cases a strict central-peripheral orientation, at which the anterior daughter nucleus (oriented to the cell’s center) becomes the macronucleus anlage, while the posterior nucleus condenses to the micronucleus. Interestingly, this spindle orientation is held in position by aster-like organized microtubules radiating out to a microfilamentous cap situated at the anterior spindle pole. As an organizing center for this cytoskeletal structure again a basal body is responsible, which is localized between nuclear envelope and cap region from meta- to telophase. It is evident, that in both divisions, the orientation of the spindle axis and consequently the fate of the division products depends completely on the existence of a cytoskeletal organizing structure, which is comparable to the asters of mitotic spindles in some higher eukaryotes.

Egg maturation and parthenogenetic recovery of diploidy in the scorpionLiocheles australasiae (Fabricius) (Scorpions, Ischnuridae)

In the scorpion Liocheles australasiae, egg maturation and parthenogenetic recoveries of chromosome number and nuclear DNA content were examined by histological, karyological observations and quantitative measurements of DNA. The primary oocyte becomes mature through two successive maturation divisions. The first maturation division takes place in the primary oocyte to produce a secondary oocyte and a first polar body. The second maturation division soon occurs in the secondary oocyte, in which the nucleus is divided into a mature egg nucleus and a second polar body nucleus, not followed by cytoplasmic fission. The first polar body, in one case, was successively divided into two second polar bodies; in the other case it was not divided. In either case, these polar bodies remained attached to the early embryo. The fate of these polar bodies during further embryogenesis were studied. In the karyological analysis, the chromosome number was divided into two groups, one from 27-32, the other was 54-64. The former was presumably the metaphase chromosome number at the meiotic division; the latter was presumably the metaphase chromosome number at the mitotic division. DNA content in the diploid nucleus of the primary oocyte, doubled before the maturation divisions, was reduced through the maturation divisions by one-half in the nuclei of the secondary oocyte and the first polar body and by one-fourth in the nuclei of the egg and the second polar bodies. The first reduction of DNA content corresponded to halving the number of the chromosomes in the first maturation division and the second to the nuclear division in the secondary oocyte. These reductions represent a common process of egg maturation, except the final production of the mature egg with two haploid nuclei, an egg nucleus, and a second polar body nucleus. These two nuclei, which were formed apart in the mature egg, drew near to fuse into a zygote nucleus. The chromosome number and nuclear DNA content were doubled in the zygote and each blastomere in embryos, supporting the hypothesis that the egg nucleus fuses with the second polar body nucleus and this conjugation initiates subsequent embryonic development.

1-Methyladenine: a starfish oocyte maturation-inducing substance

1-Methyladenine (1-MeAde) in starfish was the first compound to be identified as an oocyte maturation-inducing substance (MIS) among invertebrates in 1969 by Kanatani and co-workers. In starfish, the ripe ovary contains a huge number of fully grown oocytes of almost equal size. Each oocyte possesses a large nucleus (germinal vesicle, GV), which is arrested in late prophase of the first maturation division. The oocyte is surrounded by a single follicle layer. Such immature oocytes are not fertilisable. Resumption of meiosis in immature oocytes can be induced by 1-MeAde, and the mature oocytes thus become fertilisable (Kanatani et al., 1969; Kanatani, 1985). 1-MeAde is produced by ovarian follicle cells upon stimulation with a gonad-stimulating substance (GSS) released from the radial nerves (Fig. 1).It has been demonstrated that GSS is a peptide hormone (Kanatani et al., 1971). The action of GSS on 1-MeAde production in follicle cells appears to be mediated by its receptor, G-proteins and adenylyl cyclase (Mita & Nagahama, 1991). These findings suggest that a G-protein coupled (seven transmembrane type) receptor is involved in GSS signal transduction, similarly to the pituitary-gonadal axis in vertebrates.Thus, using degenerate probes derived from consensus sequences of the mammalian glycoprotein hormone (GTH and TSH) receptors, cDNA was cloned from mRNA of ovaries of Asterina pectinifera. The cDNA showed striking structural homology with members of the glycoprotein hormone receptor family in the transmembrane region, and contained a very large extracellular region. Expression was observed in isolated ovarian follicle cells. Thus, it seems likely that the glycoprotein hormone receptor (GTHR) family gene is related to GSS receptor in ovarian follicle cells. The phylogenic relatedness of the starfish GTHR was also assessed in relation to other vertebrate GTHRs. The analysis showed that the starfish gene diverged before differentiation of the gonadotropin (LH and FSH) and TSH receptors in vertebrates.

Centrosome inheritance in starfish zygotes: behaviour and duplicating capacity of the meiotic centrosomes in maturation division

In zygotes of almost all animals, it has been believed that only the sperm centrosome acts as the mitotic spindle poles. As first proposed a century ago by Boveri (1887), this uniparental (paternal) inheritance of the centrosome must depend on the selective loss of the maternal centrosomes. To trace the fate and duplicating capacity of all the maternal centrosomes/centrioles, including those cast off into polar bodies, we used two kinds of procedures: (1) suppression of polar body (PB) extrusion and (2) transplantation of PB centro-somes into artificially activated eggs.Gametes used in this study were from the starfish, Asterina pectinifera. Oocyte maturation was induced with 1-methyladenine (Kanatani, 1969). Suppression of PB extrusion and artificial activation were done according to Washitani-Nemoto et al. (1994). Micromanipulation was performed by the method of Saiki & Hamaguchi (1993). Behaviour of the centrosomes was examined by staining with an antibody against α-tubulin, polarisation and differential interference-contrast microscopy and transmission electron microscopy.In starfish oocytes, no centriole duplication occurs in meiosis II, hence each pole of a meiosis II spindle is formed by the splitting of paired centrioles in the inner centrosome of a meiosis I spindle into singles. Eventually, each of a second PB (PB2) and a mature egg inherits only one centriole from a meiosis II spindle (a PB1 inherits a pair of centrioles). So, either PB2 and the mature egg inherit a single centriole (Fig. 1; cf. Sluder et al., 1989; Kato et al., 1990). When mature eggs were artificially activated with the Ca2+-ionophore A23187, a single monaster was formed.

Spermatogenese beim Menschen: Grundlagenforschung und Klinik

Histological evaluation of human spermatogenesis suffers from the hazy border line between normal and pathological germ cell development. This border line needs better definition for histological fertility diagnosis and the early detection of germ cell tumors. Testicular biopsies from more than 2,900 patients with fertility disturbances and more than 1,900 patients with testicular tumors were investigated by means of semithin sectioning, different immunocytochemical methods and transmission electron microscopy. Cellular systems of the human testes possess a degree of autonomy from the body. Their morphological and functional heterogeneity reveals characteristics of cells that are not terminally differentiated. In the testis of an adult, fertile man not only the proliferation of spermatogonia, maturation divisions of spermatocytes and differentiation of spermatids take place, but also abortive germ cells, as well as apoptotic and degenerative cells appear. Disturbances of spermatogenesis are defined by the evaluation of quantity and quality of germ cell alterations. Compensatory and non compensatory defects of spermatogenesis may be distinguished. Deficiency of spermatogonial cell types, multilayered spermatogonia, megalospermatocytes, malformed spermatids and single tumor cells in the face of sufficient development of mature spermatids are considered compensatory defects of spermatogenesis. Dominating malformed germ cells or tumor cells accompanied by an arrest or lack of spermatogenesis, however, represent non-compensatory defects of spermatogenesis. In addition, normal organization and function of the microvasculature, Leydig cells and compartmentalizing cells in the intertubular space are prerequisites for spermatogenesis. The neuroendocrine function of Leydig cells may be responsible for regulating the blood flow rate and the permeability to hormones and nutritive substances. Finally, for patients a successful definition of the border line between normal and pathological events of germ cell development may be essential for early detection of germ cell tumors. Therefore, anatomical sciences not only contribute to basic research, advanced diagnostics and therapeutic concepts related to diseases of the male gonad, but also to the improvement of assisted reproduction.

Early embryonic development of the mayfly Ephemera japonica McLachlan (Insecta: Ephemeroptera, Ephemeridae).

In the newly laid egg of the mayfly Ephemera japonica, an egg nucleus (oocyte nucleus) at metaphase of the first maturation division is in the polar plasm at the mid-ventral side of the egg, and a male pronucleus lies in the periplasm beneath a micropyle situated just opposite the polar plasm or at the mid-dorsal side of egg. The maturation divisions are typical. An extensive and circuitous migration of the male pronucleus is involved in the fertilization process: it first moves anteriad in the periplasm from beneath the micropyle to the anterior pole of the egg and then turns posteriad in the yolk along the egg's long axis to the site of syngamy, near the center of the egg. Cleavage is superficial. The successive eight cleavages, of which the first five are synchronized, result in the formation of the blastoderm, and about ten primary yolk cells remain behind in the yolk. Even in the newly formed blastoderm, the thick embryonic posterior half and the thin extraembryonic anterior half areas are distinguished: the former cells are concentrated at the posterior pole of the egg to form the germ disc, and the latter cells become more flattened, forming serosa. Time-lapse VTR observations reveal a yolk stream that is in accord with the migration of the male pronucleus in time and direction. The yolk stream is also generated in activated unfertilized eggs, and it is probable that the migration of the male pronucleus in association with the fertilization may be directed by the yolk stream. J. Morphol. 238:327-335, 1998. © 1998 Wiley-Liss, Inc.

Division of Polar Bodies Induced by Their Enlargement in the Starfish Asterina pectinifera

The first polar body (FPB), which is formed at the first meiotic division during oogenesis, does not generally divide. We made a hypothesis that the amount of cytoplasm was not sufficient for the FPB to perform cell division, in spite of the same amount of genomes and centrosome as those of the secondary oocyte. To examine this hypothesis, hexylene glycol (HG) at a low concentration was applied to oocytes of the starfish Asterina pectiniferaduring the first meiotic division. Hence, FPBs were enlarged in their diameters, some of them divided once, and the division rate increased in proportion as their diameter extended. We further hypothesized that the difference between the second polar body (SPB) and the egg would be only the amount of cytoplasm and that if SPBs were enlarged, they would become eggs. When the secondary oocytes were treated with HG, large SPBs were obtained. Some of them, however, divided, and resultant daughter cells divided moreover, whereas eggs would not cleave unless they were fertilized. We discuss here the reason why the centrosome distributed during maturation division began to function in enlarged PBs.

Reorganization of the Cortical Actin Cytoskeleton during Maturation Division in theTubifexEgg: Possible Involvement of Protein Kinase C

Tubifexeggs undergo a drastic reorganization of the cortical actin cytoskeleton during metaphase of the second meiosis. At the end of the first meiosis, the egg cortex displays only scattered actin filaments and tiny dots of F-actin; during the following 90 min, cortical F-actin gradually increases in amount, becomes organized into foci that are interlinked by actin bundles, and generates a geodesic dome-like organization. In this study, we have characterized this reorganization of the cortical actin cytoskeleton. In living eggs injected with rhodamine–phalloidin at the beginning of the second meiosis, cortical actin assembly (i.e., formation of actin foci and bundles) proceeds normally, but labeled F-actin is not found to be included significantly in the formed cortical actin network, suggesting that the increase in cortical F-actin is not simply ascribable to the recruitment of preexisting actin filaments. Cortical actin assembly can be induced precociously not only by calcium ionophore A23187 but also by a phorbol ester PMA, an agonist of protein kinase C (PKC). Conversely, the formation of actin foci and bundles is inhibited by PKC antagonists, although cortical F-actin increases to some extent in the presence of these inhibitors. Similar inhibition of the cortical reorganization is elicited in eggs whose intracellular free calcium level ([Ca2+]i) has been clamped low by microinjection of a calcium chelator BAPTA. The treatment of BAPTA-injected eggs with PMA results in the formation of actin foci and bundles. An experiment with eggs injected with fluo-3 shows that [Ca2+]iincreases during metaphase of the second meiosis. These results suggest that the reorganization of cortical actin during metaphase of the second meiosis requires activation of PKC, which depends on increases in [Ca2+]i.