A light- and electron-microscopic study on the migration of primordial germ cells in the teleost,Oryzias latipes

Summary The primordial germ cells (PGCs) of Oryzias latipes in migration to the gonadal anlage have been investigated by light and electron microscopy. The ultrastructure of the PGCs, which occur in the subendodermal space on the syncytial periblast, differ conspicuously from that of the surrounding endodermal cells. After the PGCs move to the cavity between lateral plate and ectoderm, they are taken into the somatomesodermal layer and transferred to the dorsal mesentery where they form gonadal anlage with mesodermal cells. During their translocation to the dorsal mesentery through the somatic mesoderm, apparently without formation of pseudopods, the PGCs are completely surrounded by mesodermal cells. Since these conditions seem unfavorable to the active translocation of the PGCs to the dorsal mesentery, it is more likely that the PGCs are transferred passively by the morphogenic activity of the lateral-plate mesoderm.Counts of the number of the PGCs revealed that they are mitotically dormant during the migratory period. After the completion of the migration, they regain their proliferative activity. The PGCs in the female proliferate more actively than those in the male, which provides the first morphological indication of sex differentiation in this species of fish.     

Origin of the gonad in the juvenile of a solitary ascidian, Ciona intestinalis

In the just-metamorphosed juveniles of Ciona intestinalis, a round mass of tissue debris derived from the resorbed tadpole tail is situated in the broad space enclosed by the peritoneal membrane and the epidermis around the ventral side of the esophagus. In living juveni es, the origin of the gonad rudiment was traced back to the mass of tissue debris. Electron microscopically, the round mass was a clump of irregular-shaped phagocytotic cells engulfing degenerated cell fragments. On the surface of the cell clump, a small number of singly occurring round cells were found and identified as primordial germ cells on the basis of morphological continuity to obvious germ cells in later stages. Presence of nuage around the nucleus characterized the germ cells. In a few days the germ cells assembled to form a solid slender body (gonad rudiment) together with smaller somatic cells. The gonad rudiment left the space around the esophagus, moving into the narrow mesenteric space connecting the stomach and intestine on the fourth day after metamorphosis. It gradually increased in size by proliferation of the germ cells and somatic cells. The solid gonad rudiment changed into an oval vesicle with an eccentrically located cavity on about the seventh day after metamorphosis. The vesicle comprised a thinner wall made of a simple epithelium without germ cells and a thicker wall containing germ cells and somatic cells.     

A study on cell lineage,especially the germ cell line,in embryos of the teleost fish,Barbus conchonius

Summary Lucifer Yellow-Dextran labelling of lower layer cells (LLC), sometimes together with upper layer cells (ULC), of the 64-cellBarbus conchonius embryo resulted in labelled primordial germ cells (PGCs) at 12 h after fertilization (a.f.) in about 25% of cases. The presence of labelled PGCs was independent of the location of the injected blastomere with respect to the later orientation of the embryonic axis. After injection of an ULC alone, however, labelled PGCs were never found. Also, the distribution of labelled somatic cells differed between the ULC- and LLC-injected embryos. When we found fluorescent PGCs, only a few of them were labelled, suggesting that either a single predecessor exists earlier than the 64-cell stage or that the formation of germ cells is a polyclonal process. Tracing the fluorescent cells at successive stages of development shows an extensive mixing with unlabelled cells during the epiboly stage, which might well be the cause of partly unpredictable cell lineages. The chance of being committed to a specific fate is different for the ULC and LLC descendants. This might be due to relatively limited cell mixing between these two cell populations.     

Resorption of unemitted gametes in Lithognathus mormyrus (Sparidae,Teleostei): a possible synergic action of somatic and immune cells

The resorption of unemitted gametes during the post-spawning period of the male and female reproductive cycles in Lithognathus mormyrus was studied by histochemical, histological and cytological methods. The resorption of residual spermatozoa involved the phagocytotic activity of Sertoli cells bounding the seminiferous cysts of spermatozoa, and those associated with spermatogonia lining the lobular lumen. Spermatozoa remaining in the sperm duct were phagocytozed by the lining epithelial cells. Eosinophilic granulocytes and macrophages were identified in the vicinity of residual spermatozoa. The remnants of oocytes underwent an atretic phenomenon in which follicle cells were firstly involved, inducing a progressive fragmentation of the oocyte cytoplasm. Subsequently, eosinophilic granulocytes invaded oocyte degenerative areas and clung to the remaining vitelline inclusions ensuring their biotransformation into waste products (brown bodies). The analogy of the resorption processes of both male and female unemitted gametes during the post-spawning period of natural reproductive cycle, involving first the enveloping somatic cells and then immune cells, is emphasized.     

Spermatozoa in the reproductive system of a hermaphroditic marine tardigrade,Orzeliscus belopus (Tardigrada: Arthrotardigrada)

Orzeliscus belopus has long been regarded as the only hermaphroditic marine tardigrade, yet there has been no published detailed information on its internal anatomy. Our study elucidates the ultrastructure of the ovotestis and the spermatozoa of Orzeliscus cf. belopus from Bermuda. The ovotestis had no septum to separate male and female germ cells and, while early stages of spermatogenesis were not observed, many spermatozoa were found at the periphery in both anterior and posterior areas of the gonad. The nucleus and the mitochondria of the spermatozoa in the ovotestis extend backward from the centriole region, forming a half-headed arrow-shape with the nucleus on the outer side of the ‘arrowhead’. The cross section of a long vesicular body (the paranuclear vesicle) is dumbbell- or horseshoe-shaped and attached to almost the entire length of the nucleus. In the seminal receptacle, we found both a complete spermatozoon and some which started to degrade. There is an indication that sperm is further modified after discharge as within the receptacle duct the sperm is no longer half-headed arrow-shaped but has a straight nucleus. This modification might be correlated with the degeneration of the paranuclear vesicle. Our observations clearly show that O. belopus is a simultaneous hermaphrodite, and suggests that the reproductive mode includes copulation and cross-fertilization.     

Early ontogeny of Adinia xenica (Pisces,Cyprinodontiformes): 1. The development of embryos in hiding

Synopsis Examination of living and stained preserved embryos of the fundulid Adinia xenica from the intertidal salt marshes of the northern Gulf of Mexico, revealed a saltatory pattern of development. Embryonic structures indicate an ability to tolerate low oxygen conditions, primarily by extensive vascularization of the large yolksac and pericardial headfold, and capillaries covering the entire body. Adhesive fibres of the chorion hold the egg to the chosen substrate, away from anoxic sediments. Prior to hatching most embryos enter a resting interval of up to four weeks, with a mean duration of 8 days (in the laboratory). Growth ceases, development is limited to some skeletal calcification, and metabolic rate, as indicated by bradycardia and yolk resorption, is greatly reduced. This step distributes hatching and recruitment over time in a variety of environmental conditions. Hatching is considered to result from a variable summation of internal and external factors; ease of its induction increases during the resting interval. Upon hatching most embryos are well developed and ready to feed exogenously, there being essentially no free-embryo phase. The considerable individual variation in timing and relative order of some ontogenic events may provide opportunities for environmentally induced heterochronies and thus assist A. xenica to develop in a range of habitats and conditions. Reproductive style includes hiding the eggs in protected locations, and a fractional spawning pattern. Consideration of this plus morphological adaptations, egg size and adhesivity places A. xenica in the reproductive guild of brood-hiding cavity spawners.     

Number of preoptic GnRH-immunoreactive cells correlates with sexual phase in a protandrously hermaphroditic fish, the dusky anemonefish (Amphiprionmelanopus)

The populations of gonadotropin-releasing hormone (GnRH)-producing cells within the preoptic area (POA) and terminal nerve (TN) of the brain have been suggested as the neuronal systems mediating social control of sex and gonadogenesis in sequentially hermaphroditic teleosts. In the present study, the number and soma size of GnRH-immunoreactive (GnRH-ir) cells in the POA and TN were studied in male, female and juvenile individuals of the dusky anemonefish (Amphiprionmelanopus), a species which displays both male to female sex change and socially controlled sexual maturation. The results showed that the number of POA (but not TN) GnRH-ir cells differ significantly between sexual phases, with males displaying higher cell numbers than both females and juveniles. Soma sizes of POA and TN GnRH-ir cells were larger in females than in males and juveniles. However, this relationship was fully explained by differences in body size. The results indicate that high POA GnRH cell numbers are part of a masculinizing mechanism and support the hypothesis that the POA GnRH cell population plays a central role in initiating or mediating the process of socially induced gonadal and/or behavioural transformations in sequential hermaphrodites. Accepted: 9 June 1997     

Early ontogeny of Adinia xenica (Pisces,Cyprinodontiformes): 2. Implications of embryonic resting interval for larval development

Synopsis Larval development commences with first exogenous feeding, and ends with final remodelling of caenogenetic structures into the definitive organs of juvenile and adult. For the intertidal cyprinodontid Adinia xenica this generally corresponds to the interval between hatching and completion of scalation. The final step of the embryo period is a resting interval of variable duration. Embryos were induced to hatch after 2 and 10 days of this near arrest. Although the general pattern of larval development was the same for both groups, differences were observed in the rates and order of calcification of skeletal elements, fin differentiation and growth, and scalation. For example, embryos hatching 8 days later in the resting interval were already pattially calcified, but completed calcification at a slower rate than the group hatching after 2 days. These differences may be due to effects of the duration of the resting interval itself; or they may reflect genetic variation of which age at hatching is only one manifestation.     

The ontogeny of search behavior in the white crappie,Pomoxis annularis

Synopsis Animals that forage for discrete, isolated resources are often characterized as either ambush (sit-and-wait) or cruise (active) searchers. Juvenile white crappie, Pomoxis annularis, search for zooplankton prey using a saltatory search (SS) strategy. Unlike ambush and cruise search, SS involves scanning for prey only during the brief stationary periods that punctuate repositioning movements. If prey are not found, these fish swim a short distance, stop, and scan again. In this paper, we describe the ontogeny of prey search in the white crappie and compare the search pattern that they employ with that of juveniles. White crappie larvae searched for prey throughout the search space and only during the pauses that punctuated swimming movements. Prey location distances increased with fish size, as did several other components of the predation cycle. We conclude that white crappie larvae employ a search strategy similar to that exhibited by juveniles. We emphasize that, to obtain an accurate assessment of the feeding ecology of early life history stages, the search pattern that they employ must be characterized, and its components quantified.     

Mammalian testis: a target of in vivo electroporation

Mammalian spermatogenesis consists of three biologically significant processes: stem cell self-renewal and differentiation, meiosis, and haploid cell morphogenesis. Understanding the molecular mechanisms behind these processes might provide clues to the puzzle of species preservation and evolution, and to treatments for male infertility. However, few useful in vitro systems exist to investigate these processes at present. To elucidate these mechanisms, in vivo electroporation of the testis might be a convenient option. Since DNA solution can be injected into the seminiferous tubule via the rete testis, similar to germ cell transplantation, it is easy to transfect expression vectors into various differentiated germ cells and supporting Sertoli cells with adequate electric shock. Unfortunately, it is difficult to create transgenic animals using this method because of its low efficiency. However, gain- and loss-of-function assays, promoter assays, and tagged-protein behavior assays can be conducted with this technique, as in in vitro culture systems.     

Early development of the northern logperch,Percina caprodes semifasciata,according to the theory of saltatory ontogeny

Synopsis The early development of northern logperch, Percina caprodes semifasciata, was examined from an ecological perspective and in relation to the theory of saltatory ontogeny. Steps, the intervals of relative homeostasis, separated by thresholds, rapid switches to new form and function resulting from canalization of changes in the preceeding step, are described and related to environmental factors. Adaptive shifts in growth allometries provided further evidence of the saltatory nature of ontogeny. Logperch eggs are small (1.2 mm diameter), demersal, and adhesive. The simple embryonic respiratory system reflects high oxygen availability on the lotic spawning grounds. Hatching occurs early, relative to other darters, and free embryos are pelagic. The small free embryos presumably drift downstream to lentic areas, where small planktonic food is more abundant. Logperch are therefore nonguarding, open substrate spawning lithopelagophils and, along with other Percina species, have not advanced from ancestral percid reproductive styles. Percina species have generally remained in ancestral habitats-rivers and lakes. The close correlation between developmental patterns of logperch and environmental factors is therefore interpreted as a restriction based on retention of ancestral characteristics, not as evidence of novel adaptations to new habitats.     

Motile iridophores of a freshwater goby,Odontobutis obscura

Summary Reflecting chromatophores in the dermis of the skin of a freshwater goby, Odontobutis obscura, are of an iridophore type. These chromatophores contain numerous reflecting platelets, which are similar to those in iridophores of other fish and amphibian species. It was found that these iridophores are motile, i.e., these cells respond to certain stimuli with translocation of the platelets within the cells. K⁺ ions induced dispersion of the platelets in excised scale preparations, but not in excised scales from chemically denervated fish. Norepinephrine and melatonin also induced dispersion of the platelets. Alpha-MSH was effective in aggregating these organelles into the centrospheres of the cells. The conclusions reached are: (1) iridophores of O. obscura are motile; (2) the movement of the iridophores is under nervous and hormonal control.     

Changes in developmental profiles of brain gangliosides during ontogeny of a teleost fish (Sarotherodon mossambicus,Cichlidae)

Summary Ontogenetic changes of brain ganglioside concentration and composition have been followed in the teleost fishSarotherodon mossambicus Cichlidae) from the 1st day post hatching to the adult stage, in order to correlate these with findings in higher vertebrates. During the developmental period from hatching to the transition to free swimming, which comes along with maximum rate of synaptogenesis, a sharp rise in the brain ganglioside content occurs, which is mainly due to the trisialoganglioside GT1b. In the following phase of myelination (characterized in birds and mammals by an increase in GM1, GM1 and GM4) accretion of the novel and so far unidetified fraction (GM2) occurs, which is highly enriched in the brain stem. The results obtained are discussed with respect to gangliosides as useful biochemical markers for brain development and maturation in all vertebrates.     

Germ cell nuclear antigen (GCNA1) expression does not require a gonadal environment or steroidogenic factor 1: Examination of GCNA1 in ectopic germ cells and in Ftz-f1 null mice

The germ cell lineage is first recognized as a population of mitotically proliferating primordial germ cells that migrate toward the gonadal ridge. Shortly after arriving at the gonadal ridge, the germ cells begin to initiate a commitment to gamete production in the developing gonad. The mechanisms controlling this transition are poorly understood. We recently reported that a mouse germ cell nuclear antigen 1 (GCNA1) is initially detected in both male and female germ cells as they reach the gonad at 11.5 days postcoitum (dpc). GCNA1 is continually expressed in germ cells through all stages of gametogenesis until the diplotene/dictyate stage of meiosis I. Since GCNA1 expression commences soon after primordial germ cells arrive at the gonadal ridge, we wanted to determine whether the gonadal environment was essential for induction of GCNA1 expression. By examining GCNA1 expression in germ cells that migrate ectopically into the adrenal gland, we determined that both the gonadal and adrenal gland environments allow GCNA1 expression. We also examined GCNA1 expression in Ftz-F1 null mice, which are born lacking gonads and adrenal glands. During embryonic development in the Ftz-F1 null mice, the gonad and most germ cells undergo apoptotic degeneration at about 12.5 dpc. While most of the germ cells undergo apoptosis without expressing GCNA1, a few surviving germs cells, especially outside the involuting gonad clearly express GCNA1. Thus, although the Ftz-F1 gene is essential for gonadal and adrenal development, induction of GCNA1 expression in germ cells does not require Ftz-F1 gene products. The finding that germ cell GCNA1 expression is not restricted to the gonadal environment and is not dependent on the Ftz-F1 gene products suggests that GCNA1 expression may be initiated in the germ cell lineage by autonomous means. Mol. Reprod. Dev. 48:154–158, 1997. © 1997 Wiley-Liss, Inc.