Gonadotropin-releasing hormone (GnRH) is well known as the central regulator of the reproductive system through its stimulation of gonadotropin release from the pituitary. Studies on GnRH have demonstrated that GnRH has both stimulatory and inhibitory effects on cell proliferation depending on the cell type; however, the mechanisms of these effects remain to be elucidated. Against this background we used four human cell lines, TSU-Pr1, Jurkat, HHUA and DU145, and newly found that GnRH increased or decreased the colony-formation depending on the cell line. Moreover, we demonstrated that the stimulatory and inhibitory effects of GnRH exhibit distinct ligand selectivities. In order to investigate the molecular basis of these phenomena, analyses of the expression of human GnRH receptors were performed and, moreover, the effects of GnRH were analyzed under conditions in which human GnRH receptors were knocked down by the technique of RNA interference. Consequently, it was found that human type II GnRH receptor, which had been suspected of being nonfunctional because of alterations in its sequence, is involved in the effects of GnRH on cell proliferation. In this article, the influence of the autocrine activities of the cells is also reviewed, focusing on the characteristics of substances secreted from the four cell lines. Based on recent studies of GnRH and its receptors and our up-to-date findings, the evolutionary implications of GnRH action are discussed.

The sex-determining gene in Oryzias latipes and O. curvinotus has been proved to be DMY. Although O. curvinotus has the DMY gene on the Y chromosome which is homologous to the Y chromo-some of O. latipes, the sex-determining mechanism of other Oryzias fishes has not been identified. In order to uncover the sex-determining mechanism of O. luzonensis and O. mekongensis, which are most closely related species to O. latipes and O. curvinotus, we analyzed the sex ratio of the progeny of sex-reversed fish. We were able to obtain sex-reversed males by the administration of methyltestosterone, and found that these yielded all-female offspring in both species. These results indicate that O. luzonensis and O. mekongensis have the XX-XY sex-determination system.

Resting eggs of Daphnia, a key crustacean zooplankton of freshwater food chains, can remain viable for more than a century. These eggs are able to withstand freezing and drying, and can survive the harsh environment of a predator's digestive system. Until recently little was known about the chemical composition, microanatomy, and physical properties of the resting eggs.

The current study utilized a physical technique, the X-ray analytical microscope, to identify and localize component elements of the Daphnia resting egg. The analysis demonstrated that phosphorus, sulfur, potassium, and calcium were detected as elemental components of the resting egg, and detection intensities of the four elements differed according to the position of the eggs. Phosphorus and calcium were mostly detected in regions of the eggshell that surrounded the two embryos. In addition, sulfur was distributed throughout the eggshell whereas potassium was localized to the areas that corresponded to where the embryos were encased. Through the use of X-ray analytical microscopy, the current study identifies elemental characteristics in relationship to the structure of the Daphnia resting eggs.

To know whether behavior of pigment cells correlates the process of gastrulation or not, gastrulating embryos of several species of regular echinoids (Anthocidaris crassispina, Mespilia globulus and Toxopneustes pileolus) and irregular echinoids (Clypeaster japonicus and Astriclypeus manni) were examined. In M. globulus and A. crassispina, the archenteron elongated stepwise like in well-known sea urchins. In the embryos of both species, fluorescent pigment cells left the archenteron tip and migrated into the blastocoel during gastrulation. In T. pileolus, C. japonicus and A. manni, on the other hand, the archenteron elongated at a constant rate throughout gastrulation. In these species, no pigment cell was observed at the archenteron tip during invagination processes; pigment cells began to migrate in the ectoderm from the vegetal pole side toward the apical plate without entering the blastocoel. These results clearly indicate that the behavior of pigment cells closely correlated the manner of gastrulation. Further, it was examined whether the archenteron cells are rearranged during invagination, by comparing the number of cells observed on cross sections of the archenteron at the early and late gastrula stages. The rearrangement was not conspicuous in A. crassispina and M. globulus, in which archenteron elongated stepwise. In contrast, the archenteron cells were remarkably rearranged in C. japonicus, alothough the archenteron elon-gated continuously. Thus, neither the behavior of pigment cells nor the manner of gastrulation matches the current taxonomic classification of echinoids.

Most pandalid shrimps exhibit protandric hermaphroditism, and detailed information on ovarian development of pandalid species is important for a better understanding of vitellogenesis in crustacean species. In the present study, we characterized ovarian development under light and electron microscopy and examined the hemolymph vitellogenin levels in the coonstriped shrimp, Pandalus hypsinotus under laboratory conditions. To measure vitellogenin levels, a time-resolved fluoroimmunoassay (TRFIA) was developed after purification of vitellin and production of the anti-vitellin antiserum. The TR-FIA showed wide assay range (0.98–2000 ng/ml), high sensitivity (0.5 ng/ml), and low assay variability (0.9–6.4% of intraassay coefficients, 1.4–5.1% for interassay coefficients). Female P. hypsinotus had non-vitellogenic ovaries in March after the eggs attached to the abdomen hatched, and started yolk accumulation in the ovaries during April-October. During yolk accumulation, yolk globules appeared and increased in the ooplasm. After yolk accumulation, gonadosomatic index (GSI) reached 8.3–8.5 just before oviposition. Females spawned and were ovigerous during June-July of the next year. Hemolymph vitellogenin levels were low (0.006±0.008 mg/ml, mean±SD) before the yolk accumulation, and became significantly higher (2.66±0.93 mg/ml) during yolk accumulation (GSI, 2–8). Just before oviposition, levels declined to low levels (0.040±0.012 mg/ml). Vitellogenin levels were significantly correlated to GSI during the yolk accumulation. The obtained results show that the process of vitellogenesis during the female phase of P. hypsinotus is similar to other crustacean species that do not change sex.

Diapause pupae of Papilio xuthus show color polymorphism, represented by diapause-green, orange, and brownish-orange types that are each associated with specific pupation sites. We investigated the role of the site of pupation on the induction of the development of orange types (or brownish-orange types), and the endocrine mechanism underlying the control of color polymorphism in short-day pupae. All short-day larvae of the wandering stage developed into orange or brownish-orange type pupae when they were placed in rough-surfaced containers after gut-purge. Utilizing a pharate pupal ligation between the thorax and abdomen, the endocrine mechanism underlying the control of color polymorphism was shown to involve a head-thorax factor (Orange-Pupa-Inducing Factor: OPIF) that induced orange types in short-day pupae. OPIF was bioassayed using the ligated abdomens of short-day pharate pupae. OPIF was extractable with 2% NaCl solution from 5th-instar larval ganglia complexes following the mesothoracic complex (TG_2,3-AG_1–7), but it could not be extracted with either acetone or 80% ethanol solution. OPIF may not exist in the brains of day-0 pupae or in brain-subesophageal ganglion and prothoracic ganglion complexes of 5th-instar larvae. The short-day pharate pupae responded to OPIF in a dose-dependent manner.

The echinoid, Asthenosoma ijimai belonging to the order Echinothurioida from Japanese waters shows the geographical variation in morphological and ecological characters. The echinothurioid from Ryukyu Islands in southern Japan is cleary different from that of Sagami Bay and Suruga Bay in the middle part of Japan at non-molecular level. Their phylogenetic and taxonomic relationships were studied at the molecular level by allozyme analysis. The results demonstrated that the two echinothurioids from Ryukyu Islands and Sagami Bay do not share gene pools with each other, and they were fixed for different alleles at five genetic loci (Mdh, G6pd, Po, Alk-3 and Est-7) in a total of 23 enzyme genetic loci scored. This indicates no gene flow between the two echinothurioids, and is a molecular evidence for that they are reproductively isolated and genetically distinct species. The Nei's genetic distance (D=0.181) between the two were significantly higher than those between conspecific local populations, and comparable to those between closely related species in many other animals containing echinoderms. The present molecular data are well consistent with the non-molecular evidence from morphology, developmental biology and ecology. Putting these data together, we propose that the two echinothurioids should be classified as two sibling species of the genus Asthenosoma and would like to give the following scientific names: the echinothurioid species from Sagami Bay is Asthenosoma ijimai and that from Ryukyu Islands is A. ijimai R.

Four new species of freshwater rhabdocoel flatworms from ephemeral rock pools in southeastern Botswana are described and discussed. Two of them, Syringoplana kolasai n. gen. n. sp. and Mesostoma thamagai n. sp. belong to the Typhloplanidae Graff, 1905. The unique construction of the excretory system is the main characteristic of S. kolasai. M. thamagai can be separated from other Mesostoma Ehrenberg, 1837 species by the presence of a bundle of eosinophilic glands at the transition from oviduct to seminal receptacle. The other two taxa, Gieysztoria isoldeae n. sp. and G. faubeli n. sp. belong to the Dalyelliidae Graff, 1905. G. isoldeae is characterised by the presence of four separate hollow spines in the male atrium, which are connected to two accessory glandular organs. G. faubeli can be separated from other Gieysztoria Ruebush and Hayes, 1939 species by the detailed construction of the stylet. Apart from these two species the occurrence of an unidentified Microdalyellia Gieysztor, 1938 species is mentioned.