Early vertebrates may have had genes that would later produce hormones concerning Ca homeostasis in advanced vertebrates. These genes may possibly have expressed the respective product for the local control of Ca in some tissues, which must be a paracrine control. When vertebrates acquired the jaw, however, delicate but systemic Ca control system may have been needed to cope with fluctuations in blood Ca levels resulting from the digestion of food. Furthermore, when vertebrates were transferred from freshwater to seawater or to land, new control systems must have been required. Therefore, such vertebrates might have coped with two ways in their evolution. One is the increase of the number of local production sites of hormones, for the delicate control of Ca in various tissues, which must be the paracrine control. The other is the creation of new glands for the systemic control of Ca homeostasis by making paracrine cells independent from tissues. This style must be an endocrine control. Therefore, in modern vertebrates, in various tissues, calcitonin, stanniocalcin, parathyroid hormone-related protein, and maybe also prolactin, or at least substances immunoreactive to antibodies against these hormones, are expressed locally as paracrine tissues. At the same time, these hormones or similar molecules are also produced in special endocrine glands such as the ultimobranchial glands, Stannius corpuscles, the parathyroid glands and the pituitary gland, respectively. This supposition is summarized in ^{Fig. 1}.

The Japanese dace (Tribolodon hakonensis) is the only teleost species that inhabits Lake Osorezan, a remarkable acid lake in Japan with the water pH of 3.4–3.8. In the present study, physiological changes following transfer of the dace acclimated to neutral stream water (pH 6.8–7.2) to acid lake water (pH 3.6–3.7) were examined with special reference to changes in gill chloride cell morphology. The dace survived direct transfer to acid lake water for 3 days. Plasma [Na ] and [Cl⁻] showed transient decreases at 1 hr after transfer; however, the decrease in [Na ] was greater than that in [Cl⁻]. The recovery of [Cl⁻] was more evident than that of [Na ]. The transient decreases of plasma [Na ] and [Cl⁻] were followed by acidosis. Blood pH was decreased by 0.13 unit at 6 hr, but partially restored by 24 hr. In the dace acclimated to neutral stream water, chloride cells were scattered in the gill filament. Following transfer to acid lake water, however, well-developed chloride cells were arranged in a radial fashion, forming follicular or gland-like structures in the gills. Each chloride cell was equipped with an apical pit, which faced a common lumen leading to the external environment. These findings provide morphological evidence for a significant role(s) of gill chloride cells in ion and acid-base regulation in the acid-tolerant dace.

To understand the perception of an odor mixture by the slug Limax marginatus, a mixture of two odors (carrot and cucumber) was used to condition the slugs, and internal representation of the odor mixture and its components was determined by cooling-induced retrograde amnesia. Slugs conditioned with the odor mixture showed aversive behavior, not only towards the mixture, but also towards the individual odor components. When the conditioned slugs were cooled after presentation of one of the odor components, odor preferences for both components recovered, suggesting that the slugs perceived the odor mixture as an entity. However, when the slugs were exposed to the components of the odor mixture after conditioning with the mixture, cooling treatment resulted in amnesia, which was specific towards the odor presented before the cooling treatment. This suggests that slugs exposed to odor components after conditioning were able to recognize the odor components individually. Thus, slugs learn a binary mixture as an entity as long as they have no experience about the individual components of the mixture. These results are discussed in relation to other conditioning strategies, such as second-order conditioning or blocking, where a mixture of cues is used.

It is known in Paramecium that the fate of the micronucleus is determined by the intracellular localization of the nucleus and that protoplasmic streaming named cyclosis is very active during vegetative phase. The active streaming, if it occurs during conjugation, may interfere with the correct positioning of micronuclei and/or cytoplasmic determinants in a cell. In the present work, the velocity of protoplasmic streaming was measured during conjugation of P. caudatum. The results showed that the velocity changed remarkably at 3 stages. (1) It increased with the expansion of its active area 3 to 5 minutes after the mating reaction and then decreased again to the ordinary speed within 3 hours. The results of micro- or macronuclear removal showed that the increase in the velocity was not mediated either by the micronucleus or by the macro-nucleus. Moreover, injection of the anti-α-tubulin antibody at the mating reaction did not suppress the protoplasmic streaming effectively, although a role of microtubules on the protoplasmic streaming is not ruled out completely. (2) At the stages of disintegration of meiotic products and the exchange of gametic nuclei, the cytoplasm ceased streaming and then resumed it soon after synkaryon formation. (3) The streaming ceased again at the critical stage of macro-and micronuclear differentiation after the separation of mating pair, although the streaming was very active before and after this stage. Both cessations of (2) and (3) were not influenced by removal of the micronucleus during conjugation. The velocity of protoplasmic streaming during conjugation does not directly correlated with the micronuclear behavior. The cessation seems to be very convenient for intracellular positioning of the cytoplasmic determinants and/or of the micronuclei for the nuclear differentiation.

Three phenotypic color pattern genes of the guppy (Poecilia reticulata), i.e., black caudalpeduncle (Bcp), red tail (Rdt) and variegated tail patterning (Var), were genetically analyzed and mapped. Crosses between the Tuxedo (TUX) and Green Variegated (GV) guppy strains commercially cultured in Singapore were used to determine the gene control of these color patterns. F₁ progenies were produced by single-pair reciprocal crossing between TUX and GV, while the F₂ generation was obtained from full-sib mating between F₁ males and females. F₁ and F₂ data were segregated according to color phenotypes and sex, and tested by chi-square analyses. The Bcp, Rdt and Var color pattern genes, located at different loci on the X- and Y-chromosomes, showed single gene inheritance and dominant expression in both sexes. Their corresponding recessive alleles, Bcp , Rdt and Var , do not produce any color patterns. Genotypes of Tuxedo males are proposed to be X_Bcp,Rdt,Var Y_Bcp,Rdt,Var (type I), X_{Bcp ,Rdt,Var} Y_Bcp,Rdt,Var (type II) and X_Bcp,Rdt,Var Y_{Bcp ,Rdt,Var} (type III) while females are X_Bcp,Rdt,Var X_Bcp,Rdt,Var . Green Variegated males and females have the X_{Bcp ,Rdt ,Var}Y_{Bcp ,Rdt ,Var} and X_{Bcp ,Rdt ,Var}X_{Bcp ,Rdt ,Var} genotypes, respectively. Close linkages of 3.1, 2.3 and 2.2 map units were estimated for the sex-determining region (SdR)–Rdt, Rdt–Bcp, and SdR–Var gene pairs, respectively, while Bcp was approximately 5.1 map units from the SdR. The phenotypic map order of the guppy Y-chromosome is inferred to be Var–SdR–Rdt–Bcp.

Many color varieties of the guppy, Poecilia reticulata, are commercially cultured in Singapore for the aquarium industry. In the group of guppy varieties called Snakeskin, males characteristically have snakeskin-like reticulations on the body and caudal fin. The snakeskin pattern on the body of male Snake-skin guppies is due to a Y-linked gene (Ssb). Female guppies, being homogametic (XX), do not carry the Ssb gene. About 90% of Yellow Snakeskin males have the typical snakeskin pattern on their bodies and tails. The remaining males are different in that the snakeskin body pattern has been modified into four or five vertical bars on the caudal-peduncle region. F₁ and F₂ results of single-pair reciprocal matings of the Yellow Snakeskin variety show that a single gene is responsible for the vertical bar pattern. This gene, bar, is autosomal recessive. In the homozygous condition (barbar), it interacts with the Y-linked Ssb gene to give vertical barring patterns on the caudal-peduncle of Yellow Snakeskin males. This pattern is not expressed when the dominant allele, bar , is present.

The silver crucian carp, Carassius langsdorfii has three reproductive characteristics: gynogenesis, polyploidy (triploid or tetraploid), and genetic homogeneity within a family. In natural water, the silver crucian carp populations consist of multiple clonal lines. In the present study, three microsatellite DNA loci were used to distinguish several clonal lines of the silver crucian carp sampled from natural water. Progeny and the maternal fish had the same genotype in the three loci. In 237 fish collected from the wild, nine alleles were observed in GF1*, sixteen alleles in GF17*, and nine alleles in GF29*. Ten genotypes were observed in GF1*, seventeen in GF17* and eight in GF29*. The proportion of heterozygotes was very high in each locus (1.000). Sixteen clonal lines were distinguished by the combined genotypes of three microsatellite loci. Two subtypes were also detected within the clonal line KOC-011.

To study the dynamics of the microtubules throughout the sexual cycle of Paramecium caudatum, we employed a monoclonal antibody (mAb) N356, that specifically recognized the 50-57 kDa axonemal proteins, equivalent to the tubulins of P. caudatum. The mAb decoration was observed on the micronuclei, as well as structures that consisted of microtubules not only in P. caudatum but also in other species P. bursaria, P. tetraurelia and P. trichium. While the micronucleus was consistently decorated with mAb, the macronucleus was decorated only during morphological changes, that is, at binary fission of the vegetative phase and at skein-formation during conjugation. During conjugation, the oral apparatus, with its associated microtubular structures, disappeared at meiosis I then reappeared at the end of the third division of the synkaryon. The current study led to several major findings on the behavior of the germinal micronucleus during conjugation: 1) the microtubules seemed to connect some of the haploid nuclei to the paroral region at the end of meiosis II; 2) mAb decoration disappeared from degenerating haploid nuclei, while it remained in the surviving nucleus; 3) mAb decoration also disappeared from macronuclear anlagen after the initiation of nuclear differentiation, while it remained in the other four post-zygotic micronuclei. The mAb decoration pattern could be used as an indicator of the fate of a micronucleus and suggests that both the intra-micronuclear and the cytoplasmic microtubules might have an important role in determining of the fate of micronuclei by translocating them to specific areas of the cytoplasm.

We examined the morphological features of spermatozoa from Conger myriaster by scanning electron microscopy (SEM) and transmission electron microscopy (TEM). Spermatozoa consisted of three regions, a head, neck and flagellum. Total length was approximately 40 μm, with the head 3 μm, neck 0.5 μm and flagellum about 37 μm. The head was crescent-shaped, with a mitochondrion on the concave surface. The neck, with an attached rootlet, consisted of two constrictions close to the base of the flagellum. Two bundles extending from the proximal centrioles existed on both the convex and concave surfaces of the sperm head. The flagellum showed a 9 0 axonemal pattern. Some flagella showed a coiled form, the developmental mechanism of which was unclear. These features were compared with those of other Anguilliformes.

2, 22, 25-Trideoxyecdysone (5β-ketodiol) has been shown to be derived from cholesterol in eggs of the silkworm, Bombyx mori. In order to investigate the difference in ecdysteroid metabolism between diapause eggs and non-diapause eggs of the silkworm, ³H-5β-ketodiol was microinjected into eggs at several stages of early embryogenesis, and the metabolites were characterized using high-performance liquid chromatography. The injected ³H-5β-ketodiol was metabolized not only to free ecdysteroids, but also to conjugates (phosphoric esters of ecdysteroids), in both diapause eggs and non-diapause eggs. Among these metabolites, 20-hydroxyecdysone, which is considered as the active hormone in silkworm eggs, was detected in non-diapause eggs. However, in diapause eggs, various radioactive putative precursors of 20-hydroxyecdysone, such as 2, 22-dideoxyecdysone, 2-deoxyecdysone and ecdysone, were detectable, but ³H-20-hydroxyecdysone was not found. These results suggest that the 20-hydroxylation of ecdysone, which is catalyzed by ecdysone 20-monooxygenase, may be a rate-limiting step in the formation of 20-hydroxyecdysone from ketodiol in the silkworm eggs.

Juveniles of chum salmon (Oncorhynchus keta) held at high population density were apparently smaller than those held at medium and low population densities. The effects of high population density on pituitary growth hormone (GH) cells in juvenile chum salmon were examined using immunocytochemical and in situ hybridization techniques. The ratio of GH-immunoreactive (ir) area to the whole pituitary was almost constant in all of the high, medium and low population density groups, although the number and sizes of GH-ir cells were decreased in the high population density group. Image-analysis of GH-ir cells indicated the presence of a population of heterogenous cells, in which medium or rather strongly stained smaller cells and as extreme weakly stained larger cells. The medium or rather strongly stained smaller cells predominated in the high population density group, while weakly stained larger cells in the low population density group. In situ hybridization study showed somewhat different distributions and intensities of hybridization signals for mRNAs encoding GH I and II precursors. The area showing signals for GH II mRNA in the high population density group was significantly smaller than those in the medium and low population density groups. In contrast, the sizes of areas showing signals for GH I mRNA did not differ among the groups, although the intensity was slightly higher in the high population density group. These results indicate that high population density decreased the number of weakly immunoreactive larger GH cells, and also suppressed expression of the gene encoding GH II precursor, which may result in retarded somatic growth.

Members of the family Bruchidae (Insecta, Coleoptera) were examined for Wolbachia infection. Out of seven species investigated, which represented three subfamilies and five genera, amplified bands were detected only from Callosobruchus chinensis by diagnostic PCR with Wolbachia-specific primers. Bacterial 16S rDNA was amplified by PCR, cloned and sequenced from the total DNA of C. chinensis. Molecular phylogenetic analysis demonstrated that the sequence belongs to a monophyletic group of the genus Wolbachia in the α-Proteobacteria. The Wolbachia was detected from all tissues and body parts such as ovary, gut, fat body, muscle, wing, leg, head and antenna. In six geographic populations from central Japan, all the individuals examined, more than 400 in total, possessed the Wolbachia, indicating high prevalence of Wolbachia among natural populations of C. chinensis.

The retroposon SINE-R.C2 was first identified as a human-specific insertion in the complement C2 gene and the insertion of a closely related element has been implicated as a cause of Fukuyama type muscular dystrophy. In a previous study, we found SINE-R-type retroposons, derived from the endogenous retrovirus HERV-K family, have been found to be hominoid specific. In this report, we identified twelve SINE-R-type retroposons from the human Y chromosome by a PCR approach. Compared with other human retroposons (SINE-R.C2, 11, 14, 19, and HS307/HS408), the Y-chromosomal retroposon showed a high degree of sequence homology (86–96%). Phylogenetic analysis using the neighbor-joining method revealed that SINE-R-type retroposons on the Y chromosome were inter-related with hominoid primates, suggesting that SINE-R elements on the Y chromosome have evolved progressively in the hominoid radiation. Two elements SINE-R.Y-2 and Y-7 are more closely related to the human-specific retroposon SINE-R.C2 than other Y-chromosomal retroposons. Such elements thus are likely to be of great potential relevance to recent events in hominoid evolution and to structural change or genetic variation connected to various disease.

The least weasel Mustela nivalis and the ermine M. erminea of Japan are considered relicts of the last glacial period. To study phylogeographic variation in these mustelines, fragments of the mitochondrial DNA control region were sequenced. In both species, the control region included tandem repeats of 10base motifs at the 3′ portion specific to the genus Mustela. Phylogenetic trees of the 5′ portion (581–584 bases) of M. nivalis indicated that haplotypes in the Hokkaido population clearly diverged from those in the Honshu population. Geographic pattern of intraspecific variations illustrated by this result is concordant with that by the result of a previous cytogenetic study, which revealed the karyotypic differentiation between the two populations. These findings might suggest that the Hokkaido and Honshu populations of M. nivalis were geographically isolated in advance of the formation of the Tsugaru strait between Hokkaido and Honshu. Distribution pattern of the mitochondrial DNA haplotypes within Hokkaido suggests that M. nivalis experienced the repeated changes of environments during glacial and inter-glacial periods of the Quaternary, or that this species immigrated from Sakhalin/Siberia into Hokkaido so recently. On the other hand, M. erminea exhibited a smaller sequence divergence between the Hokkaido and Honshu populations, suggesting a shorter geographic isolation than in M. nivalis.

Phylogenetic relationships among Asian and African lygosomine skinks of the Mabuya group were inferred from 825 base pairs of DNA sequences of mitochondrial 12S and 16S rRNA genes. Results indicated the presence of two distinct lineages within this group, of which one consisted of Lamprolepis and Lygosoma, and the other of Apterygodon, Dasia, and Asian and African Mabuya. Within the latter, African species of Mabuya first diverged from the remainder, leaving the Asian congeners together with the Apterygodon–Dasia clade. Our results, while suggesting the non-monophyly of the genus Mabuya, do not support the currently prevailing phylogeographical hypothesis which assumes the independent origins of Lamprolepis and Lygosoma from the Asian Mabuya-like stock. On the other hand, our results suggest that morphological and karyological similarities between the Apterygodon–Dasia clade and Lamprolepis are attributable to symplesiomorphy, while their ecological similarity to convergence. Morphological and karyological character states unique to Apterygodon are supposed to have evolved from those exhibited by Dasia.

Shell morphology and length of the nonrepetitive region of the foot protein 1 (adhesive protein) amplified by the polymerase chain reaction (PCR) using oligonucleotide primers, Me 15 and Me 16, were examined in mussels collected from Asamushi water, northern Japan, and compared to those of M. galloprovincialis Lamarck from Maruishi, Hiroshima, Japan. The Asamushi mussels were collected from a suspended rope substrate and M. galloprovincialis of Maruishi from a floating dock. The mussels of the Asamushi area have been referred to as M. edulis Linnaeus or M. edulis galloprovincialis Lamarck, and were recently supposed to be M. galloprovincialis and/or M. trossulus Gould based on the geographical distributions of these species. Although 14 of 19 variables of shell characters differed significantly between mussels of Asamushi and M. galloprovincialis from Maruishi, both canonical variates analysis using eighteen shell characters and PCR product analysis showed that the mussels from Asamushi were M. galloprovincialis. The result of PCR product analysis also indicated no hybridization between M. galloprovincialis and M. trossulus in the Asamushi area.

Some Formicid ants have symbiotic intracellular bacteria in the epithelial cells of their midgut. These endosymbionts are believed to be derived from a common ancestor. A recent study revealed that endosymbionts of the ant genus Camponotus are closely related to Enterobacteriaceae, but their relationship to endosymbionts of other genera of ants is unknown. In this study, the nucleotide sequences of 16S ribosomal RNA (rRNA) of endosymbionts and mitochondrial cytochrome oxidase subunit I (COI) of their host were determined in five genera of the subfamily Formicinae (Hymenoptera: Formicidae). Based on these molecular data, we constructed phylogenetic trees in order to characterize the systematic position of the symbionts and to estimate the relationship of symbionts and hosts. The analysis showed that the endosymbionts were all connected with the Enterobacteriaceae but did not constitute a monophylitic group, while the three genera belonging to the tribe Camponotini, the endosymbionts and their hosts made a clade. The topologies of these trees were identical for the most part. These results suggest that the endosymbionts of ants have plural origins, and that in the Camponotini, ancestral symbionts have coevolved with their host ants, which are so divergent to several genera as to construct one tribe.