Zoological Science

Western sand lance, Ammodytes japonicus, is known to have an estivation period, in which they cease feeding and stay in the sand from early summer to late autumn, followed by gonadal maturation. During the feeding period prior to estivation, they swim in daytime and spend the night in the sand. Before they start swimming, they show a typical behavior of head-exposing from the sand, which is likely to be related to foraging and predation avoidance. Our previous study revealed that melatonin regulates such diel behavior of this species. To elucidate the mechanisms of behavioral regulation throughout the life cycle of this sand lance, the present study examined the changes in behavior and melatonin secretion toward the estivation period. Both head-exposing and swimming behaviors were frequently observed at the transition period toward estivation. On the other hand, occurrence of these behaviors was suppressed just before entering estivation. Subsequently, it was found that plasma melatonin concentration was about three times higher at night than in daytime in the non-estivation period, while it was retained at high levels throughout the day in the estivation period. These results indicate that diurnal swimming behavior of sand lance from the feeding to estivation periods is associated with the daily cycle of melatonin secretion.

The east coast of the Indochinese Peninsula is a well-known transition zone from subtropical to tropical systems, yet only a small number of studies have been conducted on the biogeography and phylogeography of aquatic organisms in this region. The Hau Giang medaka, Oryzias haugiangensis, was originally described from the Mekong Delta in southern Vietnam, and later reported also from southeastern Thailand, west of the Mekong Delta region. However, the species' full geographic range and population genetic structures remain unknown. Field surveys showed a widespread distribution of this species along the east coast of the Indochinese Peninsula, as far as northern Vietnam. A mitochondrial gene phylogeny and population genetic structure analysis using genome-wide single nucleotide polymorphisms revealed that the populations of O. haugiangensis are highly structuralized along the east coast of Vietnam, with the southernmost Mekong Delta population clearly separated from three populations north of central Vietnam. Further field collections are necessary to determine the boundary between the southern and northern populations, and the presence or absence of a hybrid zone.

The grey-headed lapwing (Vanellus cinereus) is a wading species in East Asia. However, examples of regional population dynamics and genetic research are limited. To reconsider the natural history and current status of the grey-headed lapwing in Japan, we analyzed the genetic diversity of the Japanese grey-headed lapwing population. We collected 77 grey-headed lapwing samples from 12 locations across Japan during the breeding season and three individuals during the wintering season and extracted DNA; 496-bp sequences of the ND2, which form part of the mitochondrial DNA, were determined for genetic analysis of the population. Consequently, 10 haplotypes were detected in 80 individuals, and 67 individuals, 84% of the total, shared two haplotypes, namely Vc1 and Vc2. Furthermore, the results showed that the prevalence of Vc1 was higher mainly in northern Japan, while that of Vc2 was higher mainly in southern Japan. Genetic diversity analysis showed that the overall haplotype diversity in Japan was 0.617, which is not particularly low. The sequence of Vc1 was exactly the same as that of grey-headed lapwing in China. Our study revealed the genetic structure of the grey-headed lapwing, suggesting that as the grey-headed lapwing expanded its distribution area into southern Japan, many Vc2-positive individuals migrated southward, resulting in a higher detection rate of Vc2 in southern Japan.

cytochrome P-450, 21-hydroxylase (cyp21a2), encodes an enzyme required for cortisol biosynthesis, and its mutations are the major genetic cause of congenital adrenal hyperplasia (CAH) in humans. Here, we have generated a null allele for the medaka cyp21a2 with a nine base-pair insertion which led to a truncated protein. We have observed a delay in hatching and a low survival rate in homozygous mutants. The interrenal gland (adrenal counterpart in teleosts) exhibits hyperplasia and the number of pomca-expressing cells in the pituitary increases in the homozygous mutant. A mass spectrometry-based analysis of whole larvae confirmed a lack of cortisol biosynthesis, while its corresponding precursors were significantly increased, indicating a systemic glucocorticoid deficiency in our mutant model. Furthermore, these phenotypes at the larval stage are rescued by cortisol. In addition, females showed complete sterility with accumulated follicles in the ovary while male homozygous mutants were fully fertile in the adult mutants. These results demonstrate that the mutant medaka recapitulates several aspects of cyp21a2-deficiency observed in humans, making it a valuable model for studying steroidogenesis in CAH.

Aposematic coloration plays a crucial role in animal defense, and it is shaped by a complex interplay of factors such as physiological limitations and sexual and natural selection. Warty newts within the genus Paramesotriton exhibit significant variation in ventral coloration. In this study, we quantified the percentage of red ventral area to investigate aposematic ventral coloration in Paramesotriton deloustali and P. guangxiensis across eight populations in northern Vietnam. To assess the interaction between predators and the aposematic signals, we conducted experiments employing three types of clay replicas of newts: dorsal, red ventral, and black ventral models. Our findings revealed a significant variation in the red ventral area among different populations. Additionally, a significant correlation was detected between the red ventral area of the newt and the annual temperature range. In clay model experiments, a significant difference in predator attack rates was observed between dorsal and ventral clay models. Interestingly, there was no significant difference in attack rates between red and black ventral types. Our study suggested that the variation in the red ventral area of warty newts is probably influenced by multiple factors, including genetic constraints, sex, ambient environment, and diet. Furthermore, our results supported the effectiveness of displaying aposematic coloration as an antipredator defense mechanism in warty newts. However, variations in body size and the pressure of mammal predation might not play a significant role in determining aposematic coloration.

Platyhelminthes are a phylum of simple bilaterian invertebrates with prototypic body systems. Compared with non-bilaterians such as cnidarians, the bilaterians are likely to exhibit integrated free-moving behaviors, which require a concentrated nervous system “brain” rather than the distributed nervous system of radiatans. Marine flatworms have an early cephalized ‘central’ nervous system compared not only with non-bilaterians but also with parasitic flatworms or freshwater planarians. In this study, we used the marine flatworm Stylochoplana pusilla as an excellent model organism in Platyhelminthes because of the early cephalized central nervous system. Here, we investigated the three-dimensional structures of the flatworm central nervous system by the use of X-ray micro-computed tomography (micro-CT) in a synchrotron radiation facility. We found that the obtained tomographic images were sufficient to discriminate some characteristic structures of the nervous system, including nerve cords around the cephalic ganglion, mushroom body-like structures, and putative optic nerves forming an optic commissure-like structure. Through the micro-CT imaging, we could obtain undistorted serial section images, permitting us to visualize precise spatial relationships of neuronal subpopulations and nerve tracts. 3-D micro-CT is very effective in the volume analysis of the nervous system at the cellular level; the methodology is straightforward and could be applied to many other non-model organisms.

Among the six mitochondrial DNA lineages of the black rat (Rattus rattus Complex; RrC), lineages II and IV are widespread in Southeast and East Asia. This study explored their demographic history using 17 new sequences from the Miyako Islands in the Ryukyu archipelago, together with 178 publicly available cytochrome b sequences. We defined six and two haplotype groups showing rapid population expansion signals in Lineages II and IV, respectively. The six haplotype groups of Lineage II were represented by haplotypes from 1) Myanmar/Bangladesh/Northeast India, 2) Laos, 3) Thailand, 4) Indonesia/Philippines, 5) Vietnam/southern China, and 6) the Ryukyu archipelago. These expansion times were estimated using time-dependent evolutionary rates to be 115,300 years ago (ya), 128,500 ya, 9600 ya, 10,600 ya, 7200 ya, and 1400 ya, respectively, although all had large confidence intervals. The two groups of Lineage IV were recovered from the mainland and islands of Southeast Asia with predicted expansion times of 197,000 ya and 5800 ya, respectively. These results suggest that climatic fluctuations during the last 200,000 years of the Quaternary, affected the population dynamics in subtropical areas at different times. Furthermore, the results of the younger rapid expansion events of RrC suggest the possibility of agricultural advancement and dispersal of Neolithic farmers to different areas within the mainland and islands of Southeast Asia during the Holocene. A subset of rats from the Miyako Islands were found to have the same lineage IV haplotypes as those in Southeast Asia, suggesting a recent introduction of these new lineages.

Sperm-specific cation channel (CatSper), sperm-specific Na ⁺ /H ⁺ exchanger (sNHE), and soluble adenylyl cyclase (sAC) are necessary in the signaling pathways to control sperm motility in many animals, whereas some animals have lost some or all of them. In the present study, we examined CatSper-uninvolved signaling for vigorous undulation of the undulating membrane that is attached to the sperm tail and gives thrust for forward motility in the internally fertilizing newt Cynops pyrrhogaster. Reverse-transcription PCR failed to detect sNHE in the newt sperm. However, the pH of sperm cytoplasm was raised under a high extracellular pH equivalent to that of egg jelly, where sperm motility is initiated by sperm motility-initiating substance (SMIS). Carbonic anhydrase XII/ XVI and SLC4A4/8 were suggested to be present in the sperm, and transported bicarbonates raised the intracellular pH. In egg jelly extract that contained SMIS, the anion transporter inhibitor DIDS weakened the undulation of the undulating membrane, while bicarbonates enhanced it. The cyclic AMP concentration was found to increase in sperm cytoplasm in the egg-jelly extract. An inhibitor of sAC (KH7) weakened the undulation of the undulating membrane, and dibutyryl cyclic AMP blocked the inhibitory effect. Inhibitor of transmembrane AC (DDA) limitedly affected the undulation. The undulation was weakened by an inhibitor of protein kinase A (H89), and by an inhibitor of transient receptor potential (TRP) channels (RN1747). Our results support the conclusions that the high pH of the egg jelly triggers a signaling pathway through sAC, PKA, and TRP channels, and coacts with SMIS to induce forward sperm motility.

Formation of the synaptonemal complex (SC) is a prerequisite for proper recombination and chromosomal segregation during meiotic prophase I. One mechanism that ensures SC formation is chromosomal movement, which is driven by the force derived from cytoskeletal motors. Here, we report the phenotype of medaka mutants lacking the telomere repeat binding bouquet formation protein 1 (TERB1), which, in combination with the SUN/KASH protein, mediates chromosomal movement by connecting telomeres and cytoskeletal motors. Mutations in the terb1 gene exhibit defects in SC formation in medaka. Although SC formation was initiated, as seen by the punctate lateral elements and fragmented transverse filaments, it was not completed in the terb1 mutant meiocytes. The mutant phenotype further revealed that the introduction of double strand breaks was independent of synapsis completion. In association with these phenotypes, meiocytes in both the ovaries and testes exhibited an aberrant arrangement of homologous chromosomes. Interestingly, although oogenesis halted at the zygotene-like stage in terb1 mutant, testes continued to produce sperm-like cells with aberrant DNA content. This indicates that the mechanism of meiotic checkpoint is sexually different in medaka, similar to the mammalian checkpoint in which oogenesis proceeds while spermatogenesis is arrested. Moreover, our results suggest that spermatogenesis is mechanistically dissociable from meiosis.

Species in the anthurid isopod genus Mesanthura have specific, dorsal dark pigmentation patterning on the body. Though Mesanthura species have traditionally been distinguished mainly by differences in the dorsal pigmentation pattern in females, the stability of the pigmentation pattern within species had not been investigated, and information was lacking on ontogenetic variation in the pattern. Our study showed the following for M. miyakoensis. (1) Mancae begin to show dorsal pigmentation in the marsupium roughly 9 days before their release. (2) The pigmentation pattern in the first-instar mancae (first free-living stage) differs from that in later instars. (3) The pigmentation pattern in females is discrete and stable from putative second-instar mancae through females lacking oostegites, and distorted but recognizable in ovigerious females. (4) The pattern in males is different from and less discrete than that in females; it remains similar through the molt from subadult to adult male but changes markedly with age, leading to heavy pigmentation of the body. (5) The pigmentation pattern in mancae and females remains stable and observable after storage in ethanol for at least 13.7 months. Our results suggest that comparisons of pigmentation pattern across species in Mesanthura taxonomy should be restricted to females in the post-manca or later stages.