Zoological Science

Sperm motility is generated by a highly organized, microtubule-based structure, called the axoneme, which is constructed from approximately 250 proteins. Recent studies have revealed the molecular structures and functions of a number of axonemal components, including the motor molecules, the dyneins, and regulatory substructures, such as radial spoke, central pair, and other accessory structures. The force for flagellar movement is exerted by the sliding of outer-doublet microtubules driven by the molecular motors, the dyneins. Dynein activity is regulated by the radial spoke/central pair apparatus through protein phosphorylation, resulting in flagellar bend propagation. Prior to fertilization, sperm exhibit dramatic motility changes, such as initiation and activation of motility and chemotaxis toward the egg. These changes are triggered by changes in the extracellular ionic environment and substances released from the female reproductive tract or egg. After reception of these extracellular signals by specific ion channels or receptors in the sperm cells, intracellular signals are switched on through tyrosine protein phosphorylation, Ca²⁺, and cyclic nucleotide-dependent pathways. All these signaling molecules are closely arranged in each sperm flagellum, leading to efficient activation of motility.

Bats, Chiroptera, constitute the second largest order of the class Mammalia and vary greatly in habitats, available foods and mating systems. The timing, duration and patterns of reproduction in bats vary considerably among species and different localities. Though much is known about the reproductive phenomena and associated endocrine characteristics of various species, the central mechanism regulating the peculiar delay and asynchrony in reproductive activity remains to be elucidated. The current understanding on the endocrine characteristics and possible mechanism of regulation of the hypothalamo-adenohypophysial-gonadal axis of bats will be reviewed, based mainly on our own studies in hibernating rhinolophid bats.

We found that erythrocytes of zebrafish have cytoplasmic peroxidase activity. Blood in the zebrafish brain was visualized using a standard peroxidase staining method after formaldehyde fixation. The erythrocytes in the brain were heavily stained, but neurons and glias were not stained at all. This easy method enables the distribution of erythrocytes in the whole brain to be determined, and enables the actual number of erythrocytes in each area in the brain to be calculated.

The paths of major, thick blood vessels in zebrafish brain are similar to those in higher vertebrates, however, the distribution of thin blood vessels is different. We also found that the erythrocytes were unevenly distributed in the brain. For example, the density of erythrocytes in the surface layer of the tectum was more than 30-fold higher than in the deeper granular layer. Very few erythrocytes were found in bundles of axons like cranial nerves and the medial longitudinal fascicle. In general, fewer erythrocytes were found in areas near the ventricle, whereas many more were found closer to the surface of the brain.

The distribution of erythrocytes in the brains of sleeping, awake and actively moving fish were compared. In the brains of sleeping fish, most of the erythrocytes were present in large vessels. This was not observed in brains of awake or actively moving fish. We found that the blood supply to motor neurons in the ventral horn of the spinal cord increased during active movement compared to that in awake or sleeping fish.

In white-rumped munia, early auditory experience is critical for normal song development. New neurons are constantly added to the telencephalon in juveniles. We examined the potential role of auditory experience in regulating the developmental changes in the song nuclei and the survival of newborn cells. We chose two special days, postnatal day 23 and 37, at which we deafened the birds through bilateral cochlea removal. All birds were injected with the cell birth marker BrdU two weeks before the lesion surgeries, and then were killed two weeks or one month later. The BrdU-positive cells were distributed throughout the brain, including the high vocal center (HVC), Lobus parolfactorius and the ventricle zone (VZ) in telencephalon, the granular cell layer (GCL) of cerebellum. Moreover, these BrdU-positive cells in the GCL could self-renew. However, the nucleus robustus archistriatalis (RA) did not sprout new neurons in juvenile. In telencephalon except the VZ, 41 percent of BrdU-positive cells were NeuN-positive, too. Deafness had no significant effect on development of HVC and RA, the distribution of new cells, and the survival of new cells in telencephalon. From these data, we propose that auditory deprivation could not affect the survival of new cells of telencephalon within one month. Surprisingly, we found deafness had a complex and dramatic effect on the number of new cells in cerebellum. Deafness at postnatal day 23 could increase the number of new cells in the GCL, while deafness at postnatal day 37 decreased the number.

The physiological significance of the position and shape of the oxygen equilibrium curve (OEC) of horse hemoglobin (Hb) is considered from the viewpoint of oxygen (O₂) transport efficiency and the effectiveness of the Bohr effect. In horse fetal and maternal bloods, their physiological O₂ affinities are nearly optimized with respect to the effectiveness of the Bohr shift occurring at the O₂ release site, when it is measured by the change in O₂ saturation per unit change in P ₅₀. With relatively low cooperativity (n=2.69) of horse Hb under physiological conditions, the effectiveness of the Bohr shift for fetal blood at O₂ uptake site and maternal blood at O₂ release site is high. These facts imply that the position and the cooperativity of horse Hb OEC are optimized to receive maximal benefit from the double Bohr shift. Before exercise, the position of the OEC for adult mares is nearly optimized for the effectiveness of the Bohr shift occurring at the O₂ release site, whereas, at maximal exercise, the position of the OEC tends to become advantageous for O₂ transport efficiency.

Embryonic development of the Pacific lamprey, Entosphenus tridentatus, from Japan is described. Egg sizes averaged 1.249 mm (longest axis) and 1.145 mm (shortest axis), the time required for hatching being 11 days at 18°C, shorter than previously reported for a lower water temperature (19 days at 15°C). Early development in E. tridentatus proceeded at a similar rate to that in other lampreys, in spite of different rearing water temperatures for the latter, indicating possible specific differences in basic developmental rates.

The midgut of Apis mellifera is remodeled during metamorphosis. The epithelium and, to a lesser extent, the muscular sheath degenerate between the end of the last larval instar and the onset of pupation (prepupa). The larval epithelium is shed to the midgut lumen and digested, while a new epithelium is reconstructed from larval regenerative cells. During pupation, some reorganization still occurs, mainly in brown-eyed pupae. In pharate adult, the midgut wall shows the characteristics of adult, although some cells have pycnotic nuclei. The localization of alkaline and acid phosphatases showed that these enzymes were not involved in the reabsorption of the midgut wall.

The effects of fasting at two water temperatures (2 and 10°C) on plasma leptin- and ghrelinimmunoreactive peptide concentrations and energy metabolism were investigated in the burbot (Lota lota), a freshwater gadoid, which lives in cold waters and tolerates long periods of fasting. Burbot were assigned to fed and fasted groups at 2 and 10°C. Leptin- and ghrelin-immunoreactive peptides were identified in burbot plasma. Fasting at 2°C reduced the plasma leptin- and ghrelin-immunoreactive peptide concentrations and reduced the relative liver weights and the liver and muscle glycogen concentrations. The concentrations of the leptin- and ghrelin-immunoreactivities correlated positively with each other. At 10°C there were decreased plasma thyroxine levels in both sexes, and slightly lower plasma testosterone concentrations in males. The results support previous studies in stating that immunoreactivities resembling mammalian leptin and ghrelin can be detected in burbot plasma and liver, but the specific functions of these peptides will have to be determined in future studies.

Recent studies have shown that there are multiple genes encoding gonadotropin-releasing hormone receptor (GnRH-R) in single species. In salmonids, however, only a single gene has been identified in the rainbow trout. We therefore isolated partial cDNAs from the brain and the pituitary of masu salmon Oncorhynchus masou by reverse transcription-polymerase chain reaction and 5′-rapid amplification of cDNA ends, using primers corresponding to conserved transmembrane domains (TMs). Five different partial cDNAs were isolated from an individual and termed as msGnRH-R1, R2, R3, R4 and R5. They are divided into two groups, msGnRH-R1, R2, R3 and msGnRH-R4, R5. Two groups share 59-71% nucleotide sequence identities. Phylogenetic analysis showed that the former group is closely related to the goldfish GnRH-R GfA, and the latter to GfB. All five msGnRH-R genes were expressed in the brain and msGnRHR1, R3 and R5 were expressed in the pituitary. In addition, we found mRNA for msGnRH-R1 in the kidney and ovary, and R2 in the ovary, whereas msGnRH-R5 gene was widely expressed in the muscle, heart, kidney and testis. Differences in the expression of msGnRH-R genes between maturing and spawning fish were observed in the brain and pituitary, except for the constantly expressed msGnRH-R5. A splicing variant of msGnRH-R1 mRNA that is capable of generating a truncated GnRH-R that consists of 5TMs was also expressed in the brain, pituitary and kidney. These results indicate that five different types of putative GnRH-R gene are present and expressed in the brain of masu salmon.

The aim of this study was to obtain basic knowledge of the plasma concentrations and interactions of weight regulatory hormones in juvenile minks (Mustela vison). Ghrelin, leptin, and growth hormone (GH) levels were validated and determined by radioimmunoassay methods from the plasma of 30 female and 30 male minks. The female minks had higher plasma ghrelin and GH levels than the males. The plasma ghrelin concentrations of the females correlated positively with their body masses (BMs). The plasma leptin levels did not differ between sexes, but there was a positive correlation between the plasma leptin concentrations and BMs in the male minks. When the data from the male and female minks were combined, the correlation between the leptin levels and the BMs was still clear, but this was not observed in the females alone. In the male minks, the plasma GH levels correlated positively with the BMs and with the plasma leptin concentrations. However, there was no correlation between the plasma ghrelin and GH or leptin concentrations. The hormone concentrations were quite similar to earlier measurements in other carnivores.

To investigate the role of serotonin (5-HT) receptor 1A or 7 in regulating lordosis behavior in female rats, ovariectomized rats were treated with 3 kinds of receptor agonists and lordosis behavior was observed. The injected agents were the selective 5-HT_1A receptor agonist, buspirone (BUS), the highly selective 5-HT_1A receptor agonist, 8-hydroxy-2-(di-n-propylamino)tetralin ((±)8-OH-DPAT), and the 5-HT_1A and 5-HT₇ receptor agonist, (R)-8-hydroxy-2-(di-n-propylamino)tetralin ((+)8-OH-DPAT). A behavioral test was performed after ovariectomy and subcutaneous implantation of a silicon tube containing estradiol. Female rats in which the lordosis quotient (LQ) was over 70 were intraperitoneally injected with several doses of these agents. As a result, in the BUS group, the dose of 3 mg/kg bw, but not 1 mg/kg was effective for suppressing lordosis. On the other hand, an inhibitory effect was observed from 0.25 mg/kg and 0.5 mg/kg in the (+)8-OH-DPAT and (±)8-OH-DPAT groups, respectively. In the time-course experiment, in all drug-treated groups, LQ decreased to lower than 20 after 15 min and low LQ continued for 1 hr at least. Measurement of locomotor activity using an infrared sensor system showed no relation between the decrease in lordosis by these agents and spontaneous locomotion. These results indicate that 5-HT_1A is strongly involved in the lordosis-inhibiting circuit of the serotonin neurons.

We cloned cDNAs encoding PC1 and PC2 from a cDNA library constructed for the anterior pituitary gland of the bullfrog (Rana catesbeiana) and sequenced them. The bullfrog PC1 cDNA consisted of 2972 base pairs (bp) with an open reading frame of 2208 bp and encoded a protein of 736 amino acids, including a putative signal peptide of 26 amino acids. The protein showed a high homology to R. ridibunda PC1 (95.1%) and mammalian PC1 (72.6%). The bullfrog PC2 cDNA consisted of 2242 bp with an open reading frame of 1914 bp and encoded a protein of 638 amino acids, including a putative signal peptide of 23 amino acids. This protein showed a high homology to R. ridibunda PC2 (95.5%) and mammalian PC2 (84.8%). The catalytic triad of serine proteinases of the subtilisin family was found at Asp-168, His-209, and Ser-383 in the PC1 protein and at Asp-167, His-208, and Ser-384 in the PC2 protein. In situ hybridization staining revealed that PC2 mRNA was detected in corticotrope cells of the tadpoles, but not in those of the adults. In the adult, only PC1 mRNA was detected in the pars distalis but both PC1 and PC2 mRNAs were detected in the pars intermedia. The data also showed that PC1 mRNA was expressed in gonadotrope cells.

The morphology and life history of a strange and unidentified botryllid ascidian were investigated. This ascidian was first collected from the stony shore of Ebisu Island in Shimoda, a city on Izu peninsula in central Japan. Unlike other botryllid ascidians, whose colonies are flat and smooth, this ascidian's colonies are rugged. In each colony, zooids are arranged into several oval systems, each of which has a thick part containing zooids and very thin parts that do not. The arrangement of ovary and testis in this species is the same as in other species of the genus Botryllus; the ovary is situated anterior to the testis. The embryo of this ascidian develops in the peribranchial cavity of its mother zooid without any brooding organs, as is the case with Botryllus scalaris and Botryllus puniceus. Meanwhile, the results of cut colony assay experiments did not show the existence of colony specificity in this ascidian. Even when two syngeneic colonies were brought into contact at their growing edges, none fused together. On the other hand, when two colonies were brought into contact with each other at their cut surfaces, they always fused into a single colony, regardless of their origin. Therefore, this species may be the only species that lacks colony specificity among the botryllids studied so far.

A new species of salamander, Hynobius yangi, is described from Kori (=Hyoam-ri), Busanshi, southeastern Korea. It is a lentic breeder belonging to the H. nebulosus species group, and has long been confused with another Korean species H. leechii. The new species, however, is genetically substantially differentiated from Korean and Japanese relatives. Hynobius yangi is morphologically very similar to H. leechii, but could be differentiated from it by the tail shape, degree of limb separation, shape of vomerine teeth series, shape of egg sac, and dorsal coloration.

A new opheliid polychaete, Euzonus japonicus sp. nov., is described. This species was collected from subtidal zones in Japanese coasts, while most Euzonus species inhabit intertidal sandy beaches. E. japonicus sp. nov. is morphologically most similar to another subtidal species, E. flabelliferus (Ziegelmeier, 1955) collected from northern Europe, but different from that in the form of branchiae. They share a unique characteristic on setiger 10, i. e., a pair of lateral transverse rows of conical cirri, instead of a pair of lateral smooth ridges that are common to all intertidal Euzonus species.