Dynamics of testis-ova in a wild population of Japanese pond frogs, Rana nigromaculata.

Although many studies have reported the occurrence of testis-ova in wild frog populations, the origin and trigger of testis-ova differentiation/development remain unclear. A high frequency of testis-ova has been previously reported for wild populations of the Japanese pond frog, Rana nigromaculata (cf. Iwasawa and Asai, '59). In the present study, we aimed to clarify the dynamics of testis-ova in this frog species, including the origin and artificial induction of testis-ova. Testis-ova were observed in both mature frogs and puberty-stage frogs (i.e., 0- and 1-year-old frogs). However, the early stages of testis-ova (~pachytene stage) were mostly observed in puberty-stage male frogs at the onset of spermatogenesis. The early stages of testis-ova were observed in the cysts of early secondary spermatogonia and the single cysts of the primary spermatogonium. This finding indicates that testis-ova differentiation occurs during spermatogonial proliferation and that it is correlated with the initiation of spermatogenesis. We also examined whether estrogen exposure induced testis-ova differentiation and how it is correlated with the progression of spermatogenesis. When 1-year-old frogs were exposed to estradiol-17ß during spring (i.e., when spermatogenesis was initiated), testis-ova differentiation was induced in a dose-dependent manner. However, this phenomenon did not occur in 1-year-old frogs during summer, (i.e., when the transition from spermatogonia to spermatocytes mainly occurs). These results present the first evidence that testis-ova of the Japanese pond frog are derived from primary and early secondary spermatogonia, and that estrogen exposure induces testis-ova differentiation accompanied by the initiation of spermatogenesis.

Unstimulated amylase secretion is proteoglycan-dependent in rat parotid acinar cells.

It is well-known that amylase is secreted in response to extracellular stimulation from the acinar cells. However, amylase is also secreted without stimulation. We distinguished vesicular amylase as a newly synthesized amylase from the accumulated amylase in secretory granules by short time pulse and chased with (35)S-amino acid. The newly synthesized amylase was secreted without stimulation from secretory vesicles in rat parotid acinar cells. The secretion process did not include microtubules, but was related to microfilaments. p-Nitrophenyl beta-xyloside, an inhibitor of proteoglycan synthesis, inhibited the newly synthesized amylase secretion. This indicated that the newly synthesized amylase was secreted from secretory vesicles, not via the constitutive-like secretory route, which includes the immature secretory granules, and that proteoglycan synthesis was required for secretory vesicle formation.

Existence of subtypes of gustducin-immunoreactive cells in the vallate taste bud of guinea pigs.

Vallate taste buds in the guinea-pig tongue were immunohistochemically investigated with regard to the colocalization of gustducin with calbindin-D28K (=spot 35 protein) and type III inositol triphosphate receptor (IP(3)R-3) in order to characterize gustducin-immunoreactive cells. Individual taste bud cells ranged from totally immunopositive to totally immunonegative for these three molecules. Among the immunoreactive cells, gustducin-immunoreactive cells were divided into two cell populations: one immunopositive and the other immunonegative for calbindin-D28K. Applying our previous data to the present results, the former cells should belong to Type III cells designated by electron microscopy. This finding provides new evidence regarding the taste bud types of cells expressing gustducin in the guinea pig.

Seasonal expression of LHbeta and FSHbeta in the male newt pituitary gonadotrophs.

Seasonal changes in LHbeta and FSHbeta mRNA levels were examined in the pituitary gland of the adult male newt, Cynops pyrrhogaster, using in situ hybridization histochemistry and a quantitative real-time RT-PCR method. The annual fluctuation of LHbeta mRNA and FSHbeta mRNA levels in the pituitary gland displayed a close relationship with seasonal changes in testicular function. The values obtained by both methods showed similar fluctuation. The levels of LHbeta mRNA were always exceeded those of FSHbeta. The present immunoelectron microscopic observations support the data on the gene expression levels of the beta-subunits of LH and FSH. Gonadectomy in the summer increased the LHbeta and FSHbeta mRNA levels. Testosterone replacement inhibited the expression of LHbeta mRNA, but not of FSHbeta mRNA, suggesting that the expression of FSHbeta is regulated by some non-steroid factor, probably inhibin. In the case of gonadectomy during any other season, the LHbeta mRNA level increased, but not to the same extent as in summer, and androgen concentrations decreased to the minimum of the year. This finding provides new information about the regulation of annual changes in LHbeta and FSHbeta expression in the pituitary gonadotrophs.

Comparative morphological studies on the stereo structure of the lingual papillae of selected primates using scanning electron microscopy.

A scanning electron microscope was used to observe the lingual papillae and their connective tissue cores (CTCs) in five primates (tupai, tamarin, crab-eating monkey, mandrill, and human). There were some slender protrusions rising from the top of the filiform papilla in all five types of primate. After removing the epithelium the filiform CTC from the tupai, tamarin and crab-eating monkey displayed a U-shaped arrangement of rod-shaped protrusions. The filiform CTC from the crab-eating monkey also had a columnar base. The human filiform CTC consisted of a primary columnar base, numerous short rod-shaped secondary protrusions from its upper periphery, and a few central protrusions. The filiform CTC from the Mandrill was fundamentally similar to that of the human, however, its base was shorter. The fungiform CTC from the tupai was column shaped, with several depressions for taste buds on the top. There were three vallate papillae in the tupai, tamarin, and mandrill, approximately four in the monkey, and between five and twelve in the human. Moderately developed foliate papillae were found in the tamarin, monkey, mandrill and human. The tupai, however, possessed a finger-like lateral organ instead. The lingual root area of the tupai, tamarin, crab-eating monkey and mandrill was relatively small with a smooth surface. Only the human had a tonsil-structure, which was located on the surface of its larger lingual root.

Villin is a possible marker of receptor cells in frog taste organs.

We investigated lingual taste organs of four frog species mainly by means of fluorescence immunohistochemistry for villin, calbindin, and serotonin. Cells immunoreactive for villin appeared in the taste organs of all the species used. These villin-immunostained cells were basoapically elongated in shape and extended up to the apical surface. They were also immunoreactive for calbindin. On the other hand, serotonin-immunoreactive cells, identified as Merkel-like basal cells, were immunonegative for villin. Considering the present results combined with those of studies by other research groups, the villin-immunostained cells were postulated to function as taste receptors.

Comparative morphological study of the lingual papillae and their connective tissue cores of the koala.

The stereo structure of each lingual papilla of the koala has a similar structure to that of various other animal species: the koala has a lingual prominence (intermolar prominence) with larger filiform papillae. (A lingual prominence is a characteristic in herbivorous animals.) The external form and connective tissue core (CTC) of the filiform papillae of koalas consist of one large main process and several smaller accessory processes. (These are similar to carnivorous animals.) Fungiform CTC have a thick dome-like structure, with several taste buds on the top. There are three vallate papillae: one central midline and two laterally located vallate papillae. The central vallate papilla has a posterior pouch lined with ciliated and non-ciliated epithelial cells. Long conical papillae are distributed in the posterior lateral area where foliate papillae are distributed in many other animal species. (Finger-like papillae are seen in dog and cat instead of foliate papillae.) It may be suggested that the tongue of the koala evolved in a special environment in Australia. Even though it has still retained special features similar to those of carnivorous cats and dogs it has evolved to resemble the tongues of herbivorous animals.