KISS1 Gene Expression in the Developing Brain of Female Pigs in Pre- and Peripubertal Periods

Puberty is associated with an increase in gonadotropin secretion as a result of an increase in gonadotropin-releasing hormone (GnRH) secretion. Kisspeptin is considered to play a key role in puberty onset in many mammalian species, including rodents, ruminants and primates. The present study aimed to determine if changes in hypothalamic expression of the KISS1 gene, encoding kisspeptin, are associated with the onset of puberty in pigs. The animals (n=4 in each group) were perfused with 4% paraformaldehyde at 0, 1, 2, 3 and 4 months old, as prepubertal stages, and at 5 months old, as the peripubertal stage, following each blood sampling. KISS1 gene expressions in coronal sections of brains were visualized by in situ hybridization. Plasma luteinizing hormone (LH) was measured by radioimmunoassay. KISS1 mRNA signals were observed in the arcuate nucleus (ARC) at all ages examined without any significant difference in the number of KISS1-expressing cells, indicating that the KISS1 gene is constantly expressed in the ARC throughout pubertal development in pigs. The plasma LH concentration was the highest in 0-month-old piglets and significantly decreased in the 1- and 2 month-old groups (P<0.05), suggesting a developing negative feedback mechanism affecting gonadotropin release during the prepubertal period. Considering the potent stimulating effect of kisspeptin on gonadotropin release in prepubertal pigs, kisspeptin secretion rather than kisspeptin synthesis may be responsible for the onset of puberty in pigs.     

Differential Effects of Continuous Exposure to the Investigational Metastin/Kisspeptin Analog TAK-683 on Pulsatile and Surge Mode Secretion of Luteinizing Hormone in Ovariectomized Goats

The aim of the present study was to determine if the estradiol-induced luteinizing hormone (LH) surge is influenced by the constant exposure to TAK-683, an investigational metastin/kisspeptin analog, that had been established to depress the pulsatile gonadotropin-releasing hormone (GnRH) and LH secretion in goats. Ovariectomized goats subcutaneously received TAK-683 (TAK-683 group, n=6) or vehicle (control group, n=6) constantly via subcutaneous implantation of an osmotic pump. Five days after the start of the treatment, estradiol was infused intravenously in both groups to evaluate the effects on the LH surge. Blood samples were collected at 6-min intervals for 4 h prior to the initiation of either the TAK-683 treatment or the estradiol infusion, to determine the profiles of pulsatile LH secretion. They were also collected at 2-h intervals from –4 h to 32 h after the start of estradiol infusion for analysis of LH surges. The frequency and mean concentrations of LH pulses in the TAK-683 group were remarkably suppressed 5 days after the start of TAK-683 treatment compared with those of the control group (P<0.05). On the other hand, a clear LH surge was observed in all animals of both groups. There were no significant differences in the LH concentrations for surge peak and the peak time of the LH surge between the TAK-683 and control groups. These findings suggest that the effects of continuous exposure to kisspeptin or its analog on the mechanism(s) that regulates the pulsatile and surge mode secretion of GnRH/LH are different in goats.     

Effects of STX,a Novel Estrogen Membrane Receptor Agonist,on GnRH-Induced Luteinizing Hormone Secretion from Cultured Bovine Anterior Pituitary Cells

STX is an agonist for a recently characterized membrane estrogen receptor whose structure has not been identified. We evaluated whether STX suppresses gonadotropin-releasing hormone (GnRH)–induced luteinizing hormone (LH) release from bovine anterior pituitary (AP) cells. We cultured AP cells (n=12) for 3 days in steroid-free conditions, followed by increasing concentrations (0.001, 0.01, 0.1, 1 and 10 nM) of 17β-estradiol or STX for 5 min before GnRH stimulation until the end of the experiment. Estradiol (0.001 to 0.1 nM) significantly suppressed GnRH-stimulated LH secretion, whereas STX did not affect GnRH-stimulated LH secretion at any of the tested concentrations. In conclusion, STX, unlike estradiol, possesses no suppressive effect on GnRH-induced LH release from bovine AP cells.     

Changes in the Immunolocalization of Steroidogenic Enzymes and the Androgen Receptor in Raccoon (Procyon lotor) Testes in Association with the Seasons and Spermatogenesis

The raccoon is a seasonal breeder with a mating season in the winter. In a previous study, adult male raccoons exhibited active spermatogenesis with high plasma testosterone concentrations, in the winter mating season. Maintenance of spermatogenesis generally requires high testosterone, which is produced by steroidogenic enzymes. However, even in the summer non-mating season, some males produce spermatozoa actively despite low plasma testosterone concentrations. To identify the factors that regulate testosterone production and contribute to differences in spermatogenetic activity in the summer non-mating season, morphological, histological and endocrinological changes in the testes of wild male raccoons should be known. In this study, to assess changes in the biosynthesis, metabolism and reactivity of testosterone, the localization and immunohistochemical staining intensity of four steroidogenic enzymes (P450scc, P450c17, 3βHSD, P450arom) and the androgen receptor (AR) were investigated using immunohistochemical methods. P450scc and P450c17 were detected in testicular tissue throughout the year. Seasonal changes in testosterone concentration were correlated with 3βHSD expression, suggesting that 3βHSD may be important in regulating the seasonality of testosterone production in raccoon testes. Immunostaining of P450arom and AR was detected in testicular tissues that exhibited active spermatogenesis in the summer, while staining was scarce in aspermatogenic testes. This suggests that spermatogenesis in the raccoon testis might be maintained by some mechanism that regulates P450arom expression in synthesizing estradiol and AR expression in controlling reactivity to testosterone.