Comparative changes in the serum concentrations of inhibin-B, prolactin, gonadotropins and steroid hormones at different reproductive States in domestic Turkey hens.

The present study was undertaken to compare the changes in circulating levels of inhibin-B, prolactin, follicle-stimulating hormone (FSH), luteinizing hormone (LH), estradiol-17beta, progesterone and testosterone during the different reproductive states of turkey hens. Blood samples were collected during different reproductive states, at laying, incubating and out of lay. Inhibin-B was measured by ELISA, while other hormones were determined by Chemiluminescent Microparticle Immunoassay (CMIA). The results revealed highly significant differences among the hen's states for all serum hormone concentrations. The highest levels of inhibin-B and prolactin were observed in incubating hens, while the lowest values were observed in laying hens. In contrast, the highest levels of FSH, LH, estradiol-17beta, progesterone and testosterone were found in the laying group, while the lowest values were found in the incubating group. The progesterone level was higher in the laying group compared with the other groups. These results clearly demonstrate that negative correlation was found between both the inhibin-B and prolactin levels and the gonadotropin and steroid hormone concentrations during the different reproductive states of the turkey hens. In addition, the results suggest that inhibin-B may be involved in control of FSH and LH secretion.

Effect of echo-guided high-intensity focused ultrasound ablation on localized experimental tumors.

We fabricated a prototype 3.25-MHz split-focus therapeutic transducer combined with a small 6.5-MHz imaging ultrasonic probe for transrectal treatment of prostate cancer and evaluated the feasibility of using split-focus high-intensity focused ultrasound (HIFU) to ablate localized tumor tissue without injuring the surrounding organs. We therefore established a localized tumor model by inoculating VX2 tumor into rabbit livers. The localized VX2 tumors of nine rabbits were transdermally treated with split-focus ablation at a peak intensity in water of 6 kW/cm2 for 4 s (6 shots) under the guidance of ultrasonic B-mode imaging. Necropsy a day after treatment found the surface of the livers and gastrointestinal tracts to be grossly normal. The VX2 tumors were completely coagulated and were surrounded by ablated liver tissue. The six shots of split-focus HIFU destroyed the VX2 tumors without injuring the liver surfaces or the surrounding organs. These results suggest that split-focus HIFU ablation could be an effective treatment of localized tumors.

Immunization of goats against inhibin increased follicular development and ovulation rate.

In the present study, two experiments were conducted to induce superovulation in goats using passive and active immunization against inhibin. In the first experiment, two groups of goats were given an intravenous injection of either 10 ml normal goat serum (control; n=6) or inhibin antiserum developed against [Tyro30]-inhibin alpha (1-30) (passively immunized; n=6) 48 h before treatment with PGF2alpha. In the second experiment, two groups of goats were immunized with inhibin vaccine (actively immunized; n=5) or Freund's adjuvant (control; n=5) followed by three booster immunizations at 4 week intervals. Blood samples were collected for determination of FSH, LH, estradiol-17beta, and progesterone. Ultrasonography was used to determine ovarian activity at PGF2alpha injection and ovulation rate one week after estrus. In both experiments, there was a significant increase in plasma FSH concentration compared with the controls. However, the pattern of the FSH levels was different between the passively and actively immunized goats. The numbers of follicles in passively and actively immunized goats (22.4 +/- 2.3 and 18.6 +/- 2.1, respectively) were significantly greater than those in the controls (2.6 +/- 0.4 and 2.3 +/- 0.4, respectively). In addition, the ovulation rate was greater in the immunized animals compared with the controls. Therefore, either passive or active immunization against inhibin could be used to induce superovulation in goats.

Changes in serum inhibin levels and immunolocalization of inhibin/activin subunits during the breeding season in the wild male Japanese black bear (Ursus thibetanus japonicus).

The objective of this study was to investigate the changes in secretion of inhibin and cellular localization of inhibin alpha and inhibin/activin (beta(A) and beta(B)) subunits during the breeding season in the wild male Japanese black bear. Histological observations of testes were performed and seminiferous tubule diameters were measured. The sections of the testes were immunostained by the avidin- biotin-peroxidase complex method (ABC) using polyclonal antisera raised against porcine inhibin alpha, inhibin/ activin beta(A), and inhibin/activin beta(B) during the breeding season. Serum concentrations of immunoreactive (ir-)inhibin, testosterone, and luteinizing hormone (LH) were measured by radioimmunoassay. Higher values of seminiferous tubule diameters and all types of spermatogenic cells including mature-phase spermatozoa were found during the breeding season. There were seasonal changes in serum concentrations of ir-inhibin, testosterone, and LH. Ir-inhibin was positively correlated with testosterone, and LH. In addition, immunoreactivity of inhibin alpha, beta(A), and beta(B) subunits were also detected in Sertoli and Leydig cells during the breeding season. These results suggest that Japanese black bear testes may secrete bioactive inhibins during the breeding season and that the circulating inhibin may be a useful indicator of the testicular function in wild male Japanese black bears.

Seasonal changes in immunolocalization of inhibin/activin subunits and testicular activity in wild male raccoon dogs (Nyctereutes procyonoides).

Thirty-four pairs of testes from wild adult raccoon dogs (Nyctereutes procyonoides) were obtained between September 2000 and May 2003. The cellular localization of the inhibin alpha and inhibin/activin (betaA and betaB) subunits in wild raccoon dog testes was investigated. The testicular weight and size and seminiferous tubule diameters were measured. There were marked seasonal variations in testicular weight and size and seminiferous tubule diameters, with values relatively low in September and high in March. Spermatogonia and primary spermatocytes were observed in September, and spermatogonia, spermatocytes, and round spermatids were present in January. All types of spermatogenic cells, including mature spermatozoa, were found in March, indicating that the breeding season is around March in Japan. Thereafter, spermatogonia and degenerating spermatocytes were observed in April. The sections of testes were immunostained by the avidin-biotin-peroxidase complex method (ABC) using polyclonal antisera raised against porcine inhibin alpha, inhibin/activin betaA and inhibin/activin betaB. The inhibin alpha and inhibin/activin (betaA and betaB) subunits were only expressed in Leydig cells in September. On the other hand, the inhibin alpha, betaA, and betaB subunits were observed in Leydig cells and Sertoli cells, but not in germ cells, in March. These results suggest that the testes of wild raccoon dogs have the ability to synthesize inhibins, and the cellular localization of inhibin/activin subunits showed season-related changes in the breeding and non-breeding seasons.

Effects of experimental cryptorchidism on sperm motility and testicular endocrinology in adult male rats.

The effect of induced cryptorchidism on testicular function and sperm motility was investigated. Bilateral cryptorchidism was created surgically in adult male rats (treated group), and sham-operated rats were used as a control group. Five rats from each group were sacrificed on days 1, 3, 5, and 7 after surgery. The percentage of motile spermatozoa began to decrease 1 day after the operation, followed by an abrupt decline 3 and 5 days later in cryptorchid rats. Furthermore, there were significant decreases in the other sperm motility parameters 5 days after inducement of cryptorchidism. In cryptorchid rats, plasma concentrations of LH, FSH, testosterone, and inhibin B were significantly lower than in the control group 1 day after the operation. Thereafter, plasma concentrations of LH, FSH, and testosterone gradually increased in the cryptorchid rats. On the other hand, plasma concentrations of inhibin B showed a further decline from day 3 after the operation onward. Concentrations of immunoreactive (ir)-inhibin, but not testosterone, in testicular interstitial fluid were remarkably increased until 3 days after surgery in the cryptorchid rats, and declined thereafter. Testicular response to human chorionic gonadotropin (hCG) for testosterone release was decreased in the cryptorchid rats compared with the control rats, indicating that heat stress to testes resulted in a reduction of the activity of Leydig cells and Sertoli cells. These results clearly indicate that heat stress to the testes resulted in a significant reduction of sperm activity within 3 days, and this was followed by changes in testicular endocrine function.

The effect of active immunization against inhibin on gonadotropin secretions and follicular dynamics during the estrous cycle in cows.

The hypothesis of the present study is that active immunization of cows against inhibin would neutralize endogenous inhibin, increase circulating levels of follicle stimulating hormone, and subsequently affect follicular dynamics and the ovulation rate during the estrous cycle. Thirteen cows were immunized against inhibin alpha-subunit and, 6 cows were immunized with a placebo. Both groups were given 4 booster immunizations 7, 14, 21, and 34 weeks after the primary injection. Ovaries were examined daily after the 2nd, 3rd, and 4th booster immunizations by transrectal ultrasonography for 25 days. After the 4th booster immunization, blood samples were collected daily for one complete estrous cycle to measure FSH and LH. The results showed that the immunized cows generated antibodies against inhibin, and that they had higher FSH levels compared with the controls. The number of follicular waves during the estrous cycle was higher in the immunized cows (3 or 4 waves) than in the controls (2 or 3 waves). Moreover, the immunized cows had a greater number of follicles during the estrous cycle compared with the control cows. The maximum number of follicles was 14.8 +/- 1.7 vs 5.4 +/- 0.2 in inhibin-immunized and control cows, respectively, during the first follicular wave and 13.9 +/- 1.9 vs 5.6 +/- 0.7, respectively, during the ovulatory wave. Multiple ovulations were increased in the immunized cows. However, the ovulation rate varied greatly in the immunized animals. In conclusion, immunization against inhibin increased FSH secretions during the estrous cycle in the cows. Moreover, the immunized cows had a greater number of follicular waves during the estrous cycle and a greater number of follicles, and this could be used as a potential source of oocytes for use in IVF/embryo transfer programs.

Effects of bilateral efferent duct ligation on sperm motility and secretion of FSH, LH, inhibin, and testosterone in adult male rats.

Effects of bilateral efferent duct ligation (EDL) on sperm motility and testicular endocrinology were investigated in adult male rats. Bilateral EDL was created surgically in adult male rats (EDL group) and shamoperated rats were used as control (control group). Five rats from each group were killed on d 3, 5, 7, 14, and 35 after the surgery. The sperm motility parameters were determined by a computer-assisted sperm analysis system using sperm collected from the cauda epididymis. Concentrations of spermatozoa in epididymis and testis were counted. The motility of sperm decreased remarkably in EDL rats compared with controls on 5 d after the operation. Four sperm motility parameters-straight velocity (VSL), deviation of the sperm head from the mean trajectory (ALH, mean), the maximum amplitude of lateral head displacement (ALH, max) and curvilinear velocity (VCL)-increased on 3 d after the operation, and followed by a subsequent decline 5 and 7 d later. Concentrations of sperm significantly decreased in both testes and epididymis from 3 and 5 d after the operation. Plasma concentrations of FSH and LH increased significantly in EDL rats from 5 and 7 d after the operation, whereas plasma concentrations of immunoreactive (ir)-inhibin, inhibin B, and testosterone decreased. Testicular content of irinhibin showed an initial increase on 3 d after the operation, followed by a subsequent decline to levels significantly below controls by d 7 postoperation. On the other hand, testicular contents of testosterone were significantly higher in the EDL group than the control group on d 7-35 after the operation, whereas circulating levels of testosterone remained low. In the EDL testes, marked degenerative changes in the Sertoli cells and spermatogonia were observed, whereas Leydig cells showed clear hyperplasia. These results demonstrated that bilateral EDL induced a rapid reduction of sperm motility parameters during a short time. Present results also suggest that EDL first induces impairment of Sertoli cells function and this leads to reduction of sperm activity and secretion of inhibins. On the other hand, circulating levels of testosterone reduced after EDL and this leads to hypersecretion of LH. A large amount of LH resulted in a stimulation of Leydig cells hyperplasia.

Immunolocalization of steroidogenic enzymes in the fetal, neonatal and adult testis of the Shiba goat.

The localizations of steroidogenic enzymes (P450scc, 3 beta HSD, P450c17 and P450arom) in testes of Shiba goats were investigated by immunohistochemistry. P450scc, 3 beta HSD, P450c17 and P450arom were detected in all Leydig cells of adults. P450scc and P450c17 were observed in most Leydig cells in the fetus (90 days) and neonate (15 days). 3 beta HSD and P450arom were found in some Leydig cells of the fetus with weak immunostaining but the numbers of immunopositive Leydig cells and intense immunostaining were increased in Leydig cells of the neonate. These results suggest that Shiba goat testes have the ability to synthesize progestin, androgen and estrogen in the fetus, neonate and adult, and synthesis of these steroid hormones showed an age-related rise.

Follicular and hormonal dynamics during the estrous cycle in goats.

Transrectal ultrasonography of ovaries was performed daily in 6 goats for 3 consecutive estrous cycles. Blood samples collected daily were measured for concentrations of FSH, inhibin A, and estradiol-17beta. Follicular and hormonal data were analyzed for associations between the follicular waves and hormonal concentrations. During the interovulatory intervals, follicular growth and regression occurred in a wave like pattern (2-5 waves), and the predominant patterns were three and four follicular waves. In addition, there was no significant difference among the diameters of dominant follicles during the growth phase of the follicular waves. The number of 3 mm follicles peaked on days 0, 7, and 11 in interovulatory intervals that had three follicular waves and on days -1, 5, 11, and 15 in those that had four follicular waves. Plasma concentrations of FSH increased around the day of follicular wave emergence and declined with the growth of follicles. Circulating FSH increased again concomitant with regression of dominant follicles in the anovulatory wave, whereas FSH levels remained low in the ovulatory wave. Inhibin A was negatively correlated with FSH, while it was positively correlated with estradiol-17beta, suggesting that inhibin A is a product of healthy growing follicles and that it contributes to the suppression of FSH secretion. In conclusion, the growth of ovarian follicles in goats exhibits a wave-like pattern, and follicular dominance is less apparent in goats. Moreover, inhibin A may be a key hormone for regulation of the follicular wave through suppression of FSH secretion in goats.

Immunolocalization of inhibin/activin subunits in the shiba goat fetal, neonatal, and adult testes.

The objective of this study was to investigate the cellular immunolocalization of inhibin alpha and inhibin/activin (betaA and betaB) subunits in the fetal, neonatal and adult testes of Shiba goats. The testes were obtained from a fetus at 90 days, a neonate at 15 days, and two adult Shiba goats (both of 3 years old). The sections of testes were immunostained by the avidin-biotin-peroxidase complex method (ABC) using polyclonal antisera raised against porcine inhibin alpha, inhibin/activin betaA, and inhibin/activin betaB. Inhibin alpha and inhibin/activin (betaA and betaB) subunits were expressed in Leydig cells, but not in the Sertoli cells of the fetus with a weak immunostaining. An increase in the number of positive cells and a more intense immunohistochemical signal for inhibin alpha and inhibin/activin (betaA and betaB) subunits were observed in the Leydig cells of neonatal testes. Moreover, inhibin alpha, betaA, and betaB subunits were expressed in the Sertoli cells and Leydig cells of adult testes, respectively. These results suggest that Shiba goats testes have the ability to synthesize inhibins in the fetus, neonate, and adult, and the cellular localization of inhibin/activin subunits showed age-related changes in fetal, neonatal, and adult testes of Shiba goats.

Developmental changes of plasma inhibin, gonadotropins, steroid hormones, and thyroid hormones in male and female Shao ducks.

Plasma samples from developing male and female Shao ducks were assayed for immunoreactive (ir-) inhibin, follicle-stimulating hormone (FSH), luteinizing hormone (LH), steroid hormones, and thyroid hormones. In the male, plasma ir-inhibin significantly increased between 75 and 155 days posthatch, and then decreased slightly at day 165. Plasma FSH of male ducks decreased from day 35 to day 55, followed by progressive elevation until day 95. Plasma FSH of male ducks fell significantly at days 135 and 165, while plasma ir-inhibin rose to high level. In female ducks, plasma ir-inhibin remained low until the start of lay, and thereafter significantly increased at day 135. Plasma FSH fluctuated before day 95 and significantly rose at day 115, and decreased thereafter. In males, plasma LH did not vary significantly before day 135, however, plasma testosterone significantly increased from day 95 onwards. No changes in plasma LH were observed during development of female ducks. Plasma estradiol-17beta gradually increased reaching a peak level at day 135. Plasma progesterone did not vary significantly before day 135 and thereafter significantly increased. Both sexes showed a similar pattern in changes of plasma thyroid hormones during sexual development. There was a significant increase in plasma thyroxine (T4) at day 95, thereafter decreased. Plasma triiodothyronine (T3) was at high level at the earlier stage of development and significantly decreased at day 55. Significant increase in plasma T3 in male and female ducks was observed at 135 and 115 days, respectively. In conclusion, these results demonstrated that the rise in inhibin is correlated with age of sexual maturity in the female while the rise in inhibin significantly precedes sexual maturity in the male. There was a progressive increase in plasma steroid hormones towards sexual maturity, and there was no sex difference in the time course of thyroid hormones.

Secretion of leptin throughout pregnancy and early postpartum period in Japanese monkeys: placenta as another potential source of leptin.

Leptin is one of the most important factors linking nutrition and reproduction. In the present study, plasma concentrations of leptin during pregnancy and early lactation in the Japanese monkey were determined. Plasma concentrations of gonadotropins, immunoreactive (ir-)inhibin, and steroid hormones were also measured. Plasma concentrations of leptin significantly increased during the second quarter of pregnancy and progressively elevated throughout pregnancy. During the fourth quarter of pregnancy, leptin levels reached up to 89 and 64 times of those during prepregancy and first quarter of pregnancy periods, respectively. After parturition, the circulating leptin level abruptly decreased. During the first 10 d of lactation, its average level decreased to the levels of the second quarter of pregnancy. Plasma irinhibin and estradiol-17beta were elevated throughout the pregnancy and decreased after parturition, and both of them were positively correlated with leptin levels during the whole pregnancy and early lactation. Plasma concentrations of progesterone significantly increased during the first quarter of pregnancy and kept at a higher level compared with prepregnancy and sharply decreased after parturition. Placental homogenates contain a large amount of leptin protein. These results suggest that placenta secretes a large amount of leptin and may be another source of leptin during pregnancy in Japanese monkeys. In addition, high correlations among leptin, irinhibin, and estradiol-17beta during these stages suggest that these hormones may have important regulating roles on leptin secretion during pregnancy in the Japanese monkey.

Induction of superovulation by immunoneutralization of endogenous inhibin in immature rats.

The effects of passive immunoneutralization of endogenous inhibin on ovulation rate in immature rats were investigated. Efficiency of superovulation on production of fertilized oocytes was compared between the inhibin antiserum (inhibin-AS) and equine chorionic gonadotropin (eCG) protocols. Immature female Wistar strain rats were superovulated with a single injection of 100-200 microl inhibin-AS, with and without an injection of human chorionic gonadotropin (hCG). A total of 77.8% of the 26-30-day-old rats treated with a single injection of 100-200 microl inhibin-AS ovulated 72 h after treatment, while rats given normal goat serum (NGS; 200 microl) did not ovulate. At 28 days of age, all of the inhibin-AS treated rats ovulated when additional hCG treatment was given, whereas the number of ovulated oocytes was not affected. The number of ovulated oocytes in the inhibin-AS-hCG treated groups was significantly higher than that of the NGS-hCG treated group. In addition, plasma concentrations of FSH in the inhibin-AS-hCG treated group significantly increased compared with the NGS treated group. While the percentage of mated rats in the 200 microl inhibin-AS-hCG treated group was significantly lower than that of the 15 IU eCG-hCG treated group, the fertilization rate was comparable between the two groups. The number of fertilized oocytes in the 200 microl inhibin-AS-hCG treated group was significantly higher in comparison with the 15 IU eCG-hCG treated group. These results suggest that immunoneutralization of endogenous inhibin could be a reliable method for induction of superovulation to collect a large number of normally fertilized oocytes in immature rats.

Plasma concentrations of immunoreactive (ir)-inhibin, gonadotropins and steroid hormones during the ovulatory cycle of the duck.

The present study was undertaken to determine changes in circulating levels of immunoreactive (ir)-inhibin, FSH, LH, estradiol-17beta, progesterone, and testosterone during the ovulatory cycle of Shao ducks. Serial blood samples were taken from two groups of laying ducks for measurement of ir-inhibin, gonadotropins, and steroid hormones at 2 h intervals for 24 h. Plasma concentrations of ir-inhibin did not change significantly during the ovulatory cycle. The highest level of plasma ir-inhibin was observed 6 h prior to ovulation, which coincided with a decreased level of plasma FSH. One FSH surge was found 12 h after ovulation. Estradiol-17beta, progesterone, and testosterone were also determined during the ovulatory cycle. Two peak values were detected for estradiol-17beta 8 h before ovulation and 4 h after ovulation, while progesterone started to increase 4 h before ovulation and reached a peak at ovulation. The highest level of plasma testosterone was detected around the time of ovulation. These results suggest that inhibin may be involved in the control of FSH secretion during the ovulatory cycle. In addition, both LH and progesterone are of importance in the ovulation process of Shao ducks.

Active immunization against inhibin improves superovulatory response to exogenous FSH in cattle.

The effect of active immunization against inhibin on the response to superovulatory treatment by porcine FSH (pFSH) was investigated in cattle. Japanese black cows were sc injected with 1 mg of porcine inhibin alpha-subunit fragment (1-26) conjugated with rabbit serum albumin (inhibin-immunized group; n=14) or rabbit serum albumin alone (control group; n=12) in Freund's complete adjuvant. Booster injections (half the amount of the primary injection) were given 35 and 70 days after the primary injection. All cows were superovulated three times with pFSH. Three days after each injection of the antigen, a progesterone-releasing intravaginal device (CIDR-B) was inserted vaginally into all animals and left in place for 10 days. Forty-eight hours before CIDR-B removal, all animals were sc injected with 30 mg pFSH dissolved in 40% polyvinylpyrrolidone, and im injected with 750 microg of PGF2alpha at CIDR-B removal. Cows were artificially inseminated twice during estrus, and ova or embryos were collected 7 or 8 days after estrus. The number of corpora lutea, the number of ova or embryos and the number of transferable embryos in inhibin-immunized cows (12.1+/-1.2, 11.1+/-1.3 and 6.2+/-1.0, respectively) were significantly greater than those in the controls (8.2+/-1.0, 5.7+/-1.1 and 3.1+/-0.7, respectively). These results indicate that active immunization against inhibin enhanced ovarian response to the usual superovulatory treatment in cattle. Therefore, immunization against inhibin may be a useful approach for improving the response to superovulation in cattle.

Immunolocalization of nerve growth factor (NGF) and its receptors (TrkA and p75LNGFR) in the reproductive organs of Shiba goats.

The objective of this study was to determine the immunolocalization of NGF and its receptors (TrkA and p75LNGFR) in the reproductive tract of the Japanese Shiba goats. Five adult goats were used in this study and sections of ovaries, uteri and oviducts were immunostained by the avidin-biotin-peroxidase complex method (ABC). The results showed that NGF and its receptors (TrkA and p75LNGFR) were expressed in granulosa cells, theca cells, interstitial cells and lutein cells in ovaries. Immunoreactions for NGF, TrkA and p75LNGFR were also detectable in epithelial cells and muscle cells of the ampulla and isthmus of the oviduct, and in epithelial cells and uterine glands of the uterus. These results strongly suggest autocrine and paracrine regulation of reproductive function by NGF in the reproductive tract of female Shiba goats.

Immunolocalization of steroidogenic enzymes in the corpus luteum and placenta of the Japanese Shiba goat.

In this study, we performed immunohistochemistry of cholesterol side-chain cleavage cytochrome P450 (P450scc), 3beta-hydroxysteroid dehydrogenase (3betaHSD), cytochrome 17alpha-hydroxylase P450 (P450c17), and cytochrome P450 aromatase (P450arom) in the corpus luteum and placenta of Shiba goats. The aim was to clarify the steroidogenic capability of the corpus luteum and placenta of Shiba goats. Ovaries containing corpora lutea were obtained from four adult Shiba goats during the luteal phase (day10; n=2) and pregnancy (90 and 120 days of gestation). Placenta was obtained from one Shiba goat on day 120 of gestation. The sections of the ovaries and placentae were immunostained using the avidin-biotin-peroxidase complex method (ABC) with polyclonal antibodies generated against steroidogenic enzymes of mammalian origin. All luteal cells expressed P450scc, 3betaHSD, P450c17 and P450arom. The distribution of P450scc, 3betaHSD, P450c17 and P450arom were not different during the luteal phase and pregnancy. P450arom showed a weak positive staining in late pregnancy (120 days). In addition, immunoreactions for P450c17 and P450arom were observed in syncytiotrophoblast of the placenta of one Shiba goat. These results indicate that, in Shiba goats, corpus luteum is not only an important source of progesterone but also has the ability to synthesize androgen and estrogen during the luteal phase and pregnancy. Also the placenta has the ability to synthesize androgen and estrogen in late pregnancy.

Immunolocalization of steroidogenic enzymes P450scc, 3betaHSD, P450c17, and P450arom in Göttingen miniature pig testes.

The objective of this study was to investigate immunolocalization of steroidogenic enzymes in Göttingen miniature (GM) pig testes. Testes of 6 adult GM pigs were obtained in September 1996 (n=2), February (n=2) and June (n=2), 1997. Steroidogenic enzymes were immunolocalized using polyclonal antisera raised against bovine adrenal cholesterol side-chain cleavage cytochrome P450 (P450scc), human placental 3beta-hydroxysteroid dehydrogenase (3betaHSD), porcine testicular 17alpha-hydroxylase cytochrome P450 (P450c17), and human placental aromatase cytochrome P450 (P450arom). Histologically, all types of spermatogenic cells including mature-phase spermatozoa in seminiferous tubules were observed in all testes throughout the year. Moreover, P450scc, 3betaHSD, P450c17and P450arom were identified in Leydig cells but not in Sertoli cells of all testes. These results suggested that adult GM pig testes have the ability to produce germ cells throughout the year, and the synthesis of progestin, androgen and estrogen occurs in the Leydig cells of GM pig testes.

Secretion of inhibin and testicular expression of inhibin subunits in male duck embryos and newly hatched ducks.

Concentrations of immunoreactive (ir-) inhibin in circulation, amniotic fluid, and testes of embryos and newly hatched ducks were determined from d 21 of incubation to d 1 of age. Plasma concentrations of FSH and LH were also determined by chicken radioimmunoassay (RIA) systems. In addition, gene expression and cellular source of inhibin were investigated by in situ hybridization and immunohistochemistry. The results showed that plasma ir-inhibin gradually declined from d 21 to d 24, followed by an increase on d 25 and remained high until d 1 after hatching. FSH in plasma was high on d 21 followed by a sharp decline toward d 25 after which FSH levels stabilized. A reverse relationship was observed between inhibin and FSH during the late stage of incubation. Embryonic testes contained high ir-inhibin levels. Testicular ir-inhibin levels were relatively high at early time points with a peak on d 23, and significantly decreased from d 23 to d 24 and stabilized thereafter. Amniotic fluid concentrations of ir-inhibin were relatively low and remained constant between d 21 and d 25. In situ hybridization demonstrated that the expression of inhibin alpha- and betaA-subunit mRNA was coexisted in the cells in the seminiferous tubules of testes on d 25. The immunoreactivity of inhibin betaA- and betaB-subunits was colocalized in the cells in the seminiferous tubules of testes on d 25. The results of dimeric inhibins determined by the ELISA method showed that inhibin B can be measured in embryonic testicular homogenate and pooled embryonic plasma. Although inhibin A was detected in testicular homogenate, it was under the detection limit in pooled embryonic plasma. In conclusion, these results indicate that cells in the seminiferous tubules of embryonic testes in ducks may secrete dimeric (bioactive) inhibins to circulation and that the FSH-inhibin feedback loop may become operational during the late stage of the incubation.