Effects of testosterone on testicular inhibin and fluid production in intact and hypophysectomized adult rats

Rats were given s.c. implants of high (HT) or low (LT) doses of testosterone and 10 days later hypophysectomy or sham-operation was performed. The rats were killed after 50 days. Unilateral efferent duct ligation was performed 16 h before death to measure seminiferous tubule fluid production and the increment in testicular inhibin values (inhibin production). Inhibin levels in testis cytosols were measured by a pituitary cell culture bioassay. The LT implants maintained serum testosterone at control values and decreased testicular weight whereas HT implants raised serum testosterone 3-fold and maintained testicular weight at 75-85% of pretreatment levels. In intact rats, LT implants caused no change in testicular inhibin content but decreased inhibin production; no significant changes occurred with HT implants. After hypophysectomy both values were significantly suppressed and could not be maintained by HT or LT implants. However, the HT implants partly restored inhibin production despite their inability to influence testicular inhibin content. In contrast, tubule fluid production depended mainly on intratesticular testosterone levels and occurred normally in intact or hypophysectomized rats with HT but not LT implants. These results indicate that inhibin and seminiferous tubule fluid production, both functions of the Sertoli cell, are under different hormonal control. The maintenance of inhibin production by the testis requires the support of pituitary hormones, presumably FSH, while seminiferous tubule fluid production requires testosterone, presumably through LH stimulation of Leydig cells. These findings are consistent with the hypothesis that inhibin is produced in response to trophic stimulation by FSH.     

An in-vivo method for estimating inhibin production by adult rat testes

The concentrations of inhibin in samples of rat testicular venous and arterial blood and interstitial fluid were measured by an in-vitro bioassay using pituitary cells in culture in which the standard was an ovine testicular lymph preparation (assigned potency 1 unit/mg). Inhibin levels were undetectable (less than 2 U/ml) in both blood samples but reached a mean concentration of 120 +/- 7 U/ml in testicular interstitial fluid. After unilateral efferent duct ligation the rate of inhibin accumulation in seminiferous tubules was determined by the difference in the inhibin content of the ligated and unligated testes. Additionally, the rate of seminiferous tubule fluid production was obtained from the difference in weight between the ligated and non-ligated testes. In the 24 h after efferent duct ligation there were linear increases in inhibin (18.5 +/- 1.0 U/h) and in seminiferous tubule fluid production (26 +/- 1 microliter/h), but there were no changes in serum FSH and LH levels. Experimental induction of bilateral cryptorchidism led to a decrease in the inhibin content of the testis after 10 days. The rate of inhibin accumulation after efferent duct ligation declined more rapidly than the inhibin content, being significantly depressed in cryptorchid testes after 3 days, suggesting that this measurement is a more sensitive index of inhibin production than the determination of testicular inhibin content.     

Effect on gonadotrophin secretion of blockage of the ductuli efferentes in the normal and androgen-deprived rat

Serum LH and FSH concentrations were measured in adult male rats. Blockage of the ductuli efferentes by surgical ligation or treatment with the anti-fertility drug alpha-chlorohydrin did not produce any changes in serum LH or androgen-dependent organ weights. Serum FSH in both groups was significantly raised by 4 days after treatment and remained so throughout most of the experimental period (42 days) but did not attain the values obtained in castrates. The gonadotrophin changes were accompanied by an initial increase in the weight and turgidity of the testes which then became flaccid and atrophied. These changes were similar but not identical after both treatments. Androgen secretion was impaired in adult rats treated with ethylene dimethane sulphonate; serum LH values were similar to those of 14-day castrates, and serum FSH was also elevated but not to the level found in the castrates. Ligation of the ductuli efferentes did not produce any further changes in concentrations of either hormone in spite of the retention of the testicular products within the testes as reflected by an increase in weight. The results are consistent with the view that changes in serum FSH after blockage of the ductuli efferentes are related to disturbances in the production of inhibin from the seminiferous epithelium rather than retention of inhibin with the testicular tubules.     

Production and endocrine role of inhibin during the early development of bull calves.

This study investigated the ontogeny of control of FSH secretion by inhibin during early prepubertal development of bulls by 1) measurements of circulating levels of inhibin and FSH from 1 to 13 wk of age, and 2) immunoneutralization of endogenous inhibin at 7, 21, 60, and 120 days of age. In addition, production and localization of inhibin in testes were examined by immunohistochemistry and Western blots at 7, 21, 60, and 120 days of age. Plasma immunoreactive inhibin levels were relatively low between 1 and 3 wk of age and then showed a tendency to rise (P < 0.1) from 4 wk of age. Circulating concentrations of FSH were low during 3 wk after birth and increased at 5 wk, remained high (P < 0.05) until 16 wk of age. Treatment with inhibin antiserum resulted in a significant (P < 0.05) increase in plasma FSH at 7, 21, 60, and 120 days of age compared to those following injection of control serum; however, the magnitude of the FSH rise after inhibin immunization was greater as bulls aged. There were no significant changes in plasma LH after inhibin immunization. An intense staining of inhibin alpha subunits was found in Sertoli cells within the solid seminiferous cords from 7 to 120 days of age, while no specific immune reaction was found in interstitial cells. Western blot analysis of testicular homogenates isolated from bulls 7-120 days of age revealed presence of a 28.5-kDa molecule that cross-reacted with inhibin alpha subunit and beta(B) subunit-specific antibodies. In this study, before 13 wk of age in bull calves, there was no inverse relationship between plasma concentrations of immunoreactive inhibin and FSH. However, the present immunization study clearly indicates that inhibin participates in the regulation of FSH secretion from infancy to early prepubertal stage, although the endocrine significance of inhibin becomes greater in older bulls. The results also indicate that the major production site of inhibin in the testis is Sertoli cells and that these cells produce inhibin that exerts a negative feedback effect on FSH secretion from early stages of development.     

Evidence for a major role of inhibin in the feedback control of FSH in the male rat

Adult rats (16-18/group) received a single intratesticular injection of 25, 100 or 400 microliters glycerol solution (7:3 in distilled water, v/v). Half of the rats in each group were given implants of testosterone, a testosterone-filled Silastic capsule (1.5 cm length) to provide serum values of testosterone within the normal range. After 1 week all animals were killed by decapitation. Serum concentrations of gonadotrophins, testosterone and immunoactive inhibin as well as testicular concentrations of testosterone and bioactive inhibin were determined. Testicular histology was studied in Paraplast-embedded tissue stained with PAS and haematoxylin-eosin. Glycerol treatment caused a dose-dependent ablation of spermatogenesis in a distinct area around the site of injection. Serum concentrations of FSH increased proportionally with increasing spermatogenic damage while serum LH and testosterone remained unaltered except with the highest glycerol dose. The rise in serum FSH was significantly correlated with serum (r = -0.70, P less than 0.001) and testicular (r = -0.66, P less than 0.001) concentrations of inhibin. A less pronounced correlation was found between LH and serum inhibin (r = 0.48). No correlation was found between the concentrations of LH and testicular inhibin or between serum concentrations of FSH and serum testosterone in the 25 and 100 microliters groups. Maintenance of low to normal serum testosterone concentrations by means of Silastic implants blocked the elevation of FSH in glycerol-treated animals but failed to affect significantly serum FSH in untreated rats. In all testosterone treated rats testicular inhibin concentrations were markedly reduced in the presence of lowered concentrations (7-14%) of testicular testosterone and unaltered serum FSH concentrations.     

Measurement of dimeric inhibins and effects of active immunization against inhibin alpha-subunit on plasma hormones and testis morphology in the developing cockerel

Inhibins and activins are implicated as endocrine regulators of follicle-stimulating hormone production and of testicular steroidogenesis and spermatogenesis in mammals. The potential involvement of these proteins in cockerels was investigated by measurement of circulating inhibin A, inhibin B, total inhibin alpha-subunit immunoreactivity (ir-alpha), activin A, LH, FSH, and testosterone from the juvenile state through to sexual maturity. Plasma inhibin A remained low between 6 to 12 wk of age and increased approximately threefold (P < 0.05) to a prepubertal peak between Weeks 14 to 18, followed by a gradual decline to the end of the study (Week 24). Although plasma FSH levels were not correlated to inhibin A before Week 16 (r = -0.17), they were negatively correlated from Week 18 (r = -0.49; P < 0.005). Inhibin B levels were below the assay detection limit until 16 wk of age but thereafter rose steadily in parallel with FSH (r = 0.27; P < 0.02) and testosterone (r = 0.35; P < 0.005). Thus, inhibins A and B showed divergent profiles during sexual maturation. Plasma ir-alpha levels were much higher than dimeric inhibin levels throughout, although the relative difference varied with age. Plasma activin A levels were below the assay detection at all times. Juvenile cockerels were actively immunized against a synthetic chicken inhibin alpha-subunit peptide conjugate to determine effects on plasma hormones and on testicular weight, morphology, and activin A content. Immunization generated circulating antibodies that bound (125)I-bovine 32-kDa inhibin but did not affect plasma FSH or testosterone levels at any stage of development. However, immunization reduced postpubertal plasma LH levels (P < 0.05) and promoted increased testicular weight (24%; P < 0.01) and total testicular activin A content (42%; P < 0.001) at 24 wk. Testis weight of immunized birds was positively correlated with inhibin antibody titer (r = 0.61; P < 0.05). Live weight gain was not affected by immunization. Morphometric analysis of testis sections showed that inhibin immunization had no effect on the fractional volume of the seminiferous tubule wall, seminiferous tubule lumen, or interstitial tissue area. Likewise, seminiferous tubule surface area and surface area:volume ratios were not different from controls. These findings support differential roles for inhibins A and B in regulating the pituitary-testicular axis during sexual maturation in the cockerel but highlight the need for more detailed studies to distinguish between potential endocrine and local intragonadal roles of inhibin-related peptides and to elucidate the mechanism by which immunization against inhibin alpha-subunit promotes testis enlargement without raising plasma FSH.     

Weaning and the developmental changes in follicle-stimulating hormone, pituitary adenylate cyclase-activating polypeptide, and inhibin B in the male rat

Pituitary Fshb concentrations increase markedly and selectively beginning on Postnatal Day 20 in the male rat. To evaluate the factors potentially responsible for this rise in FSH, we adjusted the time of weaning, which is generally also on Day 20. Male rat pups were provided nutrients by suckling only and were weaned to laboratory chow earlier (Day 17) or later (Day 23) than normally performed in animal facilities (Day 20). Between ages 17 and 29 days, significant increases were seen in serum LH (1.4-fold) and FSH (2.4-fold) levels; pituitary expression of Lhb (5.4-fold), Fshb (21.3-fold), and inhibin beta B (Inhbb, 2.26-fold) mRNAs; and testicular expression of Inhbb (10-fold) mRNA. Concurrently, significant decreases occurred in serum inhibin B levels (1.8-fold); pituitary adenylate cyclase-activating polypeptide (Adcyap1, 4.2-fold), total follistatin (Fst, 3.5-fold), and Fst isoform 288 (5.6-fold) mRNAs; and testicular expression of inhibin beta A (8.2-fold) mRNA. Early weaning significantly increased serum FSH but not LH and increased pituitary expression of Fshb and GnRH receptor (Gnrhr) mRNAs but not Lhb. Early weaning also significantly decreased serum inhibin B but increased testicular expression of the Inhbb subunit. Early weaning also caused pituitary expression of Fst and Adcyap1 to decline earlier than in the control group. Immediately after weaning, growth accelerated substantially, and the time of weaning produced significant and differential effects on circulating leptin levels that were not related to indices of FSH production. From these observations, we propose the novel hypothesis that the increase in growth rate subsequent to weaning signals circulating inhibin B levels to fall and pituitary Adcyap1 and consequently Fst expression to decrease, and that these events together facilitate the rise in Fshb and Gnrhr expression by increasing pituitary activin signaling.     

Immunoneutralization of inhibin modifies hormone secretion and sperm production in bulls.

The objective of this study was to examine the role of endogenous inhibin in the regulation of FSH, LH, and testosterone secretion and sperm production in bulls. Bulls were actively immunized against bovine inhibin alpha 1-26 gly-tyr (bINH) conjugated to human alpha globulin (HAG) or HAG alone (controls) and emulsified in Freund's complete adjuvant. Primary immunization was at 14 wk of age, followed by booster immunizations in Freund's incomplete adjuvant at 28, 30, and 34 wk of age. Ten days after each booster immunization, scrotal circumferences and body weights were measured, and blood was sampled for determination of bINH antibody titer. Ten days after the third booster, blood was sampled at 1-h intervals for 8 h to quantify serum concentrations of FSH, LH, and testosterone. After this blood sampling period, bulls were castrated and testicular sperm production was determined. Serum diluted 1:4,000 from bINH-immunized bulls bound 36%, 52%, and 53% of radioiodinated bINH after the first, second, and third boosters, respectively. Serum from controls bound less than 1% radioiodinated bINH. After the third booster, serum concentrations of FSH and testosterone were increased (p less than 0.05) and LH concentrations were decreased (p less than 0.001) in bINH-immunized bulls compared with controls. After the third booster, daily sperm production per gram of testicular parenchyma was increased (p less than 0.05) in bINH-immunized bulls compared with controls. Scrotal circumferences and body weights were similar between treatment groups throughout the experiment. We concluded that inhibin has a role in regulation of secretion of gonadotrophins and testosterone and testicular sperm production, but not testicular growth, in bulls.(ABSTRACT TRUNCATED AT 250 WORDS)     

Changes in plasma inhibin levels following pulsatile gonadotrophin-releasing hormone therapy in a man with idiopathic hypogonadotrophic hypogonadism

Changes in circulating inhibin levels were related to changes in testosterone (T) and the gonadotrophins luteinizing hormone (LH) and follicle-stimulating hormone (FSH) in a hypogonadotrophic hypogonadal man before and during pulsatile gonadotrophin-releasing hormone therapy which resulted in normal spermatogenesis. Before treatment, the plasma inhibin levels in the patient (210 +/- 50 U/l; mean +/- SD of four samples) were lower than in normal controls (552 +/- 150 U/l; p less than 0.01), as were T (1.1 nmol/l) and gonadotrophin (less than 1.0 IU/l) levels. Within 1 week of gonadotrophin-releasing hormone treatment, plasma LH (14.1 +/- 0.7 IU/l) and FSH (14.4 +/- 0.6 IU/l) reached supraphysiological levels. In response, T and inhibin concentrations increased progressively to reach high normal levels (27.7 +/- 1.6 nmol/l and 609 +/- 140 U/l) at 4 weeks, by which time the gonadotrophin levels stared to decline and gradually returned to the normal range between 12 and 24 weeks of treatment. There was a concomitant decrease in T and inhibin levels which remained within the normal range. The decline in the FSH level following the rise in testicular hormones was earlier and steeper than that of LH (37.5% decrease at 4 weeks vs. 30.4% at 12 weeks), suggesting that T and inhibin may act together to inhibit pituitary FSH secretion as opposed to LH secretion which is primarily controlled by T. It is concluded that, in man, during maturation of the pituitary-testicular axis, changes in circulating inhibin parallel those of T, and quantitatively normal inhibin secretion is dependent on gonadotrophin stimulation. FSH secretion may be regulated through negative feedback control, by both T and inhibin.     

Effects of weaning on concentrations of inhibin in follicular fluid and plasma of sows.

Changes in plasma and follicular fluid concentrations of inhibin were examined in sows after weaning at 28-32 days post partum. From 0 to 48 h after weaning, inhibin concentrations were 200-300 times higher in follicular fluid from small (less than 4 mm) and medium-large (greater than or equal to 4 mm) follicles than in ovarian venous plasma. Inhibin concentrations increased in follicular fluid from medium-large follicles at 24 and 48 h after weaning; concentrations in ovarian venous plasma were positively correlated with the number of medium-large follicles (r = 0.40) and with ovarian venous plasma concentrations of oestradiol (r = 0.61). Blood samples were collected for 30 days from sows (n = 6) that exhibited oestrus within 5 days after weaning and from sows (n = 5) that remained anoestrous for 11 days after weaning. Plasma inhibin concentrations rose in oestrous and anoestrous sows by 12 h and continued to rise for 60 h after weaning. Plasma inhibin concentrations rose further and were higher at 3.5-4.5 days after weaning in oestrous sows than in sows that remained anoestrous. After oestrus, plasma inhibin concentrations declined. At weaning, plasma concentrations of follicle-stimulating hormone (FSH) were higher in sows that subsequently exhibited oestrus than in sows that remained anoestrous. After weaning, plasma concentrations of FSH declined in both groups, reached a nadir at 2.5 days, and increased gradually in anoestrous sows; oestrous sows exhibited an FSH surge at oestrus. Plasma FSH returned to preweaning concentrations in both groups of sows at Days 7-8.(ABSTRACT TRUNCATED AT 250 WORDS)     

Circulating concentrations of inhibin-related proteins during the ovulatory cycle of the domestic fowl (Gallus domesticus) and after induced cessation of egg laying

Circulating inhibin A, inhibin B, activin A, total immunoreactive inhibin alpha-subunit (ir-alpha inhibin), LH, FSH and progesterone concentrations were measured throughout the normal ovulatory cycle and after cessation of egg laying induced by feed restriction to investigate the potential involvement of inhibins and activins in the ovulatory cycle of the domestic hen. Plasma inhibin A varied significantly (P < 0.05) during the ovulatory cycle; the concentration was highest at the preovulatory LH surge and reached a nadir 10 h later, at about the time the F(2) follicle makes the transition to become the new F(1) follicle. Plasma FSH concentrations did not change significantly throughout the cycle and showed no correlation with inhibin A. Total ir-alpha inhibin concentrations were much higher than those of inhibin A at all stages of the ovulatory cycle and showed no correlation with inhibin A or FSH. Plasma concentrations of inhibin B and of activin A were below the detection limit of the assays in all plasma samples analysed. In the feed restriction study, plasma inhibin A and total ir-alpha inhibin showed little change until the last day of oviposition (day 0) after which they fell significantly (P < 0.05) and remained low to the end of the experiment (approximately 70-78% decrease relative to day -4). Conversely, plasma FSH increased after cessation of laying and was significantly higher (P < 0.05) from day 3 to the end of the study (approximately 50% increase on day 6 relative to day -4). Plasma FSH values were negatively correlated with inhibin A (r = -0.39; P < 0.005) and total ir-alpha inhibin (r = -0.36; P < 0.005). Plasma LH and progesterone also decreased (P < 0.05) during feed restriction. The decrease in LH preceded the terminal oviposition and the associated fall in inhibin A by 2 days; there was a positive correlation between LH and inhibin A (r = 0.35; P < 0.005). Taken together these findings support (i) a role for LH in promoting inhibin A secretion by preovulatory follicles and (ii) an endocrine role for inhibin A secreted by preovulatory follicles in the maintenance of tonic FSH secretion in laying hens.     

Inhibin-like effects of interstitial testis fluid: effects on serum FSH and androgen-binding protein in epididymis

Application of rat interstitial testis fluid (ITF) to orchidectomized rats diminishes significantly the rise of FSH in serum following castration. The effect is seen for 12-24 hours on day 3 and 50 after orchidectomy but not at 48 hours in rats treated 50 days after orchidectomy. LH levels are not significantly affected. These observations are assumed to be related to an inhibin-like activity in ITF. Therefore ITF (after extraction with ether) is given to intact animals and the testicular parameter of FSH, the androgen-binding protein (ABP), is measured by "steady-state" PAGE. Unexpectedly, ABP concentrations increase. At present, this effect cannot be explained. It is concluded that ITF contains only small amounts of inhibin and will be no good source for further studies.     

Effect of testosterone on serum immunoactive inhibin concentrations in intact and hypophysectomized male rats

Intact and hypophysectomized rats were treated with graded doses of testosterone via subcutaneous Silastic implants over a 13-week period. Serum inhibin concentrations fell 50% (P less than 0.001) after 2 weeks of hypophysectomy, remaining suppressed at this level for 13 weeks. The administration of testosterone to hypophysectomized rats (serum testosterone values 2-12 ng/ml; control values 5.5 ng/ml) was without effect on serum inhibin values. In contrast, administration of testosterone to intact animals for 7 weeks resulted in an initial fall (P less than 0.05) in inhibin levels to 50-70% of controls then increasing to reach control levels at higher doses. Serum FSH concentrations were similarly biphasic with increasing dose of testosterone and values for these two hormones were significantly correlated (r = 0.44, P less than 0.01). Segments of seminiferous tubules in culture from rats after various times of hypophysectomy showed a partly suppressed secretion of inhibin. The administration of testosterone did not modify inhibin production although inhibin production was sensitive to FSH. It is concluded that (1) serum inhibin concentrations are partly suppressed after hypophysectomy and testosterone has no effect on serum inhibin values; and (2) the suppression of serum inhibin in intact rats treated with increasing doses of testosterone is attributable to the concomitant fall in serum FSH concentrations.     

Immunoactive inhibin as a marker of Sertoli cell function following cytotoxic damage to the human testis

Sertoli and Leydig cell functions were evaluated in men with testicular damage due either to cytotoxic chemotherapy (CCT) or radiotherapy (XRT). Serum immunoactive inhibin, follicle-stimulating hormone (FSH), luteinizing hormone (LH) and testosterone concentrations were measured in 15 men (19-50 years) who had received 6-10 courses of combination CCT (mustine, vinblastine, procarbazine and prednisolone) for Hodgkin's disease 1-8 years earlier and 18 men (21-49 years) who had undergone unilateral orchidectomy for testicular seminoma followed by XRT (30 Gy) to the remaining testis, 1-4 years earlier. Normal men (n = 16, 19-36 years) acted as controls. Median inhibin (422 U/l) and testosterone (16.0 nmol/l) levels in the CCT-treated group were not significantly different from controls, whereas median FSH (14.5 IU/l) and LH (10.0 IU/l) levels were higher (p less than 0.0001 and p less than 0.001) than normal (2.9 and 5.5 IU/l). The median inhibin/FSH (I/FSH) ratio in the patients was lower (p less than 0.0001) than in the controls (33.8 vs. 187.0) as was the testosterone/LH (T/LH) ratio (1.7 vs. 3.8, p less than 0.001). In the XRT-treated group, both median inhibin (194.5 U/l) and testosterone (12.7 nmol/l) levels were lower (p less than 0.0001 and p less than 0.01) than normal (532.8 U/l and 20.0 nmol/l) in the presence of greatly elevated FSH (26.0 IU/l) and LH (14.5 IU/l) levels. In conclusion, CCT-induced testicular damage is associated with subtle Sertoli and Leydig cell dysfunction demonstrated by the reduced I/FSH and T/LH ratios; however, compensatory mechanisms maintain normal testosterone and inhibin levels.(ABSTRACT TRUNCATED AT 250 WORDS)     

Short term treatments with prolactin, HMG or testosterone fail to affect testicular inhibin content in rats

To investigate the effect of prolactin (PRL) on testicular function, especially on spermatogenesis, testicular inhibin content in male rats treated with PRL was compared with those treated with HMG and testosterone. Mature Wistar male rats were given 10 or 50 IU of ovine PRL, 10 IU of HMG and 5 mg of testosterone, i. m. for 5 consecutive days and testes were removed for assessing inhibin content. Inhibin content was measured by a FSH suppressing activity in cultured rat anterior pituitary cells using aquous extract of testes. Five days' treatment with PRL, HMG, or testosterone did not influence testicular inhibin content in male rats. The possibility that these treatments had transiently affected testicular inhibin content, or that inhibin content did not reflect inhibin production was not ruled out.     

Sexual maturational changes circulatory inhibin concentration in relation to FSH concentration and testicular size in Suffolk and DLS rams

Developmental patterns in immunoactive inhibin and FSH concentrations in peripheral blood were determined for Suffolk and DLS (Dorset x Leicester x Suffolk) rams born in January Blood samples were taken every 3 to 4 wk when testes were developing during puberty (5 to 44 wk of age) and redeveloping in early adulthood (17 to 23 months of age). Suffolk lambs had a greater average daily gain (195 vs. 143 g/day, P<0.01), and they developed larger testes (P<0.01) than DLS lambs. Inhibin and FSH concentrations peaked at about the same pubertal (8 wk) and early adult (19 or 20 months) ages in both breeds. Elevations in FSH were greater (P< 0.05) in Suffolk than DLS rams at each stage of development. The pubertal inhibin peak was nearly 70% larger (P<0.01) in DLS than Suffolk rams, and the early adult peak was comparable in rams of both breeds, but much smaller (P<0.01) than the pubertal peak. Nonetheless, inhibin was positively correlated (r=0.48 to 0.57) with FSH in both breeds during each developmental stage. Inhibin and testicular size were negatively correlated in Suffolk (r=-0.74) and DLS (r=-0.86) rams during puberty, and positively correlated in DLS rams (r=0.46) in early adulthood. We conclude that 1) inhibin concentrations are higher in juvenile rams at the time Sertoli cell numbers are being established than in adult rams during testicular recrudescence and 2) rises in FSH concentration participate in regulating corresponding rises in inhibin concentration in both stages of testicular development.     

Changes in plasma inhibin A levels during sexual maturation in the female chicken and the effects of active immunization against inhibin alpha-subunit on reproductive hormone profiles and ovarian function

Inhibins and activins are firmly implicated in the control of pituitary FSH secretion and ovarian follicular development in mammals. As in mammals, inhibin A and activin A are expressed in the preovulatory follicles of birds, and a defined ovulation cycle for inhibin A has recently been demonstrated in the laying hen. To investigate further the role of inhibin-related proteins in developing pullets, circulating concentrations of inhibin A, inhibin B, total immunoreactive inhibin alpha-subunit (ir-alpha), activin A, LH, FSH, and progesterone were measured from the juvenile state through to sexual maturity in 22 birds. In the 11 birds assigned to control groups, plasma inhibin A levels were low from 7 to 13 wk of age rising about threefold to a peak at Week 19 after which levels fell slightly to a plateau level characteristic of adult hens. Plasma inhibin A levels were negatively correlated with FSH (r = -0. 33; P: < 0.001) and positively correlated with progesterone (r = 0. 67; P: < 0.001) and ir-alpha (r = 0.53; P: < 0.001). Plasma ir-alpha levels were much higher than inhibin A levels although the relative differences varied with age. Plasma levels of inhibin B and activin A were below assay detection limits at all times. The remaining group of 11 birds was actively immunized (IMM) against a synthetic chicken inhibin alpha-subunit peptide (amino acids 1-26). The IMM generated circulating antibodies that bound native bovine inhibin A but altered neither plasma FSH nor progesterone levels relative to control birds at any stage of development nor the timing of first oviposition in week 19. Apart from a transient decline 1 wk after primary IMM, plasma LH concentrations did not differ from controls. Comparison of the numbers and size-class distribution of ovarian follicles at 29 wk showed an approximate twofold increase in the number of 8- to 9.9-mm-diameter follicles (control; 1.82 +/- 0.44 vs. IMM; 3.91 +/- 0.89; P: < 0.05), a size class that corresponds to follicles that have just joined the preovulatory hierarchy. The numbers of growing follicles in other size-classes and the sizes of hierarchical F(1)-F(7) follicles were not altered by IMM. However, the number of postovulatory follicles increased (control 3.73 +/- 0. 20 vs. IMM 5.55 +/- 0.28; P: < 0.01), and significantly more (P: < 0. 02) immunized hens laid two eggs within a 24-h period on at least one occasion (control 1 of 11 vs. IMM 9 of 11). The IMM increased (P: < 0.05) activin A content of F(1) and F(2) theca layers and decreased (P: < 0.05) activin A content in F(3) and F(4) granulosa layers, raising the possibility of a local intraovarian role of activin in mediating the response to IMM. These findings support a role for inhibin A in regulating the entry of follicles into the preovulatory hierarchy in the chicken, although further studies are required to establish the mechanism by which inhibin IMM increases the rate of follicle selection and ovulation without raising plasma FSH.     

Initiation of follicular maturation during mid-pregnancy by the administration of LH in the rat

Pregnant rats were injected twice daily for 1-3 days (Days 13-16 of pregnancy) with various doses of ovine LH. Follicular maturation was determined by the ability of the follicles to ovulate in response to 10 i.u. hCG as well as by endogenous production of oestradiol-17 beta and inhibin. In control animals, no ovulation was induced by hCG given on Day 16 of pregnancy. An injection of hCG on Day 16 of pregnancy, however, induced ovulation in LH-treated animals (6.25-50.0 micrograms LH per injection, s.c. at 12-h intervals from Days 13 to 16). Concentrations of oestradiol-17 beta and inhibin activity in ovarian venous plasma increased after the administration of LH, indicating that development of ovulatory follicles had been induced. Abolishing the decline in plasma LH values therefore induced maturation of a new set of follicles or prevented the atresia of large antral follicles usually seen at this time of pregnancy. Plasma and pituitary concentrations of FSH decreased in LH-treated animals compared with those in control animals. Concentrations of progesterone, testosterone and oestradiol-17 beta in the peripheral plasma were not significantly different between the two groups. These results suggest that the increase in inhibin secretion from the ovary containing maturing follicles after LH treatment may suppress the secretion of FSH from the pituitary gland. These findings indicate that (1) the development of ovulatory follicles can be induced by the administration of exogenous LH during mid-pregnancy in the rat and (2) basal concentrations of FSH are enough to initiate follicular maturation even in the presence of active corpora lutea of pregnancy, when appropriate amounts of plasma LH are present.     

Inhibin secretion by the sheep ovary

An in-vitro bioassay for inhibin based on FSH content or release by rat pituitary cells was validated for measuring inhibin activity in ovine plasma and lymph. Dose-dependent increases in inhibin activity were detected in peripheral plasma of 4 ovariectomized ewes 1 min after i.v. injections of ovine follicular fluid, and the half-life of inhibin in plasma for 2 ewes was 45 and 50 min, respectively. Inhibin was detected in ovarian lymph but not in ovarian or jugular venous plasma, even after treatment of ewes with PMSG to induce folliculogenesis. Destruction of visible follicles (greater than 0.5 mm diameter) on the ovaries of 4 PMSG-treated ewes by electrocautery was followed by a rapid and sustained decline in secretion of inhibin in ovarian lymph for up to 4 h. Ovarian lymph flow rates were either unchanged or slightly increased after cautery. Oestrogen concentrations in peripheral venous plasma declined within 15-30 min of cautery, but concentrations remained well above baseline. There was a significant decrease in peripheral progesterone concentrations in these same samples, but not until 2-3 h after cautery. FSH in peripheral plasma was depressed or non-detectable in PMSG-treated ewes and neither FSH nor LH concentrations in peripheral plasma were significantly altered up to 4 h after cautery of ovarian follicles. It is concluded that (a) antral follicles (greater than 0.5 mm) are the source of inhibin present in ovarian lymph, and (b) the ovarian lymphatic system is a route by which inhibin could reach the peripheral circulation, particularly in the luteal phase when ovarian lymph flow rates are high.