Effect of active immunization of pony mares against recombinant porcine inhibin alpha subunit on ovarian follicular development and plasma steroids and gonadotropins.

Two pony mares were immunized against recombinant porcine inhibin alpha subunit three times with 39 day intervals. Clinical findings and endocrinological changes before immunization were taken as the control. The first significant rise in the anti-inhibin titre (P<0.05) in the circulation was found 27 days after the first injection. Maximum binding activity was reached by the 12th day after the second booster dose. The number of small, medium and large sized follicles had increased significantly compared to before immunization (11.75 +/- 4.30, 2.75 +/- 0.69 and 2.51 +/- 0.63 vs 6.50 +/- 1.43, 1.83 +/- 0.44 and 1.33 +/- 0.38, respectively), but the ovulation rate remained unchanged after immunization. The average plasma concentration of FSH and estradiol-17beta during the estrous cycle increased significantly (P<0.05) after immunization. These results suggest that immunization against inhibin is a useful tool to increase the number of ovarian follicles during the estrous cycle of pony mares. Moreover, the present study supported the concept that inhibin plays a major role in the control of follicular growth through its inhibitory effect on FSH secretion synergistically with steroid hormones.

Effects of castration and recombinant human inhibin administration on circulating levels of inhibin and gonadotropins in adult male monkeys.

Inhibin has been suggested to play a role in gonadal feedback regulation of follicle-stimulating hormone (FSH) secretion; however, neither the half-life nor the time course of action of recombinant inhibin has been reported in any primate species. We sought to determine the disappearance half-life of circulating endogenous inhibin following castration in adult male monkeys, Macaca fascicularis, and to determine the half-life of administered recombinant human inhibin A and its effect on bioactive FSH and luteinizing hormone (LH) levels in castrate monkeys. Endogenous inhibin fell from 8,122 +/- 2,077 U/L (mean +/- SEM, n = 5) prior to castration to 383 +/- 84 U/L at 24 hours and 269 +/- 44 U/L at day 21 (P < 0.05 at 24 hours vs. day 21) (detection limit of assay 234 U/L). The early phase half-life of endogenous inhibin was 34 minutes (between 8 and 60 minutes) and a later phase half-life of 75 minutes was observed between 1 and 4 hours following castration. Recombinant inhibin exhibited a 14-minute early phase half-life between 8 and 60 minutes following the 5 micrograms intravenous (i.v.) recombinant inhibin dose, and a later phase half-life of 70 minutes between 1 and 4 hours in castrate monkeys (n = 3). Serum inhibin levels were maintained within or above the precastration range for 15 minutes. Single dose recombinant inhibin, 100 micrograms subcutaneous (SC) or intramuscular (IM) administered to castrate monkeys (n = 3), achieved and maintained normal serum inhibin levels for 6 hours.(ABSTRACT TRUNCATED AT 250 WORDS)

Pharmacokinetic profile of recombinant human (rh) inhibin A and activin A in the immature rat. I. Serum profile of rh-inhibin A and rh-activin A in the immature female rat.

The serum pharmacokinetics of recombinant human inhibin A (rh-inhibin A) and rh-activin A were examined in immature female Sprague Dawley-derived rats after iv and sc injection of the drugs. After iv administration of rh-inhibin A (120 micrograms/kg), the serum concentrations were described by a biexponential equation. The weight-normalized clearance was 21.3 ml/min.kg, and the initial (t1/2 alpha) and terminal (t1/2 beta) half-lives were 2.9 min and 37.9 min, respectively. Subcutaneous administration of 120 micrograms/kg rh-inhibin A resulted in a peak serum concentration of 10.6 ng/ml at 30.8 min after injection. Approximately 24% of the sc administered material was absorbed. Serum concentrations of rh-activin A also declined biexponentially after iv injection of the drug (120 micrograms/kg). The clearance of rh-activin A was 5.1 ml/min.kg, the t1/2 alpha was 6.1 min, and the t1/2 beta was 46.3 min. The peak serum concentration of rh-activin A (104.7 ng/ml) was achieved 24.7 min after sc delivery of the drug. The bioavailability of the sc dose was 38%. Iodinated rh-inhibin A and rh-activin A were used to examine the serum forms and metabolites of the drugs. [125I]rh-inhibin A and [125I]rh-activin A associated with two serum-binding proteins. Within 2 min of iv injection, the labeled hormones bound follistatin and alpha-2-macroglobulin. Even though rh-inhibin A and rh-activin A are structurally similar and appear to bind to the same serum proteins, their disposition in the immature rat differ.

Pharmacokinetic profile of recombinant human (rh) inhibin A and activin A in the immature rat. II. Tissue distribution of [125I]rh-inhibin A and [125I]rh-activin A in immature female and male rats.

The tissue distribution of recombinant human inhibin A (rh-inhibin A) and rh-activin A was determined in immature female Sprague Dawley-derived rats after iv administration of radiolabeled proteins. [125I]rh-Inhibin A and [125I]rh-activin A diverge in their distribution to tissues of the immature female rat as examined histologically (whole body autoradiography and thin section analysis) and by computing the percent dose and tissue to blood ratios for individual tissues. [125I]rh-inhibin A accumulated in the spleen, adrenal, bone marrow, and ovary after iv injection. Iodinated rh-inhibin A was also found in the anterior and posterior pituitary. [125I]rh-activin A was found in the ovary and pituitary after iv injection. Little specific binding was found in the spleen or adrenal. The bone marrow accumulated some [125I]rh-activin A which was competed by rh-activin A. The primary route of excretion for radioactivity was the kidney, with the label appearing in the bladder by 10 min after iv injection. Not only do rh-inhibin A and rh-activin A have different pharmacokinetics, but fewer tissues accumulate radioactive rh-activin A than rh-inhibin A.

Highly polarized secretion of inhibin by Sertoli cells in vitro.

We have studied the regulation of inhibin secretion by rat Sertoli cells grown on extracellular matrix-impregnated porous filters in a twin chamber assembly. Previous studies have established that rat Sertoli cells cultured under these conditions reproduce the morphological and functional polarization observed in the Sertoli cell in situ. Sertoli cells isolated from 18- to 22-day-old Wistar rats were cultured for up to 8 days with daily changes of fully defined supplemented Eagle's Minimum Essential Medium (MEM). Rat inhibin was measured by RIA and pituitary cell bioassay, and transferrin by RIA. Inhibin measured by immunoassay or bioassay was always readily detectable in the upper, but not the lower, chamber. Inhibin secretion into the upper chamber exhibited a dose-dependent stimulation of up to 3.7-fold by ovine FSH, with a medium effective dose of 2.2 micrograms/liter and a constant bio- to immunoreactive ratio (3.6 +/- 0.4). Apically directed secretion accounted for over 80% of inhibit output under basal conditions and over 94% with FSH stimulation. Insulin also stimulated upper chamber inhibin secretion at a high dose (5 mg/liter) but not at lower doses or in conjunction with FSH exposure of Sertoli cells. Testosterone augmented FSH-induced stimulation of inhibin secretion, but was ineffective without FSH exposure. In contrast to inhibin secretion, for which FSH is the principal regulator, transferrin secretion by Sertoli cells is more evenly bidirectional (overall mean upper to lower chamber ratio of 1.5) and requires exposure to other stimuli (insulin, retinoic acid, and testosterone) in addition to FSH to achieve maximal secretion. Both submaximal and maximal FSH stimulation of inhibin output were augmented by a phosphodiesterase inhibitor, isobutylmethylxanthine, and these effects were fully reproduced by forskolin, which suggests the involvement of cAMP in the vectorial secretion of inhibin. The marked polarization of Sertoli cell inhibin secretion in vitro could not be explained by restricted transmembrane passage of inhibin. It is, therefore, suggested that the bulk of inhibin secretion by the immature rat Sertoli cell in vivo may be directed primarily into the seminiferous tubular lumen. Thus, in addition to its role in endocrine negative feedback signaling to the pituitary, inhibin may also have important functions in seminiferous tubular function and the support of spermatogenesis.