Male voles were reared from birth to the age of 56 days in photoperiods of 16L:8D or 6L:18D. In 16L:8D testes increased 10-fold in size between the age of 14 and 56 days, and there were concomitant increases in diameter of seminiferous tubules and seminal vesicle weight. Spermatozoa were present in tubules by 35 days. In 6L:18D no significant changes with age occurred in testis size, diameter of seminiferous tubules or seminal vesicle weight. LH secretion increased with age in 16L:8D, but not in 6L:18D, and pituitary and plasma levels were higher in the former than in the latter by 35 days. Pituitary levels of FSH were high up to 21 days in 16L:8D, and up to 35 days in 6L:18D. At 35 days the mean pituitary level was significantly higher in the 6L:18D than in 16L:8D. Plasma levels of FSH were significantly higher in 16L:8D at 14 and 21 days, and then gradually declined. In 6L:18D plasma FSH rose to a maximum at 35 days, but did not reach the levels occurring at an earlier age in voles in 16L:8D. The relationship between pituitary and plasma levels of FSH in the two photoperiods is compatible with the idea of a storage and a readily releasable pool of the hormone within the pituitary gland. In 16L:8D changing levels of LH and FSH seem to be due to the onset of testicular activity with consequent feedback effects of testicular hormones upon the anterior pituitary gland.

A monoclonal antibody was made after immunization of mice with the 1-32 amino terminal peptide of the alpha subunit of 32K human ovarian inhibin. The IgG2a mouse antibody reacted 6 times better with bovine 1-32 peptide than it did with 32K bovine inhibin. By contrast sheep polyclonal antibodies made by a similar method had a 29 fold bias in reactivity towards the immunizing peptide. Relative to homologous 1-32 peptide standards, the monoclonal antibody measured apparently higher amounts of immunoreactive material(s) in human (13.5 fold) and bovine (27 fold) follicular fluids than did the polyclonal anti 1-32 peptide antibodies. Immunochemical studies revealed that the epitope recognized by the monoclonal antibody was different from the major epitope recognized by the polyclonal antibodies. The monoclonal antibody reacted much better with human inhibin 1-32 sequences than with bovine (73 fold) or porcine (23 fold). Although the 32K form of human inhibin has not yet been purified, it can be inferred that the monoclonal antibody would be able to detect as little as 2 ng/ml of 32K human inhibin in competitive radioimmunoassays. The antibody must also react with some of the multiple molecular forms of inhibin found in human follicular fluids, and it was shown to function well in the quantitative immunoaffinity extraction of inhibin-like immunoreactivity from follicular fluid. It seems likely that this monoclonal antibody will prove a useful tool for research on human inhibin.

The investigation of the role of inhibin in the regulation of fertility is hindered by the lack of a routine, specific assay. The pituitary cell bioassay is time-consuming and the existing RIAs, based on either purified bovine 32-kDa inhibin or synthetic alpha-subunit peptides, are not specific for the biologically active inhibin molecules. We have used monoclonal antibodies, one specific for the N-terminal region of the human inhibin alpha chain, and the other raised to a peptide sequence close to the C-terminal of the human beta A-inhibin chain, to create a two-site sandwich ELISA specific for alpha beta-inhibin molecules. This was used to estimate levels of inhibin in crude bovine and human follicular fluids and fractions concentrated from them. Comparison of the values obtained with the ELISA and those obtained with the pituitary cell bioassay, suggests that the ELISA measures biologically active inhibin. Compared with the peptide-based RIA, the ELISA gave much lower (as little as 100-fold lower) values for the inhibin content of these samples, e.g., bovine follicular fluid 0.375 micrograms/ml (ELISA) compared with 41.0 micrograms/ml (RIA). Such large differences, possibly due to the presence of relatively large amounts of biologically inactive forms of inhibin such as the pro-alpha c or free alpha forms, suggest that those RIAs, which essentially measure the level of alpha-inhibin, considerably overestimate the levels of the active forms of inhibin in the samples and that results obtained using these assays may need reinterpretation.

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Male voles were raised from birth to 100 days of age in photoperiods of 16L:8D or 6L:18D. In the long photoperiod testes increased in size between 15 and 80 days of age, and there was an increase in seminal vesicle weight from 60 days of age. Spermatozoa were present in the testes at 60 days of age. In the short photoperiod testicular growth did not begin until 50 days of age with the seminal vesicles beginning to increase at 80 days of age. Spermatozoa were present in testes at 100 days of age. Pituitary secretion in vitro of LH and FSH in response to 1 pmol GnRH, as well as hypothalamic GnRH content, rose to peaks at 50 and 80 days of age respectively in animals exposed to long photoperiods. There was no change in pituitary secretion of FSH in response to GnRH stimulation in animals from the short photoperiod. However, pituitary release of LH in response to 1 pmol GnRH rose to a peak at 80 days of age. Hypothalamic GnRH content rose to a peak at 50 days of age and then declined. The relationship between the hypothalamic GnRH and the sensitivity of the pituitary to GnRH stimulation is compatible with the idea that GnRH can mediate its own receptor numbers.