In this study we used an isolation/restraint stress to test the hypothesis that stress will affect the secretion of LH differently in gonadectomised rams and ewes treated with different combinations of sex steroids. Romney Marsh sheep were gonadectomised two weeks prior to these experiments. In the first experiment male and female sheep were treated with vehicle or different sex steroids for 7 days prior to the application of the isolation/restraint stress. Male sheep received either i.m. oil (control rams) or 6 mg testosterone propionate injections every 12 h. Female sheep were given empty s.c. implants (control ewes), or 2 1 cm s.c. implants containing oestradiol, or an intravaginal controlled internal drug release device containing 0·3 g progesterone, or the combination of oestradiol and progesterone. There were four animals in each group. On the day of application of the isolation/ restraint stress, blood samples were collected every 10 min for 16 h for the subsequent measurement of plasma LH and cortisol concentrations. After 8 h the stress was applied for 4 h. Two weeks later, blood samples were collected for a further 16 h from the control rams and ewes, but on this day no stress was imposed. In the second experiment, separate control gonadectomised rams and ewes (n=4/ group) were studied for 7 h on 3 consecutive days, when separate treatments were applied. On day 1, the animals received no treatment; on day 2, isolation/restraint stress was applied after 3 h; and on day 3, an i.v. injection of 2 µg/kg ACTH 1-24 was given after 3 h. On each day, blood samples were collected every 10 min and the LH response to the i.v. injection of 500 ng GnRH administered after 5 h of sampling was measured. In Experiment 1, the secretion of LH was suppressed during isolation/ restraint in all groups but the parameters of LH secretion (LH pulse frequency and amplitude) that were affected varied between groups. In control rams, LH pulse amplitude, and not frequency, was decreased during isolation/restraint whereas in rams treated with testosterone propionate the stressor reduced pulse frequency and not amplitude. In control ewes, isolation/restraint decreased LH pulse frequency but not amplitude. Isolation/restraint reduced both LH pulse frequency and amplitude in ewes treated with oestradiol, LH pulse frequency in ewes treated with progesterone and only LH pulse amplitude in ewes treated with both oestradiol and progesterone. There was no change in LH secretion during the day of no stress. Plasma concentrations of cortisol were higher during isolation/restraint than on the day of no stress. On the day of isolation/restraint maximal concentrations of cortisol were observed during the application of the stressor but there were no differences between groups in the magnitude of this response. In Experiment 2, isolation/restraint reduced the LH response to GnRH in rams but not ewes and ACTH reduced the LH response to GnRH both in rams and ewes. Our results show that the mechanism(s) by which isolation/restraint stress s...

This minireview considers the state of knowledge regarding the interactions of testicular hormones to regulate the secretion and actions of GnRH in males, with special focus on research conducted in rams and male rhesus monkeys. In these two species, LH secretion is under the negative feedback regulation of testicular steroids that act predominantly within the central nervous system to suppress GnRH secretion. The extent to which these actions of testicular steroids result from the direct actions of testosterone or its primary metabolites, estradiol or dihydrotestosterone, is unclear. Because GnRH neurons do not contain steroid receptors, the testicular steroids must influence GnRH neurons via afferent neurons, which are largely undefined. The feedback regulation of FSH is controlled by inhibin acting directly at the pituitary gland. In male rhesus monkeys, the feedback regulation of FSH secretion is accounted for totally by the physiologically relevant form of inhibin, which appears to be inhibin B. In rams, the feedback regulation of FSH secretion involves the actions of inhibin and testosterone and interactions between these hormones, but the physiologically relevant form of inhibin has not been determined. The mechanisms of action for inhibin are not known.

Plasma follistatin (FS) concentrations were determined after castration (n = 5) or sham castration (n = 4) of mature rams. Both treatments resulted in a prolonged increase in FS between 7 and 19 h after surgery, which returned to pretreatment concentrations by 24 h. Tumour necrosis factor-alpha (TNF-alpha), a sensitive maker of an acute-phase response, was undetectable in plasma, indicating that the FS response was not induced by trauma due to surgery. In a second experiment, injection of castrated rams (n = 4) with ovine recombinant interleukin-1 beta, an acute-phase mediator, resulted in a sustained rise in FS concentrations within 4 h of injection. Plasma TNF-alpha concentrations increased transiently within 1 h of interleukin-1 beta injection, indicating that an acute-phase response had been initiated. Plasma follicle-stimulating hormone (FSH) concentrations were significantly decreased at 8 and 24 h after interleukin-1 beta injection, strongly suggestive of an inhibitory effect of increased FS concentrations on the secretion of FSH. Injection of castrated rams (n = 2) with a control preparation of recombinant interleukin-2 did not induce an acute-phase response, and plasma FS and FSH concentrations were unaffected. These data show that the testis is not a major source of circulating FS, that the increase in circulating FS following sham castration/castration is not due to an acute-phase response, but that conversely FS concentrations are modulated by the acute-phase mediator, interleukin-1 beta.

This study tested the hypothesis that inhibin is a major negative feedback regulator of FSH secretion but has minimal effects on LH secretion in rams. In experiment 1, castrated rams (wethers) were given either vehicle or human recombinant inhibin A (hr-inhibin) as three s.c. or three i.v. 50-micrograms injections 6 h apart or as one 50-micrograms i.v. injection followed by 100-micrograms infusion over 12 h. Human recombinant inhibin suppressed plasma FSH while the vehicle had no effect. The greatest suppression in plasma FSH was achieved following i.v. administration of hr-inhibin given either by repeated injection or by infusion. In experiment 2, wethers were given vehicle or a 50-micrograms i.v. injection followed by 800-micrograms infusion of hr-inhibin over 12 h. Infusion of hr-inhibin suppressed plasma FSH with a maximal suppression of 53.3% occurring between 15 and 24 h after the start of treatment. During this period, the plasma concentrations of FSH and inhibin were in the range of values for intact rams. Human recombinant inhibin did not influence plasma LH in either experiment. This study demonstrated that physiological treatment with inhibin, in the absence of testosterone, has the capacity to suppress plasma concentrations of FSH in wethers to the levels found in intact rams.

Three groups of mature rams were maintained on diets of hay, hay + 2% lupin or hay + 2% cowpea for 11 weeks. Serial blood samples were taken at 15-min intervals for 12 h for the determination of GH and IGF-I content by radioimmunoassay and for IGF-binding protein-3 (IGFBP-3) levels by Western blotting. The rams were killed after 77 days of supplementary feeding and their pituitary glands analysed for content of GH and GH mRNA. Mean plasma GH and baseline GH levels were significantly (P < 0.01) decreased in the rams fed lupin and cowpea compared with controls fed hay and GH pulse amplitude was significantly (P < 0.001) decreased in the group fed the cowpea diet. The frequency of GH pulses was not significantly altered by either treatment. Plasma concentrations of IGF-I were elevated in rams fed lupin (P < 0.001) or cowpea (P < 0.05). IGFBP-3 levels were not significantly (P > 0.05) altered by either treatment. There were no significant differences in pituitary content of GH mRNA but pituitary content of GH was increased in rams fed lupin (P < 0.05) and cowpea (P = 0.07). In conclusion, a high-protein diet decreases plasma GH levels and increases IGF-I without changing plasma IGFBP-3 levels in rams. Thus ongoing synthesis of GH, as indicated by the mRNA levels, may cause a build up of GH stores in the pituitary gland.