The intrapituitary endocrine events during maturation and timing of puberty in the female sheep.

The aim of this study was to determine the maturational activity of gonadotroph cells, the site of synthesis, storage and release of luteinizing hormone (LH) and follicle-stimulating hormone (FSH) in Polish Merino female sheep born after the summer solstice. The actual time of puberty of these lambs was delayed until the following breeding season, when they were 14 months old. Changes were examined in 12 peripubertal (30-, 52-week-old) and pubertal (Days 15 and 17 of the second ovarian cycle) females. Histomorphological and functional changes in the gonadotroph population were assayed with hybridohistochemistry, immunohistochemistry, computer-assisted image analysis and radioimmunoassay. The percentage of the adenohypophyseal area (PAA) occupied by gonadotrophs containing LHbeta-mRNA was higher and the LH plasma concentration and pulse frequency were lower in the 52-week-old sheep in comparison with the 30-week-old sheep (P<0.05). The PAA occupied by immunoreactive (ir)-LHbeta-cells remained stable at the 30th and 52nd weeks of age and then increased at the pubertal follicular phase. The PAA occupied by ir-FSHbeta-cells was higher in the 52-week-old sheep compared with the 30-week-old sheep and then lower at the pubertal follicular phase (P<0.05). The PAA occupied by gonadotrophs containing LHbeta-mRNA or FSHbeta-mRNA was lower at the pubertal follicular phase in comparison with the 52nd week of age (P<0.05). In pubertal sheep, the PAA occupied by gonadotrophs containing LHbeta-mRNA or FSHbeta-mRNA was higher and the PAA occupied by ir-LHbeta or ir-FSHbeta-cells was lower at the preovulatory phase in comparison with the follicular phase of the cycle (P<0.05). In conclusion, the photoperiodic suspension of gonadotroph population's maturational functions has been observed at the level of LH storage and release but not at the level of LH synthesis during the expected time of puberty in female sheep of an aseasonal breed such as Merino. The findings show the heterogeneity in the patterns of LH and FSH post-transcriptional processing during the period of peripubertal/pubertal transition, explained by the different intrapituitary regulation at the level of post-transcriptional synthesis and storage rather, than at the level of release. Altogether, intrapituitary mechanisms of ovine maturation could have the histomorphological feature. Our observations prompt the hypothesis that the female lamb may be able to transduce changes in day length into the appropriate endocrine cues for sexual maturation after attainment by the pituitary gonadotroph population the full peripubertal efficiency, manifested by the sufficient storage of LH.

The effects of prolonged, intracerebroventricular prolactin treatment on luteinizing hormone secretion, catecholaminergic activity and estrous behavior in ewes.

Searching for the role of prolactin (PRL) in controlling gonadotropic axis activity in sheep, we studied the effects of prolonged, intracerebroventricular (i.c.v.) PRL infusion on luteinizing hormone (LH) secretion and catecholaminergic activity in the hypothalamic infundibular nuclei/median eminence (IN/ME) in sexually active ewes during the periovulatory period. Three groups of animals received the following treatments: 1). i.c.v. infusion of PRL at a dose of 200 microg/day (Lower dose, n = 5); 2). i.c.v. infusion of PRL at a dose of 400 microg/day (Higher dose, n = 6), and 3). i.c.v. infusion of the vehicle (control, n = 5). Each dose of PRL was infused in a pulsatile manner, 4 x 50 microg/h and 4 x 100 microg/h, in 30-min intervals, respectively, during four consecutive days before oncoming ovulation. The estrous behavior of ewes following treatments was also monitored as a determinant of the GnRH/LH surge. Two series of blood collections were made in every ewe, the first on the day preceding the infusion (day 0 of the experiment), the second on the day after the infusion (day 5 of the experiment). In addition, on day 5 of the experiment, perfusions of the IN/ME were made by the push-pull method, either in control or lower dose-treated animals. It was shown that a significant (p < 0.01, p < 0.001) increase in tonic LH secretion during the periovulatory period remained in ewes irrespective of the kind of infusion. No statistical differences were found in LH pulse frequency, amplitude, or in the length of the pulse when compared with values from day 0 and 5 of the experiment within each group. A significant (p < 0.001) increase in IN/ME perfusate concentrations of dopamine and noradrenaline metabolites was noted in PRL-treated ewes in comparison with those in the control. The estrous behavior in PRL-treated animals was delayed for a few days, 3.80 +/- 0.80 days at the lower dose (p < 0.01), and 2.83 +/- 0.98 days at the higher dose (p < 0.05) in comparison with the control, 0.20 +/- 0.20 days. These data indicate that maintenance of an increased PRL concentration within the central nervous system (CNS) for a few days before oncoming ovulation has no inhibitory effect on tonic LH secretion. A few-day shift of the preovulatory GnRH/LH surge, as determined by estrous behavior, might, however, be a consequence of the PRL-induced increase in catecholamine turnover in the IN/ME.

The effect of dietary protein restriction on the secretion of LH and FSH in pre-pubertal female lambs.

The effect of restricted dietary protein on the synthesis, storage and release of LH and FSH was studied in pre-pubertal female lambs. The experiment started when the lambs were aged 12 weeks and weighed 26.0+/-1.6 kg. It was conducted for 25 weeks. The lambs were fed isocaloric diets containing either a restricted level of crude protein (8% CP; n=6; treatment R) or an elevated one (18% CP; n=4; treatment E). At 37 weeks of age and before the first oestrous cycle, blood samples were collected over 6 h at 10 min intervals for LH assay. The lambs were slaughtered and their brains recovered and fixed in situ. Immuno-reactive (IR) LH and FSH cells were localised by immunohistochemistry techniques. Messenger RNA analyses used by non-isotope in situ hybridisation with sense and anti-sense riboprobes from beta subunits of LH and FSH cDNA clones. Data were generated using computer analysis to measure the proportion of IR and/or hybridising cells and their optical density for immuno-staining and hybridisation signal. Plasma LH was measured by RIA. The daily live-weight gains were 56.5+/-13.1 g and 97.8+/-14.3 g for R and E lambs, respectively (P<0.05), so that final weights at slaughter were 36.1+/-1.97 kg and 39.1+/-3.44 kg, respectively (P<0.05). The number of cells expressing LH beta mRNA and the optical density of this hybridisation signal was significantly (P<0.001) lower in the R lambs but the number of IR LH positive cells was higher (P<0.001) than for the E lambs. The concentration of LH in the plasma of R sheep was lower (P<0.05) than the E group and this response was associated with a decrease (P<0.05) in LH pulse frequency and amplitude. Dietary protein concentration appeared to have no effect on the IR in FSH cells or on the expression of FSH beta mRNA. In summary, the low protein diet influenced the body weight and weight gain of growing lambs and exerted an inhibitory effect on the synthesis and the release of LH in the pituitary gonadotrophs. No such effect was observed for FSH. It was concluded that the protein concentration of the diet consumed during the growth of female lambs may be an important modulator of processes leading to the pre-pubertal rise in LH.