Identification of luteinizing hormone releasing hormone-like immunoactivity in ovine cotyledons

Luteinizing hormone releasing hormone (LHRH)-like immunoactivity has been identified in cotyledonary extracts prepared from pregnant ewes. This activity displayed similar physico-chemical properties as synthetic LHRH, as determined by reverse-phase HPLC and size-exclusion HPLC. Under reverse-phase conditions, cotyledonary LHRH-like immunoactivity displayed a retention time (10.5 +/- 0.1 min) which was not significantly different from that of synthetic LHRH. When subjected to size-exclusion HPLC, cotyledonary LHRH-like immunoactivity eluted in fractions which corresponded to a molecular weight range of 1100-1200 Da, which was not significantly different from the elution profile observed for synthetic LHRH. The cotyledonary tissue content of LHRH-like immunoactivity averaged 94 +/- 24 pg/mg (n = 6). The results of this study demonstrate the presence of LHRH-like immunoactivity in ovine cotyledons. Although placental synthesis of LHRH-like immunoactive material has been demonstrated in other species, it remains to be established whether this activity, demonstrated in ovine placenta, is the consequence of de novo placental synthesis or represents uptake from the maternal (and/or fetal) circulation.     

Absence of specific luteinizing hormone releasing hormone receptors in ovine, bovine and porcine ovaries

Crude and membrane-enriched homogenates of unfrozen follicular and luteal tissue from cows, ewes and sows were assayed for the presence of specific luteinizing hormone releasing hormone (LHRH) receptors by one-point saturation analysis using [D-Ser-(TBU)6, des-Gly-NH2(10)] LHRH-EA as the labeled and unlabeled ligand. Pituitaries from cows, ewes, sows and rats, and rat ovaries served as positive controls and were assayed with each ovarian tissue assay. Scatchard analysis was used to determine binding affinity of pools of ovarian and pituitary tissue. Specific high-affinity LHRH receptors were found in the pituitaries of cows, ewes, sows and rats and in the rat ovary. In contrast, no specific LHRH binding was detected in follicular or luteal tissue of cows, ewes or sows. Thus, unlike the rat ovary which contains LHRH receptors, ovaries from these domestic species lack specific LHRH receptors.     

Leydig cell receptors for luteinizing hormone releasing hormone and its agonists and their modulation by administration or deprivation of the releasing hormone

Leydig cells isolated from adult rat testes bound ¹²⁵I-labelled luteinizing hormone releasing hormone (LHRH) agonist with high affinity (K_A=1.2 × 10⁹M) and specificity. LHRH and the 3–9 and 4–9 fragments of LHRH agonist competed for binding sites with ¹²⁵I-LHRH agonist but with reduced affinities, whereas fragments of LHRH, and oxytocin and TRH were largely inactive. Somatostatin inhibited binding at high (10^?4M) concentrations but was inactive at 10^?6M and less. Pretreatment of rats for 7 days with 5 μg/day of LHRH agonist reduced binding of ¹²⁵I-LHRH agonist to Leydig cells $\text{in vitro}$ by 25%, whilst inhibition of endogenous LHRH by antibodies for 7 days caused a 40% decrease.     

Short-term pulsatile administration of luteinizing hormone releasing hormone in male adolescents with multiple idiopathic pituitary hormone deficiencies

In order to define both level and severity of defect in patients with idiopathic multiple pituitary hormone deficiencies (MPHD) and to find out which patient might benefit from pulsatile LHRH substitution therapy, the effect of short-term pulsatile LHRH infusion in 6 affected male adolescents was studied. Controls were 9 boys with constitutional delay of puberty (CD). During a spontaneous nocturnal plasma profile LH and FSH levels were prepubertal with little evidence of pulsatile secretory LH activity in all MPHD patients. During short-term pulsatile LHRH stimulation (36 h), however, all showed a significant rise in mean LH and FSH levels (p less than 0.0001). Linear regression analysis revealed significant continuous increases of FSH (p less than 0.001) in all patients and of LH (p less than 0.01) in all but one patient. These changes were not accompanied by an increase of testosterone, androstenedione and DHAS levels. Since all MPHD patients showed steadily increasing gonadotropin levels if stimulated in a pulsatile manner, we conclude that the defect might only in part be located at the pituitary level. Long-term pulsatile substitution therapy with LHRH is likely to be successful in these patients as has been demonstrated in patients with known hypothalamic defect.     

The induction of divergent gonadotropin secretion in vitro by variations in luteinizing hormone releasing hormone pulse regimen

The effects of luteinizing hormone releasing hormone (LHRH) pulse amplitude, duration, and frequency on divergent gonadotropin secretion were examined using superfused anterior pituitary cells from selected stages of the rat estrous cycle. Cells were stimulated with one of five LHRH regimens. With low-amplitude LHRH pulses (regimen 1) in the presence of potentially estrogenic phenol red, LH response in pituitary cells from proestrus 1900, estrus 0800, and diestrus 1,0800 were all significantly larger (P less than 0.05) than the other stages tested. In the absence of phenol red, responsiveness at proestrus 1900 was significantly larger than proestrus 0800, proestrus 1500, and estrus 0800 (P less than 0.01, 0.05, and 0.05, respectively); other cycle stages tested were smaller. No significant differences were observed between cycle stages for follicle-stimulating hormone (FSH) secretion in the presence or absence of phenol red. Because pituitary cells at proestrus 1900 were the most responsive to low-amplitude 4 ng LHRH pulses, they were also used to study the effects of LHRH pulses of increased amplitude or duration and decreased frequency. Increasing the amplitude (regimen 2) or the duration (regimens 3 to 5) increased FSH secretion; this effect was greatest with regimens 3 and 5. When regimens 3 and 5 were studied in pituitary cells obtained at proestrus 1500, FSH was significantly increased by both regimes, but most by regimen 5; furthermore, LH release was significantly reduced. When regimens 3 and 5 were studied in pituitary cells obtained at estrus 0800, FSH release was elevated most significantly by regimen 5. Thus, variations in LHRH pulse regimen were found to be capable of inducing significant divergence in FSH release from superfused anterior pituitary cells derived from specific stages of the estrous cycle.     

Exposure to ram wool stimulates gonadotropin-releasing hormone pulse generator activity in the female goat

In the sheep and goat, exposure of anestrous females to a conspecific male odor enhances reproductive activity. Interestingly, a previous report indicated that male goat hair stimulated pulsatile luteinizing hormone (LH) secretion in the ewe. In the present study, we addressed whether ram wool affects the gonadotropin-releasing hormone (GnRH) pulse generator activity in the female goat. Five ovariectomized (OVX) goats were chronically implanted with recording electrodes in the mediobasal hypothalamus, and manifestations of the GnRH pulse generator were monitored as characteristic increases in multiple-unit activity (MUA volleys). Wool or hair samples were collected from a mature ram, ewe and male goat, and their effects on the MUA volley were examined. The exposure to ram wool induced an MUA volley within 1 min in all five OVX goats, as did the exposure to male goat hair. The ewe wool had no effect on the timing of an MUA volley occurrence. An invariable association of MUA volleys with LH pulses in the peripheral circulation was also confirmed in two OVX goats exposed to ram wool. The present results clearly indicate that exposure to ram wool stimulates pulsatile GnRH/LH release in the female goat. Since exposure to male goat hair enhances pulsatile LH secretion in the ewe, it is likely that very similar, if not identical, molecules are contained in the male-effect pheromone in the sheep and goat.     

Effect of luteinizing hormone releasing hormone pulse characteristics on comparative luteinizing hormone and follicle stimulating hormone secretion from superfused rat anterior pituitary cell cultures

We have shown that 4 ng luteinizing hormone releasing hormone (LHRH) pulses induced significantly greater luteinizing hormone (LH) release from proestrous rat superfused anterior pituitary cells with no cycle related differences in follicle stimulating hormone (FSH). Current studies gave 8 ng LHRH in various pulse regimens to study amplitude, duration and frequency effects on LH and FSH secretion from estrous 0800, proestrous 1500 and proestrous 1900 cells. Regimen 1 gave 8 ng LHRH as a single bolus once/h; regimen 2 divided the 8 ng into 3 equal 'minipulses' given at 4 min intervals to extend duration; regimen 3 gave the 3 'minipulses' at 10 min intervals, thereby further extending duration: regimen 4 was the same as regimen 2, except that the 3 'minipulses' were given at a pulse frequency of 2 h rather than 1 h. In experiment 1, all four regimens were employed at proestrus 1900. FSH was significantly elevated by all 8 ng regimens as compared to 4 ng pulses; further, 8 ng divided into 3 equal 'minipulses' separated by 4 min at 1 and 3 h frequencies (regimens 2 and 4) resulted in FSH secretion that was significantly greater than with either a single 8 ng bolus (regimen 1) or when the 'minipulses' were separated by 10 min (regimen 3). In experiment 2, at proestrus 1500, FSH response to the second pulse of regimen 4 was significantly greater than in regimen 2; LH release was significantly suppressed at pulse 2 compared to regimen 2 accentuating divergent FSH secretion. At estrus 0800, FSH response to the second pulse of regimen 4 was significantly stimulated FSH at proestrus 1900, 1500 and estrus 0800, FSH divergence was most marked at proestrus 1500. These data indicate a potential role for hypothalamic LHRH secretory pattern in inducing divergent gonadotropin secretion in the rat.     

In vivo regulation of adrenocorticotrophin secretion in the immature ovine fetus. Modulation by ovine corticotrophin releasing hormone and arginine vasopressin

This study investigates the in vivo regulation of ACTH secretion in the immature ovine fetus by AVP and oCRH. Previously we have demonstrated that whilst AVP-containing neurones are present from 42 days, oCRH-containing neurones cannot be detected in the fetal paraventricular nucleus or median eminence until after 90 or 100 days respectively. In acutely exteriorized fetuses aged between 64-90 days (n = 5), a haemorrhagic stress elicited a significant increase (P less than 0.01) in ACTH values. There was also a significant correlation between plasma ACTH and AVP concentrations in these fetuses. In chronically cannulated fetuses less than 100 days (n = 6) injection of AVP (200 ng) significantly elevated fetal plasma ACTH values at 10 min (P less than 0.01) post injection. Simultaneous injection of AVP (200 ng) and oCRH (10 micrograms) into these fetuses produced a plasma ACTH value that was significantly greater at 10 min (P less than 0.05) than the summed response obtained with separate injection of oCRH and AVP. When AVP and oCRH were injection in equimolar amounts to fetuses between 101-118 days, AVP (2 micrograms) was found to have a greater effect on ACTH than was oCRH (2 micrograms). Pretreatment with a specific vascular antagonist of AVP-d(CH2)5Tyr(Me)AVP failed to significantly inhibit the increase in mean arterial pressure associated with AVP (2 micrograms) injection but partially antagonized (P = 0.04) the decrease in fetal heart rate. The antagonist however completely abolished any effect of AVP on fetal plasma ACTH values. This study suggests that AVP most likely acts through receptors in the fetal pituitary with V1 characteristics.     

Effects of luteinizing hormone releasing hormone on gonadotrophins in the hamster

The changes in serum gonadotrophins in male hamsters following one injection of 15 μg luteinizing hormone releasing hormone (LHRH) (Group A) were compared with those following the last injection of LHRH in animals receiving an injection approximately every 12 hr for 4 days (Group B) or 12 days (Group C). Peak follicle stimulating hormone (FSH) levels (ng/ml) were 1776±218 (Group A), 2904±346 (Group B), and 4336±449 (Group C). Peak luteinizing hormone (LH) values (ng/ml) were 1352±80 (Group A), 410±12 (Group B), and 498±53 (Group C). Serum FSH:LH ratios, calculated from the concentrations measured 16 hr after the last LHRH injections, were higher in Groups B and C than in Group A. Similar injections of LHRH (100 ng or 15 μg/injection) for 6 days elevated the serum FSH:LH ratio in intact males. Five such LHRH injections (100 ng/injection) blunted the rise in serum LH in orchidectomized hamsters. Direct effects of LHRH on gonadotrophin secretory dynamics or altered brain-pituitary-testicular interactions may alter the ratio of FSH to LH in the hamster.     

On the existence and separation of the follicle stimulating hormone releasing hormone from the luteinizing hormone releasing hormone.

Porcine hypothalamic fragments were extracted by 2M AcOH at 4°C, and the extractives were subsequently processed in the presence of one protease inhibitor and one anti-oxidant. Gel filtration was performed on Bio-Gel P-2, and supplementary [³H]-LHRH and [¹⁴C]- 3H]-LHRH, and was differentiated from [¹⁴C]- 相似文献   

Changes in prolactin levels caused by luteinizing hormone releasing hormone

The acute effects of luteinizing hormone releasing hormone (LHRH) on the release of prolactin (PRL) were investigated in 12 normal cycling women and 42 women with various menstrual disorders. LHRH (100 micrograms) was bolusly injected intramuscularly and PRL levels were measured immediately before the injection and at 30 minutes and 60 minutes after the injection. LHRH elicited an increase of more than 25% in PRL levels in 15 cases (27.8%) at both 30 minutes and 60 minutes after the injection, whereas PRL levels were decreased by more than 25% in 7 cases (13.0%). The PRL response to LHRH seemed to be related to basal PRL levels. Especially when the PRL concentration was 20 ng/ml or more, LHRH decreased PRL levels in 7 cases out of 16. On the other hand, LHRH increased PRL levels in the majority of cases with a PRL concentration less than 20 ng/ml. In conclusion, the LHRH injection occasionally alters PRL levels in either a positive or negative manner, depending upon the basal PRL levels.     

N-methyl-d, l-aspartate stimulates growth hormone but not luteinizing hormone secretion in the sheep

The objective of this experiment was to determine the effects of N-methyl-d, l-aspartate (NMA) on luteinizing hormone (LH) and growth hormone (GH) secretion in castrated male sheep. Blood was sampled from Hampshire wethers every 15 min for 8 hr on day 1. At 4 and 6 hr after the initiation of the experiment, wethers were treated i.v. with NMA at a dose of 12 mg/kg body weight (n = 5) or .9% saline (n = 5). The dosage of NMA was within the range of doses that was previously demonstrated to stimulate LH secretion in monkeys. Blood samples were also collected every 15 min for 1 hr on day 2, beginning 24 hr after the first injection of NMA or saline. Treatment with NMA had no effect on mean LH concentrations, LH pulse frequency or LH pulse amplitude during the 4 hr period following the first injection on day 1. On day 2, however, mean LH concentrations were lower (p less than .01) in NMA versus saline-treated wethers. Conversely, administration of NMA evoked a dramatic increase (p less than .02) in mean GH concentrations on day 1. The mechanisms responsible for the effects of NMA described herein and whether or not these effects are relevant to the physiological control of LH and GH release in the sheep warrants further scrutiny.     

Effects of prolactin on the luteinizing hormone response to gonadotropin- releasing hormone in primary pituitary cell cultures during the ovine annual reproductive cycle

In the sheep pituitary, the localization of prolactin (PRL) receptors in gonadotrophs and the existence of gonadotroph-lactotroph associations have provided morphological evidence for possible direct effects of PRL on gonadotropin secretion. Here, we investigated whether PRL can readily modify the LH response to GnRH throughout the ovine annual reproductive cycle. Cell populations were obtained from sheep pituitaries during the breeding season (BS) and the nonbreeding season (NBS), plated to monolayer cultures for 7 days, and assigned to receive one of the following treatments: 1) nil (control), 2) acute (90- min) bromocriptine (ABr), 3) chronic (7-day) bromocriptine (CBr), 4) ABr and PRL, 5) CBr and PRL, 6) PRL alone, or 7) thyrotropin-releasing hormone. Cells were treated as described above, with the aim of decreasing or increasing the concentrations of PRL in the culture, and simultaneously treated with GnRH for 90 min. The LH concentrations in the medium were then determined by RIA. GnRH stimulated LH in a dose-dependent manner during both stages of the annual reproductive cycle. During the NBS, single treatments did not significantly affect the LH response to GnRH. However, when PRL was combined with bromocriptine, either acutely or chronically, GnRH failed to stimulate LH release at all doses tested (P < 0.01). In contrast, during the BS, the LH response to GnRH was not affected by any of the experimental treatments. These results reveal no apparent effects of PRL alone, but an interaction between PRL and dopamine in the regulation of LH secretion within the pituitary gland, and a seasonal modulation of this mechanism.