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Peripheral plasma levels of luteinizing hormone-releasing hormone-like immunoreactivity and luteinizing hormone in anestrous and ovariectomized sheep.

Low levels of endogenous LHRH-like immunoreactivity (IR) were consistently detected in methanolic extracts of plasma samples from anestrous and ovariectomized sheep. Mean concentrations were similar in magnitude in the intact and castrate ewes (7.7 +/- 7.0 and 6.1 +/- 0.8 pg/ml, respectively) but revealed greater within animal fluctuations in the ovariectomized sheep (CVs: anestrous ewes = 20 +/- 3%; ovariectomized ewes = 45 +/- 10%). Patterns of plasma LHRH-like IR and LH levels were not obviously related. Differences in levels of LHRH-like IR did exist between ewes and in the ovariectomized sheep the levels were positively correlated (r = .83, P less than ,05) with the pituitary LH response to synthetic LHRH injection (0.25 micrograms). Large plasma volumes (250 ml) were extracted using chloroform, methanol and acetic acid and then chromatographed using Sephadex LH-20. Plasma LHRH-like IR could be concentrated and was found to have a column elution volume coincidence with that of synthetic LHRH. These data indicate that there is a substance in ovine plasma which is close in size, solubility and IR to sLHRH. It does not reflect LH levels in either magnitude or pattern but does exhibit a coincident increase in variation with plasma LH levels following ovariectomy and appears to be associated with the pituitary responsiveness.

Animals↗

Effects of applying gamma-aminobutyric acid(B) drugs into the medial basal hypothalamus on basal luteinizing hormone concentrations and on luteinizing hormone surges in the female sheep.

Prior investigations have shown that localized infusion by microdialysis of gamma-aminobutyric acid(B) (GABA(B)) agonists into the medial basal hypothalamus of male sheep rapidly increases GnRH and LH pulse amplitude. The objectives of these studies were to determine if infusion of GABA(B) agonists SKF 97541 or baclofen into the medial basal hypothalamus of female sheep would affect basal LH secretion and if infusion of a potent antagonist would alter expression of LH surges induced by injection of estrogen. Infusion of either SKF 97541 (10 or 40 microM) or baclofen (1 mM) into estrogen-treated ovariectomized ewes did not alter basal LH secretory patterns, whereas both drugs significantly elevated mean LH and LH pulse amplitude in ovariectomized ewes during the nonbreeding season. Infusion of the antagonist CGP 52432 (250 or 500 microM) did not affect expression of estrogen-induced LH surges in ovariectomized ewes. These observations support the concept that GABA(B) receptors in the medial basal hypothalamus regulate basal LH secretion but do not regulate the surge mode of LH secretion in the female sheep.

Animals↗

Effects of chronic treatment with a potent luteinizing hormone releasing hormone agonist on serum luteinizing hormone and steroid levels in the male rhesus monkey.

The effects of chronic administration of [D-Ser(TBU)6, des-Gly-NH2 10]LHRH ethylamide (Buserelin, Hoe 766) on plasma LH, pregnenolone, 17-hydroxyprogesterone, testosterone and dihydrotestosterone levels were studied in a nonhuman primate species, the rhesus monkey. After 1 week of daily administration of 25 micrograms of Hoe 766, the LH response measured after 3 h and 7 h is markedly inhibited when compared to the previous week's response and a slight decrease in basal LH levels is observed. A small rise of plasma LH is, however, always seen 3 h after administration of the peptide during the whole treatment period. Serum testosterone levels display a rapid stimulation from 7 to 35 ng/ml 3 h after the first administration of Buserelin and increase progressively up to at least 12 h (44 ng/ml). Plasma concentrations of 17-hydroxyprogesterone increase more slowly from 2.0 to 3.4 ng/ml during the first 3 h and reach 9.5 ng/ml 9 h later. By contrast, there are no or minimal changes or serum pregnenolone and dihydrotestosterone levels following the first injection of Hoe 766 and during the course of the study. While the acute testosterone response to Buserelin measured at 3 h is not affected in up to seven weeks of treatment with the peptide, the duration of testosterone and 17-hydroxyprogesterone response measured at later time intervals (7 h, 12 h) is decreased to 20% to 50% of control. The data suggest that the testicular steroidogenic pathway in rhesus monkeys retain its ability to respond to LH stimulation during treatment with the LHRH agonist while the basal testosterone levels are reduced. The close correlation observed between reduced serum LH and testosterone concentrations suggest that pituitary desensitization is the main factor responsible for the partial inhibition of basal testosterone concentration in rhesus monkeys.

Animals↗

Effect of estrogen administration on endogenous and luteinizing hormone-releasing-hormone-induced luteinizing hormone secretion and follicular development in the lactating sow.

The objectives of this study were to investigate whether estradiol treatment during lactation modifies 1) the patterns of endogenous LH, FSH, and prolactin (PRL) release; 2) the sensitivity of the pituitary to exogenous injections of LHRH; and 3) the responsiveness of the ovarian follicles to gonadotropin. Plasma LH, FSH, and PRL were determined in samples taken repeatedly from 18 sows on Days 24-27 of lactation. Ovaries were then recovered, and follicular development was assessed by measuring the follicular diameter (FFD) and follicular fluid estradiol-17 beta concentration (FFE) of the ten largest follicles dissected from each ovary. Sows were randomly allocated to one of four treatments: 1) Group C (4 sows) received no treatment; 2) Group LHRH (5 sows) received 800 ng of LHRH every 2 h throughout the sampling period; 3) Group E2 (4 sows) received subcutaneous implants containing estradiol-17 beta 24 h after start of sampling; 4) Group LHRH + E2 (5 sows) were administered a combination of LHRH and estradiol-17 beta implants. Between-animal variability for plasma LH, FSH, and PRL was considerable. LH concentration and LH pulse frequency increased (p less than 0.05) after LHRH treatment in the LHRH and LHRH + E2 groups; however, an acute inhibition of LH secretion was observed in the latter group immediately after estradiol implant application. In the absence of LHRH treatment, estradiol caused chronic inhibition of LH secretion. Follicular development was greater in the LHRH and LHRH + E2 groups compared to the C and E2 groups (p less than 0.05 for both FFD and FFE).(ABSTRACT TRUNCATED AT 250 WORDS)

Animals↗

Agonist-induced release of gonadotropin-releasing hormone, luteinizing hormone, and follicle-stimulating hormone and their associations with basal secretion of luteinizing hormone and follicle-stimulating hormone throughout lactation in sows.

Our working hypothesis was that the normal increase in basal secretion of LH and FSH during lactation in sows is paralleled by an increase in readily releasable pools of GnRH, LH, and FSH. Sows fitted with indwelling catheters and nursing 9-11 piglets were sampled on Days 1, 7, 14, and 21 (primiparous sows; n = 10; experiment 1) or Days 7 and 21 (multiparous sows; n = 10; experiment 2) of lactation. On each day, blood was collected at 10-min intervals for 2 h before one member of a pair of sows was infused (i.v.) with saline (n = 5) and the other with the neuroexcitatory amino acid N-methyl-DL-aspartic acid (NMA; 10 mg/kg b.w.) to measure readily releasable GnRH as estimated by LH release. Two hours later, each sow that had received saline was given GnRH (100 micrograms, i.v.) for estimation of releasable pools of pituitary LH and FSH. Responses to NMA and GnRH were estimated by area under the curve (ng.ml-1.min) of serum LH profiles for 1 h (NMA group) or 2 h (GnRH group) after treatment. Basal LH secretion in primiparous sows decreased from Day 1 to Day 7 and then returned to Day 1 values by Day 21. Similarly, in multiparous sows, basal LH was lower on Day 7 than on Day 21. Basal FSH secretion increased linearly as lactation progressed in primiparous and multiparous sows.(ABSTRACT TRUNCATED AT 250 WORDS)

Animals↗

Effects of gonadal steroids on tonic luteinizing hormone (LH) release and luteinizing hormone-releasing hormone-induced LH release from bovine pituitary cells cultured in vitro.

The objective of this study was to investigate the direct effect of estradiol-17 beta (E2) and progesterone (P4) on LH release by pituitary cells of midluteal phase cows in vitro. Pituitaries were collected at the slaughterhouse; cells were dissociated with collagenase and hyaluronidase and maintained in a static culture system in Medium 199 (M199). Various concentrations of E2 (0.1-100 nM) and P4 (0.1 and 10 nM) were used to stimulate the cells for 2, 4, 6, 15, 24, 48, or 72 h. In addition, the synergistic action of E2 and P4 was investigated by exposure of the cells to a combination of the two hormones. At the end of each incubation, the cells were challenged with LHRH (1 nM) for 2 h. The medium was collected for LH analysis at the end of each incubation period and after the LHRH challenge; furthermore, intracellular LH content was quantified at the end of each experiment. The results indicate a positive action of E2 on basal release of LH beginning after 15 h of exposure (p < 0.01). LHRH-induced LH release was modulated by E2 in a time-dependent manner with an effect at first inhibitory, then stimulatory, and finally inhibitory again (p < 0.04). P4 alone did not affect LH release, but it negatively influenced LHRH-induced LH release. P4 also exerted a positive action on intracellular LH after 6 h of incubation. A substantial inhibitory effect (p < 0.001) on both tonic LH release and LHRH-induced LH release was observed in cells exposed for 16 h to P4 after a priming with E2 for 4 h.(ABSTRACT TRUNCATED AT 250 WORDS)

Animals↗

Regulation of luteinizing hormone-releasing hormone (LHRH) secretion by melatonin in the ewe. I. Simultaneous delayed increase in LHRH and luteinizing hormone pulsatile secretion.

This study was performed to test the hypothesis that a short-day-like melatonin treatment would induce simultaneous stimulations of LHRH and LH secretion. Simultaneous serial samples of jugular and portal blood to assay LH and LHRH, respectively, were carried out at three different periods according to the expected changes in LH secretion. Twenty-six ovariectomized ewes bearing a subcutaneous implant of estradiol and exposed to a long-day photoperiod (16L:8D) received s.c. implants of melatonin on Day 0. LHRH and LH pulsatile secretion was studied in the first group during inhibition of LH secretion by long days (on Day -1, n = 6). A second group (n = 9) was sampled on Day 39. Moreover, seven animals from this group were sampled on Day 46 to focus on the beginning of LH increase. The third group (n = 6) was sampled during maximal LH secretion (Day 74, n = 6). A fourth group (n = 5) composed of noncannulated animals was sampled for LH secretion only as a control for the effects of surgical procedures. On each occasion, blood was collected every 10 min for 6 h. Between Day -1 and Day 39, LHRH pulse frequency was low and did not differ. In the same animals, this value became 2.5-fold higher between Day 39 and Day 46. Finally, on Day 74, LHRH pulse frequency was about 6-fold higher than on Day -1.(ABSTRACT TRUNCATED AT 250 WORDS)

Animals↗

Regulation of luteinizing hormone-releasing hormone (LHRH) secretion by melatonin in the ewe. II. Changes in N-methyl-D,L-aspartic acid-induced LHRH release during the stimulation of luteinizing hormone secretion by melatonin.

Injection of N-methyl-D,L-aspartic acid (NMDA), a glutamatergic agonist, cause an acute release of LH. The amount of LH released is modulated by photoperiod in sheep; it is larger in animals in which reproductive activity has been inhibited by long days than in those photostimulated by short days. The purpose of the present experiment was to determine whether this effect of photoperiod on LH response to NMDA is mediated by melatonin and whether it reflects a change in the amount of LHRH released in response to NMDA. Twenty-one ovariectomized ewes, bearing an s.c. implant of estradiol and kept in long days (16L:8D), received s.c. implants of melatonin on Day 0. The effect of a single injection of NMDA (5 mg/kg i.v.) on LHRH and simultaneous LH secretion was studied at three different periods of the stimulation of LH secretion by the melatonin treatment: first, during inhibition of LH secretion by long days (on Day -1, n = 6); secondly, during the onset of the LH increase (on Day 39, n = 9); and thirdly, during maximum LH secretion (on Day 74, n = 6). The stimulation of LHRH secretion by administration of NMDA was characterized by a large and long increase in LHRH concentration on Day -1 and Day 39. There was no significant difference between these two periods in terms of duration and amplitude of the LHRH release. On Day 74, the stimulation was much shorter than on Day -1 and Day 39 and was followed by a phase of decreased pulsatility of LHRH secretion.(ABSTRACT TRUNCATED AT 250 WORDS)

Animals↗

Luteinizing hormone/chorionic gonadotropin bioactivity in the common marmoset (Callithrix jacchus) is due to a chorionic gonadotropin molecule with a structure intermediate between human chorionic gonadotropin and human luteinizing hormone.

Chorionic gonadotropin (CG), a pregnancy-specific heterodimeric hormone found in primates, is responsible for CL rescue with pregnancy maintenance. Of the primates, the human and baboon gene sequences are the only structures so far determined. In order to study the structure and function of CG in other primates, we have isolated and sequenced the coding regions for the two subunits of marmoset CG (mCG) by the reverse transcription/polymerase chain reaction method. Study of multiple clones confirmed a high degree of homology with the human sequences (88% and 80% for the alpha and beta nucleotide sequences, respectively). Marmoset CG alpha has an extra four amino acids compared to hCG alpha, whereas the mCG beta sequence has a 3-bp deletion that maintains the reading frame and C-terminal amino acid sequence. Most of the differences between hCG beta and mCG beta peptides occur in the C-terminal region, which includes the loss of two of the O-linked glycosylation consensus sequences and the presence of an N-linked glycosylation consensus sequence. When mCG alpha and beta were co-expressed in CHO cells, assembly of biologically active hormone was confirmed by induced steroid secretion by MA10 cells. Partially purified mCG beta was used to raise anti-mCG antibodies. To date, an antibody has been obtained that is capable of detecting recombinant mCG beta, recombinant mCG dimer, and mCG dimer secreted by cultured marmoset trophoblast. Marmoset CG alpha and beta were also detectable at the transcriptional level in cultured trophoblast by in situ hybridization. This suggests that the LH/CG bioactivity reported from marmoset placentae and embryos is due to a molecule with structural features common to hLH (glycosylation pattern) and hCG (CG beta C-terminal structure).

Amino Acid Sequence↗