LHRH-induced Luteinizing Hormone Release Research Articles

Stress and the timing of breeding: Glucocorticoid-luteinizing hormones relationships in an arctic seabird

In birds, stressful environmental conditions delay the timing of breeding but the underlying mechanisms are poorly understood. The stress hormone corticosterone appears to be a good candidate for mediating the decision to breed and when to start egg-laying, via a possible inhibition of luteinizing hormone (LH) and sex-steroids production. We used luteinizing hormone releasing hormone (LHRH) challenge in pre-laying male and female Black-legged kittiwakes ( Rissa tridactyla) to test whether LH and testosterone secretion were depressed by elevated corticosterone levels. Females bearing high baseline corticosterone levels showed reduced baseline LH levels and a low ability to release LH, following LHRH challenge. Further, females bearing low baseline LH levels and elevated baseline corticosterone levels were more likely to skip breeding. However, non-breeding females were physiologically primed for breeding, since they mounted high LHRH-induced LH release. Egg-laying date was advanced in good body condition females but was unaffected by hormones secretion. In males, corticosterone levels had no effect on LH and/or testosterone secretion and did not affect their decision to breed. Interestingly, males with high LHRH-induced testosterone release bred early. Our study highlights clear sex-differences in the HPG sensitivity to stress hormones in pre-laying kittiwakes. Because females have to store body reserves and to build up the clutch, they would be more sensitive to stress than males. Moreover, intrasexual competition could force male kittiwakes to acquire reproductive readiness earlier in the season than females and to better resist environmental perturbations. We suggest that high testosterone releasing ability would mediate behavioural adjustments such as courtship feeding, which would stimulate early egg-laying in females.

Study of Enteral versus Parenteral Application of the Gonadotropin Releasing Hormone Agonist Gonadorelin[6-D-Phe] (D-Phe6-LHRH) on LH Secretion in Goettinger Miniature Pigs

With respect to the assessment of residue situation and as a part of preclinical trials to determine the biological activities of potential gonadotropin releasing hormone (GnRH) residues in porcine organisms the GnRH agonist Gonadorelin[6-D-Phe] (D-Phe(6)-LHRH) was administered either enterally or intramuscularly (i.m.) to female Goettinger miniature pigs in order to evaluate the GnRH-induced luteinizing hormone (LH) surge. Gilts received an (i) enteral application of 10 mg D-Phe(6)-LHRH via a probang (enteral group, n=7), (ii) i.m. injection of 0.1 mg D-Phe(6)-LHRH (parenteral group, n=5), or (iii) saline injection (control group, n=4). The GnRH and saline applications were repeated every second day with up to seven repetitions. Blood samples were collected via previously fitted jugular catheters immediately before injections, over an 8 h period in 1 h intervals beginning 2 h after injections, and at 24, 26, 28 and 30 h after applications. Enteral application of D-Phe(6)-LHRH induced an LH surge in 23 of 30 treatments. All gilts in the parenteral group exhibited LH release after each D-Phe(6)-LHRH application (P<0.05), whereas no LH surges were observed after saline injection in the control group. A significant (P<0.05) LH rise to mean maximum LH concentrations of 3.25 +/- 0.43 and 3.05 +/- 0.26 ng/ml occurred in both the enteral and parenteral groups, but there was no difference in the time interval after GnRH (2.6 +/- 0.3 vs. 2.3 +/- 0.3 h) and the mean duration of LH peak (6.5 +/- 0.4 and 6.8 +/- 0.3 h) between the treatment groups. In conclusion, (i) enteral application of 10 mg D-Phe(6)-LHRH induced LH release in a physiological range from the pituitary of female minipigs, and (ii) neither an accumulative effect nor a cumulative LH response were found after repeated GnRH application. Furthermore, (iii) in regard to consumer protection and gonadotropin secretion, D-Phe(6)-LHRH residues can be excluded from having long-term effects.

Evidence Suggesting that the Potentiating Action of Neuropeptide Y on Luteinizing Hormone (LH)-Releasing Hormone-Induced LH Release Remains Unaltered in Aged Female Rats

Several lines of evidence suggest that one of the mechanisms by which the hypothalamic neuropeptide Y plays an obligatory role in the preovulatory luteinizing hormone (LH) discharge in young rats is to potentiate the action of LH-releasing hormone (LHRH) on LH release at the level of the pituitary. This study examined whether an alteration in the potentiating action of neuropeptide Y on LHRH-induced LH release may contribute to the attenuation or absence of LH surges during female reproductive ageing. Young regularly cycling (2-3-month-old) and old constant oestrous (19-20-month-old) rats ovariectomized for 7 days were primed with oestradiol-17beta-filled Silastic capsules. Two days later, rats received s.c. progesterone at 09.00 h and then were injected i.p. with either saline or pentobarbital at 13.30 hours. Pentobarbital-treated rats received i.v. pulses of neuropeptide Y, LHRH, a combination of neuropeptide Y and LHRH, or saline, every 30 min from 14.00 to 18.00 h via a jugular cannula. Hourly blood samples were collected between 11.00 and 21.00 h. In old rats, the progesterone-induced LH surge was significantly attenuated and delayed as compared to that of young rats. Pentobarbital injection completely blocked the LH surge. Neuropeptide Y pulses alone had no significant effect on LH release. In contrast, LHRH pulses increased LH release in both age groups, although the response was significantly reduced in older rats. While combined pulses of neuropeptide Y and LHRH significantly increased LH release in both young and old rats as compared to that of LHRH alone, the potentiating action of neuropeptide Y on LHRH-induced LH release remained unchanged between the two age groups. These results, together with our recent demonstration of altered hypothalamic neuropeptide Y neuronal activity in middle-aged pro-oestrous rats, suggest that a deficit in neuropeptide Y secretion and action in the hypothalamus, rather than a decrease in pituitary responsiveness to neuropeptide Y, may partially be responsible for the absence of LH surges in old rats.

Role of protein kinase C in facilitation of luteinizing hormone (LH)-releasing hormone-induced LH surges by neuropeptide Y.

In female rats, neuropeptide Y (NPY) facilitates LHRH-induced LH surges without affecting basal LH release. The signal transduction mechanisms mediating this facilitation are unknown. Here, the involvement of PKC in this process was investigated. Anterior pituitaries (APs) were removed from rats at 1400 h proestrus and perifused in vitro with M199 for 5 h. After an equilibration and baseline period, tissue received hourly 5-minute pulses of the PKC inhibitor GF109203X (GFX), 2.5 microM, followed 15 min later by a 5-minute pulse of LHRH (10(-8) M), NPY (10(-6) M), or phorbol 12-myristate 13-acetate (PMA, 50 nM), or some combination. This regimen was repeated hourly for 3 h. As shown previously, NPY had no effect on basal LH release but greatly facilitated LHRH-induced LH release. Treatment with PMA also facilitated LHRH-induced LH release, to approximately the same degree as NPY. Inhibition of PKC activity with GFX completely prevented NPY's and PMA's facilitation of LH release but did not inhibit LH release stimulated by LHRH alone. Because previous work suggested involvement of both NPY and PKC in alterations of LHRH receptor affinity or number, the in vivo effects of NPY on LHRH binding characteristics were also investigated. Although NPY treatment reliably enhanced LHRH-induced LH and FSH surges in proestrous rats, this action was not accompanied by any detectable change in the affinity or concentration of LHRH receptors in anterior pituitary cell membranes. In summary, we have found that NPY's actions are blocked by PKC inhibition, mimicked by PKC stimulation, and not associated with any overt alterations in LHRH receptor affinity or number. We conclude that PKC activation is required for NPY's facilitation of LHRH-induced LH surges, and that this mechanism likely involves PKC targets other than those which may alter LHRH receptor number or affinity.

Intraventricular injection of melatonin inhibits naloxone-induced, but not NMDA- or LHRH-induced LH release in ovariectomized estrogen-primed rats.

The present study was aimed to examine the possible functional relationship between melatonin and hypothalamic transmitters, endogenous opioids and excitatory amino acids in controlling gonadotropin secretion in ovariectomized estrogen-primed rats. An intravenous injection of naloxone (mu opioid receptor antagonist), N-methyl-D-aspartate (NMDA; NMDA receptor agonist) or luteinizing hormone-releasing hormone (LHRH) significantly elevated serum luteinizing hormone (LH) concentrations within 10 min. An intraventricular treatment with melatonin, which did not affect the basal LH concentration by itself, significantly suppressed the effect of naloxone. However, the same melatonin treatment did not inhibit the NMDA-induced or LHRH-induced LH secretion. These results support the hypothesis that melatonin has a suprapituitary site of action to inhibit LHRH release, and suggest that the site of its action may be located downstream to that of naloxone action and upstream to that of NMDA in the hypothalamic LHRH neuronal pathway.

Melatonin inhibits naloxone-induced luteinizing hormone release in ovariectomized estrogen-primed rats.

The present study aimed to examine the effect of melatonin on naloxone-induced luteinizing hormone (LH) secretion in ovariectomized estrogen-primed rats. A single intracerebroventricular (i.c.v.) injection of naloxone (mu opioid receptor blocker, 15 micrograms) or an intravenous (i.v.) injection of LH-releasing hormone (LHRH, 50 ng/kg) elicited a transient and significant increase in the serum LH concentration within 10 min. While an i.c.v. injection of 100 ng melatonin by itself did not change the basal LH release, it almost completely inhibited the naloxone-induced LH release. Melatonin (10 ng) also significantly reduced the effect of naloxone. However, an i.c.v. injection of 100 ng melatonin did not affect the LHRH-induced LH release. In separate experiments, the effect of melatonin on naloxone-induced pulsatile LH secretion was studied in estrogen-treated rats. A continuous i.v. infusion of naloxone (20 mg/kg/h) induced LH pulses in rats treated i.c.v. with saline. An i.c.v. administration of 100 ng melatonin, which by itself did not affect basal LH secretion, significantly reduced the frequency, but not the amplitude, of LH pulses induced by the naloxone infusion. These results show that melatonin has a suprapituitary site of action to inhibit naloxone-induced LH release, and suggest that melatonin has an effect in inhibiting the activity of the hypothalamic LHRH pulse generator, either directly or indirectly, in female rats.

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)

Effects of LHRH and ANG II on prolactin stimulation are mediated by hypophysial AT1 receptor subtype.

We have used the nonpeptide angiotensin II (ANG II) receptor antagonists losartan (receptor subtype AT1) and PD-123319 (AT2) to determine the participation of ANG II receptor subtypes in luteinizing hormone-releasing hormone (LHRH)-induced prolactin release in a perifusion study using intact pituitaries in vitro. LHRH (1.85 x 10(-7) M) released prolactin consistently, whereas losartan (10(-5) M) abolished prolactin response without modifying basal prolactin or luteinizing hormone (LH) and follicle-stimulating hormone (FSH) release. PD-123319 (10(-5) M) had no effect on basal or LHRH-induced prolactin, LH, or FSH release. We also determined that the effect of ANG II on prolactin release was mediated by the same receptor subtype. In adenohypophysial cells dispersed in vitro ANG II (10(-8) M) released prolactin. Losartan (10(-7) and 10(-6) M), but not PD-123319, inhibited this effect. We conclude that in intact hypophyses of 15-day-old female rats the effect of LHRH on prolactin release is readily demonstrated. LHRH-induced prolactin release appears to be mediated by ANG II acting in a paracrine manner on AT1 receptors located on lactotrophs.

A potential code of luteinizing hormone-releasing hormone-induced calcium ion responses in the regulation of luteinizing hormone secretion among individual gonadotropes

Luteinizing hormone-releasing hormone (LHRH) induces two Ca2+ responses in single gonadotropes: a Ca2+ spike/plateau or oscillation. Similar receptor-mediated Ca2+ signals have been reported in many cell types but their functional significance is obscure. Accordingly, we have determined the concentration-response properties of LHRH-induced luteinizing hormone (LH) release at the single cell level. We demonstrate a critical single cell LHRH threshold for LH release. Each gonadotrope had a particular LHRH threshold value and a range of different single cell thresholds was distributed in the gonadotrope population. The physiological significance of the threshold was demonstrated by a striking reduction (delta ED50 = 153 nM) of the LHRH threshold immediately before the preovulatory surge of LH release. The metestrous phenotype of secretion resembled a quantal process in contrast with the graded process of the proestrous phenotype. That is, the quantity of hormone secreted per metestrous gonadotrope was independent of LHRH concentration and more all-or-none than graded. The LHRH threshold and the quantal secretion process of metestrous gonadotropes was further studied by measuring cytosolic Ca2+ using fura-2 and digital imaging microscopy. We provide evidence suggesting that the Ca2+ spike/plateau and oscillation are the respective responses to subthreshold and suprathreshold concentrations of LHRH. It is proposed therefore that the Ca2+ oscillation and spike/plateau response form a binary intracellular signaling code that functions as an on-off switch. It is further proposed that this potential code unraveled here for the regulation of hormone secretion may also regulate other gonadotrope functions. Thus, while the Ca2+ spike/plateau response is strongly associated with LH release, it may be associated with reduced levels of LH-beta mRNA, and reduced numbers of LHRH receptors. Conversely, while the Ca2+ oscillation appears to be unrelated to LH release, it may be associated with increased levels of LH-beta mRNA, and increased numbers of LHRH receptors. This model may explain in molecular terms the long-standing observation that an invariant, albeit pulsatile, pattern of LHRH release is sufficient to support the preovulatory surge of LH release.

Neuropeptide-Y Enhances Luteinizing Hormone (LH)-Releasing Hormone-Induced LH Release and Elevations in Cytosolic Ca2+in Rat Anterior Pituitary Cells: Evidence for Involvement of Extracellular Ca2+Influx through Voltage-Sensitive Channels*

The present studies were designed to investigate the mechanism by which neuropeptide-Y (NPY) augments the effect of LHRH to stimulate the release of LH from cultured rat anterior pituitary cells. Anterior pituitary cells from ovariectomized rats were enzymatically dispersed, cultured for 3 days, and then exposed to various secretagogues during 3-h incubations. As reported by this laboratory previously, NPY alone (100 nM) did not affect LH release, but significantly enhanced the LH response to 1 nM LHRH. This facilitatory action of NPY was mimicked by the dihydropyridine Ca2+ channel agonist Bay K 8644 (1 microM), and the enhancement of LHRH-induced LH release by either NPY or Bay K 8644 was prevented by the dihydropyridine antagonist nitrendipine (1 microM). Nitrendipine alone reduced the response to LHRH by approximately 25%, but did not affect basal LH release. In contrast, NPY failed to amplify the release of PRL in response to TRH, another Ca2(+)-mobilizing hormone. To test whether NPY also enhances the increase in cytosolic Ca2+ induced by LHRH, anterior pituitary cells were acutely dispersed into single cell suspensions, loaded with the fluorescent Ca2+ probe Indo-1 AM, and analyzed with a UV laser in an EPICS-753 flow cytometer at a rate of 500 cells/sec for 200 sec. The ratio of intracellular fluorescence resulting from Ca2+ bound to the Indo-1 to the fluorescence from Indo-1 alone (Indo-1 ratio), which is an index of the concentration of free cytosolic Ca2+, was determined for each cell. Approximately 7% of anterior pituitary cells responded to LHRH (1 or 10 nM) with significant increases in Indo-1 ratios, indicative of an increase in the concentration of free cytosolic Ca2+. EGTA (2.5 mM) reduced the basal Indo-1 ratios and attenuated, but did not abolish, the initial increase in response to LHRH, consistent with the initial extracellular Ca2+ influx-independent phase of the response to LHRH. NPY alone (100 nM) did not affect the Indo-1 ratios in anterior pituitary cells, but pretreatment with the peptide for 10 min before the scans significantly augmented the Indo-1 ratio response to 10 nM LHRH. This effect of NPY was also blocked by EGTA. Taken together, these biochemical and pharmacological studies suggest that NPY enhances the release of LH stimulated by LHRH by increasing extracellular Ca2+ entry, possibly by selectively affecting that component of the response involving dihydropyridine-sensitive L-type voltage-sensitive Ca2+ channels during the initial stages of the cellular response to LHRH.

Biological activity of luteinizing hormone in the peripartum cow: least activity at parturition with an increase throughout the postpartum interval.

The biological (B) and immunological (I) activities of luteinizing hormone (LH) were determined during four peripartum periods in the cow. The sampling periods were PRE-5 (approximately 5 days prior to parturition), POST-3 (Day 2 or 3 postpartum), POST-12 (Day 11 or 12 postpartum) and LUTEAL (Day 11 or 12 following the first observed postpartum estrus). Blood samples were collected at 10-min intervals for either 8 h (PRE-5, POST-3, and POST-12) or 6 h (LUTEAL). Four hours prior to the end of each sampling period, 100 micrograms of luteinizing hormone-releasing hormone (LHRH) was injected i.v. Plasma concentrations of LH were analyzed by a validated bioassay (rat interstitial cell testosterone) and a radioimmunoassay. Area under the LHRH-induced LH release curve was calculated by the trapezoidal rule, and endogenous LH was determined by averaging the pre-LHRH treatment values. Paired t-analyses were used to compare LH release between periods. The periods of lowest (p less than 0.05) bioactive LH (BLH) were PRE-5 and POST-3. During POST-12, BLH was maximal. Endogenous immunoactive LH (ILH) was maximal during the POST-12 period, with PRE-5, POST-3, and LUTEAL being the least (p less than 0.05). The B:I ratio was lowest (p less than 0.05) around the time of parturition (PRE-5 and POST-3), then increased (p less than 0.05) by Day 12 postpartum. The ratio then increased (p less than 0.05) again by the LUTEAL period. These data suggest that the biological quality of LH changes throughout the puerperium. We infer from these data that the low biological activity of LH (subquality LH) may contribute to the postpartum infertility in the cow.

Enhanced luteinizing hormone release by luteinizing hormone-releasing hormone in incubating female turkeys (Meleagris gallopavo).

To determine what role pituitary responsiveness plays in the suppression of gonadotropin level during incubation in the turkey, the ability of the pituitary to release luteinizing hormone (LH) in response to luteinizing hormone-releasing hormone (LHRH) was compared in incubating, laying, and photorefractory birds. In all three groups, the i.m. injection of LHRH (4 micrograms/kg) increased serum LH levels; however, the LH response was markedly enhanced in the incubating turkeys as compared with the laying (6.6-fold increase over preinjection levels vs. 1.9-fold; p less than 0.05) or the photorefractory birds (9.7-fold vs. 3.1-fold; p less than 0.05). The LHRH-induced LH release was also determined in turkeys as they shifted from the laying to the incubating phase of the reproductive cycle. This response increased (p less than 0.05) in magnitude as the birds started to incubate. The high prolactin level of incubating turkeys does not have a depressing effect on LHRH-stimulated LH release; thus, impaired LH response to LHRH is not a mechanism involved in the diminished gonadotropin secretion of incubating turkeys.

Hypothalamic and serum factors influence on prolactin and luteinizing hormone release by the pituitary gland of the young turkey ( Meleagris gallopavo)

Intravenous administration of 1.0 or 3.0 eq hypothalamic extract (HE) to 8-week-old male and 7-week-old female turkeys, respectively, induced an increase in circulating prolactin (PRL) levels but had no effect on circulating luteinizing hormone (LH) levels. The incubation of dissociated anterior pituitary cells from 13-week-old female turkeys with HE induced a dose-related increase in PRL release; however, only the highest dose of HE induced an increase in LH release. Coincubation of a hypothalamic fragment with anterior pituitary cells from 9-week-old females induced a release of both PRL and LH. Dissociated pituitary cells from 11-week-old females initially incubated for 3 hr in medium containing charcoal-treated (stripped) turkey serum yielded a larger release of PRL and LH in the presence of HE than did cells initially incubated with turkey serum or no serum. Luteinizing hormone-releasing hormone (LHRH) induction of LH release was greatest from cells initially incubated with stripped serum for 3 or 24 hr. The LHRH-induced LH release was completely blocked in cells initially incubated for 24 hr with turkey serum. The initial incubation of cells for 24 hr with stripped serum yielded a larger release of PRL and LH in response to HE than did cells initially incubated with serum. The hypothalamus of the young turkey contains substantial PRL-releasing activity as well as LH-releasing activity. The ability of the releasing factors to stimulate pituitary hormone release is influenced by factors present in the blood of the young turkey. This is especially evident in the LHRH-induced LH release where serum factors inhibited the release.

Monosodium glutamate and pituitary gland luteinizing hormone (LH) release in response to LH-releasing hormone: an in vitro study.

We studied whether an increase in the basal LH release rate and/or the anterior pituitary gland (APG) LH response to LHRH is involved in maintaining normal or near-normal serum LH levels in monosodium L-glutamate (MSG)-treated rats which have small APGs for their body weight. Female rats were injected with MSG (4 mg/g BW) or saline on days 1, 3, 5, 7, and 9 after birth (day of birth = 0). At 8-9 weeks of age, saline-treated and MSG-treated rats were ovariectomized, and 7 days later, they were decapitated. Trunk blood was collected from 18 controls and 19 MSG-treated rats, and serum LH concentrations were measured by RIA. APGs were bisected and each hemi-APG was placed in culture medium for a 30-min preincubation period, followed by two 30-min incubation periods during which water or 10 or 30 ng LHRH were added to the medium. Despite the fact that the APGs of the MSG-treated rats were half the size of those of the saline-treated rats, the serum LH levels in the 2 groups were not different. Basal LH release rates (the response to water) and LHRH-induced LH release per mg APG were increased in MSG-treated rats. Calculation of the basal LH release rates and LHRH-induced LH release on the basis of the entire weights of the APGs showed no differences between the MSG-treated rats and the controls. In 6 additional control and 6 additional MSG-treated rats, the APG LH concentration was measured and was not different between the 2 groups. The results suggest that increases in both the basal LH release rate per mg APG and the amount of LH released per mg APG in response to LHRH are of importance in the maintenance of normal or near-normal serum LH concentrations in MSG-treated rats with small APGs.

Correlation of luteinizing hormone surges with estrogen nuclear and progestin cytosol receptors in the hypothalamus and pituitary gland. II. Temporal estradiol effects.

In these studies we examined the temporal parameters of estradiol (E2) priming required before progesterone (P4) amplifies the surge release of luteinizing hormone (LH) in short-term ovariectomized (OVX) rats. Thereafter, we correlated the time of appearance and concentrations of estrogen nuclear (E2Rn) and progestin cytosol (PRc) receptors in brain regions known to be involved in the surge release of LHRH. Steroid receptor concentrations were also measured in the pituitary gland. 1 week after OVX (day 0), Silastic capsules containing E2 (150 micrograms/ml in oil s.c.) were placed at 09.00 h. The serum E2 concentrations produced by these capsules peaked (41 +/- 2.1 pg/ml) at 10.00 h on day 0 and declined thereafter to values which ranged between 19 and 10 pg/ml on days 1 through 4. Some of these E2-treated rats also received 2 P4-containing Silastic capsules (50 mg/ml s.c. in oil) at 09.00 h on either days 0, 1, 2, 3 or 4. Serum P4 concentrations produced by such capsules were 9.3 +/- 0.5 ng/ml. Blood was collected sequentially at 09.00 h and at hourly intervals from 12.00-18.00 h on the day that the animals received the P4 capsules and 10.00 and 15.00 h samples also were taken from each group the next day. LH surges occurred in these E2-treated rats in the afternoons of days 2-4, but not on days 0 and 1. P4 treatment on day 1 elicited an LH surge and on days 2-4 it amplified plasma LH surge concentrations and advanced by 1 h the time of release of this gonadotropin.(ABSTRACT TRUNCATED AT 250 WORDS)

Inhibition of luteinizing hormone (LH)-releasing hormone-induced secretion of LH in rat anterior pituitary cell culture by testosterone without conversion to 5 alpha-dihydrotestosterone.

The role of 5 alpha-reduction of testosterone in the inhibition of LH secretion was investigated in rat anterior pituitary cell cultures. Pituitary cells were preincubated with testosterone or dihydrotestosterone (17 beta-hydroxy-5 alpha-androstan-3-one) for 17 h and then with LHRH for an additional 4 h. Dihydrotestosterone was 6-fold more potent than testosterone in the inhibition of LHRH-induced LH release. Basal LH secretion was not affected by either androgen. The inhibition curves of testosterone and dihydrotestosterone were not shifted by the presence of the 5 alpha-reductase inhibitors 17 beta-N,N-diethylcarbamoyl-4-methyl-4-aza-5 alpha-androstan-3-one (4-MA) and 17 beta-N,N-diisopropylcarbamoyl-4-aza-androstan-3-one (DIPA). Neither 4-MA nor DIPA alone had an effect on either basal or LHRH-induced LH release. When pituitary cells were incubated with [3H]testosterone for 17 h, the radioactivities were found to be unmetabolized testosterone (66.9 +/- 2.4%), dihydrotestosterone (13.3 +/- 0.5%), androstenedione (15.9 +/- 1.3%), 5 alpha-androstane-3,17-dione (2.8 +/- 0.3%), and 3 alpha (beta), 17 beta-androstanediol (less than 1%). In the presence of 4-MA or DIPA, 5 alpha-reduction of testosterone was completely inhibited; androstenedione was the only metabolite. Androstenedione was only 12% as potent as testosterone in the inhibition of LHRH stimulation of LH release, and conversion of [3H]androstenedione to testosterone and dihydrotestosterone did occur in these cells. When [3H]dihydrotestosterone was incubated with pituitary cells, the radioactivities were dihydrotestosterone (64.4 +/- 0%), 5 alpha-androstanedione (19.3 +/- 1%), 3 alpha (beta), 17 beta-androstanediol (7.7 +/- 1.7%), and unknown polar metabolites. 4-MA and DIPA had no effect on the metabolism of dihydrotestosterone. These results indicate that both testosterone and dihydrotestosterone inhibit LHRH-induced LH release and that this activity of testosterone does not depend on its 5 alpha-reduction.

Modulation of gonadotropin secretion by Sertoli cell inhibin, LHRH, and sex steroids.

Sertoli cell culture media (SCM) from 10-, 20-, 30-, 35-, and 40-day-old male Wistar rats were assayed to determine the inhibin activity in anterior pituitary cells in culture. In agreement with previous data, SCM did not affect the luteinizing hormone (LH) spontaneous release at all ages studied, whereas it inhibited specifically follicle-stimulating hormone (FSH) spontaneous release by 40% for the 40-day-old rats. Younger animals (10-, 20-, and 30-day-old) showed a 60% inhibition of the FSH basal release. The inhibin activity was also different at all stages studied, the IC50 being markedly displaced to the right as the age increased, leading to a fivefold difference between 10- and 30- to 40-day-old rats. The same pattern was observed when the LH and FSH responses to 0.3 nM LH-releasing hormone (LHRH) were studied. SCM from 35-day-old rats did not alter total LH, whereas total FSH was markedly reduced, thus suggesting a reduced FSH synthesis in the presence of inhibin. SCM exerts an additive inhibitory effect with dihydrotestosterone on the LH response to LHRH, whereas it reverses the stimulatory effect of the androgen on spontaneous and LHRH-induced FSH release. Moreover, SCM reversed the stimulatory effect of 17 beta-estradiol on both spontaneous and LHRH-induced LH and FSH release, whereas the stimulatory effect of progesterone on FSH release was 50-80% inhibited. The present data show that inhibin activity of Sertoli cell origin can exert marked interactions with sex steroids in the control of gonadotropin secretion. These data also demonstrate that the inhibin component is an important factor in sexual maturation of the rat and that high FSH levels of 10-day-old rats could suggest a modulation by a nonandrogenic factor of gonadotropin secretion in developing rats.

The effect of castration, estradiol and LHRH on LH secretion of lambs fed different levels of dietary energy.

To examine the effect of diet on luteinizing hormone (LH) secretion, basal and luteinizing hormone releasing hormone (LHRH)-induced LH release was compared in intact or castrated-estradiol-17 beta implanted Finn-Dorset lambs. Ten to 12 wk old ram (n = 20) and ewe lambs (n = 20) were maintained under a 8L:16D photoperiod and fed for high (HG, 163 to 168 g/d) or low (LG, 76 to 103 g/d) rates of gain. Eight to 10 wk later, baseline LH concentrations were determined in blood samples collected at 20 min intervals for 7 h. The following day, lambs were given an iv injection of 5 micrograms of estradiol-17 beta followed within 4 h by LHRH (.5 or 2.5 micrograms). Baseline concentrations of LH for HG ewes were threefold greater than for LG ewes (4.2 vs 1.4 ng/ml), respectively. Time to peak response was inversely related to dietary energy level (P less than .025). Basal LH levels were similar across diets in rams. Total LH release following LHRH was dose-dependent (P less than .005). Effects of gonadal feedback were tested in a second group (n = 24) of castrated lambs. Changes in LH secretion were not different between diets within 3 to 4 wk after castration. A subcutaneous silastic implant (22 mm) of estradiol-17 beta inhibited (P less than .01) LH concentrations across diets in both ewes and rams. No differences in estradiol feedback on LH secretion (at the dose of steroid tested) were detected between HG and LG lambs. Within 8 d, however, basal LH concentrations were 60% lower (P less than .01) in HG vs LG ewes. Furthermore, peak LHRH-induced LH release was greater (P less than .025) in LG vs HG lambs of both sexes. Estradiol inhibited basal LH secretion in ewes and rams but facilitated LH release in lambs with a reduced rate of gain.

Site of action for β-endorphin-induced changes in plasma luteinizing hormone and prolactin in the ovariectomized rat

A number of sites have been hypothesized as loci at which opioid substances act to alter the secretion of luteinizing hormone (LH) and prolactin (PRL) (1–8). The aim of the present study was to determine the site(s) at which the opioid peptide β-endorphin (β-END) acts to influence plasma LH and PRL levels in the ovariectomized (OVX) rat. β-END, administered into the third ventricle of conscious OVX rats fitted with jugular catheters, significantly decreased plasma LH in doses ⩾ 50 ng and increased PRL levels at all doses administered (10, 50, 100 and 250 ng) in a dose dependent fashion. To identify possible central nervous system sites of action, 250 ng β-END was unilaterally infused into various brain sites. Plasma LH was significantly decreased and plasma PRL significantly increased by infusions into the ventromedial hypothalamic area, the anterior hypothalamic area, and the preoptic-septal area. There was no significant effect of β-END infusions into the lateral hypothalamic area, amygdala, midbrain central gray, or caudate nucleus. When hemipituitaries of OVX rats were incubated in vitro with β-END (10 −7M to 10 −5M), there was no suppression of basal or LHRH-induced LH release, nor was there any alteration of basal PRL release. It is concluded that β-END acts at a medial hypothalamic and/or preoptic-septal site and not the pituitary, to alter secretion of LH and PRL.

Effects of ovarian steroids on in vitro release of LHRH from mediobasal hypothalamus.

The phasic luteinizing hormone (LH) release observed in ovariectomized (OVX), estrogen-implanted rats was further amplified and advanced when progesterone (P) was given 4 h prior to the gonadotropin surge. In contrast, an inhibitory effect of P on the daily LH surge was observed when P was administered 16-36 h prior to LH peak. In order to determine whether this biphasic action of P is primarily exerted on the release of luteinizing hormone releasing hormone (LHRH), on the pituitary response to LHRH, or on both, mediobasal hypothalamic slices or pituitary fragments of adult OVX rats or of OVX rats pretreated with estrogen alone or in combination with P were tested in a perifusion system. Mediobasal hypothalamic slices were perifused in buffered (pH 7.2) oxygenated Locke's medium containing bacitracin (2 X 10(-5) M). In the absence of estrogen pretreatment, high (56 mM) concentrations of K+ were barely effective in releasing LHRH. Subcutaneous implantation of 17 beta-estradiol for 5 days markedly increased the amplitude of the LHRH secretory response to K+ depolarization. Additional administration of P (25 mg/rat s.c.) 4 h before sacrifice further amplified the K+-induced LHRH release. In contrast, the K+-evoked LHRH secretion was significantly inhibited when P was given 16 or 36 h before. Estradiol thus appears to facilitate the LHRH secretory response to depolarizing stimuli, whereas P either enhances or blocks the induced LHRH release depending upon its time of administration. At the pituitary level, the sensitivity of LHRH-induced LH release was also increased after estrogen pretreatment.(ABSTRACT TRUNCATED AT 250 WORDS)