Increase In Luteinizing Hormone Release Research Articles

An automated system for cattle reproductive management under the IoT framework. Part II: Induction of luteinizing hormone release after gonadotropin releasing hormone analogue delivery with e-Synch

Technologies for automating animal management and monitoring tasks can improve efficiency and productivity of livestock production. We developed the e-Synch system for automated control and monitoring the estrous cycle of cattle through intravaginal hormone delivery and sensing. Thus, our objective was to evaluate luteinizing hormone (LH) concentrations after intravaginal instillation of the Gonadotropin-releasing hormone (GnRH) analogue Gonadorelin with the e-Synch system. This system consists of an intravaginal electronically controlled automated hormone delivery and sensing device integrated with an IoT platform. Lactating Holstein cows with their estrous cycle synchronized were used in two experiments (Exp). In Exp 1, at 48 h after induction of luteolysis, cows (n=5-6 per group) were randomized to receive 100 µg of Gonadorelin through intramuscular (i.m.) injection, 100 µg of Gonadorelin in a 2 mL solution delivered with e-Synch, and an empty e-Synch device. In Exp 2, at 48 h after induction of luteolysis cows (n=6-7 per group) were randomized to receive 100 µg of Gonadorelin i.m., or an intravaginal treatment with e-Synch consisting of 100 or 1,000 µg of Gonadorelin in 2 or 10 mL of solution containing 10% citric acid as absorption enhancer. Circulating concentrations of LH were analyzed with linear mixed models with or without repeated measurements. In Exp 1, cows in the i.m. Gonadorelin treatment had a surge of LH whereas cows in the other two treatments did not have a surge of LH for up to 8 h after treatment. In Exp 2, the 1,000 µg dose of Gonadorelin elicited more LH release than the 100 µg dose, regardless of solution quantity. The overall LH response as determined by area under the curve, mean, and maximum LH concentrations was similar between cows receiving 1,000 µg of Gonadorelin delivered with e-Synch and 100 μg of Gonadorelin i.m. Increasing volume of solution for delivering the same dose of Gonadorelin partially increased LH release only for the 100 µg dose. We conclude that the e-Synch system could be used to automatically release Gonadorelin in a dose and volume that induces a surge of LH of similar magnitude than after i.m. injection of 100 μg of Gonadorelin. Also, the dose of Gonadorelin delivered by e-Synch is more critical than the volume of solution used.

Central somatostatin-somatostatin receptor 2 signaling mediates lactational suppression of luteinizing hormone release via the inhibition of glutamatergic interneurons during late lactation in rats.

Reproductive function is suppressed during lactation owing to the suckling-induced suppression of the kisspeptin gene (Kiss1) expression in the arcuate nucleus (ARC) and subsequent suppression of luteinizing hormone (LH) release. Our previous study revealed that somatostatin (SST) neurons mediate suckling-induced suppression of LH release via SST receptor 2 (SSTR2) in ovariectomized lactating rats during early lactation. This study examined whether central SST-SSTR2 signaling mediates the inhibition of ARC Kiss1 expression and LH release in lactating rats during late lactation and whether the inhibition of glutamatergic neurons, stimulators of LH release, is involved in the suppression of LH release mediated by central SST-SSTR2 signaling in lactating rats. A central injection of the SSTR2 antagonist CYN154806 (CYN) significantly increased ARC Kiss1 expression in lactating rats on day 16 of lactation. Dual in situ hybridization revealed that few ARC Kiss1-positive cells co-expressed Sstr2, and some of the ARC Slc17a6 (a glutamatergic neuronal marker)-positive cells co-expressed Sstr2. Furthermore, almost all ARC Kiss1-positive cells co-expressed Grin1, a subunit of N-methyl-D-aspartate (NMDA) receptors. The numbers of Slc17a6/Sstr2 double-labeled and Slc17a6 single-labeled cells were significantly lower in lactating dams than in non-lactating rats whose pups had been removed after parturition. A central injection of an NMDA antagonist reversed the CYN-induced increase in LH release in lactating rats. Overall, these results suggest that central SST-SSTR2 signaling, at least partly, mediates the suppression of ARC Kiss1 expression and LH release by inhibiting ARC glutamatergic interneurons in lactating rats.

Optogenetic Activation of Arcuate Kisspeptin Neurons Generates a Luteinizing Hormone Surge-Like Secretion in an Estradiol-Dependent Manner.

Traditionally, the anteroventral periventricular (AVPV) nucleus has been the brain area associated with luteinizing hormone (LH) surge secretion in rodents. However, the role of the other population of hypothalamic kisspeptin neurons, in the arcuate nucleus (ARC), has been less well characterized with respect to surge generation. Previous experiments have demonstrated ARC kisspeptin knockdown reduced the amplitude of LH surges, indicating that they have a role in surge amplification. The present study used an optogenetic approach to selectively stimulate ARC kisspeptin neurons and examine the effect on LH surges in mice with different hormonal administrations. LH level was monitored from 13:00 to 21:00 h, at 30-minute intervals. Intact Kiss-Cre female mice showed increased LH secretion during the stimulation period in addition to displaying a spontaneous LH surge around the time of lights off. In ovariectomized Kiss-Cre mice, optogenetic stimulation was followed by a surge-like secretion of LH immediately after the stimulation period. Ovariectomized Kiss-Cre mice with a low dose of 17β-estradiol (OVX+E) replacement displayed a surge-like increase in LH release during period of optic stimulation. No LH response to the optic stimulation was observed in OVX+E mice on the day of estradiol benzoate (EB) treatment (day 1). However, after administration of progesterone (day 2), all OVX+E+EB+P mice exhibited an LH surge during optic stimulation. A spontaneous LH surge also occurred in these mice at the expected time. Taken together, these results help to affirm the fact that ARC kisspeptin may have a novel amplificatory role in LH surge production, which is dependent on the gonadal steroid milieu.

Rational design of triazololipopeptides analogs of kisspeptin inducing a long-lasting increase of gonadotropins.

New potent and selective KISS1R agonists were designed using a combination of rational chemical modifications of the endogenous neuropeptide kisspeptin 10 (KP10). Improved resistance to degradation and presumably reduced renal clearance were obtained by introducing a 1,4-disubstituted 1,2,3-triazole as a proteolysis-resistant amide mimic and a serum albumin-binding motif, respectively. These triazololipopeptides are highly potent full agonists of KISS1R and are >100 selective over the closely related NPFF1R. When injected in ewes with a quiescent reproductive system, the best compound of our series induced a much prolonged increase of luteinizing hormone release compared to KP10 and increased follicle-stimulating hormone plasma concentration. Hence, this KISS1R agonist is a new valuable pharmacological tool to explore the potential of KP system in reproduction control. Furthermore, it represents the first step to develop drugs treating reproductive system disorders due to a reduced activity of the hypothalamo-pituitary-gonadal axis such as delayed puberty, hypothalamic amenorrhea, and hypogonadotropic hypogonadism.

Involvement of the ganglion cholinergic receptors in gonadotropin-releasing hormone, catecholamines, and progesterone release in the rat ovary

To investigate whether cholinergic ganglionic stimulus modifies the release of gonadotropin-releasing hormone (GnRH), catecholamines, and progesterone at the ovarian level. Animal study. University animal laboratory. Six to eight virgin adult Holtzman rats. Superior mesenteric ganglion-ovarian nerve plexus-ovary system removed and placed in one cuvette with two compartments, with acetylcholine added to the ganglion in the experimental group. Measurement of ovarian liquid obtained from catecholamines by high-performance liquid chromatography; measurement of progesterone (P(4)), GnRH, and luteinizing hormone (LH) by radioimmunoassay; and measurement of gene expression of 3β-hydroxysteroid dehydrogenase (3β-HSD) and 20α-hydroxysteroid dehydrogenase (20α-HSD) by reverse-transcriptase polymerase chain reaction (RT-PCR). The study focused on the estrus and diestrus II (DII) stages. On the estrus days, the release of GnRH, NA, and 20α-HSD increased, while P(4) and 3β-HSD decreased. On the DII days, GnRH, P(4), and 3β-HSD increased, while 20α-HSD and NA decreased. The ovarian liquid with GnRH showed biologic activity, namely, an increase in LH release during the DII stage and a decrease during the estrus stage. Neural stimulus from the superior mesenteric ganglion influences the release of NA, adrenaline, and GnRH. We also have demonstrated that these neurotransmitters participate in the atretogenic processes of the ovary, thus providing evidence of the necessity of the sympathetic neural pathway.

The Control Mechanism of Gonadotropin-Releasing Hormone and Dopamine on Gonadotropin Release from Cultured Pituitary Cells of Rainbow Trout Oncorhynchus mykiss at Different Reproductive Stages

The mechanism by which gonadotropin-releasing hormone (GnRH) and dopamine (DA) control gonadotropin (GTH) release was studied in male and female rainbow trout using cultured pituitary cells obtained at different reproductive stages. The mechanisms of follicle-stimulating hormone (FSH) release by GnRH and DA could not be determined yet. However, basal and salmon-type GnRH (sGnRH)- or chicken-II-type GnRH (cGnRH-II)- induced luteinizing hormone (LH) release increased with gonadal maturation in both sexes. LH release activity was higher after sGnRH stimulation than cGnRH-II stimulation at maturing stages in both sexes. The GnRH antagonist ([Ac-3, 4-dehydro-Pro 1 , D-p-F-Phe 2 , D-Trp 3,6 ] GnRH) suppressed LH release by sGnRH stimulation in a dose-dependent manner, although the effect was weak in maturing fish. The role of DA as a GTH-release inhibitory factor differs during the reproductive cycle: the inhibition of sGnRH-stimulated LH release by DA was stronger in immature fish than in maturing, ovulating, or spermiated fish. DA did not completely inhibit sGnRH-stimulated LH release, and DA alone did not alter basal LH release. Relatively high doses (10 -6 or 10 -5 M) of domperidone (DOM, a DA D2 antagonist) increased LH release, which did not change with reproductive stage in either sex. The potency of DOM to enhance sGnRH-stimulated LH release was higher in maturing and ovulated fish than in immature fish. These data suggest that LH release from the pituitary gland is controlled by dual neuroendocrine mechanisms by GnRH and DA in rainbow trout, as has been reported in other teleosts. The mechanism of control of FSH release, however, remains unknown.

The inhibitory effect of anandamide on luteinizing hormone-releasing hormone secretion is reversed by estrogen.

Because Delta-9-tetrahydrocannabinol (THC) inhibited luteinizing hormone-releasing hormone (LHRH) in male rats, we hypothesized that the endocannabinoid, anandamide (AEA), would act similarly. AEA microinjected intracerebroventricularly (i.c.v.) decreased plasma luteinizing hormone (LH) at 30 min in comparison to values in controls (P < 0.001). The cannabinoid receptor 1 (CB1-r)-specific antagonist, [N-(piperidin-1-yl)-1-(2,4-dichlorophenyl)-5-(4-chlorophenyl)-4-methyl-1H-pyrazole-3-carboxamide] (AM251), produced a significant elevation in plasma LH (P < 0.01). AEA (10(-9) M) decreased LHRH release from medial basal hypothalami incubated in vitro. These results support the concept that endogenous AEA inhibits LHRH followed by decreased LH release in male rats. In ovariectomized (OVX) female rats, AEA i.c.v. also inhibited LH release, but in this case AM251 had an even greater inhibitory effect than AEA. In vitro, AEA had no effect on LHRH in OVX rats. It seems that endogenous AEA inhibits LHRH followed by decreased LH release in OVX rats but that AM251 has an inhibitory action in this case. In striking contrast, in OVX, estrogen-primed (OVX-E) rats, AEA i.c.v. instead of decreasing LH, increased its release. This effect was completely blocked by previous injection of AM251. When medial basal hypothalami of OVX-E rats were incubated, AEA increased LHRH release. The synthesized AEA was higher in OVX-E rats than in OVX and males, indicating that estrogen modifies endocannabinoid levels and effects. The results are interpreted to mean that sex steroids have profound effects to modify the response to AEA. It inhibits LHRH and consequently diminishes LH release in males and OVX females, but stimulates LHRH followed by increased LH release in OVX-E-primed rats.

Insulin-like growth factor I enhances gonadotropin-releasing hormone-stimulated luteinizing hormone release from bovine anterior pituitary cells

The role of insulin-like growth factor I (IGF-I) in the release of luteinizing hormone (LH) is unclear in ruminants. In the present study, the effects of IGF-I on the release of LH stimulated by gonadotropin-releasing hormone (GnRH) were examined in primary cultures of bovine anterior pituitary (AP) cells, and the interaction between estradiol-17β (E 2) and IGF-I was characterized. GnRH(100 nM)-stimulated LH release from the cultured cells was increased ( P<0.05) 12, 24 and 36 h after addition of IGF-I (250 ng/ml), with a maximum at 12 h (48.4 ng/ml media versus 35.4 ng/ml media in controls). IGF-I at concentrations of 25, 250 and 500 ng/ml increased the release by 18.7, 24.2 and 28.9%, respectively ( P<0.05), when compared with controls (37.2 ng/ml media). E 2 (10 nM), IGF-I (250 ng/ml) and combined treatment of E 2 plus IGF-I also induced significant increases in LH release ( P<0.05). The amounts of LH release after treatment with E 2 alone was 37.3% greater than with IGF-I alone (39.0 ng/ml media versus 28.4 ng/ml media) ( P<0.05). When E 2 and IGF-I were added together (45.6 ng/ml media), the release of LH was significantly greater than with either E 2 alone or IGF-I alone ( P<0.05). E 2 (10 nM) significantly ( P<0.05) increased the amount of GnRH bound to the cells by 51.6% when compared with controls, however, IGF-I (250 ng/ml) failed to increase GnRH binding. These results show that IGF-I enhances GnRH-stimulated LH release without changing the number of GnRH receptors in cattle, and IGF-I interacts with E 2 to increase the response to GnRH.

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.

Chapter 13 Non-metabolic and metabolic factors causing lactational anestrus: rat models uncovering the neuroendocrine mechanism underlying the suckling-induced changes in the mother

Follicular development and ovulation are strongly inhibited during lactation. Administration of a high dose of estrogen induces luteinizing hormone (LH) surges in ovariectomized lactating rats, suggesting that brain mechanisms regulating cyclic LH release remain intact in lactating mothers. On the other hand, tonic LH release is profoundly suppressed in lactating rats. This suggests that lactational anestrus is mainly due to suppression of the mechanism regulating pulsatile gonadotropin-releasing hormone secretion in the hypothalamus, which is responsible for follicular development and steroid production. Both metabolic and non-metabolic factors are involved in suppressing pulsatile LH secretion throughout lactation in rats. During the first half of lactation, pulsatile LH secretion is strongly suppressed, even if milk production is attenuated by pharmacological blockade of prolactin secretion in ovariectomized lactating rats. Pulsatile LH release quickly recovers by removing pups or blocking neuronal input by hypothalamic deafferentation during the period. These data suggest that the suckling stimulus itself is responsible for suppression of LH release during the first half of lactation. During the second half of lactation, negative energy balance, which is caused by the milk production, appears to play a dominant role in suppressing LH secretion. Blockade of milk production by inhibiting prolactin release causes a gradual increase in LH release even if the vigorous suckling stimulus by foster pups remains. In conclusion, the suckling stimulus itself predominantly suppresses LH pulses during the first half of lactation and metabolic factors take over the role of the suckling stimulus during the second half of lactation.

Mechanism of delayed ovulation in a vespertilionid bat, scotophilus heathi: role of gonadotropin, insulin, and insulin-like growth factor-1.

The aim of this study was to evaluate the effects of luteinizing hormone (LH), follicle-stimulating hormone (FSH), insulin, and insulin-like growth factor-1 (IGF-1) on ovarian androstenedione synthesis to understand the mechanism responsible for delayed ovulation in Scotophilus heathi. We found that LH stimulated a dose-dependent increase in androstenedione synthesis by the ovary in vitro. This study also showed a clear seasonal variation in the ability of the ovary to produce androstenedione in vitro in response to LH and FSH stimulation. In response to LH and FSH, maximum quantities of androstenedione were produced during recrudescence in November. The same doses of gonadotropins during the preovulatory period in February stimulated comparatively low androstenedione secretion by the ovary. On the basis of these data, we suggest that in S. heathi, ovarian responsiveness to LH and FSH peaks during recrudescence. This study also showed a seasonal variation in the effects of insulin and IGF-1 on ovarian androstenedione production in vitro. Peak ovarian responsiveness to insulin and IGF-1 was observed during quiescence in September. It is hypothesized that increased insulin/IGF-1 sensitivity during September may be responsible for increased responsiveness to LH. Increased LH release, if coincident with the period of enhanced ovarian responsiveness to LH, may result in the excessive androstenedione production responsible for delayed ovulation in S. heathi.

In vitro pituitary responsiveness to LHRH in young and old female rats. Influence of melatonin

The effect of aging and melatonin on in vitro pituitary responsiveness to luteinizing hormone-releasing hormone (LHRH) was studied. Young cyclic (3-months-old) control (cyclic-control, N=15), and melatonin (MEL) treated for 2 months (150 μg/100 g BW) (cyclic-MEL, N=15), old acyclic (23-months-old) control (acyclic-control, N=13), and MEL-treated (acyclic-MEL, N=18) rats were used. The hormones analyzed were luteinizing hormone (LH), follicle stimulating hormone (FSH) and prolactin (PRL). The results showed a different influence of the reproductive status as well as of melatonin on the basal secretion rate of both gonadotropins, i.e. LH and FSH. Only the basal FSH release was significantly reduced in cyclic-MEL and acyclic-controls compared to cyclic-controls. The hemipitutary FSH content raised to values similar to those observed for FSH secretion and only the cyclic-MEL group showed significantly higher FSH pituitary content than for release. LHRH addition to the incubation medium resulted in increased LH release for both cyclic and acyclic rats, but FSH release was only stimulated in acyclic rats. Melatonin treatment blunted this response in both cases. In addition, melatonin treatment inhibited prolactin release in acyclic-MEL group after LHRH stimulation but not the basal levels. Pituitary LH and prolactin contents, were significantly higher than the pituitary LH and prolactin levels released from all groups studied, and were not affected by reproductive senescence nor by exogenous melatonin. These data indicate that aging influences more the secretory than the biosynthetic processes. Melatonin influences is endocrine status-dependent, being inhibitory when pituitary hormones reach their higher values.

The effect of blockade of κ-opioid receptors in the medial preoptic area on the luteinizing hormone surge in the proestrous rat

The present study examined whether blockade of κ-opioid receptors in the medial preoptic area (MPOA) prior to the critical period on the afternoon of proestrus could prematurely evoke an ovulatory luteinizing hormone (LH) surge, and if so, whether norepinephrine (NE) is involved in mediating this response. In the first experiment, push–pull perfusion of the MPOA with nor-binaltorphimine (nor-BNI), a specific κ-opioid receptor antagonist, was done in rats between 10.30 and 13.50 h on proestrus. To determine whether any resulting ovulation was due to a nor-BNI-induced increase in LH release, rats were injected with pentobarbital at 13.55 h to block the afternoon LH surge. In 7 of 10 rats, nor-BNI in the MPOA produced a large increase in LH release beginning between 12.30 and 13.30 h, and 5 of 7 ovulated. During MPOA perfusion with cerebrospinal fluid in our normal colony between 14.00 and 17.00 h, surges of LH release began in the majority of rats between 15.30 and 16.30 h. Thus blockade of MPOA κ-opioid receptors advanced the LH surge by 3 h. The next experiment examined the effect of NE synthesis inhibition with bis(4-methyl-1-homopiperazinylthiocarbonyl) disulfide (FLA-63), or α-adrenergic receptor blockade with phenoxybenzamine (PBZ), on the nor-BNI-induced LH response. In 5 of 6 vehicle-treated rats, blockade of MPOA κ-opioid receptors elicited a large increase in LH release and all 5 ovulated. In contrast, only 3 of 8 rats pretreated with FLA-63 had a large increase in LH release and ovulated, and PBZ prevented the nor-BNI-induced LH increase and ovulation in 4 of 4 rats. PBZ also prevented the afternoon LH surge and ovulation in 4 of 4 rats in our normal colony. Finally, HPLC measurement of NE levels in MPOA push–pull perfusate indicated no increase in NE release during the nor-BNI-induced or normal afternoon LH surges. These results indicate that antagonism of κ-opioid receptors in the MPOA can prematurely evoke an ovulatory LH surge prior to the critical period on the afternoon of proestrus. Furthermore, the nor-BNI-induced as well as the normal afternoon LH surges are dependent on the proper functioning of central noradrenergic neurons, but do not involve increased NE release within the MPOA.

Neural levels of cholecystokinin peptide and mRNA during dietary stimulation of growth and LH secretion in growth-retarded lambs

Chronic feed restriction of prepubertal male lambs adversely affects reproductive development by inhibiting the pulsatile release of luteinizing hormone (LH). Because this effect can be reversed by ad libitum feeding, the associations between diet-induced increases in LH release and concurrent changes in body weight gain, serum glucose, CCK peptide, and CCK mRNA were examined. Neonatally castrated male lambs received a restricted ration to maintain their respective weaning weights beginning at 8 wk of age. At 23 wk of age, lambs were assigned randomly to receive additional feed equivalent to 0%, 50%, or 100% of their previous daily intake. Serum profiles of LH and glucose were determined 2 and 4 wk after onset of the increased intakes. At 27 wk of age, lambs were euthanized and both hypothalamic and cerebral cortical tissues were collected for analysis of CCK peptide and CCK mRNA. With additional intakes, body weight gain increased (P < 0.001) proportional to the graded increases in feed intake. Mean serum LH concentrations, LH peak frequencies, and serum glucose concentrations also increased (P < 0.05) progressively among the 0%, 50%, and 100% dietary intake groups. Neither CCK peptide nor CCK mRNA differed (P > 0.05) among dietary groups suggesting that endogenous CCK in the whole hypothalamus did not change with the feeding-induced increase in LH release. Concentrations of CCK peptide in cerebral cortex were greater (P < 0.05) than hypothalamic concentrations, but there were no differences between hypothalamus and cerebral cortex in the relative abundance of CCK mRNA. In summary, dietary stimulation of growth-retarded male lambs resulted in progressive increases in body weight gain, mean serum LH, serum glucose, and LH peak frequencies. Because hypothalamic levels of CCK peptide and CCK mRNA did not change during feeding-induced secretion of LH, endogenous CCK in the hypothalamus seems unlikely as a chronic mediator of nutrition-sensitive LH release.

Effects of Prostaglandins E2, F2.ALPHA., and D2 on the Release of Growth Hormone, Prolactin, and Luteinizing Hormone from Cultured Bovine Anterior Pituitary Cells.

The objectives of research were to characterize the effects of prostaglandins (PG) on the release of growth hormone (GH), prolactin (PRL), and luteinizing hormone (LH) in cultured bovine anterior pituitary cells. PGE2 significantly increased the media concentrations of GH by 141% (P<0.01) and 27% (P<0.01) at the concentrations of 10-7 and 10-8 M, respectively, compared with controls. The PGE2 significantly increased media concentrations of PRL compared with controls at the concentrations of 10-7 and 10-8 M by 120% (P<0.01) and 22% (P<0.05), respectively. PGE2 also significantly increased LH release at the concentration of 10-7 M by 22% (P<0.05), but not at a concentration of 10-8 M. PGF2α significantly increased the release of LH compared with controls at the concentrations of 10-8 and 10-9 M by 15% (P<0.05) and 19% (P<0.05), respectively, while PGF2α failed to stimulate the release of GH and PRL. In contrast to PGE2 and PGF2α, PGD2 had no effect on the release of GH, PRL, and LH. These results suggest that PGE2 is the most potent prostaglandin among the PGs for the release of GH, PRL, and LH by acting directly on bovine pituitary cells.

Lack of catecholamine involvement in the increased luteinizing hormone release due to blockade of κ-opioid receptors in the medial basal hypothalamus during midpregnancy in the rat

Blockade of κ-opioid receptors in the medial basal hypothalamus (MBH) with nor-binaltorphimine (nor-BNI) stimulates luteinizing hormone (LH) release during midpregnancy in the rat [48]. The objective of this study was to determine whether norepinephrine (NE) or dopamine (DA) mediates the LH response to blockade of MBH κ-opioid receptors on days 13–17 of pregnancy in the rat. Two experiments were conducted. In the first, push-pull perfusion in conjunction with HPLC was used to monitor in vivo NE release in the MBH occuring in response to (a) artificial CSF followed by CSF containing nor-BNI (40 μg/h), (b) desipramine (DMI, a NE reuptake blocker, 10 μM) in CSF followed by DMI, and (c) DMI followed by DMI + nor-BNI. Blood samples were taken at 12 min intervals concurrent with push-pull perfusate samples. Plasma LH levels were determined by RIA. Nor-BNI significantly increased LH release compared to CSF alone, but perfusate NE was undetectable in either perfusion period. However, perfusion with CSF containing 100 mM K + in these rats markedly increased perfusate NE levels, indicating noradrenergic nerve terminals were present at the perfusion sites in the MBH. Addition of DMI to the CSF significantly increased perfusate NE levels, but produced no changhe in LH release. Nor-BNI + DMI perfusion increased LH secretion similar to nor-BNI alone, but produced no additional increase in MBH perfusate NE levels compared to perfusion with DMI alone. In the second experiment, push-pull perfusion in the MBH with nor-BNI was done in rats pretreated either with the NE synthesis inhibitor FLA-63 (25 mg/kg, s.c.), the α-adrenergic receptor blocker phentolamine (5 mg/kg, i.v.), the DA receptor antagonist d-butaclamol (1 mg/kg, s.c.), or vehicle. None of these drug treatments affected the increased LH release occuring during MBH perfusion with nor-BNI. The present results demonstrate that neither NE nor DA mediates the increased LH release occuring in response to blockade of κ-opioid receptors in the MBH during midpregnancy in the rat.

Effect of naloxone and β-casomorphin on the hypothalamic-pituitary-luteinizing hormone axis in vitro

The effect of naloxone and β-casomorphin on luteinizing hormone (LH) release from pituitary cell aggregates, obtained by three-dimensional culture, with or without mediobasal hypothalamic fragments was studied in vitro. Short-term naloxone perifusion at a concentration of 10 −5M did not modify either basal or LHRH-stimulated LH release from the pituitary cell aggregates. In contrast, a 12-min naloxone perifusion at the same concentration caused an increase in LH release in the mediobasal hypothalamic-pituitary cell aggregate axis. This increase was rapid (12–16 min after time pulse), marked [up to 10 times (p<0.004) the initial base line], short (return to the base line secretion 32–40 min after the beginning of the time pulse) and dose-dependent, with a rise >1000% at a concentration of 10 −4M (p<0.006). The same effect was observed when a second pulse was applied 48 min after the first one. LH release induced by naloxone was antagonized 56 ± 2% (p<0.03) by β-casomorphin (an exogenous opiate) at a concentration of 10 −5M. β-casomorphin alone did not modify LH basal secretion, but inhibited 25.1 ± 2.4% (p p<0.008) LH release enhanced by LHRH. These results indicate that naloxone, an opiate antagonist, markedly increases LH release via a μ-type opioid receptor mechanism at the hypothalamic level only, during short-term exposure.

The effects of age on the postreceptor regulation of luteinizing hormone secretion by gonadotropin-releasing hormone

We studied the effects of age on the roles of phosphoinositide (PI) and protein kinase C (PKC) in luteinizing hormone (LH) release by gonadotropin-releasing hormone from mouse pituitaries. Pituitary cells from intact and 14-day ovariectomized (OVX) mice aged 4–8 months, 10–12 months and 14–18 months were cultured at a dilution of 3×10 5 cells/ml of M199-bovine serum albumin medium for 3 days prior to stimulation with either buserelin or phorbol ester (phorbol myristate acetate, PMA), while LH was assayed by radioimmunoassay using anti-rat LH antibody (NIDDK-5-10). In intact young mice, buserelin and PMA specifically induced time- and dose-dependent increases in LH release with specific mean ED 50 of 0.82×10 −11 M (buserelin) and of 1.6×10 −8 M (PMA) and a maximal LH release of 138±15 ng/10 6 cells after a 3 h stimulation period. Age did not affect the ED 50 of either agonist but significantly reduced their ability to release LH. This reduction was more pronounced for buserelin than for PMA and was evident as early as middle-age. OVX resulted in a significant increase in both basal and stimulated LH release, but did not affect the age-related reduced secretion rate of LH by either agonist. Buserelin stimulated the incorporation of [ 3H]inositol into [ 3H]inositol phosphates (IP) in a dose-dependent manner, which was unaffected by either age or OVX. We conclude that, with aging, there occurs a reduced LH release rate to both buserelin and PKC stimulations, uncoupled to changes in PI-IP cycle. Furthermore our results suggest that these cellular defects are first manifested at middle-age and are unaffected by the status of the gonad.

Ovine Brain Areas Sensitive to Naloxone-Induced Stimulation of Luteinizing Hormone Release

Previous work established that intravenous administration of the opioid receptor antagonist naloxone abruptly increased release of luteinizing hormone (LH) and decreased release of prolactin (PRL) in suckled anestrous ewes and also increased LH release in cyclic luteal ewes. The goal of the present research was to identify brain sites at which local unilateral infusions of naloxone would consistently duplicate the previously noted effects of intravenous naloxone. Intracerebral guide tubes were surgically implanted into the brain of 13 nonpregnant and 16 pregnant ewes at least 4 weeks prior to experimentation. Intracerebral infusion (20-40 microliters each through an inner cannula) was performed once daily during postpartum anestrus in suckled fall-lambing ewes and during recurring luteal states of the estrous cycle. Naloxone infusion (n = 142) usually consisted of 50 or 100 micrograms naloxone, although 5 ewes received 200 and 400 micrograms per infusion. Control infusions (n = 103) consisted of the vehicle for naloxone (i.e., 0.9% NaCl). Serum concentrations of LH and PRL were quantified at 10-min intervals from 90 min before to 100 min after infusion. Hormone data from individual ewes were grouped for least-squares analysis of variance based upon postmortem neuroanatomical identification of each infusion site. Unilateral intracerebral administration of naloxone consistently induced an increase in LH release within 20 min in the following two neuroanatomical groups:basal forebrain (n = 9 ewes) and chiasmatic area (n = 4 ewes).(ABSTRACT TRUNCATED AT 250 WORDS)

Changes in plasma levels of testosterone during male-male interactions in the song sparrow, Melospiza melodia: time course and specificity of response

Previous investigations have shown that malemale interactions over territory boundaries result in an increase in plasma levels of testosterone (T). In order to determine the time course of this increase following an agonistic challenge, male song sparrows, Zonotrichia (= Melospiza) melodia, were exposed to simulated territorial intrusions. Responding males were captured at intervals after onset of the intrusion for up to 60 min. Plasma levels of T were elevated by 10 min after onset of the challenge, and remained high in birds sampled 10–60 min after onset of the intrusion. In addition, plasma levels of luteinizing hormone (LH) increased and were significantly elevated after 10 min of stimulation. The results suggest that at least the prolonged elevation of T secretion precipitated by a male-male interaction was mediated via an increase in LH release. However, the initial surge of T levels in plasma may be LH independent. p Investigations of captive birds confirmed that circulating LH and T levels increased when male song sparrows were challenged by a conspecific male. Challenges involving a male house sparrow, Passer domesticus, had no effect suggesting that responses to male intrusions were species characteristic and not a result of general arousal. Further, auditory stimuli (vocalizations) alone, and visual stimuli (a devocalized male) alone were less effective in stimulating an increase in secretion of T than a combination of auditory and visual stimuli.