Prolactin Secretion In Rats Research Articles

Effects of chronic bromocriptine treatment of an estrone-induced, prolactin-secreting rat pituitary adenoma.

Bromocriptine (BROM), a dopamine (DA) agonist, is commonly and successfully used for long-term treatment of human prolactinomas. We have studied the effects of chronic BROM administration to female 344 Fisher/Lis rats bearing an estrone-induced, prolactin (PRL)-secreting pituitary tumor recently characterized as a model for human prolactinoma. The animals were injected twice daily with BROM (2.5 mg/kg) or with diluent. After 1 month of treatment, the animals were sacrificed, and plasma collected and stored at -20 degrees C for PRL radioimmunoassay. The pituitary tumors were removed and tumoral mammotrophs dispersed enzymatically for studies of DA receptor binding and PRL release in vitro. BROM treatment significantly reduced tumor weight, cell size, rough endoplasmic reticulum, Golgi complexes and plasma PRL levels. [3H]-spiroperidol binding to tumoral mammotrophs was also evaluated. BROM induced a significant decrease in the number of DA binding sites without any changes in affinity. These results indicate that chronic BROM treatment of an animal model of prolactinoma induces tumor involution, reduction of PRL release and probably synthesis, and down regulation of dopaminergic binding sites.

Presence of High Affinity Dopamine Receptors in Estrone-Induced, Prolactin-Secreting Rat Pituitary Adenomas: A Model for Human Prolactinomas

Adenomatous cells obtained from a pituitary tumor induced in Fisher 344/Lis rats by the subcutaneous implantation of estrone (E1) were found to secrete large amounts of prolactin (PRL). The secretion of PRL was stimulated by thyrotropin-releasing hormone (TRH) and low concentrations of dopamine (DA), while micromolar concentrations of DA were inhibitory. High affinity binding sites for 3H-spiroperidol (3H-SPIR) were found to be present on the cells and to conform to the criteria of dopaminergic receptors. An adenylate cyclase (AC) present in the cells could be activated by a guanyl nucleotide and was inhibited by DA in the presence of guanosine 5'-triphosphate (GTP). Fractionation of the adenomatous cells by Percoll gradients identified two groups of cells capable of secreting PRL and bearing 3H-SPIR binding sites. These data indicate that this rat pituitary adenoma may be a model for human prolactinomas that might be utilized for the study of the mechanism of action of dopaminergic drugs.

Novel 8 α-ergolines with inhibitory and stimulatory effects on prolactin secretion in rats

Four derivatives of the ergot dopamine (DA) agonist lisuride (LIS), namely 6-n-propyl-lisuride (6-n-propyl-LIS), transdihydrolisuride (TDHL), 6-n-propyl-transdihydrolisuride (6-n-propyl-TDHL) and 2-bromolisuride (2-Br-LIS) were investigated in female rats with regard to their influence on hyperprolactinaemia induced by pretreatment with reserpine (2 mg/kg i.p., 24 h) at various intervals following their subcutaneous or oral administration (0.05 mg/kg). Two hours after administration, LIS, 6-n-propyl-LIS, and 6-n-propyl-TDHL caused a statistically significant inhibition of reserpine-induced hyperprolactinaemia of about the same extent. Eight hours after administration 6-n-propyl-LIS and 6-n-propyl-TDHL were as active as after 2 h in inhibiting prolactin (PRL) secretion whereas LIS was almost ineffective in this respect. TDHL caused a statistically significant inhibition of PRL secretion at 2 and 8 h after oral administration; this effect was less pronounced after s.c. administration. In contrast to the aforementioned derivatives 2-Br-LIS further increased the reserpine-induced hyperprolactinaemia. In normal male rats pretreatment with 2-Br-LIS (0.025–6.25 mg/kg s.c., 2 h) dose-dependently stimulated PRL secretion. The present data support the assumption of the longlasting DA agonistic action of 6-n-propyl-LIS and 6-n-propyl-TDHL and of the antidopaminergic properties of 2-Br-LIS recently derived from behavioural studies.

Effects of a luteinizing hormone-releasing hormone analog on the growth of estrogen-induced prolactin-secreting rat pituitary tumors and its influence on their dopamine receptors.

Effects of a luteinizing hormone-releasing hormone analog on the growth of estrogen-induced prolactin-secreting rat pituitary tumors and its influence on their dopamine receptors.

Dopamine agonistic potency of two novel prolactin release-inhibiting ergolines

Two novel 8α-amino ergolines (CH 29-717 and CU 32-085) have been shown to inhibit secretion of prolactin in rats in vivo. However, when tested for dopaminomimetic potency on pituitary cell culture preparations in vitro, CH 29-717 inhibited prolactin release with an IC50 = 4 × 10−9 M. In the same model the 1-methyl-derivative CU 32-085 was only weakly active even at high concentrations. When both compounds were incubated together, CU 32-085 displayed an nhibitory effect on the prolactin release restriction caused by CH 29-717. The activities were not time-dependent since similar results were obtained with incubation for 3 or 24 h. These results support the hypothesis that the 1-methyl ergoline derivative CU 32-085 is a partial agonist and becomes fully active in vivo only after metabolic conversion.

Effect of 8-hydroxy-2-(di-n-propylamino) tetralin on rat prolactin secretion.

8-OH- DPAT (8-hydroxy-2-[di-n-propylamino] tetralin) is a novel aminotetralin derivative which has been proposed to be a serotonin (5-HT) agonist devoid of dopamine agonist effects. We now report that the administration of 8-OH- DPAT , like known 5-HT agonists, produced a rapid elevation of serum prolactin concentrations in male rats. The prolactin response to 8-OH- DPAT , like that induced by other 5-HT agonists, was greatly potentiated in animals pretreated with the tryptophan hydroxylase inhibitor, para-chlorophenylalanine. However, the 8-OH- DPAT -induced elevation of serum prolactin concentrations in untreated rats was not dose-dependent and was modest in magnitude compared to that produced by known 5-HT agonists. In contrast to the stimulatory effects of 8-OH- DPAT on prolactin secretion in vivo 8-OH- DPAT suppressed the secretion of prolactin from anterior pituitary tissue in vitro, and this effect was blocked by haloperidol. The results of the present study are supportive of the view that 8-OH- DPAT has dopamine agonist, as well as 5-HT agonist, properties.

Some 6-substituted 1-[(5R,8S,10R)-8-ergolinyl]-3,3-diethylureas

6-Substituted 1-[(5R,8S,10R)-8-ergolinyl]-3,3-diethylureas II-VII were prepared from hydrazides of (5R,8S,10R)-6-alkyl-8-ergolinecarboxylic acids (VIIIa, VIIIb) via the corresponding azides IXa and IXb and isocyanates Xa and Xb, and/or from 1-[(5R,8S,10R)-6-methyl-8-ergolinyl]-3,3-diethylurea (I) via XI and XII. Two of the by-products arising in the latter route have been identified (XIII, XIV). Some of the ureas prepared, especially II and III, had strong inhibitory effects in vivo on the secretion of prolactin in rats.

Depending on the stimulus, central serotoninergic activation by fenfluramine blocks or does not alter growth hormone secretion in man.

The role of serotonin (5-HT) in the hypothalamic regulation of human growth hormone (GH) was reassessed through the use of fenfluramine, which selectively releases 5-HT from presynaptic terminals. Oral administration of L-dopa plus propranolol induced a potent and sustained GH release in the subjects tested (26 +/- 6 ng/ml). The administration of fenfluramine (20 mg i.v. as bolus plus 20 mg/30 min i.v.) completely suppressed the L-dopa-induced GH secretion (2 +/- 09.5 ng/ml). On the other hand, when arginine (30 g/30 min i.v.) was used as a GH stimulant of medium intensity (12.7 +/- 2.8 ng/ml), fenfluramine at the same dose was not able to alter the pattern of pituitary secretion (11.5 +/- 4.2 ng/ml). Fenfluramine alone induced a slight nonsignificant decrease in GH values with a parallel and significant increase in prolactin (PRL) secretion in accordance with the proposed serotoninergic activity of the drug. Rat PRL secretion by pituitaries incubated in vitro was inhibited by dopamine. Fenfluramine added to the system did not counteract the dopaminergic reduction of PRL release, making unlikely the possibility of an antagonism at the dopaminergic receptor as mechanism of action of fenfluramine on PRL secretion. In conclusion, depending on the stimulus under study, serotoninergic activation by fenfluramine either inhibits or does not alter GH secretion in man. No proof of a serotoninergic stimulatory component on GH regulation has been detected in this study. Fenfluramine is a valuable tool in neuroendocrinological studies, dealing with serotoninergic mechanisms.

Thyroliberin stimulates rapid hydrolysis of phosphatidylinositol 4,5-bisphosphate by a phosphodiesterase in rat mammotropic pituitary cells. Evidence for an early Ca2+-independent action.

Thyrotropin-releasing hormone (TRH; thyroliberin) stimulated rapid hydrolysis of phosphatidylinositol 4,5-bisphosphate [PtdIns(4,5)P2] by a phosphodiesterase (phospholipase C) in GH3 cells, a prolactin-secreting rat pituitary tumour cell line. TRH caused a rapid decrease in the level of PtdIns(4,5)P2 to 60% of control and stimulated a marked transient increase in inositol 1,4,5-trisphosphate, the unique product of phosphodiesteratic hydrolysis of PtdIns(4,5)P2, to a peak of 410% of control at 15 s. TRH also caused decreases in phosphatidylinositol 4-monophosphate (PtdIns4P) and phosphatidylinositol (PtdIns) to 65% and 93% of control at 15 s respectively. Inositol 1,4-bisphosphate was increased to a peak of 450% at 30 s; inositol 1-monophosphate and inositol were not elevated until 30 s and 1 min respectively after TRH addition. To study whether PtdIns(4,5)P2 hydrolysis may be caused by an elevation in cytosolic Ca2+ concentration, the changes induced by TRH in the levels of inositol sugars were compared with the effects of membrane depolarization by high extracellular [K+]. The elevation in cytosolic [Ca2+] induced by K+ depolarization did not change the level of inositol 1,4,5-trisphosphate. These data suggest that phosphodiesteratic hydrolysis of PtdIns(4,5)P2 may be the initial event in TRH stimulation of inositol lipid metabolism in GH3 cells and that PtdIns(4,5)P2 hydrolysis is not stimulated by an elevation in cytosolic Ca2+ concentration. The decreases in PtdIns4P and PtdIns may be due to enhanced conversion of PtdIns into PtdIns4P into PtdIns(4,5)P2 or to their direct hydrolysis by phosphomonoesterases and/or phosphodiesterases. These results are consistent with the hypothesis that TRH-stimulated PtdIns(4,5)P2 breakdown causes Ca2+ mobilization leading to prolactin secretion.

Biphasic effect of 5-methoxy-N,N-dimethyltryptamine on rat prolactin secretion

5-Methoxy-N,N-dimethyltryptamine (5-MeODMT), a potent serotonin (5-HT) receptor agonist, exerts a biphasic effect on rat prolactin (PRL) secretion. 5-MeODMT (2.5–10 mg/kg) produces a marked, dose-related but short-lasting (less than 30 min) rise in serum PRL levels. At intervals longer than 30 min, 5-MeODMT (1–15 mg/kg) inhibits the stimulation of PRL secretion by another 5-HT agonist, 5-methoxytryptamine (5-MeOT, 10 mg/kg), by α-methylparatyrosine (50 mg/kg) or by haloperidol (0.15 mg/kg). 5-MeODMT did not significantly alter the PRL-releasing effect of γ-butyrolactone (500 mg/kg) or a higher dose of haloperidol (1 mg/kg). The biphasic effect of 5-MeODMT on rat PRL secretion is shared by the centrally-acting 5-HT agonist quipazine, but not by 5-MeOT, an indole derivative excluded by the blood-brain barrier. The initial stimulation of PRL secretion by 5-MeODMT is probably due to its ability to activate postsynaptic 5-HT receptors. The subsequent inhibitory effect of 5-MeODMT appears to be due to increased functional activity of tuberoinfundibular dopamine neurons. The possible mechanisms underlying the inhibitory effect of 5-MeODMT on PRL release are discussed.

Influence of ergot derivatives on prolactin secretion in rats. Mechanisms of action and clinical implications

Ergot derivatives are the most effective compounds in the treatment of migraine attacks. It has been proposed that these compounds exert this effect by direct action on skull arteries or arteriovenous anastomoses, or by interfering with peripheral or central serotonin receptors. It is also possible that these compounds influence monoaminergic neurotransmission and, thereby, endorphins modulating the threshold for pain or sensory perception. In testing these hypotheses, changes in receptor sensitivity or function have to be considered since we have observed simultaneous tolerance and supersensitivity development in the same animal treated chronically with lisuride. We therefore propose that ergot derivatives can reduce the well documented higher sensitivity of migraine sufferers to various stimuli. In humans, prolactin-levels can be used for determining bioavailability of some ergot derivatives and for studying whether dopaminergic mechanism plays a role in their antimigraine effect.

Effects of pirenperone and ketanserin on rat prolactin secretion in vivo and in vitro

Pirenperone, which is chemically related to ketanserin, has been reported to be a selective serotonin 2 (5-HT 2) antagonist and a specific d-LSD antagonist. We now report that pirenperone markedly stimulates prolactin (PRL) secretion in vivo at low doses and blocks the dopamine (DA)-induced inhibition of PRL release from rat pituitary glands in vitro, suggesting it acts as an antagonist at DA 2 receptors in the anterior pituitary gland. Ketanserin, also a purported selective 5-HT 2 receptor blocker, has no effect on rat PRL secretion in vivo or in vitro, but at high doses, it inhibits the increase in serum PRL levels produced by the two 5-HT agonists, quipazine and 5-methoxy-N, N-dimethyl-tryptamine (5-MeODMT). It had a weak ability to antagonize the PRL-releasing effect of the 5-HT precursor, 5-hydroxytryptophan. These results suggest that serotonergic stimulation of rat PRL secretion by quipazine and 5-MeODMT may be partially mediated by 5-HT 2 receptors. The inability of ketanserin to effectively block the effect of 5-HTP suggests its mechanism of stimulating PRL secretion is more complicated than that of the direct acting agonists.

Effects of a series of substituted benzamides on rat prolactin secretion and 3H-spiperone binding to bovine anterior pituitary membranes

The potency of seven substituted benzamine drugs (AHR-5531B, AHR-5645B, AHR-6092, AHR-8764, bromopride, sultopride and tiapride) to stimulate rat prolactin (PRL) secretion in vivo was found to be three orders of magnitude greater than that of non-benzamide neuroleptic drugs relative to their respective abilities to inhibit 3H-spiperone binding to bovine anterior pituitary membranes. Nevertheless, the IC 50 values for the inhibition of 3H-spiperone binding by the seven substituted benzamide drugs was significantly correlated with their high potency to stimulate rat PRL secretion in vivo . Further, the slope of the regression line for these substituted benzamides paralleled that of a series of butyrophenone, phenothiazine, morphanthridine and dibenzodiazepine neuroleptic drugs. Two benzamide (sulpiride and metoclopramide) and three non-benzamide neuroleptic drugs gave intermediate results. This data suggests that blockade of different subgroups of dopamine receptors in the anterior pituitary gland labeled by 3H-spiperone may be responsible for the in vivo stimulation of PRL secretion by the benzamide and non-benzamide neuroleptioc drugs.

Characterization of bombesin receptors in a rat pituitary cell line.

Bombesin is a tetradecapeptide which stimulates prolactin secretion in rats and man and in cultures of GH4C1 cells, a clonal strain of rat pituitary tumor cells. We have utilized [125I-Tyr4]bombesin to identify and characterize specific high affinity receptors in GH4C1 cells. Scatchard analysis of equilibrium binding data at 4 degrees C indicated the presence of a single class of non-interacting binding sites for bombesin (RT = 3600 +/- 500 sites/cell). The value for the equilibrium dissociation constant (Kd = 1.2 +/- 0.4 nM) agreed closely with the ED50 (0.5 nM) for bombesin stimulation of prolactin release. [125I-Tyr4]Bombesin binding at steady state at 37 degrees C was inhibited by increasing concentrations of unlabeled bombesin in a dose-dependent manner, with an ID50 = 1.4 +/- 0.2 nM. However, binding of [125I-Tyr4] bombesin was not inhibited by 100 nM thyrotropin-releasing hormone, vasoactive intestinal peptide, epidermal growth factor, or somatostatin. Therefore, [125I-Tyr4]bombesin binds to a receptor distinct from the receptors for other peptides which regulate hormone secretion by GH4C1 cells. The analog specificity for high affinity binding showed that the receptors for bombesin recognize the COOH-terminal octapeptide sequence in the molecule. Among five pituitary cell strains tested, two which contained saturable binding sites for [125I-Tyr4]bombesin (GH4C1 and GH3) had previously been shown to respond to bombesin with increased hormone secretion, whereas three which lacked receptors (GC, F4C1, and AtT20/D16v) were unresponsive. Therefore, the [125I-Tyr4]bombesin binding sites appear to be necessary for the biological actions of bombesin. Examination of the processing and metabolism of receptor-bound peptide demonstrated that at 4 degrees C [125I-Tyr4]bombesin binds to receptors on the surface of GH4C1 cells. At 37 degrees C, receptor-bound peptide is rapidly internalized and subsequently degraded in lysosomes. In summary, we have characterized for the first time specific, high affinity pituitary bombesin receptors which are necessary for the biological action of bombesin.

(5R,8S,10R)-6-Alkyl-8-ergolinecarboxylic acids and some of their derivatives

(5R,8S,10R)-6-Methyl-8-methoxycarbonylergoline (VIIb) was converted via Ib and IIb into its 6-analogues IIIb-VIb, which, in turn, were converted into acids IIIa-VIa and hydrazides IIIc-VIc. Some of the esters prepared, mainly IVb, had inhibitory effects on the secretion of prolactin in rats.

Inhibition of prolactin secretion delays extinction of circadian LH surges in ovariectomized rats bearing estradiol implants.

Ovariectomized rats bearing Silastic capsules containing estradiol exhibit a daily afternoon surge of luteinizing hormone (LH) which decreases with time until it is undetectable by Day 10 after implantation of estradiol. Increases in basal prolactin levels as well as afternoon surges are also observed. To determine if increased prolactin secretion contributed to the extinction of the circadian LH surges, we examined the patterns of LH and prolactin secretion in rats in which prolactin was suppressed by bromocriptine treatment. In vehicle-treated control rats, the magnitude of the LH surges decreased with time. Large LH surges were observed on Days 2 and 4. A significant decrease in the surge occurred on Day 6, and it disappeared by Day 10. Animals treated with bromocriptine also exhibited large LH surges on Days 2 and 4, and in addition, secreted a greater amount of LH than the control group on Days 6, 8, and 10. In ovariectomized rats bearing estradiol implants, large afternoon surges in prolactin were observed and by Day 6, basal prolactin levels were also elevated. Bromocriptine treatment completely suppressed prolactin secretion through Day 6, but a small afternoon rise was observed on Days 8 and 10. These findings suggest that elevated prolactin secretion may be one factor contributing to the extinction of circadian LH surges in the estrogen-treated rat.

156 Establishment of estrogen responsive clonal cell lines in tissue culture from an estrogen dependent, prolactin secreting rat pituitary tumor

156 Establishment of estrogen responsive clonal cell lines in tissue culture from an estrogen dependent, prolactin secreting rat pituitary tumor

Non-opiate β-endorphin fragments and dopamine—V γ-type endorphins and prolactin secretion in rats

The effects on prolactin secretion of three peptide-derivatives of β-endorphin which show neuroleptic-like activities in rats were studied. Intravenous administration of γ-endorphin (β-endorphin (βE) 1–17) enhanced plasma prolactin levels. γ-Endorphin did not affect the prolactin secretion by hemipituitary glands incubated in vitro and by cultured pituitary cells, but it reversed, in a dose-dependent way, the dopamine-induced inhibition of prolactin release. This effect of γ-endorphin could be prevented by co-incubation with the opiateantagonist naloxone. In vivo studies with non-opiate-like fragments of γ-endorphin, des-Tyr 1-γ-endorphin (DTγE, βE 2–17) and des-enkephalin-γ-endorphin (DEγE, βE 6–17), showed that these peptides suppressed the plasma prolactin levels, when prolactin release was slightly stimulated. However, DEγE dose-dependently enhanced plasma prolactin levels, when prolactin release was low. Subchronic treatment with DEγE for 4 days made prolactin release more sensitive to the inhibitory action of small doses of apomorphine. Neither DTγE nor DEγE affected prolactin release by hemipituitary glands and cultured pituitary cells. They did also not affect the dopamine-induced inhibition of prolactin release in vitro. It is suggested that γ-endorphin increases prolactin secretion by interfering with dopaminergic systems in the pituitary and that DTγE and DEγE have a modulatory action on prolactin secretion.

Androgen metabolism and mechanism of action in prolactin secreting rat pituitary cells in culture

The effects of different androgens on prolactin (PRL) synthesis were studied in clonal strains (GH 3, GH 4) of rat pituitary tumour cells. PRL synthesis was measured as the amount of hormone which accumulated in the culture medium during 24 h (μg/mg cell protein/24 h). The GH 3 cells are known to metabolize testosterone rapidly and extensively (9̃0%). 5α-Dihydrotestosterone (DHT) (55%) and 5α-androstane-3β,17β-diol (3β-diol) (31%) constituted the major metabolites, while no conversion of testosterone to oestrogens was observed (Aakvaag and Haug J. steroid Biochem. 11, 1979, 1341–1346). Testosterone, DHT, 3β-diol, 5α-androstane-3α,17β-diol (3α-diol) and 4-androstene-3,17-dione (Δ4-A) all stimulated PRL synthesis in a dose-dependent manner. The stimulatory effects were observed at about 10 −6 M and the maximum effects were found at about 5 × 10 −7 M. 3β-Diol was invariably the most potent stimulator and caused a 2–3 fold increase in PRL synthesis. DHT and testosterone were almost equally potent, but they were on a molar basis only half as effective as 3β-diol in stimulating PRL synthesis. 3α-Diol was almost as potent as the 3β-epimer, while Δ4-A caused only a very slight increase in PRL synthesis. None of the androgens influenced cell growth measured as cell protein content per culture dish. The synthetic 5α-reductase inhibitor 4-androsten-3one,17β-carboxylic acid (androstene-17β-C OH) inhibited the conversion of testosterone to its 5α-reduced metabolites (DHT, 3β-diol and 3α-diol) without any effect on PRL synthesis or cell growth. After 40 h incubation more than 90% of the added radioactivity was recovered as intact [ 3H]-testosterone in the presence of androstene-17β-COOH (4 × 10 −5 M) as compared to only 35% in the control cultures. The finding that testosterone was equally potent in stimulating PRL synthesis in the absence or presence of androstene-17β-COOH suggested that the effect of testosterone on PRL synthesis was independent of its 5α-reduction. The stimulating effect of testosterone and DHT on PRL synthesis was equal to that of the synthetic androgen methyltrienolone (R 1881). This study shows that testosterone and its 5α-reduced metabolites DHT, 3β-diol and 3α-diol stimulated PRL synthesis dose-dependently in the GH 3 cells, and the effect of testosterone was not dependent on its 5α-reduction.

Estrogen induced growth of human breast cancer cells (MCF-7) in athymic nude mice is enhanced by secretions from a transplantable pituitary tumor

MCF-7, a human breast carcinoma cell line, was maintained s.c. in female athymic nude mice for a period of 5–6 weeks. Administration of estrogen (s.c. pellet of 17β-estradiol and estrone in drinking water, 0.5 mg/l) to these mice resulted in sustained ( P < 0.001) growth of MCF-7 tumors. Grafting of a prolactin and growth hormone secreting rat pituitary tumor to the estrogen-treated mice resulted in an increased ( P < 0.05) rate of MCF-7 tumor growth. MCF-7 did not grow in athymic nude mice grafted with rat pituitary tumor alone or in mice without hormone treatment (controls). Thus, secretions of pituitary hormones alone are not capable of promoting in vivo growth of MCF-7 although such secretions significantly enhance estrogen-induced growth of this cell line. A synergism between pituitary hormones and estrogen for in vivo growth of a human breast carcinoma has been demonstrated in this study.