Modulatory effect of steroid hormones on GnRH-induced LH secretion by cultured rat pituitary cells

1992 ◽  
Vol 70 (7) ◽  
pp. 963-969 ◽  
Author(s):  
Gabriela T. Pérez ◽  
Marta E. Apfelbaum
Keyword(s):  
Steroid Hormones ◽  
Pituitary Cells ◽  
Short Term ◽  
Stimulatory Action ◽  
Rat Pituitary ◽  
Lh Release ◽  
Gonadotrophin Releasing Hormone ◽  
Rat Pituitary Cells ◽  
Lh Secretion

The purpose of the present experiments was to examine the short- and long-term effects of estradiol-17β (E2), progesterone (P), and 5α-dihydrotestosterone (DHT), alone and in combination, on the gonadotrophin-releasing hormone (GnRH)-induced luteinizing hormone (LH) secretion, using an ovariectomized rat pituitary cells culture model. After 72 h in steroid-free medium, pituitary cells were further cultured for 24 h in medium with or without E2 (1 nM), P (100 nM), or DHT (10 nM). Cultures were then incubated for 5 h in the absence or presence of 1 nM GnRH with or without steroids. LH was measured in the medium and cell extract by radioimmunoassay. The results show that the steroid hormones exert opposite effects on the release of LH induced by GnRH, which seems to be dependent upon the length of time the pituitary cells have been exposed to the steroids. In fact, short-term (5 h) action of E2 resulted in a partial inhibition (64% of control) of LH release in response to GnRH, while long-term (24 h) exposure enhanced (158%) GnRH-induced LH release. Similar results were obtained with DHT, although the magnitude of the effect was lower than with E2. Conversely, P caused an acute stimulatory action (118%) on the LH released in response to GnRH and a slightly inhibitory effect (90%) after chronic treatment. GnRH-stimulated LH biosynthesis was also influenced by steroid treatment. Significant increases in total (cells plus medium) LH were observed in pituitary cells treated with E2 or DHT. While the stimulatory effect of E2 was evident after both acute (133%) and chronic (119%) treatment, that of DHT appears to be exerted mainly after long-term priming (118%). These results suggest that the steroids modulate GnRH-induced LH secretion by acting on both synthesis and release of LH. On the other hand, total hormone content was not affected by P. The acute (5 h) effects of E2, P, and DHT on the GnRH response in E2-primed (24 h) cells during a short-term incubation, were also tested. Addition of P to the pituitary cells primed with E2 led to an acute potentiation of the stimulatory effect of E2 on GnRH-induced LH release and total content. Conversely, the augmentative E2 effect on pituitary responsiveness to GnRH was abolished by DHT. Taken together, these findings suggest that the physiological significance of the stimulatory action of progesterone could be to define the final magnitude of the LH preovulatory surge, while the inhibition by DHT could be required to limit the LH surge to that day of proestrus.Key words: luteinizing hormone, gonadotrophin-releasing hormone, steroid hormones, cultured pituitary cells.

Ca2+ entry in gonadotrophs and αT3–1 cells: does store-dependent Ca2+ influx mediate gonadotrophin-releasing hormone action?

1996 ◽  
Vol 149 (1) ◽  
pp. 155-169 ◽  
Author(s):  
C A McArdle ◽  
W Forrest-Owen ◽  
J S Davidson ◽  
R Fowkes ◽  
R Bunting ◽  
...  
Keyword(s):  
Transient Increase ◽  
Pituitary Cells ◽  
Free Medium ◽  
Excitable Cells ◽  
Rat Pituitary ◽  
Intracellular Ca ◽  
Lh Release ◽  
Gonadotrophin Releasing Hormone ◽  
Rat Pituitary Cells

Abstract In pituitary gonadotrophs GnRH causes biphasic (spike and plateau) increases in cytosolic Ca2+ ([Ca2+]i) and gonadotrophin release. The spike phases reflect mobilization of stored Ca2+ and the plateau responses are attributed, in part, to Ca2+ influx via voltage-sensitive Ca2+ channels. In recent years, store-dependent Ca2+ influx (SDCI), in which depletion of the intracellular inositol 1,4,5-trisphosphate-mobilizable pool stimulates Ca2+ influx, has emerged as a major form of Ca2+ entry activated by phosphoinositidase C-coupled receptors in non-excitable cells. More recent evidence also indicates a role for SDCI in excitable cells. We have used dynamic video imaging of [Ca2+]i, in αT3–1 cells (a gonadotroph-derived cell line) and manipulation of the filling state of the GnRH-mobilizable Ca2+ pool to test the possible role of SDCI in GnRH action. In Ca2+-containing medium, GnRH caused a biphasic increase in [Ca2+]i whereas in Ca2+-free medium only a transient increase occurred. The response to a second stimulation with GnRH in Ca2+-free medium was reduced by >95% (demonstrating that Ca2+ pool depletion had occurred) and was recovered after brief exposure to Ca2+-containing medium (which enables refilling of the pool). Ionomycin (a Ca2+ ionophore) and thapsigargin (which inhibits the Ca2+-sequestering ATPase of the endoplasmic reticulum) also transiently increased [Ca2+]i, in Ca2+-free medium and depleted the GnRH-mobilizable pool as indicated by greatly reduced subsequent responses to GnRH. Pool depletion also occurs on stimulation with GnRH in Ca2+-containing medium because addition of ionomycin and Ca2+-free medium during the plateau phase of the GnRH response caused only a reduction in [Ca2+]i rather than the transient increase seen without GnRH. To deplete intracellular Ca2+ pools, cells were pretreated in Ca2+-free medium with thapsigargin or GnRH and then, after extensive washing, returned to Ca2+-containing medium. Pretreatment with thapsigargin augmented the increase in [Ca2+]i seen on return to Ca2+-containing medium (to two- to threefold higher than that seen in control cells) indicating the activation of SDCI, whereas pool depletion by GnRH pretreatment had no such effect. To ensure maintained pool depletion after Ca2+ re-addition, similar studies were performed in which the thapsigargin and GnRH treatments were not washed off, but were retained through the period of return to Ca2+-containing medium. Return of GnRH-treated cells to Ca2+-containing medium caused an increase in [Ca2+]i which was inhibited by nicardipine, whereas the increase seen on return of thapsigargin-treated cells to Ca2+-containing medium was not reduced by nicardipine. The quench of fura-2 fluorescence by MnCl2 (used as a reporter of Ca2+ influx) was increased by GnRH and thapsigargin, indicating that both stimulate Ca2+ influx via Mn2+ permeant channels. The GnRH effect was abolished by nicardipine whereas that of thapsigargin was not. Finally, depletion of intracellular Ca2+ pools by pretreatment of superfused rat pituitary cells with GnRH or thapsigargin in Ca2+-free medium did not enhance LH release on return to Ca2+-containing medium. The results indicate that (a) thapsigargin stimulates SDCI in αT3–1 cells via nicardipine-insensitive Ca2+ channels, (b) in spite of the fact that GnRH depletes the hormone-mobilizable Ca2+ pool, it fails to stimulate SDCI, (c) GnRH stimulates Ca2+ entry predominantly via nicardipine-sensitive channels, a route not activated by SDCI and (d) in rat gonadotrophs, GnRH-stimulated LH release is not mediated by SDCI. Journai of Endocrinology (1996) 149, 155–169

Short-term effects of oestradiol and 4-hydroxyoestradiol on LH-secretion by cultured rat pituitary cells

1985 ◽  
Vol 110 (1_Suppla) ◽  
pp. S7-S8 ◽  
Author(s):  
G. EMONS ◽  
O. ORTMANN ◽  
U. FINGSCHEIDT ◽  
P. BALL ◽  
R. KNUPPEN
Keyword(s):  
Pituitary Cells ◽  
Short Term ◽  
Rat Pituitary ◽  
Rat Pituitary Cells ◽  
Lh Secretion ◽  
Short Term Effects

Action and formation of inositol bisphosphate and inositol trisphosphate in rat anterior pituitary cells

1990 ◽  
Vol 123 (4) ◽  
pp. 459-463 ◽  
Author(s):  
Andrzej F. Przylipiak ◽  
Ludwig Kiesel ◽  
Thomas A. Karenberg ◽  
Maria S. Przylipiak ◽  
Benno Runnebaum
Keyword(s):  
Inositol Phosphates ◽  
Inositol Trisphosphate ◽  
Pituitary Cells ◽  
Permeabilized Cells ◽  
New Findings ◽  
Rat Pituitary ◽  
Lh Release ◽  
Rat Pituitary Cells ◽  
Lh Secretion

Abstract. Inositol 4,5-bisphosphate and inositol 1,4,5-trisphosphate, administered exogenously at a concentration of 3× 10−5 mol/l increased LH release in superfused rat pituitary cells by 950±267% and 281±83%, respectively. This stimulatory effect was reversible and dose-dependent. Other inositol phosphates (inositol 1-monophosphate, inositol 1,4,5,6-tetrakisphosphate, inositol 1,3,4,5,6-pentakisphosphate and inositol 1,2,3,4,5,6-hexakisphosphate), tested in vitro, did not significantly influence LH release. In saponin-permeabilized cells, the rate of basal and stimulated LH release was twice that in non-permeabilized cells. Penetration of inositol bisphosphate and inositol trisphosphate into saponin-treated pituitary cells did not increase the secretory potency of these agents compared with their effect on non-permeabilized cells. The new findings document that inositol trisphosphate formation occurs within 5-45 s after GnRH (10−7 mol/l) administration and seems to be involved in mediating the rapid, first phase of LH release, whereas inositol bisphosphate formation occurs after 3-15 min and is probably related to later phases of LH secretion. Our results suggest that inositol bisphosphate and inositol trisphosphate are important regulators of the release of luteinizing hormone and can exert their effects not only intracellularly, but also extracellularly.

Acute effects of oestradiol and progesterone on melittin- and gonadotrophin-releasing hormone-induced LH secretion

1992 ◽  
Vol 132 (2) ◽  
pp. 251-259 ◽  
Author(s):  
O. Ortmann ◽  
K. Johannsen ◽  
R. Knuppen ◽  
G. Emons
Keyword(s):  
Arachidonic Acid ◽  
Control Pulse ◽  
Pituitary Cells ◽  
Short Term ◽  
Releasing Hormone ◽  
Progesterone Treatment ◽  
Gonadotrophin Releasing Hormone ◽  
Lh Secretion ◽  
Kinetics Of

ABSTRACT It is well established that oestradiol and progesterone modulate gonadotrophin-releasing hormone (GnRH)-induced LH secretion from cultured rat pituitary cells. Short-term oestradiol and long-term progesterone treatment exert inhibition, while short-term progesterone and long-term oestradiol treatment lead to enhancement of GnRH-stimulated LH secretion. There are several lines of evidence to suggest that the steroid effects might be mediated via a mechanism involving modulation of the GnRH signal-transduction system. To evaluate the role of arachidonic acid, which serves as an intracellular signal transducer by itself or its lipoxygenase metabolites, in the mediation of oestradiol and progesterone actions, we examined their effects on melittin (activator of phospholipase A2)-stimulated LH secretion. When pituitary cells from adult female rats were treated for 48 h with 1 nmol oestradiol/l or 1 nmol oestradiol/l plus 100 nmol progesterone/l, GnRH (1 nmol/l)-induced LH secretion was stimulated or inhibited respectively. However, melittin (10–300 nmol/l)-stimulated LH secretion remained unaffected after such treatment. Short-term treatment with oestradiol inhibited GnRH-induced LH secretion while progesterone treatment of oestradiol-primed cells led to a stimulatory effect. Interestingly, melittin-stimulated LH secretion was influenced in the same way after the short treatment paradigm. Perifusion studies were performed to assess the kinetics of these acute steroid actions further. Four separate perifusion chambers were continuously perifused with medium and stimulated for 2 min with 1 nmol GnRH/l or 1 μmol melittin/l every 50 min in a pulsatile fashion. When 1 nmol oestradiol/l was added to the perifusion medium after the application of an initial control pulse, GnRH- and melittin-stimulated LH secretion were inhibited by 69 and 61% respectively. This effect was present after 50 min. When oestradiol-primed cells were treated with 100 nmol progesterone/l starting after the initial GnRH or melittin pulse, an acute stimulatory effect was observed in response to both stimuli after 50 min. LH release was enhanced by up to 279 (GnRH) or 419% (melittin) compared with the control pulse. The kinetics of inhibited or stimulated pulsatile LH secretion were virtually identical when GnRH or melittin were used as stimuli. These results demonstrate that short-term oestradiol or progesterone treatment modulate arachidonic acid-mediated LH secretion in a similar fashion to GnRH-induced LH secretion, while long-term oestradiol or progesterone treatment only affected GnRH-induced LH secretion. Journal of Endocrinology (1992) 132, 251–259

Melatonin inhibits luteinizing hormone releasing hormone (LHRH) induction of LH release from fetal rat pituitary cells

1995 ◽  
Vol 184 (2) ◽  
pp. 109-112 ◽  
Author(s):  
Atsuhiko Hattori ◽  
Damon C. Herbert ◽  
Mary K. Vaughan ◽  
Ken Yaga ◽  
Russel J. Reiter
Keyword(s):  
Luteinizing Hormone ◽  
Pituitary Cells ◽  
Luteinizing Hormone Releasing Hormone ◽  
Releasing Hormone ◽  
Fetal Rat ◽  
Rat Pituitary ◽  
Lh Release ◽  
Rat Pituitary Cells

In vitro evidence of a role for inhibin in female rabbits

1984 ◽  
Vol 246 (3) ◽  
pp. E243-E248
Author(s):  
A. L. Goodman
Keyword(s):  
Granulosa Cells ◽  
Pituitary Cells ◽  
Dependent Manner ◽  
C Cells ◽  
Rat Pituitary ◽  
Reference Preparation ◽  
Lh Release ◽  
Rat Pituitary Cells ◽  
I Cells

To examine a regulatory role for inhibin in female rabbits, an in vitro bioassay for inhibin activity was modified to use cultured rabbit pituitary cells and charcoal-extracted porcine follicular fluid (pFFx) as a reference preparation. pFFx inhibited follicle-stimulating hormone (FSH) release in a dose-dependent manner in cultures from both intact (I) and castrate (C) does at doses that also inhibited FSH release by cultured rat pituitary cells. Basal FSH release by I cells was inhibited greater than 10% by 0.02% (vol/vol) and greater than 90% by greater than or equal to 0.2% pFFx, whereas in C cells maximal inhibition of FSH release plateaued at only approximately 75%. FSH secretion was restored after removal of pFFx in day 2 media. Luteinizing hormone (LH) release by C cells was not inhibited at any dose of pFFx, but in I cells LH was progressively inhibited to approximately 60% of control levels during day 2 (but not day 1). Charcoal-extracted media (0.25-1%) in which 5 X 10(5) rabbit granulosa cells had been earlier cultured for 72 h produced a parallel inhibition of FSH release. The present findings demonstrate that 1) rabbit pituitary cells are responsive to inhibin, i.e., pFFx preferentially inhibited FSH secretion in a direct, graded, and reversible manner and 2) rabbit follicular granulosa cells secrete an inhibin-like substance.

Progesterone together with estradiol promotes luteinizing hormone β-subunit mRNA stability in rat pituitary cells cultured in vitro

1996 ◽  
Vol 134 (2) ◽  
pp. 236-242 ◽  
Author(s):  
Deokbae Park ◽  
Minseok cheon ◽  
Changmee Kim ◽  
Kyungjin Kim ◽  
Kyungza Ryu
Keyword(s):  
Mrna Stability ◽  
Actinomycin D ◽  
Pituitary Cells ◽  
Mrna Levels ◽  
Ovarian Steroids ◽  
Subunit Mrna ◽  
Rat Pituitary ◽  
Lh Release ◽  
Rat Pituitary Cells

Park D, Cheon M, Kim C, Kim K, Ryu K. Progesterone together with estradiol promotes luteinizing hormoneβ-subunit mRNA stability in rat pituitary cells in vitro. Eur J Endocrinol 1996;134:236–42. ISSN 0804–4643 The present study examined the role of ovarian steroids, estradiol and/or progesterone in the regulation of luteinizing hormone β-subunit (LH-β) mRNA levels and LH release in the rat anterior pituitary cells cultured in vitro. When estradiol (10 nmol/l and/or progesterone (100 nmol/l) were added to the cultures, neither estradiol or progesterone nor both together altered the basal LH-β mRNA levels or LH release. Continuous exposure to gonadotropin-releasing hormone (GnRH, 0.2 nmol/l) for 24 h markedly induced LH-β mRNA accumulation, and in this experimental condition, progesterone alone and progesterone + estradiol further augmented GnRH-induced LH-β mRNA levels and LH release. Then we explored further the possibility that ovarian steroids are involved in modulating LH-β mRNA stability in cultured rat pituitary cells where transcription was inhibited by actinomycin D. Anterior pituitary cells were preincubated with GnRH (0.2 nmol/l) for 16 h and, after removing GnRH from culture medium, the cells were incubated further in the presence of actinomycin D (5 μmol/l) for 24 h. The LH-β mRNA levels gradually declined to about 30% of the control values (zero time point after GnRH removal) in a time-dependent manner. During this period, either progesterone alone or progesterone + estradiol clearly blocked the degradation of LH-β mRNA species. These results indicate that ovarian steroids promote LH-β mRNA stability, thereby contributing to the maintenance of GnRH-stimulated LH-β mRNA levels. Kyungza Ryu, Department of Pharmacology, College of Medicine, Yonsei University, 120-749, Seoul, Korea

Influence of melatonin and oestradiol on the opioidergic regulation of LH and prolactin release in pony mares

1997 ◽  
Vol 154 (2) ◽  
pp. 241-248 ◽  
Author(s):  
C Aurich ◽  
J Lange ◽  
H-O Hoppen ◽  
J E Aurich
Keyword(s):  
Prolactin Secretion ◽  
Opioid Antagonist ◽  
Short Term ◽  
Melatonin Treatment ◽  
Prolactin Release ◽  
Oestradiol Treatment ◽  
Oestradiol Benzoate ◽  
Lh Release ◽  
Lh Secretion

Abstract The aim of this study was to investigate the influence of oestradiol, melatonin and season on the opioid regulation of LH and prolactin release. Effects of the opioid antagonist naloxone (0·5 mg/kg) on LH and prolactin secretion were determined in ovariectomized pony mares. In experiment 1, mares in January (n=6) were pretreated with oestradiol benzoate (5 μg/kg) for 20 days. In experiment 2, beginning in May, mares (n=7) received melatonin (15 mg) for 15 days and subsequently a combination of melatonin plus oestradiol for 20 days. In experiment 3, beginning in May, mares (n=6) were pretreated with oestradiol for 30 days, left untreated for 12 days and then given melatonin for 35 days. In all experiments the animals were injected with the opioid antagonist naloxone and saline on 2 consecutive days prior to treatment. In experiment 1, animals received naloxone and saline on days 10 and 11 and 20 and 21 following oestradiol treatment. In experiment 2, naloxone and saline were administered on days 15 and 16 following melatonin treatment and on days 10 and 11 and 20 and 21 of melatonin plus oestradiol treatment. In experiment 3, the animals received naloxone and saline on days 10 and 11, 20 and 21 and 30 and 31 of oestradiol treatment, prior to melatonin treatment and on days 15 and 16, 25 and 26 and 35 and 36 following melatonin. In January (experiment 1), naloxone evoked a significant (P<0·05) LH release at all times, however the LH increment in response to naloxone increased during oestradiol pretreatment (P<0·05) During the breeding season (experiments 2 and 3), naloxone induced a significant (P<0·05) increase in plasma LH concentrations when mares had not been pretreated with oestradiol or melatonin and after oestradiol pretreatment. Basal LH concentrations and the LH increment in response to naloxone increased significantly (P<0·05) during the 30-day oestradiol pretreatment. Melatonin decreased the naloxone-induced LH release and the LH release in response to naloxone and saline no longer differed after 25 and 35 days of melatonin pretreatment. When melatonin was given together with oestradiol for 20 days, again a significant (P<0·05) LH release in response to naloxone occurred. Prolactin release was significantly (P<0·05) increased by naloxone when mares had been pretreated with only melatonin. The opioid antagonist did not affect prolactin release in mares that had not been pretreated or received oestradiol either alone or in combination with melatonin. In conclusion, in long-term ovariectomized mares, opioids inhibit LH secretion independent from ovarian factors. This opioid inhibition of LH secretion is enhanced by oestradiol and reduced by melatonin. Although short-term melatonin treatment in-activates the opioid regulation of LH release, a prolonged influence of melatonin as occurs in winter does not prevent activation of the opioid system. This indicates that effects of melatonin on the opioid regulation of LH release change with time. An opioid inhibition of prolactin secretion is activated by melatonin given for 15–35 days but is lost under the prolonged influence of a short-day melatonin signal in winter. Journal of Endocrinology (1997) 154, 241–248

scholarly journals Effects of activin on hormone secretion by single female rat pituitary cells: analysis by cell immunoblot assay

1999 ◽  
Vol 161 (3) ◽  
pp. 375-382 ◽  
Author(s):  
S Miyamoto ◽  
M Irahara ◽  
K Ushigoe ◽  
A Kuwahara ◽  
H Sugino ◽  
...  
Keyword(s):  
Hormone Secretion ◽  
Activin A ◽  
Female Rats ◽  
Pituitary Cells ◽  
Female Rat ◽  
Single Female ◽  
Immunoblot Assay ◽  
Rat Pituitary ◽  
Rat Pituitary Cells ◽  
Lh Secretion

We investigated the effect of activin A on secretion of LH, FSH, and prolactin (PRL) by female cultured rat pituitary cells at the single-cell level by means of the cell immunoblot assay. Anterior pituitary cells from 8-week-old female rats were preincubated with or without activin A for 24 h, after which they were monodispersed and immediately used for cell immunoblot assay. The percentages of LH-, FSH- and PRL-immunoreactive cell blots detected were 5.5, 5.3 and 43.1%, respectively, of all pituitary cells applied to the transfer membrane. The percentage of LH-secreting cells and mean LH secretion per cell did not change after treatment with activin. In contrast, activin significantly increased the percentage of FSH-secreting cells and mean FSH secretion per cell to 136.0 and 114. 5% respectively. In addition, activin significantly decreased the percentage of PRL-secreting cells and mean PRL secretion per cell to 52.2 and 72.0% respectively. These results suggest that (1) activin A has effects on female rat pituitary cells that increase not only the amount of FSH secretion per cell but also the number of FSH-secreting cells, and (2) activin A decreases both the amount of PRL secretion per cell and the number of PRL-secreting cells.

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