Gonadotrophin releasing hormone receptors and the response of pituitary gonadotrophs in culture

1983 ◽  
Vol 98 (3) ◽  
pp. 391-399 ◽  
Author(s):  
A. Bérault ◽  
M.-T. Jansem de Almeida Catanho ◽  
M. Théoleyre ◽  
M. Jutisz
Keyword(s):  
Hormone Receptors ◽  
Binding Capacity ◽  
High Capacity ◽  
Biological Response ◽  
Cell Number ◽  
Culture Period ◽  
Pituitary Cells ◽  
Cell Content ◽  
Releasing Hormone ◽  
Gonadotrophin Releasing Hormone

We have investigated the effect of the time of culture on cell number, cell content of LH, cell responsiveness to gonadotrophin releasing hormone (GnRH) and binding parameters of GnRH in rat anterior pituitary cells in culture. Although a decrease in the cell number was observed during the culture period, the cell content of LH remained unchanged. The receptor affinity (Ka) in acutely dispersed cells was 0·86 × 107l/mol for [3H]GnRH and 1·36 × 1010l/mol for a highly potent agonist, d-Ser(But)6]GnRH(1–9)nonapeptide-ethylamide (GnRH-A). The affinity and binding capacity (0·3 fmol/106 cells) for iodinated GnRH-A did not change significantly during the 6-day culture period. On the contrary, the values of Ka and binding capacity (257 fmol/106 cells) for tritiated GnRH decreased by about 50% betweeen days 1 and 6 of culture. Our results suggest that 125I-labelled GnRH-A binds mostly to high-affinity and low-capacity receptor sites, while [3H]GnRH, which must be used at a higher concentration, also binds to low-affinity, high-capacity binding sites and is therefore useless for the measurement of GnRH receptor binding affinity and binding capacity. Since the biological response of the cells to GnRH increased with the time of culture, it is concluded that although GnRH action is receptor-mediated, binding capacity and biological activity are not necessarily correlated.

Effect of chronic exposure to a gonadotrophin-releasing hormone agonist on in vitro responsiveness of ovine pituitary cells to inhibin, activin and oestradiol

1994 ◽  
Vol 140 (3) ◽  
pp. 483-493 ◽  
Author(s):  
S Muttukrishna ◽  
P G Knight
Keyword(s):  
Gnrh Agonist ◽  
Inhibitory Effect ◽  
Pituitary Cells ◽  
Cell Content ◽  
Releasing Hormone ◽  
Basal Release ◽  
Lh Release ◽  
Gonadotrophin Releasing Hormone

Abstract To investigate the extent to which the direct actions of inhibin, activin and oestradiol on pituitary output of FSH and LH are dependent on the presence of functional gonadotrophin-releasing hormone (GnRH) receptors, we have compared the effects of these agents on cultured ovine pituitary cells derived from control and GnRH agonist-suppressed ewes. Chronic treatment with GnRH agonist reduced plasma LH and FSH levels (P<0·01) and abolished GnRH-induced release of LH and FSH both in vivo and in vitro. As expected, basal LH release and LH cell content in vitro were drastically reduced in GnRH agonist-suppressed cells (P<0·001). However, basal FSH release and FSH cell content were approximately twofold higher than in control cells (P<0·001). Irrespective of whether the cells had been desensitized to GnRH, inhibin and oestradiol were both found to suppress basal FSH release and FSH cell content in a dose-dependent fashion (P<0·001). Although inhibin had no effect on basal release of LH from control cells, it markedly enhanced GnRH-induced release (P<0·001). In contrast, inhibin increased (P<0·001) basal LH release from GnRH agonist-suppressed cells (which were unresponsive to the GnRH challenge). Inhibin had no overall effect on total LH content/well for either control or GnRH agonist-suppressed cells. Treatment with oestradiol, on the other hand, reduced total LH content/well, an effect which was more pronounced with GnRH agonist-suppressed cells (−44%; P<0·001) than with control cells (−14%, P<0·01). Whereas in control cells activin had no significant effect on any aspect of FSH production examined, in GnRH agonist-treated cells activin enhanced basal FSH release, residual cell content and total FSH content/well (P<0·001). Altering GnRH receptor status also modified the LH response to activin. With control cells activin increased basal release (P<0·001), decreased GnRH-induced release (P<0·001) and increased total LH content/well (P<0·001). With GnRH agonist-treated cells, however, activin had a uniform inhibitory effect on each aspect of LH production examined (P<0·001 in each case). It was concluded that desensitization of ovine gonadotrophs to GnRH by chronic agonist treatment results in a paradoxical enhancement of FSH output in vitro but has little effect on the responsiveness of the cells (in terms of gonadotrophin release and content) to either inhibin or oestradiol. In contrast, GnRH agonist treatment leads to qualitative changes in cellular reponsiveness to activin. Journal of Endocrinology (1994) 140, 483–493

Estimating the rate of externalization of gonadotrophin-releasing hormone receptors in ovine anterior pituitary cells in vitro

1988 ◽  
Vol 117 (1) ◽  
pp. 97-107
Author(s):  
L. Starling ◽  
J. E. A. McIntosh ◽  
R. P. McIntosh
Keyword(s):  
Plasma Membrane ◽  
Anterior Pituitary ◽  
Hormone Receptors ◽  
Membrane Receptors ◽  
Second Phase ◽  
Pituitary Cells ◽  
Releasing Hormone ◽  
Anterior Pituitary Cells ◽  
Rate Of Increase ◽  
Gonadotrophin Releasing Hormone

ABSTRACT We report an estimate of the rate of externalization of unstimulated receptors for gonadotrophin-releasing hormone (GnRH), and derive from this the turnover time of the unstimulated receptor. The binding of the GnRH antagonist acetyl-d-pCl-Phe1,2,d-Trp3,d-Lys6,d-Ala10]-GnRH to dispersed sheep anterior pituitary cells was non-saturable at 37 °C. Further experiments showed that the binding had two distinct phases. We suggest that these phases correspond to the initial, saturable binding to existing plasma membrane receptors, followed by binding to receptors as they are inserted into the surface membrane. The two processes are temporally distinct, and can be inhibited independently by pharmacological manipulations. The initial phase was inhibited by treatments that could be expected to reduce the number of active receptors on the cell surface (preincubation of the cells for 30 min with 100 μg neuraminidase/ml or 50 μmol GnRH/ml), and was complete in less than 30 min after the addition of the antagonist tracer. The second phase occurred continuously in the presence of tracer, and was reduced or abolished by inhibitors of microtubule function (100 μmol vinblastine/l), protein synthesis (25 μg cycloheximide/ml), or energy metabolism (0·25 mmol 2,4-dinitrophenol/l). The rate of insertion of receptors into the plasma membrane was calculated from the rate of increase of the second phase of binding. The calculated rate implies a 100% turnover of unstimulated receptors every 150 min. In contrast, previously published estimates of the rate of internalization of the GnRH–receptor complex in the rat pituitary suggest that the stimulated receptor is turned over much faster. J. Endocr. (1988) 117, 97–107

Varying the patterns and concentrations of gonadotrophin-releasing hormone stimulation does not alter the ratio of LH and FSH released from perifused sheep pituitary cells

1986 ◽  
Vol 109 (2) ◽  
pp. 155-161 ◽  
Author(s):  
J. E. A. McIntosh ◽  
R. P. McIntosh
Keyword(s):  
Dose Response ◽  
Dose Interval ◽  
Follicular Phase ◽  
Pituitary Cells ◽  
Ovulatory Cycle ◽  
Short Term ◽  
Releasing Hormone ◽  
Dissociated Cells ◽  
Gonadotrophin Releasing Hormone

ABSTRACT Our aim was to determine whether release of LH and FSH can be controlled differentially by the characteristics of applied signals of stimulatory gonadotrophin-releasing hormone (GnRH) alone, free of the effects of steroid feedback or other influences from the whole animal. The outputs of both gonadotrophins were significantly correlated (r≈0·90; P<0·0005) when samples of freshly dispersed sheep pituitary cells were perifused in columns for 7 h with medium containing a range of concentrations of GnRH in various patterns of pulses. Hormone released in response to the second, third and fourth pulses from every column was analysed in detail. Dose–response relationships for both LH and FSH were very similar when cells were stimulated with 5–8500 pmol GnRH/1 in 5-min pulses every hour. When GnRH was delivered in pulses at a maximally stimulating level, the outputs of both hormones increased similarly with increasing inter-pulse intervals. Efficiency of stimulation (release of gonadotrophin/unit stimulatory GnRH) decreased (was desensitized) with increasing pulse duration in the same way for both hormones. Thus, varying the dose, interval and duration of GnRH pulses did not alter the proportions of LH and FSH released in the short-term from freshly dissociated cells. However, the same cell preparations released more LH relative to FSH when treated with maximally stimulating levels of GnRH for 3 h in the presence of 10% serum from a sheep in the follicular phase of its ovulatory cycle compared with charcoal-treated serum. Because there was no gonadotrophin synthesis under the conditions used in vitro these results suggest that changes in the LH/FSH ratio seen in whole animals are more likely to result from differential clearance from the circulation, ovarian feedback at the pituitary, differential synthesis in intact tissue or another hormone influencing FSH secretion, rather than from differences in the mechanism of acute release controlled by GnRH. J. Endocr. (1986) 109, 155–161

Milk production in concurrently pregnant and lactating goats mated out of season

1988 ◽  
Vol 55 (4) ◽  
pp. 487-493 ◽  
Author(s):  
Christopher H. Knight ◽  
Colin J. Wilde
Keyword(s):  
Milk Production ◽  
Milk Yield ◽  
Hormone Treatment ◽  
Cell Number ◽  
Releasing Hormone ◽  
Control Value ◽  
Lactating Goats ◽  
Mammary Cell ◽  
Gonadotrophin Releasing Hormone ◽  
Stage Yield

SummaryFive lactating goats which had kidded normally in March were mated during seasonal anoestrus in May, at the time of peak milk production, after ovulation had been induced using gonadotrophin-releasing hormone (Knight et al. 1988). Milk yield was unaffected by the hormone treatment, and decreased at the same rate as that of control (non-pregnant) goats for the first 8 weeks of the pregnancy. Thereafter yield declined more quickly in the test goats and just before parturition (in October) was 57% of the control value. Following parturition in the test animals, yield rose rapidly as the second lactation was established. None became ‘dry’ at any stage. Yield continued to decline with advancing lactation in the controls, which were mated normally in October or November and dried-off in December. During their second (‘extra’) lactation in the winter the test animals produced 12% less than in a normal second lactation in summertime; during the year the extra lactation meant that the test animals produced 73% more milk than the controls. In some, a second concurrent pregnancy was established during the extra lactation, with the resuit that three lactations were obtained in the time normally taken for two. Mammary cell number and proliferation rate were both higher in the pregnant animals than in the controls in week 23 of the first lactation.

Effects of the daily administration of gonadotrophin-releasing hormone on the anterior pituitary gland of rats with restricted feeding

1992 ◽  
Vol 134 (2) ◽  
pp. 177-NP ◽  
Author(s):  
F. Kotsuji ◽  
K. Hosokawa ◽  
T. Tominaga
Keyword(s):  
Weight Loss ◽  
Weight Reduction ◽  
Anterior Pituitary ◽  
Serum Levels ◽  
Female Rats ◽  
Daily Administration ◽  
Pituitary Cells ◽  
Restricted Feeding ◽  
Releasing Hormone ◽  
Gonadotrophin Releasing Hormone

ABSTRACT To investigate the influence of weight reduction on pituitary function and its modulation by gonadotrophin-releasing hormone (GnRH), female rats were restricted to 10 g food/day for 60 days. GnRH (5 μg) or saline (0·2 ml) were administered daily between days 31 and 60 of the period of underfeeding. Underfeeding brought about a decrease in the pituitary gonadotrophin content, serum levels of gonadotrophins and oestradiol, and the number and size of both LH- and FSH-positive pituitary cells. The administration of GnRH to underfed rats produced an increase in the pituitary and serum gonadotrophin levels and the number and size of both LH- and FSH-positive pituitary cells. These observations suggest that underfeeding and/or weight loss diminish the number and activity of the pituitary gonadotrophs, and that daily administration of GnRH both increases the number of gonadotrophs and augments their activity. Journal of Endocrinology (1992) 134, 177–182

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