Adrenergic regulation of growth hormone secretion in the ewe.

This study examined the role of the adrenergic system in the regulation of growth hormone (GH) secretion in sheep. Intravenous infusion of noradrenaline (0.5 microgram/kg per min for 2 hr) totally suppressed plasma GH concentrations. Concomitant treatment of animals with the beta-adrenergic antagonist propranolol completely blocked the noradrenaline-induced suppression of GH. In contrast, intravenous injection of the centrally acting alpha 2-agonist clonidine (2 micrograms/kg) elicited a release of GH. To further investigate the central adrenergic regulation of GH secretion 10 micrograms of noradrenaline or adrenaline was microinjected (1 microliter) directly into the preoptic area of the hypothalamus of ovariectomized ewes. When the time of injection coincided with a GH trough period, both noradrenaline and adrenaline caused an increase in plasma GH concentrations, whereas if the injection coincided with an endogenous pulse of GH no additional GH response was obtained. In conclusion, these results provide evidence for the involvement of the adrenergic system in the regulation of GH secretion in sheep. Centrally, adrenergic pathways exert a stimulatory effect on GH release via an alpha 2-adrenergic system, whereas peripherally adrenergic pathways exert an inhibitory effect via beta-adrenergic mediated mechanisms. Furthermore, adrenergic stimulation of the preoptic area may inhibit somatostatin activity and directly facilitate a GH pulse. Alternatively, adrenergic innervation of the preoptic area may influence neurons (somatostatin or other) that project to the arcuate nucleus and stimulate the release of GH-releasing factor.

An epithelial cyst of the suprasellar region.

A case of a 26 year old woman with increasing headaches due to a suprasellar cyst is described. Histology was consistent with an epithelial cyst with features resembling both Rathke's cleft cysts and enterogenous cysts. The differential diagnosis and treatment of suprasellar cysts is discussed.

Human growth hormone releasing factor (hGRF) modulates calcium currents in human growth hormone secreting adenoma cells.

Electrophysiology of human growth hormone secreting tumour cells and its modification by hGRF has been studied using on-cell and Nystatin-perforated whole-cell recording configurations. Local application of hGRF (10 nM) produced an increase in the frequency of action potentials. Ca2+ currents were isolated by a ramp depolarizing pulse from -120 mV to +60 mV in the presence of tetrodotoxin (1 microM). Human GRF increased the Ca2+ currents which could be blocked by Ni+ (300 microM). We conclude that an increase in Ca2+ current is integral to the action of hGRF on these cells.

The negative feedback effects of testicular steroids are predominantly at the hypothalamus in the ram.

This study aimed to delineate the hypothalamic and/or pituitary actions of testosterone and its primary metabolites 5 alpha-dihydrotestosterone and estradiol (E) in adult castrated rams (wethers) during the breeding season. In Exp 1, wethers were treated for a week with twice daily injections (im) of peanut oil, 8, 16 or 32 mg/day testosterone propionate (TP) or dihydrotestosterone benzoate (DHTB) or an sc silastic implant containing 1 or 3 cm E. TP decreased plasma LH concentrations, increased (P less than 0.05) LH interpulse interval, did not have consistent effects on LH pulse amplitude, and had minimal effects on plasma FSH concentrations. DHTB decreased LH and FSH concentrations and increased (P less than 0.05) LH interpulse interval. E reduced (P less than 0.05) plasma LH and FSH concentrations and increased LH interpulse interval but had no effects on LH pulse amplitude. In Exp 2, hypothalamo-pituitary disconnected wethers given 125 ng GnRH every 2 h, were treated with either peanut oil, 32 mg/day TP or DHTB or 3 cm E. None of the treatments affected plasma LH or FSH concentrations or LH pulse amplitude. Exp 3 investigated the effects on GnRH of treatment of wethers either with peanut oil or TP. TP reduced GnRH concentrations (P less than 0.05) and pulse amplitude (P less than 0.01) and increased interpulse interval (P less than 0.05). These data provide evidence that, during the breeding season, the principal site of negative feedback of testicular steroids in the ram is the hypothalamus, resulting in decreased GnRH secretion; feedback effects at the pituitary are minimal.

Circulating half-lives of follicle-stimulating hormone and luteinizing hormone in pituitary extracts and isoform fractions of ovariectomized and intact ewes.

Previous studies have shown that the circulating half-life (t 1/2) of serum FSH in ewes after hypophysectomy (HPX) increased 10-fold after ovariectomy (OVEX). The basis for this difference was examined in this study by determining the circulating half-life of serum FSH and LH in HPX ewes after administration of pituitary extracts and gonadotropin isoform fractions. High-speed supernatants of pituitaries from gonadal-intact and OVEX ewes were fractionated by electrofocusing in sucrose gradients and based on the pI distribution of FSH and LH divided into four pools, pH 4.3-4.8, 4.8-5.55, 5.8-6.7, and 6.7-10. These extracts were administered by iv bolus injection to HPX gonadal-intact ewes and blood samples collected between 15-1000 min later. The clearance pattern for both serum FSH and LH was heterogenous, indicative of a major rapid and a minor slow dissociating component. A significant (P less than 0.05) difference in circulating half-lives (rapid component) was observed between pituitary extracts from intact and OVEX ewes for FSH (t 1/2 = 32.8 +/- 8.6 min vs. 89.9 +/- 32.3 min) but not LH (31.3 +/- 9.2 min vs. 39.3 +/- 6.1 min, respectively), whereas no significant difference was observed between the corresponding FSH or LH isoform preparations. To establish if the difference in circulating half-lives obtained after HPX and bolus iv injection was due to mode of delivery, an extract of pituitaries from OVEX ewes was infused for 12 h into HPX sheep and the t 1/2 values determined after cessation of treatment and compared to those after a bolus injection. The clearance of both FSH and LH from plasma after infusion was significantly prolonged than after a bolus injection. It is concluded that the difference in circulating half-lives of FSH between pituitary extracts from intact and OVEX ewes after bolus administration is due to a difference in pituitary FSH composition. However, the prolonged clearance with infusion compared to bolus administration suggests that extrapituitary factors are also responsible.

Effect of restricted feeding on the relationship between hypophysial portal concentrations of growth hormone (GH)-releasing factor and somatostatin, and jugular concentrations of GH in ovariectomized ewes.

These studies characterized the secretion of GH-releasing factor (GRF) and somatostatin (SRIF) into the hypophysial portal circulation in ewes after long term restricted feeding. In addition, we examined the temporal relationship between the concentrations of these two hypothalamic peptides in portal blood and the concentration of GH in jugular blood. Six sheep were fed 1000 g hay/day (normal feeding) and 6 sheep were fed 400-600 g hay/day (restricted feeding). This resulted in a wt loss of 35% in restricted animals compared with 6% in control animals after 20 weeks. Fluctuations in portal levels of GRF indicated a pulsatile pattern of secretion with approximately 60% of pulses coincident with, or immediately preceding, a GH pulse. Similarly, 65% of GH pulses were associated with GRF pulses. Restricted feeding increased (P less than 0.01) mean ( +/- SEM) plasma GH levels (9.8 +/- 1.4 vs. 2.9 +/- 0.6 ng/ml) and mean GH pulse amplitude (7.9 +/- 1.8 vs. 2.8 +/- 0.3 ng/ml) but did not affect mean GH pulse frequency (6.0 +/- 1.1 vs. 5.7 +/- 1.1 pulses/8 h). The level of feeding had no effect on mean portal concentration of GRF (restricted: 5.5 +/- 0.8, normal: 6.6 +/- 1.4 pg/ml), GRF pulse amplitude (14.7 +/- 2.3 vs. 13.5 +/- 0.7 pg/ml), or GRF pulse frequency (5.3 +/- 1.1 vs. 6.7 +/- 0.9 pulses/8 h). Portal concentrations of SRIF in sheep on a restricted diet were half (P less than 0.01) those of sheep fed a normal diet (10.2 +/- 2.3 vs. 19.6 +/- 1.6 pg/ml). Pulses of SRIF were not significantly associated with changes in GH or GRF concentrations. These data indicate a functional role for hypothalamic GRF in initiating GH pulses. Furthermore, the increase in GH secretion in underfed sheep was most probably due to a decrease in the release of SRIF into hypophysial portal blood. Restricted feeding had no affect on GRF secretion, but because of the reduced exposure of the pituitary gland to SRIF, it is possible that responsiveness to GRF is enhanced.

Morphine decreases LH secretion in ovariectomized ewes only after steroid priming and not by direct pituitary action.

To determine whether opiates directly modulate pituitary LH secretion in vivo, morphine was administered to hypothalamo-pituitary-disconnected (HPD) ewes which were receiving exogenous pulses of GnRH. To define the steroidal background which is permissive to a morphine-induced decrease in LH secretion, ovariectomized (OVX) ewes were treated as follows in groups of four: group 1, no implant; group 2, small 17 beta-estradiol (E2) (1 cm long x 0.33 diameter) and progesterone (P) implants; group 3, medium E2 (1 cm long x 0.46 diameter) and P implants, and group 4, medium E2 implants. Jugular blood samples were taken at 10-min intervals for 9 h, during which there was a 3-hour pretreatment period, a 3-hour treatment period when the sheep were given six intravenous injections of 10 mg morphine every 30 min, and a 3-hour run-off period. Morphine inhibited the mean plasma concentrations of LH and LH pulse frequency in group 3 only, and in 2/4 ewes in this group LH secretion was abolished and did not return to a pulsatile mode during the 3-hour run-off sampling period. In a second experiment designed to test the pituitary action of morphine, OVX-HPD ewes were primed with medium E2 and P implants and were given hourly pulses of 250 ng GnRH intravenously. Jugular blood samples were taken around each GnRH pulse over an 8-hour period. The first three pulses served as a control sampling period, after which the sheep were treated with morphine (six intravenous injections of 10 mg morphine every 30 min).(ABSTRACT TRUNCATED AT 250 WORDS)

Analysis of the ratio of biological to immunological LH secreted during the oestrogen-induced LH surge in the ewe.

Plasma concentrations of in-vitro biological and immunological LH were measured throughout the LH surge in cyclic ewes and in ovariectomized ewes treated i.m. with oestradiol benzoate. Both activities increased in parallel during the LH surge in both groups, although the ratio of biological to immunological activities (B/I ratio) was highest at the peak of the LH surge. The two activities were highly correlated (r = 0.86-0.92), with similar slopes from their regression analysis for the cyclic and ovariectomized groups (1.15 and 1.16 respectively). However, the intercepts of the regression lines did not pass through the origin, but intersected the y (radioimmunoassay) axis, suggesting that these serum samples contained immunoactivity not associated with LH bioactivity. In conclusion, an increase in the LH B/I ratio was observed during the LH surge in oestrogen-treated ovariectomized ewes and in cyclic ewes. This increase was not attributable to a change in the relationship between these two LH activities during the LH surge, but rather to the detection of bioinactive immunoactive material in plasma of unknown composition.

Effects of modifying gonadotrophin-releasing hormone input before and after the oestrogen-induced LH surge in ovariectomized ewes with hypothalamo-pituitary disconnection.

The patterns of gonadotrophin-releasing hormone (GnRH) input to the pituitary gland that affect the expression of a positive-feedback event by oestrogen on LH secretion were investigated in ovariectomized ewes with hypothalamo-pituitary disconnection (HPD). In experiment 1, ovariectomized HPD ewes were given hourly i.v. pulses of 250 ng GnRH and an i.m. injection of 50 micrograms oestradiol benzoate (OB). The ewes were given a bolus pulse of 2.25 micrograms GnRH 16 h after injection of OB, followed by half-hourly pulses of 250 ng GnRH for 14 h (treatment A). The LH surge response was significantly (P less than 0.05) greater in these ewes compared with that in ewes given a continuous infusion of GnRH (250 ng/h) after the OB injection, followed by a continuous infusion of 500 ng GnRH/h after the bolus pulse of GnRH (treatment B). When no GnRH was administered after the OB injection, except for the bolus pulse of GnRH (treatment C), the surge response was significantly (P less than 0.05) reduced compared with that in treatment A, and was reduced compared with treatment B. These data suggest that GnRH pulses are important in the generation of the OB-induced LH surge, but that a baseline secretory component can prime the pituitary to some extent. In experiment 2, a doubling of the continuous infusion dose of GnRH used in treatment B to 500 ng/h before the bolus pulse of GnRH and to 1 micrograms/h afterwards (treatment D) gave a similar response compared with treatment A, suggesting that if the baseline input of GnRH is of sufficient magnitude, it can overcome the lack of pulsatile input. In experiment 3, halving the GnRH pulse amplitude used in treatment A from 250 to 125 ng (treatment E) did not reduce the LH surge response, implying that when the GnRH input is in a pulsatile mode, the amplitude of GnRH pulses is less important than the pulsatile nature per se. In experiment 4, removal of GnRH input after the bolus pulse of GnRH (treatment F) significantly (P less than 0.05) reduced the surge response compared with when pulses were maintained (treatment A), indicating that GnRH input is still required once the LH surge has been initiated. Collectively, these experiments show that several forms of GnRH delivery, both pulsatile and baseline, can result in the full expression of a positive-feedback response in ovariectomized ewes treated with oestrogen.

Effect of restricted feeding on the concentrations of growth hormone (GH), gonadotropins, and prolactin (PRL) in plasma, and on the amounts of messenger ribonucleic acid for GH, gonadotropin subunits, and PRL in the pituitary glands of adult ovariectomized ewes.

The effects of long term restricted feeding on the synthesis, storage, and release of GH, LH, FSH, and PRL were examined in adult ovariectomized ewes. Two groups of six ewes were fed a diet of either 1000 g/day (normal feeding) or 400-600 g/day (restricted feeding) hay for 20 weeks. Restricted feeding increased mean plasma GH concentrations and the amplitude of GH pulses, but did not affect GH pulse frequency. In contrast, mean plasma LH and FSH concentrations and LH pulse frequency were decreased by restricted feeding. Mean plasma PRL concentrations were unaffected by treatment. The levels of mRNA for GH in pituitary cytosol were increased by restricted feeding, but no changes were seen in mRNA levels of alpha-subunit, LH beta, FSH beta, or PRL. The pituitary contents of hormones measured did not change with the level of feeding. In conclusion, these data show that long term restricted feeding affects anterior pituitary function in adult ewes, presumably reflecting alterations in the secretion of hypothalamic releasing and inhibiting factors.

The effect of chronic ethanol on glutamate binding in human and rat brain.

Quantitative autoradiographic techniques demonstrate that chronic alcohol administration causes a decrease in [3H]-glutamate binding to hippocampal N-methyl-D-aspartate (NMDA) receptors. A 14% decrease in [3H]-glutamate binding in the hippocampal CA1 region is seen both in the rat after five days of ethanol administration and in postmortem hippocampal tissues from alcoholics. In the rat, 24 hr ethanol withdrawal values are intermediate between control and alcohol binding levels. There was no significant effect of ethanol on [3H]-glutamate binding in the cortex or caudate.

Direct pituitary inhibition of prolactin secretion by dopamine and noradrenaline in sheep.

The effects of dopamine, noradrenaline and 3,4-dihydroxyphenylacetic acid (DOPAC) on the release of prolactin were examined in ovariectomized ewes. Infusion of dopamine (0.5 or 1 microgram/kg per min for 2 h i.v.) reduced plasma prolactin concentrations in a dose-dependent manner, whereas DOPAC (5 or 10 micrograms/kg per min for 2 h i.v.) had no effect. In a further series of experiments, ovariectomized hypothalamopituitary disconnected ewes were given dopamine or noradrenaline (each at 0.5 or 1 microgram/kg per min for 2 h i.v.), and both amines reduced mean plasma concentrations of prolactin with similar potency in a dose-dependent manner. These effects were blocked by treatment with pimozide and prazosin respectively. During the infusion of dopamine, the peripheral plasma concentrations of DOPAC and 3,4-dihydroxyphenylethyleneglycol (DHPG) were increased (DOPAC, 22 +/- 7 (S.E.M.) to 131 +/- 11 nmol/l; DHPG, 2.9 +/- 0.3 to 6.4 +/- 0.2 nmol/l), but plasma concentrations of dopamine and noradrenaline did not change. Finally, administration of domperidone, a specific dopamine receptor antagonist that does not cross the blood-brain barrier, resulted in a sustained increase in plasma prolactin concentrations in ovariectomized ewes. We conclude that the secretion of prolactin from the pituitary gland is under dual inhibitory regulation by both dopamine and noradrenaline in the sheep.

The posterior pituitary regulates prolactin, but not adrenocorticotropin or gonadotropin, secretion in the sheep.

The aim of this study was to determine the role of the posterior pituitary gland in the control of PRL, LH, FSH, and ACTH secretion in sheep. Posterior pituitary function was removed in ovariectomized ewes by electrical lesioning of the hypothalamo-neurohypophysial tract immediately posterior to the stalk-median eminence (LESION); controls were subjected to sham surgery (SHAM). LESION caused a 2-fold increase in plasma PRL concentrations on days 1-3 after surgery. Thereafter, concentrations gradually declined until they were similar to those in SHAM ewes. There was no change in plasma concentrations of LH, FSH, or ACTH after LESION. Plasma PRL responses to insulin in SHAM ewes were completely abolished, and the plasma PRL response to chlorpromazine was reduced to almost half by LESION. In contrast, audiovisual stress (barking dog) and serotonin challenge caused an immediate release of PRL in both LESION and SHAM ewes, with the amplitude of the responses indistinguishable between groups. LESION had no effect on the plasma ACTH responses to audiovisual stress, insulin, or serotonin. We conclude that the posterior pituitary gland is involved in the regulation of PRL under some circumstances, but not of LH, FSH, or ACTH secretion in the sheep. Accordingly, changes in PRL release after hypothalamopituitary disconnection in this species may reflect a loss of posterior lobe function rather than the removal of hypothalamic inputs. In addition, the PRL response to insulin is dependent on a functional posterior pituitary gland, whereas responses to audiovisual stress and serotonin appear to rely on inputs to the pituitary gland via the median eminence and the long hypothalamo-hypophysial portal blood vessels.

The oestrogen-induced surge of LH requires a 'signal' pattern of gonadotrophin-releasing hormone input to the pituitary gland in the ewe.

Two experiments were conducted with ovariectomized and hypothalamo-pituitary disconnected (HPD) ewes to ascertain the pattern of inputs, to the pituitary gland, of gonadotrophin-releasing hormone (GnRH) necessary for the full expression of an oestrogen-induced LH surge. The standard GnRH replacement to these sheep was to give pulses of 250 ng (i.v.) every 2h; at the onset of experimentation, pulses were given hourly. In experiment 1, groups of sheep (n = 7) were given an i.m. injection of 50 micrograms oestradiol benzoate, and after 10 h the GnRH pulse frequency or pulse amplitude was doubled. Monitoring of plasma LH concentrations showed that a doubling of pulse frequency produced a marked increase in baseline values, whereas a doubling of amplitude had little effect on the LH response. In a second experiment, ovariectomized HPD sheep that had received hourly pulses of GnRH for 16 h after an i.m. injection of oil or 50 micrograms oestradiol benzoate were given either a 'bolus' (2.25 micrograms GnRH) or a 'volley' (500 ng GnRH pulses 10 min apart for 30 min, plus a 500 ng pulse 15 min later). Both groups then received GnRH pulses (250 ng) every 30 min for the next 13 h. Oestrogen enhanced the LH responses to the GnRH treatments, and the amount of LH released was similar in ovariectomized HPD ewes given oestrogen plus bolus or volley GnRH treatments and ovariectomized hypothalamo-pituitary intact ewes given oestrogen. These results suggest that the oestrogen-induced LH surge is initiated by a 'signal' pattern of GnRH secretion from the hypothalamus.

Concentrations of dopamine and noradrenaline in hypophysial portal blood in the sheep and the rat.

The concentrations of dopamine, noradrenaline and their respective primary neuronal metabolites 3,4-dihydroxyphenylacetic acid (DOPAC) and 3,4-dihydroxyphenylethyleneglycol (DHPG) were measured in the hypophysial portal and peripheral plasma of sheep and rats by combined gas chromatography-mass spectrometry. Hypophysial portal and jugular blood samples were taken at 5- to 10-min intervals for 3-7 h from six conscious ovariectomized ewes. Blood was also collected for 30 min under urethane anaesthesia from the cut pituitary stalk from 16 pro-oestrous female and five intact male rats. In ovariectomized ewes, noradrenaline concentrations were higher in hypophysial portal plasma than in peripheral plasma (6.6 +/- 0.8 vs 2.2 +/- 0.4 nmol/l). In contrast, dopamine was undetectable (less than 1 nmol/l) in the portal and peripheral plasma of all ewes. Plasma levels of DOPAC and DHPG in portal and jugular samples were similar. In all pro-oestrous female rats, plasma concentrations of dopamine were higher in portal blood than in jugular blood (8.0 +/- 1.4 vs 4.8 +/- 0.6 nmol/l). Detectable concentrations of dopamine were measured in the portal plasma of two out of five male rats. Noradrenaline concentrations were higher in portal plasma than in peripheral plasma of both female (8.3 +/- 1.7 vs 3.7 +/- 0.6 nmol/l) and male (14.8 +/- 2.7 vs 6.1 +/- 1.2 nmol/l) rats. These data show that noradrenaline, but not dopamine, is secreted into the long portal vessels in sheep. The results suggest that there are species differences in the secretion of hypothalamic dopamine into hypophysial portal blood.

Long-term negative feedback effects of oestrogen and progesterone on the pituitary gland of the long-term ovariectomized ewe.

The effects of long-term treatment with physiological doses of oestradiol or oestradiol plus progesterone on plasma gonadotrophin levels and pituitary content of LH and gonadotrophin-releasing hormone (GnRH) receptors were studied in ovariectomized-hypothalamo-pituitary disconnected ewes given 250 ng pulses of GnRH every 2 h (i.v.). A pilot experiment showed that 3 cm long Silastic implants (s.c.) reduced both LH pulse frequency and pulse amplitude in long-term (greater than 6 months) ovariectomized ewes. The main experiment was conducted over 3 weeks in ovariectomized-hypothalamo-pituitary disconnected ewes that had received pulsatile GnRH replacement for 1 week after pituitary surgery. Group 1 (n = 5) received GnRH pulses alone throughout the study. Group 2 (n = 6) received oestradiol in week 2 and oestradiol plus progesterone in week 3 and in group 3 (n = 6) the steroid treatments were reversed. Oestradiol reduced (P less than 0.05) the mean (+/- S.E.M.) amplitude of LH in pulses in group 2 (from 8.2 +/- 1.6 to 5.0 +/- 0.5 micrograms/l) and group 3 (from 11.6 +/- 1.2 to 9.3 +/- 1.0 micrograms 1): an additional effect of progesterone was seen in group 2 but not group 3. The amplitudes of the LH pulses did not change in the control ewes. Plasma concentrations of FSH were reduced by approximately 50% by the oestradiol treatments with no additional effects of progesterone. There was no effect of steroidal treatment on pituitary content of LH or pituitary levels of GnRH receptors.(ABSTRACT TRUNCATED AT 250 WORDS)

Pharmacodynamics of gonadotropin-releasing hormone. 1. Effects of gonadotropin-releasing hormone pulse contour on pituitary luteinizing hormone secretion in vivo in sheep.

The gonadotropin-releasing hormone (GnRH) waveform arrives at the pituitary gonadotropes via the pituitary portal blood and provides the immediate suprapituitary stimulus to luteinizing hormone (LH) secretion. Despite their importance, nature and influence of the physiological GnRH waveform in vivo have been difficult to study. Recent pharmacological and in vitro studies have suggested the potential importance of the wave contour as a specific and independent factor in the pharmacodynamic effects of GnRH on pituitary gonadotrope LH secretion in vivo, and it has been hypothesized that the steepness of the rising edge of the GnRH wave contour is a specific determinant of pituitary LH secretion. In order to investigate the pharmacodynamic influence of GnRH pulse wave contour on pituitary LH secretion in vivo, variations in plasma LH responses to alterations in GnRH wave contour were measured in chronic ovariectomized, hypothalamopituitary-disconnected sheep undergoing physiological pulsatile GnRH maintenance regimen at a fixed dose (250 ng/pulse) and frequency (interpulse interval 120 min). Variable wave contours were then generated by administration of the same total GnRH pulse dose over various lengths of time from near-instantaneous bolus to increasing lengths of constant-rate infusion time up to 8 min. This model allowed specific examination of pulse wave contour in the absence of concurrent changes in endogenous GnRH or sex steroid secretion and holding constant GnRH pulse dose, frequency, and route of administration.(ABSTRACT TRUNCATED AT 250 WORDS)

Gonadotropin-releasing hormone associated peptide (GAP) and putative processed GAP peptides do not release luteinizing hormone or follicle-stimulating hormone or inhibit prolactin secretion in the sheep.

A series of experiments was performed to monitor plasma luteinizing hormone (LH), follicle-stimulating hormone (FSH), and prolactin responses to human gonadotropin-releasing hormone (GnRH) associated peptide (GAP) and related peptides. Ovariectomized hypothalamo-pituitary disconnected (HPD) ewes were challenged with injections (1-10 micrograms i.v.) of GAP, or given, with and without estradiol, hourly 500- or 1,000-ng pulses of GAP for 5-7 days. In all cases GAP failed to cause the release of LH or FSH from the pituitary gland or to alter mean plasma prolactin concentrations. When the same HPD ewes were given hourly or 2-hourly pulses of 250 ng GnRH, LH responded in a a pulsatile manner, and FSH secretion was maintained, thus confirming the functional integrity of the pituitary gland after HPD. Fragments of the GAP molecule (pro-GnRH 14-36, 28-36, 38-49, and 51-66) and GAP dimer did not stimulate LH or FSH or inhibit prolactin release in HPD ewes. GAP and GAP dimer did not affect pituitary responsiveness to GnRH administration. GAP also failed to inhibit the thyrotropin-releasing hormone-induced rise in prolactin. Finally, GAP injections (100 micrograms i.v.) given to lactating ewes did not cause any change in plasma prolactin concentrations. These data show that human GAP, GAP dimer, or putative processed GAP peptides do not act on the sheep pituitary gland in a variety of physiological states to regulate gonadotropin or prolactin secretion.

Pituitary receptors for gonadotropin-releasing hormone in relation to changes in pituitary and plasma gonadotropins in ovariectomized hypothalamo/pituitary-disconnected ewes. II. A marked rise in receptor number during the acute feedback effects of estradiol.

Ovariectomized (OVX), hypothalamo/pituitary-disconnected (HPD) ewes were used to ascertain the short-term effects of estradiol on the number of gonadotropin-releasing hormone (GnRH) receptors in the pituitary gland. The time course of the study was such that measurements were made during the period of short-term negative feedback and positive feedback. Groups of 4 OVX-HPD ewes were given 250-ng pulses of GnRH each hour and an i.m. injection of oil (Group 1) or 50 micrograms estradiol benzoate in oil (Groups 2-4). Blood samples were collected from each ewe prior to treatment with estradiol or oil and again immediately before slaughter. Groups 2, 3, and 4 were killed 6, 16, and 20 h, respectively, after administration of estradiol. Amplitudes of luteinizing hormone (LH) pulses and average plasma concentrations of LH were reduced 6 h after estradiol treatment. Sixteen and 20 h after injection, the average plasma LH levels were elevated, but pulse amplitudes were similar to preinjection values. The number of GnRH receptors was significantly (p less than 0.01) increased within 6 h of estrogen treatment and further increased 16 and 20 h after treatment. Pituitary content of LH was similar in all groups. These data indicate that the number of GnRH receptors in the pituitary gland of ewes can be acutely influenced by a direct effect of estradiol. However, the magnitude and direction of the change in receptors number does not account for the changes in pituitary responsiveness to GnRH, suggesting estradiol also modifies post-receptor mechanisms that influence secretion of LH.

Effect and site of action of hypothalamic neuropeptides on prolactin release in sheep.

In an attempt to identify a physiological prolactin-releasing factor in the sheep, ovariectomized ewes were given intracarotid injections (10(-8)-10(-7) mol/animal) of thyrotropin-releasing hormone (TRH), vasoactive intestinal polypeptide (VIP), peptide histidine-isoleucine amide (PHI), oxytocin (OT), arginine vasopressin (AVP), substance P (SP), bombesin (BB), neurotensin (NT) and neuropeptide Y (NPY). Administration of TRH, AVP, NT and OT resulted in immediate and significant increases in plasma prolactin concentrations, the greatest stimulatory effect being obtained after TRH; other peptides had no effect in ovariectomized hypothalamo-pituitary intact ewes. AVP, NT and OT failed to release prolactin in ovariectomized ewes. These results suggest that (1) AVP, NT and OT may act via the hypothalamus to regulate prolactin secretion in hypothalamo-pituitary intact ewes; (2) VIP, PHI, SP, BB and NPY appear to have no direct roles at the pituitary level to control prolactin secretion in sheep, and (3) TRH stimulates prolactin secretion in ovariectomized ewes by a direct pituitary action.