bluuebunny/crossref_metadata_2025 huggingface dataset:
Autoradiographical localization of oxytocin-binding sites in the guinea-pig ovary at different stages of the oestrous cycle
Authors: Zhang L. , Dreifuss J. J. , Dubois-Dauphin M. , Tribollet E.
Published: December 2008

The discovery that oxytocin is synthesized and stored in corpora lutea of ruminants has fostered a renewed interest in the possible roles of oxytocin in ovarian function. In the present study we describe the distribution of binding sites for oxytocin in the guinea-pig ovary. Sections were reacted with a radioiodinated oxytocin antagonist (125I-labelled OTA) to yield autoradiograms on film. Specific binding sites for oxytocin were defined as those which bound 0·05 nmol 125I-labelled OTA/l and where 1 μmol non-radioactive oxytocin/1 displaced the radioactivity. 125I-Labelled OTA consistently labelled the ovarian stroma and the theca interna, but not the corpora lutea, the granulosa cells or the theca externa. The amount of 125I-labelled OTA bound to ovarian stroma and theca interna was high in animals killed during dioestrus, and low during and shortly after oestrus. These data suggest that the binding sites are regulated by steroid hormone levels and that in the guinea-pig ovary oxytocin could exert a role in follicular steroidogenesis, maturation or ovulation rather than in luteal function. Oxytocin-binding sites were also shown in the uterus but their numbers varied only slightly during the cycle.

Journal of Endocrinology (1991) 131, 421–426
Steroid control of uterine motility via γ-aminobutyric acidA receptors in the rabbit: a novel mechanism?
Authors: Majewska M. D. , Vaupel D. B.
Published: December 2008

γ-Aminobutyric acid (GABA) is a regulator of uterine motility. Stimulation of GABAA receptors tonically inhibited contractions of rabbit uterine strips, while stimulation of GABAB receptors enhanced contractions. Steroids appeared to interact with GABAA receptors to modulate uterine contractility: tetrahydroprogesterone (THP) inhibited while pregnenolone sulphate (PS) increased contractions. THP rapidly antagonized the stimulatory effect of PS, but progesterone inhibited the contractions after a delay, suggesting that the known 'silencing' actions of progesterone on the uterus could be mediated via the metabolite THP, which potentiates the inhibitory function of GABAA receptors. This novel mechanisms may play a role in uterine function during pregnancy and parturition.

Journal of Endocrinology (1991) 131, 427–434
Steroidogenic enzyme activity in the rat testis following Leydig cell destruction by ethylene-1,2-dimethanesulphonate and during subsequent Leydig cell regeneration
Authors: O'Shaughnessy P. J. , Murphy L.
Published: December 2008

Ethylene-1,2-dimethanesulphonate (EDS) rapidly destroys Leydig cells in the rat testis, although repopulation occurs within 5–7 weeks. In this study we have examined the activity of testicular steroidogenic enzymes after Leydig cell destruction and during regeneration. This was designed to measure the contribution of cells, other than Leydig cells, to steroidogenic activity in the testis, and to determine whether changes in steroidogenic enzyme activity during Leydig cell regeneration after EDS parallel those which occur during normal Leydig cell development. The enzymes studied are those responsible for androgen synthesis and metabolism in the testis.

Adult male Wistar rats (300–350 g) were injected with EDS (100 mg/kg, i.p.) and testicular steroidogenic enzyme activity was measured on days 0, 3, 7, 14, 21 and 35. On day 3, when no Leydig cells remain in the testis, cholesterol side-chain cleavage (CSCC) activity, per testis, declined to undetectable levels, while 3βhydroxysteroid dehydrogenase (3βHSD) and 17α-hydroxylase retained only 0·04 and 0·15% of control activity respectively. In contrast, 17-ketosteroid reductase (17-KSR) and 5α-reductase retained 33 and 10% of control activity respectively. On day 7 there was a further loss of 17-KSR activity (to 20% of control) but no change in other enzymes. The 17-KSR activity remaining on day 7 after EDS was contained almost exclusively in the seminiferous tubules, while the low 3β-HSD activity remaining was confined largely to the interstitial tissue. Other enzymes showed a more even distribution between the two compartments.

On day 14 after EDS there was a tenfold increase in 3β-HSD activity (compared with day 7), with no change in CSCC, 17α-hydroxylase or 5α-reductase and a further loss of 17-KSR (to 11% of control). Between days 14 and 21 there were marked increases in the activities of CSCC, 3β-HSD, and 17α-hydroxylase, while 17-KSR showed no change in activity from day 14. Activity of 5α-reductase increased between days 14 and 21 to levels greater than those seen in control animals. By day 35 the activities of all enzymes had returned to control levels.

The results show that, in the adult rat testis, the activities of CSCC, 3β-HSD and 17α-hydroxylase are confined, almost entirely, to the Leydig cells, and only 17-KSR shows significant activity in another cell type. During regeneration of Leydig cells after EDS the pattern of changes in enzyme activity is very similar to that seen in the normal development of the adult population of Leydig cells.

Journal of Endocrinology (1991) 131, 451–457
ADRENALINE STIMULATES CHOLESTEROL SIDE-CHAIN CLEAVAGE CYTOCHROME P450 mRNA ACCUMULATION IN BOVINE ADRENOCORTICAL CELLS
Authors: Ehrhart-Bornstein M. , Bornstein S.R. , Trzeclak W.H. , Usadel H. , Güse-Behling H. , Waterman M.R. , Scherbaum W.A.
Published: December 2008

The effect of adrenaline on the accumulation of mRNA encoding cholesterol side-chain cleavage cytochrome P450 (P450scc) and cortisol secretion was studied in bovine adrenocortical cells in primary culture. Treatment of cultured cells with adrenaline resulted in a 2-fold increase in mRNA encoding P-450scc, as revealed by Northern blot analysis. Under these conditions the maximal stimulation with ACTH resulted in a 6-fold accumulation of mRNA encoding P450scc. The effect of adrenaline on the expression of P450scc was abolished by the ß-blocker propranolol, while propranolol had no effect on ACTH-induced P450scc mRNA accumulation. Adrenaline stimulated the secretion of cortisol in a dose-dependent manner with a median effective dose of 0.5 μmol/l. The adrenaline-stimulated cortisol secretion amounted to 42% of the effect of ACTH (0.1 nmol/l). Upon adrenaline treatment, cAMP concentration in the culture medium increased about 50-fold over the basal value. It is concluded that the stimulatory action of adrenaline upon cortisol formation requires ß-adrenergic receptors and is due, at least in part, to a cAMP-mediated increase in the accumulation of mRNA encoding P450scc.
Seasonal changes in plasma androgens, glucocorticoids and glucocorticoid-binding proteins in the marsupial sugar glider Petaurus breviceps
Authors: Bradley A. J. , Stoddart D. M.
Published: December 2008

An investigation spanning two breeding seasons was carried out to examine endocrine changes associated with reproduction in a wild population of the marsupial sugar glider Petaurus breviceps, a small arboreal gliding possum.

Using techniques of equilibrium dialysis and polyacrylamide gel electrophoresis at steady-state conditions, a high-affinity, low-capacity glucocorticoid-binding protein was demonstrated in the plasma of Petaurus breviceps.

Equilibrium dialysis at 36 °C using cortisol gave a high-affinity binding constant of 95 ± 5·2 litres/μmol for a presumed corticosteroid-binding globulin (CBG) while the binding constant for the cortisol-albumin interaction was 3·5 ± 0·4 litres/mmol.

There was no difference between the sexes in the affinity of binding of cortisol to CBG; however, the cortisol-binding capacity underwent seasonal variation in both sexes. Progesterone was bound strongly to the presumed CBG while neither oestradiol nor aldosterone appeared to be bound with high affinity to P. breviceps plasma.

In the males, peaks in the plasma concentration of testosterone coincided with the July–September breeding season in both years. A significant inverse relationship was shown to exist between the plasma testosterone concentration and the CBG-binding capacity.

In both sexes an increase occurred in the plasma concentration of free cortisol during the first breeding season, a pattern which was not repeated in the subsequent breeding season, possibly due to a lower population density in that year.

Journal of Endocrinology (1992) 132, 21–31
Preferential release of tri-iodothyronine following stimulation by thyrotrophin or thyrotrophin-releasing hormone in sheep of different ages
Authors: Peeters R. , Buys N. , Vanmontfort D. , Van Isterdael J. , Decuypere E. , Kühn E. R.
Published: December 2008

The influence of TRH and TSH injections on plasma concentrations of tri-iodothyronine (T3) and thyroxine (T4) was investigated in neonatal (injection within 0·5 h after delivery) and growing lambs and in normal, pregnant and lactating adult ewes (all 2 years old and originating from Suffolk, Milksheep and Texal cross-breeds). Neonatal lambs had higher levels of T3, T4 and GH compared with all other groups, whereas prolactin and TSH were higher in lactating ewes. In all animals, injections of TRH increased plasma concentrations of prolactin and TSH after 15 min but not of GH at any time. Small increases in T3 and T4 were observed in neonatal lambs, without any effect on the T3 and T4 ratio, after prolactin administration, whereas prolactin did not influence plasma concentrations of T3 or T4 in all other experimental groups. Similar results for thyroid hormones were obtained after TRH or TSH injections. It was therefore concluded that the effects observed after TRH challenge were mediated by the release of TSH. With the possible exception of neonatal lambs, plasma concentrations of T3 after administration of TRH or TSH were always increased before those of T4; the increase in T3 occurred within 0·5–1 h compared with 2–4 h for T4 in all experimental groups. This resulted in an increased ratio of plasma T3 to T4 up to 4 h after injection. It is concluded that, in sheep, TRH and TSH preferentially release T3 from the thyroid gland probably by a stimulatory effect of TSH on the intrathyroidal conversion of T3 to T4.

Journal of Endocrinology (1992) 132, 93–100
Serum half-life and in-vivo actions of recombinant bovine placental lactogen in the dairy cow
Authors: Byatt J. C. , Eppard P. J. , Veenhuizen J. J. , Sorbet R. H. , Buonomo F. C. , Curran D. F. , Collier R. J.
Published: December 2008

The clearance rate of recombinant bovine placental lactogen (rbPL) from the blood serum of four lactating dairy cows was measured using a specific radioimmunoassay. Two animals were non-pregnant, while the other two were at approximately 120 days of gestation. The rbPL was administered as an i.v. bolus injection (4 mg total) via an indwelling jugular catheter. Blood samples were taken periodically for 180 min and assayed for rbPL. Analysis of the clearance curves for the bolus injection suggested a single-compartment model and a serum half-life of 7·25 min. In a second experiment with the same animals, following cessation of lactation, rbPL or bovine GH (bGH) were administered by s.c. injection (50 mg/day) for 5 consecutive days. Blood samples were taken twice per day during the treatment period and a 3-day pretreatment period. Samples were analysed for glucose, blood urea nitrogen (BUN), non-esterified fatty acids (NEFA), creatinine, insulin, insulin-like growth factor-I (IGF-I) and IGF-II, tri-iodothyronine (T3), progesterone and IGF-binding protein-2 (IGFBP-2) to determine whether rbPL mediates similar metabolic effects to those of bGH. Administration of bGH stimulated an increase in NEFA, glucose, T3 and insulin, whereas none of these variables was affected by rbPL. The plasma concentrations of IGF-I and IGF-II were both increased by treatment with rbPL but, to a lesser extent than occurred with bGH. Interestingly, BUN and IGFBP-2 concentrations were reduced equally by bGH and rbPL. These results suggest that rbPL does not necessarily act as a GH agonist but, rather, may have distinct effects on intermediary metabolism that could be mediated through another specific receptor.

Journal of Endocrinology (1992) 132, 185–193
Effects of C19 steroids on adrenal steroidogenic enzyme activities and their mRNA levels in guinea-pig fasciculata-glomerulosa cells in primary culture
Authors: Provencher P. H. , Tremblay Y. , Bélanger A.
Published: December 2008

The present study examined the effects of steroids on steroidogenic enzyme activity in adrenal glands. Guinea-pig fasciculata-glomerulosa (FG) cells maintained in primary culture were exposed to steroids for 48 h. Although the treatment with androstenedione alone had no effect on 3β-hydroxysteroid dehydrogenase 4-ene-5-ene-isomerase (3β-HSD), 17-hydroxylase and 17,20-lyase activities, there was inhibition of 11-hydroxylase and 21-hydroxylase activities. When FG cells were exposed to 10 nmol ACTH/l for the last 24 h of incubation, ACTH alone had no effect on steroidogenic enzymes but, while combined with androstenedione, it further decreased 21-hydroxylase activity and stimulated 17-hydroxylase and 17,20-lyase activities. Cortisol, corticosterone, oestradiol and 11β-hydroxy androstenedione had no effect on steroidogenic enzyme activities while the inhibitory effect on 21-hydroxylase activity was only observed with androstenedione, testosterone and dihydrotestosterone. Addition of hydroxyflutamide, a pure antiandrogen, did not block the inhibitory effect of androstenedione on 21-hydroxylase and 11-hydroxylase activities. The reduction in oxygen tension from 19 to 2% which was aimed at examining the oxygen-mediated effects on steroidogenic enzymes, revealed that the reduction in 21-hydroxylase activity induced by androstenedione could not be prevented by low oxygen tension. An interaction of C19 steroids at the level of the enzymes is also suggested by our finding that androstenedione had no effect on basal and ACTH-stimulated steady-state 11-hydroxylase, 17-hydroxylase, 17,20-lyase and 21-hydroxylase mRNA levels. These results indicate that C19 steroids alter the adrenal steroidogenic enzyme activities in such a manner that C19 steroid synthesis is increased while glucocorticoid production is inhibited. The mechanism of action of C19 steroids does not involve gene expression for steroidogenic enzymes but probably a direct interaction with steroidogenic enzymes, namely 21-hydroxylase, 17-hydroxylase and 17,20-lyase. Our data suggest that C19 steroids may reduce the amount of 21-hydroxylase in the microsomal fraction which may have a major impact on the levels of microsomal P450 reductase available for 17-hydroxylase and 17,20-lyase activities.

Journal of Endocrinology (1992) 132, 269–276
Oxytocin stimulates LH production by the anterior pituitary gland of the rat
Authors: Robinson G. , Evans J. J. , Catt K. J.
Published: December 2008

Gonadotrophin-releasing activity of oxytocin has previously been demonstrated in vitro and in vivo. This study investigated whether oxytocin is also able to induce LH accumulation in pituitary cells. Following trypsin digestion and mechanical dispersion, pituitary cells from female rats were incubated with oxytocin (100 nmol/l) for 24 h. LH release stimulated by oxytocin increased (P < 0·001) progressively during the incubation indicating a different secretory pattern from the more rapid but less sustained secretion stimulated by gonadotrophin-releasing hormone. Oxytocin also enhanced (P < 0·01) total LH accumulation in the incubation system (released plus cell contents) which was apparent after 7–11 h of stimulation. The release of LH stimulated by oxytocin was reduced by the protein synthesis inhibitor cycloheximide (10 μmol/l). However, cycloheximide did not completely block oxytocin-stimulated LH release; there remained some LH release above that seen in non-stimulated controls (P < 0·01) revealing the presence of a cycloheximide-resistant component in the release mechanism. Furthermore, accumulation of total LH in 24 h incubations was suppressed (P < 0·01) by cycloheximide.

The advancement in LH release which oxytocin has been shown to induce in vivo in pro-oestrous rats was accompanied by an early reduction of pituitary LH stores. However, the fall normally observed in LH content during the surge was markedly attenuated by the oxytocin treatment. Thus, loss of pituitary LH stores was less in oxytocin-treated rats than in saline-treated controls, even though net LH release into plasma was increased. Therefore, oxytocin stimulated the replenishment of LH stores.

Although the mechanism(s) remains to be defined and the relationships between in-vitro and in-vivo results are as yet uncharacterized, the present study demonstrates that oxytocin treatment stimulates LH production in both dispersed cells and intact pituitaries in situ.

Journal of Endocrinology (1992) 132, 277–283
Requirement of oestrogens for the sensitivity of prolactin cells to vasoactive intestinal peptide in rats and man
Authors: Pizzi M. , Rubessa S. , Simonazzi E. , Zanagnolo V. , Falsetti L. , Memo M. , Spano P. F.
Published: December 2008

Vasoactive intestinal peptide (VIP) is a prolactin-releasing hormone which is involved in the multifactorial modulation of prolactin secretion in mammals. Intravenous injection of VIP (1 μg/kg) to fertile women increased plasma prolactin levels and heart rate and reduced diastolic pressure. The same treatment to menopausal women caused similar cardiovascular effects but did not modify plasma prolactin levels. In contrast, TRH (200 μg, i.v.) induced a significant increase in plasma prolactin levels in both fertile and menopausal women. The relevance of oestrogens in affecting VIP-stimulated prolactin secretion was evaluated in vitro by measuring prolactin release from pituitary cells of control and ovariectomized rats. The sensitivity of rat mammotrophs to VIP, but not to TRH, was completely suppressed 3 or 4 weeks after ovariectomy. Furthermore, implantation of rats with a silastic capsule containing oestradiol-17β during ovariectomy, preserved the cell responsiveness to VIP. The prolactin-releasing property of VIP was also restored when pituitary cells from ovariectomized rats were cultured for 3 days in the presence of 10 nmol oestradiol-17β/l before being used for prolactin release experiments. The present study shows that the ability of prolactin-secreting cells to respond to the stimulatory action of VIP requires high levels of circulating oestrogens, both in man and rats.

Journal of Endocrinology (1992) 132, 311–316
Effects of deglycosylation of human thyroperoxidase on its enzymatic activity and immunoreactivity
Authors: Giraud A. , Franc J.-L. , Long Y. , Ruf J.
Published: December 2008

Thyroid peroxidase (TPO) is a glycoprotein enzyme which catalyses the iodination of thyroglobulin and the coupling of iodinated tyrosines. Human TPO (hTPO) is the microsomal antigen recognized by the autoantibodies in the serum of patients with autoimmune thyroid disease.

An active detergent-solubilized immunoaffinitypurified hTPO was deglycosylated, either by peptide N-glycosidase F (PNGase F) or by endo-β-N-acetylglucosaminidase H (endo H), and the enzymatic activity and immunoreactivity of the native and degylcosylated forms were compared. Electrophoretic controls and affinoblotting with concanavalin A showed that deglycosylation was not total and that it was more pronounced with endo H than with PNGase F. The enzymatic activity of hTPO was inhibited by endo H deglycosylation, but not by PNGase F deglycosylation; this inhibition was not due to aggregation and/or insolubilization of the molecule subsequent to deglycosylation. Immunoreactivity was monitored by enzyme-linked immunosorbant assay (ELISA) with 13 mouse monoclonal antibodies, rabbit polyclonal antibodies and antibodies from serum of patients with Hashimoto's thyroiditis. In contrast with enzymatic activity, immunoreactivity was not modified or was slightly enhanced (with four monoclonal antibodies) by deglycosylation.

The results indicate that strong, if not total, deglycosylation induces a modification of the tertiary structure of hTPO, which affects the enzymatic site but does not modify markedly the epitopes implicated in the recognition of the molecule by the antibodies tested.

Journal of Endocrinology (1992) 132, 317-323
Parathyroid hormone-related peptide might stimulate calcium secretion into the milk of goats
Authors: Barlet J.-P. , Champredon C. , Coxam V. , Davicco M. J. , Tressol J. C.
Published: December 2008

Parathyroid hormone-related peptide (PTHrP) has been shown to stimulate the placental calcium pump in pregnant ewes. Recent studies also suggest a physiological role for this peptide during lactation. In the present work, we have studied the possible role of synthetic human PTHrP(1–34), (1–86) and (140–173) fragments on calcium, inorganic phosphorus and magnesium secretion into milk of four adult goats in early lactation. This was done by measuring differences in calcium, inorganic phosphorus and magnesium concentrations in the carotid artery and superficial epigastric vein, the concentration of these minerals in milk and milk production. Both PTHrP(1–34) and (1–86) fragments stimulated calcium, inorganic phosphorus and magnesium uptake by the mammary gland and secretion into milk, without any significant effect on milk production. Therefore PTHrP might have important physiological roles in the pregnant and/or lactating female, by regulating calcium transport through the placenta and mammary gland.

Journal of Endocrinology (1992) 132, 353–359
In-vitro and in-vivo responses to chicken LHRH-I and chicken LHRH-II in male turkeys (Meleagris gallopavo)
Authors: Guémené D. , Williams J. B.
Published: December 2008

Stimulation of male turkey hypophyses in vitro with chicken (c)LHRH-I, cLHRH-II or porcine (p)LHRH (0·1 μmol/l) using a perifusion technique caused an increase in the release of LH. In this system, cLHRH-II was approximately 2·5-fold more potent than cLHRH-I and pLHRH which were equipotent. The difference was due to a greater amplitude of the response but not to a prolonged action. Hypophyseal desensitization to a subsequent stimulation was induced when the interval between stimulations was 30 min, but did not occur when lengthened to 60 or 120 min.

Injection of a single dose of cLHRH-I or -II in vivo at doses of 10 and 0·1 nmol/kg body weight stimulated increases in the plasma concentration of LH and testosterone initiated within 1 or 10 min after injection respectively. As in vitro, cLHRH-II induced greater responses, which were dose-related, than did cLHRH-I. However, this difference could be attributed to a greater potency of cLHRH-II and to a more prolonged action. At the 10 nmol/kg dose, the shape of the LH response to cLHRH-II changed; it consisted of an initial increase during 10 min after injection, followed by a more sustained phase during which LH levels were still increasing between 20 and 60 min after injection. In contrast, after an injection of cLHRH-I at doses of 10 or 0·1 nmol/kg or cLHRH-II at a dose of 0·1 nmol/kg, LH levels were at a peak within 5 min and thereafter declined gradually. This decrease in LH may therefore simply be related to the disappearance of the LHRH from the circulation or to other unknown actions of cLHRH-II, when high doses are used.

Journal of Endocrinology (1992) 132, 387–393
Differential dopamine-induced prolactin mRNA levels in various prolactin-secreting cell (sub)populations
Authors: Kazemzadeh M. , Velkeniers B. , Herregodts P. , Collumbien R. , Finné E. , Derde M. P. , Vanhaelst L. , Hooghe-Peters E. L.
Published: December 2008

We have examined the effects of dopamine on prolactin gene expression using quantitative in-situ hybridization histochemistry in different pituitary cell (sub)populations separated according to their density on a discontinuous Percoll gradient. Administration of dopamine resulted in a drastic reduction in hybridization of 35S-labelled DNA probe complementary to prolactin mRNA in total pituitary cells and in lactotrophs with low density. In contrast, dopamine significantly stimulated mRNA accumulation in prolactin-secreting cells with high density compared with other cell layers. The combined use of Percoll gradient and quantitative in-situ hybridization is a valuable and sensitive method with which to examine prolactin-secreting cell response to a given stimulation. Prolactin-secreting cells with high and low density clearly show functional heterogeneity in their response to dopamine.

Journal of Endocrinology (1992) 132, 401–409
Human Recombinant Activin-A Modulates the Steroidogenesis of Cultured Bovine Adrenocortical Cells
Authors: Nishi Y. , Haji M. , Tanaka S. , Yanase T. , Takayanagi R. , Etoh Y. , Nawata H.
Published: December 2008

The effect of human recombinant activin-A on adrenal steroidogenesis was studied in cultured bovine adrenocortical cells. Activin-A significantly reduced cortisol output from ACTH (10nmol/l)-stimulated adrenocortical cells incubated for 24 hours in a dose-dependent manner (10, 100 and 500ng activin-A /ml suppressed cortisol secretion by 19, 33 and 40%), although no significant effect was observed in the case of 3 h incubation. Dehydroepiandrosterone (DHEA) secretion from ACTH-stimulated adrenocortical cells incubated for 24 h was also decreased by the addition of activin-A in a dose-dependent manner. (10, 100 and 500ng activin-A /ml suppressed DHEA secretion by 22, 56 and 58%).

These inhibitory effects of activin-A (100ng/ml) on cortisol and DHEA secretion were partially blocked by the addition of follistatin / FSH-Suppressing Protein (200ng/ml). In contrast, activin-A treatment resulted in no significant decrease in aldosterone secretion. There were no significant effects of activin-A on basal secretions of cortisol, DHEA or aldosterone from adrenocortical cells. These results suggest that activin-A has a direct inhibitory effect on ACTH-stimulated bovine adrenocortical steroidogenesis.
The identification of 125I-labelled iodomelatonin-binding sites in the testes and ovaries of the chicken (Gallus domesticus)
Authors: Ayre E. A. , Yuan H. , Pang S. F.
Published: December 2008

The existence of 125I-labelled iodomelatonin-binding sites in chicken ovaries and testes was investigated. The specific binding of 125I-labelled iodomelatonin to chicken ovarian and testicular tissue satisfies all the criteria for a binding site. It was rapid, stable, saturable, reversible, specific and of high affinity. Equilibrium studies showed that total and non-specific binding increased over a range of 5–150 pmol 125I-labelled iodomelatonin/1 tested, with specific binding reaching saturation towards the middle range of radioligand concentrations. Scatchard analyses indicated a dissociation constant (Kd) of 36·5 ± 5·3 pmol/l (means ± s.e.m.) in the membrane preparations of chicken testes at the middle point in the period of light and a maximum number of binding sites (Bmax) of 0·93 ±0·40 fmol/mg protein (n = 6). In membrane preparations of chicken ovaries, the Kd was 102·2 ± 27·3 pmol/l and the Bmax was 2·77± 0·38 fmol/mg protein (n= 6). Equilibrium and kinetic dissociation constants in the picomolar range indicate high-affinity and physiologically relevant 125I-labelled iodomelatonin-binding sites. Competitive inhibition studies determined the following order of relative potency for inhibition of 125I-labelled iodomelatonin-binding to chicken gonadal membranes: 6-chloromelatonin > melatonin > N-acetylserotonin > > > 5-hydroxytryptamine, tryptamine, 5-methoxytryptophol, l-acetylindole-3-acetic acid, 5-hydroxyindole-3-acetic acid and l-tryptophan. The presence of 125I-labelled iodomelatonin-binding sites suggests a direct pinealgonadal connection in the chicken.

Journal of Endocrinology (1992) 133, 5–11
Opioid modulation of FSH, growth hormone and prolactin secretion in the prepuberal gilt
Authors: Barb C. R. , Kraeling R. R. , Rampacek G. B.
Published: December 2008

The role of endogenous opioid peptides (EOP) in modulating GH, prolactin (PRL) and FSH secretion was evaluated in prepuberal (P) gilts. In experiment I, P gilts received 1 (n = 2), 3 (n = 3) or 6 (n = 3) mg naloxone (NAL)/kg body weight i.v. Blood was collected every 15 min for 2 h prior to and 2 h after NAL and an additional 1 h after 100 μg gonadotrophinreleasing hormone (GnRH) i.v. In experiment II, P and mature (M) gilts were ovariectomized. Three weeks after ovariectomy, P and M gilts were injected twice a day for 10 days with either 0·85 mg progesterone (P4)/kg body weight or oil vehicle (V), resulting in the following groups: PP4 (n=11), PV (n = 10), MP4 (n=11) and MV (n=10). All gilts received 1 mg NAL/kg body weight on the last day of treatment. Blood samples were collected every 15 min for 4 h before and 2 h after NAL and an additional 1 h after 100μg GnRH i.v. In experiment III, six P and five M gilts were ovariectomized and surgically implanted with intracerebroventricular (i.c.v.) cannulae. Blood was collected every 15 min for 3 h before and 3 h after i.c.v. injection of 500 μg morphine in artificial cerebrospinal fluid (CSF) or 250 μl CSF. In experiment I, all doses of NAL failed to alter PRL secretion, while NAL increased (P<0·05) GH secretion in three out of eight gilts. However, NAL suppressed (P <0·05) FSH concentrations. In experiment II, NAL treatment increased (P<0·01) serum PRL concentrations and suppressed (P <0·05) FSH secretion in MP4 gilts. Serum PRL and FSH concentrations were unaltered by NAL in PV, PP4 and MV gilts. In experiment III, mean serum GH, PRL and FSH concentrations were unaffected by CSF injections. Morphine treatment evoked a rapid increase in serum GH and PRL concentrations in both P and M gilts. In contrast, morphine failed to influence FSH secretion in P gilts but did suppress FSH concentrations in M gilts. These data suggest that EOP receptors are functionally coupled to the GH and PRL secretory systems. There is an age-related change in EOP modulation of PRL secretion, while EOP modulation of FSH secretion is an age- and ovarian-dependent process.

Journal of Endocrinology (1992) 133, 13–19
Hypophysectomy prevents the castration-induced increase in porphyrin concentrations in the Harderian glands of the male golden hamster: a possible role for prolactin
Authors: Buzzell G. R. , Hoffman R. A. , Vaughan M. K. , Reiter R. J.
Published: December 2008

The Harderian glands of golden hamsters contain high concentrations of porphyrin pigments, with female hamsters having considerably higher porphyrin concentrations than males. Castration of male hamsters leads to a rapid increase in porphyrin concentrations; testosterone treatment of females has the opposite effect, suggesting a central role for androgens in inhibiting the realization of high porphyrin concentrations by this organ. Previous studies in our laboratories have shown, however, that administration of a dopamine agonist to castrated hamsters prevents the normal increase in Harderian porphyrins from occurring. This suggests that prolactin is necessary for low androgen levels to lead to maximal increases in Harderian porphyrin concentrations. The present study tested the hypothesis that prolactin is involved in the control of Harderian porphyrin levels in the golden hamster.

Although hypophysectomy of male hamsters reduced serum testosterone to levels in castrated hamsters, the resultant increase in Harderian porphyrin concentrations was much less than that seen after a similar period of castration. Furthermore, combining the two procedures (castration and hypophysectomy) also led to a blunted increase in Harderian porphyrin, suggesting that a pituitary hormone is necessary for low testosterone levels to lead to increased porphyrins.

Evidence that this pituitary hormone is prolactin comes from the observations that eliminating all pituitary hormones except prolactin, by severing the connection of the pituitary with the hypothalamus or transplanting the pituitary to a distant site (beneath the kidney capsule) led to greatly augmented Harderian porphyrin levels, in intact or castrated male hamsters.

Hyperprolactinaemia in female hamsters did not counter the effects of testosterone in reducing Harderian porphyrin concentrations, suggesting that, although prolactin is needed for porphyrin production in a low androgen milieu, it does not stimulate porphyrin maintenance when testosterone levels are increased.

Journal of Endocrinology (1992) 133, 29–35
N-Methyl-d-aspartate does not prevent effects of melatonin on the reproductive and thyroid axes of male Syrian hamsters
Authors: Hoover P. A. , Vaughan M. K. , Little J. C. , Reiter R. J.
Published: December 2008

The reproductive and thyroid status of male Syrian hamsters maintained on long days (14 h light, 10 h darkness) were assessed after 10 weeks of daily injections of pharmacological doses of melatonin (25 μg s.c.) and/or N-methyl-dl-aspartic acid (NMDA, 0·025–6 mg i.p.), a compound with receptor sites in the central nervous system which are known to affect reproduction. Melatonin given during the late light phase decreased reproductive organ weights and levels of serum and pituitary prolactin and serum thyroxine (T4); these results are similar to published reports on the effects of chronic short photoperiod treatment of this species. Reproductive organ weights, T4 levels and values for prolactin did not differ significantly between groups receiving only melatonin and those receiving NMDA in addition to melatonin; likewise these variables did not differ significantly between groups receiving only either NMDA or saline. NMDA alone and in combination with melatonin increased serum tri-iodothyronine (T3). The brown adipose tissue enzyme T4 5′-deiodinase demonstrated an increased activity in the presence of NMDA, with the lowest dosage eliciting the most significant effect.

Previous studies have demonstrated that NMDA reverses the reproductive effects of short photoperiod. The results of this study show that NMDA is incapable of preventing the inhibitory reproductive effects of exogenously administered melatonin. These observations are consistent with the proposal for a site of action for NMDA on neural regions more proximal than those altered by melatonin; alternatively, NMDA may interfere with neurotransmitter actions in the pathway controlling melatonin production.

Journal of Endocrinology (1992) 133, 51–58
Intraventricular administration of thyrotrophin-releasing hormone (TRH) suppresses prolactin secretion and synthesis: a possible involvement of dopamine release by TRH from rat hypothalamus
Authors: Ikegami H. , Jikihara H. , Koike K. , Morishige K. , Kurachi H. , Yamamoto N. , Hirota K. , Miyake A. , Tanizawa O.
Published: December 2008

The administration of thyrotrophin-releasing hormone (TRH) causes a variety of dopamine-related biological events. To understand the specific role of TRH on rat hypothalamic dopamine neurones, we examined the in-vivo effects of intraventricular (i.c.v.) infusion of TRH on the release and synthesis of prolactin in the rat pituitary gland and on the changes in binding of [3H]MeTRH and dopamine turnover rates in rat hypothalamus. We have also examined the in-vitro effects of TRH on the release of [3H]dopamine from dispersed tuberoinfundibular dopamine neurones.

Female rats were treated with i.c.v. infusions of 1 μmol TRH/l daily for 1, 3 and 7 days using Alzet osmotic pumps. Following 7 days of treatment the serum prolactin concentrations were significantly decreased. A reduction in hypothalamic TRH-binding sites (Bmax) was also apparent but the dissociation constant (Kd) was unaffected. Northern blot analysis of total RNA isolated from the pituitary glands of control animals using 32P-labelled prolactin cDNA as a probe indicated the presence of three species of prolactin gene transcripts of approximately 3·7, 2·0 and 1·0 kb in size, and these were decreased by TRH treatment. We examined the turnover rate of dopamine in the rat hypothalamus when TRH was administered i.c.v. for 7 days. There was a significant increase in 3,4-dihydroxyphenylacetic acid/dopamine ratio with TRH treatment. Moreover, exposure to TRH stimulated [3H]dopamine release from rat tuberoinfundibular neurones in a time- and dose-dependent manner. Dopamine receptor antagonists such as SCH23390 and (−)sulpiride, and other neuropeptides such as vasoactive intestinal peptide and oxytocin did not affect TRH-stimulated [3H]dopamine release.

These data suggest that i.c.v. administration of TRH might decrease both prolactin secretion and accumulation of prolactin gene transcripts in the pituitary by stimulating dopamine release from tuberoinfundibular neurones.

Journal of Endocrinology (1992) 133, 59–66
Distribution of the nuclear thyroid-hormone receptor in extraocular and skeletal muscles
Authors: Schmidt E. D. , Schmidt E. D. L. , van der Gaag R. , Ganpat R. , Broersma L. , de Boer P. A. J. , Moorman A. F. M. , Lamers W. H. , Wiersinga W. M. , Koornneef L.
Published: December 2008

The correlation between the occurrence of Graves' ophthalmopathy and Graves' hyperthyroidism may indicate a role for tri-iodothyronine (T3) hormone in the pathogenesis of Graves' ophthalmopathy. In Graves' ophthalmopathy the recti eye muscles are greatly enlarged whereas skeletal muscles seem unaffected. The distribution of the nuclear T3 receptor was studied in normal human and rat eye and skeletal muscles with immunohistochemistry using mouse (monoclonal) antibodies, and by in-situ hybridization for the detection of mRNA encoding the T3-receptor protein.

Nuclear staining with T3-receptor antibodies was found in all types of tissues studied. Cytoplasmic staining occurred predominantly in the muscle fibres of the orbital layer of the eye muscles and was generally absent or very low in skeletal muscle fibres and hepatocytes. Immunostaining could be inhibited by preabsorbing the antibodies with bacterially expressed T3-receptor protein, implying specificity. The presence of nuclear and cytoplasmic hormonefree T3 receptor sites was indicated after preincubation of sections with T3 hormone: T3-receptor immunostaining decreased and T3-hormone staining increased. In-situ hybridization clearly revealed the presence of α-1 and β-1 forms of the T3-receptor mRNA in liver, skeletal muscles, and orbital and intermediate layers of the eye muscles.

The data demonstrate the presence of T3 hormone-receptor molecules in the extraocular and skeletal muscles. The different susceptibilities of these muscles to Graves' hyperthyroidism may relate to the quantitative differences in T3 hormone-receptor distribution.

Journal of Endocrinology (1992) 133, 67–74
Effect of food deprivation and altered thyroid status on the hypothalamic-pituitary-thyroid axis in the rat
Authors: Blake N. G. , Johnson M. R. , Eckland D. J. A. , Foster O. J. F. , Lightman S. L.
Published: December 2008

Propylthiouracil (PTU) was administered to rats for different lengths of time with or without food deprivation on the last 2 days. Within 4 days of PTU treatment peripheral 3,5,3′-tri-iodothyronine (T3) fell to low levels and β-subunit of thyroid-stimulating hormnone (β-TSH) mRNA increased significantly in the anterior pituitary. Pro-thyrotrophin-releasing hormone (pro-TRH) mRNA in the hypothalamic paraventricular nucleus (PVN) increased significantly in the control group of animals by 8 days and in the food-deprived group by day 12; the increment of pro-TRH mRNA in the food-deprived group on day 12 was significantly less than that in the control group. In a second study, animals were treated with intraperitoneal injections of T3 with or without the food deprivation. After 4 days of T3 treatment, peripheral T3 levels were markedly increased and pro-TRH mRNA in the PVN and β-TSH mRNA in the anterior pituitary were significantly reduced. Food deprivation had no additional suppressive effect. These studies confirm that the predominant effect of food deprivation on the thyroid axis is at the hypothalamic or suprahypothalamic level and that it can, at least in part, overcome the increase in TRH mRNA due to diminished T3 feedback.

Journal of Endocrinology (1992) 133, 183–188
Testicular 3β-hydroxysteroid dehydrogenase/Δ5–4 isomerase in the hypophysectomized rat: effect of treatment with 5α-dihydrotestosterone
Authors: Fanjul L. F. , Quintana J. , González J. , Santana P. , Estévez F. , Ruiz de Galarreta C. M.
Published: December 2008

The in-vivo regulatory effect of androgens on steroidogenesis was investigated. Adult (2 to 3 months old) hypophysectomized rats were treated intratesticularly with increasing doses of 5α-dihydrotestosterone (DHT; 10–200 μg/100 g body weight) or vehicle (50 μl dimethyl sulphoxide; DMSO) in the contralateral testis. Intratesticular testosterone concentrations were extremely low in hypophysectomized rats 15–20 days after surgery. Treatment with DHT caused a dose-dependent inhibition of testicular 3β-hydroxysteroid dehydrogenase/Δ5–4 isomerase (3β-HSD) 2 h later, and this effect was apparent at the dose of 20 μg/100 g body weight (P < 0·01). The inhibitory effect of 3β-HSD was not due to a possible interference of DHT in the enzyme assay, since various concentrations of the androgen (0·1–100 μmol/l) were ineffective as inhibitors of 3β-HSD. The highest dose of DHT used in this study (200 μg/100 g body weight) resulted in a rapid (1–2 h) and transient (4–6 h) inhibition (approximately 80%) of 3β-HSD activity. Pretreatment of rats with the antiandrogen cyproterone acetate (5 mg/rat) or the protein synthesis inhibitor cycloheximide (10 mg/rat) did not affect the enzyme activity of testes injected with DMSO, but counteracted the inhibitory effect of DHT on 3β-HSD activity in the contralateral testis. The results presented suggest that the inhibitory effect of the non-aromatizable androgen DHT is receptor-mediated and involves the synthesis of a factor(s) that modulates 3β-HSD activity.

Journal of Endocrinology (1992) 133, 237–243
Functional maturation of arginine vasotocin secretory responses to osmotic stimulation in the chick embryo and the newborn chicken
Authors: Klempt M. , Ellendorff F. , Grossmann R.
Published: December 2008

The time-course of appearance of plasma vasotocin and the ontogeny of the vasotocin response to osmotic stimulation during embryonic development was examined in the chicken. In the first experiment, blood was obtained by heart puncture of chicken embryos on embryonic days (E) 15–20, and by decapitation of chickens on the day of hatching (D1 = E21). Plasma was separated for measurement of osmolality and vasotocin. Basal plasma osmolality remained constant until E20 (285–291 mosmol/kg) and was significantly increased at D1 (304± 2 mosmol/kg). Plasma vasotocin was first detectable at E16 (5·5 ± 0·8 (s.e.m.) pmol/l) and increased thereafter with a peak at E18 (21·8 ± 2·3 pmol/l). The concentration decreased again at D1 (9·0 ± 1·7 pmol/l). Osmotic stimulation with hypertonic NaCl 60 min before the animals were killed in the second experiment increased plasma osmolality, at all time points, by 9± 0·2 mosmol/kg in all animals, but increased vasotocin in an age-dependent manner. The vasotocin response was greatest in D1 chickens, and the increase in vasotocin concentration relative to the control group value was 61 pmol/l, whereas the corresponding value in young (E15–E18) and older (E19–E20) embryos was only approximately 9 and 18 pmol/l respectively.

In a third experiment the time-course of vasotocin secretion after osmotic stimuli in catheterized chickens (E18 and D1) was established. The basal plasma osmolality was significantly increased after 15 min in both groups. Vasotocin levels were significantly raised after 15 min in E18 and after 30 min in D1 chickens and further increased thereafter.

These data suggest a relatively late appearance of plasma vasotocin at E16 compared with the early synthesis and storage in the hypothalamus, at E6 and E8 respectively, as reported in the literature. Although an osmotic challenge could trigger vasotocin release as early as E15, a late maturation of release mechanisms might be the case. The secretion mechanisms are far more developed in the hatched chicken compared with the embryo. The lack of correlation between plasma osmolality and vasotocin levels during ontogeny suggests a distinct function of vasotocin other than on fluid transfer during embryonic development.

Journal of Endocrinology (1992) 133, 269–274
Effect of type 1 transforming growth factor-β on the level of aromatase cytochrome P-450 in human fetal hepatocytes
Authors: Rainey W. E. , Price T. M. , Means G. D. , Carr B. R.
Published: December 2008

Aromatase cytochrome P-450 (P-450AROM) is the enzyme in the steroidogenic pathway controlling the formation of oestrogens from 19 carbon steroid precursors. Aromatase is present in various tissues of the human fetus. The liver is second only to the placenta in the level of P-450AROM activity in the fetus. In this study we examined the effects of type 1 transforming growth factor-β (TGFβ) on P-450AROM expression in human fetal (HF) hepatocytes. The HF hepatocytes were dispersed into single cells which were placed into monolayer cell culture until confluent. Cells were then rinsed and treated in serum-free media with dibutyryl cyclic AMP (Bu2cAMP) for 72 h. Treatment with Bu2cAMP (2 mmol/l) caused a fivefold increase in aromatase activity in hepatocytes. The increase in aromatase activity apparently represented an increase in P-450AROM enzyme as determined by immunoblotting using an antibody directed against human placental aromatase. TGFβ blocked basal, as well as Bu2cAMP increases, in aromatase activity by over 50%. The effect of TGFβ was dose-dependent with maximal inhibition observed using 2–5 ng TGFβ/ml. Immunodetectable P-450AROM decreased in parallel with activity following TGFβ treatment. The mechanism of TGFβ action was not through increasing phosphodiesterase (PDE) breakdown of cAMP since inhibition of PDE had no effect on TGFβ action. Finally we examined the level of P-450AROM mRNA using competitive polymerase chain reaction amplification. Bu2cAMP increased mRNA levels of P-450AROM by 2·5-fold, while TGFβ inhibited this induction by 35%. The results of this investigation demonstrated that TGFβ is a potent regulator of P-450AROM expression in HF hepatocytes.

Journal of Endocrinology (1992) 133, 311–320
Somatolactin, a novel pituitary protein: purification and plasma levels during reproductive maturation of coho salmon
Authors: Rand-Weaver M. , Swanson P. , Kawauchi H. , Dickhoff W. W.
Published: December 2008

Somatolactin (SL), a newly discovered fish pituitary protein belonging to the GH/prolactin family, was isolated from coho salmon (Oncorhynchus kisutch). Antibodies were raised to purified coho SL, and a homologous radioimmunoassay was developed and validated. The assay was specific for SL as indicated by the absence of cross-reactivity with coho salmon GH, gonadotrophins I and II and less than 0·2% cross-reaction to prolactin. Serial dilutions of plasma and pituitary extracts from Oncorhynchus species including coho salmon, chinook salmon and rainbow trout were parallel to the coho salmon SL standard curve. Displacement curves for dilutions of Atlantic salmon (Salmo salar) plasma, but not pituitary extract were parallel to the standards. Plasma levels of SL were measured in coho salmon throughout the final year of reproductive maturation. During the period of gonadal growth, plasma SL levels increased and were highly correlated to oestradiol levels in females and 11-ketotestosterone levels in males. Peak levels of SL were observed at the time of final maturation and spawning in both sexes. It is hypothesized that SL may regulate some physiological aspect of reproduction.

Journal of Endocrinology (1992) 133, 393–403