bluuebunny/crossref_metadata_2025 huggingface dataset:
Differential regulation by thyroid hormones of myosin heavy chain α and β mRNAs in the rat ventricular myocardium
Authors: Green N. K. , Franklyn J. A. , Ahlquist J. A. O. , Gammage M. D. , Sheppard M. C.
Published: December 2008

The effect of tri-iodothyronine (T3) treatment on myocardial levels of α and β myosin heavy chain (MHC) mRNAs in the rat was defined in vivo and in vitro. Dose–response experiments were performed in intact hypothyroid and euthyroid rats; in addition, studies in vitro examined the effect of T3 on MHC mRNAs in neonatal cardiac myocytes in primary culture. Specific α and β MHC mRNAs were determined by Northern blot and dot hybridization to oligonucleotide probes complementary to the 3′ untranslated regions of the MHC genes. An increase in myocardial β MHC mRNA was demonstrated in hypothyroidism, accompanied by a reduction in α MHC mRNA. Marked differences in the sensitivity of α and β MHC mRNAs to T3 replacement were found; a dose-dependent increase in α mRNA was evident at 6 h after T3 treatment, in the absence of consistent effects on β mRNA, whereas 72 h after T3 replacement was commenced, stimulatory effects of T3 on α MHC mRNA, evident at all doses, were accompanied by a dose-dependent inhibition of β MHC mRNA. No effect of thyroid status on actin mRNA was found, indicating the specificity of MHC gene regulation. T3 treatment of cardiac myocytes in vitro exerted similar actions on MHC mRNAs to those found in vivo, with a more marked influence on α than β MHC mRNA. These studies of the action of T3 in vivo and in vitro have thus demonstrated specific effects of T3 on pretranslational regulation of the α and β MHC genes, influences which differ not only in terms of stimulation or inhibition, but also in magnitude of effect.

Journal of Endocrinology (1989) 122, 193–200
The hypothalamus and photoperiodic control of FSH secretion by melatonin in the male Syrian hamster
Authors: Bonnefond C. , Walker A. P. , Stutz J. A. , Maywood E. , Juss T. S. , Herbert J. , Hastings M. H.
Published: December 2008

In the photoinhibited castrated male Syrian hamster, removal of the pineal gland or transfer to long photoperiods was followed by a rapid increase in the serum concentration of FSH. Levels were significantly above those of controls within 10 days. Central passive immunization of pineal-intact photoinhibited castrated animals against melatonin had a stimulatory effect on serum FSH levels, comparable with that observed following pinealectomy or transfer to short days. The effects of pinealectomy were blocked by programmed s.c. infusions of melatonin in a time-dependent manner. Serum FSH levels remained low in animals receiving 100 ng melatonin delivered over 10 h but the same mass of melatonin delivered over 4 h had no effect on the response to pinealectomy. Lesions of the anterior hypothalamus had no effect on the pinealectomy-induced increase of serum FSH in animals receiving saline infusions. However, in lesioned animals, programmed infusions of melatonin were no longer able to suppress the rise in FSH following pinealectomy. These results demonstrated that pineal melatonin is the mediator of central photoperiodic control of FSH secretion. The duration of the melatonin signal determines its effectiveness and an intact anterior hypothalamus is necessary for the signal to be read and/or the appropriate neuroendocrine response expressed.

Journal of Endocrinology (1989) 122, 247–254
Photoperiodic control of the development of the LHRH neurosecretory system of European starlings (Sturnus vulgaris) during puberty and the onset of photorefractoriness
Authors: Goldsmith A. R. , Ivings W. E. , Pearce-Kelly A. S. , Parry D. M. , Plowman G. , Nicholls T. J. , Follett B. K.
Published: December 2008

The development of the reproductive system was studied in juvenile starlings during the acquisition of photosensitivity, the attainment of sexual maturation after photostimulation and the subsequent onset of photorefractoriness, using immunohistochemistry for LHRH and radioimmunoassay measurements of hypothalamic, pituitary and plasma hormone concentrations. The first stage of sexual development induced by exposure of photorefractory immature starlings to short days (8 h light:16 h darkness; 8L:16D) was characterized by a decrease in pituitary prolactin content within 1 week and an increase in hypothalamic LHRH content, in the size of the LHRH perikarya and in the intensity of immunostaining in the median eminence in 4–6 weeks. Sexual maturation occurring after exposure to long days (18L:6D) was associated with further increases in LHRH content and cell size, and increases in LH and prolactin concentrations. During testicular regression, LHRH perikarya were reduced in size and staining intensity but LHRH immunostaining in the median eminence and content in the hypothalamus remained high until gonadal regression was almost complete. Prolactin levels were maximal during testicular regression. These results suggest that gonadal regression is initiated by a reduction in LHRH synthesis and possibly, in addition, an external inhibitory influence on LHRH release. Hypothalamic LHRH content eventually declined and LHRH immunostaining in the median eminence was much reduced in fully photorefractory starlings maintained under long days.

Journal of Endocrinology (1989) 122, 255–268
Purified FSH stimulates production of inhibin by the human ovary
Authors: Buckler H. M. , Healy D. L. , Burger H. G.
Published: December 2008

Ovarian inhibin production is stimulated by the administration of human menopausal gonadotrophins or following a rise in endogenous LH and FSH. In order to determine whether FSH specifically stimulates inhibin secretion in vivo, immunoassayable serum inhibin levels were measured following the administration of a highly purified preparation of urinary FSH free of significant contamination with LH. Ten anovulatory women underwent a protocol of induction of ovulation with purified FSH and human chorionic gonadotrophin (hCG). During the induction of ovulation, blood samples were taken for radioimmunoassay of FSH, LH, oestradiol, progesterone and inhibin. During the administration of FSH there were increases in plasma concentrations of FSH, oestradiol and inhibin (P < 0·01) but no significant change in the concentration of LH. Oestradiol and inhibin concentrations rose in parallel and were closely correlated (τ = 0·920, n = 110, P < 0·001). There was also a direct correlation between the measured level of FSH and inhibin (τ = 0·512, n = 110, P < 0·05), but there was no correlation between LH and oestradiol, inhibin or FSH. Inhibin (τ− = 0·702, n = 10, P < 0·01) and oestradiol (τ− = 0·691, n = 10, P < 0·01) were correlated with the number of follicles seen on ovarian ultrasound. Levels of oestradiol and inhibin reached a peak on the day of hCG administration or on the following day. Inhibin levels then fell over the next 2 days in all cycles. In an ovulatory cycle resulting in conception, inhibin and oestradiol then rose in parallel with progesterone. We conclude that inhibin appears to be a follicular product which, in the follicular phase of the cycle, is stimulated by FSH alone, with granulosa cells being the probable site of production.

Journal of Endocrinology (1989) 122, 279–285
Ovine placental steroid 17α-hydroxylase/C-17,20-lyase, aromatase and sulphatase in dexamethasone-induced and natural parturition
Authors: Mason J. I. , France J. T. , Magness R. R. , Murry B. A. , Rosenfeld C. R.
Published: December 2008

Parturition in the sheep is preceded by an abrupt alteration in placental steroid metabolism causing a shift from progesterone to oestrogen production. This change is believed to be a consequence of the prepartum rise in cortisol in the fetal circulation and involves increases in activities of the enzymes steroid 17α-hydroxylase (cytochrome P-45017α), steroid C-17,20-lyase, and possibly aromatase and steroid sulphatase. The activity levels have been determined of steroid 17α-hydroxylase, aromatase and steroid sulphatase in placental microsomes in late pregnancy, dexamethasone-induced labour and in natural labour at term. Over the gestational period of 118-140 days, basal levels of placental aromatase were relatively constant (mean value (±s.e.m.) of 5·6 ± 0·5 pmol/min per mg microsomal protein (n = 10)). Pregnenolone and progesterone 17α-hydroxylase activities were undetectable (< 0·5 pmol/min per mg microsomal protein (n = 7)). In six animals in labour induced with infusion of dexamethasone into the fetus, placental aromatase activity increased to a value of 14·0± 1·0 pmol/min per mg protein; placental pregnenolone 17α-hydroxylase, measured in four of the animals, also increased to 453 ± 77 pmol/min per mg microsomal protein. In five animals in natural spontaneous labour with vaginal delivery, aromatase activity was 26·7 ± 5·2 pmol/min per mg microsomal protein and pregnenolone 17α-hydroxylase activity was 141 ±14 pmol/min per mg microsomal protein. Steroid sulphatase activity was barely detectable (< 1·5 pmol/min per mg microsomal protein) during late pregnancy, dexamethasoneinduced labour or natural parturition. Immunoblotting of placental microsomal preparations with a specific antibody to cytochrome P-450 17α indicated that the glucocorticoid-induced activity of steroid 17α-hydroxylase was associated with an increased content of cytochrome P-45017α. The cotyledon can then convert pregnenolone to dehydroepiandrosterone and thereby generate aromatizable C19 steroids for the aromatase system to facilitate oestrogen synthesis. Importantly, the ovine placental steroid 17α-hydroxylase inefficiently metabolizes 17α-hydroxyprogesterone to androstenedione and thus progesterone is not a principal precursor of oestrogen. The data are consistent with a mechanism of glucocorticoid action in natural ovine parturition to increase cytochrome P-45017α synthesis either by stabilizing the mRNA for cytochrome P-45017α or else acting as a stimulus to transcription of the steroid 17α-hydroxylase gene.

Journal of Endocrinology (1989) 122, 351–359
The use of cells doubly labelled with [14C]inositol and [3H]inositol to search for a hormone-sensitive inositol lipid pool with atypically rapid metabolic turnover
Authors: Maccallum S. H. , Barker C. J. , Hunt P. A. , Wong N. S. , Kirk C. J. , Michell R. H.
Published: December 2008

Some, though not all, previous studies have suggested that the inositol lipid which is hydrolysed during transmembrane signalling in response to receptor activation might be drawn from a metabolically discrete and relatively small hormone-sensitive lipid pool that turns over more rapidly than the bulk of membrane inositol lipid. In order to seek evidence for the existence of this putative hormone-sensitive lipid pool, we have double-labelled cells by growing them for 3 days in a medium containing [14C]inositol and then supplying them with [3H]inositol for the final 2 h before stimulation. We anticipated that stimulation of these doubly labelled cells might provoke the formation, from the postulated hormone-sensitive pool, of small quantities of relatively 3H-enriched inositol phosphates, and that these could be harvested from cells (provided that the cytosolic inositol monophosphatase and inositol 1,4-bisphosphate/inositol 1,3,4-trisphosphate 1-phosphatase activities are first inhibited by Li+). Experiments of this type, using both vasopressin-stimulated WRK1 rat mammary tumour cells and 3T3 mouse fibroblasts stimulated by prostaglandin F2α, have largely failed to demonstrate the formation of relatively 3H-enriched inositol phosphates. There was a tendency for phosphatidyl-inositol 4-phosphate and phosphatidylinositol 4,5-bisphosphate to have slightly higher 3H: 14C ratios than phosphatidylinositol, but the 3H: 14C ratios of the inositol phosphates formed in stimulated cells were not substantially greater than the 3H: 14C ratios of the inositol lipids. We therefore conclude, at least for the two cell lines that we studied, that hormone-stimulated inositol lipid hydrolysis can call, either directly or indirectly, upon the majority of the inositol lipid complement of the stimulated cell.

Journal of Endocrinology (1989) 122, 379–389
Cytosolic free calcium oscillates in single bovine adrenal glomerulosa cells in response to angiotensin II stimulation
Authors: Johnson E. I. M. , Capponi A. M. , Vallotton M. B.
Published: December 2008

The characteristics of the change in cytosolic free Ca2+ concentration ([Ca2+]i) in response to agonist stimulation were studied in individual cells of the bovine adrenal zona glomerulosa.

Following digestion and dispersion, the cells were loaded with the fluorescent Ca2+ indicator fura-2. The cells adhered to Concanavalin A-coated coverslips and were studied using dual excitation wavelength microfluorimetry. In this procedure individual cells under constant perfusion are visualized by microscopy and excited with light alternating rapidly between 340 and 380 nm. The ratio of fluorescence (F) emitted from the cell (F340/F380) correlates directly with [Ca2+]i.

Continuous stimulation with angiotensin II (AII; 10 nmol/l) resulted in a brisk transient rise in [Ca2+]i within 8 s of application of the stimulus. In 50% of cells studied, this initial peak was followed by a series of oscillations in [Ca2+]i lasting up to 13 min, with an average period of 33·0 ± 5·9 (s.e.m.) seconds. [Ca2+]i did not return to prestimulation levels and, subsequent to the oscillatory phase, the [Ca2+]i remained increased for several minutes. Upon removal of extracellular Ca2+ the oscillations ceased almost immediately although [Ca2+]i remained increased. However, in Ca2+-free medium, a single peak of [Ca2+]i still occurred in response to AII. Cells remained refractory to restimulation over a 15-min period.

In contrast, stimulation with K+ (8 mmol/l) rapidly increased [Ca2+]i to a level similar to that induced by AII but without inducing oscillations. Moreover, the effect lasted only while K+ was present and was highly reproducible over multiple stimulations during a 15-min period.

These results corroborate, at the single cell level, the known action of AII of causing release of intracellular Ca2+, but reveal a more complex mechanism of action on Ca2+ influx than previously recognized, possibly invoking a role for a putative second messenger-operated membrane Ca2+ channel.

Journal of Endocrinology (1989) 122, 391–402
Absence of circadian patterns of secretion of melatonin or cortisol in Weddell seals under continuous natural daylight
Authors: Barrell G. K. , Montgomery G. W.
Published: December 2008

Blood samples were obtained from adult female Weddell seals in McMurdo Sound, Antarctica, utilizing four different seals every 2 h for 24 h on two occasions in November 1987. Plasma levels of melatonin were mostly near zero, whereas the values for cortisol were high (2101 ±54 and 1908 ± 108 (s.e.m.) nmol/l, n = 52). For both hormones there was no evidence of a diurnal pattern of secretion. Juvenile seals sampled in December 1985 also had no evidence of a diurnal secretion pattern of melatonin, but blood levels of melatonin were higher than in adults. The results indicate that secretion of melatonin in adult Weddell seals ceases during the Antarctic summer.

Journal of Endocrinology (1989) 122, 445–449
[3H]Angiotensin II binding to basolateral membranes from rat proximal renal tubule: effect of sodium intake and captopril
Authors: Lewis N. P. , Ferguson D. R.
Published: December 2008

Basolateral membranes were prepared from rat renal cortex by density gradient centrifugation. Their purity was confirmed by electron microscopy and by marker enzyme enrichment. The basolateral membrane preparation was shown to be derived predominantly from the proximal renal tubule by measurement of hormone-stimulated adenylate cyclase; marked stimulation of adenylate cyclase was found with parathyroid hormone, but not with isoprenaline, antidiuretic hormone or calcitonin. A single class of specific high-affinity [3H]angiotensin II-binding site was identified in the basolateral membrane preparation which, after correction of results for tracer degradation, showed equilibrium dissociation constant of 0·23 nmol/l and binding site concentration of 485·8 fmol/mg protein.

Binding sites for [3H]angiotensin II were measured in basolateral membranes prepared from rats fed diets with a low, normal or high sodium content. A trend of increased binding site density with reduced sodium intake was found which did not reach statistical significance. No effect on affinity was demonstrated. Treatment of rats on a low-sodium diet with captopril (500 mg/l drinking water) caused a significant reduction in binding site density; no effect on affinity was demonstrated.

These findings suggest that the density of angiotensin II receptors at this site is altered by the activity of the renin-angiotensin system.

Journal of Endocrinology (1989) 122, 499–507
Circadian rhythms of calcitonin gene expression in the rat
Authors: Lausson S. , Segond N. , Milhaud G. , Staub J. F.
Published: December 2008

The daily changes in rat thyroid calcitonin and its specific mRNA concentrations, and the relationship between their dynamics and the plasma levels of calcitonin, calcium and phosphate over a 24-h period were investigated. The circulating calcitonin concentration rose during the daily dark period when plasma calcium and phosphate levels were minimal, indicating that plasma calcitonin rhythm cannot be generated directly by a linear effect of calcium on hormone secretion. Moreover, we established that the expression of the calcitonin gene also exhibited periodic dynamics observable at the pretranslational level: the gland content of hybridizable specific calcitonin RNA displayed daily rhythms; specific RNA levels peaked during the light period and were minimal during the first part of the dark period. Significant changes in thyroid calcitonin concentrations also occurred over a 24-h period. Statistical analyses which distinguished between variations over the 24-h period and residual variations were performed to test the relationships between the various parameters. The daily rhythms of hybridizable RNA, thyroid calcitonin and plasma minerals appeared to be in phase, while the plasma calcitonin concentration displayed variations out of phase with these rhythms. The implication of the correlations observed on the residual variations is discussed in comparison with the temporal relationship between the daily variations. The results fit the hypothesis that hormone production and secretion are self-oscillating processes. Plasma concentrations of calcium and phosphate might play a role in the synchronization of the calcitonin metabolism periodicity.

Journal of Endocrinology (1989) 122, 527–534
Glucocorticoids act rapidly in vitro to attenuate second messenger responses to ACTH secretagogues in rats
Authors: Nicholson S. A. , Gillham B.
Published: December 2008

Fragments of rat anterior pituitary glands incubated in vitro and challenged with either of two ACTH secretagogues were used to investigate the extent to which the acute, biphasic, feedback-like inhibitory effects on hormone secretion exerted by the synthetic glucocorticoid dexamethasone were related to alterations in second messenger responses. Addition of dexamethasone was shown to cause both an immediate inhibition (fast inhibition) of the release of ACTH-like immunoreactivity induced by arginine vasopressin (AVP) or corticotrophin-releasing factor (CRF-41), and also an inhibition that occurred after removal of the steroid and was maximal 90 min after its introduction (early delayed inhibition).

The accumulation of adenosine 3′,5′-monophosphate (cAMP) in the tissue was enhanced in a dose-related manner by CRF-41, as was that of phosphate esters of inositol (inositol phosphates) by AVP. The dose–response curve for the effect of CRF-41 on cAMP production was markedly shifted to the right by dexamethasone acting in the time-domain of fast inhibition (i.e. the response was attenuated, but not abolished). Application of the steroid during the same time-period reduced significantly the inositol phosphate response induced by the higher concentration of AVP tested (3000 mmol/l), but had no effect on the action of a lower concentration (30 mmol/l).

In contrast, the cAMP and inositol phosphate dose–response curves to CRF-41 and AVP respectively were unaffected by the glucocorticoid when it was applied at the time which generated early delayed inhibition of ACTH release.

It is concluded that part of the fast inhibitory action of dexamethasone on ACTH secretion in vitro (but not its delayed effects) involves a rapid alteration of second messenger responses to secretagogues, but the precise mechanism underlying this process remains to be established.

Journal of Endocrinology (1989) 122, 545–551
Studies on the mechanisms of ACTH-induced inhibition of aldosterone biosynthesis in the rat adrenal cortex
Authors: Abayasekara D. R. E. , Vazir H. , Whitehouse B. J. , Price G. M. , Hinson J. P. , Vinson G. P.
Published: December 2008

In rats, chronic treatment with high doses of ACTH (10–40 μg/100 g body weight per day) leads to a marked reduction in aldosterone synthesis by adrenal capsules. The possibility that this inhibition is secondary to a decrease in plasma potassium levels or in renin angiotensin system (RAS) activity has been explored.

The effects of chronic ACTH treatment were compared in (I) animals in which the endogenous RAS activity was stimulated by restricting dietary sodium intake, (II) animals in which plasma angiotensin II was increased by infusion from implanted osmotic minipumps and (III) animals which received supplementary potassium and in which hypokalaemia was prevented. In all cases, rates of aldosterone biosynthesis in vitro by adrenal capsules were decreased in ACTH-treated animals to an extent similar to those in untreated controls. In addition, ACTH treatment of hypophysectomized rats resulted in a similar inhibition of aldosterone biosynthesis to that found in sham-operated controls. It may be concluded that the ACTH-induced reduction of aldosterone biosynthesis is independent of the secretion of other pituitary hormones, and cannot be simply ascribed to either a reduction in RAS activity or in plasma potassium levels. The results are consistent with the view that the effects of chronic ACTH treatment are mediated by a direct action on the zona glomerulosa cell, which leads to its transformation into a zona fasciculata-like form.

Journal of Endocrinology (1989) 122, 625–632
Effects of altered photoperiod or maternal melatonin administration on plasma prolactin concentrations in fetal lambs
Authors: Bassett J. M. , Curtis N. , Hanson C. , Weeding C. M.
Published: December 2008

During winter (December to March), when late-pregnant ewes were maintained under an artificial long-day photoperiod (16 h light) for 3 weeks or more before insertion of fetal vascular cannulae between 118 and 120 days of gestation (full term, 147 days), plasma prolactin concentrations in their fetal lambs were significantly increased throughout the last 3 weeks of gestation in comparison with values in similar aged fetuses from ewes experiencing only the natural short-day (<9 h light) winter photoperiod. When additional lighting was given only after vascular cannulation, fetal plasma prolactin increased steadily from low values, characteristic of winter pregnancies, to high values, characteristic of long-day (16 h light:8 h darkness) pregnancies. Maternal plasma prolactin concentrations changed in a similar way. During summer pregnancies (>16 h light), plasma prolactin in fetal lambs was significantly reduced within 48 h when ewes were given melatonin by i.v. infusion for 14 h each night to simulate the winter duration of the nocturnal increase in plasma melatonin. Maternal plasma prolactin concentrations also decreased significantly when melatonin was given for 3 weeks, but not in a shorter experiment. Increases in fetal plasma prolactin were proportional to the basal prolactin concentration in fetuses injected i.v. with TRH or a dopaminergic antagonist, metoclopramide, to assess how photoperiod influenced the responsiveness of prolactin secretion to acute stimulation. The results confirm that photoperiod, rather than developmental maturity, is the principal determinant of plasma prolactin in the fetal lamb during the last third of gestation, and provide evidence that photoperiodic information is transmitted to the fetus through the diurnal rhythm of melatonin in the ewe.

Journal of Endocrinology (1989) 122, 633–643
Effects of FSH and testosterone on highly purified rat Sertoli cells: inhibin α-subunit mRNA expression and inhibin secretion are enhanced by FSH but not by testosterone
Authors: Toebosch A. M. W. , Robertson D. M. , Klaij I. A. , de Jong F. H. , Grootegoed J. A.
Published: December 2008

The effects of FSH and testosterone on inhibin mRNA expression and inhibin production by highly purified Sertoli cell preparations were examined. Sertoli cells were isolated from testes of 22-day-old rats by sequential trypsin, collagenase and hyaluronidase treatments, with subsequent osmotic shock treatment on day 3 of culture. Contamination by peritubular and germ cells was <0·5 and 1–3% respectively. Intracellular and secreted inhibin levels were measured by radioimmunoassay, using Sertoli cells which were incubated for 24 h in the absence or presence of FSH and testosterone from days 4 to 5 of culture. FSH stimulated the cellular inhibin content and the secreted inhibin level by four- and sevenfold respectively, with a half-maximal effective dose of 5–50 ng/ml. Under the present incubation conditions, testosterone (1 μmol/l) had no effect on immunoreactive inhibin levels in either the presence or absence of FSH. Similarly, the expression of inhibin α-subunit mRNA was increased following FSH stimulation, whereas testosterone had no effect. The expression of inhibin βB-subunit mRNAs was not influenced by FSH or testosterone.

It is concluded that highly purified Sertoli cell preparations, with a very low number of peritubular or germ cells, are fully responsive to FSH with respect to inhibin mRNA expression and inhibin production.

Journal of Endocrinology (1989) 122, 757–762
Regulation by ovarian factors of the LHRH-induced LH response in pituitary glands in situ or grafted under the kidney capsule in intact and ovariectomized rats
Authors: van Dieten J. A. M. J. , de Koning J. , van Rees G. P.
Published: December 2008

When pituitary glands from intact female rats are incubated with LHRH, the resulting LH release shows a biphasic pattern: an initial low rate of LH release (lag phase) is followed by a high rate. When pituitary glands from long-term ovariectomized rats are incubated, the rate of LH release is high throughout stimulation with LHRH. The disappearance of the lag phase might be due to increased LHRH release after ovariectomy and/or the disappearance of ovarian factors.

To distinguish between these possibilities, pituitary glands which had been exposed to endogenous LHRH (pituitary glands in situ) or which had been unexposed to endogenous LHRH (pituitary glands transplanted under the kidney capsule) were incubated in the presence or absence of LHRH.

Biphasic LH secretion patterns were observed during incubation with LHRH with the animal's own pituitary gland and with the transplanted pituitary gland from intact, but not from ovariectomized rats. Thus the disappearance of the lag phase after ovariectomy results from the absence of ovarian secretory products, rather than from increased release of LHRH.

Journal of Endocrinology (1989) 123, 41–45
Suppression of pituitary-testis function in rats treated neonatally with a gonadotrophin-releasing hormone agonist and antagonist: acute and long-term effects
Authors: Kolho K.-L. , Huhtaniemi I.
Published: December 2008

The acute and long-term effects of pituitary-testis suppression with a gonadotrophin-releasing hormone (GnRH) agonist, d-Ser(But)6des-Gly10-GnRH N-ethylamide (buserelin; 0·02, 0·1, 1·0 or 10 mg/kg body weight per day s.c.) or antagonist, N-Ac-d-Nal(2)1,d-p-Cl-Phe2,d-Trp3,d-hArg(Et2)6,d-Ala10-GnRH (RS 68439; 2 mg/kg body weight per day s.c.) were studied in male rats treated on days 1–15 of life. The animals were killed on day 16 (acute effects) or as adults (130–160 days; long-term effects). Acutely, the lowest dose of the agonist decreased pituitary FSH content and testicular LH receptors, but with increasing doses pituitary and serum LH concentrations, intratesticular testosterone content and weights of testes were also suppressed (P< 0·05–0·01). No decrease was found in serum FSH or in weights of accessory sex organs even with the highest dose of the agonist, the latter finding indicating continuing secretion of androgens. The GnRH antagonist treatment suppressed pituitary LH and FSH contents and serum LH (P< 0·05–0·01) but, as with the agonist, serum FSH remained unaltered. Testicular testosterone and testis weights were decreased (P <0·01) but testicular LH receptors remained unchanged. Moreover, the seminal vesicle and ventral prostate weights were reduced, in contrast to the effects of the agonists.

Pituitary LH and FSH contents had recovered in all adult rats treated neonatally with agonist and there was no effect on serum LH and testosterone concentrations or on fertility. In contrast, in adult rats treated neonatally with antagonist, weights of testis and accessory sex organs remained decreased (P <0·01–0·05) but hormone secretion from the pituitary and testis had returned to normal except that serum FSH was increased by 80% (P <0·01). Interestingly, 90% of the antagonist-treated animals were infertile.

It is concluded that treatment with a GnRH agonist during the neonatal period does not have a chronic effect on pituitary-gonadal function. In contrast, GnRH antagonist treatment neonatally permanently inhibits the development of the testis and accessory sex organs and results in infertility. Interestingly, despite the decline of pituitary FSH neonatally, neither of the GnRH analogues was able to suppress serum FSH values and this differs from the concomitant changes in LH and from the effects of similar treatments in adult rats.

Journal of Endocrinology (1989) 123, 83–91
Effects of 1,25-dihydroxyvitamin D3 and cortisol on bovine and human parathyroid cells
Authors: Karmali R. , Farrow S. , Hewison M. , Barker S. , O'Riordan J. L. H.
Published: December 2008

Incubation of bovine parathyroid cells with 1,25-dihydroxyvitamin D3 (1,25-(OH)2D3) decreased both preproparathyroid mRNA levels and parathyroid hormone (PTH) secretion. There was a fall to 56·6 ± 13·7% (mean ± s.e.m.) and 65·1 ± 9·3% in mRNA levels and PTH secretion respectively at 1 nmol 1,25-(OH)2D3/l, and 41·1 ± 13·6% and 42·0 ± 12·1% at 10 nmol 1,25-(OH)2D3/l after 24 h. After 48 h in 0·1 nmol 1,25-(OH)2D3/l, mRNA levels had fallen to 35·3 ± 12·6% and PTH secretion to 32·1 ± 5·0%. In human adenomatous cells, however, incubation with 1,25-(OH)2D3 (10 nmol/l) had no effect on either mRNA levels or PTH secretion even after 48 h. This lack of sensitivity of adenomatous cells to 1,25-(OH)2D3 was not due to an absence of receptors (3847 ± 39 receptors/ng cytosolic protein in adenomatous cells compared with 4068 ± 371 in bovine cells) or receptors being of low affinity.

Cortisol (1 μmol/l) caused a reduction in the number of receptors for 1,25-(OH)2D3 in bovine parathyroid cells of approximately 20% within 24 h of incubation, but no change in affinity. This decrease was accompanied by abolition of the response to 1,25-(OH)2D3 and was reversible, in that withdrawal of cortisol for the final 24 h of incubation was sufficient for the response to return, the number of receptors having returned to control values.

These results suggest that only a small percentage of receptors for 1,25-(OH)2D3 in bovine parathyroid cells may be functional at any one time. Furthermore, the insensitivity of human adenomatous cells to 1,25-(OH)2D3 does not seem to be due to a lack of receptors but may be due to a defect in the interaction between the receptor protein and the PTH gene.

Journal of Endocrinology (1989) 123, 137–142
Specific effect of oestrone sulphate on protein synthesis and secretion by cultured epithelial cells from guinea-pig endometrium
Authors: Chaminadas G. , Alkhalaf M. , Rémy-Martin J. P. , Propper A. Y. , Adessi G. L.
Published: December 2008

Patterns of induced protein synthesis and secretion in guinea-pig endometrial epithelial cell cultures in response to oestrone sulphate alone and oestrone sulphate plus progesterone were investigated. Epithelial cells were cultured for 3 days in growth medium, then washed three times in a steroid-free medium. For each experiment, anticytokeratin immunostaining was used to discriminate the epithelial cells from the stromal cells. Only experiments in which the control dishes displayed more than 80% of anticytokeratinimmunostained cells were further processed. After this period oestradiol-17β (20 nmol/l; control), oestradiol-17β (20 nmol/l) plus progesterone (0·5 μmol/l), oestrone sulphate (1 μmol/l) or oestrone sulphate (1 μmol/l) plus progesterone (0·5 μmol/l) were added to the medium for 48 h. An immunocytochemical progesterone receptor assay showed that oestradiol-17β increased the progesterone receptor content of cells, and progesterone added to cultured cells in the presence of oestradiol-17β induced a significant increase in oestrogen sulphotransferase activity assessing the hormone responsiveness of the cultured cells.

In these culture conditions and after 16 h of incubation, oestradiol-17β induced a 1·7-fold increase in [3H]thymidine incorporation into DNA, and [35S]methionine incorporation into cellular proteins was linearly increased up to 8 h. Biochemical changes induced by the different hormone treatments were studied by labelling the proteins with a 6-h pulse of [35S]methionine. The proteins present in the medium and in cells were analysed by two-dimensional polyacrylamide gel electrophoresis, followed by fluorography. Addition of oestrone sulphate alone or with progesterone produced a change in the patterns of cellular and secreted proteins compared with those in cells cultured with either oestradiol-17β or oestradiol-17β plus progesterone. Three cellular proteins (Mr < 14 000, isoelectric point (pI) 5·2 and 5·3; Mr 75 000, pI 4·9) and one secreted protein (Mr 155 000, pI 5·6–5·9) were specifically induced and could serve as markers of oestrone sulphate action.

Journal of Endocrinology (1989) 123, 233–241
Direct pituitary inhibition of prolactin secretion by dopamine and noradrenaline in sheep
Authors: Thomas G. B. , Cummins J. T. , Doughton B. W. , Griffin N. , Smythe G. A. , Gleeson R. M. , Clarke I. J.
Published: December 2008

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 μg/kg per min for 2 h i.v.) reduced plasma prolactin concentrations in a dose-dependent manner, whereas DOPAC (5 or 10 μg/kg per min for 2 h i.v.) had no effect. In a further series of experiments, ovariectomized hypothalamo-pituitary disconnected ewes were given dopamine or noradrenaline (each at 0·5 or 1 μg/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.

Journal of Endocrinology (1989) 123, 393–402
Osmoregulation of vasopressin secretion in the insulin-withdrawn streptozotocin-treated diabetic rat
Authors: Charlton J. A. , Thompson C. J. , Palmer J. M. , Thornton S. , Baylis P. H.
Published: December 2008

To investigate whether hyperglycaemic ketoacidotic diabetic rats continue to osmoregulate the secretion of arginine vasopressin (AVP), male Wistar rats were injected with streptozotocin (150 mg/kg body weight). Rats rendered diabetic were maintained on protamine–zinc insulin (PZI) for 11 days (insulin-treated rats; n = 35), after which PZI was withdrawn for 72 h in half the rats (insulin-withdrawn rats). Insulin-withdrawn and -treated rats were divided into two groups; one was injected i.p. with distilled water (20 ml/kg) and the other with hypertonic saline (500 mmol NaCl/l; 20 ml/kg), and killed 30 min after injection. Insulin-withdrawn rats (water loaded and osmotically stimulated) were hyperglycaemic (16·5 ± 0·8 and 16·5 ± 0·9 mmol glucose/l respectively) and ketotic (2077 ± 664 and 1474 ± 170 μmol acetoacetate/l respectively). Insulin-treated rats were euglycaemic and non-ketotic. Osmotic manipulation caused similar changes in plasma sodium in both insulin-withdrawn and -treated rats. Plasma AVP was low in the water-loaded rats (0·6 ± 0·1 and 4·5 ± 0·9 pmol/l in the insulin-treated and -withdrawn rats respectively) and increased in rats injected with hypertonic saline (1·2 ± 1·8 and 35·2 ± 17·9 pmol/l respectively). There was no evidence of hypotension and hypovolaemia in any group of rats. Linear regression analysis defined the functions: plasma AVP = 2·56 (plasma Na – 141), r = +0·63, P < 0·01 for hyperglycaemic ketotic rats; plasma AVP = 0·83 (plasma Na – 146), r = +0·78, P < 0·001 for insulin-treated animals. The slopes and abscissal intercepts were significantly (P < 0·05) different. We conclude that the hyperglycaemic ketotic diabetic rat retains the ability to osmoregulate AVP secretion.

Journal of Endocrinology (1989) 123, 413–419
Ontogeny of ACTH(1–24) receptors in rat adrenal glands during the perinatal period
Authors: Chatelain A. , Durand P. , Naaman E. , Dupouy J. P.
Published: December 2008

Binding of ACTH to receptors was studied on crude adrenal membranes from fetal and newborn rats. 125I-Labelled ACTH(1–24) was used as the radioligand, the steroidogenic potency of which was 100-fold lower than that of unlabelled ACTH(1–24). Binding was specific, rapidly equilibrated and temperature dependent. Scatchard analysis of the binding data revealed a single class of binding sites with a dissociation constant of about 100 nmol/l at all stages of development studied. The concentration of ACTH receptors expressed per mg membrane proteins decreased in fetuses between days 17 and 21 of gestation and remained stable in newborn rats from weeks 1 to 4. The number of ACTH receptors expressed per adrenal increased regularly in fetal and newborn rats. The perinatal evolution of these concentrations of ACTH receptors is related to the increase in the size of the adrenals and the changes in cytoplasmic structures of the adrenocortical cells. When the number of ACTH-binding sites was expressed per μg DNA, maximum values occurred in fetuses on day 19 of gestation, and minimum values in newborn rats, 1 week after birth. There was an excellent correlation between the plasma levels of immunoreactive ACTH and corticosterone and the number of ACTH receptors per μg DNA during the perinatal period. Other results suggest that ACTH is able to up-regulate the number of its own receptors.

Journal of Endocrinology (1989) 123, 421–428
Human growth hormone treatment of hypophysectomized rats increases the proportion of type-1 fibres in skeletal muscle
Authors: Ayling C. M. , Moreland B. H. , Zanelli J. M. , Schulster D.
Published: December 2008

The studies describe alterations after hypophysectomy in the proportion of the type-1 and type-2 fibres in rat skeletal muscles, and the effects of replacement treatment with pituitary human (h) GH.

Cytochemical analysis of myosin ATPase, succinate dehydrogenase and lactate dehydrogenase activities in sections of rat hind limb muscles were used as markers of fibre type and revealed that hypophysectomy reduced the proportion of type-1 fibres by 50% in soleus and in extensor digitorum longus muscles. This reduction in the proportion of type-1 fibres was accompanied by the appearance of transitional fibres (type 2C/1B).

Following seven daily injections of hGH (60 mIU/day) to hypophysectomized rats, the proportion of type-1 fibres in both soleus and in extensor digitorum longus was increased with a concomitant reduction in the number of transitional fibres. After 11 days of treatment, all these transitional fibres had reverted back to type-1 fibres. Only hGH was observed to elicit this effect; injections of other pituitary hormones had no effect on the proportions of these transitional fibres.

These alterations in fibre type occurred more rapidly than the changes reported after prolonged electrical stimulation of muscle or following extended exercise.

These findings suggest that hypophysectomy and GH injection can result in a rapid alteration in the fibre composition of skeletal muscle, which may have important implications in terms of the resistance to fatigue and speed of contraction of the muscle.

Journal of Endocrinology (1989) 123, 429–435
Metabolic clearance of insulin-like growth factor-II in sheep
Authors: Hodgkinson S. C. , Davis S. R. , Moore L. G. , Henderson H. V. , Gluckman P. D.
Published: December 2008

The metabolic clearance of ovine insulin-like growth factor-II (IGF-II) was examined in sheep using 131I-labelled IGF-II. Following i.v. administration the tracer was distributed in a volume similar to that of the vascular space (58-5 ±3.3 ml/kg; mean ± s.e.m., n = 5) and demonstrated a triphasic pattern of clearance. Size-exclusion chromatography of a plasma sample collected 1 min after injection revealed peaks of radioactivity corresponding to hormone complexed to binding proteins of 150 and 40–50 kDa (relative abundance 21 and 65% respectively), a high molecular weight binding protein (>200 kDa; 5%) and 'free' tracer (9%). Chromatography of sequential plasma samples revealed different patterns of clearance for these constituents. Half-lives of 131I-labelled IGF-II complexed to the 150 and 40–50 kDa binding proteins, as calculated from rate constants for their decay, were 351 ± 30 and 9.6 ± min respectively (n = 5). These differ markedly from estimates for the clearance of IGF-I (545 ± 25 min, n = 8, and 34 ± 2.3 min, n = 6) associated with carrier proteins of the same apparent molecular weights. This was reflected in calculated metabolic clearance rates for IGF-I (3.9 ± 0.5 ml/min) and IGF-II (7.8 ±1.0 ml/min). Chromatography also revealed that free IGF-II was reduced to negligible levels by 12 min. In contrast, radioactivity eluting in the position expected for the > 200 kDa binding protein was cleared from the circulation very slowly. However, the small proportion of total radioactivity eluting in these molecular weight regions precluded calculation of decay constants for these species. Tracer degradation was monitored throughout the clearance study and estimated to be <20% at 800 min following i.v. administration. Less than 20% of tracer was cleared into urine over the 24 h of sampling, concurrent with a >90% fall in plasma radioactivity. Tracer in urine was completely degraded.

Journal of Endocrinology (1989) 123, 461–468
A role for insulin-like growth factor-I in the regulation of human thyroid cell growth by thyrotrophin
Authors: Ollis C. A. , Hill D. J. , Munro D. S.
Published: December 2008

Human thyroid epithelial cells in monolayer culture were found to release radioimmunoassayable insulinlike growth factor-I (IGF-I) over a 48-h culture period in serum-free medium. In the presence of supraphysiological concentrations of TSH (1–100 mU/ml) known to be inhibitory to DNA synthesis by human thyroid cells, the release of IGF-I was found to be inhibited in six thyroid cultures studied. In only one out of the six was IGF-I release increased in the presence of physiological mitogenic concentrations of TSH (0·1–100 μU/ml). Human thyroid fibroblasts, established by long-term culture of thyroid epithelial cells under fibroblast-selective conditions, also secreted IGF-I which was unaffected by the presence of TSH at both low and high concentrations.

Using a monoclonal antibody against human IGF-I, monolayer cultures of both human thyroid epithelial cells and human thyroid fibroblasts showed positive immunocytochemical staining for IGF peptide. However, fixed sections of intact thyroid tissue only showed positive staining for IGF peptide associated with the fibrous layers surrounding the thyroid follicle, with no staining of the follicular epithelial cells.

The growth of human thyroid epithelial cells was also found to be increased by IGF-I (25–100 ng/ml) added in medium plus 1 % fetal calf serum as assessed by the incorporation of [3H]thymidine into DNA. In the presence of a monoclonal antibody to IGF-I the increase in [3H]thymidine uptake in response to IGF-I was abolished as was that seen in response to TSH.

This study indicates a possible paracrine/autocrine role of IGF-I in the regulation of human thyroid epithelial cell proliferation by interaction with TSH.

Journal of Endocrinology (1989) 123, 495–500
AN ENZYME-LINKED IMMUNOSORBENT ASSAY FOR CORTISOL IN THE SALIVA OF MAN AND DOMESTIC FARM ANIMALS
Authors: Cooper T.R. , Trunkfield H.R. , Zanella A.J. , Booth W.D.
Published: December 2008

An enzyme-linked immunosorbent assay has been developed for the direct assay of cortisol in the saliva of man, sheep, calf and pig. The assay is sensitive (∼1 pg), highly specific and reproducible, and has the advantage over radioimmunosassay in being cheaper and quicker to carry out. The relatively-non invasive method of collecting saliva on cotton buds coupled with simple assay equipment provide a good potential for assessing cortisol status in studies on stress and welfare in farm animals as well as the possibility of monitoring cortisol status in mammals generally.
Pattern of serum concentrations of prolactin and progesterone during pregnancy and lactation in the brown hare (Lepus europaeus)
Authors: Caillol M. , Mondain-Monval M. , McNeilly A. S.
Published: December 2008

A heterologous radioimmunoassay system developed for the rabbit has been shown to measure prolactin in the hare. Concentrations of prolactin showed a significant (P<0·01) increase in the last 3 days of pregnancy (87·7±11·7 compared with 9·8 ± 1·4 (s.e.m.) μg/l; n= 10 and 9 respectively) in pregnant females isolated from males, as well as in pregnant females kept with males and mating prepartum. This rise of prolactin at the end of pregnancy was not due to mating stimuli and occurred at a time when progesterone levels were still high (159 nmol/l). The injection of a slow-release preparation of bromocriptine (5 mg s.c.), which reduces prolactin secretion at the end of pregnancy, did not impair parturition. During lactation, prolactin levels increased significantly (61·2±19·8 compared with 5·3 ± 0·1 μg/l; P<0·01) only after suckling stimuli.

Journal of Endocrinology (1990) 124, 11–17