bluuebunny/crossref_metadata_2025 huggingface dataset:
Influence of blood calcium on calcitonin receptors in isolated chick osteoclasts
Authors: Eliam M. C. , Baslé M. , Bouizar Z. , Bielakoff J. , Moukhtar M. , de Vernejoul M. C.
Published: December 2008

Isolated osteoclasts obtained from young chickens fed a normal (+ Ca) or deficient ( − Ca) calcium and vitamin D diet for 3 weeks, were studied for their ability to bind salmon calcitonin (sCT). Osteoclasts obtained from −Ca chickens, when incubated with 0·1 μmol sCT/l, doubled cyclic (c)AMP production and retracted from a glass support, as observed by scanning electron microscopy. The presence of receptors was also demonstrated by autoradiography and competition analysis of 125I-labelled sCT binding. The number of receptors per cell was 0·9 ± 0·1 × 104. In contrast, osteoclasts obtained from + Ca chickens did not increase cAMP production and did not retract in the presence of 0·1 μmol sCT/l. No specific binding of 125I-labelled sCT could be demonstrated on these osteoclasts.

Plasma levels of calcium and calcitonin were measured in +Ca and − Ca chickens. The plasma concentration of calcium was markedly lower at 3 weeks in −Ca than in +Ca chickens. The plasma concentration of calcitonin was decreased in − Ca chickens compared with +Ca chickens at the first week and kept decreasing during the 3 weeks.

These results strongly support the hypothesis that calcium and vitamin D intake regulate plasma calcitonin levels in chickens, and that calcitonin receptors can be detected on chicken osteoclasts only when blood calcium is decreased by a diet deficient in calcium and vitamin D.

J. Endocr. (1988) 119, 243–248
Testicular steroid metabolism during development in the normal and hypogonadal mouse
Authors: Sheffield J. W. , O'Shaughnessy P. J.
Published: December 2008

The patterns of testicular steroidogenesis were investigated during postnatal development in the normal mouse and in the hypogonadal (hpg) mouse from 20 days. The hpg mouse lacks GnRH and may be used to examine the function of this peptide in normal gonadal development. Testicular tissue was incubated with [3H]pregnenolone and metabolites were separated by thin-layer chromatography and high-performance liquid chromatography. In the normal mouse from 1 to 10 days, metabolism occurred predominantly through the Δ4 pathway, and progesterone, 17α-hydroxyprogesterone, androstenedione and testosterone were the main metabolites formed, together with significant amounts of an unidentified polar steroid. Between 15 and 25 days, androstenedione became the major metabolite formed from pregnenolone. There was also a marked increase in 5α-reductase activity during this age range, and 5α-dihydrotestosterone and 5α-androstane-3α, 17β-diol were significant metabolites. In normal animals older than 30 days, testosterone became the major metabolite, and between 30 days and adulthood the pattern of metabolism changed significantly due to increased formation of intermediates from the Δ5pathway. In the hpg mouse between 20 and 30 days, the pattern of steroid metabolism was unlike that of any age of the normal animal. Progesterone was the major metabolite formed and dehydroisoandrosterone was the major C19 steroid formed, although significant levels of androstenedione and testosterone were also formed. After 30 days there was a marked decrease in steroid metabolism, with androstenedione (the major androgen) being formed mainly through the Δ4 pathway. Addition of human chorionic gonadotrophin to tissue from normal or hpg animals had little effect on the pattern of metabolism, except in the 10- and 20-day-old normal animal where there was an increased accumulation of progesterone.

These results indicate that in the normal mouse the major testicular steroidogenic pathways show significant changes during development. In the hpg mouse the steroidogenic pathways are markedly different from those in normal animals, indicating the importance of the gonadotrophins in controlling the pattern of steroidogenesis. The change in the testicular steroidogenic pattern during development in the hpg mouse may also indicate, however, that these testes are under some stimulation during early postnatal development.

J. Endocr. (1988) 119, 257–264
Validation of a heterologous radioimmunoassay for insulin-like growth factor-I in bovine serum
Authors: Holland M. D. , Hossner K. L. , Niswender G. D. , Elsasser T. H. , Odde K. G.
Published: December 2008

A quantitative, repeatable, heterologous radioimmunoassay (RIA) for insulin-like growth factor-I (IGF-I) was developed for bovine serum. Untreated serum could not be assayed due to interference by IGF-I-binding protein. Serum acidified to pH 3·6 in glycine–HCl (0·1 mol/l) for 24 h at 37 °C and neutralized with NaOH produced inhibition curves non-parallel to the [Thr59]-IGF-I standard. Neutralization by 40-fold dilution of acidified serum samples with assay buffer produced inhibition curves nearly parallel to the IGF-I standard. Complete parallelism was achieved by utilizing preprecipitated normal rabbit serum–sheep anti-rabbit γ-globulin to separate antibody-bound 125I-labelled IGF-I from free 125I-labelled IGF-I. Recovery of IGF-I (1·3–52·3 fmol) added to serum was quantitative. The sensitivity of the RIA (n = 6) was 8·25 ± 0·17 (s.e.m.) fmol. Intra- and interassay coefficients of variation were 3·03 and 4·95% respectively. Serum IGF-I levels measured in beef calves at weaning were positively correlated with weaning weight, total weight gain from birth to weaning and average daily weight gain. In conclusion, a heterologous RIA for IGF-I in beef serum which is sensitive, accurate, precise and repeatable has been developed.

J. Endocr. (1988) 119, 281–285
Electrophysiological correlates of fluid transport in cultured porcine thyroid cells
Authors: Pearson J. , Bourke J. R. , Manley S. W. , Huxham G. J. , Matainaho T. , Gerard C. , Verrier B. , Mauchamp J.
Published: December 2008

Confluent monolayers of cultured porcine thyroid cells transport fluid from the apical to the basal surface, forming circumscribed zones of detachment from the culture dish substrate (domes). The transepithelial potential (TEP), positive on the basal side, was 12·9 ± 0·4 (s.e.m.) mV (n = 93) under control conditions, increasing to 38·9 ± 0·3 mV (n = 281) when fluid transport was stimulated by prostaglandin E2 (PGE2; 1 μmol/l). Forskolin (1 μmol/l) and 8-(4-chlorophenylthio) adenosine 3′,5′-cyclic monophosphate (0·5 mmol/l) were also effective in increasing TEP. Addition of amiloride in concentrations sufficient to block fluid transport (100 μmol/l) reduced the TEP to 5·8 ± 0·3 mV (n=76). Substitution of N-methyl-d-glucamine for sodium in the medium reduced the PGE2-stimulated TEP to 13·4 ± 0·8 mV (n = 32). Substitution of gluconate for chloride increased the TEP to 40·3 ± 0·4 mV (n = 160). Removal of bicarbonate or potassium from the medium, or addition of ouabain (200 μmol/l) were also effective in reducing the TEP. In media of low bicarbonate concentration (1 mmol NaHCO3/l), acetazolamide (1 mmol/l) reduced the TEP. Fluid transport by the monolayer as measured by the change in height of domes was increased by PGE2 (1 μmol/l). PGE2-stimulated fluid transport was inhibited by sodium or chloride ion substitution, bicarbonate removal or the addition of ouabain (200 μmol/l) or amiloride (100 μmol/l). It was concluded that fluid transport in thyroid monolayers is mediated by rheogenic sodium transport with chloride transport being passive, electrogenically coupled to sodium transport. Sodium entry to the apical pole of the cells occurs by an amiloride-sensitive mechanism, and sodium extrusion at the basal pole depends on the Na+/K+ ATPase.

J. Endocr. (1988) 119, 309–314
Plasma clearance in the rat of the LH bioactivity of two human LH standards of differing molecular composition
Authors: Storring P. L. , Khan S. A. , Mistry Y. G. , Diczfalusy E.
Published: December 2008

The LH biological potency of the International Reference Preparation (IRP) of Human Pituitary LH for Immunoassay (IRP 68/40) relative to that of the 2nd IRP of Human Pituitary FSH and LH for Bioassay (IRP 78/549) is markedly greater when estimated by in-vitro interstitial cell testosterone production (TICT) bioassay than by in-vivo bioassay, and by the 4-h ovarian ascorbate depletion (OAAD) assay than by the 4-day seminal vesicle weight gain assay. Other preparations of human LH which, like IRP 68/40, were highly purified, showed a similar spectrum of bioactivity in these assay systems and also contained a higher proportion of more basic LH isoforms than are found in crude pituitary extracts such as IRP 78/549.

In an attempt to explain these differences, a comparison was made of the plasma survival in rats of the LH bioactivity (by TICT assay) of these two preparations. Contrary to expectation, their relative plasma clearance rates over a 4-h period did not account for their differing bioactivities. The plasma half-life of the LH bioactivity (with 95% confidence limits) was estimated to be 42·4 (35·3–49·5) min for IRP 68/40 and 41·3 (31·5–51·0) min for IRP 78/549.

Furthermore the time-course of action in vivo of IRP 78/549 did not appear to be more prolonged than that of IRP 68/40. Thus their plasma testosterone responses during the course of these 4-h plasma clearance studies were similar, and estimates of the LH potency of IRP 68/40 relative to that of IRP 78/549 were no greater by 2-h than by 4-h OAAD assay.

The more basic isoforms of LH present in IRP 68/40 and in other purified human LH preparations may therefore differ from those in crude pituitary extracts, such as IRP 78/549, in their intrinsic activity to stimulate the different target cells in these assay systems rather than in their bioavailability in vivo.

J. Endocr. (1988) 119, 327–334
A sensitive and specific two-site enzyme-immunoassay for human calcitonin using monoclonal antibodies
Authors: Seth R. , Motté P. , Kehely A. , Wimalawansa S. J. , Self C. H. , Bellet D. , Bohuon C. , MacIntyre I.
Published: December 2008

A highly sensitive, specific and rapid two-site enzyme-immunometric assay (EIA) for the measurement of immunoreactive (ir) human calcitonin (hCT) in human plasma was developed using high-affinity monoclonal antibodies. The assay was validated in terms of sensitivity, specificity and reproducibility and its performance compared with that of a radioimmunoassay (RIA) employing a polyclonal antiserum. The sensitivity of the overnight EIA (2 pmol/l) was comparable with the long-incubation (7 days) RIA. The overnight RIA had a sensitivity of 10 pmol/l. The inter- and intra-assay variations of the EIA were less than 12%. Some related and non-related peptides were compared with synthetic hCT for cross-reactivity in the assay and were found to be negative. The mean recovery of added synthetic hCT from plasma of normal volunteers was 96%. Both RIA and EIA have been applied to the measurement of ir-hCT in normal volunteers and in patients with medullary carcinoma of the thyroid. In both groups, the level of ir-hCT measured by EIA was found to be lower than that measured by RIA, presumably due to the ability of the more specific EIA to detect only the 'mature' form of the hormone. EIA offers an attractive alternative to the more cumbersome and lengthy RIA in current usage, with the added advantage of employing a non-isotopic label.

J. Endocr. (1988) 119, 351–357
Effects of thyroxine on oestrogen receptor concentrations in anterior pituitary and hypothalamus of hypothyroid rats
Authors: Altschuler L. R. , Ceppi J. A. , Ritta M. N. , Calandra R. S. , Zaninovich A. A.
Published: December 2008

The effects of thyroxine (T4) were studied on the concentration of oestrogen receptors in the anterior pituitary gland and hypothalamus of ovariectomized euthyroid and hypothyroid rats. A group of rats was made hypothyroid by the administration of I. Seven days after ovariectomy, animals were separated into five groups: I, euthyroid controls; II, hypothyroid controls; III, hypothyroid and injected with oestradiol benzoate (10 μg/day for 10 days); IV, hypothyroid and injected with T4 (4 μg/day for 10 days) and V, hypothyroid and injected with both oestradiol and T4 as described above. In group I, oestrogen receptor levels in pituitary cytosol were 44·4 ± 3·4 (s.d.) fmol/mg protein and in the nucleus 47·7 ± 4·0 fmol/mg DNA. In group II the respective values were 12·8 ± 1·7 fmol/mg protein (P <0·01) and 12·7 ± 1·7 fmol/mg DNA (P <0·01 compared with group I). In group III, cytosolic receptor concentrations decreased when compared with those in group II (P <0·05), whereas nuclear receptor concentrations rose significantly (P <0·01). Group IV had both pituitary cytosolic and nuclear receptors increased (P <0·01 compared with group II). In group V there were no changes in cytosolic receptor concentrations but a significant (P <0·01) rise in nuclear receptors as compared with group II. Hypothalamic oestrogen receptors in untreated hypothyroid rats (group II) were unchanged in the cytosol and diminished (P <0·01) in the nucleus in relation to euthyroid controls (group I). Thyroxine, but not oestrogen, was effective in increasing the concentration of cytosolic receptors (P <0·05). Neither hormone caused changes in nuclei. The results show that there is a pronounced decrease in pituitary and hypothalamic (nuclei) oestrogen receptors in untreated hypothyroid rats and that this decrease can be reversed by T4 treatment.

J. Endocr. (1988) 119, 383–387
In-vitro control of growth hormone secretion by synthetic releasing factors in young and adult ducks (Anas platyrhynchos)
Authors: Foltzer-Jourdainne C. , Harvey S. , Mialhe P.
Published: December 2008

Release of GH from perifused duckling hemipituitaries was stimulated, in a biphasic manner, by synthetic TRH and human pancreatic GH-releasing factor (GRF). At all effective concentrations, the level of GH release was increased within 5 min of TRH or GRF perifusion and was maximal after 10 min of TRH perifusion and after 20 min of GRF perifusion. Although TRH was perifused for 20 min the level of GH release declined during the last 10 min. The most effective dose of TRH (1·0 μg/ml; 2·7 μmol/l) and GRF (0·5 μg/ml; 110 nmol/l) provoked similar (250– 300%) increases in the level of GH release. However, since the effect of TRH was only of short duration, the total release of GH induced by GRF was higher than that elicited by TRH, especially with the low dose. The increase in release of GH induced by TRH or GRF was blunted when pituitaries from adult ducks were used. As in young ducks, the GH response to GRF was higher, whereas the response to TRH was very low. The GH response of perifused adult pituitaries to GRF was, however, potentiated when TRH was perifused simultaneously. The basal release of GH from both young and adult pituitary glands was unaffected by perifusion with somatostatin-14 (SRIF-14) at doses of 1 and 2 μg/ml. The perifusion of hemipituitary glands with similar doses of SRIF-14 was also unable to suppress the stimulation of GH release induced by prior perifusion with GRF, although when SRIF-14 and TRH were simultaneously perifused TRH-induced GH release was markedly suppressed.

These results demonstrate direct effects and interactions of TRH, GRF and SRIF on the release of GH from duck pituitary glands. GRF is the most potent releasing factor for GH in both young and adult ducks although in adult ducks it is less effective. These results also provide evidence that the age-related decline in the in-vivo GH response to TRH is due to a desensitization of pituitary somatotrophs.

J. Endocr. (1988) 119, 421–429
Oestradiol synergizes with 5α-dihydrotestosterone or 3α- but not 3β-androstanediol in inducing sexual behaviour in castrated rats
Authors: Södersten P. , Eneroth P. , Hansson T.
Published: December 2008

Castrated male rats were treated with constant-release implants filled with testosterone, oestradiol-17β, 17β-hydroxy-5α-androstan-3-one (5α-dihydrotestosterone; DHT), 5α-androstane-3α,17β-diol (3α-Adiol) or 5α-androstane-3β,17β-diol (3β-Adiol). Only testosterone activated the sexual behaviour of the rats. If combined with oestradiol, DHT or 3α-Adiol induced the behaviour, but 3β-Adiol failed to have this effect. Oestradiol inhibited the in-vitro formation of [14C]Adiols from [14C]DHT by combined preoptic and hypothalamic tissue, but only when given in high doses. No effect on the formation of [14C]Adiols from [14C]DHT was found in rats treated in vivo with DHT or with the combination of DHT and oestradiol which effectively stimulated sexual behaviour. These results do not support the suggestion that oestradiol may synergize with androgens to induce sexual behaviour in castrated rats by inhibiting androgen metabolism.

J. Endocr. (1988) 119,461–465
Precopulatory behaviour in the male hamster: effect of the 5α-reductase inhibitor 17β-N,N-diethylcarbamoyl-4-aza-5α-androstan-3-one
Authors: Steel E. , Hutchison J. B.
Published: December 2008

Three independent components of hamster masculine behaviour (approaching, leaving and sniffing the female) have been shown to depend on both androgenic and oestrogenic action. The behavioural role of 5α-reduced androgens was assessed by blocking 5α-reduction of testosterone by means of 17β-N,N-diethylcarbamoyl-4-aza-5α-androstan-3-one (4-MA) through slow-release silicone elastomer implants. Three dose levels of 4-MA were given to intact, sexually active males. The probability of approaching the female and the amount of sniffing directed to her were both decreased, while the probability of leaving the female was increased. Sniffing and the tendency to approach were affected at lower dose levels than was the tendency to leave. The higher dosage of 4-MA also abolished odour-based discrimination between females shown by normal males. These effects of 4-MA on behaviour were confirmed in castrated hamsters maintained on testosterone. The suppressive effects of 4-MA on behaviour were reversed by treatment with 5α-dihydrotestosterone (DHT). We conclude that testosterone maintains sociosexual behaviour in part by conversion to its 5α-reduced metabolites; components of this behaviour differ in their relative dependence on DHT. In this species, DHT is likely to be a behaviourally active metabolite of testosterone involved in the fine control of behaviour.

J. Endocr. (1988) 119, 483–491
SECRETION OF BIOACTIVE INHIBIN BY THE OVARY OF THE BOOROOLA MERINO EWE WITH OR WITHOUT A COPY OF THE FECUNDITY (F) GENE.
Authors: Tsonis C. G. , Baird D. T. , Campbell B. K. , Downing J. A. , Scaramuzzi R. J.
Published: December 2008

The secretion rates of bioactive inhibin, oestradiol and progesterone were measured during the mid-luteal phase and at various times during the follicular phase of the cycle by a sensitive bioassay using sheep pituitary cells in culture in 12 Booroola ewes with and without copies of the Fecundity (F) gene in which the left ovary had been auto-transplanted to the neck.

Inhibin secretion was high during the luteal phase and fell in the early follicular phase in all genotypes (P < 0.01). In Booroola ewes with a F/- genotype, inhibin secretion then increased again, towards luteal rates, in the mid and late follicular phases. In Booroola ewes without a copy of the F gene (+/+) inhibin secretion remained low at all three sampling times in the follicular phase. The secretion rate of inhibin at 36 h (P < 0.1) and 48 h (P < 0.01) were significantly lower in ewes from the +/+ (no copy of the gene) ewes than in F/(one copy of the gene) ewes. Oestradiol secretion was low during the luteal phase and increased steadily during the early (24 h) to a plateau in the mid (36 h; P < 0.01) and late (48 h; P < 0.05) follicular phase. Progesterone secretion was high during the luteal phase, and decreased to a very low rate by 24 h after prostaglandin (PG) treatment (P < 0.001) and remained low. At 24 h after PG the concentration of FSH was significantly lower (P < 0.01) than that during the luteal phase and remained suppressed until the onset of the LH surge. There were no significant differences in LH concentrations. We conclude that (1) the secretion of inhibin by the ovary is highest in the luteal phase and (2) inhibin secretion is significantly raised during the mid to late follicular phase in Booroola ewes with a copy of the Fecundity gene compared with those without.
Feedback regulation by progesterone of stress-induced prolactin release in rats
Authors: Deis R. P. , Leguizamon E. , Jahn G. A.
Published: December 2008

We have previously found that modifications to serum progesterone concentration have profound inhibitory effects on prolactin release in response to ether stress. The objective of the present study was to determine the effect of ether stress on progesterone secretion and the role of this steroid in ether-induced prolactin release.

Serum progesterone concentration, 5 min after ether stress had been applied over a 2-min period, was consistently increased in male rats, in cyclic rats on the mornings of pro-oestrus and oestrus, and in androgenized rats in permanent oestrus. Ovariectomized androgenized rats showed the same response. Adrenalectomy of male and female rats abolished the progesterone increase induced by stress. Thus, the progesterone secreted by stressed rats is mostly of adrenal origin.

In groups of male and pro-oestrous rats, circulating concentrations of prolactin and progesterone were measured from 5 to 60 min after stress. In both sexes the serum prolactin concentration was significantly increased at only 5 and 10 min after stress when compared with control values. In pro-oestrous rats the serum progesterone concentration was significantly higher than in controls at 5, 10 and 20 min after stress, whilst in male rats the concentration remained significantly higher at 30 min.

Thirty minutes after the first stress, male and pro-oestrous rats were etherized for 2 min, and bled 5 min after removal from the ether container. In female rats this second stress produced only a slight but significant increase in serum prolactin concentrations, whereas in male rats prolactin concentrations did not increase. The second stress was still capable of significantly increasing circulating progesterone concentrations to levels similar to those obtained after the first stress in animals from all groups. Thus, an increased circulating progesterone concentration did not lead to regulation of further progesterone secretion. To find whether this type of response was due to a blocking effect of the previously released progesterone, animals were injected with the anti-progesterone RU 38486 (17β-hydroxy-11β-(4-dimethylaminophenyl)-17α-propinyl-oestra-4,9-dien-3-one) or with a specific antibody raised against progesterone. In both groups of treated rats the second stress induced a significant increase in serum prolactin and progesterone concentrations to give values similar to those obtained after the first stress. When the second stress was applied to female rats 60 min after the first the prolactin response was comparable to that obtained after the first exposure to ether.

In conclusion, we have demonstrated that serum prolactin and progesterone concentrations are significantly increased after ether stress, and that the latter hormone exerts an inhibitory regulatory feedback on prolactin secretion. These results provide an important new insight into the role of progesterone in the regulation of prolactin release.

Journal of Endocrinology (1989) 120, 37–43
Androgen regulation of major mRNAs for proteins of the immature and adult mouse vas deferens
Authors: Martinez A. , Berger M. , Jean Cl.
Published: December 2008

When total RNA and poly (A)+ RNA extracted from the adult mouse vas deferens were translated in the rabbit reticulocyte lysate protein synthesizing system, they showed a similar electrophoretic pattern of about 20 protein bands of widely varying molecular weights. Within the total mRNA population, three functional mRNAs coding for proteins with molecular weights of 24, 34·5 and 36 kDa showed marked changes after 1 month of castration. The time-course of the response of the translatable proteins showed that while 3 days after castration the 24 and 36 kDa bands were missing, the mRNA coding for the 34·5 kDa band was markedly reduced only 20 days after castration. The negative effect of castration on the three mRNAs could be completely reversed by treatment with testosterone. When total RNA extracted from the immature mouse vas deferens was translated, the 24, 34·5 and 36 kDa protein bands were detectable in 10-day-old males and were synthesized in significant amounts between 10 and 20 days (24 and 36 kDa bands) or between 20 and 30 days (34·5 kDa band). Based on its electrophoretic and immunological properties, the 34·5 kDa protein band, which is predominant in the translation products, was identified as the major androgen-dependent protein previously described in vivo.

Journal of Endocrinology (1989) 120, 67–74
Cortisol stimulates whole body calcium uptake and the branchial calcium pump in freshwater rainbow trout
Authors: Flik G. , Perry S. F.
Published: December 2008

Exposure of rainbow trout to a reduced ambient calcium level (from 490 to 25 μmol Ca2+/l) caused hypocalcaemia and induced a rapid increase (within 1 h) in systemic cortisol levels. Under conditions of low environmental calcium concentrations, cortisol levels remained increased for at least 8 days. After this time the in-vitro Ca2+-transport capacity of branchial basolateral membrane vesicles was increased due to stimulation of Ca2+-ATPase activity, presumably as a result of chloride cell proliferation. Pituitary prolactin cells were unaffected by low ambient calcium levels. Fish kept in water containing 490 μmol Ca2+/l and treated with cortisol for 7 days displayed an increase in whole body calcium uptake and an enhancement of the branchial calcium transport capacity; concomitantly, hypercalcaemia was observed. We conclude that, in the rainbow trout, cortisol exerts hypercalcaemic effects by stimulating Ca2+ uptake from the water and that this effect forms an intrinsic part of the established mineralocorticoid action of cortisol in fish.

Journal of Endocrinology (1989) 120, 75–82
Osteocalcin during the reproductive cycle in normal and diabetic rats
Authors: Verhaeghe J. , Van Herck E. , Van Bree R. , Van Assche F. A. , Bouillon R.
Published: December 2008

Concentrations of osteocalcin were measured in plasma and bone of normal and diabetic rats during the reproductive cycle and compared with plasma 1,25-dihydroxyvitamin D3 (1,25-(OH)2D3) concentrations. The age-dependence of osteocalcin was also examined. Plasma concentrations of osteocalcin levels were low but detectable in 21-day-old fetuses (3·7 ± 0·3 nmol/l); osteocalcin concentrations were highest in weaning rats (104 ± 9 nmol/l) and decreased thereafter. In adult rats, plasma concentrations of both osteocalcin and 1,25-(OH)2D3 increased during the last days of normal pregnancy, and even more so in rats fed a diet low in calcium and phosphate. After an early post-partum decline, osteocalcin concentrations in plasma remained at non-pregnant levels in lactating rats fed a high calcium/phosphate diet while their 1,25-(OH)2D3 concentrations were higher than in non-pregnant rats; however, lactating rats fed a low calcium/phosphate diet showed increasing osteocalcin concentrations.

In spontaneously diabetic BB rats, plasma osteocalcin concentrations were severely decreased compared with those in non-diabetic rats, more than would have been expected from their decreased 1,25-(OH)2D3 concentrations. Moreover, plasma osteocalcin did not increase during pregnancy or lactation in diabetic rats, even when fed a low calcium/phosphate diet. Fetuses of diabetic rats also had lower plasma osteocalcin levels than fetuses from non-diabetic rats or than weight-matched fetuses from semistarved rats. In contrast to plasma osteocalcin concentrations, bone osteocalcin concentrations and content were not altered by pregnancy, lactation, low calcium/phosphate diet or diabetes.

These data show that plasma osteocalcin concentrations change during the reproductive period in normal rats but not in diabetic rats, and that osteocalcin levels do not seem to be regulated solely by 1,25-(OH)2D3.

Journal of Endocrinology (1989) 120, 143–151
Vulnerability of the human Leydig cell to radiation damage is dependent upon age
Authors: Shalet S. M. , Tsatsoulis A. , Whitehead E. , Read G.
Published: December 2008

Testicular function was studied in three groups of patients previously treated for malignant disease, and a control group of adult males. The adult patients in groups one and two underwent unilateral orchidectomy for a testicular tumour but only in group two was this followed by post-operative high-dose irradiation (30 Gy) to the remaining testis. Four of the five boys in group three had a unilateral orchidectomy between the ages of 1 and 4 years and all five received a similar dose of irradiation (27·5–30 Gy) to the scrotum as in group two. The five subjects in group three were studied between the ages of 12 and 34 years.

In group one the median basal testosterone level (16·0 nmol/l) was normal and the basal gonadotrophin levels mildly but significantly increased, reflecting a resetting of the pituitary-testicular axis following unilateral orchidectomy. In group two the median basal testosterone level (12·5 nmol/l) was significantly lower and the median basal FSH and LH levels were significantly higher than the respective values in group one, indicating that irradiation to the testis in adult life may damage both the germinal epithelium and the Leydig cells. All five males in group three showed grossly increased FSH and LH levels, with a median basal testosterone level (< 2·5 nmol/l) significantly lower than groups one and two. None of the five showed a testosterone response to a stimulation test with human chorionic gonadotrophin or underwent puberty spontaneously.

The severe reduction in testosterone levels in group three compared with group two, despite the similarity of radiation dose received, suggests a much greater vulnerability to radiation-induced Leydig cell damage in the prepubertal boy compared with the adult male.

Journal of Endocrinology (1989) 120, 161–165
Multiple forms of rat prolactin and growth hormone in pituitary cell subpopulations separated using a Percoll gradient system: disulphide-bridged dimers and glycosylated variants
Authors: Bollengier F. , Velkeniers B. , Hooghe-Peters E. , Mahler A. , Vanhaelst L.
Published: December 2008

Prolactin and GH cells from rat pituitary glands were separated into three main fractions on discontinuous Percoll gradient layers. SDS-PAGE and subsequent immunoblotting of these fractions revealed that: (1) multiple rat prolactin (rPRL) molecular variants were present in total culture, Percoll layer 1 and 2; four variants were clear-cut: Mr ∼23 000, Mr doublet ∼25 000–26 000, Mr ∼40 000 and Mr ∼42 000; (2) cell cytosol from Percoll gradient layer 1 was particularly enriched in prolactin; (3) cells from gradient layer 1 secreted into the culture medium only prolactin in detectable amounts; (4) three distinct molecular forms of rat growth hormone (rGH) were recorded in layer 3: Mr ∼36 000,24 000 and 20 000; the 20 000 variant was paramount; and (5) cells from layer 3 secreted both rPRL and rGH into the culture medium.

Reduction experiments showed that, on the one hand, 42 000 and 40 000 rPRL variants and, on the other hand, 36 000 rGH variants are disulphide-bridged dimers. An important finding was the presence of glycosylated rPRL and rGH: indeed Concanavalin A–Sepharose 4B affinity chromatography indicated that 26 000 rPRL and 24 000 rGH display a very strong affinity for lectin. Competitive inhibition tests showed that this affinity is specific and not due to hydrophobic binding. When rPRL was submitted to deglycosylation in conditions specific for O-linked glycoproteins, the 26 000 rPRL variant disappeared. The biological role of glycosylated rPRL is as yet unknown.

Journal of Endocrinology (1989) 120, 201–206
Interstitial fluid volume in the rat testis: androgen-dependent regulation by the seminiferous tubules?
Authors: Maddocks S. , Sharpe R. M.
Published: December 2008

Regulation of testicular interstitial fluid (IF) volume has been investigated in adult male rats in which the Leydig cells were selectively destroyed with a single i.p. injection of ethane dimethane sulphonate (EDS). Following this treatment, some animals also received testosterone supplementation by s.c. injection every 3 days, beginning either from the time of EDS injection, or 3–12 days afterwards. The volume of IF obtained by drip collection was determined, and testosterone and gonadotrophin concentrations measured in blood and in IF. Testosterone levels in IF and serum became undetectable by 3 days after EDS treatment. IF volume was reduced by 50% (P < 0·01) to reach a minimum level between 6 and 9 days after treatment. However, this decline was prevented in the absence of Leydig cells by supplementation with testosterone from the time of EDS injection, a treatment which also kept gonadotrophins at minimum or undetectable levels. Furthermore, the reduced IF volume seen up to 9 days after treatment with EDS alone could be restored to control levels within 3 days by a single injection of testosterone. The results obtained demonstrate that androgens, but not Leydig cells or gonadotrophins, are required for the maintenance of interstitial fluid volume in the adult rat testis. It is suggested that the seminiferous tubules may mediate this response, through an androgen-dependent mechanism.

Journal of Endocrinology (1989) 120, 215–222
An acid-stable insulin-like growth factor (IGF)-binding protein from pig serum inhibits binding of IGF-I and IGF-II to vascular endothelial cells
Authors: Gopinath R. , Walton P. E. , Etherton T. D.
Published: December 2008

The effects of a porcine insulin-like growth factor (IGF)-binding protein on binding of IGF-I and IGF-II to porcine aortic endothelial cells (PAEC) were determined. Binding of 125I-labelled IGF-I and -II to IGF receptors was inhibited by IGF-binding protein. IGF-binding protein inhibited binding of IGF-I and -II in a dose-dependent manner with half-maximal inhibition occurring at 5·43 and 108 μg/l respectively. A125I-labelled IGF-I–IGF-binding protein complex, formed by incubating 125I-labelled IGF-I with IGF-binding protein overnight at 4 °C, did not effectively bind to endothelial IGF receptors. Addition of IGF-binding protein to PAEC previously incubated with IGF-I caused a marked dissociation of bound IGF-I (47% dissociation within 12 h). These results indicate that the acid-stable IGF-binding protein which appears to be a part of the 150 kDa GH-dependent binding protein, blocks binding of IGF-I and -II by the IGF receptors and appears to exhibit a higher affinity for IGF-I than the endothelial type-I IGF receptor. The ramifications of this latter point with respect to transfer of circulating IGFs (bound to their IGF-binding proteins) across the vascular endothelium are not clear.

Journal of Endocrinology (1989) 120, 231–236
Role of the adrenal cortex in chronic stress-induced inhibition of prolactin secretion in male rats
Authors: López-Calderón A. , Ariznavarreta C. , Calderón M. D. , Tresguerres J. A. F. , Gonzalez-Quijano M. I.
Published: December 2008

The response of prolactin to chronic stress in intact, adrenalectomized and adrenomedullectomized male rats was studied. Immobilization stress in intact animals induced a significant increase in plasma concentrations of prolactin after 20 and 45 min and a significant decrease when the rats were submitted to chronic restraint (6 h daily for 4 days). Five weeks after adrenomedullectomy, plasma prolactin and corticosterone responses to chronic stress were not modified. In contrast, the inhibitory effect of chronic stress on prolactin secretion was totally suppressed by adrenalectomy. When treated with dexamethasone during the 4 days of restraint, adrenalectomized stressed rats showed similar plasma concentrations of prolactin to the intact stressed rats. These data indicate that the adrenal cortex is able to play an inhibitory role on prolactin secretion during stress only through a prolonged release of glucocorticoids.

Journal of Endocrinology (1989) 120, 269–273
Circulating reproductive hormones and hypothalamic oestradiol and progestin receptors in infertile Zucker rats
Authors: Whitaker E. M. , Robinson A. C.
Published: December 2008

Plasma concentrations of LH, FSH, prolactin and progesterone were measured during the oestrous cycle in obese (fa/fa) and non obese (Fa/?) Zucker rats. In obese rats the mid-afternoon surge of LH during prooestrus was reduced compared with that in non-obese rats (P<0.05), and the maximum concentrations of FSH and prolactin declined more slowly during oestrus. Progesterone concentrations were higher during most of the oestrous cycle in obese rats. Oestradiol and progestin receptors were measured in the hypothalamus of female Zucker rats. Lower concentrations of oestradiol receptors were found in the preoptic area of obese rats (P<0.05). Concentrations of oestradiol receptors in the medial basal hypothalamus were also lower in obese rats, though the difference was not statistically significant. Concentrations of progestin receptors were similar in both phenotypes in the preoptic area and media basal hypothalamus. It seems likely that the abnormalities in reproductive hormones and oestradiol receptors contribute to the infertility of obese female Zucker rats. The underlying mechanism has still to be determined.

Journal of Endocrinology (1989) 120, 331–336
The effect of a 72-h fast on plasma levels of pituitary, adrenal, thyroid, pancreatic and gastrointestinal hormones in healthy men and women
Authors: Beer S. F. , Bircham P. M. M. , Bloom S. R. , Clark P. M. , Hales C. N. , Hughes C. M. , Jones C. T. , Marsh D. R. , Raggatt P. R. , Findlay A. L. R.
Published: December 2008

Seventeen human subjects fasted without electrolyte replacement for 3 days and hormone levels were measured before, during and after the fast. Immediate consequences of the fasting state in healthy human subjects include a marked increase in plasma cortisol, ACTH, β-endorphin, β-lipotrophic hormone, adrenaline, noradrenaline and dopamine. Levels of all these hormones were much greater on the first morning of the fast than in the post-prandial state, even though the plasma glucose level was no lower than that observed on the morning before the fast began. A clear fall in TSH and tri-iodothyronine (T3) levels was observed, but thyroxine levels did not change significantly. Insulin levels fell whereas proinsulin levels did not fall during the fast, though they did rise markedly upon re-feeding. An increase in GH levels was particularly apparent in male subjects, but was also seen in females when evening samples were compared. Pancreatic glucagon showed a modest rise during the fast, but fell again on refeeding; total glucagon also rose as the fast proceeded, but increased markedly upon re-feeding. Levels of gastrin and peptide YY remained low during the fast. Plasma electrolyte levels were unchanged. The following were closely correlated: cortisol with ACTH, T3 with log10TSH, dopamine with noradrenaline, and (negatively, during the fast) pancreatic glucagon with glucose.

Journal of Endocrinology (1989) 120, 337–350
Characterization of insulin-like growth factor binding proteins in ovine tissue fluids
Authors: Hodgkinson S. C. , Moore L. , Napier J. R. , Davis S. R. , Bass J. J. , Gluckman P. D.
Published: December 2008

Competitive tracer binding studies using radio-iodinated insulin-like growth factor-I and -II (125I-labelled IGF-I and 125I-labelled IGF-II) together with size exclusion chromatography and IGF-I affinity chromatography have been used to characterize IGF binding protein activity in ovine tissue fluids. Binding proteins of >200, 150 and 40–50 kDa were revealed in these studies and shown to be widely distributed in body fluids. Thus, the > 200 kDa binding protein, which is IGF-II specific, is present in plasma from mature sheep, colostrum and follicular fluid as well as fetal sheep plasma. This may be the ovine equivalent of the soluble type-2 IGF receptor recently identified in rat serum. The presence of a 150 kDa binding protein, of mixed specificity for IGF-I and IGF-II, in fetal and mature sheep plasma was confirmed in these studies. This protein, previously believed to be restricted to vascular fluids, was also identified in mammary lymph, follicular fluid and as a minor component in vitreous humor. Binding proteins of 40–50 kDa were revealed in every fluid tested and multiple variants with distinct specificities were also suggested. This was investigated by IGF-I affinity chromatography using mature sheep plasma. Following passage through the affinity adsorbent, binding of 125I-labelled IGF-I to proteins in the 40–50 kDa region was abolished but when 125I-labelled IGF-II was used as tracer, a binding protein of 40–50 kDa was still observed.

Thus sheep plasma contains at least two 40-50 kDa binding proteins. The competitive tracer binding studies indicated that one such protein demonstrates mixed specificity for IGF-I and -II while the other strongly favours IGF-II.

Journal of Endocrinology (1989) 120, 429–438
Maternal pinealectomy abolishes the diurnal rhythm in plasma melatonin concentrations in the fetal sheep and pregnant ewe during late gestation
Authors: McMillen I. C. , Nowak R.
Published: December 2008

We have investigated the effect of pinealectomy of ewes in pregnancy on the presence of the diurnal rhythm in fetal and maternal plasma concentrations of melatonin. Six ewes were pinealectomized between 104 and 118 days of gestation. Fetal and maternal blood samples were collected during 24-h periods between 125 and 140 days of gestation in the pinealectomized ewes and in an intact control (n = 4). There was a significant diurnal rhythm in both fetal and maternal plasma concentrations of melatonin in the control group. In this group, the fetal and maternal plasma melatonin concentrations were significantly higher in the dark (128·4±6·2 and 192·2± 10·7 pmol/l respectively) than in the light (46·2 ± 4·2 and 25·8 ± 2·1 pmol/l respectively). However there was no diurnal rhythm in either the fetal or maternal plasma melatonin concentrations in the pinealectomized group between 125 and 140 days of gestation. In contrast to the control animals, there was also no light–dark difference in the fetal or maternal plasma melatonin concentrations in four pinealectomized animals sampled frequently in the 3–7 days preceding delivery (mean length of gestation 146·5 days). However, in the pinealectomized sheep there was a gradual increase in the combined light–dark fetal plasma melatonin concentrations during late gestation from 27·9 ± 2·8 pmol/l (at 15–20 days before delivery) to 95·2± 14·1 pmol/l on the day of delivery.

We have therefore demonstrated that the maternal pineal is the major source of the diurnal rhythm in maternal and fetal plasma melatonin concentrations. However maternal pinealectomy does not appear to remove all the melatonin immunoreactivity from the maternal and fetal plasma in late gestation.

Journal of Endocrinology (1989) 120, 459–464
Effects of ethanol and inhibitors of its metabolism on the circulating levels of Met-enkephalin in greyhounds
Authors: Medbak S. , Mason D. F. J. , Rees L. H.
Published: December 2008

The mechanisms involved in the release of Metenkephalin-like immunoreactivity (MLI) into the circulation following oral administration of ethanol and chlorpropamide were investigated in dogs. The origin of plasma MLI and the sites where it may be metabolized were also studied. Moreover, the molecular nature of circulating MLI was characterized.

In conscious animals oral administration of ethanol (0·15 ml/kg) led to a significant (P<0·01) rise in plasma MLI concentrations in chlorpropamidepretreated animals from a basal level of 43 ± 6 (mean ± s.e.m.) to a peak of 66 ± 8 ng/l. Similar rises in MLI concentrations were observed following administration of ethanol with disulfiram and ethanol with chlorpropamide and captopril. In contrast, the administration of ethanol alone or ethanol with 4-methylpyrazole resulted in a decrease in plasma MLI concentrations.

Comparisons of two different doses of i.v. acetaldehyde, the first metabolite of ethanol, showed that plasma MLI concentrations rose significantly (P<0·05) only after the larger dose (8 mg/kg), rising from 45±7 to 81 ± 18 ng/l. These results suggest that acetaldehyde is the active component in the chlorpropamide+ ethanol-induced MLI secretion.

Plasma MLI was also measured following acetaldehyde infusion in adrenalectomized dogs with and without hexamethonium treatment. Acute bilateral adrenalectomy resulted in a decrease (P<0·05) in plasma MLI concentrations, but the levels remained detectable. Moreover, subsequent acetaldehyde infusion led to rises in plasma MLI similar to those observed in animals with intact adrenals. These MLI responses were not altered by the concurrent i.v. administration of hexamethonium. Gel filtration chromatography revealed that Met-enkephalin exists in the circulation predominantly in larger molecular forms with approximate sizes of 18 000 and 8000 Da in the basal state, after stimulation and following adrenalectomy.

Journal of Endocrinology (1989) 120, 473–480
Effect of the body condition of ewes on the secretion of LH and FSH and the pituitary response to gonadotrophin-releasing hormone
Authors: Rhind S. M. , McMillen S. , McKelvey W. A. C. , Rodriguez-Herrejon F. F. , McNeilly A. S.
Published: December 2008

The effects of body fat content (body condition) of ewes on hypothalamic activity and gonadotrophin-releasing hormone (GnRH) secretion and on pituitary sensitivity to GnRH were investigated using Scottish Blackface ewes. Two groups of 12 ewes were fed so that they achieved either a high body condition score (2·98, s.e.m. = 0·046; approximately 27% of empty body weight as fat) or a low body condition score (1·94, s.e.m. = 0·031; approximately 19% of empty body weight as fat) by 4 weeks before the period of study. Thereafter, they were differentially fed so that the difference in mean condition score was maintained. Oestrus was synchronized, and on day 11 of the subsequent cycle half of the ewes of each group were ovariectomized. On day 12, the remaining ewes were injected (i.m.) with 100 μg prostaglandin F2α analogue and ovariectomized 30 h later. Numbers of large ovarian follicles and corpora lutea present at ovariectomy were recorded. Blood samples were collected at 15-min intervals for 12 h on day 10 of the cycle (luteal phase) and at 10-min intervals from 24 to 30 h after prostaglandin injection (follicular phase). At days 2 and 7 after ovariectomy, samples were collected at 15-min intervals for 8 h and ewes were then injected with 10 μg GnRH and samples were collected for a further 3 h. Samples were assayed for LH and FSH. Ewes in high body condition had more more large follicles than ewes in low body condition during the luteal phase (15·3 vs 6·5; P < 0·05) and follicular phase (11·5 vs 7·0; NS) and a slightly higher mean ovulation rate (1·50 vs 1·20; NS). However, during the luteal and follicular phases of the cycle before ovariectomy there was no effect of condition score on mean LH or FSH concentrations or mean LH pulse frequency or pulse amplitude. Two days after ovariectomy, ewes of high body condition had a higher mean LH pulse frequency than ewes of low body condition (P < 0·05) and higher mean FSH concentrations (P < 0·05). Mean LH concentration and pulse amplitude were not affected. LH and FSH profiles were not affected by body condition on day 7. GnRH-induced increases in LH and FSH concentrations on days 2 and 7 were not affected by body condition. At day 7, but not day 2, ewes ovariectomized during the luteal phase of the cycle had a significantly (P < 0·05) greater GnRH-induced LH release compared with ewes ovariectomized during the follicular phase. It is concluded that body condition directly affects hypothalamic activity and GnRH secretion, but not pituitary sensitivity to GnRH, and that effects on reproductive performance are also mediated through changes in ovarian hormones or in hypothalamo-pituitary sensitivity to ovarian hormones.

Journal of Endocrinology (1989) 120, 497–502