bluuebunny/crossref_metadata_2025 huggingface dataset:
Characteristics and developmental changes of corticotrophin-releasing hormone-binding sites in the fetal sheep anterior pituitary gland
Authors: Lü F. , Yang K. , Challis J. R. G.
Published: December 2008

The responses of the fetal sheep pituitary to corticotrophin-releasing hormone (CRH) change during gestation with maximum output of ACTH around days 120–130, and decreased ACTH output near term. However, there is no information available concerning the extent to which these responses may be modulated by alterations in the number of CRH receptors. Therefore we measured specific CRH-binding sites, and changes in binding characteristics in membrane preparations from fetal sheep anterior pituitaries collected at days 65–70, 85–88, 100–110, 125–130 and at term (approximately 145 days). Binding assays were carried out using 125I-labelled Tyr-ovine CRH (125I-Tyr-oCRH), incubated with crude membrane fractions for 90 min at 22 °C. Binding was time- and temperature-dependent, linear with protein concentration, saturable and specific for oCRH. Scatchard analysis of binding data for individual tissues revealed a single class of CRH-binding sites with high affinity (Kd ≃1 nmol/l) that did not change significantly with gestational age. However, the number of CRH-binding sites increased progressively from days 65–70 to a maximum at days 125–130, then decreased at term. These results demonstrate the presence of specific CRH-binding sites in the fetal sheep anterior pituitary. Furthermore, the change in CRH receptor number with advancing pregnancy follows a similar time-course to the changes reported previously in responsiveness of the fetal sheep anterior pituitary to exogenous CRH stimulation in vivo. These results suggest that alterations in CRH receptor number may contribute to changes in responsiveness of the fetal sheep anterior pituitary to CRH during gestation.

Journal of Endocrinology (1991) 130, 223–229
Regulation by thyroid status of c-myc, c-fos and H-ras mRNAs in the rat myocardium
Authors: Green N. K. , Gammage M. D. , Franklyn J. A. , Sheppard M. C.
Published: December 2008

Effects of thyroid status on expression of a variety of myocardial genes, such as those encoding contractile proteins, have been reported, as well as interactions between thyroid hormones and developmental and haemodynamic regulation of contractile protein synthesis. In addition, it is clear that developmental and haemodynamic factors regulate expression of specific proto-oncogenes, including c-myc, c-fos and H-ras, in the myocardium but the effect of thyroid status on such proto-oncogene products, which are proposed to play a critical signal-transducing role in the heart, has been previously unexplored.

In order to determine whether changes in thyroid status are associated with changes in expression of these putative intracellular signals, we examined the effect of hypothyroidism and tri-iodothyronine (T3) treatment on myocardial levels of c-myc, c-fos and H-ras mRNAs in the rat. The induction of hypothyroidism was associated with a marked increase in myocardial c-myc, c-fos and H-ras mRNAs, changes reversed by 72 h of T3 replacement. Administration of T3 to euthyroid rats had no significant effect on myocardial c-myc or c-fos mRNAs, but inhibition of H-ras mRNA by T3 was evident. These observations demonstrating influences of thyroid status on expression of specific proto-oncogenes suggest that thyroid hormones, as well as exerting direct effects on expression of functionally important myocardial genes, also interact with the cellular transduction pathways mediated by the products of the c-myc, c-fos and H-ras genes.

Journal of Endocrinology (1991) 130, 239–244
Reduced serum concentrations of insulin-like growth factor-I (IGF-I) in protein-restricted growing rats are accompanied by reduced IGF-I mRNA levels in liver and skeletal muscle
Authors: VandeHaar M. J. , Moats-Staats B. M. , Davenport M. L. , Walker J. L. , Ketelslegers J.-M. , Sharma B. K. , Underwood L. E.
Published: December 2008

The serum concentration of insulin-like growth factor-I (IGF-I) is reduced in growing rats fed a low-protein diet, and this decrease is age-dependent, being more pronounced in younger animals. To determine whether this decrease in serum IGF-I is related to a decrease in IGF-I mRNA, growing female rats were given free access to either a 15% protein-sufficient or a 5% protein-deficient diet for 1 week. Protein restriction in 4-week-old rats decreased body weight gain by 44% (P < 0·001) compared with 4-week controls), serum IGF-I concentration by 67% (P < 0·001) and liver IGF-I mRNA abundance by 51% (P < 0·001). During week 6, protein restriction for 1 week resulted in a 20% increase in food intake with no change in weight gain, a 38% reduction in serum IGF-I (P < 0·001 compared with 6-week controls) and a 39% decrease in liver IGF-I mRNA (P < 0·001). The serum IGF-I concentration was highly correlated (r = 0·80; P < 0·001) with the hepatic IGF-I mRNA concentration. Skeletal muscle IGF-I mRNA abundance was also decreased significantly by protein restriction (37% at week 4, P<0·001, and 24% at week 6, P < 0·01) and was closely correlated (r = 0·71; P < 0·001) with body weight gain. Liver GH-binding protein and GH receptor mRNA abundance were reduced by 1 week of protein deprivation at week 6 but not at week 4. We conclude that the reduced serum IGF-I of young rats fed a low-protein diet is due, in part, to reduced liver IGF-I mRNA, and that these changes are not dependent on GH binding. Decreased skeletal muscle IGF-I mRNA during protein restriction is consistent with an autocrine/paracrine action of IGF-I in muscle.

Journal of Endocrinology (1991) 130, 305–312
Influence of nutritional status on growth hormone-dependent circulating somatomedin-C activity in mature sheep
Authors: Wynn P. C. , Stuart M. C. , Wallace A. L. C. , Kirby A. C. , Annison E. F.
Published: December 2008

The effect of daily administration of ovine GH for a period of 4 weeks on somatomedin-C biological activity in plasma was investigated in mature Merino sheep fed a maintenance energy intake (low plane; LP) or 1·6 times this amount (high plane; HP). The GH treatment resulted in a significant (P < 0·05) increase in plasma GH levels in blood samples collected 23·5 h after each daily injection in both LP and HP groups. Plasma concentrations of somatomedin-C activity and insulin were significantly stimulated to a maximum level by the third GH injection and remained at this level for 7 days. Subsequently, circulating levels of both hormones fell to 40–50% of the peak response to GH and returned to basal levels within 48 h of the cessation of GH injections. In the HP group the response of plasma insulin and somatomedin-C activity to GH injection was greater than in the LP group.

Journal of Endocrinology (1991) 130, 313–320
Roles of progesterone and oestradiol in determining the temporal sequence and quantitative expression of sexual receptivity and the preovulatory LH surge in the ewe
Authors: Fabre-Nys C. , Martin G. B.
Published: December 2008

In a series of experiments using a quantitative method for measuring receptivity and implants that allowed rapid and controlled changes in the blood concentrations of oestradiol-17β and progesterone, we have re-examined the roles of these steroids in the induction of sexual behaviour and the LH surge in ovariectomized ewes.

Progesterone priming was found to increase the proportion of ewes showing oestrus, reduce the latency to the onset of oestrus, and increase the 'intensity' of the behaviour as measured by the receptivity index, but it did not affect the size of the LH surge. Progesterone was able to facilitate the expression of oestrus even when it was withdrawn 8 days before oestrogen treatment, suggesting that it exerts its effect by restoring the sensitivity of the oestrogen-refractory animal to oestrogen. When it was present at the time oestrogen was administered, progesterone inhibited the stimulatory effect of oestrogen, but this effect disappeared as soon as the progesterone was withdrawn. Thereafter, expression of both the behavioural and endocrine responses was delayed by 24–30 h. These data show that the timing of the preovulatory behavioural and endocrine events is determined primarily by the time of progesterone withdrawal. The amount of oestradiol and the timing of any rise in concentration serve only to modulate these effects.

Journal of Endocrinology (1991) 130, 367–379
The cell-adhesive effect of basic fibroblast growth factor on pituitary cells in vitro
Authors: Inoue K. , Sakai T. , Hattori M.
Published: December 2008

Both native and recombinant basic fibroblast growth factor (FGF) exerted a marked effect on the morphology of a pituitary clonal somatotroph-like cell line (MtT/S) in vitro. Addition of 10 pg–20 ng bovine basic FGF/ml caused the cells to flatten and spread out over the culture dish, the cells showing strong surface contact. Those in contact with the culture dish were epithelial in appearance, being polygonal, closely apposed and forming a pavement-like monolayer. Basic FGF inhibited cell proliferation in a dose-dependent manner at the higher concentrations tested (0·5–20 ng/ml). This cell-adhesive effect of basic FGF was also observed in normal dispersed pituitary cells. The observed stimulation of pituitary cell adhesion is the first report of a morphological effect of basic FGF on pituitary cells and could explain the physiological significance of basic FGF and its high concentration in the pituitary gland.

Journal of Endocrinology (1991) 130, 381–386
Diurnal changes in the plasma concentrations of LH and hypothalamic contents of LHRH-I and LHRH-II in the domestic hen
Authors: Wilson S. C. , Gladwell R. T. , Cunningham F. J.
Published: December 2008

Diurnal changes of LH secretion in sexually immature hens of 9, 11, 13 and 15 weeks of age consisted of 25–40% increases in the mean concentrations of LH in plasma between 15.00 and 18.00 h, i.e. between 2 h before and 1 h after the onset of darkness. During this time there was a tendency for the mean contents of LHRH-I in the anterior hypothalamus and posterior hypothalamus to increase by 21–74% and 20–56% respectively. In hens of 9 and 15 weeks, diurnal changes in the plasma concentration of LH closely paralleled those of LHRH-I content in the posterior hypothalamus. In contrast, the diurnal rhythm of LH secretion in hens of 11 and 13 weeks was more marked and plasma concentrations of LH continued to rise steeply between 18.00 and 21.00 h, i.e. between 1 and 4 h after the onset of darkness. At 11 weeks, this was associated with a reduction (P<0·01) in the contents of LHRH-I and LHRH-II, particularly in the anterior hypothalamus. In laying hens, a diurnal decline (P<0·01) in the plasma concentration of LH between 1 and 4 h after the onset of darkness was preceded by a fall (P<0·05) in the content of LHRH-I in the posterior hypothalamus and in the total hypothalamic content of LHRH-II (P<0·01). In all groups of hens, irrespective of the times of day at which tissue was taken, significant (P<0·05–<0·001) correlations between the contents of LHRH-I and LHRH-II in the anterior hypothalamus were observed.

It is concluded that a diurnal rhythm of release of LHRH-I may drive the diurnal rhythm of LH secretion. Thus, in sexually immature hens of 9 and 15 weeks and laying hens in which diurnal changes in plasma LH were small there were parallel changes in the content of LHRH-I in the posterior hypothalamus. However, where the plasma concentration of LH was increased substantially, as at 11 weeks, there was a decline in the hypothalamic contents of LHRH-I. A simultaneous fall in the hypothalamic content of LHRH-II raises the possibility of a causal relationship between the activities of LHRH-II, LHRH-I and the release of LH.

Journal of Endocrinology (1991) 130, 457–462
Galanin in the normal human pituitary and brain and in pituitary adenomas
Authors: Bennet W. M. , Hill S. F. , Ghatei M. A. , Bloom S. R.
Published: December 2008

Galanin-like immunoreactivity (IR) was measured by radioimmunoassay in extracts of non-tumorous and tumorous human pituitaries and in multiple sites in the human brain. Galanin-IR was present in considerable quantities in the non-tumorous pituitaries (21·4±1·2 pmol/g wet weight; mean ± s.e.m., n = 30). In 25 pituitary tumours, galanin-IR was detectable in extracts of only nine, with a mean concentration of 11·5±4·4 pmol/g. Galanin-IR was undetectable in the remaining 16. Of ten brain sites, galanin-IR was detected only in the hypothalamus, where the concentration was 9·1±1·8 pmol/g (n = 5). On fast protein liquid chromatography of the non-tumorous pituitary extracts, galanin-IR mostly eluted in a peak with a retention time similar to that of synthetic porcine galanin. On gel permeation chromatography, galanin-IR eluted as a peak with an elution coefficient (Kav) of 0·72, also similar to that of porcine galanin, with additional preceding (Kav 0·62) and following (Kav 0·77) peaks of galanin-IR. These results show that healthy human pituitary and hypothalamus contain substantial amounts of galanin, whereas it is present in variable amounts or not at all in pituitary tumours. Chromatographic analysis suggests that pituitary galanin is present in three molecular forms, with the majority corresponding to synthetic porcine galanin.

Journal of Endocrinology (1991) 130, 463–467
Effects of exogenous growth hormone on growth and body composition in genetically selected mice
Authors: Bootland L. H. , Hill W. G. , Sinnett-Smith P. A.
Published: December 2008

The effects of exogenous GH on growth and body composition were investigated in lines of mice selected for high or low body weight (P-lines) or high or low body fat (F-lines). Mice from all lines were given daily injections of recombinant bovine GH or a placebo for 21 days from 4 weeks old. They were killed and various organ weights measured. There was no consistent effect of GH on organ weights. In all lines of mice the rate of weight gain and final weight increased in response to GH. In both lines selected for body fat, GH treatment decreased fat content. The low body weight mice also became less fat, but in the high body weight mice GH treatment increased fat percentage. The results indicate that the differences in growth rate and body composition observed in these lines are not due to differences in responsiveness to GH.

Journal of Endocrinology (1991) 131, 19–24
Thyroidal inhibition of chicken pituitary growth hormone: alterations in secretion and accumulation of newly synthesized hormone
Authors: Denver R. J. , Harvey S.
Published: December 2008

Hypothyroidism reduces GH synthesis and release in several mammalian species, in which thyroid hormone directly stimulates GH gene transcription. In contrast, hypothyroidism stimulates GH secretion in birds, in which thyroid hormone directly inhibits pituitary GH release. We have, therefore, investigated the effects of thyroid status on the accumulation of newly synthesized GH in the pituitaries of 8- to 10-week-old Leghorn cockerels in vitro and in vivo. The incorporation of [35S]methionine into immunoprecipitable GH ([35S] GH) was increased, over a 4-h incubation period, in glands from birds made hypothyroid by injections of methimazole (50 mg/kg day for 10 days) in comparison with glands from vehicle-injected controls. Treatment with tri-iodothyronine (T3, 100 μg/kg per day for 10 days) in vivo did not significantly alter the accumulation of [35S]GH in vitro but did block the release of [35S]GH in response to a GH secretagogue (thyrotrophin-releasing hormone; exposure to 280 nmol/l for 30 min) and reduced immunoassayable pituitary GH content. Pretreatment of glands from euthyroid birds with T3 (100 nmol/l) in vitro (for 20 h) reduced the basal accumulation of [35S]GH as well as that induced by another GH secretagogue (GH-releasing factor; 100 nmol/l) during a 6-h labelling period. These results show that, unlike the generally stimulatory action of thyroid hormone in mammals, in birds, T3 exerts a direct inhibitory effect on the accumulation of newly synthesized pituitary GH.

Journal of Endocrinology (1991) 131, 39–48
Modulation of immunoreactive epidermal growth factor levels in the submandibular gland, pancreas, liver, kidney and gastrointestinal tract of suckling rats by cortisone and tri-iodothyronine
Authors: Schaudies R. P. , Grimes J. , Davis D. , Koldovský O.
Published: December 2008

Suckling rats exhibit age-dependent differences in epidermal growth factor (EGF) levels in several organs. The present studies evaluated the effects of two hormones known for their maturative effect on suckling rats, cortisone and tri-iodothyronine (T3), on immunoreactive EGF levels in specific organs. Suckling rats were administered cortisone (5 mg/100 g body weight per day) or T3 (50 μg/100 g body weight per day) on days 8, 9, 10 and 11 after birth, and killed on day 12. Submandibular glands, kidneys, pancreas, liver and gastrointestinal tract mucosa and lumen were assayed for immunoreactive EGF by a speciesspecific radioimmunoassay. Low levels of EGF in the submandibular glands were increased slightly by both T3 and cortisone treatment. Cortisone evoked a tenfold increase in EGF in the pancreas, but had no effect on levels in the kidney or liver. In contrast, T3 evoked a sixfold increase in the EGF level in the kidney, but had no effect on levels in the pancreas or liver. Hormonal administration had no effect on EGF levels in the stomach. Within the intestinal tract, cortisone had no effect on the luminal EGF content of the duodenum, jejunum or ileum, but caused a decrease in the midjejunum. T3 evoked a decrease in the luminal EGF content of the ileum. The effect of cortisone on mucosal EGF content varied between regions; an increase was seen in the duodenum with a decrease in the midjejunum and ileum. T3 administration resulted in a significant decrease in EGF only in the mucosa of the ileum. The EGF-degradative capacity of the luminal contents of the jejunum and ileum, as studied in vitro, were increased by treatment with both T3 and cortisone. However, no direct correlation was observed between alterations of the EGF content in the lumen or mucosa and the increased degradative capacity. We therefore conclude that modulation of EGF levels within the intestine cannot be explained exclusively by alterations in the degradation rates. The results indicate that both adrenal and thyroid glands may play an important role in the modulation of EGF levels in the developing rat.

Journal of Endocrinology (1991) 131, 95–100
Purification and characterization of calcium-calmodulin kinase II from human parathyroid glands
Authors: Kato M. , Hagiwara M. , Nimura Y. , Shionoya S. , Hidaka H.
Published: December 2008

Calmodulin has been identified in parathyroid cells and is thought to play an important role in the production or secretion of parathyroid hormone. However, a detailed investigation of calmodulinbinding proteins in parathyroid glands has not been conducted. In this study, we attempted to determine the presence of calmodulin-binding protein in human parathyroid adenoma by affinity chromatography. The eluted protein from a calmodulin-coupled Sepharose 4B column with EGTA was analysed by sodium dodecyl sulphate-polyacrylamide gel electrophoresis which revealed a major protein band of Mr 50 000. A Ca2+/calmodulin-dependent protein kinase activity was detected at the protein peak using dephosphorylated casein as a substrate. The 50 kDa band was identified as calcium/calmodulin-dependent protein kinase II (CaM-kinase II) by immunoblotting. The substrate specificity, pH dependency and affinity for calmodulin of this enzyme were identical to those of CaM-kinase II from rat brain. Also, the kinase activity was sensitive to KN-62, a specific inhibitor of CaM-kinase II. In total, 0·48 mg of this kinase was purified from 3 g human parathyroid adenoma.

Journal of Endocrinology (1991) 131, 155–162
Transferrin inhibits aromatase activity of rat granulosa cells in vitro
Authors: Li Y. D. , Zhang Z. W. , Li W. X.
Published: December 2008

The effect of transferrin on basal and FSH-stimulated aromatase activity of granulosa cells from immature female rats treated with diethylstilboestrol (DES) was examined in vitro by a radiometric method. The basal activity of the enzyme was very low after 3 days of incubation. Treatment with FSH (20 ng/ml) resulted in a 9·6-fold increase in activity, whereas coincubation with increasing doses of transferrin (3–300 μg/ml) produced a dose-dependent inhibition of FSH-stimulated aromatase activity with a projected minimal effective dose of < 2 μg/ml. A time-course study showed that the inhibitory effect of transferrin on aromatase activity has become significant at 48 h of incubation.

The inhibitory action of transferrin on the enzyme complex was further confirmed by showing that the FSH dose–response curve was significantly suppressed by concomitant treatment with 100 μg transferrin/ml with a maximum suppression of 54·1 % at a dose of 30 ng FSH/ml.

The possibility that transferrin may act through a non-specific inhibitory effect seems unlikely, as no changes in cell number and DNA content per well were observed. In fact, protein synthesis was enhanced after treatment with transferrin. Aromatase activity, stimulated by several promoters of cyclic AMP (cAMP), such as prostaglandin E2 (PGE2), forskolin and 8-bromo-cAMP, was significantly suppressed by 100 μg transferrin/ml (36·6, 47·4 and 23·4% inhibition respectively), suggesting that the effect of transferrin on FSH action may involve a site(s) distal to cAMP generation.

These findings indicated that transferrin, present in follicular fluid, may play an important role in the regulation of granulosa cell differentiation.

Journal of Endocrinology (1991) 131, 245–250
Oral insulin in diabetic dogs
Authors: Saffran M. , Field J. B. , Peña J. , Jones R. H. , Okuda Y.
Published: December 2008

Bovine crystalline insulin, mixed with an absorption enhancer, was loaded by hand into gelatin capsules, which were then coated with an azopolymer designed to deliver the insulin in the upper colon. In 34 experiments with 14 pancreatectomized mongrel dogs of both sexes, the coated capsules were administered orally after a pre-dose period of 1 h. The dogs had cannulae in the portal vein, hepatic vein and femoral artery and Doppler flow probes on the portal vein and hepatic artery. Insulin and food were withdrawn the day before an experiment. Responses measured were plasma glucose, plasma insulin, hepatic glucose production rate, hepatic plasma flow rate and plasma glucagon-like immunoactivity (GLI). Control experiments, with capsules without insulin, produced small changes from 'pre-dose' values. Insulin-containing capsules, without the azopolymer coating, resulted in some early changes consistent with upper gastrointestinal absorption. Single oral doses (66 to 400 nmol/kg) of insulin in completely coated capsules produced peaks of portal plasma insulin and transient decreases in plasma glucose, hepatic glucose production, hepatic plasma flow and plasma GLI. The changes usually began 1·5–2 h after administration of a single dose, and lasted for up to 3 h, but were not significantly related to the dose of insulin. Multiple oral doses of insulin, given at 1·5-h intervals, resulted in multiple peaks of plasma insulin, a continuing dose-dependent fall in plasma glucose to near-euglycaemia with the highest dose, and profound decreases in hepatic glucose production and plasma GLI. These data demonstrate that insulin absorbed from the gastrointestinal tract causes changes in glucose metabolism in the diabetic dog that are consistent with the action of insulin primarily on the liver and that repeated oral doses are necessary to correct the hyperglycaemia.

Journal of Endocrinology (1991) 131, 267–278
Regulation of testicular insulin-like growth factor-I in pubertal growth hormone-deficient male rats
Authors: Spiteri-Grech J. , Bartlett J. M. S. , Nieschlag E.
Published: December 2008

GH plays a major role in pubertal growth, effects mainly mediated by stimulation of insulin-like growth factor-I (IGF-I) production by the liver. However, the role of GH in the regulation of pubertal onset, spermatogenesis and fertility is still under debate. GH and FSH have, in addition, been implicated in the regulation of IGF-I production by Sertoli cells in a number of studies, although conflicting results have been reported. The interpretation of studies using GH-deficient mutant mice has been complicated by the presence of additional defects in the hypothalamic-pituitary-gonadal axis of these animals. We have therefore used GH-deficient mutant male rats with no other documented hormonal deficiencies to study the effect of GH administration on somatic and testicular development, circulating and testicular IGF-I concentrations and testicular histology.

Body weights in GH-deficient rats substituted with GH were not significantly different from untreated or GH-treated normal rats and were significantly higher than body weights in untreated dwarf rats. Similarly, circulating IGF-I concentrations in GH-treated GH-deficient rats were not significantly different from those in untreated or GH-treated normal rats but were significantly higher than circulating IGF-I concentrations in untreated dwarf rats. No differences in testicular IGF-I concentrations were observed in any of the groups studied. Testicular weights remained low in both untreated and GH-treated GH-deficient animals compared with control animals but spermatogenesis was qualitatively and quantitatively normal in all groups at the end of the observation period.

We conclude that GH does not play a major role in the regulation of testicular IGF-I production at puberty although we cannot exclude the possibility that the low but detectable levels of GH in the blood of mutant rats is sufficient to augment testicular responsiveness to gonadotrophins and therefore result in normal gonadal development.

Journal of Endocrinology (1991) 131, 279–285
The insulin-like growth factors and their binding proteins in a case of non-islet-cell tumour-associated hypoglycaemia
Authors: Cotterill A. M. , Holly J. M. P. , Davies S. C. , Coulson V. J. , Price P. A. , Wass J. A. H.
Published: December 2008

Non-islet-cell tumours which induce hypoglycaemia are rare. Insulin-like growth factor-II (IGF-II) produced by some tumours is thought to be responsible for the hypoglycaemia and other systemic effects, despite normal or even low serum IGF-II levels. We studied a 44-year-old woman presenting with symptomatic hypoglycaemia associated with a large intraabdominal haemangiopericytoma. The serum IGF-II level was 455 μg/l when measured after acid-ethanol extraction (normal range (NR) 450–750 μg/l) and 1063 μg/l after acid chromatography (normal human serum pool 1068 μg/l). Levels of fasting plasma insulin, C-peptide, glucose and serum IGF-I levels were low before the operation (< 2 mU/l (NR <2-14), 0·23 nmol/l (NR 0-4-1-2), 3-1 mmol/l, (NR 3-7-5-9) and 002 U/ml respectively). After tumour removal, the symptoms resolved rapidly and the patient made a full recovery. Secretion of both insulin and growth hormone was suppressed before the operation in response to a 75 g glucose meal and to an infusion of 100 μg GH-releasing hormone (GHRH) respectively in comparison with studies after the operation. Serum IGF-II levels 6 weeks and 12 weeks after the operation fell to 385 μg/1 (777 μg/1; acid chromatography) and 280 μg/1 (647 μg/1; acid chromatography) and serum IGF-I levels increased to 0-35 U/ml and 0-26 U/ml. Serum before the operation and tumour extract contained chiefly a large molecular weight precursor IGF-II (molecular weight 15 000–20 000) which disappeared from the serum after the operation. The IGF-binding proteins (IGFBP-1, IGFBP-2, IGFBP-3 and IGFBP-4) were examined. The preoperation fasting serum IGFBP-1 level was lower than expected (31 μg/l (NR 20–70 μg/l)) and similar to levels at 6 weeks after the operation (33 μg/l). This was surprising given the differences in plasma insulin levels before and after the operation (< 2 mU/l versus 13 mU/l). Using Western ligand blotting techniques, serum IGFBP-3 levels were found to be low and IGFBP-2 appeared to be the dominant IGFBP before the operation. Serum IGFBP-3 levels after the operation fell further. This further decrease appeared, in part, to be due to the presence of a cation-dependent IGFBP-3-specific protease which has previously only been described in late pregnancy.

We conclude that in this subject, despite normal serum IGF-II levels, the hypoglycaemia and systemic effects on insulin and GH secretion were due to increased bioavailability of a circulating tumour-produced precursor form of IGF-II. This increased bioavailability appears to be due to alterations in the circulating levels and perhaps affinities of the IGFBPs.

Journal of Endocrinology (1991) 131, 303–311
Effect of acute and chronic administration of clonidine on hypothalamic content of growth hormone-releasing hormone and somatostatin in the rat
Authors: Gil-Ad I. , Laron Z. , Koch Y.
Published: December 2008

Hypothalamic GH-releasing hormone (GHRH) and somatostatin contents were determined in male rats at pre- and mid-pubertal stages following acute (0·15 mg/kg) or chronic (0·1 mg/kg twice daily for 7 days) administration of clonidine. Hypothalamic GHRH content increased markedly during transition from pre- to mid- and late-pubertal stages (2·05±0·17vs 3·13±0·26 and 3·46±0·20 ng/hypothalamus respectively, means ±s.e.m.). Hypothalamic GHRH content was markedly decreased 1 h after clonidine administration in mid-pubertal but not in prepubertal rats (3·67±0·23 vs 2·65±0·22 ng/hypothalamus). Hypothalamic somatostatin content, on the other hand, decreased in both age groups, although the decrease in the mid-pubertal group was more pronounced (51% and 38% respectively). In addition, the GH responsiveness to clonidine was higher at mid-puberty than at prepuberty. Determination of hypothalamic somatostatin and GHRH content in the chronic experiment 4 h after the last drug administration revealed a marked decrease in GHRH in the mid-pubertal rats and a slight decrease in the prepubertal rats (1·78±0·32 vs 3·15±0·31 and 2·01±0·30 vs 2·35±0·15 ng/hypothalamus; P < 0·005 and P<0·05 respectively) whereas somatostatin levels were not altered. It is suggested that the clonidineinduced GH secretion is modulated by a chain of events which involves primary stimulation of GHRH release resulting in increased GH secretion which, via a negative-feedback mechanism, triggers an enhancement of somatostatin release which ultimately normalizes the system. This mechanism is age-dependent and reaches full maturity only at the onset of puberty.

Journal of Endocrinology (1991) 131, 381–385
Sex differences in the ultradian pattern of plasma growth hormone concentrations in mice
Authors: MacLeod J. N. , Pampori N. A. , Shapiro B. H.
Published: December 2008

Ultradian patterns of plasma GH concentration were determined in adult male and female mice. Serial blood samples were collected every 15 min over 8 h through surgically placed chronic indwelling right atrial catheters and assayed for GH content by an homologous radioimmunoassay. In both sexes, GH concentrations fluctuated episodically from baseline values that were often in the range of 2 μg/l which approached the limit of assay sensitivity, to peak values sometimes reaching 100 μg/l. Male mice, however, demonstrated a regular periodicity of GH peaks approximately every 2·5 h with interposed stable baseline concentrations that were significantly longer in duration than in females. The absence of extended baseline concentrations in females reduced cycle length to an average of 1·4 h and significantly increased the overall mean GH concentration. The duration, height and area of GH peaks and the average concentration between peaks were comparable in males and females. Sexual dimorphism in the ultradian patterns of serum GH concentrations have been shown in both mice and rats to regulate the expression of several sex-specific phenotypes. Comparing endogenous GH patterns in mice and rats demonstrates that males of both species have less frequent peaks than females over the same interval of time. This separation of GH peaks appears to be an essential element for the expression of masculine GH-dependent traits.

Journal of Endocrinology (1991) 131, 395–399
Testicular steroidogenesis in the testicular feminized (Tfm) mouse: loss of 17α-hydroxylase activity
Authors: Murphy L. , O'Shaughnessy P. J.
Published: December 2008

Testicular feminized (Tfm) mice are totally insensitive to androgen and may be used to study the role of the androgen receptor in normal development and function. We have examined testicular and Leydig cell steroidogenesis in Tfm mice. Serum bioactive LH was high in Tfm mice but serum testosterone was low and this was associated with a severe reduction in testicular testosterone production in vitro. Examination of [3H]pregnenolone metabolism by testes of Tfm mice indicated that progesterone, rather than testosterone, was the major steroid produced. Leydig cells were isolated from normal and Tfm mice and from normal mice in which testicular descent was surgically prevented before puberty. As in whole testes, androgen production in response to human chorionic gonadotrophin was severely reduced in Leydig cells from testes of Tfm mice compared with normal or cryptorchid groups. In contrast, progesterone production by Leydig cells from testes of Tfm mice was markedly increased in comparison with other groups. Total steroid production (progesterone plus androstenedione plus testosterone), however, was only 24% of normal in Leydig cells from Tfm mice. The pattern of steroid production by Leydig cells from cryptorchid testes was similar to control, although total steroid production was reduced to about 50% (this was significantly higher than the Tfm group, P<0·05). The high progesterone/androgen ratio in testes from Tfm mice suggested that 17α-hydroxylase was depleted in these animals. To confirm this, activity of the four major steroidogenic enzymes associated with the smooth endoplasmic reticulum was measured. Activities (per testis) of 3β-hydroxysteroid dehydrogenase and 5α-reductase were normal in Tfm and cryptorchid mice but, as expected, 17α-hydroxylase activity was only 2·4% of control and 4·5% of cryptorchid testes. 17-Ketosteroid reductase activity was markedly reduced in cryptorchid testes (14·4% of control) but there was a further reduction in testes from Tfm mice to 0·1% of control. Results show that the Tfm mutation is associated with marked loss of 17α-hydroxylase and 17-ketosteroid reductase activities. This suggests that these enzymes may require receptor-mediated androgen stimulation during development to express normal activity.

Journal of Endocrinology (1991) 131, 443–449
Comparison of the galactopoietic response to pituitary-derived and recombinant-derived variants of bovine growth hormone
Authors: Eppard P. J. , Bentle L. A. , Violand B. N. , Ganguli S. , Hintz R. L. , Kung L. , Krivi G. G. , Lanza G. M.
Published: December 2008

Two studies were designed to examine the differences in galactopoietic potency of molecular variants of pituitary- and recombinant-derived bovine GH (bGH). The recombinant bGH molecules included aminoterminal and position-127 amino acid substitutions which are representative of two of the four natural pituitary variants or of partially degraded bGH molecules. Amino-terminal variants of bGH included methionine (Met1), alanine (Ala1), serine (Ser1) or deletion of four amino acids (Δ1−4). The Δ1–4 variants were representative of degradation products previously isolated in pituitary bGH preparations. In the first study, 54 lactating Holstein cows received i.m. injections of a buffer solution (control), pituitaryderived bGH, or recombinant-derived [Met1, Leu127]-bGH, [Met1, Val127]-bGH, [Ala1, Leu127]-bGH, or [Ala1, Val127]-bGH. Cows received 25 mg bGH/day for 21 days. Substitution of the amino-terminal alanyl residue with methionine did not affect milk response. GH variants with Val127 elicited a greater milk response (8·5 kg/day) than Leu127 bGH variants (6·5 kg/day). The average milk response to the four recombinant bGH variants was 7·5 kg/day greater than controls compared with 4·4 kg/day for pituitary-derived bGH. In contrast, blood bGH concentrations were equivalent for pituitary and recombinant bGH treatments, approximately 20 μg/l more than control levels at 3 h after injection. Blood free fatty acid concentrations were increased, but insulin and glucose levels were unaffected by bGH treatment. In the second study, 54 lactating Holstein cows received i.m. injections of a buffer control solution or recombinant-derived [Met1,Leu127]-bGH, [Ser1,Leu127]-bGH, [Ser1,Val127]-bGH, [Δ1–4,Leu127]-bGH or [Δ1–4,Val127]-bGH. Cows received 25 mg bGH/day for 28 days. The milk response to full-length bGH variants was 6·6 kg/day greater than the response to the amino-terminal deletion variants (P<0·05). Substitution of valine for leucine did not affect milk response to either the deletion (Δ1–4) or full-length (Met1 or Ser1) bGH molecules. In conclusion, the lowered galactopoietic potency of pituitary bGH preparations was demonstrated, at least in part, to be due to the presence of amino-terminal amino acid deletions rather than differences in amino acid sequences of recombinant bGH. Ala1 bGH variants with valine at postiion 127 elicited a greater milk response than Leu127 variants.

Journal of Endocrinology (1992) 132, 47–56
Arachidonic acid-induced LH release is ATP-independent and insensitive to N-ethyl maleimide
Authors: Kaye P. V. , van der Merwe P. A. , Millar R. P. , Davidson J. S.
Published: December 2008

The mechanism of arachidonic acid (AA)-induced LH release was characterized using sheep pituitary cells in primary culture permeabilized with Staphylococcal α-toxin. In intact cells, exogenous AA evoked release of LH in a manner which was partially dependent on extracellular Ca2+. At similar concentrations, AA also caused cell permeabilization as monitored by efflux of [3H]2-deoxyglucose metabolites. In α-toxin-permeabilized cells where cytosolic Ca2+ was clamped at resting levels, AA retained its ability to cause LH release. Unlike the stimulation of exocytosis produced by Ca2+, phorbol ester or cyclic AMP, AA-evoked release was independent of ATP and was not inhibited by pretreatment with N-ethyl maleimide. These findings indicated that exogenous AA does not cause LH release by Ca2+ influx or mobilization or by activating protein kinase C. The results suggest that LH release induced by exogenous AA is probably due to its detergent-like properties, and does not represent true exocytosis.

Journal of Endocrinology (1992) 132, 77–82
Different effects of the antiprogesterone RU486 on progesterone secretion by the corpus luteum of rats with 4- and 5-day oestrous cycles
Authors: Sánchez-Criado J. E. , Uilenbroek J. Th. J. , Karels B.
Published: December 2008

Administration of the antiprogesterone RU486 (2 mg/day) for 14 days to rats with a 5-day reproductive cycle resulted in an increase in both ovarian and pituitary weight in contrast with rats with a 4-day oestrous cycle. Luteal progesterone production decreased earlier in 4-day than in 5-day cyclic rats. Treatment of 5-day cyclic rats with antiprogesterone from the day of metoestrus onwards resulted in the advancement of the preovulatory prolactin surge by 24 h. Progesterone production by the corpus luteum was, however, not affected, indicating that in 5-day cyclic rats the corpora lutea are still functionally active at the time of the preovulatory surge of prolactin. They become, therefore, stimulated both in size and progesterone production. In contrast, the corpora lutea in 4-day cyclic rats are functionally inactive at the time of the preovulatory surge of prolactin, and prolactin acts luteolytically. In conclusion, the advancement of the preovulatory surge of prolactin by 24 h accounts, at least in part, for the increase in ovarian weight in 5-day cyclic rats after treatment with antiprogesterone. The results of these experiments do not agree with a direct effect of the antiprogesterone RU486 on progesterone secretion by the corpus luteum.

Journal of Endocrinology (1992) 132, 115–122
Effects of oestradiol-17β on FSH-stimulated steroidogenesis in cultured marmoset granulosa cells
Authors: Shaw H. J. , Hodges J. K.
Published: December 2008

A role for oestradiol in ovarian follicular development is well recognized. However, a number of disparate effects have been reported for the action of oestradiol in the primate ovary. To investigate this further, we have examined the effects of oestradiol on the differentiation of granulosa cells isolated from small (0·5–1 mm) antral follicles obtained from the ovaries of prepubertal marmoset monkeys (Callithrix jacchus). Granulosa cells were co-cultured with oestradiol and human FSH (hFSH) for 48 h, or were pretreated with oestradiol for 48 h before addition of gonadotrophin for a further 48 h. Oestradiol (0·01–100 nmol/l) had no effect on basal or hFSH-stimulated progesterone accumulation and aromatase activity when the hormones were added concurrently. Furthermore, oestradiol did not influence the ability of hFSH to induce LH/human chorionic gonadotrophin (hCG) responsiveness in immature granulosa cells. The absence of synergism between oestradiol and hFSH in the induction of marmoset granulosa cell differentiation was independent of the presence of phenol red in culture medium. However, gonadotrophin-stimulated steroidogenesis was attenuated when cells were cultured in the presence of phenol red compared with in its absence; this effect was more pronounced for gonadotrophin-stimulated aromatase activity but was evident for LH/hCG-stimulated progesterone accumulation at higher doses of hCG (10 and 100 ng/ml). An effect of phenol red on basal steroidogenesis was less obvious.

In contrast, pretreatment of granulosa cells with oestradiol (0·1 μmol/l) alone for 48 h resulted in a greater steroidogenic response to hFSH (P<0·001) and dibutyryl cyclic AMP (P<0·01) treatment during a second 48-h culture period compared with cultures pretreated with dihydrotestosterone (0·1 μmol/l), hFSH (30 ng/ml) or without hormones. This stimulatory effect of oestradiol appeared to be specific in that it was antagonized (P<0·05) by the anti-oestrogen tamoxifen (1 μmol/l).

In conclusion, this study provides evidence for a stimulatory autocrine role of oestradiol in the primate ovary. However, the action of oestradiol appears not to synergize with FSH directly, but rather to increase FSH-stimulated steroidogenesis by an independent mechanism.

Journal of Endocrinology (1992) 132, 123–131
Translational status of the insulin-like growth factor-I mRNAs in liver of protein-restricted rats
Authors: Thissen J.-P. , Underwood L. E.
Published: December 2008

Dietary protein restriction in young rats decreases serum insulin-like growth factor-I (IGF-I) concentrations and the amount of liver IGF-I mRNA, suggesting that regulation for the liver IGF-I gene expression in this model occurs at a pretranslational level. To determine whether there is also translational control, we assessed the association of the liver IGF-I mRNA transcripts with polysomes in livers of normally fed (15% dietary protein) versus protein-restricted (5% dietary protein) rats. One week of dietary protein restriction reduces serum IGF-I concentrations by 54% and the amount of liver IGF-I mRNA by 35%, with the 7·5 kb size-class of IGF-I mRNA being the most affected (−48%). Protein restriction reduces the amount of the polysomal IGF-I mRNAs by 30%, a value in close agreement with the changes in total IGF-I mRNAs. Protein restriction is not associated with changes in the distribution of IGF-I mRNAs between the polysomal and non-polysomal fractions. All major size-classes of IGF-I mRNA transcripts (7·5, 4·7, 1·7, 0·9–1·2 kb) are associated with the polysomes in both dietary groups, suggesting involvement in the initiation phase of the IGF-I translation. We conclude that no untranslatable pool of IGF-I mRNAs is present in the liver of protein-restricted animals. Protein restriction, however, decreases slightly the mean size of polysomes. This decrease in ribosomal number associated with the IGF-I mRNA could decrease translational efficiency. Our data suggest that dietary protein restriction does not impair the initiation of IGF-I mRNA translation.

Journal of Endocrinology (1992) 132, 141–147
Reproductive effects of placing micro-implants of melatonin in the mediobasal hypothalamus and preoptic area in rams
Authors: Lincoln G. A. , Maeda K.-I.
Published: December 2008

The reproductive effects of placing micro-implants of melatonin in the mediobasal hypothalamus (MBH) and preoptic area (POA) were monitored in Soay rams. Groups of animals were initially conditioned to alternating 16 weekly periods of long days (16 h light:8 h darkness; 16L:8D) and short days (8L:16D) for at least 9 months to entrain the seasonal reproductive cycle. All experiments were then initiated at 10 weeks under long days when the animals were sexually inactive. In experiment 1, rams were exposed to short days for 14 weeks or maintained on long days to illustrate the photoperiodically induced re-activation and regression of the reproductive axis. In experiments 2–4, rams received micro-implants of melatonin in the MBH or POA, or received control treatments (sham-operated or no surgery) for 12–14 weeks while maintained on long days (total of 12 animals/treatment). The melatonin implants consisted of 22-gauge stainless-steel cannulae with melatonin fused inside the tip and were placed bilaterally in the brain. Incubation of the implants in Tricine-buffered saline (pH 8·0) at 37 °C showed that the release rate of melatonin was relatively constant after an initial peak in week 1 (means ± s.e.m.: 3·42 ± 0·43 μg/24 h).

Rams with melatonin implants placed in the MBH, but not in the POA, showed a consistently earlier re-activation of the reproductive axis compared with the control animals in all three experiments (12/12 for MBH vs 2/12 for POA). The mean time to maximum testicular diameter was 12·2 ± 0·9, 21·6 ± 1·8 and 22·3 ± 1·2 weeks for the MBH, POA and combined control groups respectively (MBH vs control, P < 0·01; analysis of variance). The premature growth of the testes in the MBH group was associated with an earlier increase in the blood plasma concentrations of FSH and testosterone, and the appearance of the sexual skin coloration. Removal of the implants resulted in a decline in all reproductive parameters. The melatonin treatments did not cause a detectable increase in the peripheral concentrations of melatonin, or affect the diurnal rhythm in melatonin which reflected the long-day photoperiod. When implants containing 125I-labelled melatonin were introduced into the brain the associated radioactivity was localized to within 1 mm of the implants.

The overall results demonstrate that the constant administration of melatonin into the MBH blocks the effect of the endogenous long-day melatonin signal and induces gonadal redevelopment. This provides the first evidence that melatonin acts within or close to the MBH to relay effects of photoperiod and influence the timing of the reproductive cycle in the ram.

Journal of Endocrinology (1992) 132, 201–215