bluuebunny/crossref_metadata_2025 huggingface dataset:
Horse plasma corticotrophin-releasing hormone (CRH): characterisation and lack of a late gestational rise or a plasma CRH-binding protein
Authors: Ellis M J , Livesey J H , Donald R A
Published: December 2008

Immunoreactive corticotrophin-releasing hormone (irCRH) was present in methanolic extracts of equine peripheral blood and showed no elevation in maternal peripheral serum in late gestation (0·54 ±0·25 pmol/l; mean ± s.d.) compared with control horses (0·41 ± 015 pmol/l). The irCRH of methanolic extracts of pituitary venous plasma had a similar elution position following reverse-phase HPLC to synthetic human CRH(1–41) and to irCRH released from horse stalk-median eminence tissue incubated in vitro. Gel chromatographic studies showed no evidence for a plasma CRH-binding protein (CRHBP) analogous to that found in human plasma in either peripheral blood from normal or pregnant horses or in pituitary venous plasma sampled from a cannulated horse. CRH-binding activity was detectable in peripheral plasma from one horse, however the molecular size of this was indicative of a γ-globulin rather than the 37 kDa CRHBP.

These studies suggest that, unlike in the human, CRH does not rise to high values in late gestation nor circulate in a bound form in equine plasma.

Journal of Endocrinology (1994) 143, 455–460
Effects of insulin-like growth factor-I (IGF-I) and IGF-II on the growth of antler cells in vitro
Authors: Sadighi M , Haines S R , Skottner A , Harris A J , Suttie J M
Published: December 2008

The effects of insulin-like growth factors -I and -II (IGF-I and -II) on the growth of undifferentiated (fibroblast zone) cells from the growing tip of red deer velvet antlers and from cells 1·5 cm distal to the growing tip (cartilage zone) were investigated in primary cell culture. The addition of IGF-I or IGF-II to the medium of cultures preincubated in serum-free medium for 24 h increased the rate of [3H]thymidine uptake in a dose-dependent manner in both cell types, with maximal stimulation occurring when 1 nm–30 nm was added. The addition of IGF-II to the incubation medium containing IGF-I did not cause a further increase in [3H]thymidine uptake in either cell type over and above each growth factor alone, indicating that there were unlikely to be synergistic effects of IGF-II on the mitogenicity of IGF-I. Binding studies were carried out using 3 × 105 fibroblast zone cells and cartilage zone cells after they had been incubated in serum-free medium for 24 h. 125I-Labelled IGF-I (10−9 m) in a final volume of 200 μl was added to each culture and incubation carried out at 4 °C for a further hour. 125I-Labelled IGF-I bound specifically to both fibroblasts and cartilage zone cells; binding was displaced by both unlabelled IGF-I and by IGF-I antibody. These findings indicate that IGF-I and IGF-II are important mediators for antler growth in vitro and suggest that in view of correlations between IGF-I and antler growth, IGF is functionally significant in controlling velvet antler growth in vivo.

Journal of Endocrinology (1994) 143, 461–469
The effect of germ cell complement on the presence of oxytocin in the interstitial and seminiferous tubule fluid of the rat testis
Authors: Nicholson H D , Greenfield H M , Frayne J
Published: December 2008

In the rat testis oxytocin has been localized to the Leydig cells, and these cells have been shown to produce oxytocin in vitro. The present study was performed to determine whether oxytocin is present in the interstitial fluid (IF) and seminiferous tubule fluid (TF) of the rat and whether concentrations of the peptide vary within the two compartments following germ cell destruction.

In order to destroy germ cells adult male rats were anaesthetized and their scrotal regions placed in a water bath at 43 °C for 20 min. Control animals were subjected to anaesthesia alone. Groups of 6 animals were killed 3, 7 and 21 days after heat treatment and their testes removed for histological examination or fluid extraction. IF and TF were separated and the oxytocin content of the fluids measured by radioimmunoassay.

Immunoreactive oxytocin was detected in both the IF (100 ±11 pg/ml) and TF (27± 4 pg/ml) of control rats and this immunoreactivity co-eluted with the authentic peptide following HPLC. Three days after heat treatment IF levels of oxytocin were significantly reduced but TF levels of the peptide were significantly increased. These changes were associated with a lack of pachytene spermatocytes in the histological sections. Seven and 21 days after heat treatment the levels of oxytocin in the IF and TF were not significantly different from control levels. Similar changes in IF and TF oxytocin levels were seen in a second experiment when pachytene spermatocytes were removed using the testicular toxicant methoxyacetic acid.

To investigate whether oxytocin passes from the IF into the TF, 3H-oxytocin was infused into the testicular arteries of both control and heat-treated rats and the rats killed at regular intervals over the next 24 h. In both groups of animals <5% of the 3H-oxytocin passed from the IF into the TF.

These data suggest that the seminiferous tubule as well as the Leydig cells may secrete oxytocin and that this secretion may be influenced by the presence of germ cells, particularly pachytene spermatocytes.

Journal of Endocrinology (1994) 143, 471–478
An investigation of the ability of TSH and Graves' immunoglobulin G to increase intracellular calcium in human thyroid cells, rat FRTL-5 thyroid cells and eukaryotic cells transfected with the human TSH receptor
Authors: MacNeil S , Munro D S , Metcalfe R , Cotterell S , Ruban L , Davies R , Weetman A P
Published: December 2008

The purpose of this study was to determine if immunoglobulin G preparations (IgGs) from patients with Graves' disease can increase intracellular calcium in thyroid cells, as has been reported for TSH. Both TSH and Graves' IgGs (prepared by protein G affinity chromatography) increased calcium in a range of thyroid cells; however, the response seen, using Fura-2-loaded coverslips of cell monolayers, varied considerably. Chinese hamster ovary (CHO/JPO9) cells transfected with a high number of human TSH receptors showed the greatest response: TSH (10 mU/ml) increased calcium in 46% of experiments and 18 out of 25 (72%) Graves' IgGs increased calcium at 0·1 mg/ml (significantly greater, P<0·001, than for control IgGs where cells responded to 2 out of 13 preparations). Rat FRTL-5 cells only responded to TSH in 22% of experiments and to 2 out of 8 (25%) of Graves' IgGs. Similarly, human thyroid cells responded to TSH in 22% of experiments and to 2 out of 9 (22%) of Graves' IgGs. (When studying cyclic AMP responses in JPO9 cells, much higher concentrations of Graves' IgGs were required (1–3 mg/ml).) However, higher concentrations (03 mg/ml) of both Graves' IgGs, and to a lesser extent of control IgGs, were capable of increasing calcium in cells both with and without TSH receptors (control CHO cells and normal human dermal fibroblasts). We conclude that relatively low concentrations of patient IgGs can be distinguished from control IgGs in JPO9 cells on the basis of their ability to increase calcium, but that additionally all IgG preparations possibly contain another factor which can increase calcium in a range of cells independent of the presence of the TSH receptor.

Journal of Endocrinology (1994) 143, 527–540
Binding of thyroid hormone to the goat testicular Leydig cell induces the generation of a proteinaceous factor which stimulates androgen release
Authors: Jana N R , Bhattacharya S
Published: December 2008

Leydig cells isolated from goat testis were sonicated and pure nuclear preparations obtained for 125I-3,5,3′-triiodothyronine (T3)-binding assay. Under optimum assay conditions of pH 7·2 at 37 °C and 90 min of incubation, binding of 125I-T3 to Leydig cell nuclei reached saturation at 1·2 nmol/l concentration. A Scatchard analysis of T3 binding exhibited a Kd of 0·535 × 10−9 mol/l and a maximum binding capacity of 1·25 pmol/mg DNA. Competitive inhibition studies showed T3 binding to be analogue specific. The physiological relevance of T3 binding to goat Leydig cell was examined by adding increasing concentrations of T3 to the Leydig cell incubation (1×10 cells/incubation). T3 (10, 25 and 50 ng/ml or 4, 10 and 20 ng/incubation) resulted a dose dependent increase in androgen release and in all cases stimulation of androgen release was statistically significant (P<0·01) compared with control. Stimulation of Leydig cell androgen release by T3 was significantly inhibited by actinomycin-D (P<0·01) and cycloheximide (P<0·01). T3 had additive stimulatory effects on LH-augmented androgen release from Leydig cells. T3 (50 ng/ml or 20 ng/incubation) effected a more than twofold increase in Leydig cell protein synthesis compared with control and both actinomycin-D and cycloheximide (50 μg/ml) inhibited it completely. The data indicated that the stimulatory effect of T3 on androgen release is mediated via T3-induced protein(s). Sub-cellular fractions obtained from T3-treated Leydig cells showed an increase in protein synthesis in mitochondrial and soluble supernatant fractions (100 k sup) and it was only 100 k sup which stimulated androgen release from Leydig cells in separate incubations. Treatment of 100 k sup with trypsin or heat abolished its stimulatory effect. Incubation of Leydig cells with T3 for different times showed an increase in protein synthesis prior to the stimulation of androgen release. The results therefore indicated that T3 binding to Leydig cells induced the generation of a proteinaceous factor(s) which in turn stimulated androgen release.

Journal of Endocrinology (1994) 143, 549–556
The effect of oleic acid on the secretion of thyrotrophin and growth hormone by cultured rat anterior pituitary cells
Authors: Kennedy J A , Nicolson R , Wellby M L
Published: December 2008

Elevation of non-esterified fatty acids (NEFA) in vivo is associated with abnormal control of TSH. To determine whether TSH secretion is directly inhibited by NEFA, as has been reported for GH, cultured rat anterior pituitary cells were exposed for 20 h to oleic acid in medium containing 77 × 10−5 mol/l bovine serum albumin (BSA). In a molar ratio with albumin of 1·2 (total oleic acid 9× 10 mol/l), or greater, oleic acid inhibited basal GH secretion (maximum inhibition to 40% of control) while basal TSH was less affected, a ratio of 3 (2·3 × 10−4 mol/l oleic acid) or greater causing a smaller degree of inhibition (maximum inhibition to 80% of control). In the presence of 10−9 mol/l growth hormone-releasing hormone or 108 mol/l TRH, inhibition was achieved at a ratio of 12 (9 × 10−4 mol/l oleic acid) or greater. Basal TSH was less sensitive to inhibition by thyroxine (T4) in the presence of oleic acid/albumin at a ratio of 6 or greater, and inhibition by oleic acid was less than additive with T4 at a ratio of 6 or greater. Responses to tri-iodothyronine (T3) were unaffected at a ratio of 6 (4·6 × 10−4 mol/l oleic acid), but a ratio of 12 inhibited the effects of both T3 and T4 on TSH. Oleic acid had less effect in the presence of TRH, a ratio of 12 causing a small increase in the threshold concentration of T3 and T4 for TSH inhibition. Further studies are required to determine the mechanism by which oleic acid inhibits the response of basal TSH to T4 as well as the reason for a reduced effect of oleic acid in the presence of TRH. In some critically ill patients, total serum NEFA/albumin ratios from 1·5 to 6 have been reported, indicating that the direct inhibitory effects on TSH observed in vitro occur at free NEFA concentrations achieved in vivo. However, the direct inhibitory effect on TSH may be offset to some extent by reduced responsiveness to T4 at higher oleic acid concentrations. Hence other sites of action of NEFA in vivo may also be important in limiting TSH secretion. Further studies should examine the hypothalamic hormones like TRH and somatostatin, which control the thyrotrophs, as potential sites of action of NEFA.

Journal of Endocrinology (1994) 143, 557–564
Opioid peptides and testicular activity in the lizard Podarcis s. sicula Raf
Authors: Ciarcia G , Facchinetti F , Vallarino M , Pestarino M , Paolucci M , Cardone A , Fasano S , Pierantoni R , Genazzani A R
Published: December 2008

In mammals endorphinergic systems have been shown to modulate reproductive processes and β-endorphin (β-EP) has been found to influence sexual functions, acting at the hypothalamus-pituitary-gonadal axis level. Using immunocytochemical and in vitro studies, evidence for a diffuse pro-opiomelanocortin-related opioid system in the lizard Podarcis s. sicula was produced. In the testis, β-EP immunoreactivity showed seasonal variation, being most pronounced in the interstitial cells of sexually quiescent lizards (December). Reverse-phase high-performance liquid chromatography, coupled with radioimmunoassay and immunocytochemistry, showed that β-EP and acetyl β-EP increased during December, while their concentrations were low during April, when the highest testicular activity occurred.

Using in vivo studies, it was found that naltrexone treatment, blocking pituitary opioid receptor, increased androgen levels in the plasma and in the testis. It was also found with in vitro studies that the endogenous opioid system inhibits gonadotrophin release and therefore androgen production by the testis.

The data reported here provide evidence for the physiological role played by opioid peptides at the pituitary level to regulate the seasonal reproductive activity of the lizard Podarcis s. sicula.

Journal of Endocrinology (1994) 143, 565–571
Effects of basic fibroblastic growth factor on the function and proliferation of human clinically non-functional pituitary adenomas which secreted glycoprotein hormones in vitro
Authors: Atkin S L , Jeffreys R V , Foy P M , Hipkin L , Radcliffe J , White M C
Published: December 2008

The effects of human recombinant basic fibroblastic growth factor (bFGF) on the secretion, viability, proliferation, attachment and morphology of ten dispersed human clinically non-functional (NF) adenomas were examined in vitro. Four clinically NF adenomas secreting FSH and/or LH in vitro were unaffected by 10 nm bFGF over a 4-h period. Over 4 days 10 nm bFGF stimulated LH secretion (66% and 72%, P<0·01) from two out of seven clinically NF adenomas secreting LH, whilst FSH (three tumours) and α-subunit secretion (three tumours) were unaffected. One adenoma co-secreting LH and α-subunit and one secreting LH alone were studied over 21 days; LH secretion fell progressively, but the decline was significantly less (P<0·05) with bFGF (10 nm) treatment after 14 and 21 days in both adenomas, whilst the fall in α-subunit secretion was unaffected by bFGF treatment. A 24-h GnRH test performed at the start and end of the 21-day period in one of these tumours showed an increase in both basal and stimulated LH secretion in the bFGF-treated group over control (124%, P<0·001). There was no effect of bFGF (10 nm) on viability, S-phase proliferation, attachment or morphology of adenoma cells over a 4-day period. These results suggest that bFGF has a role in tumorous LH secretion from these adenomas, but is not mitogenic (at least over 4 days) and is without effect on other parameters of in vitro differentiated function.

Journal of Endocrinology (1995) 144, 173–178
In vivo responsiveness of thyroid glands to TSH in normal and novel 'growth-retarded' mice: transient elevation in normal mice and impairment in 'growth-retarded' mice
Authors: Tomita K , Yoshida T , Morita J , Atsumi S , Totsuka T
Published: December 2008

The in vivo responsiveness of thyroid glands to TSH at various ages in novel 'growth-retarded' (grt/grt) mice derived from Snell's dwarf (DW/J) mice and in their normal counterparts were analysed by determining serum T4 concentrations before and after the administration of exogenous TSH. The serum T4 concentration in normal mice increased in response to TSH at 2, 4 and 12 weeks of age but not at 1 week of age. A transient augmentation of such thyroidal responsiveness to TSH was apparent in normal mice at 2 weeks of age, when the serum T4 level exhibits a peak and the pubertal growth of mice starts. In contrast to normal mice, at any age examined from 2 to 12 weeks after birth, exogenous TSH did not influence serum T4 concentrations in the grt/grt mice at all. On the other hand, serum TSH concentrations in young grt/grt mice were highly elevated compared with those in normal mice and they were normalized by a 2–3 week's treatment with T3. Morphological studies demonstrated degenerated thyroid glands in the grt/grt mice. These results suggest that the severe hypothyroidism and consequent growth retardation in growth-retarded mice are due to impairment of the thyroid glands of the mutant mice in producing and/or secreting thyroid hormones in response to TSH.

Journal of Endocrinology (1995) 144, 209–214
Changes in IGF-I and -II expression and secretion during the proliferation and differentiation of normal rat osteoblasts
Authors: Birnbaum R S , Bowsher R R , Wiren K M
Published: December 2008

IGF-I and -II have potent effects on proliferation and differentiation of osteoblasts in vitro. These cells secrete both IGFs and expression of these peptides is regulated by several of the hormones and growth factors that promote bone resorption and/or formation. However, the physiological role(s) of IGFs in the remodelling process of adult bone is still unclear. Some confusion may arise from results influenced, in part, by differences in the state of osteoblast development of in vitro cultures. Several laboratories have demonstrated that murine osteoblast cultures progress from proliferating preosteoblasts, to mature differentiated osteoblasts that form an extracellular matrix, to cultures that form a mineralized matrix. We have recently documented changes in IGF-binding protein expression and secretion in these cultures. To complement and extend this work, we have examined IGF-I expression and secretion and IGF-II expression during in vitro osteoblast development.

Steady-state mRNA levels of both IGF-I and -II increased from the earliest time examined, day 5 in culture, to a maximum at day 11 and, thereafter, declined. IGF-I secreted into the medium also changed in a biphasic manner, but IGF-II could not be quantitated due to the sensitivity of our assay. Secretion of IGF-I was lowest between days 8 and 14. IGF-I secretion on day 5 was significantly greater than day 8. Similarly, IGF-I secretion from day 17 to 26 was also greater than observed for days 8 to 14. If differentiation of the cells was inhibited, this late rise in IGF-I secretion was abolished. We conclude that IGF-I is an autocrine mitogen of the proliferating preosteoblasts. Further, we also suggest that the rise in IGF-I secretion, late in osteoblast development, may lead to sequestration of this mitogen in the extracellular matrix for release during a subsequent remodelling cycle.

Journal of Endocrinology (1995) 144, 251–259
Inhibin forms in human plasma
Authors: Robertson D M , Sullivan J , Watson M , Cahir N
Published: December 2008

In order to identify the molecular weight forms of bioactive and immunoactive inhibin in human plasma, plasma/serum was sequentially fractionated by immunoaffinity chromatography (using immobilised inhibin α subunit antiserum), reversed phase HPLC and preparative SDS-PAGE. The electroeluted gel fractions were assayed for inhibin in vitro bioactivity and immunoactivity, the latter by RIA. Initial experiments examined human follicular fluid as an inhibin-rich source. Bioactive and immunoactive fractions of 30, 35, 53, 65 and ∼120 kDa were identified in addition to bio-inactive, immunoactive fractions of 26 kDa and 32 kDa. These molecular weights correspond to those of known inhibin forms and are attributed to differing degrees of glycosylation of the inhibin α subunit and variable processing of the α and β inhibin subunits.

Fractionation of male plasma pools revealed the presence of higher molecular weight immunoactive forms (55–120 kDa) as well as 28–31 kDa forms although the molecular weight distribution of activity between pools varied. To assess if the molecular weight pattern was modified by storage and/or subsequent fractionation, protease inhibitors were added initially to plasma and fractionated as above. The molecular weight distribution of immunoactivity was largely unaffected by the treatment, indicating that minimal processing had occurred. Postmenopausal serum itself showed low to undetectable activity. The addition of recombinant human 31 kDa inhibin to postmenopausal serum resulted in a molecular weight profile of inhibin immunoactivity consistent with the presence of 31 kDa inhibin. Fractionation of a serum pool from women undergoing gonadotrophin stimulation, in which inhibin levels were elevated, showed a range of bioactive and immunoactive inhibin forms over the 30–120 kDa range. A good correspondence between activities was observed.

It is concluded that: 1. inhibin exists in plasma/serum as a range (28–120 kDa) of molecular weight forms. 2. In female serum, the majority of inhibin isoforms appear to be bioactive. 3. This fractionation procedure provides a basis for investigating the forms of inhibin in plasma and provides a means of assessing the specificity of new assay methods.

Journal of Endocrinology (1995) 144, 261–269
Inhibition of Leydig cell activity in vivo and in vitro in hypothyroid rats
Authors: Antony F F , Aruldhas M M , Udhayakumar R C R , Maran R R M , Govindarajulu P
Published: December 2008

Leydig cell steroidogenic activity under basal and stimulated conditions was studied in hypothyroid rats. Hypothyroidism was induced at a prepubertal age (30 days postpartum) by surgical thyroidectomy, and l-thyroxine (T4) supplementation (6 μg/100 g body weight/day for 30 days) to hypothyroid rats was begun after 30 days. Hypothyroidism for 60 days reduced serum LH and FSH without affecting prolactin. Serum and intratesticular testosterone and the specific activity of Leydig cell 3β- and 17β-hydroxysteroid dehydrogenases diminished in hypothyroid rats. The stimulatory effect of LH on Leydig cell steroidogenic activity and cAMP was also adversely affected in hypothyroid rats. All these changes were reversed by T4 supplementation. The present results suggest that prepubertal hypothyroidism suppresses both basal and stimulated Leydig cell activity in adult rats.

Journal of Endocrinology (1995) 144, 293–300
Effects of intranasal glucocorticoids on endogenous glucocorticoid peripheral and central function
Authors: Knutsson U , Stierna P , Marcus C , Carlstedt-Duke J , Carlström K , Brönnegård M
Published: December 2008

Glucocorticoids are among the most potent anti-inflammatory agents that can be used in the treatment of rhinitis. Their mechanisms of action are multiple and complex and a number of reports describe significant systemic effects of locally administered glucocorticoids. In order to evaluate the short-term systemic effects of intranasally administered glucocorticoids, 14 normal healthy subjects were treated with two doses of either budesonide (BUD) or fluticasone propionate (FP) for 2 weeks. Before treatment, at regular intervals during the treatment, 1 week and finally 6 weeks after termination of treatment, the effects on glucocorticoid receptor (GR) and methallothionein (MTIIa) mRNA expression levels were examined in peripheral lymphocytes using a solution hybridization assay. Serum cortisol, osteocalcin and urinary cortisol levels were also determined. An insulin tolerance test (ITT) was performed at the end of the second week of treatment and at the end of the 6-week washout period with no statistically significant change in cortisol response. In peripheral lymphocytes, GR mRNA levels were significantly down-regulated. MTIIa mRNA levels increased significantly. Serum osteocalcin decreased significantly during treatment with both BUD and FP. Serum cortisol decreased after 1 week of treatment whereas urinary cortisol was not affected until the second week of treatment. In conclusion, intranasal glucocorticoids at clinically recommended doses have not only significant systemic effects on adrenal function, but also have an effect on specific gene expression in peripheral lymphocytes. These effects are receptor-dependent, reversible, and according to serum and urinary cortisol levels and ITT, leave the hypothalamic-pituitary-adrenal function intact. Finally, these short-term systemic effects were not associated with any of the noticeable side-effects usually observed during long-term treatment with glucocorticoids.

Journal of Endocrinology (1995) 144, 301–310
IGF feedback effects on growth hormone secretion in ewes: evidence for action at the pituitary but not the hypothalamic level
Authors: Fletcher T P , Thomas G B , Dunshea F R , Moore L G , Clarke I J
Published: December 2008

The putative negative feedback effects of IGF-I and IGF-II on GH secretion were tested by intracerebroventricular (icv) and intrapituitary administration to sheep. Over two consecutive days, serial jugular blood samples were taken at 10 min intervals for 6 h from ewes (n=3/group) fitted with indwelling stainless steel cannulae into the lateral or third cerebral ventricles. The sheep were injected (icv) with either vehicle or purified ovine IGF-I (2, 4 or 8 μg). IGF-I injection had no effect on plasma GH secretion. Serial blood samples were taken from a second group of nine ewes in which ovine or recombinant human (rh) IGF-I was infused (2·5 μg/h for 2 h) into the third ventricle; once again, IGF-I failed to affect the episodic pattern of GH secretion. Three ewes fitted with indwelling stainless steel cannulae placed in the anterior pituitary gland were consecutively infused with either ovine or rhIGF-I (2·5 μg/h for 2 h) or vehicle. Plasma GH concentrations were suppressed in 3/3 sheep from 1–1·5 h after the commencement of infusion and GH levels remained low for the remainder of the sampling period. In another group of five ewes synergistic effects of IGF-I and IGF-II on GH secretion were tested by icv infusion of rhIGF-I, rhIGF-II, or rhIGF-I+rhIGF-II (5 μg/h for 2 h) or vehicle (sterile 10 mm HCl/saline). Each sheep received each treatment in a randomised design. Infusion (icv) of IGF-I and IGF-II alone or in combination failed to alter GH secretion.

These observations suggest that IGF-I derived from peripheral tissues may modulate GH release at the pituitary level but that IGF-I acts neither alone nor in conjunction with IGF-II as a negative feedback regulator of GH secretion via the hypothalamus in the ewe.

Journal of Endocrinology (1995) 144, 323–331
Type 1 angiotensin II receptors in rat and human sperm
Authors: Vinson G P , Puddefoot J R , Ho M M , Barker S , Mehta J , Saridogan E , Djahanbakhch O
Published: December 2008

The physiological factors which induce and maintain mammalian sperm maturation and motility generally remain unclear, although several agents are known to be involved. We describe here the application of immunocytochemical and immunoblotting methods to identify the angiotensin II type 1 (AT1) receptor in the tails of ejaculated rat and human sperm. Motility data on stimulated and unstimulated sperm from volunteers and patients attending fertility clinics showed that angiotensin II may increase both the percentage of motile sperm and their linear velocity, while the specific AT1 receptor antagonist DuP753 inhibited the action of angiotensin II on the percentage of motile sperm. In rat seminiferous tubules, AT1 receptors were present in primary spermatogonia and in spermatid tails, but immunoreactivity was not seen in sperm contained in caput or cauda epididymis, showing that AT1 receptor function is regulated during transit through the reproductive tract. Since local tissue reninangiotensin systems are present in both male and female tracts, the data suggest that angiotensin II has a role in the maintenance of sperm function and fertility.

Journal of Endocrinology (1995) 144, 369–
Reduced liver insulin-like growth factor-I gene expression in young zinc-deprived rats is associated with a decrease in liver growth hormone (GH) receptors and serum GH-binding protein
Authors: Ninh N X , Thissen J-P , Maiter D , Adam E , Mulumba N , Ketelslegers J-M
Published: December 2008

Zinc depletion attenuates growth and decreases circulating IGF-I. To investigate the mechanisms responsible for the IGF-I decline, we determined the effects of dietary zinc (Zn) deficiency on body and organ growth, serum IGF-I, serum GH-binding protein (GHBP), liver GH receptors and liver expression of their corresponding gene. After 1 week of adaptation to a normal zinc diet, a zinc-deficient diet (ZD; Zn, 0 p.p.m.) or a zinc-normal diet (CTR; Zn, 75 p.p.m.) was available ad libitum to 4-week-old Wistar rats for 4 weeks. Pair-fed animals (PF) received the zinc-normal diet in the same absolute amount as that consumed the day before by the ZD group. The food intake of ZD and PF rats was reduced by 32% (P<0·001) compared with the CTR group. Zinc depletion specifically reduced body weight gain (−22%, P<0·05), serum IGF-I concentrations (−52%, P<0·001), hepatic GH receptors (−28%; P<0·05) and serum GHBP levels (−51%; P<0·05), compared with the PF group. GH concentrations were reduced in ZD animals compared with CTR rats (P<0·01). The caloric restriction of PF animals also decreased body weight gain (−50%, P<0·001), serum IGF-I concentrations (−21%, P<0·05), liver GH receptors (−38%, P<0·001) and serum GHBP levels (−38%, P<0·01), when compared with the CTR group. Both ZD and PF groups had reduced liver IGF-I and GH receptor/GHBP mRNA levels in comparison with the CTR group (P<0·01). However, only liver IGF-I mRNA levels were specifically reduced by zinc deficiency (ZD vs PF rats; P<0·05). Our observations suggest that beside the decline of GH secretion, decreased hepatic GH receptors and/or GHBP concentrations might be responsible for the decline of circulating IGF-I in ZD animals.

Journal of Endocrinology (1995) 144, 449–456
Differential effect of selective block of α2-adrenoreceptors on plasma levels of tumour necrosis factor-α, interleukin-6 and corticosterone induced by bacterial lipopolysaccharide in mice
Authors: Haskó G , Elenkov I J , Kvetan V , Vizi E S
Published: December 2008

The effect of selective block of α2-adrenoreceptors on plasma levels of tumour necrosis factor-α (TNF-α), interleukin-6 (IL-6) and corticosterone induced by bacterial lipopolysaccharide (LPS) was investigated in mice using ELISA and RIA. It was found that the LPS-induced TNF-α response was significantly blunted in mice pretreated with CH-38083, a novel and highly selective α2-adrenoreceptor antagonist (the α2/α1 ratio is >2000). In contrast, LPS-induced increases in both corticosterone and IL-6 plasma levels were further increased by CH-38083. Since it has recently been shown that the selective block of α2-adrenoreceptors located on noradrenergic axon terminals resulted in an increase in the release of noradrenaline (NA), both in the central and peripheral nervous systems, and, in our experiments, that propranolol prevented the effect of α2-adrenoreceptor blockade on TNF-α plasma levels induced by LPS, it seems likely that the excessive stimulation by NA of β-adrenoreceptors located on cytokine-secreting immune cells is responsible for this action. Since it is generally accepted that increased production of TNF-α is involved in the pathogenesis of inflammation and endotoxin shock on the one hand, and corticosterone and even IL-6 are known to possess anti-inflammatory properties on the other hand, it is suggested that the selective block of α2-adrenoreceptors might be beneficial in the treatment of inflammation and/or endotoxin shock.

Journal of Endocrinology (1995) 144, 457–462
Effects of indigestible dextrin on glucose tolerance in rats
Authors: Wakabayashi S , Kishimoto Y , Matsuoka A
Published: December 2008

A recently developed indigestible dextrin (IDex) was studied for its effects on glucose tolerance in male Sprague–Dawley rats. IDex is a low viscosity, water-soluble dietary fibre obtained by heating and enzyme treatment of potato starch. It has an average molecular weight of 1600.

An oral glucose tolerance test was conducted with 8-week-old rats to evaluate the effects of IDex on the increase in plasma glucose and insulin levels after a single administration of various sugars (1·5 g/kg body weight). The increase in both plasma glucose and insulin levels following sucrose, maltose and maltodextrin loading was significantly reduced by IDex (0·15 g/kg body weight). This effect was not noted following glucose, high fructose syrup and lactose loading.

To evaluate the effects of continual IDex ingestion on glucose tolerance, 5-week-old rats were kept for 8 weeks on a stock diet, a high sucrose diet or an IDex-supplemented high sucrose diet. An oral glucose (1·5 g/kg body weight) tolerance test was conducted in week 8. Increases in both plasma glucose and insulin levels following glucose loading were higher in the rats given a high sucrose diet than in the rats fed a stock diet. However, when IDex was included in the high sucrose diet, the impairment of glucose tolerance was alleviated. Moreover, IDex feeding also significantly reduced accumulation of body fat, regardless of changes in body weight.

These findings suggest that IDex not only improves glucose tolerance following sucrose, maltose and maltodextrin loading but also stops progressive decrease in glucose tolerance by preventing a high sucrose diet from causing obesity.

Journal of Endocrinology (1995) 144, 533–538
Central effects of growth hormone-releasing hexapeptide (GHRP-6) on growth hormone release are inhibited by central somatostatin action
Authors: Fairhall K M , Mynett A , Robinson I C A F
Published: December 2008

Growth hormone (GH) release is stimulated by a variety of synthetic secretagogues, of which growth hormone-releasing hexapeptide (GHRP-6) has been most thoroughly studied; it is thought to have actions at both pituitary and hypothalamic sites. To evaluate the central actions of this peptide, we have studied GH release in response to direct i.c.v. injections in anaesthetized guinea pigs. GHRP-6 (0·04–1 μg) stimulated GH release >10-fold 30–40 min after i.c.v. injection. The same GH response required >20-fold more GHRP-6 when given by i.v. injection. GH release could also be elicited by a non-peptide GHRP analogue (L-692,585, 1 μg i.c.v.), whereas a growth hormone-releasing factor (GRF) analogue (human GRF 27Nle(1–29)NH2, 2 μg, i.c.v.) was ineffective. A long acting somatostatin analogue (Sandostatin, SMS 201–995, 10 μg i.c.v.) (SMS) given 20 min before 200 ng GHRP-6 blocked GH release. This was unlikely to be due to a direct effect of SMS leaking out to the pituitary, since central SMS injections did not affect basal GH release, nor did they block GH release in response to i.v. GRF injections. We conclude that the hypothalamus is a major target for GHRP-6 in vivo. Since the GH release induced by central GHRP-6 injections can be inhibited by a central action of somatostatin, and other data indicate that GHRP-6 activates GRF neurones, we suggest that somatostatin may block this activation via receptors known to be located on or near the GRF cells themselves. Somatostatin may therefore be a functional antagonist of GHRP-6 acting centrally, as well as at the pituitary gland.

Journal of Endocrinology (1995) 144, 555–560
Increasing gestational age and cortisol alter the ratio of ACTH precursors:ACTH secreted from the anterior pituitary of the fetal sheep
Authors: McMillen I C , Merei J J , White A , Crosby S , Schwartz J
Published: December 2008

We have used a perifusion system and slices of the anterior pituitary of the fetal sheep combined with specific immunoradiometric assays to investigate the effect of increasing gestational age and cortisol infusion on the output of ACTH(1–39) and the ACTH precursors, proACTH and pro-opiomelanocortin, from the fetal sheep pituitary. Two slices from each fetal anterior pituitary at 106–113 days (n=3), 120–136 days (n=5) and 140–143 days (n=5) of gestation were used. Slices from each anterior pituitary were perifused with the perifusion buffer for at least 120 min prior to the infusion of cortisol (100 nm) for 30 min or buffer alone (control). The anterior pituitary output (fmol/5 min per mg pituitary) of ACTH(1–39) and the ACTH precursors were measured using specific immunoradiometric assays. There was a significant increase in the anterior pituitary secretion rate of ACTH(1–39) between 120 and 136 days (1·04 ±0·23 fmol/5 min per mg) and between 140 and 143 days of gestation (3·08 ±0·33 fmol/5 min per mg). In contrast, there was no change in the secretory rate of the ACTH precursors between 105 and 143 days of gestation. The ratio of the anterior pituitary output of the ACTH precursors:ACTH(1–39) therefore decreased between 120 and 143 of days gestation from 19·10 ±2·05 to 6·36 ± 0·58. There was no effect of cortisol infusion on the anterior pituitary secretion of either ACTH(1–39) or the ACTH precursors before 116 days of gestation. After 120 days, the anterior pituitary output of ACTH(1–39) was significantly decreased by cortisol with the maximal change (43 ± 7%) occurring 10–15 min after the start of cortisol inclusion in the perifusate. Cortisol also altered the secretion of ACTH precursors. Although there was no significant effect with respect to baseline secretion rates, precursor secretion was elevated at the beginning of perifusion with cortisol, compared with precursor secretion after cortisol. The ratio of the anterior pituitary output of ACTH precursors:ACTH(1–39) increased from basal values of 16 ±4 and 12 ±4 (precortisol infusion) to 48 ± 14 at 15 min after the start and 40 ± 14 at 45 min after the end of the cortisol infusion. The differential effects of increasing gestational age and cortisol infusion on the output of ACTH(1–39) and the ACTH precursors may be explained by a change in the functional populations of corticotrophs in the fetal sheep anterior pituitary. These changes may be important in the stimulation of the fetal adrenal cortex which occurs before delivery.

Journal of Endocrinology (1995) 144, 569–576
Oxytocin antagonists delay the initiation of parturition and prolong its active phase in rats
Authors: Antonijevic I A , Douglas A J , Dye S , Bicknell R J , Leng G , Russell J A
Published: December 2008

The physiological importance of oxytocin for the initiation and maintenance of labour and delivery is controversial. We investigated the effects of two novel peptide oxytocin antagonists on the onset and the progress of delivery in rats implanted with a jugular vein cannula one day before term. During delivery rats were given either an oxytocin antagonist (OVT16, n=10, or F382, n=7, 30 μg/kg) or vehicle (n=10, 9) after the birth of the second pup and the time to deliver five more pups was recorded. Other rats were given an injection of F382 (30 μg/kg, n=7) or vehicle (n=9) after the birth of the fourth pup and the time to deliver three more pups was recorded. In another experiment rats were given repeated injections of F382 (30 or 60 μg/kg, n=13, 11) or vehicle (n=32) prepartum on the day of expected term and the time of onset and the progress of delivery was recorded.

Rats given an antagonist after the second pup delivered the next five pups in 100 ± 8 min (F382) and 83 ± 12 min (OVT16), significantly slower than the respective controls (51 ± 6 and 49 ± 6 min, U-test, P<0·05). Four of the 7 rats given F382 after the fourth pup showed no prolongation of delivery (time between pups 4–7: 24·7±2·9 vs 27·5±3·1 min in controls), while in the other three rats delivery was prolonged (time between pups 4–7: 86 ± 4·3 min). The onset of delivery was significantly delayed in rats given repeated injections of F382 at 60 μg/kg on the afternoon of expected delivery but before labour had begun (median shift 14·5 h vs controls, U-test, P<0·05). These results suggest that in rats oxytocin is involved in both the initiation and the maintenance of parturition and that it is more important during the early rather than the later stages of delivery.

Journal of Endocrinology (1995) 145, 97–103
Epitope mapping and analysis of a growth-enhancing monoclonal antibody by limited tryptic digestion of porcine GH
Authors: Shieh H-M , Bass R T , Wang B S , Corbett M J , Buckwalter B L
Published: December 2008

In this study, the epitope of a murine PS-7·6 monoclonal antibody (mAb) which was raised against the recombinant porcine GH (pGH) and subsequently shown to enhance the growth-promoting activity of pGH in a hypophysectomized rat model, was mapped by the limited tryptic digestion of pGH. A pGH fragment corresponding to amino acid residues 70–95 was separated by reverse-phase HPLC and also immunoprecipitated by PS-7·6 mAb. This fragment was found in an RIA to compete with radiolabelled pGH for the binding of PS-7·6 mAb in a dose-dependent fashion. Several peptides covering this potential epitope region of pGH(70–95) were synthesized and assayed by competitive RIA. The results suggested that pGH(75–90) was the optimal sequence recognized by PS-7·6 mAb. Sequential alanine substitution of each residue of pGH(75–90) revealed that the side chains of Leu76, Ile83 and Leu87 were critical for binding to PS-7·6 mAb. Other residues could be replaced by alanine without substantially altering the binding affinity. The region of amino acids 75–95 comprises the C-terminal end of the second helix of pGH and the repeating pattern of i and i+3 (i+7) of the critical amino acids appears consistent with PS-7·6 mAb binding to the hydrophobic side of the helix. The sequence and the helical structure of the epitope of PS-7·6 mAb provide the basis for designing the effective peptide vaccines to enhance the growth performance of animals.

Journal of Endocrinology (1995) 145, 169–174
Corpuscles of Stannius and blood flow regulation in freshwater North American eels, Anguilla rostrata LeSueur
Authors: Butler D G , Oudit G Y
Published: December 2008

Cardiac output (CO), heart rate (HR), stroke volume (SV), dorsal aortic blood flow (DABF), dorsal aortic blood pressure (PDA) and plasma electrolytes were monitored in stanniectomized and sham-operated freshwater eels over a 3-week period; branchial shunting and systemic resistance (RSYS) were estimated. DABF was significantly reduced by 45% from 11·72±0·48 (control) to 6·55±0·41 (n=6; day 21) ml.min−1.kg−1 within 3 weeks after the removal of the corpuscles of Stannius. This large reduction in blood flow was due to a 25% decrease in CO and a 100% increase in estimated branchial shunting which preceded the fall in CO. CO was decreased from 16·07 ±0·31 (control) to 11·91 ±1 (n=6; day 21) ml.min−1.kg−1 through a reduction in SV; there was no significant change in HR. Estimated branchial shunting, a relative measure of branchial arterio-venous blood flow, corresponded to 2·53±0·18 ml.min−1.kg−1 (control; n=12), which represents 16% of baseline CO. Ventral and dorsal aortic pulse flows also decreased following stanniectomy. The decrease in DABF occurred in conjunction with a reduction in PDA which was measured for 12 days in a separate group of eels. Baseline PDA (3·03 ±0·1 kPa) significantly decreased by 15% to 2·55 ±0·13 kPa 4 days after stanniectomy. However, this fall in PDA was transient and accompanied by an elevation in derived RSYS. These results support the hypothesis that the corpuscles of Stannius are closely linked to cardiovascular regulation in freshwater eels. Electrolyte changes (hypercalcemia, hypomagnesia, hyperkalemia and hyponatremia) were temporally coupled to the changes in blood flows. Impaired cardiovascular function and altered patterns of blood flow to osmoregulatory organs such as the gills, kidney and skin may have led to some or all of the electrolyte disturbances which followed stanniectomy.

Journal of Endocrinology (1995) 145, 181–194
Regulation of gill prolactin receptors in tilapia (Oreochromis niloticus) after a change in salinity or hypophysectomy
Authors: Auperin B , Rentier-Delrue F , Martial J A , Prunet P
Published: December 2008

Prolactin (PRL) receptors in gill tissue have been analyzed in tilapia (Oreochromis niloticus) after transfer from fresh water (FW) to brackish water (BW). This study has indicated the presence of only one class of tilapia PRL (tiPRL) receptor whatever the salinity. After transfer, however, the percentage of specific binding of the two forms of tiPRL (tiPRLI and tiPRLII) increased significantly. Scatchard analysis of tiPRLI binding indicated an increase in receptor affinity, an effect which was not accompanied by any change in receptor specificity. Transfer to BW also caused the number of tiPRL receptors to increase rapidly, remaining high in fish adapted to BW for 28 days. Based on the sharp reduction in plasma tiPRLI and tiPRLII levels after transfer to BW, one possible explanation may be that tiPRL itself is an important factor regulating the number of free receptors. This hypothesis finds support in the fact that the number of tiPRL receptors also increased in hypophysectomized fish reared in FW. However, the absence of change in receptor affinity after hypophysectomy suggested that yet other factors are involved in tiPRL receptor regulation during the transfer from FW to BW. The paradoxically high numbers of tiPRL receptors in the gills of BW-adapted tilapia, even though PRL is known to be a FW-adapting hormone, is discussed with regard to the environment in which tilapia live.

Journal of Endocrinology (1995) 145, 213–220
Effects of perinatal thyroid hormone deprivation on the growth and behaviour of newborn mice
Authors: Wallace H , Pate A , Bishop J O
Published: December 2008

Experiments were designed to distinguish between neonatal effects due to maternal thyroxine (T4) deprivation and those due to autonomous (fetus/pup) T4 deprivation, employing mice heterozygous for the bTG-tk transgene TG66·19 which specifically directs high-level expression of herpes virus type I thymidine kinase to the thyrocytes. Heterozygous TG66·19 females were either untreated or Ganciclovir was administered to destroy their thyrocytes and so render them T4-deficient. When mated to normal males these heterozygous females are expected to produce on average 50% normal and 50% heterozygous transgenic conceptuses. Ganciclovir was administered to the dams (both untreated and Ganciclovir-pretreated) during days 14–18 of gestation. At optimum levels of in utero Ganciclovir administration the non-transgenic pups showed no discernible effect while the transgenic pups were rendered athyrocytic and completely T4-deficient. The dams pretreated with Ganciclovir are hypothyroid throughout gestation, while the dams to which Ganciclovir was administered for the first time during gestation are not expected to become hypothyroid until about the time of parturition. In this way four sets of pups were generated for purposes of comparison: hypothyroid (transgenic) and euthyroid (non-transgenic) pups born to euthyroid dams and hypothyroid and euthyroid pups born to hypothyroid (Ganciclovir-pretreated) dams. Normal growth during days 1–10 after birth was dominated by the T4 status of the dam during gestation. Growth during days 11–21 and the correct timing of eye opening and ear elevation were dominated by the autonomous T4 status of the fetus/pup. The timely development of the surface-righting reflex (relative to weight gain) was shown to require both maternal and fetus/pup T4. The development of the cliff-avoidance reflex was independent of the T4 status of both pup and dam and of pup weight. The size of the pups at birth depended primarily on a normal T4 status in the dam but surprisingly T4 deficiency in fetuses/pups partly compensated for maternal T4 deficiency. The results presented here clearly demonstrate the utility of the HSV-tk-transgene-Ganciclovir-administration protocol in studying the interplay of maternal and fetal T4 deprivation in rodents.

Journal of Endocrinology (1995) 145, 251–262