bluuebunny/crossref_metadata_2025 huggingface dataset:
Radioimmunoassay of insulin-like growth factor-binding protein-6 in human serum and other body fluids
Authors: Baxter R. C. , Saunders H.
Published: December 2008

A radioimmunoassay has been established for the insulin-like growth factor-binding protein, IGFBP-6, isolated from a human transformed fibroblast cell-line. The binding proteins IGFBP-I and IGFBP-3 did not cross-react, but both IGF-I and IGF-II markedly inhibited IGFBP-6 tracer binding to antiserum. This inhibition, greater for IGF-II than for IGF-I, was fully reversed by the addition of IGFBP-3 to sequester the IGFs. After fractionation of human serum and follicular fluid samples by gel chromatography, interference in the radioimmunoassay by fractions corresponding to the 150 kDa IGF-IGFBP complex could be eliminated by IGFBP-3. The equivalent fractions from cerebrospinal fluid and amniotic fluid fractionation did not interfere in the assay. The mean IGFBP-6 level in adult human serum was 0·221 ±0·110 mg/l, with values significantly higher in men than women, and slightly decreased in pregnancy. Similar values were seen in umbilical cord serum and in amniotic and follicular fluid samples, while the mean level in cerebrospinal fluid was slightly lower, 0·152±0·049 mg/l. This assay will facilitate studies on the regulation of IGFBP-6 production, and its role as an IGF carrier.

Journal of Endocrinology (1992) 134, 133–139
Effects of the daily administration of gonadotrophin-releasing hormone on the anterior pituitary gland of rats with restricted feeding
Authors: Kotsuji F. , Hosokawa K. , Tominaga T.
Published: December 2008

To investigate the influence of weight reduction on pituitary function and its modulation by gonadotrophin-releasing hormone (GnRH), female rats were restricted to 10 g food/day for 60 days. GnRH (5 μg) or saline (0·2 ml) were administered daily between days 31 and 60 of the period of underfeeding. Underfeeding brought about a decrease in the pituitary gonadotrophin content, serum levels of gonadotrophins and oestradiol, and the number and size of both LH- and FSH-positive pituitary cells. The administration of GnRH to underfed rats produced an increase in the pituitary and serum gonadotrophin levels and the number and size of both LH- and FSH-positive pituitary cells. These observations suggest that underfeeding and/or weight loss diminish the number and activity of the pituitary gonadotrophs, and that daily administration of GnRH both increases the number of gonadotrophs and augments their activity.

Journal of Endocrinology (1992) 134, 177–182
Lung to blood passage of human growth hormone after intratracheal instillation: stimulation of growth in hypophysectomized rats
Authors: Folkesson H. G. , Hedin L. , Weström B. R.
Published: December 2008

The passage from the lower respiratory tract into the blood of human GH (hGH; Mr = 22 000) and bovine serum albumin (BSA; Mr = 67 000) was assessed after intratracheal instillation in adult rats. The plasma level of immunoreactive hGH reached its maximum at 0·5–1 h after instillation and had almost disappeared within 24 h. Higher plasma levels were obtained in male rats than in female rats resulting in a higher total lung passage of hGH in male rats than in female rats (means±s.d.; 6·0± 1·7% vs 3·3±1·2%, P <0·01). The plasma level of BSA showed a different pattern, with a maximum at 16–24 h after instillation and a total lung passage of 4·3 ± 1·7% of the given dose for both sexes. The plasma levels of hGH increased non-linearly with increasing dose instilled in the dose range 36–720 μg/kg body weight. When hGH was instilled daily at a dose of 720 μg/kg body weight to hypophysectomized rats for 1 week, they responded with a significant increase in body weight when compared with hypophysectomized control rats (16·8±4·2 g vs −1·8±2·4 g, P <0·001).

The results demonstrate that, despite their different molecular weights, hGH and BSA pass through the lower respiratory tract into the circulation with similar efficiencies in the rat. However, the lung passage of hGH, unlike that of BSA, showed sexual dependency, an earlier plasma concentration maximum and a tendency of the passage to saturation with increasing dose instilled. Since hGH was shown to be biologically active after the passage through the lungs, the lung may be a potential new route for hGH administration.

Journal of Endocrinology (1992) 134, 197–203
Pituitary-testis function in rats treated neonatally with a gonadotrophin-releasing hormone agonist: short- and long-term effects
Authors: Garnelo P. , Pinilla L. , Gaytan F. , Aguilar E.
Published: December 2008

Acute and long-term effects of neonatal and prepubertal treatments with an LH-releasing hormone agonist (LHRH-A) were studied in Wistar male rats. Animals injected with d-Ala6-d-Gly10-LHRH ethylamide (2 μg/kg per day) or vehicle from days 1 to 15 or from days 16 to 29 were killed at different ages. Treatment between days 1 and 15 induced a decrease in both pituitary FSH and LH content as well as a reduction in plasma FSH and blockade of the response to LHRH. These effects were apparent on day 16 after treatment. Basal and human chorionic gonadotrophin (hCG)-stimulated progesterone and testosterone secretion in vitro was similar in testes from male rats treated with LHRH-A or vehicle. Reduced testicular weight was observed until day 90, whereas puberty, spermatogenesis and fertility were unaffected. The decrease in plasma FSH concentrations after neonatal treatment with LHRH-A was also found in groups of animals killed on day 10 and was possibly the cause of reduced testicular weight, since treatment with FSH from day 1 to day 15 blocked the effect of LHRH-A. Likewise, treatment with LHRH-A from day 1 to day 15 also reduced FSH and LH secretion in males orchidectomized on day 1 of life.

Animals injected with LHRH-A from day 15 to day 29 exhibited, at the end of the treatment period, reduced testicular weight, and decreased pituitary gonadotrophin content and plasma FSH concentrations, whereas LH plasma concentrations were normal. In adulthood, the pituitary-testis function did not vary from normal. Our results demonstrate that: (1) administration of LHRH-A from day 1 to day 15 of life desensitized the gonadotrophs, which in turn lowered plasma gonadotrophin concentrations and caused a long-term reduction in testis weight without changes in the quality of spermatogenesis or reproductive activity in adulthood; (2) chronic treatment with LHRH-A during the neonatal period did not result in a steroidogenic lesion; and (3) administration of LHRH-A from day 15 to day 29 produced only a transient reduction in testicular weight and in spermatogenesis.

Journal of Endocrinology (1992) 134, 269–277
Bioactive and immunoreactive FSH and immunoreactive inhibin concentrations in the ovine fetus
Authors: Phillips D. J. , Smith P. R. , Heath D. A. , Condell L. A. , McNatty K. P.
Published: December 2008

The bioactive (B) and immunoreactive (I) pituitary contents/concentrations of FSH, together with the plasma concentrations of B-FSH, I-FSH and I-inhibin were determined in ovine fetuses at days 55, 75, 90 and 135 of gestation (day 145 = term). The pituitary contents and concentrations of B-FSH and I-FSH increased in both sexes with gestational age. The female fetuses had significantly (P <0·01) higher pituitary contents/concentrations of B-FSH and I-FSH than the male fetuses at days 75 and 135. The pituitary B/I ratios of FSH were not significantly different with age or sex. The plasma concentrations of B-FSH remained relatively constant from days 75 to 135, with no significant differences between sexes or with age. In contrast, the plasma concentrations of I-FSH reached a peak at day 90 and then declined towards term in both sexes. At all gestational ages except day 55, the female fetuses had significantly (P <0·05) higher plasma concentrations of I-FSH than the males. In both sexes, the plasma B/I ratios of FSH were lowest at day 90 and had increased again by day 135, with the male fetuses having significantly (P <0·05) higher B/I ratios compared with the female group at days 75 and 135 but not at day 90. At all gestational ages, the plasma concentrations of I-inhibin declined throughout gestation in the female fetuses, whereas in the males they reached a nadir at day 75 and then increased towards term. The concentrations of I-inhibin were significantly (P <0·01) higher in the male fetuses compared with the females. Collectively, these data suggest that there are changes in the forms of FSH present in the pituitary gland and plasma throughout gestation in the ovine fetus. Moreover, they infer that the difference between the sexes in FSH synthesis and/or secretion may be attributed in part to the circulating concentrations of inhibin.

Journal of Endocrinology (1992) 134, 287–295
Inhibitory action of somatostatin-14 on hormone-stimulated cyclic adenosine monophosphate induction in porcine granulosa and luteal cells
Authors: Rajkumar K. , Kerr D. E. , Kirkwood R. N. , Laarveld B.
Published: December 2008

Somatostatin-14 (SRIF-14) inhibited, in a concentration-dependent manner, LH- and forskolin-stimulated cyclic adenosine monophosphate (cAMP) induction in porcine granulosa and luteal cells. The inhibitory effect of SRIF-14 on hormone-induced cAMP generation was more potent in porcine ovarian cells than in the GH-3 pituitary cell line. The inhibitory effect of SRIF-14 was impeded by neutralizing its biological activity with specific antiserum. Preincubation of luteal and granulosa cells with phorbol 12-myristate 13-acetate (PMA) enhanced LH- and forskolin-stimulated cAMP levels. SRIF-14 failed to inhibit LH- or forskolin-stimulated cAMP levels in cells preincubated with PMA. It is concluded that SRIF-14 inhibits hormone-stimulated cAMP induction in the porcine ovary. LH-induced protein kinase C activation may be physiologically important to alleviate the inhibitory effects of SRIF-14.

Journal of Endocrinology (1992) 134, 297–306
Distribution of relaxin between human maternal and fetal circulations and amniotic fluid
Authors: Johnson M. R. , Abbas A. , Nicolaides K. H. , Lightman S. L.
Published: December 2008

Relaxin was measured in maternal blood and amniotic fluid samples at 9–40 weeks and in fetal blood samples at 19–41 weeks of pregnancy. In amniotic fluid, concentrations of relaxin rose from 58 ng/1 (geometric mean) at 10 weeks to 142 ng/l at 14 weeks and declined subsequently to 55 ng/l at 22 weeks. In maternal blood, mean relaxin concentrations were ten times greater than in amniotic fluid, and concentrations decreased with gestation. Since there was no significant association between the relaxin concentrations in the two compartments, relaxin in the amniotic fluid may be derived from the decidualized endometrium rather than the maternal circulation, alternatively its metabolism may be different in the two compartments. The absence of detectable concentrations of relaxin in any of the fetal blood samples demonstrates that there is no significant placental transfer or fetal synthesis of this peptide.

Journal of Endocrinology (1992) 134, 313–317
The effect of gonadotrophin surge-inhibiting factor on the self-priming action of gonadotrophin-releasing hormone in female rats in vitro
Authors: Koppenaal D. W. , Tijssen A. M. I. , de Koning J.
Published: December 2008

The present study was designed to explore further the functional antagonism between gonadotrophin-releasing hormone (GnRH) and the ovarian factor, gonadotrophin surge-inhibiting factor (GnSIF). In all experiments, pituitary tissue was exposed to various amounts of GnSIF, after which the self-priming action of GnRH was studied. GnSIF was increased in vivo by FSH treatment and increased in vitro by adding various amounts of follicular fluid (FF) to cultured pituitary cells.

Treatment with 3 or 10 IU FSH suppressed the initial LH response and delayed the maximally primed LH response to GnRH. Treatment with FSH was only effective in intact rats on days 1 and 2 of dioestrus. There was no difference in the rate of maximal LH release irrespective of treatment with either FSH or saline. Since FSH treatment was ineffective in long-term ovariectomized rats, it was concluded that the initial suppressive effect of FSH on LH release was mediated by GnSIF.

Cycloheximide prevented the self-priming action of GnRH by inhibiting GnRH-induced protein synthesis. The initial protein synthesis-independent GnRH-stimulated LH release, which was already suppressed by FSH treatment, remained suppressed in the presence of cycloheximide. Pretreatment with GnRH in vivo increased the protein synthesis-independent GnRH-induced LH release during subsequent incubation of the glands. This increase did not occur after FSH treatment.

Pituitary cells, cultured for 20 h in medium only, failed to elicit the self-priming effect of GnRH. Preincubation with FF maintained the self-priming effect. This was independent of the concomitant presence of various amounts of oestradiol. Preincubation with bovine FF suppressed the initial GnRH-stimulated LH release dose-dependently. Porcine FF, human FF and testicular extract suppressed the release of LH in a similar way.

It was concluded that GnSIF suppresses the initial LH response to continuous GnRH stimulation. Increased levels of GnSIF caused by FSH treatment also delayed the primed LH release. The mechanism of functional antagonism between GnSIF and GnRH could give rise to the occurrence of the phenomenon of GnRH self-priming.

Journal of Endocrinology (1992) 134, 427–436
The effects of systemic insulin, insulin-like growth factor-I and growth hormone on bone growth and turnover in spontaneously diabetic BB rats
Authors: Verhaeghe J. , Suiker A. M. H. , Visser W. J. , Van Herck E. , Van Bree R. , Bouillon R.
Published: December 2008

Spontaneously diabetic BB rats have a markedly depressed longitudinal bone growth and bone formation/turnover. In this study, male diabetic BB rats were infused intraperitoneally or subcutaneously for 2 weeks with hormones that are believed to stimulate skeletal growth and/or trabecular bone formation: insulin (3 or 4 U/day), human GH (hGH; 400 mU/day), recombinant human insulin-like growth factor-I (rhIGF-I; 300 or 600 μg/day) and testosterone (80 μg/100 g body weight per day).

Saline-treated diabetic BB rats had decreased plasma concentrations of IGF-I and osteocalcin (OC) (OC, 3·7 ±0·3 vs 13·1 ± 0·8 (s.e.m.) nmol/l in controls); bone histomorphometry showed decreased epiphyseal width, osteoblast surface (0·04±0·04 vs 1·5±0·3%) and osteoid surface, and mineral apposition rate (MAR) (1·8±0·5 vs 7·9±0·6 μm/day).

Testosterone and hGH infusions had no effect on weight loss or on decreased skeletal growth and bone formation of diabetic rats, nor did they increase plasma IGF-I concentrations. Insulin infusions into diabetic rats resulted in hyperinsulinaemia and accelerated weight gain. The epiphyseal width, osteoblast/osteoid surfaces and OC levels of insulin-treated rats were normalized or stimulated well above control values (osteoblast surface, 4·3 ±0·8%; plasma OC, 16·1 ± 1·4 nmol/l); the MAR (4·0 ± 0·9 μm/day) was only partly corrected after the 2-week infusion. Infusions of rhIGF-I into diabetic rats doubled but did not restore plasma IGF-I levels to normal; weight gain, however, was similar to that in control rats. IGF-I treatment had no effect on epiphyseal width, osteoblast/osteoid surfaces and OC concentrations, but improved the decreased MAR (4·6±1·2 μm/day).

These results indicate (1) that the decreased epiphyseal width and osteoblast recruitment of diabetic BB rats are directly related to their insulin deficiency, and (2) that IGF-I, administered systemically, corrects, in part, the decreased MAR in diabetes, suggesting a role in osteoblast function and/or mineralization.

Journal of Endocrinology (1992) 134, 485–492
11β-Hydroxysteroid dehydrogenase in the rat ovary: high expression in the oocyte
Authors: Benediktsson R. , Yau J. L. W. , Low S. , Brett L. P. , Cooke B. E. , Edwards C. R. W. , Seckl J. R.
Published: December 2008

The enzyme 11β-hydroxysteroid dehydrogenase (11β-HSD) catalyses the conversion of physiological glucocorticoids to inactive products, thus modifying the access of glucocorticoids to glucocorticoid and mineralocorticoid receptors. Glucocorticoids may affect ovarian function both indirectly and via binding to ovarian receptors. We have demonstrated 11β-HSD bioactivity and mRNA expression in rat ovary in vitro. The enzyme was localized to oocytes and luteal bodies immunohistochemically using two antibodies raised against purified rat liver 11β-HSD. These data are supported by in-situ hybridization studies, which also localized 11β-HSD mRNA expression to oocytes and luteal bodies. The results suggest that 11β-HSD may modulate the effects of glucocorticoid on ovarian function.

Journal of Endocrinology (1992) 135, 53–58
Fluid balance, electrolyte profiles and plasma parathyroid hormone concentrations in ewes treated with epidermal growth factor
Authors: Gow C. B. , Wilkinson M. , Silvapulle M. J. , Moore G. P. M.
Published: December 2008

The infusion of low doses of epidermal growth factor (EGF) into lactating ewes stimulates water intake and urine volume. The plasma concentrations and daily output of various electrolytes in milk and urine are also affected. We have investigated this further by recording the effects of EGF infusion on fluid balance, electrolyte profiles and plasma concentrations of glucose and parathyroid hormone (PTH) in non-pregnant, non-lactating ewes. Twenty-four animals (n= 8 per group) received infusions of 100 ml saline/day into the jugular vein for 10 days (days 1–10) followed by EGF at a dose rate of either 1 (low dose), 5 (medium dose) or 10 (high dose) μg/kg liveweight per day in 100 ml saline for 5 days (days 11–15). All ewes then received an infusion of 100 ml saline/day for 10 days (days 16–25).

Most plasma and urine samples had undetectable concentrations of EGF-immunoreactive material during the periods of saline infusion. During EGF infusion, the highest amounts of EGF infusate excreted in urine were 1·6, 5·9 and 5·6% for ewes in low, medium and high dose groups respectively. Water intake increased by 17% (0·5 kg), 88% (2·5 kg) and 89% (2·3 kg) and urine volume increased by 29% (0·5 kg), 108% (2·2 kg) and 134% (2·1 kg) for the three groups respectively. Fluid balance and feed intake were not affected by EGF infusion, but the output of faecal dry matter was reduced in ewes receiving the two higher doses of EGF.

All levels of EGF resulted in hypocalcaemia, increased plasma PTH concentrations and hypermagnesaemia. There was no effect of EGF on plasma concentrations of K+ and glucose or on daily urinary excretion of K+ and Mg2+. The only response to the low dose was a reduced plasma concentration of Na+ and an increased daily urinary urate excretion. The two higher doses increased the daily urinary excretion of Na+, PO43− and urate, but had no effect on the respective concentrations in plasma. Urinary Ca2+ excretion was reduced only during infusion of the medium dose of EGF.

The responses of most variables were similar during infusion of the medium and high doses of EGF. All three doses of EGF induced polydipsic and diuretic responses in ewes, and infusions of 5–10 μg EGF/kg liveweight per day affected renal excretion of Ca2+, Na+ and PO3−4. We interpret the responses of the kidney and plasma PTH concentrations as a means of maintaining the homeostasis of plasma profiles of electrolytes.

Journal of Endocrinology (1992) 135, 91–101
Comparison of specific immunoassays for detection of the β-core human chorionic gonadotrophin fragment in body fluids
Authors: de Medeiros S. F. , Amato F. , Matthews C. D. , Norman R. J.
Published: December 2008

We have validated two new methods, one radioimmunoassay (RIA) and one immunoradiometric assay (IRMA), for the detection of β-core hCG fragment (βC-hCG) in body fluids. In addition, we have compared their performance with two other assays designed for βC-hCG quantification. The RIA uses a rabbit polyclonal antibody raised against pure βC-hCG which has a high affinity constant, is sensitive to 5 pmol/l, and has significant cross-reaction only with the free βLH subunit. The IRMA, designed in a liquid phase, uses the same polyclonal antibody associated with a 125I-labelled mouse monoclonal antibody (32H2) raised against βhCG, is sensitive to 1·5 pmol/l, and does not cross-react significantly with any related glycoprotein. Comparison between these two assays and two others previously published was made by measuring βC-hCG in urine from healthy pregnant women (n = 47) and gave correlation coefficients higher than r= 0·960 with any combination. Analysis of βC-hCG in urine of non-pregnant subjects (n= 238) showed measurable βC-hCG in 8·8% (levels ranged from 5 to 34 pmol/l) with the IRMA and 88·3% with the RIA (n=30; ranging from 28·4 to 228 pmol/l) (P = 0·05). We concluded that, despite different affinities of the antibody involved and different cross-reactivities with related glycoproteins, the four assays we examined may be equally employed to detect βC-hCG in pregnancy urine. However, the IRMA appears to be more appropriate for βC-hCG analysis in non-pregnant individuals, specifically in postmenopausal women because of the high cross-reactivity of the RIA with free βLH or β fragments of other glycoproteins. These studies have significance for our understanding of the physiology of βC-hCG in cancer, pregnancy and after the menopause,

Journal of Endocrinology (1992) 135, 161–174
Ectopic transcription of the parathyroid hormone gene in lymphocytes, lymphoblastoid cells and tumour tissue
Authors: Handt O. , Reis A. , Schmidtke J.
Published: December 2008

Using reverse transcription polymerase chain reaction, steady-state levels of parathyroid hormone (PTH) mRNA were investigated in a number of human, bovine and rat tissues. Transcripts were consistently detected in parathyroid glands as well as in lymphocytes, lymphoblastoid cells and several tumours. Levels of transcription were not measurably increased in lymphoblastoid cells and tumour tissues compared with unstimulated peripheral lymphocytes. The level of 'ectopic' transcription of the PTH gene in lymphoblastoid cells appeared to be resistant to the administration of both vitamin D and phorbol esters.

Journal of Endocrinology (1992) 135, 249–256
Milking frequency alters the milk yield and mammary blood flow response to intra-mammary infusion of insulin-like growth factor-I in the goat
Authors: Prosser C. G. , Davis S. R.
Published: December 2008

The milk yield and mammary blood flow responses to close-arterial, intra-mammary infusion of IGF-I were investigated in five Saanen goats milked frequently or normally the day before. Animals were infused for 6 h with recombinant human IGF-I (1·3 nmol/min) and milked hourly following i.v. injection of oxytocin beginning 2 h before infusion and then every 2 h. On one occasion animals were milked five times (after i.v. injection of oxytocin) on the day before infusion and on the other they were milked twice, without oxytocin. The ratio of milk yield from the infused to that from non-infused gland increased by 17 ±4% (mean ± s.e.m.) in goats milked twice the day before infusion and by 6 ± 2% when the infusion was preceded by frequent milking. Maximal responses were obtained 4 h after the start of the infusion and differed significantly (P<0·05), according to pretreatment milking. Blood flow through the infused gland rose in parallel to the milk yield response. At 5 h, when maximal levels were achieved, blood flow was 182 ±23% of the pre-infusion flow rate following twice-daily milking and 139 ± 3% of the pre-infusion flow rate following more frequent milk removal. Thus, more frequent milk removal on the day before close-arterial infusion of IGF-I attenuated both the milk yield and mammary blood-flow response to the infusion of IGF-I.

Journal of Endocrinology (1992) 135, 311–316
The identification and characterization of 125I-labelled iodomelatonin-binding sites in the duck kidney
Authors: Song Y. , Ayre E. A. , Pang S. F.
Published: December 2008

The melatonin-binding sites in membrane preparations of duck kidney were demonstrated by utilizing 125I-labelled iodomelatonin as a radioligand. Binding at these sites was found to be reversible, saturable, specific and of high affinity. Scatchard analysis of the specific binding revealed an equilibrium binding constant (Kd) of 44·6±4·4 pmol/l (n= 6) and a total number of binding sites (Bmax) of 6·43 ±0·60 fmol/mg protein (n= 6) at the mid-point of the light period (mid-light). The Hill coefficient approached 1·0, suggesting a single class of 125I-labelled iodomelatoninbinding site in the duck kidney. Diurnal variation in 125I-labelled iodomelatonin binding showed that the Bmax was 53·4% higher at mid-light than at mid-point of the dark period (P<0·05), with no significant variation in Kd. The Kd value determined from kinetic analysis was 22·5 pmol/l in birds at mid-light, which was comparable with values determined from equilibrium studies. The order of pharmacological affinity for 125I-labelled iodomelatonin-binding sites in the duck kidney membrane preparations was: 2-iodomelatonin > melatonin > 6-chloromelatonin > 6-hydroxymelatonin > N-acetylserotonin » 5-methoxytryptamine, 5-methoxytryptophol, 5-hydroxytryptamine, tryptamine, 1-acetylindole-3-carboxaldehyde, 5-hydroxyindole-3-acetic acid, l-tryptophan, 5-methoxyindole-3-acetic acid, 3-acetylindole, acetylcholine, epinephrine, norepinephrine and harmaline. The pharmacological characteristics indicated that 125I-labelled iodomelatonin-binding sites are highly specific for melatonin. Our finding of 125I-labelled iodomelatonin-binding sites in the kidney suggests that melatonin may regulate kidney function. The possibility of a direct effect of melatonin on renin secretion and daily urine production is hypothesized.

Journal of Endocrinology (1992) 135, 353–359
Noradrenergic peripheral denervation of the female rat accelerates the positive feedback mechanisms resulting in pubertal ovulation, and blocks the modifications induced by administration of testosterone propionate at birth
Authors: Flores A. , Domínguez R.
Published: December 2008

The noradrenergic innervation of the ovary of prepubertal rats causes an inhibitory response of the follicles to gonadotrophins, leading to ovulation. We investigated the possibility that noradrenergic peripheral denervation at birth, produced by treatment with guanethidine, modifies the positive feedback effects of gonadotrophins and oestradiol in prepubertal rats, and also the possibility that peripheral denervation can modify the anovulatory syndrome induced by androgenization at birth.

Noradrenergic peripheral denervated rats of 18 days of age treated with pregnant mare serum gonadotrophin (PMSG) ovulated 96 h later, while normal animals did not ovulate (4/9 vs 0/12, P < 0·05) and the number of ova shed was lower than in rats which ovulated spontaneously at first vaginal oestrus (3·5 ± 0·6 vs 8·3 ± 0·4 (s.e.m.), P < 0·01). Oestradiol benzoate (10 μg) did not induce ovulation in either normal or denervated animals (0/11 and 0/11). The anovulatory syndrome induced by the administration of testosterone propionate (75 μg) at birth was partially blocked by noradrenergic peripheral denervation (4/7 ovulated vs 0/10).

The results suggest that some neural information arising from the ovary modulates, in an inhibitory way, the stimulatory feedback mechanisms required to induce ovulation. Partial inhibition of the anovulatory syndrome resulting from androgenization caused by peripheral noradrenergic denervation suggests that noradrenergic neural information sent by the ovary to the hypothalamus results in a decreased concentration of noradrenaline in the hypothalamus and in the aromatization of androgens to oestrogens.

Journal of Endocrinology (1992) 135, 415–420
Oestradiol increases the extracellular levels of amine metabolites in the ewe hypothalamus during anoestrus: a microdialysis study
Authors: Gayrard V. , Malpaux B. , Thiéry J. C.
Published: December 2008

Giving a subcutaneous oestradiol implant during anoestrus to ovariectomized ewes inhibits pulsatile LH secretion. This effect results from an increased negative feedback of oestradiol and depends on the synthesis of biogenic amines, mainly from the mediobasal hypothalamus. In the present study, we examined the effect of oestradiol on the extracellular levels of amines and their metabolites. Eight ewes were sampled by microdialysis from the lateral retrochiasmatic area, including the dopaminergic A15 nucleus, during inhibition of LH secretion by long days. Two dialysis sessions were carried out on each ewe; one after a 10-day oestradiol treatment and the other one after 10 days without oestradiol treatment. Half of the ewes were first oestradiol-treated then untreated, the other half received the treatment in the reverse order.

Oestradiol caused a decline in pulsatile LH secretion without affecting the secretion of prolactin. This steroid also led to a significant increase in the levels of amine metabolites: 3,4-dihydroxyphenylacetic acid, homovanillic acid and 5-hydroxyindolacetic acid in the extracellular medium. These results demonstrate the effect of oestradiol on aminergic activity as related to changes in hormonal secretions during long days (16 h of light per 24 h). Thus our data support the hypothesis that amines inhibit gonadotrophic secretion during anoestrus in the ewe and suggest that there is an activation of the aminergic neurones from the retrochiasmatic area in this regulatory mechanism.

Journal of Endocrinology (1992) 135, 421–430
The processing of β-endorphin and α-melanotrophin in the pars intermedia of Xenopus laevis is influenced by background adaptation
Authors: Maruthainar K. , Peng-Loh Y. , Smyth D. G.
Published: December 2008

β-Endorphin-and α-melanotrophin (α-MSH)-related peptides were extracted from the pars intermedia of Xenopus laevis maintained for 2, 4 or 6 weeks on a white background and for the same periods on a black background. The peptides were resolved under dissociating conditions by gel exclusion chromatography on Sephadex G-50 and they were detected by radioimmunoassay with antibodies to β-endorphin, α,N-acetyl β-endorphin and α-MSH. The β-endorphin-related peptides separated into two fractions of different molecular size. Further purification of the peptides in each fraction was by ion exchange chromatography on SP-Sephadex C-25 and by high-pressure liquid chromatography. The α-MSH-related peptides were resolved by gel exclusion and ion exchange chromatography. The purified β-endorphin- and α-MSH-immunoreactive peptides were identified by comparison of their chromatographic properties with the corresponding peptides from porcine pituitary or by comparison with synthetic peptides.

The major form of β-endorphin in the pars intermedia of the frog adapted to a white background was identified as α,N-acetyl β-endorphin (1–8); it was accompanied by a small quantity of acetylated peptides with molecular size similar to β-endorphin. In contrast, the pars intermedia of the frogs adapted to a black background contained approximately equal amounts of α,N-acetyl β-endorphin (1–8) and the larger forms of β-endorphin. The higher molecular weight forms were identified as the α,N-acetyl derivatives of β-endorphin (1–26), (1–27) and (1–31); however after 6 weeks of white adaptation the sole remaining peptide in this group was the 26-residue peptide. An additional β-endorphin immunoreactive peptide, provisionally identified as β-endorphin (10–26), was present in both black- and white-adapted animals; the amounts of this peptide increased during white adaptation. Major differences in the processing of α-MSH were also observed. In the frogs adapted to a black background des-acetyl α-MSH greatly predominated over the acetyl form whereas after 6- weeks adaptation to a white background the acetylated peptide proved to be the principal component.

The results demonstrate that the proteolytic processing of β-endorphin and the acetylation of α-MSH in Xenopus laevis are influenced by background adaptation. The formation of β-endorphin (1–8) appears to reflect the action of an endopeptidase that acts at the single arginine residue present at position 9. This cleavage does not appear to take place in mammalian β-endorphins where position 9 is occupied by lysine.

Journal of Endocrinology (1992) 135, 469–478
Paradoxical hypertrophy and plasticity of the testis in rats recovering from early thyroid deficiency: a growth study including effects of age and duration of hypothyroidism
Authors: Meisami E. , Sendera T. J. , Clay L. B.
Published: December 2008

The effects of various durations of postnatal hypothyroidism followed by recovery were studied on testicular growth in Long–Evans and Sprague–Dawley rats from birth to 7 months. Hypothyroidism was induced by adding propylthiouracil (PTU) in drinking water (0·1%, w/v). Recovery was induced by withdrawal of PTU. Testicular growth was reduced in rat pups by 20, 65 and 90% at days 10, 25 and 50. Upon withdrawal of PTU at weaning (25 days), testicular growth resumed and became compensatory; catch-up growth occurred by day 65. Paradoxically, testicular growth progressively increased, surpassing the control weights by 40, 50 and 100% at days 75, 90 and 210. Maximal testicular growth rate in the recovery group was 35% higher, occurred 2 weeks later and lasted 2 weeks longer than controls. Testes of rats subjected to prolonged postnatal hypothyroidism (60 or 120 days) also showed recovery and hypertrophy, amounting to nearly twice the normal maximal growth levels, after at least 6 months of recovery. Body weights of recovering rats remained always significantly below those of controls. When the suckling pups were exposed to short, week-long regimes of PTU treatment, only rats treated during the early postnatal weeks (days 1–8 or 9–16) had enlarged testes; PTU treatment during the late suckling period (days 17–24) or later had no effects. Total duration of hypothyroidism in the suckling period was positively correlated with testicular enlargement. The results indicate that hypothyroidism early in life is stimulatory to testicular growth, resulting in a paradoxical twofold increase in final testicular size which occurs even if hypothyroidism is prolonged. These effects occur similarly in different strains of rat with differing sized testes. It is suggested that there is a sensitive period for this effect, i.e. during the first 2 weeks after birth. The marked plasticity of testicular growth as shown by its recovery and hypertrophy, even after long periods of hypothyroid retardation, is unique in the body and may be a useful model for studying hormonal factors regulating testicular growth and for animal breeding and research into infertility.

Journal of Endocrinology (1992) 135, 495–505
Transforming growth factor-β is a potent inhibitor of basal and stimulated relaxin release by porcine luteal cells maintained in monolayer culture
Authors: Taylor M. J. , Clark C. L.
Published: December 2008

The effect of transforming growth factor-β (TGF-β) on relaxin release by porcine large luteal cells (LLC) was examined by use of a reverse haemolytic plaque assay. In this assay, mixed luteal cells were co-cultured in monolayers with protein A-coupled sheep erythrocytes. In the presence of complement and porcine relaxin antiserum, a zone of haemolysis (a plaque) developed around relaxin-releasing LLCs. The rate of plaque development in time-course experiments and the average size of plaque areas were used to monitor the rate of relaxin release and cumulative amounts of hormone respectively. Monolayers were bathed in medium containing TGF-β alone, or in the co-presence of a stimulatory secretagogue (prostaglandin E2; PGE2).

Exposure of luteal cell-containing monolayers to TGF-β (1 ng/1–100 μg/l) elicited a dose-related inhibition in the rate of basal relaxin release. Minimal and maximal concentrations were approximately 10 ng/l and 10 μg/l respectively. Treatment with 1 μg TGF-β/l reduced the cumulative amount of relaxin released to 63 ± 6% of control values (mean ± s.d., P < 0·05, n = 6; averaged over the whole course of the experimental incubation). Exposure of monolayers treated with TGF-β to the relaxin-stimulatory secretagogue PGE2 (0·1 μmol) resulted in a significant (P < 0·05) increase in the amount of relaxin released by TGF-β-suppressed LLCs, and restored rates of hormone release to control levels. This is evidence that TGF-β and PGE2 interact antagonistically in the modulation of relaxin. The effect of TGF-β was strictly time-dependent. A period of at least 16 h of treatment with TGF-β was required to induce an inhibitory effect on relaxin.

These results implicate TGF-β as a novel inhibitor of relaxin release, and the presence of TGF-β in porcine luteal tissue suggests that such regulation may be achieved via paracrine or autocrine routes. It is possible that TGF-β interacts functionally during luteal life with other secretagogues to achieve integrated control of hormone release.

Journal of Endocrinology (1992) 135, 543–550
Human chorionic gonadotrophin and subunit composition of maternal serum and coelomic and amniotic fluids in the first trimester of pregnancy
Authors: Iles R. K. , Wathen N. C. , Campbell D. J. , Chard T.
Published: December 2008

Sixteen matched samples of first trimester amniotic fluid (AF), extraembryonic coelomic fluid (EECF) and maternal serum (MS) were assayed for intact human chorionic gonadotrophin (hCG) and free subunits. Total β-hCG (free β-subunit and intact hCG) levels in the EECF (median 410 kIU/l) were 61 times greater than levels in AF (median 6·73 kIU/l) and 2·8 times greater than in MS (median 141·5 kIU/l). Levels of intact hCG in the EECF (median 245 kIU/l) were 142 times greater than in AF (median 1·73 kIU/l) and 1·6 times greater than in MS (median 157 kIU/l). Free α-subunit levels in EECF (median 17·3 mg/l) were 66 times greater than in AF (median 0·262 mg/l) and 12 times greater than in MS (median 1·3 mg/l). Virtually all of the total β-hCG immunoreactivity in MS can be attributed to intact hCG, but only 60% of total β-hCG in the EECF and 20% of that in AF can be accounted for by the intact hormone. In both EECF and AF the free α-subunit was a major constituent; on a molar basis the ratio of free α:free β:intact hCG was 1:1·2:0·3 in AF, 1:0·6:0·5 in EECF and 1:0:5 in MS. Chromatography of MS, EECF and AF on Sephadex G-100 confirmed the hCG and subunit composition of the fluids. On the basis of these findings it seems likely that previous studies showing very high levels of hCG in AF during the first trimester may have incorrectly sampled the EECF. In reality, the levels of total hCG (and free subunits) are low in the AF, and only 20% is intact hCG. In both AF and EECF the free subunits may have been derived by dissociation of intact hormone, or possibly by independent synthesis. These and other findings suggest that either the amnion acts as a barrier to the transfer of proteins or that there may be dynamic removal from this compartment. By contrast, the EECF might act as a relatively stable reservoir for these proteins.

Journal of Endocrinology (1992) 135, 563–569
Immunohistochemical localization of 17α-hydroxylase/C17–20 lyase and aromatase cytochrome P-450 in the human ovary during the menstrual cycle
Authors: Tamura T. , Kitawaki J. , Yamamoto T. , Osawa Y. , Kominami S. , Takemori S. , Okada H.
Published: December 2008

Immunohistochemical localization of 17α-hydroxylase/C17–20 lyase (P-45017α,lyase) and aromatase cytochrome P-450 (P-450arom) in normal human ovaries during the menstrual cycle was studied using specific polyclonal antibodies which were raised against corresponding enzymes. In the follicular phase of matured follicles, P-45017α,lyase was localized in theca interna cells and P-450arom in granulosa cells. P-45017α,lyase was expressed in theca interna cells before P-450arom was expressed in granulosa cells. The corpus luteum showed immunoreactivity to both enzymes and, after menstruation, immunoreactivity decreased gradually until it could not be detected in the corpus albicans. In corpus luteum graviditatis the immunoreactivity continued to be expressed strongly. In some atretic follicles, P-45017α,lyase and/or P-450arom continued to be expressed. In the stromal layer, P-45017α,lyase was detected in secondary interstitial cells, which originated from the theca interna of atretic follicles, and P-450arom was detected in hilar cells. Immunoreactivity to both enzymes was also detected in oocytes of developing follicles. These results are consistent with the two cell theory in the human ovary. They also suggest that androgens and oestrogens are produced not only by follicles and corpora lutea but also by stroma and oocytes.

Journal of Endocrinology (1992) 135, 589–595
Russell's viper venom stimulates insulin secretion from rat islets of Langerhans
Authors: Jones P. M. , Mann F. M.
Published: December 2008

Burmese Russell's viper venom (RVV) caused a dose-and temperature-dependent stimulation of insulin secretion from islets of Langerhans isolated from rat pancreas by collagenase digestion. RVV stimulated both basal and glucose-induced insulin secretion at concentrations which did not compromise islet cell viability as assessed by exclusion of trypan blue dye. The effects of RVV on insulin secretion could not be attributed to the activation of protein kinase C (PKC), since down-regulation of PKC by prolonged exposure to a tumour-promoting phorbol ester did not abolish subsequent secretory responses to RVV. However, RVV-induced insulin secretion was inhibited in the absence of extracellular Ca2 +, and RVV did not stimulate insulin secretion from Ca2+-clamped electrically permeabilized islets at either substimulatory (50 nmol/l) or stimulatory (10 μmol/l) concentrations of Ca2 +, suggesting that changes in cytosolic Ca2+ are important in the stimulation of insulin secretion by RVV. The phospholipase A2 (PLA2) inhibitor quinacrine caused a dose-dependent inhibition of RVV-induced insulin secretion, suggesting that the activation of PLA2, perhaps in response to Ca2+ influx, may be partially responsible for RVV-induced insulin secretion.

Journal of Endocrinology (1993) 136, 27–33
Distribution, molecular characterization of pituitary adenylate cyclase-activating polypeptide and its precursor encoding messenger RNA in human and rat tissues
Authors: Ghatei M. A. , Takahashi K. , Suzuki Y. , Gardiner J. , Jones P. M. , Bloom S. R.
Published: December 2008

The distribution of a novel neuropeptide, pituitary adenylate cyclase-activating polypeptide (PACAP), was studied in the brain of the rat and man and a variety of other rat tissues using Northern blot hybridization and two radioimmunoassays for PACAP 1–38 and PACAP 1–27. The assay, using PACAP 1–38 as standard and an antibody to PACAP 21–38 and radiolabelled tracer, revealed immunoreactive PACAP in all brain regions examined, with the highest concentrations in the rat being in the hypothalamus, nucleus accumbens and substantia nigra (380 ± 34, 310 ± 37 and 346 ± 30 pmol/g wet tissue, means±s.e.m., n = 5 respectively), whilst in man the highest concentrations were found in the pituitary gland (15·8 ± 4·7 pmol/g). Immunoreactive PACAP 1–38 was also detected in the rat gastrointestinal tract, adrenal gland and testis. The assay using PACAP 1–27 as standard and label and an antibody to PACAP 1–27 detected immunoreactive PACAP only in the rat hypothalamus (12·6 ± 1·8 pmol/g wet tissue, n = 5). PACAP mRNA of approximately 2·7 kb in size was detectable in all brain regions of both rat and man, and its distribution paralleled that of the immunoreactive peptide.

Gel permeation chromatography of different regions of human and rat hypothalamus, and also rat spinal cord and small intestine, showed a broad immunoreactive peak corresponding to PACAP 1–38. Fast protein liquid chromatography (FPLC) resolved this peak into two immunoreactive peaks, the majority eluting in the position of synthetic PACAP 1–38. Presence of immunoreactivity corresponding to PACAP 1–27 was also confirmed in rat hypothalamic extracts using FPLC. The presence of immunoreactive PACAP and its precursor encoding mRNA in various neural and other tissues is in accord with a role for PACAP as a neurotransmitter, neuromodulator or neurohormone.

Journal of Endocrinology (1993) 136, 159–166
Immunoreactive and receptor-active insulin-like growth factor-I (IGF-I) and IGF-binding protein in blood plasma from the freshwater fish Macquaria ambigua (golden perch)
Authors: Anderson T. A. , Bennett L. R. , Conlon M. A. , Owens P. C.
Published: December 2008

The presence of insulin-like growth factor-I (IGF-I)-related molecules and IGF-binding factors in blood from golden perch, Macquaria ambigua, an Australian native freshwater fish, was investigated.

Serum was acidified to dissociate IGF and IGF-binding protein complexes that might be present, and fractionated by size-exclusion high-performance liquid chromatography at pH 2·8. Fractions were neutralized and their activities assessed by (i) an immunoassay for mammalian IGF-I which also detects chicken IGF-I but in which all known forms of IGF-II react very poorly, (ii) a receptor assay for IGF-II in which all known forms of IGF-I react poorly, and (iii) a type-I IGF receptor assay in which mammalian IGF-I and IGF-II polypeptides are almost equivalent. No IGF-II-like activity was detected. Three peaks of IGF-I-like activity were detected by IGF-I immunoassay and type-I IGF receptor assay. The major peak of activity was similar in molecular size to human IGF-binding protein-3, 45–55 kDa ('large IGF'), and a minor peak of activity which was similar in size to mammalian IGFs, 7·5 kDa. A third peak of activity was observed eluting at a time which indicates that it is a smaller molecule than any previously described IGF. The large IGF was temperature-sensitive, but was not a binding protein for 125I-labelled human IGF-I (hIGF-I). This material therefore was able to bind to anti-hIGF-I antibodies and to human type-I IGF receptors, and may represent the fish equivalent of mammalian prepro-IGFs.

The two smallest forms of IGF activity identified by IGF-I radioimmunoassay and type-I radioreceptor assay following acidic size-exclusion chromatography were able to stimulate protein synthesis by L-6 myoblasts in culture, although large IGF did not.

When fresh (but not frozen and thawed) golden perch serum was incubated with 125I-labelled hIGF-I and then fractionated by size-exclusion liquid chromatography at pH 7·4 through Sephadex G-100, the radioactivity became associated with a complex, intermediate in size between free IGF-I and the major IGF-binding protein in human serum. The association of 125I-labelled hIGF-I with the complex was inhibited by the presence of unlabelled hIGF-I in the incubation.

These studies show that receptor-active, immunoreactive and bioactive IGF-I-like activity is present in golden perch serum, and demonstrate the presence of an IGF-I-binding factor in this species.

Journal of Endocrinology (1993) 136, 191–198