bluuebunny/crossref_metadata_2025 huggingface dataset:
Vasoactive intestinal peptide inhibits the steroid-induced LH surge in the ovariectomized rat
Authors: Weick R. F. , Stobie K. M.
Published: December 2008

The LH surge was induced in ovariectomized rats by sequential treatment with oestradiol benzoate and progesterone. Vasoactive intestinal peptide (VIP) or saline was infused into the third cerebral ventricle from 13.30 to 16.30 h on the afternoon of the anticipated LH surge. Two blood samples were taken by jugular puncture from each animal, one at 12.00 h as a control sample and the other at 16.00, 18.00, 20.00 or 22.00 h. Saline-infused animals showed a normal LH surge, with mean plasma LH concentrations reaching a peak at 18.00 h, declining by 20.00 h and reaching control (12.00 h) levels by 22.00 h. Plasma LH in animals infused with VIP was not significantly higher than control levels at 16.00 or 18.00 h. By 20.00 h, mean LH levels in VIP-infused rats had risen to the levels seen at that time in saline-infused rats, and by 22.00 h LH had returned to control levels in VIP-infused animals. We interpret these findings to mean that VIP inhibits LH secretion during the LH surge. It does not block the surge completely, as pentobarbital during the critical period would have done; nor does VIP appear to affect the timing of the LH surge. Rather, VIP inhibits the increased LH secretion rates of the LH surge only during the period of VIP treatment and for a short time afterward.

Journal of Endocrinology (1992) 133, 433–437
Immunoreactive β-core-like material in normal postmenopausal urine: human chorionic gonadotrophin or LH origin? Evidence for the existence of LH core
Authors: Iles R. K. , Lee C. L. , Howes I. , Davies S. , Edwards R. , Chard T.
Published: December 2008

Material with the immunochemical properties of the β-core of human chorionic gonadotrophin (hCG) can be found in the urine of normal postmenopausal women. However, we have been unable to detect intact hCG (using an assay which is specific for the α–β heterodimer of intact hCG) in serum of such subjects. The levels of serum LH and urinary β-core were compared in matched samples from 28 women (serum LH: median 27 U/l, range 4-70 U/l, urinary β-core: median 0·27 μg/l, range < 0·05–0·645 μg/l). Urine (4 litres) from three postmenopausal women was concentrated, dialysed and subjected to gel exclusion chromatography on Sephadex G-100. Fractions were analysed by specific assays for LH, intact hCG, total β-hCG (free β-subunit and intact hCG), free α-subunit and β-core. Material eluting at the expected position of the β-core fragment of hCG was detected in all three samples by the β-core, β-hCG and LH assays, despite the fact that the LH antibody does not recognize the authentic β-core of pregnancy. Electrophoresis and Western blotting of the concentrated urines revealed that material of the same molecular size as β-core was recognized by the antibody to LH but not by a monoclonal antibody raised to free β-hCG which also recognizes the β-core molecule of hCG. We conclude that the predominant core-like material identified in postmenopausal urine is probably derived from the β-subunit of LH.

Journal of Endocrinology (1992) 133, 459–466
A comparison of the clearance of ovine and human corticotrophin-releasing hormone (CRH) in man and sheep: a possible role for CRH-binding protein
Authors: Saphier P. W. , Faria M. , Grossman A. , Coy D. H. , Besser G. M. , Hodson B. , Parkes M. , Linton E. A. , Lowry P. J.
Published: December 2008

A specific binding protein for human corticotrophin-releasing hormone (hCRH), which does not bind to the ovine hormone (oCRH), has recently been demonstrated in human plasma. No such binding protein has been found in sheep plasma. We have investigated the half-life of human and ovine CRH in man and in sheep. Peptides were measured directly in plasma with two-site immunoradiometric assays, as these assays are unaffected by the presence of inactivated peptide fragments. In man, the half-life of hCRH (30·5 ± 3·3 min; mean ± s.e.m.) was significantly (P < 0·001) less than that of oCRH (42·8 ± 6·4 min). In sheep, there was no significant difference between the half-life of hCRH (46·5 ± 7·2 min) and that of oCRH (39·8 ± 10·1 min); these half-lives were also significantly (P < 0·001) longer than that of hCRH in man. One possible explanation for the shorter half-life of hCRH in man is that the clearance of hCRH is enhanced by CRH-binding protein, although other binding proteins often have the opposite effect.

Peak ACTH and cortisol responses occurred earlier in sheep than in man, although no differences were found in the response times to oCRH or hCRH within either species. The responses were more sustained in sheep than in man, and the previously reported biphasic response was only seen in some of the sheep and not in man. Absolute responses to either peptide were greater in sheep than in man; however, in man an 8·1-fold rise in ACTH was measured in response to oCRH, while hCRH gave a significantly (P = 0·043) smaller 4·4-fold response. These experiments are discussed with particular reference to the effect of hCRH-binding protein and its possible role in potentiating the clearance of the peptide.

Journal of Endocrinology (1992) 133, 487–495
Antiprogesterone RU486 induces dissociation of LH and FSH secretion in the cyclic rat: effect of anti-inhibin serum
Authors: Sánchez-Criado J. E. , Bellido C. , López F. J. , Galiot F.
Published: December 2008

Administration of the antiprogesterone RU486 to 4-day cyclic rats from metoestrus to pro-oestrus increases serum levels of LH while decreasing levels of FSH. If it is assumed that there is only one gonado-trophin-releasing hormone, there is no direct explanation for the decrease in FSH concentrations. The purpose of these experiments was to investigate the effect of RU486 on gonadotrophin secretion in cyclic rats during periods when the secretion of LH and FSH diverges. RU486 blunted the transient increase in FSH concentration on the afternoon of metoestrus and the compensatory ovarian hypertrophy on the next day of oestrus in unilaterally ovariectomized 4–day cyclic rats. In addition, bilateral ovariectomy reversed the effect of RU486 on the basal secretion of FSH. RU486 induced an increase in basal LH concentrations. Since ovarian inhibin decreases the basal release of FSH, and decreases in peripheral inhibin seem to be responsible for the transient rise in FSH during the oestrous cycle, the effect of RU486 on serum levels of LH and FSH during dioestrus in rats injected with a sheep anti-inhibin serum (AIS) were further evaluated. Treatment with AIS increased FSH levels in oil-treated rats without altering the levels of LH. In contrast, the effects of AIS on FSH secretion were blunted in RU486-treated rats. The results suggest that inhibin might be involved in the RU486-induced decrease of FSH secretion in cyclic rats.

Journal of Endocrinology (1992) 134, 43–49
Effects of steroid hormones on cyclic adenosine 3′,5′-monophosphate levels in the rat lung
Authors: Nabishah B. M. , Khalid B. A. K. , Morat P. B. , Alias A. K. , Zainuddin M.
Published: December 2008

The possible role of cyclic adenosine 3′,5′-monophosphate (cAMP) in mediating the action of steroid hormones was investigated using the rat lung. Male rats were adrenalectomized and treated with olive oil, dexamethasone, corticosterone, deoxycorticosterone (DOC) or progesterone. At the end of 10 days, 100 μg isoprenaline/kg was injected intraperitoneally 5 min before the animals were killed to stimulate cAMP production. Adrenalectomy significantly decreased cAMP levels in the rat lung. Dexamethasone and corticosterone pretreatment reversed the effect of adrenalectomy whereas progesterone pretreatment but not DOC pretreatment significantly decreased lung cAMP levels. Cyclic AMP levels in normal female rats, whether pregnant or not, were not significantly different from those in male rats. We concluded that the absence of glucocorticoid, as after adrenalectomy, decreased the cAMP levels in rat lungs and that this could be reversed by either dexamethasone or corticosterone replacement. Progesterone reduced the cAMP content in rat lungs by acting as a glucocorticoid antagonist or by acting directly via progesterone receptors.

Journal of Endocrinology (1992) 134, 73–76
The ontogenic and functional relationships between growth hormone- and prolactin-releasing cells during the development of the bovine pituitary
Authors: Kineman R. D. , Faught W. J. , Frawley L. S.
Published: December 2008

The ontogeny of GH- and prolactin-releasing cells in the developing bovine pituitary was evaluated by reverse haemolytic plaque assays which allows for the detection of hormone release from individual pituitary cells in culture. With this approach, we observed that GH-releasing cells ontogenically preceded prolactin-releasing cells. In fact, GH secretors were observed as early as 59 days of gestational age while cells that released prolactin were not identified until 98 days. The amounts of both GH- and prolactinreleasing cells increased with time to reach more than 50% and 20% of all pituitary cells near term (term ∼280 days) respectively. Interestingly, the first cells shown to release prolactin also released GH (i.e. were mammosomatotropes). This temporal and functional relationship between GH and prolactin secretors provides suggestive evidence that GH-secreting cells act as the progenitor cells for prolactin secretors via a functional intermediate, the mammosomatotrope.

Journal of Endocrinology (1992) 134, 91–96
A comparison of plasma vasopressin and oxytocin concentrations during the oestrous cycle of the ewe
Authors: Wathes D. C. , Ayad V. J. , Kumari M. , Matthews E. L. , Wathes C. M.
Published: December 2008

Blood samples were collected twice daily (09.30 and 17.00 h) via indwelling jugular-vein cannulae from five ewes throughout an entire oestrous cycle. Additional frequent samples were taken at 30-min intervals for 8 h on each of days 3 (early luteal phase), 9 (mid-luteal phase), 12 (late luteal phase) and 0 (day of oestrus). Plasma concentrations of arginine vasopressin and oxytocin were measured in all samples by radioimmunoassay and progesterone was measured in the twice-daily samples only.

Both oxytocin and progesterone showed the expected pattern of plasma concentrations, increasing during the early luteal phase, reaching a plateau and declining either preceding (oxytocin) or at (progesterone) luteolysis. Vasopressin concentrations showed a significant dependence on the day of cycle (P<0·05, analysis of variance) with concentrations lowest at oestrus and minor peaks on days 4 and 8–9. There was no correlation between the concentrations of vasopressin and progesterone. Vasopressin values were significantly higher in the morning than in the afternoon samples (1·3 cf. 0·9 pmol/l; P<0·001). Analysis of the frequent samples showed a significant (P<0·001) dependence on the time of day for vasopressin but not oxytocin. Values were high throughout the morning, declined to a trough at 15.00 h and rose again by 17.00 h.

We conclude that there is a minor variation in the vasopressin concentration during the oestrous cycle, which is not related to the circulating progesterone concentration but could be regulated by oestradiol. We also provide evidence for a diurnal rhythm in the release of vasopressin into the plasma in the ewe. The concentrations of vasopressin measured are unlikely to be sufficient to influence the activity of the reproductive tract in vivo.

Journal of Endocrinology (1992) 134, 107–113
Metabolism of dihydrotestosterone to 3α-androstanediol in brain and plasma: effect on behavioural activity in female rats
Authors: Erskine M. S. , Hippensteil M. , Kornberg E.
Published: December 2008

Six experiments were carried out to determine whether dihydrotestosterone (5α-androstan-17β-ol-3-one; DHT) acts to inhibit oestradiol (OE2)-induced lordosis behaviour after its metabolic conversion to 5α-androstane-3α,17β-diol (3α-androstanediol, 3α-Adiol). In experiments 1 and 2, ovariectomized rats were treated with several doses of DHT or 3α-Adiol, injected with OE2 and progesterone, and tested for lordosis responsiveness. Significant inhibition of lordosis occurred after a dose of 3α-Adiol which was approximately threefold less than the effective DHT dose. In experiments 3 and 4, plasma concentrations of DHT and 3α-Adiol were measured after the injection of these steroids to ovariectomized rats at doses shown to be both sufficient or insufficient to inhibit lordosis. Behaviourally effective dosages of DHT and 3α-Adiol produced circulating concentrations of 3α-Adiol greater than those produced by behaviourally ineffective doses of DHT or 3α-Adiol. At 30 min after injection of DHT (experiment 3), 78·8% of plasma androgens were in the form of 3α-Adiol, while after injection of 3α-Adiol, only 7·4% were DHT. When plasma DHT and 3α-Adiol were measured at 3, 6, 9 and 12 h after steroid injection (experiment 4), plasma levels of 3α-Adiol produced by the behaviourally subthreshold dose of DHT were significantly lower than levels produced by behaviourally sufficient dosages of DHT or 3α-Adiol. In experiments 5 and 6, concentrations of DHT and 3α-Adiol were measured in five brain regions 1 and 6 h after injection of behaviourally sufficient doses of these steroids to ovariectomized females. At 1 h after injection, similar levels of DHT and 3α-Adiol were measured in DHT- and 3α-Adiol-injected females, and DHT concentrations in the preoptic area were significantly higher in both groups than in any other brain area. At 6 h, animals injected with DHT had significantly higher levels of DHT in all brain areas combined than did 3α-Adiol-or vehicle-injected animals. Brain concentrations of 3α-Adiol were not different between groups injected with DHT, 3α-Adiol or vehicle at this time. In brain, 34·6% of DHT had been converted to 3α-Adiol after 1 h and 53·0% of 3α-Adiol had been converted to DHT. These results suggest that the inhibitory action of DHT on lordosis may be a consequence of its conversion to 3α-Adiol, and that this conversion may account for the higher behavioural potency of the latter steroid.

Journal of Endocrinology (1992) 134, 183–195
Changes in the plasma concentration of gastric inhibitory polypeptide and other metabolites in response to feeding in sheep
Authors: McCarthy J. P. , Faulkner A. , Martin P. A. , Flint D. J.
Published: December 2008

Plasma concentrations of gastric inhibitory polypeptide (GIP)-like activity were determined in sheep before and after refeeding following a 48-h fast. Plasma concentrations increased significantly after feeding, from about 250 pg/ml to about 550 pg/ml. Other metabolites in plasma also increased at this time, reflecting the absorption of nutrients from the gastrointestinal tract. Significant increases were observed in the plasma concentrations of acetate, β-hydroxybutyrate and triacylglycerol. By comparing the time-courses of the changes in concentration of GIP and other metabolites in plasma, possible sites of secretion and secretagogues of GIP in ruminant animals are proposed. The results demonstrate that GIP is secreted in response to nutrient absorption in adult ruminants and that, as in simple-stomached animals, the absorption of long-chain free fatty acids plays an important role in this secretion.

Journal of Endocrinology (1992) 134, 235–240
In-vitro DNA synthesis in Leydig and other interstitial cells of the rat testis
Authors: Moore A. , Findlay K. , Morris I. D.
Published: December 2008

Replicative DNA synthesis (125I-labelled iododeoxy-uridine incorporation) was measured in interstitial cells prepared from rat testes and separated by Percoll density gradient centrifugation. Leydig cells were identified by 125I-labelled human chorionic gonadotrophin (hCG) binding and 3β-hydroxysteroid dehydrogenase histochemistry. Continuous density gradients indicated that interstitial cell DNA synthesis was not associated with Leydig cells, and was greater in cells from the immature than from the mature rat testis. Fractionation of cells by discontinuous density gradients into Leydig cell-rich and -depleted pools did not result in a similar enrichment of DNA synthesis. Treatment of the adult rat with hCG increased DNA synthesis into both fractions but oestrogen had no effect. DNA synthesis was greater in cells from the immature rat but, in contrast to the adult, in-vivo hCG treatment had no effect, whilst oestrogen decreased synthesis. To characterize the cells synthesizing DNA further, interstitial cells were prepared from testes in which the Leydig cells were depleted by in-vivo treatment with ethane dimethanesulphonate (EDS) or depleted in their germ cells by treatment in utero with busulphan. EDS treatment had no effect on DNA synthesis by the interstitial cells in spite of the 125I-labelled hCG binding being markedly reduced. Similarly, busulphan treatment was also without effects upon DNA synthesis. Fluorescence-activated cell cycle analysis of cells from both fractions from germ cell-depleted testes indicated that only a small proportion (3%) of the interstitial cells were actively dividing and this was almost doubled in cells from the germ cell-depleted immature rat testes. The experiments showed that the majority of cells in the interstitium of the rat testes do not synthesize DNA and are not undergoing cell division. The small proportion of cells that are dividing are probably not Leydig cells. The experiments have identified a dividing interstitial cell population and, in consideration of the changes in the immature and mature rat as well as the effects of hormone treatment, they may be regarded as putative Leydig cell precursors.

Journal of Endocrinology (1992) 134, 247–255
Apparent α-inhibin subunit immunoactivity in porcine and ovine luteal extracts is due to interference by cytosolic proteases in the assay
Authors: Bramley T. A. , Menzies G. S. , Baxter G. , Webb R. , McNeilly A. S.
Published: December 2008

Immunoreactive α-inhibin (ir-inhibin) was measured in luteal homogenates and subcellular fractions of ovine and porcine corpora lutea (CL) and in pig granulosa cells (GCs), using a sensitive radioimmunoassay specific for the 1–26 amino acid sequence of the N-terminus of the α chain of porcine inhibin (p1–26 α-inhibin). Inclusion of N-ethylmaleimide (N-EM) and/or EDTA in the immunoassay had no effect on the measurement of p1–26 α-inhibin peptide standards, on ir-inhibin levels in ovine follicular fluid and serum, or on ir-inhibin in subcellular fractions of pig GC. Fractionation of porcine GC homogenates on sucrose gradients demonstrated a major particular peak of ir-inhibin (buoyant density, 1·15–1·21 g/cm3) with variable activity in the cytosol. The particulate ir-inhibin peak was released into the cytosol by pretreatment of GC homogenates with the saponin, digitonin, prior to fractionation. Porcine GC extracts contained a protein (Mr 45 000) which immunoblotted against p1–26 α-inhibin antibody.

In the absence of inhibitors of proteolysis, apparent ir-inhibin activity was very high in extracts of sheep and pig CL. However, inclusion of N-EM or EDTA in the radioimmunoassay significantly reduced ir-inhibin levels in porcine and ovine CL extracts in a dose-dependent manner. Measurements of peptide tracer integrity indicated that porcine luteal cytosol degraded 125I-labelled p1–26 α-inhibin peptide. Subcellular fractionation studies demonstrated high levels of apparent ir-inhibin in luteal cytosol fractions, with only minor activity peaks associated with particulate fractions; however, this material was not releasable by digitonin. Immunoblotting of detergent extracts of porcine luteal particulate fractions failed to demonstrate α-inhibin material, and immunocytochemical localization studies of α-inhibin in porcine and ovine luteal sections were negative.

Our results are consistent with the intracellular packaging/storage of a form of α-inhibin (Mr similar to that of α-inhibin subunit precursor) in the porcine granulosa cell. However, luteinization of the porcine follicle was associated with a dramatic fall in ir-inhibin content, and the loss of immunostaining for α-inhibin peptides. We conclude that porcine and ovine CL contain little, if any, authentic inhibin. These studies emphasize the importance of excluding proteolytic artefacts when measuring biological peptides in luteal tissue extracts by radioimmunoassay.

Journal of Endocrinology (1992) 134, 341–352
The effect of fetal hypophysectomy with or without ACTH replacement on the molecular weight profile of enkephalin-containing peptides in the adrenal medulla of the fetal sheep
Authors: Coulter C. L. , Young I. R. , Browne C. A. , McMillen I. C.
Published: December 2008

We have investigated the possible role of the fetal pituitary and ACTH in the control of the synthesis and post-translational processing of the enkephalin precursor, proenkephalin A (proEnk A), in the fetal sheep adrenal gland in late gestation.

Fetal hypophysectomy (n = 8) or sham operations (n = 4) were performed between 109 and 118 days of gestation. At 138–139 days, either ACTH(1–24) (10·5 μg/0·24 ml saline per h, n = 4) was infused intravenously for 72 h into hypophysectomized fetal sheep or 0·9% (w/v) NaCl alone (0·24 ml/h, n = 4) was infused for 72 h into hypophysectomized fetal sheep and sham-operated animals. At the end of the infusion the pregnant ewe was killed and left or right adrenal glands (n = 12) were collected from the fetal sheep that were intact and given saline (Intact + sal; n = 4), hypophysectomized and given saline (Hx + sal; n = 4) and hypophysectomized and given ACTH (Hx + ACTH; n = 4). Each adrenal was homogenized in acid (acetic acid (1 mol/l)/HCl (20 mmol/l)/2-mercaptoethanol (0·2%)). After centrifugation, the supernatant was loaded onto a Sephadex G-75 column (2·0 × 50 cm), eluted at 80 ml/24 h and fractions were collected (5 ml, n = 42). An aliquot of each fraction (2 ml) was dried down prior to enzymatic digestion (trypsin/carboxypeptidase B) and oxidation with H2O2, and assay for methionine-O-enkephalin (immunoreactive Met-O-Enk).

The total adrenal content of immunoreactive Met-O-Enk was significantly greater in the Hx + ACTH group (326·2 ±66·7 (s.e.m.)ng/adrenal) when compared with either the Intact + sal group (152·7 ±44·0 ng/adrenal) or the Hx + sal group (112·1 ±20·8 ng/adrenal). In the adrenal glands from all fetuses immunoreactive Met-O-Enk was found in four molecular weight ranges: < 12 kDa, 12–7 kDa, 7–3 kDa and < 3 kDa. There was no significant difference between the Hx + sal and Hx + ACTH groups in the proportion of immunoreactive Met-O-Enk present in each of the molecular weight ranges in the adrenals and therefore the data from these groups were combined for further statistical analysis. The proportion of immunoreactive Met-O-Enk in the > 12 kDa range was significantly less in the Intact + sal group (5·5 ±2·3%) when compared with the hypophysectomized sheep with or without ACTH replacement (18·7 ± 4·5%).

These data demonstrate that fetal hypophysectomy alters the molecular weight profile of Enk-containing peptides in the adrenal of the fetal sheep and whilst ACTH replacement in the hypophysectomized fetus does not alter the post-translational processing of the Enk-containing peptides, it stimulates an increase in the total amount of immunoreactive Met-O-Enk in the fetal adrenal in late gestation.

Journal of Endocrinology (1992) 134, 369–375
ELEVATING MATERNAL INSULIN-LIKE GROWTH FACTOR-I IN MICE AND RATS ALTERS THE PATTERN OF FETAL GROWTH BY REMOVING MATERNAL CONSTRAINT
Authors: Gluckman P.D. , Morel P.C.H. , Ambler G.R. , Breier B.H. , Blair H.T. , McCutcheon S.N.
Published: December 2008

Fetal growth is normally constrained by maternal factors. This constraint is demonstrated by the usual inverse linear relationship between litter size and mean fetal weight. Cross-breeding experiments between mice of lines selected for high or low plasma insulin-like growth factor (IGF-I) levels suggested that elevations in maternal IGF-I abolish (P <0.01) this constraining effect and reverse the usual positive relationship between fetal and placental size in late gestation. This was confirmed by treating mice and rats throughout pregnancy with IGF-I. In normal mice and in low IGF-I line mice treatment with IGF-I 10μg 8-hourly s.c. from day 1 to 19 of pregnancy) abolished maternal constraint whereas 0.9% (w/v) NaCl treatment did not. In Wistar rats osmotic pumps were implanted to deliver IGF-I (1μg/g body weight per day), bovine GH (bGH; 0.6μg/g body weight per day) or saline from day 1 to 19 of pregnancy. IGF-I therapy but not bGH or saline abolished (P < 0.01) maternal constraint and altered (P <0.01) the relationship between placental and fetal weight.

When high or low IGF-I line mice embryos were transplanted into a normal line of mice, the expected negative relationship (P <0.05) between mean fetal weight and litter size was maintained. However the embryos of the high line were heavier (P <0.05) than those from the low line irrespective of fetal number, suggesting a direct role for IGF-I in the regulation of fetal growth.

Thus both endogenous and exogenous elevations in maternal IGF-I indirectly promote fetal growth either by altering nutrient delivery to the placenta or by affecting placental function.
Interaction between myometrial relaxants and oxytocin: a comparison between relaxin, cromakalim and salbutamol
Authors: Downing S. J. , Hollingsworth M.
Published: December 2008

The influence of treatment with oestradiol on the effects of the uterine relaxants, relaxin, salbutamol (an agonist at β2-adrenoceptors) and cromakalim (a potassium channel opener) and their interactions with the uterine stimulant oxytocin were investigated in vivo in the ovariectomized rat. Oestradiol benzoate (0·4 μg/kg per day) significantly increased sensitivity to cromakalim as an inhibitor of spontaneous uterine contractions compared with vehicle-treated rats by approximately threefold. The same dose of oestradiol benzoate had no effect on uterine sensitivity to salbutamol. Previous studies have shown that this dose of oestradiol benzoate produces a twofold increase in uterine sensitivity to relaxin as an inhibitor of spontaneous contractions. Oestradiol influenced the ability of relaxin to inhibit oxytocin-stimulated uterine contractions. In corn oil-treated rats, uterine responses to relaxin were markedly reduced during oxytocin infusion compared with responses to relaxin before oxytocin; the maximum obtainable response to relaxin was less than 50% inhibition. In oestradiol-treated rats, uterine sensitivity to relaxin during oxytocin infusion was similar to that observed against spontaneous contractions. Cromakalim was able to inhibit uterine contractions during oxytocin infusion in both corn oil- and oestradiol-treated rats, uterine sensitivity to cromakalim being similar in the absence and presence of oxytocin for both hormone treatment groups. Salbutamol was also able to inhibit uterine contractions during oxytocin infusion in both corn oil- and oestradiol-treated rats. Oestradiol treatment increased the potency of salbutamol as an inhibitor of oxytocin-stimulated uterine contractions compared with corn oil treatment by 3·5-fold. The interaction of oestradiol and relaxin during late pregnancy may be important for attenuation of the myometrial response to stimulants.

Journal of Endocrinology (1992) 135, 29–36
Activation of the hypothalamo-pituitary-adrenocortical axis in the conscious rabbit by the pyrogen polyinosinic: polycytidylic acid is dependent on corticotrophin-releasing factor-41
Authors: Milton N. G. N. , Hillhouse E. W. , Milton A. S.
Published: December 2008

The pyrogenic interferon inducer polyinosinic: polycytidylic acid (Poly I: C) was shown to activate the rabbit hypothalamo-pituitary-adrenocortical (HPA) axis in vivo. The immunoreactive cortisol response to Poly I:C (2·5 μg/kg) was shown to have a corticotrophin-releasing factor-41 (CRF-41)-dependent component which was abolished by peripheral immunoneutralization using an anti-CRF41 monoclonal antibody (KCHMB001; 2·5 mg/kg i.v.). Peripheral administration of the arginine vasopressin (AVP) V1 receptor antagonist ([deaminoPen1, O-Me-Tyr2, Arg8]-vasopressin; 225 nmol/kg i.v.) had no effect on the response of immunoreactive cortisol to Poly I:C, suggesting that AVP was not involved in activation of the HPA axis. Poly I: C increased both body temperature and circulating immunoreactive prostaglandin E2; these responses were abolished by the cyclo-oxygenase inhibitor ketoprofen (3 mg/kg s.c.). The immunoreactive cortisol response to Poly I: C, however, remained after the administration of ketoprofen, indicating a prostaglandin (PG)-independent component. The immunoreactive cortisol levels in control, saline vehicle-treated, animals were reduced by both the CRF-41 receptor antagonist (α-helical CRF (9–41); 6·25 mmol/kg i.v.) and ketoprofen (3 mg/kg s.c.) indicating that this basal state is dependent on both CRF-41 and PGs.

Journal of Endocrinology (1992) 135, 69–75
Effects of basic fibroblast growth factor, transforming growth factor-β and nerve growth factor on the secretory function of the bovine corpus luteum in vitro
Authors: Miyamoto A. , Okuda K. , Schweigert F. J. , Schams D.
Published: December 2008

The effects were investigated of basic fibroblast growth factor (bFGF), transforming growth factor-β (TGF-β) and nerve growth factor (NGF) on the release of progesterone and oxytocin from the bovine corpus luteum (CL) at different stages of the oestrous cycle. A microdialysis system (MDS) of CL and a cell culture system with a reduced number of endothelial cells were used.

In the MDS of CL from the mid-luteal stage (days 8–12 of the oestrous cycle), infusion with bFGF (0·1, 1, 10 and 100 ng/ml), TGF-β (0·1, 1 and 10 ng/ml) and NGF (0·1, 1, 10 and 100 ng/ml) for 30 min induced significant acute effects on the release of progesterone. Both bFGF and NGF stimulated the release of progesterone during peptide infusion, TGF-β and also bFGF in the period thereafter. This stimulation was dose-dependent during and after the infusion only for bFGF. This response pattern was observed at all luteal stages for the three growth factors, but bFGF was more stimulatory at the early (days 5–7) and mid-luteal stages during and after peptide infusion. The release of oxytocin was stimulated by bFGF in a dose-dependent manner. At the highest dose, bFGF, TGF-β and NGF stimulated the release of oxytocin throughout all three luteal stages. When luteal cells were cultured with growth factors, only TGF-β showed a dose-dependent inhibition of both basal and LH-stimulated progesterone as well as oxytocin release (measured between 48 and 52 h of culture). NGF had an inhibitory effect only on the basal release of oxytocin. bFGF had no effect on the release of either hormone under continuous stimulation in cell culture.

The results indicate that bFGF, TGF-β and NGF act directly and acutely on the secretory function of bovine CL in the MDS but also have long-term effects as shown in cell culture. bFGF appears to be an important autocrine/paracrine regulator of CL function, since local expression of its mRNA, peptide synthesis and its mitogenic and non-mitogenic actions have now been confirmed. Endothelial cells from the CL have been identified as target cells for bFGF. Differences observed between the two systems might thus be attributed to the presence or absence of cellto-cell contact and a reduced number of endothelial cells, as well as to the duration of peptide stimulation and medium changes every 24 h compared with the flow-through conditions in the MDS.

Journal of Endocrinology (1992) 135, 103–114
Effects of tri-iodothyronine and insulin-like growth factor-I (IGF-I) on alkaline phosphatase activity, [3H]thymidine incorporation and IGF-I receptor mRNA in cultured rat epiphyseal chondrocytes
Authors: Ohlsson C. , Nilsson A. , Isaksson O. , Bentham J. , Lindahl A.
Published: December 2008

The effects of tri-iodothyronine (T3) and insulin-like growth factor-I (IGF-I) on [3H]thymidine incorporation, alkaline phosphatase (ALP) activity and IGF-I receptor mRNA levels were studied in rat epiphyseal chondrocytes cultured in monolayer.

Chondrocytes from enzymatically digested rat tibia epiphyseal growth plates were seeded in monolayer culture and precultured for 7–14 days in Ham's F-12 medium supplemented with 10% (v/v) newborn calf serum and 1% (v/v) of a serum substitute. After preculture the medium was changed to Ham's F-12 medium containing 1% (v/v) serum from hypophysectomized rats, and the effects of T3 and/or IGF-I on DNA synthesis ([3H]thymidine incorporation), ALP activity (a late marker of differentiated epiphyseal chondrocytes) and IGF-I receptor mRNA levels were studied.

ALP activity was increased by T3 in a dose-dependent manner with a maximal response at 10 μg T3/1 (678 ±86% compared with control culture). The increase in ALP activity was accompanied by a concomitant decrease in [3H]thymidine incorporation (52 ±14% compared with control culture). Human GH (hGH; 50 μg/l) and IGF-I (25 μg/l) had no stimulatory effect on ALP activity. However IGF-I (10 μg/l) exerted an inhibition on the T3 (10 μg/l)-induced increase in ALP activity (64 ± 9% compared with T3-treated culture). T3 (3 μg/l) inhibited the increase in [3H]thymidine incorporation caused by 25 μg IGF-I/l(51 ± 13% compared with IGF-I-treated culture). Furthermore, IGF-I receptor mRNA levels were increased by 10 μg T3/l (137 ±4·2% compared with control culture) while no effect of hGH (50 μg/l) or IGF-I (25 μg/l) was demonstrated.

Both T3 and IGF-I were shown to interact with epiphyseal chondrocytes and both substances seemed to affect cell proliferation and maturation and therefore longitudinal bone growth. Furthermore, the results indicated that IGF-I is important for proliferation of the cells while T3 initiates the terminal differentiation of epiphyseal chondrocytes.

Journal of Endocrinology (1992) 135, 115–123
A role for di-acetyl α-melanocyte-stimulating hormone in the control of cortisol release in the teleost Oreochromis mossambicus
Authors: Lamers A. E. , Flik G. , Atsma W. , Wendelaar Bonga S. E.
Published: December 2008

In stressed tilapia, Oreochromis mossambicus, total α-melanocyte-stimulating hormone (α-MSH) levels and di-acetyl α-MSH/mono-acetyl α-MSH (di:mono) ratios are elevated. We therefore investigated the role of α-MSH in the regulation of the pituitary–interrenal axis. The corticotrophic activities of des-acetyl α-MSH, mono-acetyl α-MSH and di-acetyl α-MSH were compared. These forms of α-MSH were isolated from neurointermediate lobes and tested in a superfusion experiment with homologous interrenal tissue. The corticotrophic activity of di-acetyl α-MSH was the highest, followed by that of des-acetyl α-MSH and mono-acetyl α-MSH. Apparently, acetylation of α-MSH is of functional significance for corticotrophic action. Di-acetyl α-MSH proved to be about 100 times less potent than ACTH(1–39): the half-maximal stimulating concentrations for ACTH and di-acetyl α-MSH were 0·89 nmol/l and 110 nmol/l respectively. Surprisingly, a superfusate from neurointermediate lobes proved to be only about three times less active than a superfusate from the pituitary pars distalis, in which the corticotrophic activity is attributable to its ACTH content. When selectively stripped of all forms of α-MSH by passage through a Sepharose column coated with an antiserum against α-MSH the neurointermediate lobe superfusate was devoid of corticotrophic activity. Thus α-MSH appears to be the corticotrophic factor in the superfusate of the neurointermediate lobe. After the same treatment, the corticotrophic activity of the pars distalis superfusate was not affected. We conclude that (in vivo) an as yet unidentified factor is co-released with α-MSH from the neurointermediate lobe, and that this potentiates its corticotrophic activity.

Journal of Endocrinology (1992) 135, 285–292
Zona radiata proteins are synthesized by rainbow trout (Oncorhynchus mykiss) hepatocytes in response to oestradiol-17β
Authors: Oppen-Berntsen D. O. , Gram-Jensen E. , Walther B. T.
Published: December 2008

The present study delineates the origin of the three major proteins constituting the rainbow trout (Oncorhynchus mykiss) zona radiata. Intraperitoneal administration of oestradiol-17β induced the appearance in the blood from juvenile fish (both sexes) of proteins immunoreactive to rabbit antisera against zona radiata proteins (zr proteins). These proteins had similar molecular weights to the zr proteins (a, 60 kDa; β, 55 kDa; and γ, 50 kDa). Primary hepatocyte cultures from fish treated with oestradiol-17β incorporated radioactive [35S] methionine into four major proteins with molecular weights of 160, 60, 55 and 50 kDa. Only the latter three proteins were specifically immunoprecipitated with antibodies to zr proteins. Furthermore, our data demonstrate that in such cultures the biosynthetic mole ratios of these secreted proteins (60, 55 and 50 kDa) are close to one. Control cultures from fish that had not been treated with oestradiol-17β failed to produce immunoreactive proteins. The data support the hypothesis that zr proteins are synthesized in a concerted manner in the liver during teleostean oogenesis and transported by the blood for deposition in ovarian follicles.

Journal of Endocrinology (1992) 135, 293–302
Different molecular forms of C-type natriuretic peptide isolated from the brain and heart of an elasmobranch, Triakis scyllia
Authors: Suzuki R. , Takahashi A. , Takei Y.
Published: December 2008

The presence of a C-type natriuretic peptide (CNP) has been reported in the brain of mammals, birds, amphibians and teleost fishes, mostly as a 22-residue peptide (CNP-22). In the present study, we attempted to isolate natriuretic peptides from an elasmobranch, Triakis scyllia, using a chick rectum-relaxant assay, and different molecular forms of CNP were found in the brain and heart. Only CNP-22 was recovered from the brain, as is the case in other vertebrates. A large amount of prohormone (proCNP or CNP-115) and small amounts of its C-terminal peptides (CNP-38 and CNP-39) were isolated from the atrium and ventricle, however. No CNP-22 was recovered from the heart. Natriuretic peptides other than CNP were not isolated from Triakis heart and brain by the rectumrelaxant assay. The 22 residues at the C-terminal region of proCNP, i.e. CNP-22, were well conserved when Triakis and mammals were compared, although the sequence homology of the N-terminal segment of proCNP was very low. Not only was CNP-22 identical but the N-terminal segments of proCNP were also quite similar when Triakis and another elasmobranch, Scyliorhinus canicula, were compared. These data suggest that, in elasmobranchs, CNP is a primary hormone in the natriuretic peptide family, and also that CNP is processed differently in the brain and heart.

Journal of Endocrinology (1992) 135, 317–323
Intracellular pH and growth hormone-releasing factor-stimulated adenosine 3′5′-monophosphate, intracellular calcium and growth hormone release from rat anterior pituitary cells
Authors: Puttagunta A. L. , Chik C. L. , Girard M. , O'Brien L. , Ho A. K.
Published: December 2008

In this study, we examined the effect of changes in intracellular pH (pHi) on basal and GH-releasing factor (GRF)-stimulated cyclic AMP (cAMP), intracellular Ca2+ and GH release using a static monolayer culture prepared from dispersed rat anterior pituitary cells. To modulate pHi, two approaches were used: variation of extracellular pH (pHo) and addition of sodium propionate and ammonium chloride which alter pHi directly. Direct pHi measurement with 2′7′-bis(carboxyethyl)-5(6)-carboxyfluorescein showed that for pHo values between 6·9 and 7·6, a change in pHo of 0·1 units resulted in a change in pHi of 0·045 units. Sodium propionate (30 mmol/l) reduced pHi by 0·06 units whereas ammonium chloride (30 mmol/l) increased pHi by 01 units. Increasing pHo from 6·6 to 7·8 enhanced the maximal GRF-stimulated cAMP and GH responses by 80% and 300% respectively, indicating that the GRF-stimulated cAMP and GH release were both pH-dependent. Acute elevation of pHo from 6·6 to 7·8 also increased basal GH release by sixfold. Reduction of pHi by sodium propionate, however, had no significant effect on GRF-stimulated cAMP levels while the corresponding GRF-stimulated GH release was reduced by up to 40%. In comparison, elevation of pHi by ammonium chloride enhanced the GRF-stimulated cAMP release by up to 75% and the corresponding increase in GH was less than 20%. When the relationship between pHi and intracellular Ca2+ was determined with the fluorescent Ca2+ indicator, Fura-2, it was found that increasing pHo and treatment with ammonium chloride increased intracellular Ca2+, while sodium propionate and reducing pHi had no effect on intracellular Ca2+. These results indicate that activation of adenylate cyclase and mobilization of intracellular Ca2+, two intracellular signalling pathways of importance to GH secretion, are both sensitive to changes in pHi.

Journal of Endocrinology (1992) 135, 343–352
Testosterone-induced suppression of self-healing Plasmodium chabaudi malaria: an effect not mediated by androgen receptors?
Authors: Benten W. P. M. , Wunderlich F. , Mossmann H.
Published: December 2008

This study investigates whether androgen receptors (AR) mediate the suppressive effect of testosterone on self-healing Plasmodium chabaudi malaria. Our data show the following. (1) Female and castrated male mice of the inbred strain C57BL/10 self-heal and survive infections when challenged with 106 P. chabaudi-parasitized erythrocytes. However, self-healing is prevented when circulating testosterone levels are high as in intact males or in females and castrated males pretreated with 0·9 mg testosterone twice a week for 3 weeks. (2) The lethal outcome of P. chabaudi in intact males is not affected by different doses of AR blockers such as cyproterone acetate, cyproterone, flutamide and nilutamide when applied at least 3 weeks before infection and during infection. Also, these AR blockers do not impair the testosterone-induced lethal outcome of infections in testosterone-treated females and castrated males. (3) Tfm mice possessing mutant non-functional ARs and normal 'male' testosterone levels succumb to infection with P. chabaudi. However, the corresponding wild-type mice possessing functioning ARs are able to resist P. chabaudi infections at low circulating testosterone levels. (4) In contrast to testosterone, testosterone metabolites such as 5α-dihydrotestosterone, 5β-dihydrotestosterone, androsterone and 1-dehydrotestosterone cannot suppress self-healing in castrated male B10 mice. Our data suggest that testosterone suppresses the development of protective immunity against P. chabaudi malaria, and that this immunosuppressive effect of testosterone is not primarily mediated by the classical AR response.

Journal of Endocrinology (1992) 135, 407–413
Expression of the growth hormone receptor gene in chicken pituitary glands
Authors: Hull K. L. , Fraser R. A. , Harvey S.
Published: December 2008

Although GH has no direct effect on GH release from chicken pituitary glands, GH receptor mRNA similar to that in the rabbit liver was identified by Northern blot analysis in extracts of adult chicken pituitaries. Complementary (c) DNA, reverse transcribed from chicken pituitary RNA, was amplified by the polymerase chain reaction (PCR) in the presence of 3′- and 5′-oligonucleotide primers coding for the extracellular domain of the chicken liver GH receptor and was found to contain an electrophoretically separable fragment of 500 bp, identical in size to that in chicken liver. Digestion of this pituitary cDNA with NcoI produced expected moities of 350 and 150 bp. Amplification of chicken pituitary cDNA in the presence of oligonucleotide primers for the intracellular sequence of the chicken liver GH receptor produced an electrophoretically separable fragment of approximately 800 bp, similar to that in chicken liver. This fragment was cut into expected moieties of 530 and 275 bp after digestion with EcoRI. These PCR fragments were identified in extracts of the pituitary caudal lobe, in which somatotrophs are confined and account for the majority of endocrine cell types, and in the cephalic lobe, in which somatotrophs are lacking. Translation of the GH receptor mRNA in the pituitary gland was indicated by the qualitative demonstration of radio-labelled GH-binding sites in plasma membrane preparations, in pituitary cytosol and in nuclear membranes. These results provide evidence for the expression and translation of the GH receptor gene in pituitary tissue, in which GH receptors appear to be widely distributed within cells and in different cell types. GH may therefore have paracrine, autocrine or intracrine effects on pituitary function.

Journal of Endocrinology (1992) 135, 459–468
Site-dependent effects of experimental hypo- and hyperthyroidism on resident macrophages in extraocular muscles of rats: a quantitative immunohistochemical study
Authors: Schmidt E. D. , van Hogerwou G. , van der Gaag R. , Wiersinga W. M. , Asmussen G. , Koornneef L.
Published: December 2008

It has been suggested that the effects of dysthyroidism on resident immunocompetent cells of the extraocular muscles may play a role in the pathogenesis of Graves' ophthalmopathy. The distribution of such cells was therefore studied in extraocular muscles of rats that were made hyper- or hypothyroid by the oral administration of thyroxine or propylthiouracil respectively. Skeletal muscles were studied for comparison. The cell distributions were analysed in cryostat cross-sections subjected to a two-step immunoperoxidase method using well-characterized monoclonal antibodies against T cells, B cells, macrophages and MHC class II antigens.

The extraocular muscles of control (euthyroid) rats contained numerous macrophages, fewer MHC-II positive cells and T cells and no B cells. Differences in the distribution of immunocompetent cells were found in control rats, between skeletal and extraocular muscles as well as within the various recti eye muscles. This particular tissue distribution resembles that previously reported for human extraocular and skeletal muscles.

Quantitative analysis showed that experimental dysthyroidism only affected cell populations in the extraocular muscles. Significant effects on the number of macrophages were observed in the inferior rectus muscle of both hypo- and hyperthyroid rats, this was most pronounced in the orbital layer of the muscles. Both hyper- and hypothyroidism appear to affect local cell distributions in a tissue-specific manner. The presently observed site-dependent effects of dysthyroidism on local immunocompetent cell populations may have relevance for the differential involvement of muscular tissues in Graves' ophthalmopathy.

Journal of Endocrinology (1992) 135, 485–493