bluuebunny/crossref_metadata_2025 huggingface dataset:
Effects of treatment with the opioid antagonists naloxone and naltrexone on LH secretion and on sexual maturation in immature female rats
Authors: Meijs-Roelofs H. M. A. , Kramer P.
Published: December 2008

The effects on sexual maturation of the opioid receptor antagonists naloxone and naltrexone were studied in the female rat. Neonatal treatment (days 1–10) with either naloxone (2·5 mg/kg at 6-h intervals) or naltrexone (20 or 50 mg/kg per day) did not advance sexual maturation as judged by age and body weight at vaginal opening and first ovulation. After treatment with naltrexone (20 mg/kg) first ovulation occurred 2·3 days earlier than in saline-treated control rats but this could be attributed to a growth-stimulating effect of naltrexone; the effect was not observed with 50 mg/kg. An effect of neonatal treatment with naloxone on serum LH levels was seen at 23 days of age (155±36 (s.e.m.) vs 14±4 μg LH/1 in controls, P<0·01), but not at 29 or 33 days of age, at 2 days before first ovulation nor at first pro-oestrus. There were no differences in the number of ova released at first oestrus, nor in the length of the first cycle. Neonatal treatment with naltrexone (50 mg/kg per day) did not alter the response to treatment with human chorionic gonadotrophin at 28–31 days of age: ovulation of a mean of 7·3 ova was induced on day 32 in both naltrexone- and saline-treated rats. Naltrexone treatment (four daily injections of 20 mg/kg at 2-h intervals) at 28–32 days of age advanced first ovulation by 4·4 days in about 40% of the rats.

Short-term effects of naloxone and naltrexone treatment on LH secretion were studied in 15-day-old rats and a clear increase in serum LH concentration was seen 15 min after injection; LH levels were similar with the various doses of antagonist used (2·5 and 20 mg naloxone/kg; 2·5, 20 and 50 mg naltrexone/kg). At 30 min after injection LH levels were still maximal if the 2·5 mg/kg dose of either antagonist was used, whereas with the higher doses a significant (P<0·01) decrease in the level of LH was observed, indicating a longer-lasting effect of the lower (2·5 mg/kg) dose of antagonist on LH secretion. No differences in effectiveness were seen between the supposedly short-lasting opioid receptor antagonist naloxone and the supposedly long-lasting antagonist naltrexone.

It was concluded that neonatal treatment with the opioid antagonists naloxone and naltrexone did not specifically influence sexual maturation. The antagonists clearly increased LH secretion in short-term experiments. Treatment with naltrexone at 28–32 days of age advanced first ovulation in 40% of the rats, a result that is in agreement with a possible opiatergic influence on the gradually increasing LH release during the final phase of sexual maturation.

The effects of the opioid antagonist naltrexone on the LH release mechanism did not parallel its reported long-lasting effects on nociceptive response, no long-lasting increase of LH release was seen, and the timecourse of the LH response was the same using the antagonists naloxone and naltrexone.

J. Endocr. (1988) 117, 237–243
Involvement of the somatostatin and cholinergic systems in the mechanism of growth hormone autofeedback regulation in the rat
Authors: Torsello A. , Panzeri G. , Cermenati P. , Caroleo M. C. , Ghigo E. , Camanni F. , Müller E. E. , Locatelli V.
Published: December 2008

The involvement of the cholinergic system in GH secretion has recently acquired increasing importance. Data have been presented suggesting that in rats the effect of cholinergic modulation on GH secretion takes place through inhibition or stimulation of hypothalamic somatostatin (SRIF) release. To investigate further the significance of cholinergic-SRIF link and its role in the regulation of GH secretion, the action of cholinergic agonist and antagonist drugs in the GH short-loop feedback mechanism mediated by SRIF was investigated. Intracerebroventricular (i.c.v.) infusion of 0·2 or 2·0 μg GH/rat into the lateral brain ventricle of adult male rats induced a significant reduction in the GH-releasing hormone (GHRH; 2 μg/kg, i.v.)-induced peak GH rise, but only the 2·0 μg dose reduced also the GH-integrated area after administration of GHRH. This effect was absent after central administration of 20·0μg GH/rat, due probably to leakage of some GH from the cerebral ventricle into the systemic circulation. Pretreatment with cysteamine (300 mg/kg, s.c.), a known depletor of hypothalamic SRIF, or with anti-SRIF serum (0·5 ml/rat) completely counteracted the lessening of the GH response to GHRH induced by 2·0μg GH injected i.c.v. Similarly, pretreatment with the cholinergic agonist pilocarpine (3 mg/kg, i.v.) completely antagonized the inhibitory effect of central infusion of GH on the GHRH-induced GH response. Atropine (1·0 mg/kg, i.v.), a muscarinic cholinergic antagonist, strikingly inhibited the GHRH-induced GH rise, but when given in combination with i.c.v. infusion of GH there was no additive inhibitory effect. These data reinforce the idea that the GH autofeedback triggered by i.c.v. infusion of GH is mediated by enhanced SRIF release, and suggest that the hypothalamic cholinergic system plays a major role in this mechanism.

J. Endocr. (1988) 117, 273–281
Subcellular fractionation of the porcine corpus luteum: sequestration of progesterone in a unique particulate fraction
Authors: Bramley T. A. , Menzies G. S.
Published: December 2008

Homogenates of porcine corpus luteum were subjected to fractionation by differential-rate centrifugation or sucrose density gradient fractionation, with or without pretreatment with digitonin. Fractions of each gradient were assayed for a number of markers characteristic of the major intracellular organelles and cell-surface membranes, and for progesterone content.

The majority of the progesterone content of homogenates of porcine corpus luteum was associated with a low-density particulate fraction which equilibrated at a buoyant density of 1·07–1·09 g/cm3. Pretreatment with digitonin increased the buoyant density of the progesterone-enriched fraction markedly (to 1·13–1·15 g/cm3) without causing release of steroid. The density distributions of progesterone content in control and digitonin-treated luteal gradient fractions were quite distinct from those of the major intracellular organelles and luteal cell-surface membranes. However, NADH–cytochrome C reductase activity (but not other endoplasmic reticulum markers) was also enriched in this fraction. The results suggest that most of the progesterone of the porcine corpus luteum is associated with a unique particulate fraction which is enriched in digitonin-reactive lipids and NADH–cytochrome C reductase activity.

J. Endocr. (1988) 117, 341–354
Characteristics of the ACTH response to repeated pulses of corticotrophin-releasing factor and arginine vasopressin in vitro
Authors: Evans M. J. , Brett J. T. , McIntosh R. P. , McIntosh J. E. A. , McLay J. L. , Livesey J. H. , Donald R. A.
Published: December 2008

A multi-column perifusion system was used to investigate the dynamics of the dose–response relationships of ACTH release by ovine pituitary cells when stimulated by both corticotrophin-releasing hormone (CRF) and arginine vasopressin (AVP) given alone and in combination. A dose–response relationship was obtained when 10-min pulses were given at 60-min intervals over the range of 0·002–2000 nmol CRF/l and 1–2000 nmol AVP/l, with a minimum effective concentration of 0·02 nmol CRF/l or 1 nmol AVP/l. When AVP was given together with CRF, the expected potentiation of the ACTH response occurred when compared with the summed response of these secretagogues given separately. At the higher concentrations of CRF and AVP used, the ACTH responses to repeated pulses decreased with time during the experiment. The rate of this loss of responsiveness was significantly correlated to the size of the response to the first pulse (for CRF: r = 0·89, P<0·01; for AVP: r = 0·95, P < 0·01), being greatest when the response was potentiated by adding the secretagogues together (for CRF plus AVP: r = 0·95, P <0·01). Reduced availability of receptors or changes in intracellular transduction processes may contribute to this desensitization. Reduced levels of secretable ACTH do not appear to be implicated because desensitization to pulses of one secretagogue did not cause equivalent desensitization to the other. In addition, cells stimulated continuously with submaximal levels of either secretagogue showed desensitization while more ACTH was still available for release to higher levels of stimulant. The potentiation of CRF plus AVP resulted primarily from a rapid increase in the height of the response since the width of the response (at one-third maximum height) was significantly (P < 0·001) less after AVP plus CRF than after CRF alone. Also the rapidity with which ACTH concentrations fell after removal of the stimulus from the perifusion medium was significantly (P <0·01) faster following AVP, and CRF plus AVP (P <0·05), than after CRF alone.

It is concluded that reduced ACTH responsiveness following repeated stimulation is dependent upon the type and concentration of the secretagogue. AVP had a shorter duration of action than CRF in vitro and potentiated the initial response to CRF rather than prolonging its action. Desensitization to each stimulant appeared to act by a mechanism independent of the other and therefore appeared to occur at or near the receptor level and be unrelated to the availability of ACTH.

J. Endocr. (1988) 117, 387–395
Concentrations of oxytocin-neurophysin prohormone mRNA in corpora lutea of sheep during the oestrous cycle and in early pregnancy
Authors: Jones D. S. C. , Flint A. P. F.
Published: December 2008

Concentrations of oxytocin in corpora lutea from pregnant and non-pregnant sheep rose during luteinization to peak between days 5 and 9 after oestrus before falling to low levels on days 12 and 14. Concentrations of the 608-base mRNA encoding the oxytocin-neurophysin prohormone, measured by Northern blotting using a cRNA probe, were highest before day 5, and fell to low levels by day 9. The decline in oxytocin during the second half of the oestrous cycle and in pregnancy could therefore be accounted for by reduced luteal concentrations of oxytocin-neurophysin mRNA. The rate of decline in prohormone mRNA concentration between days 3 and 16 followed first order kinetics, suggesting that gene expression ceased early in the cycle. The lag between peak oxytocin-neurophysin mRNA and peak oxytocin concentrations may represent the time taken for post-translational processing of the prohormone.

J. Endocr. (1988) 117, 409–414
Effect of serotonin antagonists on prolactin and progesterone secretion in rats: evidence that the stimulatory and inhibitory actions of serotonin on prolactin release may be mediated through different receptors
Authors: Jahn G. A. , Deis R. P.
Published: December 2008

The serotoninergic regulation of prolactin release was studied in female rats in different reproductive states using ketanserin, a specific S2 receptor blocker, ICS 205-930 ((3α-tropanyl)1H-indol-3-carboxylic acid ester), a specific S3 receptor blocker and p-chlorophenylalanine (pCPA), a serotonin synthesis inhibitor. Administration of ketanserin to pro-oestrous rats inhibited the afternoon prolactin surge; this inhibition was prevented by progesterone. On day 3 of pregnancy, pCPA or ketanserin blocked the afternoon prolactin surge, and administration of oestrogen (on day 2) and progesterone (on day 3) in combination, but not alone, prevented this effect. On day 9 of pregnancy, treatment with oestrogen (on day 8) and progesterone (on day 9) induced an afternoon surge of prolactin which was prevented by administration of ketanserin or pCPA. On days 9 and 16, pCPA induced a slight increase in serum prolactin in rats not treated with steroids, but ketanserin had no effect. On day 13, ketanserin and pCPA had no effect on serum prolactin levels, but after increasing serotoninergic transmission by injecting fluoxetine and 5-hydroxytryptophan, serum prolactin levels were decreased. On day 19, ketanserin produced a transient increase in the serum concentration of prolactin, probably produced by the marked decrease in the serum concentration of progesterone induced by the S2 receptor blocker. Administration of ICS 205-930 to pro-oestrous rats or rats on day 19 of pregnancy had no effect on serum concentrations of prolactin and progesterone. These results suggest that the stimulatory action of serotonin on prolactin release is mediated through its binding to type-2 receptors, while the inhibitory action seen on the second half of pregnancy may be mediated through type-1 receptors.

J. Endocr. (1988) 117, 415–422
Effects of bovine follicular fluid on gonadotrophin secretion in intact and chronically ovariectomized ewes before and after desensitization of pituitary gonadotrophs to gonadotrophin-releasing hormone
Authors: Knight P. G. , Castillo R. J.
Published: December 2008

Intact and chronically ovariectomized ewes were treated for 4 days with charcoal-treated bovine follicular fluid (FF) or charcoal-treated bovine serum during the late-anoestrous period, and the effects on basal and gonadotrophin-releasing hormone (GnRH)-induced secretion of LH and FSH observed. Subsequently, ewes received s.c. implants containing a sustained-release formulation of a potent GnRH agonist d-Ser(But)6-Azgly10-LHRH (ICI 118630) to desensitize pituitary gonadotrophs to hypothalamic stimulation, and the effects of bovine FF and bovine serum were re-assessed 2 weeks later. Chronic exposure (for 2–3 weeks) to ICI 118630 significantly reduced basal levels of LH and FSH in both intact and ovariectomized ewes and completely abolished both spontaneous LH pulses as well as exogenous GnRH-induced acute increases in plasma LH and FSH levels. Treatment with bovine FF significantly reduced plasma FSH levels, but not LH levels, in both intact and ovariectomized ewes before and after chronic exposure to ICI 118630. In intact ewes before exposure to ICI 118630, treatment with bovine FF actually enhanced pulsatile LH secretion and raised mean plasma LH levels by 240% (P <0·05). No such stimulatory effect of bovine FF on LH secretion was observed in intact ewes exposed to ICI 118630 or in ovariectomized ewes before or after exposure to ICI 118630, suggesting that the effect probably involved an alteration in ovarian steroid feedback affecting hypothalamic GnRH output. Treatment with bovine FF did not significantly affect the magnitude of GnRH-induced surges of LH or of FSH observed in either intact or ovariectomized ewes before exposure to ICI 118630. These observations indicate that charcoal-treated bovine FF, a rich source of inhibin, can directly suppress pituitary FSH secretion in vivo, irrespective of whether a functionally intact hypothalamo-pituitary-ovarian axis is present.

J. Endocr. (1988) 117, 431–439
Two vasopressin-like peptides in the pig testis?
Authors: Nicholson H. D. , Smith A. J. , Birkett S. D. , Denning-Kendall P. A. , Pickering B. T.
Published: December 2008

Vasopressin (VP)-like immunoreactivity (IR) has been located in the testes of several species of mammal. There is evidence that most of this IR in the rat does not represent authentic arginine vasopressin (AVP) and that a second AVP-like peptide may exist. We have studied testis samples from the pig, which produces lysine vasopressin (LVP) in its pituitary, and have found both LVP- and AVP-like IR.

High-performance liquid chromatography (HPLC) of testis extracts showed two peaks of VP-IR. The first peak co-eluted with authentic LVP and was recognized only by antisera which cross-reacted with LVP. The second peak co-eluted with authentic AVP and was recognized by antisera raised against AVP. Both VP-like peptides bound to a neurophysin affinity column and the HPLC elution profiles of the bound peptides were similar to those of the authentic hormones. When the LVP-like material was oxidized with performic acid, a peak of IR running in the same position as oxidized authentic LVP on HPLC was produced. Similarly, the performic acid-oxidized AVP-like material co-eluted with oxidized authentic AVP.

The presence of both LVP- and AVP-like peptides in the pig testis may mean that more than one gene is involved. A second VP-like gene could also explain the anomalies of VP-IR in other species.

J. Endocr. (1988) 117, 441–446
Receptor-binding characteristics and contractile responsiveness of the myometrium following prolonged infusion of bradykinin and oxytocin in rats
Authors: Engstrom T. , Atke A. , Vilhardt H.
Published: December 2008

Binding of [3H]oxytocin to purified myometrial plasma membranes was unaffected by continuous infusion of bradykinin over 5 days in rats pretreated with oestradiol 2 days before collection of tissue. In contrast, oxytocin treatment resulted in a 76% decrease in maximal binding of [3H]oxytocin and thereby in oxytocin receptor concentration without affecting the dissociation constant. The KM value (molar concentration giving half maximal contraction) of isolated uterine strips stimulated with oxytocin was increased and maximal contractile responses were reduced following oxytocin infusions.

The binding of [3H]bradykinin to purified plasma membranes was influenced by treatment with both oxytocin and bradykinin. Bradykinin infusions down-regulated the bradykinin receptor concentration by 19%, while the receptor affinity remained unchanged. Maximal contraction (Emax) values of isolated strips stimulated with bradykinin exhibited a slightly attenuated response and KM values were significantly enhanced. Long-term treatment with oxytocin down-regulated myometrial bradykinin receptors by 31%. In addition, oxytocin infusions caused Emax to decrease and KM to increase in experiments with isolated uterine strips stimulated with bradykinin.

It is concluded that the down-regulation of oxytocin and bradykinin receptors following prolonged exposure to oxytocin may result from changes in a common pathway for intracellular peptide receptor processing. Likewise, the increased KM values of isolated myometrial strips (despite unchanged dissociation constants) suggest that prolonged oxytocin treatment affects the coupling between receptor activation and contractile response.

J. Endocr. (1988) 118, 81–85
Reduced vasoactive intestinal polypeptide immunoreactivity in the pituitaries of hormone-deficient mutant mice
Authors: Noguchi T. , Sugisaki T. , Kanamatsu T. , Satoh I. , Nishikawa N.
Published: December 2008

The prolactin-producing cells of the bovine anterior pituitary were found to contain a vasoactive intestinal polypeptide (VIP) immunoreactive substance, thus suggesting a role for VIP in the regulation of prolactin release.

The pituitaries of the dw and lit strains of mutant mice, congenitally deficient in prolactin-producing cells, and hyt mice, which were found to have reduced numbers of prolactin-producing cells, showed a markedly reduced VIP immunoreactivity. Hypothalamic VIP immunoreactivity, however, was found to be unchanged in the three strains of mutant mice, indicating that the high concentration of VIP in the hypothalamus does not derive from the adenohypophysis through retrograde flow. The deficiency in the mutant mice seems to be due to the lack of prolactin target cells in the pituitary.

J. Endocr. (1988) 118, 179–185
Correlation between myometrial receptor affinity, lipophilicity and antagonistic potency of oxytocin analogues in the rat
Authors: Atke A. , Vilhardt H. , Melin P.
Published: December 2008

Purified myometrial plasma membrane fractions were prepared from rats treated with oestradiol to induce oestrus. The binding affinities of 11 antagonistic oxytocin analogues to the oxytocin receptor of the plasma membranes were measured. Furthermore, lipophilicity of the peptides was assessed by reversed-phase high pressure liquid chromatography. No significant correlation was found between lipophilicity of the analogues and values for antagonistic potencies or binding affinities. Also, receptor-binding affinity did not correlate with in-vitro antagonistic activity whereas a significant correlation was obtained between binding affinities and in-vivo antagonistic potency for analogues void of partial agonist properties. It is concluded that neither receptor affinity nor lipophilicity in the analogues can predict the potency of the antagonists in vitro. However, receptor affinity was found to be a relatively good predictor of the in-vivo potency, while the usefulness of measuring antagonistic potency in vitro is questioned.

J. Endocr. (1988) 118, 187–192
Effects of protein kinase C activators on the in-vitro action of thyrotrophin in pigs
Authors: Ginsberg J. , Murray P. G.
Published: December 2008

The ability of the non-phorbol protein kinase C (PKC) activator 12-hydroxy-daphnetoxin (mezerein) to modulate differentiated thyroid function was examined in vitro. A dose-dependent inhibition of TSH-stimulated iodide organification was observed in porcine thyroid cells exposed to mezerein. Under identical conditions mezerein caused the translocation of PKC from its inactive cytosolic form to an active membrane-bound form in thyroid cell extracts. The relative biological potencies of mezerein and the phorbol ester, 12-0-tetradecanoylphorbol-13-acetate (TPA), to inhibit thyroid function in vitro corresponded to their abilities to activate PKC. This effect was also observed when dibutyryl cyclic AMP was used, implying a post-receptor site of action. To provide further evidence for this concept, the effects of mezerein and TPA on receptor-related events were studied. Neither mezerein nor TPA had any effect on the binding of radiolabelled TSH to solubilized porcine thyroid membranes. However, both mezerein and TPA were capable of stimulating cyclic AMP (cAMP) production in porcine thyroid cells in the basal state but could not augment TSH or forskolin-activated cAMP release. These data provide evidence that activation of PKC plays a role in the regulation of differentiated thyroid function in vitro and suggest that the effects of PKC are complex, with independent actions on cAMP accumulation and post-receptor events.

J. Endocr. (1988) 118, 199–203
Ontogeny of catecholamine-synthesizing enzymes and enkephalins in the sheep adrenal medulla: an immunocytochemical study
Authors: McMillen I. C. , Mulvogue H. M. , Coulter C. L. , Browne C. A. , Howe P. R. C.
Published: December 2008

An immunocytochemical staining technique was used to investigate the development of the sheep adrenal medullary cells containing enkephalins and the catecholamine synthetic enzymes dopamine β-hydroxylase (DBH) and phenylethanolamine N-methyl transferase (PNMT). No staining was observed in the adrenocortical cells with any of the antisera used in this study. Positive staining with anti-DBH was observed throughout the medulla in both adult and fetal adrenal glands from 80 days of gestation. Positive staining with anti-PNMT was observed in all glands from as early as 80 days of gestation, and staining with this antiserum was mainly confined to the peripheral medullary cells, which were adjacent to, and interdigitated between, the cells of the adrenal cortex. In the fetus between 80 and 120 days of gestation, staining for the enkephalins was observed in both the peripheral columnar and the central polygonal adrenal medullary cells. After 125 days of gestation and in the adult ewe, the peripheral columnar cells were uniformly stained with anti-enkephalin whereas many unstained cells were present in the central medullary region. Therefore, enkephalin-containing peptides are present in the catecholamine cells of the fetal and adult sheep adrenal and there appears to be a changing pattern in the distribution of the enkephalins in the fetal adrenal in late gestation.

J. Endocr. (1988) 118, 221–226
The mitogenic effects of corticotrophin-releasing factor on the anterior pituitary gland of the rat
Authors: McNicol A. M. , Kubba M. A. G. , McTeague E.
Published: December 2008

The immunohistochemically defined corticotroph population in the anterior pituitary gland of the adult male Sprague–Dawley rat has been quantified at 2 and 6 weeks after bilateral adrenalectomy using the stereological measurement, volume density (Vv). An approximately twofold increase in corticotroph Vv was demonstrated at 2 weeks in adrenalectomized rats compared with that in sham-operated controls and this was maintained at 6 weeks. Daily i.p. injections of ovine corticotrophin-releasing factor (CRF-41) induced a significant dose-related increase in corticotroph Vv when administered at doses of 25 and 50 μg/kg body weight, but this was less than the increase following adrenalectomy.

Assessment of changes in mitotic activity of the total anterior lobe of adult Sprague–Dawley rats and of the corticotroph population was also made after daily i.p. injections of 50 μg CRF-41/kg for 2 and 7 days. There was no increase in overall mitotic index at either time. However, the numbers of mitotic corticotrophs were significantly increased in CRF-injected animals compared with those in saline-injected rats. These results indicate a role for CRF-41 in the regulation of corticotroph growth.

J. Endocr. (1988) 118, 237–241
The influence of oestrogen and progesterone on the actions of two calcium entry blockers in the rat uterus
Authors: Downing S. J. , Hollingsworth M. , Miller M.
Published: December 2008

The potency and maximum effect of the calcium entry blocker nifedipine as an inhibitor of uterine contractions in vivo are increased in rats in late pregnant compared with non-pregnant rats. The influence of ovarian steroids produced during pregnancy (oestrogen and progesterone) on the potency and maximum effect of two calcium entry blockers (nifedipine and diltiazem) against uterine contractions during i.v. infusion was therefore investigated in anaesthetized non-pregnant rats. The influence of pregnancy on the relationship between serum concentrations of diltiazem during i.v. infusion and uterine and cardiovascular effects was also investigated.

A twofold increase in the potency of nifedipine as an inhibitor of uterine contractions was observed in rats treated with oestrogen or oestrogen plus progesterone compared with rats treated with corn oil. There was no change in potency in rats receiving progesterone alone. Maximum inhibition of uterine contractions by nifedipine was significantly increased by all three hormone treatments.

A twofold increase in the potency of diltiazem and a significant increase in maximum inhibition of uterine contractions was observed in rats in late pregnancy compared with non-pregnant rats. No increase in potency of diltiazem in reducing blood pressure or heart rate was observed in rats in late pregnancy. No significant difference in potency of diltiazem against uterine contractions was observed in rats treated with oestrogen, progesterone or oestrogen plus progesterone.

In order to determine if the hormone-induced changes in the potency of nifedipine against contractions in vivo were due to a direct effect of the ovarian steroids on voltage-operated calcium channels of the uterus or were mediated by extra-uterine mechanisms, the potency of nifedipine as a relaxant of uterine spasms in vitro was investigated. The potency of nifedipine in the isolated uterus as a relaxant of the spasm induced by KC1 did not differ between rats which received oestrogen, progesterone or oestrogen plus progesterone or corn oil. Additionally, the potency of nifedipine to inhibit oxytocin-driven phasic tension development was not affected by treatment with oestradiol.

These findings suggest that the increase in potency and maximum effect of nifedipine against uterine contractions observed previously in rats in late pregnancy can be ascribed, in part, to the action of the ovarian steroids (oestrogen and progesterone). The increase in potency of diltiazem observed in rats in late pregnancy, however, is due to factors other than oestrogen and progesterone. The lack of effect of hormonal manipulations on the potency of nifedipine on the isolated uterus suggests that the changes in potency in vivo due to oestrogen and progesterone are mediated by extra-uterine mechanisms.

J. Endocr. (1988) 118, 251–258
Relationships between prolactin, LH and broody behaviour in bantam hens
Authors: Sharp P. J. , Macnamee M. C. , Sterling R. J. , Lea R. W. , Pedersen H. C.
Published: December 2008

The interactions between broody behaviour and changes in concentrations of plasma prolactin and LH were investigated in bantam hens. Adoption of newly hatched chicks caused incubating hens to leave their nests and prevented plasma prolactin decreasing as rapidly as in hens deprived of their nests and not given chicks. Further, the hens allowed to rear chicks came back into lay later (P< 0·001) than the hens not allowed chicks. Plasma prolactin decreased and plasma LH increased in hens deprived of their nests: these changes were reversed when the hens re-nested. The changes in plasma LH and prolactin in nest-deprived and re-nesting birds were not always synchronous; this was particularly clear immediately after nest deprivation when the increase in plasma LH preceded the decrease in the plasma prolactin. Readiness to incubate disappeared between 48 and 72 h after nest deprivation and corresponded with the time when plasma prolactin decreased to baseline values. Administration of ovine prolactin depressed (P<0·01) the initial increase in plasma LH after nest deprivation, but repeated administration of prolactin for up to 72 h failed to suppress plasma LH to the values found in incubating hens. Repeated administration of ovine prolactin at 5- to 8-h intervals for 72 h maintained readiness to incubate in nest-deprived hens. It is concluded that the secretion of prolactin in broody hens is facilitated by the presence of chicks and that increased concentrations of plasma prolactin maintain incubation behaviour. In incubating hens the secretion of LH and prolactin may be partly regulated independently. In addition, LH secretion may also be inhibited by increased plasma prolactin.

J. Endocr. (1988) 118, 279–286
α-Amidated peptides derived from pro-opiomelanocortin in human pituitary tumours
Authors: Fenger M. , Johnsen A. H.
Published: December 2008

Human pituitary tumours, obtained at surgery for Cushing's disease and Nelson's syndrome, were extracted and the content and molecular forms of proopiomelanocortin (POMC)-derived peptides determined by radioimmunoassay, gel chromatography, reversed-phase high-performance liquid chromatography (HPLC) and sequence analysis. In the tumours from patients with Cushing's disease the mean concentrations of amidated peptides relative to the total amount of POMC were as follows: α-MSH, 1·7%; amidated γ-MSH (γ1-MSH), 8·5% and the peptide linking γ-MSH and ACTH in the precursor (hinge peptide or joining peptide) in its amidated form (HP-N), 17·1%. The same relative concentrations in the tumours from patients with Nelson's syndrome were 8·5% (α-MSH), 7·5% (γ1-MSH) and 12·2% (HP-N). More than 95% of the ACTH(1–39) immunoreactivity eluted as synthetic ACTH(1–39) by gel chromatography and HPLC. The remaining ACTH(1–39) immunoreactivity eluted as partly glycosylated high molecular weight forms. All the α-MSH and its glycine-extended precursor ACTH(1–14) were of low molecular weight, mainly non- or mono-acetylated forms, but significant amounts of diacetylated analogues were also present. γ1-MSH and γ2-MSH immunoreactivities eluted as high molecular weight forms and were partly glycosylated. No low molecular weight forms of γ1-MSH or γ2-MSH could be detected in the pituitary tumours. Amidated hinge peptide was mainly of the 30 amino acid form.

In conclusion, all the molecular forms of the amidated peptides detected in tumours from patients with Cushing's disease and Nelson's syndrome were similar to the molecular forms found in the normal human pituitary. The main difference between the tumours and the normal pituitary was the greater amount of peptides produced, particularly α-MSH and γ1-MSH.

J. Endocr. (1988) 118, 329–338
One-step purification of chicken growth hormone from a crude pituitary extract by use of a monoclonal immunoadsorbent
Authors: Berghman L. R. , van Beeumen J. , Decuypere E. , Kühn E. R. , Vandesande F.
Published: December 2008

The immunization of mice with an affinity-purified glycoprotein preparation from chicken pituitary tissue yielded several monoclonal antibodies towards the recently described glycosylated variant of chicken GH. As all these antibodies recognize the classical (non-glycosylated) GH molecule equally well, they provide a suitable tool for the development of both a specific immunoadsorbent and an assay method. This paper deals with the surprising purification power of the immunoadsorbent that was produced with one of the monoclonal antibodies. The resulting preparation was more than 99% pure as assessed by reversed phase high-performance liquid chromatography and sodium dodecyl sulphate-polyacrylamide gel electrophoresis, so that no further purification steps were needed before the determination of the amino acid sequence of the material. The efficiency of the purification protocol as determined by a homologous, monoclonal antibody-based radioimmunoassay was virtually absolute. Moreover, the affinity-purified GH preparation was a mixture representing the multiple molecular forms of pituitary chicken GH, including both oligomeres and glycosylated GH. The purified preparations were finally used to demonstrate the hepatic 5′-monodeiodinase-stimulating activity of GH in the chicken embryo (results not shown), in order to prove that the biological activity of the molecule had not been damaged by elution from the immunoadsorbent.

J. Endocr. (1988) 118, 381–387
Phorbol ester pretreatment attenuates the growth hormone (GH) response to GH-releasing factor in cultured rat pituitary cells
Authors: Ohmura E. , Okada M. , Ohba Y. , Onoda N. , Sano T. , Tsushima T. , Shizume K.
Published: December 2008

The effect of phorbol ester pretreatment on rat (r) GH release induced by GH-releasing factor (GRF) or 8-bromo-cyclic (c)AMP was investigated using rat pituitary cells cultured in monolayers. Pretreatment with 12-O-tetradecanoylphorbol-13-acetate (TPA) for 3 h significantly suppressed the rGH release induced by GRF, but not that by 8-bromo-cAMP 20 h later; this suppressive effect of TPA was concentration-dependent from 8 to 160 nmol/l, and complete suppression was observed after pretreatment with 80–160 nmol TPA/l. Production of cAMP by pituitary cells stimulated with GRF was similarly attenuated in TPA-pretreated cells. The rGH responsiveness to GRF of these cells was fully recovered on prolonged culture (40 h), suggesting that the inhibitory effect of TPA is reversible.

In contrast, pretreatment with GRF (5 nmol/l) resulted in suppression of the rGH response to subsequent exposure to GRF (5 nmol/l) or 8-bromo-cAMP (10 mmol/l), but not to TPA. These observations suggest that pretreatment with TPA modifies the rGH response to GRF at steps before the formation of cAMP.

J. Endocr. (1988) 118, 423–428
Tumour formation and insulin secretion by clonal RINm5F cells following repeated subcutaneous transplantation in NEDH rats
Authors: Flatt P. R. , DeSilva M. G. , Swanston-Flatt S. K. , Powell C. J. , Marks V.
Published: December 2008

The effects of repeated s.c. transplantation of clonal insulin-secreting RINm5F cells in NEDH rats on tumour morphology, insulin–glucose homeostasis and the function of RINm5F cells re-established in culture were examined after maintenance in vivo for periods of up to 308 days. Transplantation of RINm5F cells for ten consecutive passages consistently produced a single encapsulated vascularized tumour associated with the development in recipient rats of hyperinsulinaemia, hypoglycaemia and eventual neuroglycopenic coma. At the tenth passage, tumour-bearing rats exhibited a markedly enhanced rate of insulinstimulated glucose uptake by 19 days, with evidence of a large and variable insulin response to i.p. glucose. Survival was 22 ± 2 days, and resected RINm5F cell tumours exhibited weak immunostaining for insulin in scattered cells, with strands of fibrous tissue separating clusters of tumour cells many of which had distinct polarity. There was no detectable immunostaining for glucagon, somatostatin or pancreatic polypeptide. The insulin content and insulin secretory output of RINm5F cells re-established in culture after 20, 146, 259 or 308 days propagation in vivo were generally enhanced compared with non-passaged RINm5F cells. The magnitude of the effect was not appreciably affected by the duration of maintenance in vivo, but it was critically dependent upon the subsequent period of culture in vitro. Thus, whereas 2-day cultured RINm5F cells from the eighth tumour passage exhibited a greater than 100–fold increment of insulin content and release, with enhanced secretory responsiveness to leucine, arginine, theophylline, K + (25 mmol/l) or Ca2+ (7·6 mmol/l), RINm5F cells cultured for a further 19 days had almost completely lost the attributes resulting from 259 days of maintenance in vivo. The results indicate substantial enhancement of the functional capabilities of RINm5F cells in vivo, and suggest that the resulting tumours afford a novel model in NEDH rats of responsive trabecular-type insulinomas.

J. Endocr. (1988) 118, 429–437
Solid-phase iodothyronine-5′-deiodinase (5′-D) assays applied in production of monoclonal antibodies against 5′-D
Authors: Boye N. , Frøkiaer H. , Kaltoftt K. , Laurberg P.
Published: December 2008

Characterization of iodothyronine-deiodinating enzymes has been difficult due to loss of enzyme activity during purification. To obtain a new tool for studying these enzymes we investigated the possibility of developing monoclonal antibodies (MAbs) against iodothyronine-5′-deiodinase (5′-D). Two specific and sensitive solid-phase microassays were developed for screening hybridoma supernatants for the presence of antibodies inhibiting rat kidney 5′-D. and antibodies binding to but not inhibiting the enzyme.

BALB/c mice were immunized with a 3-((3-cholamidopropyl) -dimethylammonio) -1- propanesulphonate (CHAPS)-solubilized 5′-D-rich membrane preparation from rat kidney cortical tissue. Spleen cells were fused with NSI-Ag 4/1 mouse myeloma cells by means of polyethylene glycol.

Two hybridoma cell lines (AF5 and BE8) secreting MAbs specifically binding to without inhibiting 5′-D were produced. The AF5 antibody was of the IgG2a subclass and the BE8 antibody of the IgG2b subclass. Binding of one of the antibodies to the enzyme inhibited binding of the other in both an enzyme-linked immunosorbent assay (ELISA) and a specific enzymebinding assay. CHAPS-solubilized kidney microsomal fraction was chromatographed on a Sepharose 6B column. Elution profiles of 5′-D activity and MAb-binding antigens, as measured by ELISA with both AF5 and BE8, were identical.

Monoclonal antibodies should be valuable probes in the further elucidation of the nature of the iodothyronine-deiodinating activity in various tissues.

J. Endocr. (1988) 118, 439–445
Selective release of FSH in lactating rats during the period of follicular atresia induced by the administration of antiserum to LH-releasing hormone
Authors: Taya K. , Sasamoto S.
Published: December 2008

Passive immunoneutralization of LHRH by injecting a caprine antiserum to LHRH (LHRH-AS) in lactating rats nursing two pups on day 5 of lactation resulted in an immediate decline in concentrations of LH in the plasma during the 24-h study period, followed by a gradual increase to control levels 30 h later. Concentrations of oestradiol-17β and inhibin activity in ovarian venous plasma also decreased abruptly in LHRH-AS treated animals and recovered to control levels 36 h later. These changes were correlated with changes in concentrations of LH in the plasma. On the other hand, plasma concentrations of FSH increased abruptly in the LHRH-AS treated animals within 3 h after injection, but the concentrations declined gradually to control levels 48 h later. The ability of follicles to ovulate in response to human chorionic gonadotrophin (hCG) began to decrease within 6 h after treatment with LHRH-AS, and further decreased until 18 h after injection of LHRH-AS, when hCG induced ovulation (with two oocytes) in only one of five animals. A gradual increase in ovulation rate to control levels was noted by 36 h after injection of LHRH-AS. These results indicate that Graafian follicles present at the time of LHRH-AS injection had become atretic and that a new set of follicles had then begun to mature. Selective release of FSH could not be induced by injection of LHRH-AS in ovariectomized animals. Treatment with inhibin (porcine follicular fluid) suppressed the selective release of FSH, whereas treatment with oestradiol-17β had no inhibitory effect on the selective release of FSH.

These findings indicate (1) that tonic secretion of LH is an important factor in normal follicular maturation and maintenance in lactating rats, (2) that selective release of FSH after injection of LHRH-AS is attributed to the removal of a negative influence of inhibin from antral follicles during the period of follicular atresia and (3) that a selective surge of FSH is responsible for initiation of new follicular maturation.

J. Endocr. (1988) 118, 455–464
IDENTIFICATION OF mRNA FOR RELAXIN IN THE ENDOMETRIUM OF THE PREGNANT GUINEA-PIG
Authors: Tashima L. , Greenwood F. C. , Bryant-Greenwood G. D. , Peaker M.
Published: December 2008

The endometrium of late pregnant guinea-pigs was found to contain mRNA for relaxin. Poly(A)+RNA hybridized to 3 2P-labelled porcine relaxin-specific oligonucleotide probes. These probes corresponded to the C-peptide region of the prohormone. There was no hybridization to a 3 2P-labelled probe to the B-chain of porcine relaxin even under conditions of low stringency. The size of the relaxin mRNA was approximately 1.0 kb and similar to that found for relaxin mRNA from the pregnant sow ovary. This is the first study on relaxin mRNA from a uterine source.
Effect of aldrin on spermatogenesis, plasma gonadotrophins and testosterone, and testicular testosterone in the rat
Authors: Chatterjee S. , Ray A. , Ghosh S. , Bhattacharya K. , Pakrashi A. , Deb C.
Published: December 2008

Quantitative evaluation of the different varieties of germ cells at stage VII of the seminiferous epithelium cycle, namely type-A spermatogonia (ASg), preleptotene spermatocytes (pLSc), mid-pachytene spermatocytes (mPSc) and step 7 spermatids (7Sd), along with radioimmunoassay of plasma gonadotrophins (FSH and LH), testosterone and testicular testosterone were performed in Wistar rats following treatment with aldrin (polycyclic chlorinated hydrocarbon insecticide) for approximately one (13 days) or two cycles (26 days) of the seminiferous epithelium.

Extensive degeneration of all varieties of germ cells at stage VII, reduction in the sperm count and significant reductions in plasma concentrations of LH and testosterone were observed following aldrin treatment. The reduction in plasma concentrations of FSH was statistically significant only after treatment for two cycles. The inhibitory effect of aldrin on plasma gonadotrophins, testosterone levels, testicular testosterone content and numbers of 7Sd and ASg was maximum after treatment for two cycles. Administration of human chorionic gonadotrophin along with aldrin treatment for two cycles partially prevented the degeneration of germ cells and enhanced testosterone production.

The results indicate that aldrin may have a direct inhibitory influence on gonadotrophin release, but the possibility of a direct action of the insecticide at the level of the testes is also discussed.

J. Endocr. (1988) 119, 75–81
Comparison of plasma progesterone profiles in cyclic, pregnant, pseudopregnant and hysterectomized pigs between 8 and 27 days after oestrus
Authors: King G. J. , Rajamahendran R.
Published: December 2008

Plasma progesterone concentrations were compared in cyclic (n = 12), pregnant (n =12), oestradiol-induced pseudopregnant (n=12) and hysterectomized gilts (n=10) between days 8 and 27 after oestrus. The results were grouped into periods covering days 8–13, 14–20 and 21–27 and analysed by least-squares analysis of variance. Plasma progesterone concentrations were significantly (P<0·001) higher in hysterectomized compared with other groups between days 8 and 13. Progesterone concentrations declined rapidly after day 14 in cyclic females and gradually in the other groups. Throughout the third and fourth weeks the mean progesterone concentrations for hysterectomized animals were consistently higher than for pseudopregnant animals (P<0·05). The pregnant group means were below but not significantly different from the hysterectomized means in both of the last two periods. The greater progesterone concentrations in hysterectomized gilts indicated that secretion is high without any conceptus-produced or -mediated luteotrophin, and corpora lutea in cyclic, pregnant or pseudopregnant gilts may never reach full secretory potential.

J. Endocr. (1988) 119, 111–116
Ovarian inhibin content in immature female rats following hypophysectomy: control by exogenous gonadotrophins
Authors: Carson R. S. , McMaster J. W.
Published: December 2008

The effect of exogenous ovine gonadotrophins on ovarian inhibin content in hypophysectomized immature female rats was studied to determine which of the pituitary gonadotrophins control ovarian inhibin production in vivo. Inhibin and steroid contents of 27 000 g ovarian supernatants were measured by specific radioimmunoassay. Whereas injection of ovine (o)FSH resulted in dose-dependent increases in ovarian weight, progesterone and oestradiol content above that of control animals injected with diethylstilbestrol alone, injection of oLH was without significant effect. Similarly, ovarian inhibin content was not altered significantly by injection of oLH, but was increased in a dose-dependent manner by injection of oFSH. Concentrations of inhibin in serum were correlated directly with ovarian inhibin content. The results of this study indicate that ovarian inhibin content is increased by injection of oFSH but not oLH. It is likely, therefore, that inhibin production by small antral follicles is controlled by FSH activity alone.

J. Endocr. (1988) 119, 185–189