Showing papers in "Journal of Endocrinology in 1979"

Effects of lesions of the suprachiasmatic nuclei and of p-chlorophenylalanine on the circadian rhythms of adrenocorticotrophic hormone and corticosterone in the plasma, and on locomotor activity of rats.

Abstract: Adrenocorticotrophin (ACTH) and corticosterone in the plasma of adult female rats were measured sequentially at 4 h intervals for 24 h before and after lesions of the suprachiasmatic nuclei or treatment with p-chlorophenylalanine (to inhibit serotonin synthesis). After lesions or p-chlorophenylalanine treatment, the concentrations of ACTH were diminished relative to those in control animals and rhythmic changes could not be detected. However, injection of animals, pretreated with p-chlorophenylalanine, with 5-hydroxytryptophan (60 mg/kg) 8 h before the time when plasma ACTH is maximal in intact animals, stimulated ACTH secretion up to control values. Mean corticosterone concentrations in plasma remained unchanged (after lesions) or increased (after p-chlorophenylalanine). This increase was associated with an increased minimal concentration of corticosterone. After both treatments there was evidence of continued circadian or ultradian rhythms of corticosterone concentration. Locomotor activity of female rats given identical treatment, but without blood sampling, indicated that nocturnal activity was diminished after lesions whereas diurnal activity was enhanced after p-chlorophenylalanine treatment. Periodicity analysis detected the persistence of free-running circadian, and sometimes ultradian activity, rhythms. Adrenalectomy did not alter further the activity pattern observed in rats with lesions. These results therefore support the proposition that both the suprachiasmatic nuclei and the serotoninergic system play an irreplaceable role in the mechanism of ACTH secretory rhythms. The suprachiasmatic nuclei are also important for synchronization of locomotor activity and corticosterone rhythms, which may both persist after the suppression of ACTH rhythms.

Variations in concentrations of prolactin, luteinizing hormone, growth hormone and progesterone in the plasma of broody bantams (Gallus domesticus)

Abstract: The concentrations of prolactin, LH, progesterone and GH were measured in the blood of broody bantam hens. The concentration of prolactin was at its highest when the birds began to incubate their eggs and in six out of nine hens it tended to remain raised until the eggs hatched. The increase in the concentration of prolactin was small: in incubating hens it was only 23% higher than in hens caring for their young and 14% higher than in laying hens (P less than 0.05 for both comparisons). The concentration of GH tended to be depressed in hens caring for young but otherwise was not related to reproductive activity. The concentrations of LH and progesterone decreased at the onset of incubation and remained depressed while the hens sat on their eggs (P less than 0.001) for both comparisons). After the chicks hatched, the level of LH began to increase slowly whereas the level of progesterone remained low. The hens stopped showing broody behaviour between 4 and 10 weeks after the chicks had hatched; this corresponded to the time when the concentration of LH had increased to values found in laying hens. These observations provide some evidence that prolactin secretion increases at the onset of incubation and support the view that the hormone is not secreted at an increased rate while hens are caring for their young.

Varying response to luteinizing hormone of two luteal cell types isolated from bovine corpus luteum.

Abstract: Luteal cell suspensions obtained by enzymatic digestion of pregnant cow corpus luteum were found to be heterogenous and mainly made up of two types of cells of different sizes. The large cells (37 micrometers, average diameter) could be separated from the small ones (18 micrometers, average diameter) by sedimentation at unit gravity in a gradient of Ficoll-bovine serum albumin. A comparative in-vitro study of the synthesis of progesterone by the two types of cells indicated striking differences between them. The average content and the synthesis of progesterone in the absence and presence of a saturating dose of bovine LH after incubation for 2 h were 0.07, 0.12 and 6.9 pg/cell for the small cells and 0.65, 2 and 10 pg/cell for the large ones. Moreover, the sensitivity to low concentrations of LH was 100 to 1000 times higher for the small cells than for the large ones. Oestradiol-17 beta at concentrations ranging from 5 X 10(-10) to 5 X 10(-4) mol/l exerted a dose-dependent inhibition on the stimulation of LH in both cell types. These results suggest a possible involvement of both cell types in the synthesis of progesterone in vivo with a greater contribution by the small cells to stimulation induced by LH. Moreover, it appears that small cell suspensions could be a useful model system for in-vitro studies of the control of the synthesis of progesterone in cow corpus luteum.

Biological and binding activities of equine pituitary gonadotrophins and pregnant mare serum gonadotrophin

Abstract: The biological and binding activities of pregnant mare serum gonadotrophin (PMSG) were compared with those of highly purified FSH and LH from the pituitary gland of the same species. Pregnant mare serum gonadotrophin showed activity in bioassays considered to be specific for both FSH (e.g. the Steelman-Pohley ovarian augmentation test and cyclic AMP production by rat seminiferous tubules) and LH(androgen production by rat Leydig cells), as well as activity in a variety of radioreceptor assay systems previously considered to be specific for one of the two types of gonadotrophin. The potency of PMSG was high compared with that of purified ovine FSH or LH standards in all assays but PMSG was considerably less active than equine FSH and LH in vitro. In radioreceptor assays employing rat, pig and horse tissues, the activity of PMSG was equivalent to only 1--5% of equine FSH in competing for FSH-binding sites and only 3--35% of equine LH in competing for LH-binding sites. Pregnant mare serum gonadotrophin was least active in homologous binding assays with horse testis and equine LH as radioligand. In the rat Leydig cell bioassay, the activity of PMSG was only 2.0% that of equine LH. Furthermore, in some assays equine LH was found to resemble PMSG in exhibiting a high degree of FSH-like activity that could not be accounted for by cross-contamination. The FSH immunoactivity of equine LH was less than 0.5% that of equine FSH, but equine LH was up to 63% as potent as equine FSH in competition for FSH-binding sites and it was 20% as active in the Steelman-Pohley ovarian augmentation bioassay. Equine LH did not, however, show the expected activity in the cyclic AMP production bioassay. Thus, the FSH-binding sites and physiological receptors may not be identical. Overall, comparison of PMSG with pituitary gonadotrophins from homologous species shows that the apparent dual activity of PMSG may not be a unique feature of this pregnancy hormone since equine LH also exhibits some FSH activities. The chemical resemblance between PMSG and equine LH is noteworthy in this regard.

A radioimmunoassay for neurotensin in human plasma.

Abstract: A specific radioimmunoassay for neurotensin in human plasma has been developed capable of detecting changes of 5 pmol/l with 95% confidence. Neurotensin-like immunoreactivity has been detected in human plasma in two molecular forms and rises by 27 +/- 8 (S.E.M.) pmol/l (n = 9) after ingestion of food.

Ultradian and circadian rhythms in the plasma concentration of cortisol in sheep

Abstract: Three Merino ewes, adapted for about 3 weeks to their environment, were bled at 10 min intervals through a jugular venous cannula. Radioimmunoassay of plasma samples for cortisol revealed marked diurnal variations with peak levels just after midnight and lowest values in the afternoon. This rhythm appeared to result from a changing amplitude associated with a distinct ultradian rhythm (frequency 0.8-1.2 cycles/h) in the plasma level of cortisol. Calculation of the daily rate of secretion of cortisol from the hormone profiles gave a mean value of 8.49 mg. Arguments are put forward in favour of this method for obtaining the true rate of secretion of cortisol.

Comparison of prolactin levels in human semen and seminal plasma

Abstract: Semen samples were collected from 35 men and the levels of prolactin in semen and seminal plasma were measured. There was no significant difference in prolactin concentrations between the two fluids (t = 0.333, P greater than 0.7). There was also no correlation between the prolactin concentration and the kinematic viscosity of the semen (r = 0.065, P greater than 0.7).

Effects of chemoreceptor and baroreceptor stimulation on the discharge of hypothalamic supraoptic neurones in rats.

Abstract: Experiments have been performed to examine the effects of activating the carotid body chemoreceptors and the arterial baroreceptors on the discharge of neurones within the hypothalamic supraoptic nucleus of the rat Chemoreceptors were activated by intracarotid injection of 09% NaCl solution equilibrated with 100% CO2 The baroreceptors of the carotid sinus and aortic arch were activated by raising the blood pressure with an intravenous injection of phenylephrine Chemoreceptor stimulation activated and baroreceptor stimulation inhibited the discharge of all the phasically discharging neurones tested Neither stimulus had any consistent effect on non-phasically discharging neurones, although slight inhibition occasionally occurred Anaesthesia of the carotid bifurcation abolished the effects of cardiovascular stimulation on the supraoptic neurones Responses resumed when the anaesthesia wore off However, the anaesthesia also seemed to alter the phasic pattern of discharge The results are discussed with reference to the influence of the cardiovascular receptors upon the neurones in the supraoptic nucleus, and with reference to possible roles for the cardiovascular reflexes in control of vasopressin secretion

Dependence of spontaneous and angiotensin-induced drinking in the rat upon the oestrous cycle and ovarian hormones.

Abstract: The influence of the oestrous cycle on spontaneous and dipsogen-induced drinking was studied in female rats. Spontaneous fluid intake was lowest on the day of oestrus. Drinking induced by subcutaneous isoprenaline, and by angiotensin II (injected into the preoptic area), also showed marked cyclical variation, being lower at pro-oestrus and oestrus than at other stages of the cycle. Drinking induced by subcutaneous hypertonic NaCl or by intracranial carbachol did not vary with the oestrous cycle. Cyclicity of spontaneous and of angiotensin-induced water intake was not apparent in rats before puberty or after ovariectomy. Ovariectomy reduced drinking in response to isoprenaline. Treatment with oestradiol benzoate (20 micrograms) caused a reduction in spontaneous water intake, but a marked increase in the drinking response to isoprenaline. Treatment with oestradiol benzoate and progesterone (2.5 mg) caused a larger decrease in spontaneous water intake and an insignificant increase in isoprenaline-induced drinking. Water intake induced by subcutaneous hypertonic saline was unaffected by gonadal steroids. The results provide further evidence for the view that the thirst of extracellular origin, in which the renin-angiotensin system is involved, is brought about by mechanisms different from those that respond to cellular dehydration. Only drinking caused by activation of extracellular mechanisms appeared to be sensitive to the ovarian cycle and to ovarian hormones.

Effects of photoperiod on hypothalamic luteinizing hormone releasing hormone in the male hamster.

Abstract: Male hamsters were maintained on long (14 h light : 10 h darkness; 14L : 10D) or short (6L : 18D) photoperiods. Animals on short-days had reduced levels of LH in the serum and anterior pituitary gland, decreased androgen in the circulation and regressed testes and accessory sex organs. These same hamsters had significantly raised concentrations of hypothalamic luteinizing hormone releasing hormone (LH-RH). There was no significant difference in the response to exogenous LH-RH between groups maintained on long- and short-days. Castration significantly reduced levels of LH-RH in the hypothalamus in the long-day animals but had little effect on this parameter in short-day animals which had already undergone testicular regression. The increased levels of LH-RH in the hyothalami of both intact and castrated hamsters on non-stimulatory photoperiods is interpreted as a decreased release of the neurohormone which subsequently results in a decreased release of LH.

Photoperiodic control of seasonal breeding in the ram: participation of the cranial sympathetic nervous system

Abstract: Four mature Soay rams, cranially sympathectomized by removal of the superior cervical ganglia, were housed alongside four normal rams controlled lighting conditions of alternating 16 week periods of short days of 8 h light: 16 h darkness (8L : 16D) and long days (16L : 8D). The changes in the concentration of FSH, LH, prolactin and testosterone in the plasma, the size of the testes, the intensity of the sexual flush and the sexual and aggressive behaviour of the animals were recorded. While the control rams were able to respond to the artificial lighting conditions with synchronized cycles of reproductive activity, the ganglionectomized animals failed to respond. The treated rams had well-developed testes and relatively high levels of gonadotrophins and testosterone in the blood throughout the experiment. It is concluded that the cranial sympathetic nervous system is involved in the photoperiodic control of seasonal breeding in the ram, probably through its role in the innervation of the pineal gland.

Synthesis of prolactin by human decidua in vitro.

Abstract: To determine whether human decidua and/or chorion synthesizes and secretes prolactin, explants of decidua obtained at Caesarian section and explants of chorion from the membranes separating dizygotic twins were cultured for periods of up to 6 days. The decidual explants released 366 +/- 37 ng prolactin/100 mg tissue (mean +/- S.D.) during each day in culture and incorporated 3H-labelled amino acids into immunoprecipitable prolactin. In the radioimmunoassay for prolactin, serial dilutions of incubation medium displaced 125I-labelled prolactin parallel to the displacement by pituitary prolactin and the prolactin in the medium eluted from Sephadex G-150 in a position indentical to that of pituitary prolactin. Chorionic explants released prolactin into the incubation medium during day 1 of culture only and did not incorporate 3H-labelled amino acids into prolactin. These results demonstrate that prolactin is synthesized by the decidua and not by the chorion and suggest that the decidua is the source of prolactin in amniotic fluid.

A bioassay for inhibin using pituitary cell cultures.

Abstract: A bioassay for inhibin based on the suppression of gonadotrophin releasing hormone (Gn-RH)-stimulated secretion of FSH by primary monolayer cultures of rat anterior pituitary cells is described. The cultures were exposed to standard or test materials for 3 days. The levels of FSH in the media were measured by radioimmunoassay after exposure for 6 h to a maximally stimulating concentration of Gn-RH (10 nmol/l). The standard was prepared from ovine testicular lymph. Several preparations of proteins from gonadal tissues or secretions suppressed the levels of FSH in parallel with the standard. The levels of LH were also reduced but higher doses of active material were required. Non-specificity from cell damage and inactivation of Gn-RH have been excluded. The secretion of gonadotrophins by the pituitary cells was also inhibited by androgens, but not in parallel with the standard and secretion of LH was affected more than that of FSH. Control lymph protein preparations from castrated sheep had no detectable activity. The assay was sensitive and had adequate precision and practicability. It has proved useful for monitoring preliminary steps in the purification of inhibin.

Bioassay of inhibin-like activity using pituitary cells in vitro.

Abstract: The secretion of follicle-stimulating hormone (FSH) by the pituitary gland appears to be partially regulated by a protein hormone, inhibin, which is produced in the Sertoli cells of the testis and is also present in ovarian follicular fluid (FF). The aim of the present study was to develop a sensitive method for the detection and estimation of inhibin-like activity, using dispersed pituitary cells in culture. Pituitary cells from adult male rats were dispersed and precultured for 3 days. After renewal of the medium (2 ml), samples to be tested for inhibin-like activity were added and culture was continued for a further 3 days. A dose-dependent suppression of the concentration of FSH in the medium (lambda = 0.17--0.22) was observed after addition of FF (0.05--1 microliter) or Sertoli cell culture medium (SCCM, 0.05--1 ml). Luteinizing hormone (LH) concentrations were not affected with these doses of FF, but SCCM and higher doses of FF caused a significant increase in the concentration of LH in the medium. During an additional 6 h of culture in the presence of luteinizing hormone releasing hormone (LH-RH), FF and SCCM suppressed the release of both FSH (lambda = 0.07--0.11) and LH in a dose-dependent way. Cellular content of FSH, but not of LH, was decreased after these treatments. These results could not be explained by damage to the pituitary cells, by degradation of FSH or LH-RH, or by effects of steroids. It is concluded that this pituitary cell culture system can be used as a sensitive method for the estimation of inhibin-like activity in FF and SCCM.

Determination of tamoxifen and an hydroxylated metabolite in plasma from patients with advanced breast cancer using gas chromatography–mass spectrometry

Abstract: An assay has been established for the selective measurement of tamoxifen and its monohydroxy derivative, metabolite B, in human plasma. The assay was used to examine the concentrations of these compounds, relative to oestradiol-17 beta, in the plasma of patients undergoing tamoxifen therapy for advanced breast cancer. Oral administration of the drug (20 mg twice a day) raised the level of tamoxifen in plasma to approximately 200 ng/ml 20 days after the commencement of treatment. This level was 3000-fold higher than the corresponding concentration of oestradiol which remained within the range for post-menopausal women. Metabolite B was present in plasma at a much lower concentration than tamoxifen although in considerable excess over oestradiol. The overall results are discussed in relation to the possible mechanism of action of the drug.

Ontogeny and control of prolactin receptors in the mammary gland and liver of virgin, pregnant and lactating rats.

Abstract: Prolactin receptors were identified and partially characterized in the mammary gland of the rat. The binding of 125I-labelled ovine prolactin to a subcellular particulate fraction of rat mammary gland decreased between days 30 and 100 of age. Over the same period, binding to the liver increased and there was a significant negative correlation between prolactin binding in the two tissues. Binding to the mammary gland was low during pregnancy, increased in early lactation and declined after the litters were weaned. Binding to the liver was lower during lactation than during pregnancy or the period after weaning suggesting that tissue-specific factors may operate in the control of this receptor. In virgin rats, prolactin binding by the mammary gland was increased by oestrogen. This effect was blocked by hypophysectomy and partially restored by replacement therapy with prolactin. Hypothyroidism and treatment with progesterone also reduced the response to oestrogen. The maintenance of prolactin binding by the mammary gland of lactating rats depends on the presence of the ovaries and pituitary, thyroid and adrenal glands. Examination of the ratio epithelium: stroma suggests that prolactin acts by increasing the number of epithelial cells in the mammary gland and that thyroid, adrenal and ovarian hormones modulate the number of receptors per cell.

Priming effect of luteinizing hormone releasing factor in vitro: role of protein synthesis, contractile elements, Ca2+ and cyclic AMP.

Abstract: The mechanism of the priming effect of luteinizing hormone releasing factor (LH-RF) upon gonadotrophin secretion was studied using short-term incubation of hemipituitary glands from pro-oestrous rats. The dependence of the priming, but not the LH releasing action of LH-RF on protein synthesis in pituitary tissue was confirmed. Cytochalasin B failed to affect the first response to LH-RF, but abolished the priming effect, suggesting that the integrity of cellular microfilaments was essential. Colchicine and vinblastine did not modify the response to LH-RF. Neither inhibitors of DNA nor the inhibitor of RNA polymerase II, alpha-amanitin, significantly affected the priming action of LH-RF. Normal extracellular concentrations of Ca2+ were necessary for gonadotrophin release, but the priming effect was not significantly affected by low extracellular Ca2+ and could not be elicited by raising intracellular Ca2+ concentrations. Adenosine 3':5'-cyclic phosphate did not appear to act as a second messenger for either the gonadotrophin releasing or the priming action of LH-RF.

Increased ratio of 5 alpha-reductase: 3 alpha (beta)-hydroxysteroid dehydrogenase activities in the hyperplastic human prostate.

Abstract: The activities of 5 alpha-reductase and 3 alpha (beta)-hydroxysteroid dehydrogenase were assayed in homogenates of eight normal, 21 hyperplastic and four carcinomatous human prostates. Samples consisting of 300--500 microgram tissue protein in Tris buffer, pH 7.0, were incubated at 37 degrees C for 30 min in the presence of 50 nM-[3H]androgen and an NADPH-generating system started with 5 X 10(-4)M-NADP. The yield of 5 alpha- and 3 alpha-reduced metabolites, as established by using t.l.c. and g.l.c., gave an estimate of enzyme activity. The formation of metabolites denoting 5 alpha-reductase activity in normal, hyperplastic and carcinomatous tissue respectively was 28.8 +/- 47 (S.E.M.), 76.8 +/- 8.9 and 3.5 +/- 0.7 pmol 30 min-1 mg protein-1; similarly, that denoting 3 alpha (beta)-hydroxysteroid dehydrogenase activity was 69.3 +/- 6.7, 46.6 +/- 5.7 and 38.8 +/- 22.1 pmol 30 min-1 mg protein-1. In all normal prostates 5 alpha-reductase activity was lower than 3 alpha (beta)-hydroxysteroid dehydrogenase activity. Conversely, in 18 out of 21 hyperplastic prostates, 5 alpha-reductase activity was higher than 3 alpha (beta)-hydroxysteroid dehydrogenase activity. The effect of the increase in 5 alpha-reductase activity without a compensatory change in 3 alpha (beta)-hydroxysteroid dehydrogenase activity was to alter the mean ratio between 5 alpha-reductase and 3 alpha (beta)-hydroxysteriod dehydrogenase activities from 0.47 +/- 0.11 in the normal prostate to 1.84 +/- 0,19 in hyperplastic tissue. It is inferred that this change may predispose the hyperplastic prostate to asymmetrical rates of androgen metabolism and thereby contribute to the abnormal accumulation of dihydrotestosterone.

Dependence of testicular germ cells on hormones: a quantitative study in hypophysectomized testosterone-treated rats

Abstract: Quantitative analysis of spermatogenesis was used to investigate the dependence of testicular germ cells on hormones in Sprague-Dawley rats. Adult hypophysectomized rats were given daily injections of testosterone propionate (TP) for 30 days. Intact and hypophysectomized control rats received vehicle only. Spermatogonia were classified as undifferentiated or differentiated, using established criteria suitable for morphological identification on periodic acid-Schiff's-haematoxylin stained sections of testis. Data on cell counts showed that the undifferentiated spermatogonia may be partially dependent on TP and/or pituitary hormones. The group of differentiated spermatogonia were dependent on pituitary hormones, and TP only partially restored their number by partially protecting them from spontaneous degeneration in stages XIV to II (A3-to-Intermediate). The maturation and division of B type spermatogonia and maturation of preleptotene spermatocytes to the zygotene stage appeared to be independent of hormones. Maturation of pachytene spermatocytes was hormone dependent, and TP completely supported their development, meiotic division and spermiogenesis in the complete absence of pituitary hormones.

Large-scale preparation of highly purified pyrogen-free human growth hormone for clinical use

Abstract: A method is described for the large-scale isolation of highly purified human growth hormone for therapeutic use. The hormone was extracted from frozen pituitary glands under mildly alkaline conditions. Purification was effected by gel filtration and ion-exchange chromatography. The final product was pyrogen-free and had a potency of 2.5 i.u./mg measured by bioassay against the current international standard. The yield of growth hormone/gland was approximately 5 mg.

Plasma oxytocin and steroid concentrations during late pregnancy, parturition and lactation in the miniature pig.

Abstract: Plasma oxytocin concentrations were measured during late pregnancy, parturition and lactation in the miniature pig. Measurements were made of plasma oestradiol, oestrone and progesterone to determine whether there was any relationship between the concentrations of oxytocin and these steroids in the circulation. Plasma oxytocin concentrations were low or undetectable in late pregnancy. Rises of up to 68.8 mum./ml were seen at the time of delivery of the foetuses and at the expulsion of the placenta. The only steroid that seemed to relat to oxytocin release was progesterone. Oxytocin release was consistently seen when progesterone concentrations had fallen to below 10 ng/ml but no increase in concentration was observed while oestrone and oestradiol increased to their maximum concentrations of 3.86--11.6 and 0.43--0.70 ng/ml respectively. During lactation, when both oestrogen and progesterone concentrations were low, suckling caused the levels of oxytocin to increase to 7.4 muu./ml. These increases were greater during the first 2 weeks of lactation than later.

Acetylcholine stimulates growth hormone secretion, phosphatidyl inositol labelling, 45Ca2+ efflux and cyclic GMP accumulation in bovine anterior pituitary glands.

Abstract: Acetylcholine (25 micromol/l) in the presence of the choline esterase inhibitor physostigmine (67 micromol/l) increased the release of growth hormone and efflux of 45Ca2+ from perifused bovine pituitary slices; the time taken for the maximal response to occur was the same. In batch incubations, acetylcholine (1 micromol/l--1 mmol/l) increased pituitary cyclic GMP concentrations in the pituitary gland within 2 min, and increased incorporation of [3H]inositol and [32P]phosphate into pituitary phosphatidyl inositol within 15 min. Cyclic AMP concentrations were not significantly changed 2 or 5 min after acetylcholine addition. All the tissue responses were inhibited by prior exposure of the tissue to atropine (1 micromol/l) but not by tubocurarine (10 micromol/l--1mmol/l), indicating that the responses were mediated by receptors of the muscarinic type. The similarities between these responses and those to known hypothalamic hypophysiotrophic hormones are discussed.

Cellular localization of a prolactin-like antigen in the rat brain.

Abstract: The distribution of immunoreactive neurones and fibres was studied in rat brain using an antiserum to rat prolactin. Neurones containing the immunoreactive material were localized in the arcuate, ventromedial, premamillary, supraoptic and paraventricular nuclei of the hypothalamus. Immunoreactive nerve fibres were widely distributed within the brain. No differences were observed in labelling between male and female rats, or as a consequence of hypophysectomy.

Highly specific binding of 1,25-dihydroxycholecalciferol in bone cytosol.

Abstract: A method developed initially for the detection of high-affinity binding of glucocorticoids in the cytosol from foetal rat calvaria has been adapted for metabolites of vitamin D. Consistent displacement of [3H]1,25-dihydroxycholecalciferol ([3H]1,25(OH)2D3) by unlabelled 1,25(OH)2D3 was obtained with an apparent dissociation constant (Kd) of 2.3 x 10(-9) mol/l. The specificity of this binding was examined by competition experiments. Displacement of labelled 1,25(OH)2D3 by a 100-fold excess of unlabelled metabolites, expressing the fall with unlabelled 1,25(OH)2D3 as 100%, was as follows: 25-hydroxycholecalciferol (25(OH)D3), 61%; 24,25-dihydroxycholecalciferol, 29%; cholecalciferol, 3%. These are similar to results for the chick mucosa nuclear 1,25(OH)2D3 receptor. No displacement was obtained with corticosterone, testosterone, oestradiol or progesterone. When [3H]25(OH)D3 was used as ligand, a displacement curve with unlabelled 25(OH)D3 indicated only binding with a greater Kd (approximately 10(-7) mol/l). These data suggest a direct action of 1,25(OH)2D3 on bone which is similar to that of steroid hormones on their target tissues.

Reduction of non-specific serum responses in human pituitary gonadotrophin radioimmunoassays.

Abstract: Serum and plasma from human and domestic animals contain variable amounts of non-specific material(s) which may be mistaken for hormone in assays for human LH and FSH, based upon antisera of high sensitivity and hormonal monospecificity. The non-specific response curves are generally, but not invariably, less steep than those of the hormone standards and endogenous homologous hormones. The levels of non-specific intrusion can be of sufficient magnitude to obscure specific estimations seriously, particularly at low hormone levels, unless the assays are designed to minimize this effect. The non-specific effects could be minimized (but not abolished) by careful optimization of the assays which involved making the response curve as sensitive as possible and incorporating the serum at a final dilution of 1 : 2, since further dilution increased the relative contribution of the non-specific substance(s). The optimized assays require only 48 h of total incubation and show a sevenfold increase in the mean concentration of LH between sera from prepubertal children and adults accompanied by a mean threefold difference in the concentration of FSH.

Dopaminergic control of oxytocin release in lactating rats.

Abstract: During suckling, anaesthetized lactating rats release regular (about every 7 min) but brief pulses of oxytocin (0.5--1.0 mu.) which produce single transient increases in intramammary pressure. Drugs which selectively impair synaptic transmission were used to determine the role of dopamine and noradrenaline in regulating this natural reflex. Diethyldithiocarbamate (100--200 mg/kg, i.v.) and alpha-methylparatyrosine (100--400 mg/kg, i.v.) which inhibit the synthesis of catecholamines both blocked the suckling-induced release of oxytocin. The milk-ejection reflex was also inhibited in a dose-dependent manner by the intravenous administration of the dopamine antagonists, fluphenazine (0.7 mg/kg), pimozide (1.4 mg/kg), cis-dupenthixol (4.5 mg/kg) and metoclopramide (6.0 mg/kg), and caused a significant inhibition P less than 0.01) of the reflex in 50% of the rats tested. The alpha-adrenoceptor antagonist phenoxybenzamine (1.4 mg/kg) was similarly effective. Dopamine (40 micrograms), bromocriptine (10 micrograms), apomorphine (100 micrograms), noradrenaline (10 micrograms) and phenylephrine (2 micrograms) injected into the cerebral ventricles evoked a sustained release of oxytocin which produced multiple increases in intramammary pressure; isoprenaline (4 micrograms) was ineffective. The release of oxytocin evoked by dopamine and noradrenaline was prevented by cis-flupenthixol and phenoxygenzamine respectively. None of the drugs used affected the mammary sensitivity to exogenous oxytocin nor were their actions modified by pretreatment with propranolol (1 mg/kg). The results suggest that the neural pathway for the reflex release of oxytocin during suckling in the rat contains both dopaminergic and noradrenergic synapses, the latter acting through alpha-adrenoceptors and being distal in the pathway to the dopaminergic component.

Adrenocorticotrophin-related peptides in adult and foetal sheep pituitary glands.

Abstract: Differences in foetal and adult adrenal function may be due to qualitative as well as quantitative changes in the pituitary corticotrophic stimulus. Pituitary glands from adult and foetal sheep were freshly dissected and stored at -70 degrees C until extracted at pH 1.5. The extracts were subjected to chromatography on Sephadex G-100 superfine and fractions were assayed by multiple radioimmunoassays directed against the NH2- and CO2H-terminal sequences of ACTH and lipotrophin (LPH). Peaks corresponding to beta-melanocyte-stimulating hormone (beta-MSH), beta-LHP, gamma-LPH, beta-endorphin and ACTH were identified, with little or no evidence for the presence of alpha-MSH and corticotrophin-like intermediate lobe peptide. Three peaks of large molecular weight material, A. B and C, were identified and their relative proportions shown to be considerably greater in the foetus than in the adult. The immunoassay profile of peaks A and B suggested that they were 'stem hormones' which could give rise to a family of biologically active peptides. Since the 'family tree' which they engender varies according to the stage of development, it is proposed that the changes in the 'trophic family' may explain the different adrenal responses of the foetal and adult sheep.

Influence of androgens on the weights of the male accessory reproductive organs and on the activities of mitochondrial enzymes in the epididymis of the rat

Abstract: The influence of androgens on the male accessory glands of the rat was assessed in terms of changes in weight and of the specific activity of the mitochondrial enzymes, succinate dehydrogenase, glycerolphosphate dehydrogenase and pyruvate carboxylase, in the epididymis. In some instances, the activity of the cytoplasmic enzymes, hexokinase and phosphofructokinase, was also measured and the influence of androgens on these enzymes was found to be similar to that on the mitochondrial enzymes. After the administration of androgen to castrated rats the specific activity of enzymes reached a new steady state sooner than did epididymal weight. The time taken for the specific activity of the enzymes to reach a new steady state after the removal of androgen was variable, depending on the enzyme and the region of the epididymis. This time was generally longer, however, than the time taken for induction, and in the case of glycerolphosphate dehydrogenase, the decline of activity was slower in the cauda than in the caput. In castrated animals, about 100 times as much androgen was required to attain maximum tissue weight as was required to attain maximum enzyme activity. The epididymis, prostate and seminal vesicles responded similarly to androgen in terms of the dose-response pattern and the time taken for tissue weight to attain a new steady-state value, although the gain in weight of the epididymis relative to its weight in unstimulated control animals was less than the relative gain of the other accessory glands. Enzymes in the cauda epididymidis required lower amounts of androgen to elicit maximum activity than were required by those in the caput. The rate of change in the accessory glands in attaining new steady-state levels of tissue weight and enzyme activity was independent of the dose of androgen except during the first few days of hormone administration. Androgens were the most effective steroids in stimulating an increase of tissue weight and enzyme activity, although some changes were induced by oestradiol-3-benzoate and progesterone.

Relaxin inhibits spontaneous and prostaglandin-driven myometrial activity in anaesthetized rats.

Abstract: Porcine relaxin (250 guinea-pig units/mg) infused intravenously into anaesthetized rats at 20 micrograms/h reversibly abolished spontaneous intra-uterine pressure cycles yet left the myometrium responsive to oxytocin in doses of 4--8 mu. The inhibition was found to be primarily of the frequency, rather than of the amplitude, of pressure cycles. Relaxin (5 or 10 micrograms) was capable of completely suppressing uterine activity driven by prostaglandin F2 alpha infusion in oestrogen-treated ovariectomized rats. Whereas the beta-adrenergic blocker, propranolol, had no effect on relaxin-induced inhibition, phentolamine, an alpha-blocker, significantly delayed the relaxin effect. It is unlikely, however, that relaxin operates through an alpha-inhibitory receptor. The results show that relaxin acts primarily as a frequency modulator and is capable of antagonizing an exogenous myometrial stimulant.