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W. H. OWEN and D. R. IDLER

SUMMARY

Cortisol and cortisone were identified and their levels determined in sea raven plasma by a double isotope derivative assay involving acetylation with [1-3H]acetic anhydride, purification by thin-layer and paper chromatography, followed by recrystallization to constant 3H: 14C ratios. The mean level of cortisol in four plasma samples was 7·2±1·0 μg/100 ml (range 4·0–9·2) and the mean level for cortisone in three samples was 1·1 ± 0·2 μg/100 ml (range 0·7–1·5).

Metabolic clearance rates (MCR) were determined for both corticosteroids by the method of continuous infusion over an 8-h period. The mean MCR for cortisol in five fish was 126 ± 17 ml/kg/h, and for cortisone 449 ± 48 ml/kg/h in six fish. The mean percentage conversion of [1,2-3H]cortisol to cortisone was 9·4 ± 2·9%. There was no evidence of any significant conversion of cortisone to cortisol.

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CAROL A. HARVEY and D. A. A. OWEN

SUMMARY

The rates of uterine and ovarian blood flow during the oestrous cycle in rats were measured using radioactive microspheres. Blood flow was highest in the ovaries and uteri during pro-oestrus and lowest during metoestrus. During pro-oestrus, mean ovarian blood flow was 676·2 ± 183·6 (s.d.) ml/min/100 g wet tissue and mean uterine blood flow was 249·7 ± 120·1 ml/min/100 g. During metoestrus mean ovarian blood flow was 117·4 ± 19·8 ml/ min/100 g and mean uterine blood flow was 38·5 ± 7·4 ml/min/100 g. In ovariectomized rats, uterine blood flow was 28·7 ± 10·5 ml/min/100 g.

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F. J. Owen, M. H. Cake, and S. D. Bradshaw

A progesterone receptor system, with a high specificity for progestins, was detected in the uterine tissue of the marsupial, Setonix brachyurus (quokka), using the synthetic progestin 17α,21-dimethyl-19-nor-4,9-pregnadiene-3,20-dione (R5020). The apparent equilibrium dissociation constant of the ligand binding to the cytosolic component was 2·2 nmol/l, and to the nuclear component 4·8 nmol/l. Significant loss of binding ability of the receptor occurred when cytosol was pretreated with dextran-coated charcoal. All binding studies were performed, therefore, in the presence of endogenous steroid which was demonstrated to affect the dissociation constant but have no effect on the estimation of the concentration of binding sites. Cytosolic binding was increased sixfold by oestradiol-17β treatment in vivo, and the translocation of the bound complex into the nucleus was effected by progesterone. It is suggested that the binding component described plays a role in the action of progesterone on the uterine tissue of the quokka.

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M. H. CAKE, F. J. OWEN, and S. D. BRADSHAW

The plasma progesterone concentrations during pregnancy and the oestrous cycle of the quokka were measured daily after each had been initiated by the removal of pouch young. Progesterone levels ranged from 0·6 ng/ml in the early stages of the oestrous cycle to about 2·5 ng/ml at the peak of the luteal phase. There was no significant difference between pregnant and non-pregnant states before the removal of the pouch young nor in the latter half of the cycle. However, the plasma progesterone concentration on days 3–4 after removal of the pouch young was significantly greater in pregnant animals when compared with nonpregnant animals at the same stage and also when compared with the levels before removal of young. This early peak in the concentration of progesterone in peripheral plasma is discussed in relation to the development of the previously dormant blastocyst.

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L Morgan, J Arendt, D Owens, S Folkard, S Hampton, S Deacon, J English, D Ribeiro, and K Taylor

This study was undertaken to determine whether the internal clock contributes to the hormone and metabolic responses following food, in an experiment designed to dissociate internal clock effects from other factors. Nine female subjects participated. They lived indoors for 31 days with normal time cues, including the natural light: darkness cycle. For 7 days they retired to bed from 0000 h to 0800 h. They then underwent a 26-h 'constant routine' (CR) starting at 0800 h, being seated awake in dim light with hourly 88 Kcal drinks. They then lived on an imposed 27-h day (18 h of wakefulness, 9 h allowed for sleep), for a total of 27 days. A second 26-h CR, starting at 2200 h, was completed. During each CR salivary melatonin and plasma glucose, triacylglycerol (TAG), non-essential fatty acids (NEFA), insulin, gastric inhibitory peptide (GIP) and glucagon-like peptide-1 (GLP-1) were measured hourly. Melatonin and body temperature data indicated no shift in the endogenous clock during the 27-h imposed schedule. Postprandial NEFA, GIP and GLP-1 showed no consistent effects. Glucose, TAG and insulin increased during the night in the first CR. There was a significant effect of both the endogenous clock and sleep for glucose and TAG, but not for insulin. These findings may be relevant to the known increased risk of cardiovascular disease amongst shift workers.

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A P D Lord, L C Read, P C Owens, A A Martin, P E Walton, and F J Ballard

Abstract

Halothane anaesthesia in young sheep results in greatly increased plasma binding capacity for radiolabelled insulin-like growth factor (IGF), as demonstrated using size-exclusion chromatography. Most of the increased binding was at an estimated molecular mass range of 30–50 kDa, with a smaller increase evident at 130–150 kDa. These changes were not evident in control animals which had food withheld for the same period. The progressive increase in plasma radioligand binding during anaesthesia was the net result of a rise in circulating levels of a 29–31 kDa IGF-binding protein (IGFBP), as shown by ligand blotting, and declining plasma concentrations of IGF-I and IGF-II. Recovery from anaesthesia was accompanied by the restoration of plasma IGFs and the IGFBP towards pre-anaesthesia concentrations. The induced IGFBP was provisionally identified as IGFBP-1 because it bound anti-IGFBP-1 antiserum but not antibodies against IGFBP-2, IGFBP-3 or IGFBP-4. The elevation of plasma IGFBP-1 immunoreactivity was associated with reduced concentrations of glucose and insulin, the regulators of IGFBP-1 in humans and rats. These results suggest that IGF experiments that require anaesthesia but assume that the anaesthetised state is representative of conscious sheep should be reassessed. A similar situation may occur with other mammalian species.

Journal of Endocrinology (1994) 141, 427–437

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A A Al-Habsi, A Y A AlKindi, I Y Mahmoud, D W Owens, T Khan, and Aisha al-Abri

Circulating estradiol (E2), progesterone (Pro), testosterone, and corticosterone (B) levels were monitored in the green turtles Chelonia mydas during different nesting phases. Successful nesting includes emergence from sea, chamber and nest excavation, oviposition, burying the nest, and returning to sea. Unsuccessful nesting includes chamber and nest excavations but without oviposition. Blood samples were taken from the cervical sinus and collected within 5-min of capture to minimize stress. The samples were collected between 2000 and 0100 h during the peak season (May–October). High-performance liquid chromatography using a u.v. detection system coupled with tandem quadrupole mass spectrometry was used to measure B. Plasma B levels were significantly higher in successful and unsuccessful phases over emergence and excavation phases. However, B levels in successful versus unsuccessful or emergence versus excavation phases were not significantly different. Plasma steroid levels were measured by the Coat-A-Count RIA technique. Pro levels were significantly higher (P<0.005) in successful over unsuccessful turtles and also successful turtles over turtles in the other phases (P<0.01). The Pro levels immediately after nesting were found to be higher than that reported previously. Plasma testosterone values were higher in successful turtles but not significantly different from the turtles in other phases. Estrogen levels were undetected in all phases. Overall, the hormone values during different phases of nesting may play a major role in formulating the nesting behavior and physiology of the nesting activities in the green turtle.

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M. J. Waters, V. H. Oddy, C. E. McCloghry, P. D. Gluckman, R. Duplock, P. C. Owens, and M. W. Brinsmead

ABSTRACT

The physiological role of placental lactogen (PL; chorionic somatomammotrophin) in the ewe has been investigated by infusion of ewes (n = 3) on day 131 of pregnancy with sufficient ovine PL (oPL) antibody to neutralize circulating oPL for at least 12 h. Effectiveness of the antibody neutralization was defined both in vitro and in vivo according to rigorous criteria. Control ewes (n = 3) were infused simultaneously with an equivalent amount of pooled goat gamma globulin. Since both sets of ewes had previously been catheterized with jugular, utero-ovarian and femoral vein catheters and a femoral arterial catheter, it was possible to measure whole body glucose kinetics as well as muscle and uterine glucose, free fatty acid (FFA) and 3-hydroxybutyrate extraction. In addition, plasma levels of insulin, GH, prolactin, insulin-like growth factor-I (IGF-I), IGF-II, progesterone and cholesterol were determined in femoral arterial samples.

Neutralization of maternal oPL did not significantly affect whole body glucose metabolism, uterine and muscle glucose extraction, or 3-hydroxybutyrate extraction by muscle. A trend towards lower plasma FFA levels was observed after prolonged infusion, but was not statistically significant. However, plasma insulin levels rose significantly during antibody infusion after an early fall. These observations are rationalized in terms of the known requirements of ruminant metabolism during pregnancy, and contrasted with the accepted model for the role of human PL in the metabolic adjustments of pregnancy.

No change in plasma IGF-I, IGF-II or GH was observed, providing no support for the concept that oPL is responsible for maternal somatomedin generation during pregnancy. Similarly, plasma prolactin did not differ between antibody-treated and control groups. Finally, antibody neutralization had no influence on either plasma progesterone or cholesterol, mitigating against a role for oPL in progesterone production during late pregnancy in the ewe.

J. Endocr. (1985) 106, 377–386

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R De Matteo, D J Hodgson, T Bianco-Miotto, V Nguyen, J A Owens, R Harding, B J Allison, G Polglase, M J Black, and K L Gatford

Preterm birth is associated with increased risk of type 2 diabetes (T2D) in adulthood; however, the underlying mechanisms are poorly understood. We therefore investigated the effect of preterm birth at ~0.9 of term after antenatal maternal betamethasone on insulin sensitivity, secretion and key determinants in adulthood, in a clinically relevant animal model. Glucose tolerance and insulin secretion (intravenous glucose tolerance test) and whole-body insulin sensitivity (hyperinsulinaemic euglycaemic clamp) were measured and tissue collected in young adult sheep (14 months old) after epostane-induced preterm (9M, 7F) or term delivery (11M, 6F). Glucose tolerance and disposition, insulin secretion, β-cell mass and insulin sensitivity did not differ between term and preterm sheep. Hepatic PRKAG2 expression was greater in preterm than in term males (P = 0.028), but did not differ between preterm and term females. In skeletal muscle, SLC2A4 (P = 0.019), PRKAA2 (P = 0.021) and PRKAG2 (P = 0.049) expression was greater in preterm than in term overall and in males, while INSR (P = 0.047) and AKT2 (P = 0.043) expression was greater in preterm than in term males only. Hepatic PRKAG2 expression correlated positively with whole-body insulin sensitivity in males only. Thus, preterm birth at 0.9 of term after betamethasone does not impair insulin sensitivity or secretion in adult sheep, and has sex-specific effects on gene expression of the insulin signalling pathway. Hence, the increased risk of T2D in preterm humans may be due to factors that initiate preterm delivery or in early neonatal exposures, rather than preterm birth per se.