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Stewart (1971) showed that mice of the Peru strain produce [8-lysine]-vasopressin rather than [8-arginine]-vasopressin which is found in other strains of mice and is the usual form of the neurohypophysial hormone in mammals other than the Suina (Ferguson & Heller, 1965). Most mammals are more sensitive to [8-arginine]-vasopressin than [8-lysine]-vasopressin as an antidiuretic hormone (Sawyer, Chan & van Dyke, 1962); but in pigs [8-lysine]-vasopressin is nearly as potent as [8-arginine]-vasopressin (Ferguson & Heller, 1965), indicating some co-adaptation of the renal receptors and the chemical structure of the hormone. It was therefore decided to assess the relative sensitivity of Peru and CBA/FaCam mice to [8-arginine]-vasopressin and [8-lysine]-vasopressin.
Although stomach-loading of conscious Peru mice will induce a rapid water diuresis (Stewart, 1968), this is inhibited by anaesthesia and operational stress. Two bioassay methods using conscious animals have therefore been developed and used on CBA/FaCam and Peru adult male mice. In one method,
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SUMMARY
The possibility of genetic variation in the amino acid composition of neurohypophysial hormones of the mouse was investigated. Acetic acid extracts from acetone-dried neurohypophyses from six strains of mice were subjected to thin-layer chromatography. Extracts from two strains were tested for natriferic potency on a toad bladder preparation. Extracts from three strains of mice were purified on a carboxymethylcellulose column, and the amino acid composition determined by paper electrophoresis at pH 2.
All six strains of mice were found to elaborate an oxytocic principle with the chromatographic properties of oxytocin. All results were compatible with the view that five of the strains elaborate the usual mammalian vasopressin, [8-arginine]-vasopressin. However, chromatography, natriferic assays and analyses of amino acid composition indicated that the Peru strain of mice elaborated [8-lysine]-vasopressin. These mice would be the first mammals outside the Suina which possess this hormone, and the difference between the strains of mice is almost certainly genetic in origin. The significance of genetic variation within a species in the structure of a hormone is discussed in terms of its physiological and evolutionary consequences.
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SUMMARY
Adenyl cyclase was measured in vitro in renal medullary homogenates from male mice of the Peru and CBA/FaCam strains. The basal activity of adenyl cyclase was increased on incubation with 30 mm-NaF and with varying concentrations of [8-arginine]-vasopressin (AVP) and [8-lysine]-vasopressin (LVP) up to 100 mu./ml. In both strains of mice, the maximal hormone activation was the same whichever vasopressin was used, and the same degree of stimulation was observed on incubation of homogenates with both hormones together. It is concluded that both hormones have the same intrinsic activity in this system, and are acting on the same population of receptors within each strain of mouse.
Half-maximal adenyl cyclase activation was achieved with 240 ± 50 (s.e.m.) μu. AVP/ml and 920 ± 160 μu. LVP/ml in homogenates from CBA/FaCam mice; and 240 ± 40 μu. AVP/ml and 1900 ± 250 μu. LVP/ml in Peru mice. These results are compared with previously reported potencies in these mice of the two vasopressins as antidiuretic agents in vivo. Whilst there is good agreement between results in vitro and in vivo with CBA/FaCam mice, there is a highly significant discrepancy with Peru mice. Explanations for these results, including the possibility that cyclic AMP may not be the sole mediator of the increased water permeability of the distal tubule induced by vasopressin, are discussed.
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Desert-living kangaroo rats (Dipodomys merriami) have large neurohypophysial stores of vasopressin compared with rodents from non-arid environments (Ames & van Dyke, 1950). This presumably contributes to the animals' ability to produce very concentrated urine for extended periods (Schmidt-Nielsen, 1964). We have investigated the pituitary store of vasopressin in another rodent adapted to arid environments, the Mongolian gerbil (Meriones unguiculatus), both when allowed free access to water, and after 7 days of water deprivation. Young adult gerbils (ten males and ten females) were obtained from a commercial supplier and kept in our laboratory with free access to food and water for 2 weeks before experimentation began.
The animals were divided into two equal groups both kept at 30 °C and 30 ± 4% relative humidity. One group (control) was allowed free access to food (Edinburgh University rat cake) and water; the other group (dehydrated) was allowed food only. Vasopressin was assayed
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SUMMARY
The effects of the β-adrenergic receptor blocking agent, dl-propranolol, and of the antithyroid drug, carbimazole, upon some manifestations of thyroxine (T4)-induced changes in peripheral metabolism were studied in rats. Propranolol lowered the heart rate, but did not alter the following changes induced by T4: increment in heart rate, increase in heart or kidney weight, increase in urinary hydroxyproline, decrease in body weight gain or increase in serum T4. Carbimazole administration lowered serum T4 and reduced weight gain, but had no effect upon heart rate or hydroxyproline excretion.
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Abstract
The entire coding region of an ovine endometrial oxytocin receptor (OTR) cDNA was generated by PCR, subcloned into the SV40 major late promoter expression vector pSVLJ and transiently expressed in Cos-7 cells. A specific OTR antagonist, 125I-labelled d(CH2)5 [Tyr(Me)2,Thr4,Tyr-NH2 9]-vasotocin (OTA), was used to describe the binding kinetics of the expressed receptor which had a K d of 4·5 nm and Bmax of 2·4 nm/mg protein (6·8 × 105 receptor molecules/transfected cell). The functional properties of the expressed OTR were determined by measuring oxytocin-induced phosphoinositide (PI) hydrolysis. Oxytocin increased PI turnover in OTR transfected cells fourfold in excess of residual endogenous activity, and stimulated phospholipase C (PLC) activity in a dose- and time-dependent manner, confirming that the expressed OTR cDNA was functional. Arginine vasopressin also stimulated PI turnover in a dose-dependent manner; thresholds of responses to oxytocin and arginine vasopressin were 10−9 m and 10−7 m respectively. OTA did not increase PI turnover and competitively inhibited the oxytocin-induced response. Direct activation of the pathway by aluminium fluoride and guanosine (3′-Othio)-triphosphate (GTPγS) confirmed that the OTR was G-protein linked. Co-incubation of GTPγS with oxytocin shifted the PI-response threshold from 10−7 m to 10−9 m and significantly increased the level of response, suggesting that maximum PI turnover was agonist-dependent. The G-protein involved in mediating the signal transduction pathway was pertussis toxin-insensitive and, therefore, probably a member of the Gq subfamily. The PLC inhibitor, U73122, had no effect on oxytocin-induced PI turnover, consistent with the response in endometrial tissue. These data suggest that the signalling pathway mediated by expressed OTR is similar to that attributed to OTR occupancy in ovine endometrium.
Journal of Endocrinology (1996) 149, 389–396
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ABSTRACT
Analysis of ovine conceptus RNA by slot blotting, Northern analysis and nested polymerase chain reaction failed to detect oxytocin–neurophysin prohormone mRNA. Probes used hybridized with both the 3' end of the prohormone mRNA and the oxytocin-coding sequence. Northern analysis of bovine and porcine conceptus RNA was also negative, and polymerase chain reaction demonstrated oxytocin–neurophysin mRNA in ovine corpus luteum, but not in human corpus luteum or decidua, or in ovine endometrium. Infusion of oxytocin into the uterine lumen in cyclic ewes between days 9 and 19 or 20 after oestrus failed to prolong the luteal phase of the cycle and had no effect on endometrial oxytocin receptor concentrations or uterine prostaglandin F secretion. Oxytocin administered systematically prevented luteolysis and reduced uterine prostaglandin F secretion. Taken together, these data suggest that blastocyst-derived oxytocin is unlikely to contribute to corpus luteum maintenance in early pregnancy. They are inconsistent with a previous report that the ovine blastocyst synthesizes and secretes oxytocin.
Journal of Endocrinology (1991) 130, 443–449
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Glucocorticoids play a fundamental role in the endocrinology of pregnancy but excess glucocorticoids in utero may lead to abnormalities of fetal growth. Protection against fetal exposure to cortisol is provided by the enzyme 11β-hydroxysteroid dehydrogenase 2 (11β-HSD2) located in the human placental trophoblast. By contrast, relatively little is known concerning the function of glucocorticoid-activating 11β-HSD1, which is strongly expressed within human maternal decidua. To address this we have assessed: i) changes in decidual 11β-HSD1 expression across gestation and ii) the functional role of glucocorticoids in decidua. Human decidua was collected from women undergoing surgical termination of pregnancy in first (n = 32) and second (n = 10) trimesters, and elective caesarean sections in the third trimester (n = 9). Analysis of mRNA for 11β-HSD1 by real-time RT-PCR showed increased expression in second (9.3-fold, P < 0.01) and third (210-fold, P < 0.001) trimesters. Studies using primary cultures of decidual cells also revealed higher levels of cortisol generation in the third trimester. Changes in decidual 11β-HSD1 with gestation were paralleled by increased expression of the apoptosis markers caspase-3 and annexin-V, particularly in cluster designation (CD)10−VE non-stromal cells (20-fold in third trimester relative to first trimester). Apoptosis was also readily induced in primary cultures of third trimester decidual cells when treated with cortisol, cortisone, or dexamethasone (all 100 nM for 24 h). The effect of cortisone but not cortisol or dexamethasone was blocked by an 11β-HSD inhibitor confirming the functional significance of endogenous cortisol generation. These data show that autocrine metabolism of glucocorticoids is an important facet of the feto-placental unit in late gestation and we propose that a possible effect of this is to stimulate programmed cell death in human decidua.
