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Search for other papers by D. J. Skene in
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ABSTRACT
A sensitive, specific, reproducible and practical radioimmunoassay for the determination of 5-methoxytryptophol (ML) in pineal glands of different species has been developed. High-affinity specific antisera were produced by immunization of sheep with ML–bovine serum albumin. Iodinated ML, used as the radiolabel, was synthesized by direct iodination of ML using 1,3,4,6,-tetrachloro-3,6-diphenylglycouril as the oxidant. Sensitivity of the assay was 0·005 pmol/tube. The validity of the assay was checked using classical techniques. Cross-reactivity with other indoles was negligible. Parallel inhibition curves were obtained for rat, hamster, sheep and tortoise pineal homogenates. Using thin-layer chromatography, tortoise pineal immunoreactivity also co-chromatographed with standard ML. Samples (n = 5) with ML concentrations of 0·013, 0·052 and 0·209 pmol/tube had intra-assay coefficients of variation of 9·8, 5·7 and 7·6% respectively. Their respective interassay coefficients of variation were 17·7 16·5 and 11·4% (n = 8). The pineal concentration of ML was found to be species dependent. Afternoon ML levels were 0·052± 0·002 (s.e.m.) pmol/gland in the rat (n = 16), 0·539 ±0·089 pmol/gland in the hamster (n = 16), 1·73±0·225 nmol/g in the sheep (n = 10) and 7·15± 0·465 pmol/gland in the tortoise (n = 4). The ratio of ML:melatonin content in the pineal gland also showed a large interspecies variation with values of 0·02 in the rat, 0·22 in the sheep, 2·7 in the hamster and 17 in the tortoise.
J. Endocr. (1986) 110, 177–184
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SUMMARY
Using a protein-binding technique, progesterone concentrations in peripheral plasma (jugular vein) were measured throughout the oestrous cycle of 24 ewes. Examination of the specificity of the method by thin-layer chromatography indicated that interference from other steroids was not significant in sheep plasma. During the first 4 days of the cycle (days 0–3), plasma progesterone concentrations were below 0·4 ng./ml., increasing to a mean level of 1·5–2·5 ng./ml. between days 4 and 9, and remaining at this level for approximately 5 days, before declining rapidly on days 14 and 15 to reach a low level on the day before oestrus. The progesterone concentration on the day of oestrus was extremely low (0·1 ng./ml.), and was of the same order as that found in the plasma of wethers and anoestrous or ovariectomized ewes. Three ewes, superovulated with pregnant mare serum gonadotrophin, showed marked elevation of peripheral progesterone concentration during the luteal phase of the cycle, the concentration being proportional to the number of corpora lutea formed.
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ABSTRACT
The possible involvement of a deficit of GH and insulin-like growth factor-I (somatomedin C) (IGF-I/SMC) in mediating the effects of propylthiouracil (PTU)-induced hypothyroidism on body and skeletal growth and myelination was studied in the neonatal rat. Myelination (as assessed by 2′,3′-cyclic nucleotide 3′-phosphohydrolase (CNP) activity), skeletal growth (as assessed by tail length) and body weight of pups from PTU-treated mothers were significantly retarded compared with normal animals or euthyroid controls. At 20 days after birth, plasma GH in hypothyroid animals was undetectable (< 10 μg/l), pituitary GH content was 1 ·2% of control, and plasma, liver and kidney IGF-I/SMC concentrations were 63, 68 and 50% of control values respectively. CNP activity in hypothyroid brain was 52% of normal controls but the concentration of IGF-I/SMC was 113–154% of control. Treatment of hypothyroid animals from day 1 with GH (10 mg/kg body weight per day) restored liver and plasma IGF-I/SMC concentrations at 20 days to values above those of normal animals and euthyroid controls. The concentration of IGF-I/SMC was also significantly (P< 0·001) restored in hypothyroid kidney (79% of normal), but the concentration in brain was unaffected. These observations provide evidence that the GH treatment employed in the present experiments was adequate to restore the deficit. GH treatment had no significant effect on tail length or CNP activity, and only a small (4–24%) effect on body weight at 20 days. Only thyroxine was able fully to restore body weight and substantially restore tail length and CNP activity.
The present study provides strong evidence against an important involvement of GH or IGF-I/SMC in mediating the effects of thyroid hormone on myelination and body growth in the infant rat. It does not, however, rule out the possibility that thyroid hormone is required for the expression of the growth-promoting effects of IGF-I/SMC by other mechanisms such as the expression of the IGF-I/SMC receptor.
J. Endocr. (1988) 119, 117–125
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Search for other papers by TANA J. OXBORROW in
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Search for other papers by I. D. SMITH in
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Search for other papers by G. D. THORBURN in
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SUMMARY
The progesterone concentration in the peripheral plasma of ewes throughout pregnancy has been determined by a protein-binding method.
Plasma progesterone concentrations during the first 50 days of pregnancy (2–3 ng./ml.) were not significantly higher than peak concentrations during the luteal phase in cycling non-pregnant ewes, but there was no decrease in the concentration 15–20 days after mating as occurs in non-pregnant ewes.
Between 50 and 120 days after mating the plasma progesterone concentration increased steadily to values 2–5 times that found in early pregnancy. These high concentrations were maintained until lambing. A decrease in progesterone concentration during the week preceding lambing was usually, but not always, observed.
Mean plasma progesterone concentrations during the last 50 days of pregnancy in ewes with twins were approximately twice those in ewes with a single foetus.
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Progesterone levels in the peripheral blood of non-pregnant mares have been measured by Short (1959); progesterone was detectable only when there was a fully developed corpus luteum present in the ovaries. On the other hand, Short (1964) has shown that progesterone secretion into the ovarian vein, negligible at oestrus, increases very significantly within the first 24–36 hr. after ovulation. More sensitive methods for the measurement of progesterone concentration in peripheral plasma have since been developed. Peripheral plasma progesterone concentrations have been measured throughout the menstrual cycle of women (Neill, Johansson, Datta & Knobil, 1967a) and monkeys (Neill, Johansson & Knobil, 1967b) and throughout the oestrous cycle of the ewe (Thorburn, Bassett & Smith, 1969) by competitive protein-binding techniques and in cows, pigs and sheep by gas—liquid chromatography (Stabenfeldt, Ewing, Patton & McDonald, 1969). The present study using a protein-binding assay method was undertaken in an attempt to demonstrate
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Abstract
The present study was designed to investigate whether transport, a mild environmental stressor, could affect the oestradiol-induced LH surge in postpartum ewes and, if so, the mechanism involved. Welsh Mountain ewes, with lambs removed at parturition (day 0) and hand-milked 12 and 48 h later, were given 50 μg oestradiol benzoate intramuscularly at various times postpartum. Blood samples were taken via an indwelling jugular venous catheter every 2 h from 8 to 24 h after oestradiol injection. All results are given as means ± s.d. On day 1 oestradiol was unable to induce an LH surge in any ewe. Transport (10–14 h after oestradiol) delayed the onset of the oestradiol-induced LH surge on day 14 (17·5 ±1·7 vs 14·4±2·0 h, n=5 each; P<0·05), but not on day 28 (14·9 ±2·0 vs 14·0 ±2·4 h, n=5 out of 7). Transport had no effect on the amplitude of the surge on either day. Naloxone treatment (1 mg/kg per 2 h) was unable to prevent the delay caused by transport (18·0±1·1 vs 17·5 ± 1·7 h, n=8 each), and did not affect the amplitude of the surge (28·4±5·3 vs 28·1 ±2·3 ng/ml, n=8 each). The duration of the LH surges were not assessed. On day 7, transport from 16 to 20 h after oestradiol delayed the LH surge (22·8 ±2·0 vs 18·0 ± 2·8 h, n=8 each; P<0·05) and reduced the surge amplitude (19·7 ±1·7 vs 22·8 ±2·8 ng/ml; P<0·05), whilst transport from 10 to 14 h did not. Transport (16 to 20 h) had no effect on surge duration (6·25 ±0·7 vs 6·75 ± 1·0 h). In conclusion, transport inhibited the oestradiol-induced LH release in the early postpartum ewe by a non-opioidergic mechanism, but only if the stressor occurred within 2–3 h of the expected onset of the surge.
Journal of Endocrinology (1994) 142, 447–451
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ABSTRACT
Ovaries from pregnant and postpartum Sprague–Dawley rats were examined for content of immunoreactive β-endorphin by radioimmunoassay, and for its localization by the peroxidase-antiperoxidase technique. In addition, the molecular forms of β-endorphin immunoreactivity were separated by gel chromatography and reverse-phase high performance liquid chromatography (RP-HPLC). Ovaries from rats early in pregnancy showed intense granular cytoplasmic staining of luteal cells, with an even distribution of granular material throughout the cytoplasm. By middle to late pregnancy the staining pattern was changed, with immunoreactive material showing a less granular and unevenly distributed staining pattern and with some areas of the cytoplasm totally devoid of immunoreactive material. The concentrations of immunoreactive β-endorphin measured during pregnancy were significantly lower than levels in mature non-pregnant rat ovary. The ovarian concentration of immunoreactive β-endorphin fell progressively during pregnancy and early lactation, returning to normal cyclic rat levels at 20 days post partum. The ovarian concentration of β-endorphin-like material was lowest at 6 days post partum (0·53 ± 0·08 ng/g wet weight; mean ± s.e.m.), representing approximately 10% of the concentration found in pooled ovaries from randomly cyclic adult rats. Gel chromatography revealed only a single peak of immunoreactive β-endorphin, co-eluting with 3·5 kD molecular weight ovine β-endorphin(1–31). This contrasts with gel profiles of adult cyclic rat ovary, where large molecular weight species pro-opiomelanocortin (31 kD) and β-lipotrophin (11·5 kD) are also present. On RP-HPLC the predominant species of low molecular weight immunoreactive material co-eluted with β-endorphin(1–31).
These data show that pregnancy in the rat is associated with marked changes in the levels, cellular localization and chromatographic profiles of ovarian β-endorphin. The aetiology and physiological significance of these changes remains to be established.
J. Endocr. (1986) 108, 343–350
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SUMMARY
A biochemical comparison of the lactogenic effect of ovine prolactin and of bovine growth hormone on pregnant mouse mammary gland in organ culture was made. No qualitative differences were observed; both hormones (in the presence of insulin and corticosterone) stimulated the synthesis of casein and RNA in mouse mammary gland explants. The inhibition of RNA synthesis with actinomycin D was associated with a decrease in casein synthesis. However, quantitatively, prolactin was more efficient than growth hormone in stimulating casein synthesis.
The synthesis of casein in mouse mammary gland explants incubated in the presence of various combinations of hormones gave results which suggested that prolactin and growth hormone are operating on the same sites of action.
Analysis of, and purification by, polyacrylamide gel electrophoresis of the bovine growth hormone showed that the lactogenic effects were not due to the presence of prolactin as an impurity, but were an intrinsic property of the growth hormone.
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ABSTRACT
The bioactive (B) and immunoreactive (I) pituitary contents/concentrations of FSH, together with the plasma concentrations of B-FSH, I-FSH and I-inhibin were determined in ovine fetuses at days 55, 75, 90 and 135 of gestation (day 145 = term). The pituitary contents and concentrations of B-FSH and I-FSH increased in both sexes with gestational age. The female fetuses had significantly (P <0·01) higher pituitary contents/concentrations of B-FSH and I-FSH than the male fetuses at days 75 and 135. The pituitary B/I ratios of FSH were not significantly different with age or sex. The plasma concentrations of B-FSH remained relatively constant from days 75 to 135, with no significant differences between sexes or with age. In contrast, the plasma concentrations of I-FSH reached a peak at day 90 and then declined towards term in both sexes. At all gestational ages except day 55, the female fetuses had significantly (P <0·05) higher plasma concentrations of I-FSH than the males. In both sexes, the plasma B/I ratios of FSH were lowest at day 90 and had increased again by day 135, with the male fetuses having significantly (P <0·05) higher B/I ratios compared with the female group at days 75 and 135 but not at day 90. At all gestational ages, the plasma concentrations of I-inhibin declined throughout gestation in the female fetuses, whereas in the males they reached a nadir at day 75 and then increased towards term. The concentrations of I-inhibin were significantly (P <0·01) higher in the male fetuses compared with the females. Collectively, these data suggest that there are changes in the forms of FSH present in the pituitary gland and plasma throughout gestation in the ovine fetus. Moreover, they infer that the difference between the sexes in FSH synthesis and/or secretion may be attributed in part to the circulating concentrations of inhibin.
Journal of Endocrinology (1992) 134, 287–295
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The presence of calcium in the fluid in which intestinal or kidney tissue is incubated is required for the tissue to respond to angiotensin. Everted sacs of rat colonic mucosa exhibited an increased rate of fluid transport in the presence of angiotensin; this response was lost when the serosal fluid, but not the mucosal fluid, was calcium-free. Angiotensin-stimulated transport was maintained when calcium was replaced with strontium or barium, but was lost when calcium was exchanged for magnesium. Similarly, calcium ions were required in the incubation fluid of rat kidney cortex slices to demonstrate angiotensin-enhanced sodium transport. These observations are discussed in relation to the possible roles of divalent cations in the mechanism of action of angiotensin.
