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J Liebermann and D Schams

Abstract

In the present investigation, the effect of recombinant (BST) and pituitary-derived (bGH) bovine somatotrophin on progesterone and oxytocin release was examined. Individual copora lutea (CL) were obtained from cows at different stages of the oestrous cycle (days 5–7, 8–12 and 15–18) and also from early pregnancy (days 60–120) and were implanted with an in vitro microdialysis system (MDS).

Perfusion with BST for 60 min (005, 0·5 and 5 μmol/l) induced a dose-dependent stimulation of progesterone release. Release of oxytocin from CL was significantly stimulated by BST at all dose levels. BST (0·5 μmol/l) stimulated progesterone release most during the early and mid-luteal phases and oxytocin release especially during the early luteal stage (days 5–7) of the oestrous cycle. CL from early pregnancy (days 60–120) treated with BST showed a significant response in progesterone and oxytocin release. bGH showed comparable effects.

Our results suggest that somatotrophin acts directly on the secretory function of bovine CL in the MDS, specifically during the early luteal stage (days 5–7) of the oestrous cycle and early pregnancy (days 60–120). Somatotrophin may therefore have physiologically relevant effects associated with the development and maintenance of luteal function.

Journal of Endocrinology (1994) 143, 243–250

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R. Claus and D. Schams

ABSTRACT

Oxytocin concentrations were measured radioimmunologically in sows on the day of standing oestrus over a 6-h period (controls, n=6) or 1 h before and 5 h after mating (n=5) or transcervical infusion of either 100 ml saline (0·9% (w/v) NaCl, n=7) or saline plus 10 μg oestradiol (simulation of seminal oestrogens, n=5).

In the controls, oxytocin was low, at around 1·0 pmol/l, throughout the investigation period. Similarly, saline infusion did not lead to a noticeable change in oxytocin concentrations in six out of seven sows. In one sow, however, infusion led to a maximum of 86 pmol/l at 1 min after infusion. Oestradiol led to no immediate increase in oxytocin concentrations. Later in the post-treatment period (2–5 h) they were only slightly increased (1 pmol/l vs 3 pmol/l). All mated sows reacted with a rapid and clear increase in oxytocin. Maximal concentrations (42·0±5·1 pmol/l; mean ± s.e.m.) appeared 2 min after the onset of ejaculation. Clearly increased concentrations were found for 40 min. It was concluded that mating specifically leads to a rise in oxytocin, probably due to both mechanical and pheromonal stimuli provided by the boar.

Journal of Endocrinology (1990) 126, 361–365

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F. Ellendorff and D. Schams

ABSTRACT

The neuroendocrine reflex theory of milk ejection was investigated in the horse under natural suckling conditions. To this end 12 lactating mares were provided with acute jugular catheters and with intramammary pressure (IMP) recording catheters. The foal had free access to the contralateral mammary complex. Intramammary pressure could thus be recorded while blood was drawn simultaneously for oxytocin analysis from the undisturbed animal. Suckling periods associated with a characteristic increase in IMP lasted significantly longer than unsuccessful nursing attempts. Elements of successful sucklings involved physical stimulation of the mammary gland, a quiet phase and a sudden increase in IMP. Successful suckling took place at about 20-min intervals with a wide range from < 5 min to > 100 min. Between 5 and 10 mU oxytocin i.v. were sufficient to evoke an increase in IMP identical in shape and duration to a naturally induced increase in IMP. Mean peak oxytocin levels reached 15·8 pmol/l plasma, with a maximal release of 39 pmol/l. In the majority of cases (> 80%) peak oxytocin release did not occur until after the increase in IMP; in some cases an oxytocin surge was not detectable at all, despite a milk ejection-associated increase in IMP. In three cases increase in IMP could be observed while the foals were away from the mother with no signs of any intention to suckle. The data indicate that in the horse some elements of the neuroendocrine reflex, such as tactile stimulation of the teat and a surge of oxytocin before an increase in IMP, are facultative and not essential for normal milk ejection.

J. Endocr. (1988) 119, 219–227

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D. Schams, R. Koll and C. H. Li

ABSTRACT

The effect of insulin-like growth factor-I (IGF-I), epidermal growth factor (EGF), fibroblast growth factor (FGF) and nerve growth factor (NGF) on production of oxytocin and progesterone by cultured bovine granulosa and luteal cells was studied. Secretion of oxytocin was stimulated, in a dose-dependent manner, by IGF-I at 48 and 120 h of culture to levels much higher than those after stimulation with LH, FSH, EGF, FGF or NGF. A similar effect of IGF-I was observed for progesterone but, in contrast to oxytocin, secretion of progesterone was not increased by EGF, NGF or FGF. During primary culture, for 4 h, of dispersed bovine luteal cells obtained from corpora lutea between days 4 and 10 of the oestrous cycle, all the growth factors tested failed to stimulate secretion of oxytocin or progesterone. The data suggest the relevance of growth factors (especially IGF-I) for ovarian physiology and their possible importance for differentiation of follicles and luteinization.

J. Endocr. (1988) 116, 97–100

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D. SCHAMS, A. LAHLOU-KASSI and P. GLATZEL

Oxytocin was measured radioimmunologically during the oestrous cycle in six ewes of each of two local Moroccan breeds of sheep. Concentrations in both breeds approached the lower limit of the assay (3 pg/ml) from 2 days before oestrus, throughout heat and ovulation until day 2 of the cycle. Oxytocin concentrations then increased in both breeds, the resulting highest levels on days 5–7 were, on average, between 30 and 60 pg/ml in the D'man sheep and 13–31 pg/ml in the Timhadite breed. Oxytocin levels then decreased to about 7 pg/ml in the D'man and about 4–5 pg/ml in the Timhadite breed on days 14–15. After ovariectomy oxytocin concentrations remained at about the limit of detection for a further 19–20 days in both breeds.

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R. D. HOOLEY, D. SCHAMS and J. K. FINDLAY

Growth hormone was measured by radioimmunoassay in blood samples collected from ewes during the hormonal induction of lactation and during hand-milking post partum. Ovariectomized ewes were induced to lactate with injections of progesterone + oestradiol benzoate every 3 days for 30 days (priming phase) and then daily injections of dexamethasone for 5 days (trigger phase). The ewes were then milked daily. Immunoreactive GH levels fluctuated considerably but were generally in the range 1–15 ng/ml during all phases of lactation and were unaffected by bromocriptine treatment. Milk yield was unrelated to GH levels.

Growth hormone was also measured in blood plasma taken at frequent intervals around the time of milking in lactating ewes approximately 4 weeks post partum. Although a clear prolactin response to milking was observed, there was no indication of a GH response.

Although there is probably a minimum requirement for GH for lactation, a relationship between immunoreactive GH levels and milk yields was not established, perhaps because of limitations of the radioimmunoassay to detect all the biologically active GH.

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S Kobayashi, B Berisha, WM Amselgruber, D Schams and A Miyamoto

The newly formed corpus luteum (CL) rapidly develops after ovulation and has the features of active vascularisation and mitosis of steroidogenic cells. These stage-specific mechanisms also may contribute to gain the function of prostaglandin F2 alpha (PGF2 alpha)-resistant CL at this stage. Recent studies suggest that the vasoactive peptide angiotensin II (Ang II) regulates luteal function. Thus, this study aimed to investigate (i) the expression of angiotensin-converting enzyme (ACE) mRNA by RT-PCR and the ACE protein expression by immunohistochemistry, (ii) the effects of angiogenic growth factors, basic fibroblast growth factor (bFGF) and vascular endothelial growth factor (VEGF), on the secretion of Ang II, PGF2 alpha, progesterone and oxytocin (OT), and (iii) the effects of luteal vasoactive peptides (Ang II and endothelin-1 (ET-1)) or OT on the secretion of PGF2 alpha, progesterone and OT from bovine early CL (days 3--4 of the oestrous cycle), and evaluate a possible interaction of these substances with PGF2 alpha. The expression of mRNA for ACE was found in theca interna of mature follicle, early CL and endothelial cells from developing CL as well as pituitary and kidney, but granulosa cells of mature follicle were negative. The immunohistochemical analysis revealed that blood capillaries (endothelial cells) were stained for ACE, but luteal cells were negative in early CL. To examine the effects of substances on the secretory function of the CL, an in vitro microdialysis system was used as a model. The infusion of bFGF and VEGF stimulated Ang II and PGF2 alpha secretion as well as progesterone, but not OT secretion in early CL. The infusion of Ang II after PGF2 alpha infusion continued the stimulatory effect on progesterone and OT release within early CL until 3 h thereafter. However, the infusion of ET-1 alone had no effect on progesterone or OT release. The infusion of luteal peptides such as Ang II and OT stimulated PGF2 alpha secretion, whereas the infusion of ET-1 did not. In conclusion, the overall results of this study indicate that a functional angiotensin system exists on the endothelial cells of early CL, and that angiogenic factors bFGF and VEGF upregulate luteal Ang II and PGF2 alpha secretion, which fundamentally supports the mechanism of progesterone secretion in bovine early CL. This idea supports the concept that the local regulatory mechanism involved in active angiogenesis ensures the progesterone secretion in the developing CL in vivo.

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B Berisha, D Schams, M Kosmann, W Amselgruber and R Einspanier

Locally produced growth factors may have important modulatory roles in final ovarian follicular growth. The aim of this study was to investigate the possible participation of vascular endothelial growth factor (VEGF) and basic fibroblast growth factor (FGF2) in bovine follicles during final growth. Ovaries were collected from a slaughterhouse within 10-20 min after exsanguination. A classification of follicles into five groups (<0.5; >0.5-5; >5-20; >20-180; >180 ng/ml) was performed according to the follicular fluid (FF) oestradiol-17 beta content. For a better characterisation of classes the mRNA expressions of FSH receptor, LH receptor and aromatase cytochrome P450 in theca interna (TI) and granulosa cells (GC) were determined. Analysis of VEGF transcript by RT-PCR showed that GC and theca cells express predominantly the smallest isoforms (VEGF(121) and VEGF(165)). VEGF mRNA expression in both tissues (TI and GC) and VEGF protein concentration in total follicle tissue increased significantly (and correlated) with developmental stages of follicle growth. The expression of mRNA for VEGF receptor (VEGFR)-1 and VEGFR-2 was very weak in GC, without any regulatory change during final follicle growth. In contrast, TI showed strong expression of mRNA for both receptors in all follicle classes examined. VEGF protein concentrations in FF increased significantly and continuously to maximum levels in preovulatory follicles. As shown by immunohistochemistry, VEGF protein was clearly localised in TI and GC of preovulatory follicles. FGF2 and FGF receptor (FGFR) mRNA expression in TI increased significantly during final growth of follicles. In contrast, the FGF2 and FGFR mRNA expression in GC was very weak and without any regulatory change during follicle growth. Histological observation revealed that FGF2 protein was localised in theca tissue (cytoplasm of endothelial cells and pericytes) but not in GC. Our results suggest that VEGF and FGF families are involved in the proliferation of capillaries that accompanies the selection of the preovulatory follicle resulting in an increased supply of nutrients and precursors, and therefore supporting the growth of the dominant follicle.

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D. Schams, H. Mayer, A. Prokopp and H. Worstorff

ABSTRACT

Oxytocin secretion in dairy cows was measured during milking by means of a specific radioimmunoassay. Basal oxytocin concentrations before milking recorded from 147 milkings from 21 cows, where an assay method with enhanced sensitivity was employed, were 1·5 ± 0·6 pmol/l and there was no evidence for any conditioned release of oxytocin. A 1-min manual stimulation before milking evoked a variable but distinct increase in oxytocin concentrations in 188 out of 195 milkings performed using 29 cows. Despite the high variation in absolute concentrations five types of typical secretion pattern could be distinguished. There was no obvious relationship between patterns or absolute concentrations of oxytocin and milk-flow characteristics. Evidence is given that milk ejection seems to follow the threshold principle in that small releases of oxytocin up to a range of 3–5 pmol/l plasma are sufficient to evoke maximum milk ejection.

J. Endocr. (1984) 102, 337–343

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A. Miyamoto, K. Okuda, F. J. Schweigert and D. Schams

ABSTRACT

The effects were investigated of basic fibroblast growth factor (bFGF), transforming growth factor-β (TGF-β) and nerve growth factor (NGF) on the release of progesterone and oxytocin from the bovine corpus luteum (CL) at different stages of the oestrous cycle. A microdialysis system (MDS) of CL and a cell culture system with a reduced number of endothelial cells were used.

In the MDS of CL from the mid-luteal stage (days 8–12 of the oestrous cycle), infusion with bFGF (0·1, 1, 10 and 100 ng/ml), TGF-β (0·1, 1 and 10 ng/ml) and NGF (0·1, 1, 10 and 100 ng/ml) for 30 min induced significant acute effects on the release of progesterone. Both bFGF and NGF stimulated the release of progesterone during peptide infusion, TGF-β and also bFGF in the period thereafter. This stimulation was dose-dependent during and after the infusion only for bFGF. This response pattern was observed at all luteal stages for the three growth factors, but bFGF was more stimulatory at the early (days 5–7) and mid-luteal stages during and after peptide infusion. The release of oxytocin was stimulated by bFGF in a dose-dependent manner. At the highest dose, bFGF, TGF-β and NGF stimulated the release of oxytocin throughout all three luteal stages. When luteal cells were cultured with growth factors, only TGF-β showed a dose-dependent inhibition of both basal and LH-stimulated progesterone as well as oxytocin release (measured between 48 and 52 h of culture). NGF had an inhibitory effect only on the basal release of oxytocin. bFGF had no effect on the release of either hormone under continuous stimulation in cell culture.

The results indicate that bFGF, TGF-β and NGF act directly and acutely on the secretory function of bovine CL in the MDS but also have long-term effects as shown in cell culture. bFGF appears to be an important autocrine/paracrine regulator of CL function, since local expression of its mRNA, peptide synthesis and its mitogenic and non-mitogenic actions have now been confirmed. Endothelial cells from the CL have been identified as target cells for bFGF. Differences observed between the two systems might thus be attributed to the presence or absence of cellto-cell contact and a reduced number of endothelial cells, as well as to the duration of peptide stimulation and medium changes every 24 h compared with the flow-through conditions in the MDS.

Journal of Endocrinology (1992) 135, 103–114