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Sergio A Arispe, Betty Adams and Thomas E Adams

Plant-derived estrogens (phytoestrogens, PEs), like endogenous estrogens, affect a diverse array of tissues, including the bone, uterus, mammary gland, and components of the neural and cardiovascular systems. We hypothesized that PEs act directly at pituitary loci to attenuate basal FSH secretion and increase gonadotrope sensitivity to GnRH. To examine the effect of PEs on basal secretion and total production of FSH, ovine pituitary cells were incubated with PEs for 48 h. Conditioned media and cell extract were collected and assayed for FSH. Estradiol (E2) and some PEs significantly decreased basal secretion of FSH. The most potent PEs in this regard were coumestrol (CM), zearalenone (ZR), and genistein (GN). The specificity of PE-induced suppression of basal FSH was indicated by the absence of suppression in cells coincubated with PEs and an estrogen receptor (ER) blocker (ICI 182 780; ICI). Secretion of LH during stimulation by a GnRH agonist (GnRH-A) was used as a measure of gonadotrope responsiveness. Incubation of cells for 12 h with E2, CM, ZR, GN, or daidzein (DZ) enhanced the magnitude and sensitivity of LH secretion during subsequent exposure to graded levels of a GnRH-A. The E2- and PE-dependent augmentation of gonadotrope responsiveness was nearly fully blocked during coincubation with ICI. Collectively, these data demonstrate that selected PEs (CM, ZR, and GN), like E2, decrease basal secretion of FSH, reduce total FSH production, and enhance GnRH-A-induced LH secretion in a manner that is dependent on the ER.

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C. SERNIA and C. H. TYNDALE-BISCOE

SUMMARY

Specific binding of radio-iodinated ovine prolactin to subcellular tissue fractions of the tammar wallaby (Macropus eugenii) was investigated. Specific binding was found, in order of decreasing binding activity, in the lactating mammary gland, corpus luteum, corpus albicans, adrenal gland and ovary. Specific binding was absent in kidney, liver, brain and inactive mammary gland.

The mean association constant (Ka at 23 °C) was determined as 0·90 × 109, 2·20 × 109, 2·44 × 109, 3·38 × 109 and 10·98 × 1091/mol for mammary gland, adrenal, corpus albicans, corpus luteum and ovary respectively. The mean receptor concentration (N) varied from 92·87 × 10−14 mol/mg protein for the mammary gland to 1·03 × 10−14 mol/mg protein for the ovary. The concentration in the corpus luteum varied between tissue pools collected at different times of the annual breeding cycle.

The specificity for prolactin was shown in the mammary gland and corpus luteum by the failure of ovine FSH, LH, GH and TSH to displace 125I-labelled ovine prolactin, whereas it was displaced readily by both ovine and bovine prolactin.

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A. T. COWIE, J. S. TINDAL and A. YOKOYAMA

SUMMARY

Mammogenesis has been studied in hypophysectomized goats (nine completely and four incompletely hypophysectomized). In the absence of the pituitary, hexoestrol alone or in combination with progesterone failed to stimulate the growth of the mammary gland or to prevent regression of the already developed mammary gland. In the ovariectomized-hypophysectomized goat considerable mammary lobulo-alveolar growth can be promoted by administering hexoestrol and progesterone in combination with prolactin (ovine), growth hormone (bovine) and adrenocorticotrophin.

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P. PRUNET, L. M. HOUDEBINE, C. DELOUIS and B. BRETON

The lactogenic properties of extracts of the pituitary glands of salmon and trout were evaluated by using the organ culture technique with rabbit mammary explants. Crude extracts and fractions obtained after chromatography on Ultrogel and selected for their capacity to compete with ovine prolactin in a rabbit mammary gland radioreceptor assay were added to the culture medium. The criteria of lactogenesis were lactose synthetase activity, casein synthesis, measurements of the concentration of casein messenger RNA and the histology of mammary glands. All these tests led to the conclusion that salmon and trout pituitary glands contain a prolactin-like principle capable of initiating milk synthesis in the rabbit mammary cell.

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YOSHIHIKO NAKANISHI, JUNICHI MORI and HIROSHI NAGASAWA

SUMMARY

The effects on reproductive function of restriction to one half of the normal food intake for 30 or 60 days and subsequent unrestricted feeding were investigated in adult female Sprague–Dawley rats.

Restricted feeding resulted in the cessation of oestrous cycles within 9 to 28 days, associated with decreased pituitary, ovarian and uterine weights. Pituitary content and concentration of FSH were increased by restricted feeding and the levels of FSH in 60 day underfed rats were about twice as high as those of the controls at dioestrus. There was little difference in pituitary LH content between the underfed groups and the controls at the end of restricted feeding. Pituitary LH concentration was significantly higher in rats underfed for 60 days than in the control rats. Pituitary prolactin content was one half and one third of that in control rats in rats underfed for 30 and 60 days respectively. Pituitary prolactin concentration was also decreased by restricted feeding. At the end of restricted feeding, no differences in serum LH and prolactin levels were found between the groups, whereas the serum FSH level in 60 day underfed rats was higher than that in the controls at dioestrus.

After re-feeding, normal oestrous cycles returned within 3–5 days in almost all rats, regardless of the length of the previous cessation of oestrous cycles. Pituitary contents of FSH and LH in underfed rats decreased after re-feeding following the return of oestrous cycles. The rate of decrease was much greater in 60 day underfed rats than that in 30 day underfed rats. On the other hand, serum levels of FSH, LH and prolactin in these underfed rats were increased by re-feeding and the levels in the evening of the first pro-oestrus were higher than those in the morning of dioestrus and pro-oestrus. Serum levels of these hormones increased more in 60 day underfed rats than in the other groups at any stage. After re-feeding, pituitary, ovarian and uterine weights increased and the uterine epithelial layer was clearly repaired in both underfed groups, although not always to the control levels by the first oestrus. The end-bud system of the mammary gland which degenerated during restricted feeding was comparable to that of the controls at the first oestrus after re-feeding in 30 day underfed rats, but not in 60 day underfed rats.

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A. T. COWIE and W. R. LYONS

SUMMARY

In the hypophysectomized-ovariectomized-adrenalectomized hooded Norway rat, growth of the mammary ducts can be induced with oestrone + somatotrophin + corticoids; lobule-alveolar development ensues if in addition to the above combination progesterone + prolactin is also administered. On withdrawing these hormones the local injection of prolactin then causes secretion in the underlying mammary gland, but only when corticoids are administered systemically.

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D. S. FLUX and R. E. MUNFORD

SUMMARY

Intact mature virgin female mice of the CHI strain treated with ACTH showed no change in total mammary gland area or in the degree of branching of the mammary duct systems. Their ovaries were larger than those of control mice. Similar mice treated with ACTH in a further experiment showed more large corpora lutea in the ovaries than did control mice.

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T. Harigaya, S. Sakai, K. Kohmoto and Y. Shoda

Regulation of mammary prolactin receptors by steroid hormones was investigated in ovariectomized mid-pregnant mice. Ovariectomy increased the number of mammary prolactin receptors per cell with no effect or a slight decrease in dissociation constant (K d). The simultaneous removal of adrenals prevented this increase in numbers. A single injection of glucocorticoid (corticosterone or cortisol) in ovariectomized–adrenalectomized mice restored the number of prolactin receptors in mammary glands to the same level as that in ovariectomized controls without changing the K d. Aldosterone, deoxycorticosterone and oestradiol did not affect the number of mammary prolactin receptors after ovariectomy–adrenalectomy. Serum concentration of prolactin was not influenced by the hormone manipulation except with injections of oestradiol or cortisol and apparently did not correlate with the number of prolactin receptors. These results indicated that glucocorticoids are required for the increase in the number of mammary prolactin receptors induced by ovariectomy in mid-pregnant mice.

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K. KOHMOTO and H. A. BERN

SUMMARY

Mammotrophic (i.e. mammogenic and/or lactogenic) activity of mouse placentae of different stages (4–19 days of pregnancy) was examined using organ co-culture of placental explants with mammary tissue. The test mammary tissues were taken from midpregnant (11–12 days) nulliparous A/Crgl mice and cultured in a synthetic medium (Waymouth's) supplemented with insulin (5 μg/ml) and aldosterone (1 μg/ml). The responses of mammary gland to placental explants were judged histologically, and were compared with those seen after the addition of ovine prolactin (5 μg/ml). With placentae from 6- to 19-day pregnant animals, distinct mammotrophic activity was seen, with the appearance of eosinophilic secretion in the mammary alveolar lumina, whereas with 4- or 5-day-old 'placentae', no mammotrophic activity was detected. Inasmuch as growth hormone does not substitute for prolactin in mammary gland development and function in the A/Crgl mouse, it can be concluded that a prolactin-like factor is present in the mouse placenta. The influence of placentae on mammary gland was further analysed by transplantation of placental fragments to mammary fat pads. Local lobuloalveolar development was prominent in some instances in the area around the placental transplants.

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VK Pedchenko and W Imagawa

Proliferation and differentiation of mammary epithelia are regulated by the combined action of systemic hormones and locally derived paracrine growth factors. Keratinocyte growth factor (KGF) is a potential candidate stromal factor that may participate in the hormonal control of stromal/epithelial interactions. In this study, we have examined the in vivo effect of 17beta-estradiol (E) treatment on KGF expression in mammary glands of peripubertal (5-week-old) and mature (11-week-old) mice. Mice received subcutaneous injections of hormone after which KGF mRNA levels were assayed by ribonuclease protection analysis of mammary gland RNA. E treatment caused a dose- and time-dependent increase in KGF mRNA levels in intact mice from both age groups. Neither 17alpha-estradiol nor progesterone injection affected KGF mRNA levels. Comparison of the relative expression of KGF in parenchyma-free fat pads and in intact glands demonstrated that the basal and E-dependent KGF mRNA levels did not require the presence of mammary epithelium. ELISA assay of KGF tissue content demonstrated that concomitantly with an up-regulation of mRNA, E treatment also increased KGF protein in mammary glands from peripubertal and mature mice. These data show that E treatment stimulates both KGF mRNA and protein expression in mammary stroma in vivo and raises the possibility that KGF has a role in E-regulated mammary gland development.