Tissue-specific variation in expression of prolactin receptor gene subtypes in hypergravity-exposed rats.

Prolactin (PRL) effects are mediated by membrane receptors (PRLR) of which the long (PRLR-L) and short form (PRLR-S) predominate. Our objective was to compare the distribution pattern of PRLR-L and PRLR-S transcripts and their ratio in adipose (AD), liver (LV), mammary (MG) and pituitary (PG) tissues of stationary (SC, n = 8) and hypergravity (HG, n = 8) exposed periparturient rats. Pregnant rats were exposed to 2 g force from day 11 of gestation (G11) through post partum day 1 (P1). PRLR-L mRNA expression compared to PRLR-S was greater (P < 0.001) in AD, MG and PG but was lower (P < 0.001) in LV in both HG and SC animals at P1. The ratio of PRLR-L/PRLR-S mRNA in the AD, LV, MG and PG was not different between HG and SC rats. In summary, these data reveal that the hypergravity-induced downregulation of PRLR is not directly triggered by deranged distribution of PRLR isoforms.

Effects of hypergravity on mammary metabolic function: gravity acts as a continuum.

Mammary metabolic activity in pregnant rats is significantly increased in response to spaceflight. To determine whether changes in mammary metabolism are related to gravity load, we exposed pregnant rats to hypergravity and measured mammary metabolic activity. From days 11-20 of gestation (G), animals were centrifuged (20 rpm; 1.5, 1.75, or 2.0 x gravity) or were maintained at 1 G. On G20, five rats from each group were removed from the centrifuge and euthanized. The remaining dams (n = 5/treatment) were housed at 1 G until parturition. After 2 h of nursing by the pups, the postpartum dams were euthanized (G22). Glucose oxidation to CO2 and incorporation into lipids was measured. Mammary glands from dams euthanized on G20 revealed a strong negative correlation between metabolic rate and increased G load. Approximately 98% of the variation in glucose oxidation and 94% of the variation in glucose incorporation into lipids can be accounted for by differences in G load. Differences in metabolic activity disappeared in the postpartum dams. When we combined previous data from the microgravity with hypergravity environments and plotted the ratio of mammary metabolic rate vs. G load, there was a significant exponential relationship (r2 = 0.99). These data demonstrate a remarkable continuum of response across the microgravity and hypergravity environments and support the concept that gravitational load influences mammary tissue metabolism.

Effects of steroid hormone treatment on mammary development in prepubertal heifers.

The objective of this study was to determine the effects of steroid hormone implantation in heifer calves on the ability of mammary tissue to develop subsequently in organ culture. Twenty-four calves were paired by date of birth and assigned to groups (eight calves/group). At 4, 7, or 10 mo of age, calves were implanted subcutaneously (s.c.) with pellets containing cholesterol or cholesterol, 17 beta-estradiol, and progesterone for 9 or 18 d. The calves were euthanized and uteri and mammary glands were removed and weighed. Slices of mammary parenchymal tissue were incubated for 5 d at 37 degrees C in a 50% O2, 5% CO2 humidified atmosphere in Waymouth's 752/liter medium supplemented with insulin (5.0 micrograms/ml) or lactogenic hormone complex insulin (5.0 micrograms/ml), aldosterone (0.1 microgram/ml), hydrocortisone (0.1 microgram/ml), and prolactin (1.0 microgram/ml) in the presence or absence of epidermal growth factor (EGF) (0.06 microgram/ml) to promote lobulo-alveolar development. Tissue sections were stained and mounted on slides for morphologic and histologic analysis or prepared to evaluate expression of beta-casein mRNA. There were no morphologic differences in slices from calf mammary tissues despite age, steroid hormone priming, or hormones used in tissue culture. The 4-mo-old calves required steroid priming followed by incubation of the tissue slices with the lactogenic complex with or without epidermal growth factor to induce cytological changes associated with lactogenesis but did not express beta-casein mRNA. At 7 mo of age, steroid hormone priming was not necessary for induction of alveolar formation and secretion. Incubation of the tissue slices from 7-mo-old calves with the lactogenic complex was sufficient to induce alveolar formation and secretion. However, beta-casein mRNA was not expressed. At 10 mo of age, exposure of tissue from calves to the lactogenic hormones caused histologic changes reminiscent of the ability to secrete milk regardless of hormone priming. However, estrogen and progesterone priming was necessary before incubation of the tissue slices with the lactogenic hormones to induce beta-casein mRNA expression. When epidermal growth factor was added to the lactogenic hormone complex, beta-casein mRNA expression decreased. These data support the concept that there is a sequential development of responsiveness of the mammary gland to various hormones. By 10 mo of age, prepubertal heifers reach a stage of maturity where steroid hormone priming followed by incubation of tissue slices with the lactogenic hormones is sufficient to induce both structural and functional differentiation.

Negative effects of a high level of nutrient intake on mammary gland development of prepubertal goats.

Ten French Alpine kids were used to determine whether mammary development was altered by level of nutrient intake during the prepubertal period. At 6 wk of age, kids were paired on the basis of BW and assigned to an ad libitum or restricted diet of milk from Jersey cows. Kids on the restricted diet were fed 70% of their pair mate's milk intake for 4 wk and then 50% for 9 wk. Low quality grass hay was available for all kids. The ADG was greater for kids fed for ad libitum intake. Kids fed for ad libitum intake had larger mammary glands and increased adipose deposition. Measurement of hydroxyproline indicated that connective tissue increased with gland size. Kids on the restricted diet had more DNA and protein per gram of mammary gland, indicating greater secretory development. Goats can be used as a model to study the effect of level of nutrient intake on hormones and growth factors that regulate mammary gland development.

Characterization of binding of transforming growth factor-beta 1 to bovine mammary membranes.

This study developed a procedure to measure binding of transforming growth factor-beta 1 to bovine mammary membranes. Mammary membranes were incubated with 3 M MgCl2 to remove endogenously bound transforming growth factor-beta 1. Binding was optimized by incubation of 200 micrograms of membrane protein with 125I-labeled transforming growth factor-beta 1 in 25 mM Tris, 10 mM CaCl2, and 1.0% BSA in a total volume of .5 ml in the presence or absence of unlabeled transforming growth factor-beta 1. The reaction equilibrated in 2 h at 4 degrees C. Specific binding was linear from 142 to 1140 micrograms of membrane protein. The reaction was specific for the beta transforming growth factors; transforming growth factor-beta 1, transforming growth factor-beta 2, and transforming growth factor-beta 3 could complete effectively with 125I-labeled transforming growth factor-beta 1 for the receptor. The growth factors, epidermal growth factor, IGF-I, or transforming growth factor-alpha did not compete effectively with 125I-labeled transforming growth factor-beta 1 for binding to bovine mammary membranes. Scatchard analysis showed that the number of receptors averaged 251 pmol/mg of membrane protein and the affinity was 8.7 x 10(-11) M. Binding to mammary membranes was higher during the prepubertal and pubertal periods than during lactation. Binding to mammary membranes during early lactation averaged 24% of the binding observed during other physiological states.(ABSTRACT TRUNCATED AT 250 WORDS)