Prolactin Preparations Research Articles

Radioimmunoassay for bovine prolactin in plasma.

A sensitive radioimmunoassay for bovine prolactin in plasma has been developed. The separation of free and antibody bound 131I labeled bovine prolactin was accomplished by the double antibody method. The method was so sensitive that unlabeled bovine prolactin as little as 0.063 mμg permitted detection of bovine prolactin in plasma as low as 2 mμg/m/ under conditions employed. Plasma from lactating cows and bovine pituitary extracts produce an identical dose response curve obtained by the purified bovine prolactin preparation. Caprine prolactin in plasma and pituitary extracts can be also measured using ovine prolactin as the standard hormone. Cross reactions of pituitary extracts from various non-ruminant species with bovine prolactin antiserum were not detectable. The binding of antibody to 131I labeled bovine prolactin was not inhibited by other pituitary hormones. Bovine prolactin added to plasma was recovered quantitatively. The half-life of injected bovine prolactin in the lactating cow was 29 min.

A study of the antihormonal activity of an antiserum to ovine prolactin using the local lactogenic response in the rabbit.

SUMMARY A rabbit antiserum to a purified preparation of ovine prolactin was prepared. The capacity of the antiserum to counter the biological effect of the preparation of ovine prolactin was determined. When injected intraductally before the injection of prolactin into the same duct the antiserum inhibited the lactogenic effect of prolactin on the rabbit mammary gland. The weight, nitrogen content and reducing sugar content of the mammary glands were used as criteria to judge the effect of the antiserum. The results of specificity tests on the antiserum, using the techniques of double diffusion and immunoelectrophoresis, are also reported.

Effects of exogenous prolactin on reproduction and growth in adult males of the lizard Anolis carolinensis

Adult male Anolis were treated with 5–150 μg/day of exogenous ovine prolactin during the winter and spring under several different conditions of light and temperature. Calorimetric assays of various food insects were used to determine caloric assimilation rates. Prolactin increased food consumption markedly in the winter, when appetites were normally low, and promoted an increase in lean body weight, while fat depots in liver and abdominal fat pads were reduced. Thyroid epithelia were hypertrophied but metabolism was reduced with prolactin. Growth hormone in a dose (1 μg/day) equivalent to the maximum contamination expected in the prolactin preparation had no effect on the growth of Anolis. The natural increases in appetite during the spring obviated the action of prolactin on food consumption, but the hormone continued to exert a somatotropic effect. Prolactin-treated animals stored significantly less fat than controls. Increased body length occurred in prolactin-treated animals maintained on fluctuating temperatures (32-20°C) while the controls only fattened. Prolactin-treated lizards exposed to 32°C continuously did not grow in length but molted more frequently. Thyroid histology and metabolism were unaffected by prolactin during the spring. There was no effect of prolactin on the testes, during recrudescence in the winter or when fully active in the spring. Sexual behavior and accessory sex organs were also unaffected by prolactin, whereas 1 μg growth hormone stimulated accessory sexual structures.

Studies on hormones from the anterior pituitary gland II. Identification and isolation of prolactin from the “granular” fraction of transplanted rat pituitary tumours

An isolation procedure was recently described for bovine prolactin suitable for small quantities of pituitary tissue, based on extraction of the “granular” fraction obtained by differential centrifugation, followed by DEAE-cellulose column chromatography 1. Application of this method to the first to fourth passages of transplantable oestrone-dependent pituitary tumours, originally induced by oestrone pellet implantation and subsequently maintained in oestrone-treated orchidectomized rats, yielded approx. 2 mg of protein per g (wet wt.) of tumour tissue. p]In polyacrylamide-gel electrophoresis at pH 8.9, this protein showed a single major band moving slightly behind the indicator band. Recordings of the differentiated extinction at 277 mμ made during a sedimentation-velocity experiment performed at neutral pH in a Spinco analytical ultracentrifuge (equipped with an ultraviolet scanning system 2) showed a symmetrical peak. Prolactin activity assayed on organ cultures of mouse mammary glands 3,4 was found to be 15.6 I.U./mg. The rat preparation performed better than an ovine prolactin preparation of the stated potency of 20 I.U./mg in inducing and sustaining pseudopregnancies in female mice. An antiserum to the rat prolactin preparation was induced in rabbits. The antiserum neutralized the prolactin activity of the preparation in the assay on organ culture. Precipitation reactions on Ouchterlony plates indicated complete immunological identity between the rat-tumour preparation and prolactin in whole homogenate of normal rat pituitary, but incomplete immunological identity (indicated by spur formation) between the reaction to the latter and that to prolactin in whole homogenate of mouse prolactin-producing tumour tissue.

Isolation of internally labeled rat prolactin by preparative disc electrophoresis.

The preparation of rat prolactin labeled internally with 3H- and 14C-lysine is described. The incorporation procedure was carried out by incubating rat pituitary glands in vitro in the presence of the labeled amino acids. Estrogen pretreatment of the animals enhanced the synthesis and release of prolactin by the incubated pituitaries. Fractionation of the labeled pituitary proteins was performed on a simple preparative disc electrophoresis column capableof separating 10 mg quantities of protein. A large peak of radioactivity which emerged from the column soon after the dye front was shown to be labeled rat prolactin. A much smaller and later peak of radioactive growth hormone was also isolated. The incubation medium was found to contain most of the labeled prolactin formed during the incubation, relatively little prolactin being present in the gland after 20 hr. (Endocrinology 80: 324, 1967)

Erythopoiesis during pregnancy and lactation in the mouse. II. Role of erythropoietin.

Suppression of erythropoietin secretion in lactating and normal female mice by 60% oxygen decreased their RCV and reticulocyte output and abolished plasma erythropoietic activity. In normal mice and in nonlactating postpartum mice, administration of prolactin resulted in an increase of RCV and of PV to values equivalent to those of 15-day postpartum lactating mice. Administration of prolactin to lactating mice and to normal female mice concurrently exposed to hyperoxia prevented the decrease of RCV and of PV which resulted from exposure to hyperoxia alone. The data indicate that in mice, the prolactin preparation used acted as an erythropoietic stimulant and caused a plasma hypervolemia which was not related to the action of erythropoietin.

Prolactin assay by the decidual reaction in the mouse.

SUMMARY The potency of preparations of ovine prolactin NIH-P-S5, NIH-P-S6 and Panlitar Armour S10209 and bovine prolactin NIH-P-B1 was estimated by the luteotrophic assay in normal adult mice. The method is based on the decidual response in a damaged uterine horn. The results were compared with those obtained by the workers using the pigeon crop gland method and found to be closely similar.

Preparation of Bovine, Ovine and Porcine Prolactin1

A simple method is described for the preparation of prolactin from bovine, ovine and porcine pituitary glands, by extraction of the gland residue left from the serial extraction method of Ellis (Endocrinology 64: 554, 1961) with alkaline ethanol and precipitation of the active material at pH 5.5 and 83% ethanol. Products of specific activity 15-25 IU/mg are easily obtained in excellent yields by simple salt fractionation of this precipitate. The preparations are remarkably free of other biological activities. Bovine and ovine prolactin may be still further purified by column chromatography on DEAE-cellulose. (Endocrinology 77: 150, 1965)

THE EFFECT OF GROWTH HORMONE AND PROLACTIN PREPARATIONS ON THE CONTROL BY INTERSTITIAL CELL-STIMULATING HORMONE OF UPTAKE OF 65-ZN BY THE RAT DORSOLATERAL PROSTATE.

SUMMARY Five pituitary hormones: follicle-stimulating hormone (FSH), adrenocorticotrophic hormone (ACTH), thyroid-stimulating hormone (TSH), growth hormone and prolactin, were tested for their capacity to alter the control by interstitial cell-stimulating hormone (ICSH) of the uptake of 65Zn by the dorsolateral prostate gland of the mature hypophysectomized Sprague-Dawley rat. Only prolactin and growth hormone produced significant augmentation of the response to ICSH. Contamination with growth hormone was apparently not responsible for the augmentation of ICSH activity brought about by the NIH-prolactin preparation used. NIH-prolactin and a highly purified preparation provided by Dr C. H. Li were shown to be equally effective in their capacity to augment the 65Zn-uptake response produced by ICSH. Augmentation was detectable with total doses of prolactin as low as 10–30 μg. (0·21–0·63 i.u.). Prolactin caused an augmentation of testosterone activity on uptake of 65Zn in the hypophysectomized and castrated rat, indicating an effect of prolactin on the prostate that is not mediated by the testis. NIH-growth hormone was not as effective as the prolactin preparations in enhancing ICSH activity, but in sufficient doses produced a significant increase in the 65Zn-uptake response to ICSH. These studies showed also that the uptake of 65Zn of the dorsolateral prostate was a more sensitive and more consistent parameter than glandular weight for the detection of augmentation of ICSH response by growth hormone or prolactin preparations.

The effect of growth hormone and prolactin preparations on the intermediary metabolism of rat adipose tissue

1.1. The effects of a bovine growth hormone preparation and three ovine prolactin preparations (PLR, PLC and PLF) on adipose tissue metabolism were compared with those of insulin and adrenalin. PLR and PLC were prepared using a DEAE-cellulose column, whereas PLF was prepared using Sephadex G-75. PLF contains two minor components which are largely absent from PLC. Adrenalin, bovine growth hormone, PLC and PLR increased (a) CO2 production from [6-14C]glucose; (b) glyceride-glycerol production from [I-14C]glucose to a greater extent than from [6-14C]glucose; (c) inhibited lipogenesis and (d) stimulated lipolysis. Dilution of PLC resulted in a diminution of lipolysis with a corresponding decrease in effect upon glucose metabolism. It was concluded that the effect of these pituitary hormones on glucose metabolism by rat adipose tissue is dependent upon their stimulation of lipolysis. Conversely, insulin and PLF stimulated (a) CO2 production from [I-14C]glucose; (b) lipogenesis; and (c) more glyceride-glycerol production from [6-14C]glucose than from [I-14C]glucose. PLC and bovine growth hormone reversed the stimulation of glycogen synthesis by insulin.2.2. PLC, PLF and insulin stimulated protein synthesis from [I-14C]leucine by adipose tissue taken from either normal or hypophysectomized rats. However, bovine growth hormone stimulated leucine incorporation only in tissues taken from hypophysectomized animals.3.3. It is concluded that the mode of action of prolactin on carbohydrate and lipid metabolism is dependent upon the mode of its purification, whereas its action upon protein synthesis is common to all preparations.

IN VITRO ACTIONS OF GROWTH HORMONE ON GLUCOSE METABOLISM IN ADIPOSE TISSUE.

Epididymal adipose tissue obtained from hypophysectomized rats responds to the addition of growth hormone in vitro with increased glucose uptake, oxidation to CO2 and conversion to fatty acids. As little as 0.01 μg/ml of bovine growth hormone was found effective in this regard. In all, 10 growth hormone preparations from 5 species of animals were tested and all were found to be active. Growth hormone also produced these effects when adipose tissues from normal rats fed a high carbohydrate, fat-free diet were tested, but was ineffective on adipose tissue from normal rats fed the standard laboratory ration. The increase in glucose utilization was accompanied by a proportionate increase in appearance of both carbon 1 and carbon 6 in CO2. Corticotropin and thyrotropin, while stimulating glucose uptake and oxidation to CO2, failed to increase the incorporation of glucose carbons into fatty acids. Of the other pituitary hormones tested only oxytocin and certain prolactin preparations caused changes in glucose m...

BIOLOGICAL CHARACTERISTICS OF PITUITARY AND PLACENTAL HORMONES.

Summary. The biological characteristics of gonadotrophin and prolactin were studied by their effects upon the ovaries and the intact and damaged uterine horns of 3-month-old mice which had been hypophysectomized on the first day of dioestrus. The method, developed primarily for measuring luteotrophin by the formation of deciduomata in the damaged uterine horn, was called the deciduoma assay but it can also differentiate the activities of fsh and lh. The decidual reaction in the damaged uterine horn appears to be a suitable endpoint for measuring ovarian progestagen released by the activity of luteotrophin, human luteinizing hormone or placental hormones. These preparations are deciduotrophic but they are not all classified as luteotrophins. The term `luteotrophin' is reserved for preparations which cause hypertrophy of luteal cells accompanied by hyperaemia of the corpora lutea and which are deficient in other gonadotrophic activities. Only preparations of ovine prolactin and luteotrophin in human and bovine growth hormones and bovine thyrotrophin are therefore included in this group. The deciduoma method is possibly invalid for the assay of mixtures containing relatively large amounts of fsh because the growth of follicles results in the elimination of corpora lutea. However, the method can be used to demonstrate biological differences between similar hormones and it may be of value in studies on the purification of hormones.

THE DECIDUOMA ASSAY: A METHOD FOR MEASURING PROLACTIN.

SUMMARY Mice 3 months of age were hypophysectomized on the 1st day of dioestrus and treated with prolactin for 5 consecutive days commencing immediately after hypophysectomy. One uterine horn was traumatized on the 3rd day of the experiment and autopsy was performed on the 6th day. Prolactin maintained the function of the corpus luteum and induced the formation of deciduomata in the injured uterine horn. The increase in weight of the damaged uterine horn compared with the control horn was used as the end point. Since mechanical injury alone caused an increase of up to 10·4 mg., a positive response was taken as an increase of 11 mg. or more. The response was treated as quantal and the percentage of positive responses was transferred to probits. Parallel line assays were used with three or two dose levels of the standard and the unknown. The design permitted a simultaneous study of the effects of a preparation on both the damaged and the undamaged horns of the uterus. Prolactin caused an increase in weight of the damaged horn and gonadotrophin an increase in weight of both horns, but when prolactin was contaminated with gonadotrophin it could still be assayed by the difference in weight between the two. Three different preparations of ovine prolactin (Armour) were assayed against the 2nd International Prolactin Standard and the results compared favourably with the maker's estimate of potency. Two preparations were also assayed by the pigeon crop gland and by the dioestrus methods and the results were similar. The index of discrimination was about unity.

PREPARATION OF A LOW MOLECULAR WEIGHT COMPONENT WITH LACTOGENIC ACTIVITY FROM A LIMITED CHYMOTRYPTIC DIGEST OF OVINE PROLACTIN.

THE limited digestion of ovine pituitary lactogenic hormone by chymotrypsin has recently been investigated in this laboratory1,2. It was found that preparations of prolactin showed lactogenic activity even after 50 per cent digestion with the enzyme. Ultracentrifugation investigations revealed that the prolactin core contained a component3 which had a sedimentation coefficient close to that of the non-treated hormone. The digest also contained another protein fraction which sedim nted more slowly than the bulk of the material3. This communication is concerned with the purification and properties of this latter low-molecular-weight component.

Human Growth Hormonelike Activity of Ovine Prolactin in Man.

s1 April 1963Human Growth Hormonelike Activity of Ovine Prolactin in Man.J. C. Beck, M.D., F.A.C.P., E. E. McGarry, M.D., F.A.C.P.J. C. Beck, M.D., F.A.C.P.Search for more papers by this author, E. E. McGarry, M.D., F.A.C.P.Search for more papers by this authorAuthor, Article, and Disclosure Informationhttps://doi.org/10.7326/0003-4819-58-4-722_3 SectionsAboutPDF ToolsAdd to favoritesDownload CitationsTrack CitationsPermissions ShareFacebookTwitterLinkedInRedditEmail ExcerptGrowth hormone preparations from other than primate sources are biologically inactive in man. Recent chemical, biological, and immunological evidence suggests that human growth hormone is identical or similar to human prolactin. Therefore, the metabolic effect of a purified sheep prolactin preparation was studied in man. Three hypopituitary dwarfs on metabolic balance study were given human growth hormone and, subsequently, sheep prolactin. The sheep prolactin preparation was found to mimic the metabolic action of human growth hormone, producing nitrogen retention, hypercalciuria, and relative impairment of carbohydrate tolerance. One 18-year-old female with a bone age of 12 years and a prepubertal body... This content is PDF only. To continue reading please click on the PDF icon. Author, Article, and Disclosure InformationAffiliations: Montreal, Quebec (BS) PreviousarticleNextarticle Advertisement FiguresReferencesRelatedDetails Metrics Cited ByGrowth and Pituitary Hormones 1 April 1963Volume 58, Issue 4Page: 722-722KeywordsBoneCarbohydratesGrowth hormoneHuman growth hormoneProlactin Issue Published: 1 April 1963 PDF DownloadLoading ...

Studies on the metabolism of adipose tissue; some in vitro effects of a prolactin preparation alone and in combination with insulin or adrenalin.

The glucose uptake, net gas exchange, lactic acid production, oxygen consumption (in the presence of various carbon compounds) and free fatty acid release by rat epididymal adipose tissue in vitro have been measured in the presence of a prolactin preparation (0.2–1.0 mg/ml) alone and in combination with insulin or Adrenalin. Glucose uptake and net gas exchange are stimulated by this pituitary preparation in a manner that mimics the action of insulin. Both these processes may be stimulated further by the concomitant addition of insulin and, in the case of net gas exchange, the effect is synergistic. The prolactin preparation diminishes the production of lactic acid by the tissue and counteracts the action of insulin or Adrenalin in stimulating the formation of this metabolite. The prolactin preparation causes a slight but significant increase in oxygen consumption in the presence of glucose. More striking is the ability of the preparation to stabilize with time the burst in oxygen consumption seen upon add...

Prolongation of dioestrus in the mouse as a quantitative assay of luteotrophic activity of prolactin.

SUMMARY A method has been developed for the quantitative assay of luteotrophic activity of prolactin using the prolongation of dioestrus in mature virgin mice as the end-point of the effect. The assay is based on the principle of an 'all-or-none' response and criteria for a positive result have been specified. A modified parallel line assay has been used with three doses each of the standard and of the unknown. Four different preparations of prolactin have been tested using bovine prolactin, Armour Laboratories, Lot no. D14083–2B as the reference preparation. The assay is not specific, but it may be used for quantitative measurement of the luteotrophic activity of purified preparations of prolactin.

THE IN VITRO EFFECT OF PROLACTIN ON β-GLUCURONIDASE IN THE TESTIS OF THE RAT

SUMMARY The effect of prolactin on the activity of β-glucuronidase in the testis of the rat and mouse has been studied in vitro. Prolactin increased this activity, and a linear log dose-response relationship was found to exist with doses of between 0·3 and 1·2 mg. of a preparation of prolactin containing between 20 and 25 i.u./mg. The optimal conditions for this effect have been investigated. Twenty other substances have been tested; none increased the enzymic activity, but six decreased it. Possible mechanisms for this effect are discussed.

Hormonal specificity of the in vitro action of growth hormone on amino acid transport into rat muscle.

Various pituitary hormones were tested for their ability to stimulate in vitro the transport of AIB into the cells of isolated hypophysectomized rat diaphragms. Crude and purified TSH, purified FSH, purified LH, purified MSH, posterior pituitary extracts and synthetic oxytocin andvasopressin were all ineffective at the concentrations tested. Three ovine prolactin preparations stimulated AIB transport into muscle when present in the incubation medium at high concentrations. This activity was abolished, however, when the prolactin solutions were adjusted to pH 8 and heated for 20 minutes at 100 C. The heating procedure also eliminates the action of growth hormone on amino acid transport but does not alter the ability of prolactin to stimulate the pigeon crop sac. Thus, it is conceivable that the amino acid transport stimulating activity present in ovine prolactin is due to growth hormone contamination. These experiments indicate that growth hormone is the only pituitary hormone which interacts to any extent...

Characterization of Pituitary and Peptide Hormones by Electrophoresis in Starch Gel

SUMMARY A variety of pituitary and other protein hormone preparations exhibited characteristic patterns in starch gel electrophoresis, with the Ferguson and Wallace modification of Poulik’s discon- tinuous buffer system. Five human growth hormone prepara- tions prepared by different methods appeared similar. The four major bands in human growth hormone were inseparable from the four major bands in three different ovine prolactin preparations. Human prolactin shared two major bands in common with both human growth hormone and ovine prolactin. Human, simian, and porcine growth hormone shared three major bands in common, indistinguishable from one another, even with change in pH and prolonged electrophoresis. The simian prepa- ration had one and the porcine preparation four major compo- nents not detected in the human preparation. Purified bovine growth hormone had no bands in common with the other three preparations. Among the melanocyte-stimulating hormones (MSH) , syn- thetic (r-MSH (Hofmann) had the identical electrophoretic mobility as the major component in Schally’s preparation. The characteristic pattern of corticotropin A1 and AQ and ar-MSH and /I-MSH was established; a variety of corticotropin prepara- tions were examined, and the content of each of these peptides was assessed. Three purified parathyroid hormone preparations and two purified relaxin preparations were found not to be homogeneous