Localization Of Prolactin Research Articles

Calreticulin Outside the ER

Calreticulin is considered a resident protein of the endoplasmic reticulum (ER), where it participates in protein folding and calcium homeostasis. Shaffer et al. show that in an in vitro translation and translocation system (containing rough microsomes), a small but detectable fraction of mature processed calreticulin, but not prolactin, appears not to be translocated into the microsomes. This difference in localization of prolactin and calreticulin resulted from differences in the efficiency of translocation into the microsomes, which reflected differences in the signal sequences of the two proteins. Exchange of the signal sequences resulted in complete translocation of calreticulun with the prolactin signal sequence and incomplete translocation of prolactin with the calreticulin signal sequence. Five percent to 20% of the calreticulin had a cytoplasmic localization when calreticulin was overexpressed in transfected cells, depending on the cells and conditions. Calreticulin functions as an inhibitor of transcriptional activation by steroid hormone receptors. Expression of wild-type calreticulin, which produces a small fraction of cytosolic Crt, or the Prl-Crt fusion protein, which is essentially all translocated into the ER, in Crt —/— mouse embryo fibroblasts (MEFs) allowed the effect of cytosolic calreticulin on a glucocorticoid reporter gene to be assessed. Cells with wild-type calreticulin exhibited substantially less reporter gene expression than cells expressing the fully translocated Prl-Crt protein. Thus, inefficient translocation and the production of a small, but functionally important, population of cytosolic calreticulin appears to mediate an inhibitory effect on glucocorticoid receptor signaling. K. L. Shaffer, A. Sharma, E. L. Snapp, R. S. Hegde, Regulation of protein compartmentalization expands the diversity of protein function. Dev. Cell 9 , 545-554 (2005). [PubMed]

Effects of prolactin on the luteinizing hormone response to gonadotropin- releasing hormone in primary pituitary cell cultures during the ovine annual reproductive cycle.

In the sheep pituitary, the localization of prolactin (PRL) receptors in gonadotrophs and the existence of gonadotroph-lactotroph associations have provided morphological evidence for possible direct effects of PRL on gonadotropin secretion. Here, we investigated whether PRL can readily modify the LH response to GnRH throughout the ovine annual reproductive cycle. Cell populations were obtained from sheep pituitaries during the breeding season (BS) and the nonbreeding season (NBS), plated to monolayer cultures for 7 days, and assigned to receive one of the following treatments: 1) nil (control), 2) acute (90- min) bromocriptine (ABr), 3) chronic (7-day) bromocriptine (CBr), 4) ABr and PRL, 5) CBr and PRL, 6) PRL alone, or 7) thyrotropin-releasing hormone. Cells were treated as described above, with the aim of decreasing or increasing the concentrations of PRL in the culture, and simultaneously treated with GnRH for 90 min. The LH concentrations in the medium were then determined by RIA. GnRH stimulated LH in a dose-dependent manner during both stages of the annual reproductive cycle. During the NBS, single treatments did not significantly affect the LH response to GnRH. However, when PRL was combined with bromocriptine, either acutely or chronically, GnRH failed to stimulate LH release at all doses tested (P < 0.01). In contrast, during the BS, the LH response to GnRH was not affected by any of the experimental treatments. These results reveal no apparent effects of PRL alone, but an interaction between PRL and dopamine in the regulation of LH secretion within the pituitary gland, and a seasonal modulation of this mechanism.

Distribution of PRL, GH, TSH and ACTH cells in the pituitary distal lobe of pre‐ and prometamorphic Bufo viridis larvae

The differentiation and topographical localization of prolactin (PRL), growth hormone (GH), thyrotropin (TSH), and adrenocorti‐cotropic hormone (ACTH) cells were studied, using immunohisto‐chemical techniques, in Bufo viridis larval pituitary at different developmental stages, i.e., the beginning of premetamorphosis, the beginning of prometamorphosis, and mid‐prometamorphosis. The four cell types are distributed in different areas of the distal lobe. PRL cells mainly occur in the anterior half, while GH cells are mostly localized in the posterior half, ACTH cells in the dorsoros‐tral area, and TSH cells in the ventrorostral one. This distribution is maintained, without significant changes, during development. All the different cell types undergo a relevant increase in their cell number during premetamorphosis; in particular, TSH cells are very few at the beginning of premetamorphosis, being the least abundant cell population, but become well represented at the end of it. On the other hand, GH cells are the most numerous cell population, both during pre‐ and prometamorphosis. Finally, our observations indicate that the pituitary intermediate lobe is already well differentiated and distinct from the pars distalis at the beginning of premetamorphosis.

Cost- and Time-Saving Immunohistochemical Method for Co-localization of Prolactin with Growth Hormone or Adrenocorticotropic Hormone in Rat Pituitary

Abstract A cost-effective and time-saving immunohistochernical capillary gap method for consistent single staining of cells in pituitaries from large numbers of rats was developed for localization of prolactin (PRL), growth hormone (GH) or adrenocorticotropic hormone (ACTH). Co-localization of PRL with either GH or ACTH using rabbit polyclonal antibodies required a 5 min enzymatic digestion in 0.04% pepsin/0.1 N hydrochloric acid between antibody stains to reduce background, and an appropriate chromogen selection and application order. In the co-localization experiments, when PRL staining using AEC as chromogen was followed by GH staining using Vector SG as chromogen, a significant number of double-labeled acidophils were seen. If either antibody detection order or chromogen order was reversed, identification of double-labeled cells was more difficult. PRL and ACTH were easily distinguished in the same section of rat pituitary because both hormones were not seen within the same cell. This chromogen combin...

Cost- and Time-Saving Immunohistochemical Method for Co-localization of Prolactin with Growth Hormone or Adrenocorticotropic Hormone in Rat Pituitary

A cost-effective and time-saving immunohistochernical capillary gap method for consistent single staining of cells in pituitaries from large numbers of rats was developed for localization of prolactin (PRL), growth hormone (GH) or adrenocorticotropic hormone (ACTH). Co-localization of PRL with either GH or ACTH using rabbit polyclonal antibodies required a 5 min enzymatic digestion in 0.04% pepsin/0.1 N hydrochloric acid between antibody stains to reduce background, and an appropriate chromogen selection and application order. In the co-localization experiments, when PRL staining using AEC as chromogen was followed by GH staining using Vector SG as chromogen, a significant number of double-labeled acidophils were seen. If either antibody detection order or chromogen order was reversed, identification of double-labeled cells was more difficult. PRL and ACTH were easily distinguished in the same section of rat pituitary because both hormones were not seen within the same cell. This chromogen combination may have application with other antibodies. (The J Histotechnol 20:313, 1997)

Immune localization of prolactin receptor in the mitochondria-rich cells of the euryhaline teleost ( Oreochromis mossambicus) gill

The present work demonstrates, by Western blotting and immunofluorescent staining, the presence and localization of prolactin (PRL) receptor in tilapia (ti) Oreochromis mossambicus gills. Gill epithelial cells that reacted with PRL receptor antibody were found to be labelled concomitantly with Con A, a marker of the apical crypts in mitochondria-rich (MR) cells. No positive staining was observed in pavement cells or mucus cells with PRL receptor antibody. This indicates that PRL receptors are located specifically in the gill MR cells. Further, the tiPRL receptors were found only in the MR cells of seawater-adapted tilapia gills. The effects of salinity and ions on the expression of tiPRL receptors are discussed. © 1997 Federation of European Biochemical Societies.

Localization of prolactin and its receptor messenger RNA in the human decidua.

Prolactin (PRL) is known as an anterior pituitary hormone. On the other hand, PRL is also produced in the human decidualized endometrium. The physiological role and site of action of endometrial PRL have not yet been clarified. This study was designed to investigate the localization of PRL receptor (PRL-R) gene-expressing cells in the human decidualized endometrium using in situ hybridization histochemistry. Sense and antisense 35S-labeled RNA probes for human PRL-R mRNA were hybridized with cryostat sections of human decidua, which were obtained from patients undergoing therapeutic abortion at 8-10 weeks of gestation. Hybridization signals for PRL-R mRNA were seen over the decidual cells. No labeled cells were seen in the chorion, amnion, or trophoblast. Comparing the localization of PRL-R gene-expressing cells to that of PRL gene-expressing cells using adjacent sections, their distributions were quite similar. These results indicate that not only PRL but also PRL-R transcripts are located in the decidual cells.

Immunohistochemical Localization of Prolactin in Normal and Neoplastic Uterine Cervical Tissues

Immunohistochemical Localization of Prolactin in Normal and Neoplastic Uterine Cervical Tissues

The Effect of Epidermal Growth Factor (EGF) with Estradiol and Progesterone on Prolactin Secretion from Cultured Human Decidual Tissues of Early Pregnancy

Prolactin is now accepted as a normal product of the decidual cells of the human endometrium. We investigated the effect of epidermal growth factor (EGF) with estradiol and progesterone on prolactin secretion by the decidual tissues from the early pregnant endometrium. The decidual tissues were separated from villi, minced and cultured in collagen gel matrix with serum-free medium. Immunological staining of the cultured decidual tissues showed prolactin localization and EGF receptors on the stromal cells. Cultured media were collected every 2 days. The culture for the first 2 days was incubated with the serum-free medium alone (= preculture), and the following test culture was supplemented with/without additives. The prolactin content in cultured media was quantified by EIA. The results of the effect of steroid(s) and EGF were represented as a comparison of prolactin contents in the preculture and the test culture. An increase in prolactin secretion was found after the tissues were treated with a combination of 10(-8)M estradiol and 10(-6)M progesterone or 10(-6)M progesterone alone. After 8 days, the prolactin secretion rate increased about 3-fold over the precultured value. Estradiol alone kept the prolactin secretion at the precultured value. Prolactin secretion gradually decreased in the non-additive culture. These results indicate that progesterone was essential in the secretion of prolactin. Simultaneously, similar decidual tissues were incubated with a combination of EGF and steroid(s). The secretion of prolactin in the group treated with progesterone alone decreased dose-dependently responding to added EGF on the 8th day of culture. In the presence of estradiol and progesterone, the secretion rate decreased to the values similar to the progesterone alone group with the addition of 0.1, 1 ng/ml EGF, and the decrease in prolactin secretion was less with the addition of 10 ng/ml EGF. Mixed cultures of the decidual tissues and villi showed that the prolactin secretion rate increased in all groups treated with/without estradiol and/or progesterone. These results imply that progesterone derived from villi might control decidual prolactin secretion. The effect of high concentration EGF (50 ng/ml) on the prolactin secretion appeared similar to the isolated decidual tissues. These results suggest that decidual prolactin secretion is regulated by the combined effects of steroids and EGF.

Immunocytochemical localization of prolactin in carcinoma of the cervix

Immunocytochemical methods were used to demonstrate prolactin in both normal and malignant human cervices. Four of five cervices with epidermoid carcinoma and three of four cervices with adenocarcinoma demonstrated prolactin. One of four normal cervices stained positive for prolactin in the endocervical glands. Four cervices from pregnant patients were positive for prolactin. The results of this study show prolactin in human cervical carcinomas. The rote and source of cervical prolactin is unknown.

ULTRASTRUCTURAL LOCALIZATION OF PROLACTIN IN THE HUMAN PITUITARY PROLACTINOMAS AND ITS CHANGES BY BROMOCRIPTINE TREATMENT

Fourteen human prolactinomas, four of which received bromocriptine treatment, were studied by immunoelectron microscopy to observe the intracellular localization of prolactin (PRL). In the untreated cases two different patterns of PRL localization were noted. In the first type (Type I) PRL was seen in Golgi saccules, single stacks of rough endoplasmic reticulum (RER), and a few secretory granules and in the second type (Type II) it was present in the predominant parallel arrays of RER. Type II cells were observed more frequently in the cases with higher serum prolactin levels. In the treated cases with normalized serum prolactin levels, the tumor cells showed accumulation of secretory granules. This finding suggested secretory inhibition. RER and Golgi saccules were inconspicuous in these cells and this might indicate decreased production of PRL. In a case which was resistant to the bromocriptine treatment, most tumor cells were similar in ultrastructural localization of PRL to those cells in the untreated cases and this might suggest the lack of dopamine receptor.

506 Subcellular Localization of prolactin in long-term cultures of human prolactinomas. An immunoelectron microscopic study of two cases

506 Subcellular Localization of prolactin in long-term cultures of human prolactinomas. An immunoelectron microscopic study of two cases

Ultrastructural immunocytochemical localization of prolactin and growth hormone in the porcine pituitary.

The immunocytochemical peroxidase-antiperoxidase technique was used to identify prolactin- and growth hormone-producing cells in the porcine pituitary at the ultrastructural level. The growth hormone-producing cells contain round secretory granules (300 nm to 500 nm in diameter). The prolactin-producing cells can be identified by their distinct round and ovoid secretory granules which vary in size. Most of these cells contain large granules (450 nm to 750 nm in diameter), but some prolactin-producing cells display smaller secretory granules (250 nm to 500 nm). The two hormones were localized exclusively in the secretory granules. Staining for prolactin was observed in round and ovoid granules, as well as in small and polymorphic granules within the Golgi complex. This study confirmed (i) that the two hormones are located in different cells, and (ii) that under normal physiological conditions no one cell can synthesize and store both hormones simultaneously.

AUTORADIOGRAPHIC LOCALIZATION OF THE BINDING OF 125I-LABELLED PROLACTIN TO RAT TISSUES IN VITRO

The binding of 125I-labelled prolactin to normal rat tissues was examined with autoradiographic techniques. Tissue slices were incubated with 125I-labelled ovine prolactin in the presence or absence of excess unlabelled hormone to distinguish between the specific and the non-specific localization of grains. Specific binding was found in the liver, adrenal gland and kidney from female rats, and in the mammary gland, ovary, testis and prostate gland. Conversely, fat, muscle, heart, lung, and spleen from female rats and uterine tissue did not bind 125I-labelled prolactin appreciably, or show competition for binding in the presence of unlabelled hormone. In the testis, prolactin bound exclusively to Leydig cells; in the prostate gland, binding was localized in the secretory epithelium. Kidney tubules in the cortex displayed specific prolactin localization whereas the medulla did not bind prolactin. Adrenal medulla showed no hormone binding; however, the zona reticularis and to a lesser extent the zona fasciculata bound prolactin. In the ovary, grains were limited to the theca and to a lesser extent the corpus luteum and follicles. In all cases, binding was essentially abolished when unlabelled prolactin was included in the incubation medium. These results confirm reports of the presence of prolactin receptors in these tissues and serve to identify the cells within a particular organ that respond to prolactin.

Prolactin Cells of the Human Pituitary Gland in Old Age

Pituitary glands obtained at autopsy from 20 male and 20 female subjects, over 80 years of age, have been investigated with various staining procedures, including the immunoperoxidase technique for the cytological localization of prolactin. Prolactin cells were numerous and well developed in the anterior lobes. The incidence, distribution, granulation and immunoreactivity of prolactin cells showed no apparent difference between old male and female subjects dying of acute or chronic illnesses. It can be concluded that prolactin cells do not regress in old age. The morphologic findings are consistent with the assumption that the pituitary gland is capable of secreting prolactin in aging subjects.

Movement and localization of prolactin in the pigeon crop-sac: an autoradiographic study.

Department of Applied Pharmacology, School of Pharmacy, The Hebrew University, Jerusalem, Israel (Received 9 August 1976) Although the proliferation of the pigeon crop-sac induced by prolactin has been used during four decades for quantitative evaluation of the hormone, little is known of the molecular mechanism of this process. The crop-sac comprises layers of mucosa, sub-mucosa, muscle and skin: prolactin is known to stimulate the hyperplastic multiplication of the mucosa itself. This mucosal epithelium is a multilayer (stratified) of flat (squamous) non-cornified tissue in which cuboidal cells of the basal layer can be easily distinguished. This progressive flattening of the cells in the more superficial layers suggests that the reproductive layer is nearer the basal layer. How does prolactin stimulate this proliferation? Previous studies from this Department have demonstrated the biological activity of 125I-labelled prolactin on the pigeon crop mucosa in vivo (Shani, Givant, Sulman, Eshkol & Lunenfeld, 1972) and in

Localization of prolactin in rat kidney tissue using a double-antibody technique.

A fluorescein-labelled double-antibody technique was used for the microscopical localization of ovine prolactin in kidney tissue of rats previously injected intravenously with 50 i.u. of the hormone at varying intervals before death. Thirty seconds after the injection, the hormone could already be visualized in teh lumina of the proximal tubules. After 2 min, discrete praticulate fluorescence appeared on the brush border of the epithelium of the proximal tubules. Three minutes later prolactin was clearly localized in the apical portion of the cytoplasm of these cells as well as on the brush border. Ten minutes after the injection, the hormone appeared exclusively within the cell, highest concentrations being observed in the perinuclear region. After 20 min, however, the hormone appeared evenly distributed throughout the cytoplasm. The distributtion of prolactin was in all cases confined to the proximal tubular region of the nephron. The findings support the suggestion that the site of action of prolactin in the kidney is the epithelium of the proximal tubule. As prolactin was first visualized in the lumen of this portion of the nephron, it appers that the hormone gains access to these cells via the glomerular filtrate.

Immunohistochemical localization of prolactin in the pituitary gland of the pigeon, Columba livia.

Specific antibodies to ovine prolactin were produced in rabbits. The antisera were conjugated to fluorescein isothiocyanate and "purified" by column chromatography. Antibodies, thus labelled, were applied to fresh-frozen sections of pigeon pituitary glands. Fluorescence was observed in specific cells of the cephalic lobe. After staining, these cells were identified as the erythrosinophilic eta cells.

Prolactin localization in the primate pituitary by immunofluorescence.

Cells which contain prolactin were clearly distinguished from those which contain growth hormone in adult monkey pituitary glands by means of histologic and fluorescent antibody techniques. The results indicate that in primates, as well as in other mammals, prolactin is immunochemically distinguishable from growth homone.

Localization of prolactin in the reptilian pars distalis

The relationship between the distribution of prolactin and the various cell types in the pars distalis (PD) of two species of lizard, two species of snake and a turtle were examined in order to determine the site of prolactin production in the reptilian pituitary. Prolactin was measured by pigeon crop-sac bioassay, and, in the turtle, by polycrylamide gel (disc) electrophoresis. Pigeon crop-sac stimulating activity (prolactin) was observed in the pituitary of all species examined. In every case, most of the hormone was localized in the rostral half of the PD and in the lizards none was detected in the caudal region. Disc electrophoresis verified this finding in the turtle and also suggested that a presumptive growth hormone is concentrated in the caudal half of the chelonian gland. Thus, it appears likely that the lactotropes are located rostrally and the somatotropes caudally in the reptilian PD. Histological studies of the PD demonstrate two types of acidophils which show a marked regional distribution; a carminophil is restricted almost entirely to the rostral half of the PD and an orangeophil to the caudal half. On the basis of patterns of hormone distribution, it seems likely that the rostral carminophil is the lactotrope, and the caudal orangeophil may be the somatotrope.