Uptake Of Prolactin Research Articles

PROLACTIN BINDING SITES IN XENOPUS LAEVIS (DAUDIN) (ANURA PIPIDAE) KIDNEY: AN AUTORADIOGRAPHIC AND IMMUNOCYTOCHEMICAL STUDY

SUMMARY We have studied prolactin binding sites in Xenopus laevis (Daudin) (Anura Pipidae) kidney by autoradiography and immunocytochemistry. By the autoradiographic technique after intracardiac injection of 125I- oPRL labelling is appreciable in Xenopus laevis kidney at the level of the large vessels and capillaries, the renal corpuscle and, to a variable extent, within the lumina and epithelial cells of the proximal convoluted tubules. Compared to the autoradiographic reaction, the immunocytochemical method failed to reveal the hormone when extremely low doses of it were injected but allowed a much more exact localization of prolactin uptake by the epithelial cells of the proximal convoluted tubules. On the basis of the autoradiographs we could not ascertain wheter, or to what extent, the hormone was bound to specific renal cell PRL receptors: in fact, not only insulin and luteinizing hormone, but also unlabelled prolactin, injected as a specific competitor, failed to bring about the recognizable displa...

Milk prolactin, feed volume and duration between feeds in women breastfeeding their full-term infants over a 24 h period.

Previous studies have suggested that the uptake of prolactin from the blood into the milk may be restricted when the alveolus is distended with milk. Therefore the aim of this study was to determine the relationship between prolactin in milk and milk production by measuring the concentration of prolactin in samples of fore- and hind-milk as well as the volume of milk removed for each breast, at each breastfeed over a 24 h period. The mean (+/- S.D.) concentration of prolactin in milk for all women (n = 15) over the 24 h period was 18.5 +/- 11.6 microg l(-1) (fore-milk) and 16.8 +/- 12.8 microg l(-1) (hind-milk). The variation between women masked small changes within women in the concentration of prolactin in milk over the 24 h period, therefore a prolactin ratio (individual fore- or hind-milk value divided by the mean for all fore- or hind-milk samples collected over a 24 h period) was determined. The concentration of prolactin was highest in milk between 02.01 and 06.00 h (prolactin ratio for fore- to hind-milk, 1.5), and lowest between 10.01 and 18.00 h (prolactin ratio for fore- to hind-milk, 0.8). Furthermore, we observed that the difference in prolactin concentration between the fore- and hind-milk (fore-hind gradient) was greatest between 06.01 and 10.00 h (4 microg l(-1)). To ensure that this effect was not due to permeability in the paracellular pathway, the concentrations of serum albumin and sodium in milk were measured. No significant (P > 0.05) changes over the 24 h period, or with increasing time since last feed were observed. We therefore concluded that when the breast is most drained of milk (in the late evening), and the rate of milk synthesis is greatest, the fore-hind prolactin gradient in the milk of the following feed (in the early morning) is highest. This is consistent with the observation that prolactin uptake from the blood by the lactocyte is dependent on the fullness of the breast, such that prolactin uptake may be inhibited in full alveoli.

Immunocytochemical demonstration of prolactin interaction with choroid plexus in aging and acute hyperprolactinemia

Prolactin (PRL) exerts a direct effect on the central nervous system, reaching the PRL-responsive brain regions via cerebro-spinal fluid (CSF). The hormone enters the CSF by a specific receptor-mediated transport mechanism that is localized on the epithelium of the choroid plexus (CP) of brain ventricles. PRL interactions with the CP in aging were examined in young (3-month) and old (27-month) female Wistar rats using immunocytochemistry (immunogold technique). The enhancement of PRL uptake by the CP in animals at both ages was achieved by the modelling of acute hyperprolactinemia. A great age-related difference was found in the intensity of immunocytochemical reaction under activated conditions, the uptake of PRL by CP being significantly higher in young animals than in old. The character of the colloidal gold particle distribution in different components of CP epithelial cells appeared to be the same in both age groups. The weakening of PRL-transporting capacity in the CP of old animals may constitute one aspect of the alteration of neuroendocrine regulation in the CP-CSF system that occurs during aging.

Prolactin is transported across the epithelium of the jejunum and ileum of the suckling rat.

Milk prolactin is transferred from the gastrointestinal tract to the circulation of the suckling rat. To identify the site of prolactin penetration and to determine the mechanism by which the hormone traverses the mucosal barrier, we followed the uptake of prolactin from ligated loops of jejunum or ileum in vivo by three methods: autoradiography, transport of prolactin-gold conjugates, and immunocytochemistry. Autoradiographic studies demonstrated specific binding sites for 125I-prolactin on apical membranes of the jejunum and ileum. Excess cold prolactin reduced radiolabel in apical and basal compartments. Gel autoradiography of portal sera showed the presence of intact prolactin and a prolactin fragment following jejunal transport but only a prolactin fragment following ileal transport. Uptake of prolactin-gold conjugates demonstrated that, in the jejunum, label was present at the luminal surface, within endosomal compartments and lysosomes, in basal coated and smooth vesicles, within basal coated pits, and beyond the basolateral surface. In the ileum, label was found at the luminal surface; within the tubulocisternae, endosomal vesicles, lysosomes, and basal smooth vesicles; and beyond the basolateral surface. Immunoreactive prolactin was present throughout the transepithelial pathways. This study demonstrates that prolactin is selectively and nonselectively absorbed in the jejunum and ileum and that the hormone is directed either to the lysosome for degradation or across the epithelium by means of a transcellular pathway.

In vivo autoradiographic analysis of prolactin binding in brain and choroid plexus of the domestic ring dove

The binding of intravenously administered prolactin to choroid plexus and brain tissue was determined radioautographically in the ring dove, a species that exhibits prolactin-induced alterations in brain function. An intense autoradiographic reaction was detected over the epithelial cells of the choroid plexus 5 min after the intravenous injection of 125I-ovine prolactin. A significant reaction was also observed over the infundibulum but no significant uptake of prolactin occurred in other brain areas. The binding of radiolabelled prolactin to infundibulum appeared to be non-specific, since excess unlabelled hormone did not reduce silver grain density. In contrast, 125I-ovine prolactin binding in choroid plexus was significantly reduced by excess unlabelled ovine prolactin or human growth hormone, but not by ovine luteinizing hormone. Specific binding to choroid plexus was also detected in vitro. The lack of significant brain uptake of prolactin in vivo is discussed in relation to recent in vitro evidence for specific binding sites for prolactin in several dove brain regions. Similarities between the binding results obtained in this avian species and those reported previously in mammals suggest that the two vertebrate groups exhibit similar patterns of prolactin interaction with neural target tissues.

Secretion and Mammary Gland Uptake of Prolactin in Dairy Cows During Lactogenesis

Mammary arteriovenous differences of prolactin concentration and net mammary uptake of prolactin from blood were quantified near parturition in 9 dairy cows. Six cows were milked once daily for at least 6 d before parturition, and prepartum lactogenesis occurred in 3 of 6 cows. Prepartum milking 2 or more d before parturition abruptly increased secretion of prolactin into blood, but milkings within 1 d before or after parturition did not increase prolactin secretion. Concentrations of prolactin in whole milk sampled over 8 d before parturition (64.5ng/ml) were substantially greater than those occurring several days after parturition (19ng/ml). Milk prolactin concentrations were unaffected by the successful induction of prepartum lactogenesis, which greatly increased prepartum yields of milk (2 to 8 kg/milking). Therefore, the alveolar lumenal content of prolactin was greatest in pregnant cows with prepartum lactogenesis. This enhanced content of intraalveolar prolactin before parturition was associated with an absence of mammary uptake of prolactin immediately prior to ejection of the prolactin-containing milk from the alveoli. However, prolactin uptake was quickly restored to about 2μg/min per half udder shortly after milk ejection. During the prepartum period, an enhanced intraalveolar reservoir of 200 to 400μg prolactin, due to induction of prepartum lactogenesis, appears to saturate temporarily all putative sites for uptake of prolactin from blood.

Differential kinetics and sensitivity to chloroquine of receptor-mediated insulin and prolactin endocytosis in liver parenchymal cells

Systemically injected [ 125I]prolactin or [ 125I]insulin was accumulated and cleared from rat liver at different rates. Quantitative subcellular fractionation indicated a predominant accumulation of [ 125I]insulin in liver microsomes while [ 125I]prolactin was found in both the light-mitochondrial and microsomal fractions. The acidotropic agent chloroquine diminished the rate and extent of loss of each ligand from liver homogenates. In chloroquine treated rats, radiolabeled insulin accumulated in both the light-mitochondrial and the microsomal fractions. Subfractionation of microsomes on discontinuous sucrose gradients revealed ‘early’ endosomes in which ligand uptake was maximal at 2–5 min. In contrast, comparable subfractionation of the light mitochondrial fraction revealed ‘late’ endosomes in which ligand uptake was maximal at 10–20 min. Chloroquine-treated rats showed a more marked enhancement of insulin compared to prolactin uptake in the ‘early’ endosomes. It is suggested that ‘early’ endosomes found in the Golgi-intermediate and -heavy fractions floated from parent microsomes may selectively degrade insulin but not prolactin. This could account for the apparently different kinetics of insulin and prolactin uptake into liver parenchyma.

Prolactin uptake into liver endocytic components. Reduced sensitivity to chloroquine

Subfractionation of hepatic Golgi fractions on Percoll gradients revealed two populations of lactogen-receptor enriched components with one pool of ϱ = 1.040–1.050 and a high-density component of ϱ = 1.053–1.064. 125I-labelled prolactin uptake into Percoll gradient subfractions demonstrated rapid accumulation into low-density elements and slower accumulation in high-density structures. Electron microscope radioautography demonstrated that silver grains were largely associated with lipoprotein-filled structures. Radiolabel was highly concentrated in these components especially the very pure higher-density component where enrichment over the homogenate was 184-fold at the peak time after injection (10 min). Administered chloroquine accumulated in the higher-density component but not in low-density elements, suggesting that the former are at an acid intraluminal pH. In contrast to the marked effect of chloroquine on insulin uptake into endocytic structures found in Golgi fractions (Posner, B.I., Patel, B.A., Khan, M.N. and Bergeron, J.J.M. (1982) J. Biol. Chem. 257, 5789–5799), little effect of this pH disrupting agent was observed on prolactin uptake. The difference between the effect of chloroquine on 125I-labelled prolactin and 125I-labelled insulin uptake may reflect the greater stability of prolactin-receptor complexes in a low-pH environment.

Localization of prolactin uptake by the kidney of the teleostean, Colisa lalia

Although evidence has been presented that the hormone prolactin specifically binds to the kidneys of the teleost Sarotherodon mossambicus ( J. N. Fryer, 1979, Gen. Comp. Endocrinol. 39, 397–403.) prolactin has not previously been shown to become localized at any particular morphological sites in fishes. This problem has been investigated in the dwarf gourami, Colisa lalia. Ovine prolactin was labeled with 125I and injected intramuscularly. Fish were sacrificed after 15, 30, and 90 min. Other animals were injected with an excess of unlabeled prolactin, growth hormone, or luteinizing hormone 2 min prior to administration of the label. Paraffin-embedded tissues were sectioned, coated with Kodak NTB-3, and exposed for 3–5 weeks. After developing, samples of several body tissues were stained and examined for the presence of silver grains. The results indicate that, as in rat and frog, kidney tubules of C. lalia are sites of prolactin localization. No other sites of prolactin localization were identified.

Interaction between prolactin and zinc in the human prostate gland.

The interaction between prolactin and zinc was examined in vitro in the human prostate gland. The results indicated that prolactin did not modulate the acute uptake of zinc into benign prostatic hypertrophy tissue whereas zinc, in contrast, increased the uptake of prolactin into the prostate gland. Our study further showed that the augmented uptake of prolactin by zinc was partly due to an increase in the non-specific binding properties of the peptide hormone. We were also able to demonstrate that the specific binding of 125I-labelled human prolactin to the receptor was reduced in the presence of zinc by a competitive mechanism.

Characterization of the prolactin receptor in human prostate.

The uptake of 125I-labelled human prolactin by subcellular fractions obtained from the human hyperplastic prostate was investigated and the specific binding sites were characterized. The binding was both time- and temperature-dependent with maximum specific uptake achieved after incubation for 20 h at 4 degrees C (dissociation constant = 2.4 x 10(-10) mol/l). The present studies also indicate that the bulk of the specific prolactin binding was confined to the 105 000 and 15 000 g membrane fractions whilst the cytosol and nuclear pellet exhibited a lower capacity for the peptide hormone. Attempts to optimize the binding revealed that pretreatment of the subcellular fractions with 4 mol MgCl2/l quadrupled the binding sites; a similar effect was produced after pretreatment with dextran-coated charcoal, but the changes were less pronounced. Furthermore, our studies suggest that the binding was specific for the human prolactin, with other hormones such as ovine prolactin, human LH, human FSH and human GH showing little or no competition. The addition of magnesium and copper ions to the incubation medium also markedly increased the specific binding, whereas calcium, manganese and EDTA inhibited the prolactin uptake. Freezing and storage of tissue at -70 degrees C did not greatly affect the binding sites. The results suggest that we are clearly dealing with a specific prolactin-binding protein.

Uptake of prolactin from cerebrospinal fluid in rat brain

The fate of prolactin in the cerebrospinal fluid was studied by the use of fluorescein labelled prolactin. The distribution was compared to the immunocytochemical distribution of endogenous prolactin. High intensity fluorescence was found in the ependyma of the area postrema, rostral dorsolateral cerebral aqueduct close to the subcommissural organ, and in some cells of the floor of the cerebral aqueduct. This distribution was not seen when excess unlabelled prolactin was injected. The results suggest prolactin uptake from CSF at specific sites which correspond to sites of localization of immunoreactive prolactin.

No evidence for a peripheral effect of L-DOPA on plasma prolactin in the rat

Male and female rats with two permanently indwelling intravenous catheters were infused for 2 hours with ovine prolactin. During equilibrium conditions the effects of intravenously injected L-DOPA and benzerazide (a blocker of dopa-decarboxylase) on steady state levels of ovine prolactin were measured. A dose of 4.5 mg L-DOPA per 100 gr body weight (b.w.) caused a transient increase of plasma ovine prolactin. A dose of 0.3 mg L-DOPA/100 gr b.w. had no effect, neither in males nor in females, while benzerazide (20 mg/100 gr b.w.) had only a slight effect. The experiments suggest that L-DOPA does not affect the peripheral uptake of prolactin from the plasma.

Effect of colchicine on the uptake of prolactin and insulin into Golgi fractions of rat liver.

In previous studies we have shown that 125I-labeled prolactin is taken up by a receptor-dependent process and concentrated in an intact form in Golgi elements from female rat liver (J. Biol. Chem., 1979, 254:209-214). In this study we have examined the effect of colchicine on this uptake process into Golgi elements. Colchicine [25 mumol (10 mg)/100 gm body wt] was injected intraperitoneally in adult female rats, and hepatic Golgi fractions were prepared at 1, 2, and 3 h postinjection. The enzyme recoveries and morphological appearance of fractions from colchicine-treated and control (alcohol alone) animals were similar. At times greater than 1 h after colchicine there was a marked (greater than 60%) inhibition of uptake of 125I-ovine prolactin (125I-oPRL) into Golgi light and intermediate fractions but no inhibition of uptake into Golgi heavy and plasmalemma elements. At times from 2 to 45 min postinjection, 125I-oPRL was extracted from Golgi elements and found to be largely intact as judged by rebinding to receptors. The inhibitory effect of colchicine was seen at doses ranging from 0.25 mumol to 25 mumol/100 g body wt. Vincristine also inhibited 125I-oPRL uptake into the Golgi light and intermediate fractions but lumicolchicine had no inhibitory effect. There was a smaller effect of colchicine both at early (1 h) and later (3 h) times on the extent and pattern of 125I-insulin uptake. Colchicine treatment did not produce a significant change in lactogen receptor levels in the Golgi fractions. These results demonstrate that colchicine treatment inhibited the transfer of prolactin into Golgi vesicular elements. The much smaller effect on insulin uptake suggests that there may be differences in the manner in which the two hormones are handled in the course of internalization.

The uptake of prolactin into female rat liver. Concentration of intact hormone in the Golgi apparatus.

The uptake of prolactin into female rat liver. Concentration of intact hormone in the Golgi apparatus.

Dopamine enhances the action of prolactin on rat blood vessels. Implications for dopamine effects on plasma prolactin.

It has been claimed that dopamine enhances peripheral uptake of prolactin. Dopamine at 2.1 x 10(-9)M, a concentration which had no effect by itself, enhanced both the potentiation of rat mesenteric vascular reactivity caused by 50ng/ml ovine prolactin and the inhibition of reactivity caused by 500ng/ml prolactin. These observations are consistent with the proposal that dopamine can interact peripherally with prolactin.

Prolactin inhibition test with L-dopa: decrease and restoration of plasma prolactin levels in the rat by a peripheral process.

L-DOPA, within 30 min after administration, induced a highly significant decrease of plasma prolactin levels (phase 1) in a number of groups of rats, differing in age and/or endocrine status, apparently by direct inhibition of prolactin release from the pituitary. Three hours after administration of L-DOPA these low plasma prolactin concentrations in treated animals had increased (phase 2) and did not differ significantly from levels in control animals, indicating that the effect of L-DOPA on plasma prolactin levels is only of short duration. During this process some interesting phenomena were observed, especially in the animals treated with oestrone. The elimination rate of prolactin from plasma was very high (t 1/2 = 2.8 min), as indicated by decreasing concentrations of the hormone during phase 1. Pituitary prolactin content did not change during phase 1, suggesting that prolactin synthesis was also stopped. Notwithstanding the high elimination rate, plasma prolactin regained initial concentrations in phase 2, suggesting release of a substantial part of the pituitary prolactin content. The latter,however, remained constant during the whole experiment (i.e. before L-DOPA administration and during phase 1 as well as phase 2). The results suggested another working mechanism of L-DOPA in decreasing plasma prolactin levels, namely by stimulating the uptake of this hormone in the periphery. After the effect of L-DOPA had ceased, most of the prolactin from the periphery returned into the bloodstream, causing a rapid restoration of plasma prolactin levels without substantial release from the pituitary. The nature of the processes responsible for the peripheral uptake of prolactin is discussed.