Prolactin levels in functional hypothalamic amenorrhea: a retrospective case-control study.

PURPOSE: Functional hypothalamic amenorrhea (FHA) is due to hypothalamic dysregulation. Literature lacks data about prolactin in FHA women, although both prolactin levels and FHA are associated with stress. Moreover, polycystic ovarian morphology is common in FHA and there is an association between FHA and polycystic ovary syndrome. Thus, the aim of this study was to assess prolactin levels in FHA patients and controls with a special focus on factors influencing prolactin levels, that could be considered as "sensors" of the hypothalamic-pituitary dysregulation. METHODS: In a retrospective cohort study, 140 women with clearly defined FHA were compared to 70 healthy, normally ovulating women matched for age. The main outcome parameter was prolactin. Factors associated with prolactin levels > 12 µg/L were tested using a multivariable binary logistic regression model. RESULTS: The median prolactin level was 11.5 µg/L (interquartile range, IQR 7.5-14.4), which was similar to the control group (median 10.7, IQR 8.3-14.5; p = 0.065). Only two women had hyperprolactinemia (prolactin > 25 µg/L; 1.4%). In a multivariable binary logistic regression model eating disorder (odds ratio, OR 0.206; p = 0.040), excessive exercise (OR 0.280; p = 0.031) and TSH (OR 1.923; p = 0.020) were significantly associated with prolactin levels > 12 µg/L. CONCLUSION: Women with FHA have similar prolactin levels to healthy age-matched individuals. Eating disorders and excessive exercise where associated with prolactin levels < 12 µg/L, in contrast to TSH.

Structure and function of a dual antagonist of the human growth hormone and prolactin receptors with site-specific PEG conjugates.

Human growth hormone (hGH) is a pituitary-derived endocrine protein that regulates several critical postnatal physiologic processes including growth, organ development, and metabolism. Following adulthood, GH is also a regulator of multiple pathologies like fibrosis, cancer, and diabetes. Therefore, there is a significant pharmaceutical interest in developing antagonists of hGH action. Currently, there is a single FDA-approved antagonist of the hGH receptor (hGHR) prescribed for treating patients with acromegaly and discovered in our laboratory almost 3 decades ago. Here, we present the first data on the structure and function of a new set of protein antagonists with the full range of hGH actions-dual antagonists of hGH binding to the GHR as well as that of hGH binding to the prolactin receptor. We describe the site-specific PEG conjugation, purification, and subsequent characterization using MALDI-TOF, size-exclusion chromatography, thermostability, and biochemical activity in terms of ELISA-based binding affinities with GHR and prolactin receptor. Moreover, these novel hGHR antagonists display distinct antagonism of GH-induced GHR intracellular signaling in vitro and marked reduction in hepatic insulin-like growth factor 1 output in vivo. Lastly, we observed potent anticancer biological efficacies of these novel hGHR antagonists against human cancer cell lines. In conclusion, we propose that these new GHR antagonists have potential for development towards multiple clinical applications related to GH-associated pathologies.

The role of prolactin/vasoinhibins in cardiovascular diseases.

Prolactin (PRL) is a polypeptide hormone that is mainly synthesized and secreted by the lactotroph cells of the pituitary. There are two main isoforms of PRL: 23-kDa PRL (named full-length PRL) and vasoinhibins (including 5.6-18 kDa fragments). Both act as circulating hormones and cytokines to stimulate or inhibit vascular formation at different stages and neovascularization, including endothelial cell proliferation and migration, protease production, and apoptosis. However, their effects on vascular function and cardiovascular diseases are different or even contrary. In addition to the structure, secretion regulation, and signal transduction of PRL/vasoinhibins, this review focuses on the pathological mechanism and clinical significance of PRL/vasoinhibins in cardiovascular diseases.

Plasmin generates vasoinhibin-like peptides by cleaving prolactin and placental lactogen.

Vasoinhibin is an antiangiogenic, profibrinolytic peptide generated by the proteolytic cleavage of the pituitary hormone prolactin by cathepsin D, matrix metalloproteinases, and bone morphogenetic protein-1. Vasoinhibin can also be generated when placental lactogen or growth hormone are enzymatically cleaved. Here, it is investigated whether plasmin cleaves human prolactin and placental lactogen to generate vasoinhibin-like peptides. Co-incubation of prolactin and placental lactogen with plasmin was performed and analyzed by gel electrophoresis and Western blotting. Mass spectrometric analyses were carried out for sequence validation and precise cleavage site identification. The cleavage sites responsible for the generation of the vasoinhibin-like peptides were located at K170-E171 in prolactin and R160-T161 in placental lactogen. Various genetic variants of the human prolactin and placental lactogen genes are projected to affect proteolytic generation of the vasoinhibin-like peptides. The endogenous counterparts of the vasoinhibin-like peptides generated by plasmin may represent vasoinhibin-isoforms with inhibitory effects on vasculature and coagulation.

Thrombin Cleaves Prolactin Into a Potent 5.6-kDa Vasoinhibin: Implication for Tissue Repair.

Vasoinhibin is an endogenous prolactin (PRL) fragment with profibrinolytic, antivasopermeability, and antiangiogenic effects. The fact that blood clotting, vascular permeability, and angiogenesis are functionally linked during the wound-healing process led us to investigate whether thrombin, a major protease in tissue repair, generates vasoinhibin. Here, we have incubated human PRL with thrombin and analyzed the resulting proteolytic products by Western blot, mass spectrometry, high-performance liquid chromatography purification, recombinant production, and bioactivity. We unveil a main thrombin cleavage site at R48-G49 that rapidly (< 10 minutes) generates a 5.6-kDa fragment (residues 1-48) with full vasoinhibin activity, that is, it inhibited the proliferation, invasion, and permeability of cultured endothelial cells and promoted the lysis of a fibrin clot in plasma with a similar potency to that of a conventional 14-kDa vasoinhibin (residues 1-123). The R48-G49 cleavage site is highly conserved throughout evolution and precedes the intramolecular disulfide bond (C58-C174), thereby allowing the 5.6-kDa vasoinhibin to be released without a reduction step. Furthermore, the 5.6-kDa vasoinhibin is produced by endogenous thrombin during the clotting process. These findings uncover the smallest vasoinhibin known, add thrombin to the list of PRL-cleaving proteases generating vasoinhibin, and introduce vasoinhibin as a thrombin-activated mechanism for the regulation of hemostasis, vasopermeability, and angiogenesis in response to tissue injury.

Development of Vasoinhibin-Specific Monoclonal Antibodies.

Vasoinhibin is a protein hormone with antiangiogenic, antivasodilatatory, and antivasopermeability effects generated by the proteolytic cleavage of prolactin. The discovery of its role in diabetic retinopathy and peripartum cardiomyopathy led to the evaluation of new pharmacological treatments in clinical interventional trials. However, the quantitative evaluation of vasoinhibin in biological samples from patients has not been possible due to the lack of vasoinhibin-specific antibodies. Recently, loop 1 of vasoinhibin was identified to have a different three-dimensional structure compared to PRL, and thus to contain vasoinhibin-specific epitopes. Here, we report the development of two sets of vasoinhibin-specific monoclonal antibodies against two neighboring regions of the vasoinhibin loop 1. An experimental sandwich ELISA with two monoclonal anti-vasoinhibin antibodies was developed, which had no cross-reactivity to recombinant human full-length prolactin. The ELISA had a quantitation limit of 100 ng/ml, and intra-assay- and inter-assay coefficients of variation of 12.5% and 14%, respectively. The evaluation of 15 human serum samples demonstrated concentrations of below limit of detection (n=3), below limit of quantitation (n=1) and between 0.23 µg/ml (230 ng/ml) to 605 µg/ml (n=12) in the quantifiable range. Despite the high specificity of the monoclonal-monoclonal antibody sandwiches which discriminate vasoinhibin from PRL, there might be cross-reactivities by serum proteins other than vasoinhibin. A fully established vasoinhibin ELISA may support diagnostic and therapeutic measures in vascular diseases.

A computational approach for explaining the effect of the prl gene polymorphism on prolactin structure and biological activity in Japanese quails.

Prolactin is a versatile hormone with multiple activities, including a negative control on egg production. This study was conducted to genotype all the coding portions of the prl gene using PCR-SSCP-sequencing, and to investigate the effects of amino acid substitutions of the prl gene on the structure and function of prolactin in quails using in silico approach. Though all genotyped exons exerted homogenous PCR-SSCP patterns, a total of 12 novel SNPs were detected in the investigated exons, including three SNPs in exon-1, 8 SNPs in exon-2, and one SNP in exon-4. Three adjacent missense SNPs were detected in exon-2, namely H69P, T70P, and S71F. Computational tools indicated obvious deleterious effects of T70P, with less extent to H69P and S71F on prolactin functions and activity, which may lead to limited participation of this hormone in the negative control of egg production. In conclusion, the introduction of in silico prediction has suggested an alternative solution for the breeders to evaluate the effect of each witnessed nsSNP in protein structure and function. The current study suggests three nsSNPs, T70P, T70P, and S71F as strong candidates for the negative effect on prolactin biological activity with a consequent reversal positive effect on egg productivity traits.

Prolactin peptide (pPRL) induces anti-prolactin antibodies, ROS and cortisol but suppresses specific immune responses in rainbow trout.

Prolactin has several immune functions in fish however, the effects on innate and specific components of rainbow trout immunity are currently unknown. Therefore in this study, prolactin peptide (pPRL) injection in rainbow trout generated anti-PRL antibodies that were confirmed through Western blot assays of fish brain tissue extract. At the same time, this group of fish was immunized with a viral antigen (VP2) and the specific antibody titer generated by the rainbow trout was subsequently determined, as well as the sero-neutralizing capacity of the antibodies. Interestingly, this group of fish (pPRL-VP2) generated approximately 150% less antibodies compared with fish immunized only with the viral antigen (VP2), and pPRL-VP2 fish increased their cortisol level by 4 times compared to the control. Additionally, through qPCR assay, we determined that the pPRL-VP2 fish group decreased pro-inflammatory transcript expression, and the serum of these (pPRL-VP2) fish stimulated ROS production in untreated fish leukocytes, a phenomenon that was blocked by the pharmacological cortisol receptor inhibitor (RU486). Collectively, this is the first report that indicates that pPRL could modulate both components of immunity in rainbow trout.

Identification and structural characterization of the factors involved in vitellogenesis and its regulation in the African Osteoglossiforme of aquacultural interest Heterotis niloticus (Cuvier, 1829).

The African bonytongue (Heterotis niloticus) is an excellent candidate for fish farming because it has outstanding biological characteristics and zootechnical performances. However, the absence of sexual dimorphism does not favor its reproduction in captivity or the understanding of its reproductive behavior. Moreover, no molecular data related to its reproduction is yet available. This study therefore focuses on the structural identification of the different molecular actors of vitellogenesis expressed in the pituitary gland, the liver and the ovary of H. niloticus. A transcriptomic approach based on de novo RNA sequencing of the pituitary gland, ovary and liver of females in vitellogenesis led to the creation of three transcriptomes. In silico analysis of these transcriptomes identified the sequences of pituitary hormones such as prolactin (PRL), luteinizing hormone (LH) and follicle-stimulating hormone (FSH) and their ovarian receptors (PRLR, FSHR, LHR). In the liver and ovary, estrogen receptors (ER) beta and gamma, liver vitellogenins (VtgB and VtgC) and their ovarian receptors (VLDLR) were identified. Finally, the partial transcript of an ovarian Vtg weakly expressed compared to hepatic Vtg was identified based on structural criteria. Moreover, a proteomic approach carried out from mucus revealed the presence of one Vtg exclusively in females in vitellogenesis. In this teleost fish that does not exhibit sexual dimorphism, mucus Vtg could be used as a sexing biomarker based on a non-invasive technique compatible with the implementation of experimental protocols in vivo.

Protein secondary structure determines the temporal relationship between folding and disulfide formation.

How and when disulfide bonds form in proteins relative to the stage of their folding is a fundamental question in cell biology. Two models describe this relationship: the folded precursor model, in which a nascent structure forms before disulfides do, and the quasi-stochastic model, where disulfides form prior to folding. Here we investigated oxidative folding of three structurally diverse substrates, ß2-microglobulin, prolactin, and the disintegrin domain of ADAM metallopeptidase domain 10 (ADAM10), to understand how these mechanisms apply in a cellular context. We used a eukaryotic cell-free translation system in which we could identify disulfide isomers in stalled translation intermediates to characterize the timing of disulfide formation relative to translocation into the endoplasmic reticulum and the presence of non-native disulfides. Our results indicate that in a domain lacking secondary structure, disulfides form before conformational folding through a process prone to nonnative disulfide formation, whereas in proteins with defined secondary structure, native disulfide formation occurs after partial folding. These findings reveal that the nascent protein structure promotes correct disulfide formation during cotranslational folding.

A novel type of prolactin expressed in the bullfrog pituitary specifically during the larval period.

Prolactin (PRL) is one of the major hormones that control amphibian metamorphosis. Recently, a PRL (PRL1B) gene that is different from the known PRL (PRL1A) gene has been found in the genomes of several amphibian species. In order to ascertain whether the PRL1B gene is expressed in the bullfrog (Rana catesbeiana) pituitary, cloning of cDNA encoding PRL1B in the pituitary of the premetamorphic bullfrog tadpole was attempted. The bullfrog PRL1B amino acid sequence predicted from the obtained cDNA showed 62% identity with those of Xenopus PRL1Bs that have been presumed from the genome sequences, whereas the sequence identity between bullfrog PRL1A and PRL1B was 48%. A molecular phylogenetic tree showed that bullfrog PRL1B is most appropriately grouped with amphibian PRL1Bs. Quantitative PCR analysis revealed that the mRNA expression levels of bullfrog PRL1B in the pituitary were high during pre- and prometamorphosis, sharply declined at metamorphic climax and became undetectable after metamorphosis. In contrast, PRL1A mRNA levels were relatively low during pre- and prometamorphosis, rose at climax and remained high after metamorphosis. Immunohistochemical study using antibodies against partial peptides of PRL1A and PRL1B revealed that most of the PRL1A- and PRL1B-immunoreactive cells in the larval pituitary were distributed separately, but that some of the cells immunoreactive with both antibodies were also present. Western blot analysis with the larval pituitary extract indicated that PRL1B-immunoreactive band appeared at the position of molecular weight ca. 22.1 kDa and PRL1A-immunoreactive band at the position of ca. 22.8 kDa. The results obtained in this experiment suggest the possibility that PRL1B plays as-yet-unknown role(s) during the pre-climactic period of metamorphosis. This is the first report on the existence of PRL1B as a protein in the amphibian larval pituitary.

In silico prediction of prolactin molecules as a tool for equine genomics reproduction.

The prolactin hormone is involved in several biological functions, although its main role resides on reproduction. As it interferes on fertility changes, studies focused on human health have established a linkage of this hormone to fertility losses. Regarding animal research, there is still a lack of information about the structure of prolactin. In case of horse breeding, prolactin has a particular influence; once there is an individualization of these animals and equines are known for presenting several reproductive disorders. As there is no molecular structure available for the prolactin hormone and receptor, we performed several bioinformatics analyses through prediction and refinement softwares, as well as manual modifications. Aiming to elucidate the first computational structure of both molecules and analyse structural and functional aspects related to these proteins, here we provide the first known equine model for prolactin and prolactin receptor, which obtained high global quality scores in diverse software's for quality assessment. QMEAN overall score obtained for ePrl was (- 4.09) and QMEANbrane for ePrlr was (- 8.45), which proves the structures' reliability. This study will implement another tool in equine genomics in order to give light to interactions of these molecules, structural and functional alterations and therefore help diagnosing fertility problems, contributing in the selection of a high genetic herd.

Molecular evolution of prolactin in Chiroptera: Accelerated evolution and a large insertion in vespertilionid bats.

Pituitary prolactin (PRL) shows an episodic pattern of evolution in mammals, with a slow underlying rate (near stasis) and periods of rapid change in some groups. PRL evolution in bats, the second most speciose mammalian order, has not previously been studied, and is examined here. Slow basal evolution of PRL is seen in some bats, particularly megabats, but in most microbat groups evolution of PRL is more rapid. Accelerated evolution of PRL is particularly notable in the family Vespertilionidae, where analysis of nonsynonymous and synonymous substitutions indicates that it reflects adaptive evolution/positive selection. Remarkably, vespertilionid bats also show a large sequence insertion, of variable length, into exon 4 of PRL, giving a protein sequence 18-60 amino acids longer than normal, with the longest insertions in bats of the genus Myotis. An equivalent insertion has not been reported in PRL of any other vertebrate group. In the 3-dimensional structure of the complex between PRL and the extracellular domain (ecd) of its receptor (PRL:PRLR2) the inserted sequence is seen to be introduced in the short loop between helices 2 and 3 of PRL; it is far removed from the receptor-binding sites, and may not interfere with binding. The ecd of the receptor also shows variable rates of evolution, with a higher rate in the Vespertilionidae, but this is much less marked than for the hormone. The distribution of substitutions introduced into PRL during vespertilionid evolution appears to be non-random, and this and the evidence for positive selection suggests that the rapid evolution and insert sequence introduction were associated with a significant change in the biological properties of the hormone.

Novel reagents for human prolactin research: large-scale preparation and characterization of prolactin receptor extracellular domain, non-pegylated and pegylated prolactin and prolactin receptor antagonist.

To provide new tools for in vitro and in vivo prolactin (PRL) research, novel protocols for large-scale preparation of untagged human PRL (hPRL), a hPRL antagonist (del 1-9-G129R hPRL) that acts as a pure antagonist of hPRL in binding to hPRL receptor extracellular domain (hPRLR-ECD), and hPRLR-ECD are demonstrated. The interaction of del 1-9-G129R hPRL with hPRLR-ECD was demonstrated by competitive non-radioactive binding assay using biotinylated hPRL as the ligand and hPRLR-ECD as the receptor, by formation of stable 1:1 complexes with hPRLR-ECD under non-denaturing conditions using size-exclusion chromatography, and by surface plasmon resonance methodology. In all three types of experiments, the interaction of del 1-9-G129R hPRL was equal to that of unmodified hPRL. Del 1-9-G129R hPRL inhibited the hPRL-induced proliferation of Baf/LP cells stably expressing hPRLR. Overall, the biological properties of del 1-9-G129R hPRL prepared by the protocol described herein were similar to those of the antagonist prepared using the protocol reported in the original study; however, the newly described protocol improved yields by >6-fold. To provide long-lasting hPRL as a new reagent needed for in vivo experiments, we prepared its mono-pegylated analogue and found that pegylation lowers its biological activity in a homologous in vitro assay. As its future use will require the development of a PRL antagonist with highly elevated affinity, del 1-9-G129R hPRL was expressed on the surface of yeast cells. It retained its binding capacity for hPRLR-ECD, and this methodology was shown to be suitable for future development of high-affinity hPRL antagonists using a library of randomly mutated open reading frame of del 1-9-G129R hPRL and selecting high-affinity mutants by yeast surface display methodology.

Influence of sulpiride treatment on the level of prolactin and immunoglobulins in the peripheral blood of mares during the postpartum period.

The aim of this study was to determine the effect of increased levels of prolactin (PRL) on the concentration of immunoglobulins in the blood, colostrum and milk of mares. The study was conducted on 12 mares of the Polish Pony breed (6 in the control and 6 in the experimental group). To induce hyperprolactinaemia in mares of the experimental group, 750 mg sulpiride was administered orally once a day. The initial PRL concentration was 52.22 ± 11.21 ng/ml in the control group and 49.39 ± 10.12 ng/ml in the experimental group. In the subsequent days, the concentration of PRL dynamically changed. Statistical analysis showed highly significant differences (P < 0.01) between the groups. The concentration of immunoglobulins in the blood plasma was at the same level during the experimental period (32.97-29.08 mg/ml in the experimental group and 28.60-18.11 mg/ml in the control group). Statistical analysis showed highly significant differences between the groups in blood plasma immunoglobulin level (P < 0.01). The highest immunoglobulin concentration was obtained within 12 h after parturition in the control and the experimental group (23.49 ± 2.12 mg/ml and 26.94 ±1.72 mg/ml, respectively). The lowest values were obtained on day 12 after parturition in the experimental group (10.15 mg/ml ± 1.47 mg/ml) and on day 7 after parturition in the control group (14.30 mg/ml ± 2.48 mg/ml). In conclusion, this study did not provide evidence that the lactogenic hormone prolactin is involved in the transfer of immunoglobulins into the colostrum in horses.

The effect of different protease inhibitors on stability of parathyroid hormone, insulin, and prolactin levels under different lag times and storage conditions until analysis.

INTRODUCTION: Proteolytic cleavage through proteases affects peptide hormone levels, which is of particular significance when the time interval between sampling and analysis is prolonged. We evaluated the stability of parathyroid hormone, insulin, and prolactin molecules (i) with different protease inhibitors such as K2 EDTA, aprotinin, and protease inhibitor cocktail (PIC), (ii) with different lag times (6-72 hours), and (iii) under different storage temperatures (4°C vs room temperature [RT]) until analysis. MATERIALS AND METHODS: Blood samples were collected into 2 sets of 5 Vacutainer® tubes (Becton Dickinson) from 10 healthy adults. Tubes 1 and 2 were plain gel separator tubes. Tubes 3, 4, and 5 contained PIC (1%), aprotinin (500 KIU/mL), and K2 EDTA, respectively. After centrifugation at 1300 g for 10 minutes, PIC added to tube 2 of each set. Samples were analyzed and then one set was stored at 4°C, whereas the other at RT until analysis at 6, 24, 48, and 72 hours. Hormone levels were determined with electrochemiluminescence immunoassay (ModularE170; Roche Diagnostics). The results were compared with desirable bias limits (DBL) from Westgard QC database. RESULTS: Insulin at RT decreases exceeding the DBL starting from 24 hours and K2 EDTA preserved insulin. PTH exceeded the DBL at RT for 48 hours or longer and PIC addition after centrifugation inhibited its degradation. Prolactin remained stable in all tested conditions. All parameters in the plain gel separator tubes remained within DBL when stored at 4°C until 72 hours. CONCLUSIONS: Different proteases may degrade peptide hormones and measures should be taken to counteract these effects especially if there is a delay before analysis.

A Complex Dance: The Importance of Glycosaminoglycans and Zinc in the Aggregation of Human Prolactin.

The zinc binding hormone pituitary human prolactin (hPRL) is stored in secretory granules of specialized cells in an aggregated form. Glycosaminoglycans (GAGs) are anionic polysaccharides commonly associated with secretory granules, indicating their involvement in granule formation. Here we, for the first time, study the impact of GAGs in combination with Zn(2+) on the reversible hPRL aggregation across the pH range of 7.4-5.5. Zn(2+) alone causes hPRL aggregation at pH 7.4, while aggregation between pH 7.4 and 5.5 requires both Zn(2+) and GAGs. GAGs alone cause hPRL aggregation below pH 5.5. Comprehensive thermal stability investigations show that hPRL is particularly destabilized toward thermal denaturation at pH 5.5 and that GAGs increasingly destabilize hPRL at decreasing pH values. We propose that Zn(2+) causes hPRL aggregation through low-affinity Zn(2+) binding sites on hPRL with GAGs facilitating Zn(2+) binding by neutralizing repulsive positive charges of hPRL in the acidic environments of the TGN and mature secretory granules. In a manner independent of the aggregation-causing agent(s), the different hPRL aggregates show very similar secondary structure and amorphous morphology. We speculate that this may be a recognizable sorting signal in the formation of hPRL granular vesicles.

Urine oligosaccharide pattern in patients with hyperprolactinaemia.

Free milk-type oligosaccharides are produced during pregnancy and lactation and may have an impact on several cells in the immune system. Our aim was to investigate if patients with isolated hyperprolactinaemia, not related to pregnancy, also have increased synthesis and urinary excretion of milk-type oligosaccharides and to compare the excretion pattern with that found during pregnancy. Urine samples were collected as morning sample from 18 patients with hyperprolactinaemia, 13 healthy controls with normal prolactin levels and four pregnant women. After purification, lactose and free oligosaccharides were analysed and quantified by high-performance anion-exchange chromatography with pulsed amperometric detection. The identity of peaks was confirmed by exoglycosidase treatment and comparison with oligosaccharide standards. Prolactin was measured in serum collected between 09 and 11 a.m. by a standardized immunochemical method. Patients with hyperprolactinaemia had higher urinary excretion of lactose than normoprolactinemic controls and urinary lactose correlated positively to prolactin levels (r = 0.51, p < 0.05). Increased levels of the fucosylated oligosaccharides 2-fucosyl lactose and lacto-di-fucotetraose were found in urine from three and two patients, respectively. The acidic oligosaccharide 3-sialyl lactose was found in high amount in urine from two patients with prolactin of >10,000 mU/l. However, pregnant women in their third trimester had the highest concentration of all these oligosaccharides and excretion increased during pregnancy. This study is first to show that both lactose and certain fucosylated and sialylated milk-type oligosaccharides are increased in some patients with hyperprolactinaemia. It remains to elucidate the functional importance of these findings.

Effects of age and reproductive experience on the distribution of prolactin and growth hormone secreting cells in the anterior pituitary of a passerine.

Plasma prolactin (PRL) is released from lactotrophs in the anterior pituitary. As plasma PRL levels rise during incubation in domestic fowl, the number of lactotrophs (PRL-immunoreactive, PRL-IR cells) increases while the number of growth hormone secreting cells, somatotrophs (GH-IR cells), declines. We measured plasma PRL levels using radioimmunoassay (RIA) and examined the distribution of lactotrophs and somatotrophs in the anterior pituitary of breeding and nonbreeding zebra finches of known ages with and without prior breeding experience using fluorescent immunohistochemistry (IHC). Plasma PRL levels were higher in breeding than in nonbreeding birds, regardless of age, sex, or previous breeding history. PRL-IR cells were localized primarily, but not exclusively, to the cephalic aspect of the anterior pituitary (AP) and along the ventral margin. Birds with prior reproductive experience had more PRL-IR cells than birds with no prior reproductive experience and breeders had slightly higher PRL-IR cell counts than did nonbreeders, but there was no correlation between the number of PRL-IR cells and plasma PRL levels. GH-IR cells were concentrated in the caudal aspect of the AP with some cells in the cephalic lobe, but numbers did not differ between any of the groups studied. An increase in PRL-IR cells corresponded with an increase in GH-IR cells. An increase in lactotroph number with reproductive experience in zebra finches may facilitate future reproductive events by allowing for more robust PRL secretion and increased reproductive success.