Prolactin Gene Research Articles

Effect and Interaction of β-Lactoglobulin, Kappa Casein, and Prolactin Genes on Milk Production and Composition of Awassi Sheep.

Simple SummaryBeta-lactoglobulin (β-LG), prolactin (PRL), and Kappa casein (CSN3) all contribute to the determination of milk production and composition, but have not been assessed in local Awassi sheep. Therefore, our aim was to analyze the contribution of these genes in milk production and composition traits in commercial Awassi ewe population by genotyping and sequencing these genes. Our results showed the prevalence of the different variations (alleles) of the tested genes in the Awassi population, and no association among β-LG and CSN3 polymorphic genotypes and milk production, or PRL and fat%. Also, all 3 genes help determine the milk production potential of Awassi ewes and help assess milk components, and thus can be used in breeding programs to select for milk potential.A participatory animal-breeding program was applied to 9 commercial Awassi sheep flocks in Jordan. This study aimed to assess the influence of Beta-lactoglobulin (β-LG), Prolactin (PRL), and Kappa casein (CSN3) genes, genotypes and their interaction on milk production and composition traits of 167 genotyped Awassi ewes via Polymerase Chain Reaction (PCR) followed by sequencing. Allele frequencies for the two variants were 0.42 and 0.58 for β-LG, 0.82 and 0.18 for PRL, and 0.92 and 0.08 for CSN3. No association was found among β-LG and CSN3 polymorphic genotypes with milk production traits. However, ewes with PRL AA genotype showed higher milk production, β-LG AB was associated with lowest fat%, high solid not fat (SNF)%, protein%, and lactose%. β-LG BB was associated with highest milk density. PRL, β-LG, and CSN3 polymorphic genotypes were differentially associated with milk production and component traits. Furthermore, β-LG × PRL interaction showed the highest milk production and fat%; β-LG × PRL recorded the highest SNF%, protein%, lactose%, and milk density, while the PRL × CSN3 had the highest fat% and SNF%. The enhancing effects of these gene interactions can be incorporated in Awassi breeding programs to improve milk production and composition.

Disentangling juxtacrine from paracrine signalling in dynamic tissue.

Prolactin is a major hormone product of the pituitary gland, the central endocrine regulator. Despite its physiological importance, the cell-level mechanisms of prolactin production are not well understood. Having significantly improved the resolution of real-time-single-cell-GFP-imaging, the authors recently revealed that prolactin gene transcription is highly dynamic and stochastic yet shows space-time coordination in an intact tissue slice. However, it still remains an open question as to what kind of cellular communication mediates the observed space-time organization. To determine the type of interaction between cells we developed a statistical model. The degree of similarity between two expression time series was studied in terms of two distance measures, Euclidean and geodesic, the latter being a network-theoretic distance defined to be the minimal number of edges between nodes, and this was used to discriminate between juxtacrine from paracrine signalling. The analysis presented here suggests that juxtacrine signalling dominates. To further determine whether the coupling is coordinating transcription or post-transcriptional activities we used stochastic switch modelling to infer the transcriptional profiles of cells and estimated their similarity measures to deduce that their spatial cellular coordination involves coupling of transcription via juxtacrine signalling. We developed a computational model that involves an inter-cell juxtacrine coupling, yielding simulation results that show space-time coordination in the transcription level that is in agreement with the above analysis. The developed model is expected to serve as the prototype for the further study of tissue-level organised gene expression for epigenetically regulated genes, such as prolactin.

Effects of high and low temperature on expression of GnIH, GnIH receptor, GH and PRL genes in the male grass puffer during breeding season

Gonadotropin-inhibitory hormone (GnIH) is a multifunctional hypophysiotropic neurohormone and has a stimulatory role in the control of reproduction in the grass puffer. To clarify the neuroendocrine mechanisms underlying the effect of changes in water temperature on reproduction in fish, we previously revealed that, in parallel to gonadal regression, both low and high temperature significantly decreased the expressions of the genes encoding kisspeptin (kiss2), kisspeptin receptor (kiss2r), gonadotropin-releasing hormone 1 (gnrh1) in the brain and gonadotropin (GTH) subunits (fshb and lhb) in the pituitary of sexually mature male grass puffer. In this study, we examined the changes in expression of gnih and GnIH receptor gene (gnihr) in the brain and pituitary along with the genes for growth hormone (gh) and prolactin (prl) in the pituitary of male grass puffer exposed to low temperature (14 °C), normal temperature (21 °C, as initial control) and high temperature (28 °C) conditions for 7 days. The levels of gnih and gnihr mRNAs were significantly decreased in both low and high temperature conditions compared to normal temperature in the brain and pituitary. Similarly, the gh mRNA levels were significantly decreased in both low and high temperature conditions. The prl mRNAs showed no significant changes at high temperature, whereas drastically decreased at low temperature possibly by dysfunctional cold stress. Taken together, the present results suggest that, in addition to the inhibitory effect of temperature changes on the Kiss2/GnRH1/GTH system, the suppression of GnIH/GH system may also be involved in the termination of reproduction by high temperature at the end of breeding season.

MON-434 Familial Alactogenesis Associated with a Prolactin Mutation

BACKGROUND: Isolated prolactin deficiency is a rare disorder manifesting as absence of puerperal lactation. We identified a family with multiple members who had alactogenesis and examined genetic causes. SUBJECTS: Three women in one family (proband, sister and niece) reported puerperal alactogenesis, i.e. no milk production postpartum. All the women had regular menstrual cycles during their reproductive years. The proband, currently 66 yrs, had 7 pregnancies and 2 live births. Prolactin level was 1.16 ng/mL (2.8-29.2 ng/mL) at the age of 47 yrs. The sister, currently 73 yrs, had 3 children. Her prolactin level was 1.4 ng/mL (2.8-29.2 ng/mL) at the age of 64 yrs. Both women had menopause before age 45 years. The niece, currently 43 years old, had infertility of unknown cause and conceived 2 children with fertility treatment. She had prolactin levels of 1.13 ng/mL, 0.618 ng/mL and 0.759 ng/mL at the age of 34-36 yrs. METHODS: DNA and mRNA were extracted from blood of all three women and a fertile control with a history of normal lactation. PRL exons 1-6 were Sanger sequenced from the DNA. cDNA was subjected to RT-PCR with primers spanning intron 5 and the terminal portion of exon 6. RT-PCR was analyzed using the 2-ΔΔCT method. Prolactin mRNA expression was compared among the affected women and the control subject. RESULTS: We identified a heterozygous mutation (c.658C>T) changing CGA (arginine) to TGA (stop codon) (p.Arg220Ter) in exon 6 of the prolactin gene. PRL mRNA expression (0.011±0.003 vs. 1.0±0.0; p<0.001) and terminal exon 6 expression (0.0027±0.0028 vs. 0.99±0.05; p<0.001) was decreased compared to control mRNA expression. DISCUSSION: We have identified a genetic cause of puerperal alactogenesis resulting from a stop-gain mutation in the prolactin gene (PRL). PRL mRNA expression is decreased in these subjects. These findings suggest that the mutated PRL may undergo nonsense decay. The low circulating prolactin resulted in inability to breastfeed. It may also be the cause of infertility and early menopause, but further studies are needed. CONCLUSION: We report a stop-gain mutation in exon 6 of the prolactin gene (PRL) resulting in familial puerperal alactogenesis.

A Study on the Effect of Prolactin Gene Variants on Milk Production Traits of Holstein Cattle

Recent studies aiming to improve the daily milk production of cattle using both genetic and environmental factors are extremely important. The aim of this study is to determine the possible relationships between daily milk production and polymorphic distribution of exon 3 region of the prolactin (PRL) gene, which is an important marker for selection. With this purpose, 155 Holstein cows were included in the study, and the demographic data of the cattle were recorded. Venous blood was collected from the jugular vein of cows in 2 tubes of 2 mL each containing EDTA for DNA isolation. Genomic DNA were isolated from these whole blood samples. The prolactin gene region of the cattle were identified using PCR/RFLP and Sanger DNA sequencing method. The genotype, allele frequencies and Hardy–Weinberg equilibrium were calculated using Arlequin version 3.5.1.3 package software. Accordingly, it was found that cows with genotype AA had statistically higher milk yield as compared to those with genotype AB and BB. Missense mutation, frameshift and silent mutations were detected in the prolactin gene exon 3. It was also found that cows with missense mutation had statistically lower milk fat percentage as compared to the others. Consequently, the effects of genotypic distribution and mutation in the prolactin gene on the milk production of Holstein cows, which is a special breed in terms of milk productivity, have been revealed.

PRL -1149T allele (rs1341239) is associated with decreased risk of rheumatoid arthritis in population from southern Mexico: analysis of mRNA expression and PRL serum levels.

Prolactin (PRL) is a sex hormone with immunomodulatory properties, and it is associated with the clinical activity of rheumatoid arthritis (RA). The -1149G>T polymorphism at the prolactin (PRL) gene has been associated with autoimmune diseases, but its functional effect is unclear. To analyze the association of the PRL -1149G>T polymorphism with disease susceptibility, mRNA, and protein expression of PRL in RA patients from Southern Mexico. We included 300 RA patients and 300 control subjects (CS). Genotypes were identified by the polymerase chain reaction-restriction fragment length polymorphism (PCR-RFLP) technique, the PRL mRNA expression was determined by real-time PCR, and PRL serum levels were measured by enzyme-linked immunosorbent assay. Applying genetic models of inheritance (dominant, recessive, and additive), we found an association between the T allele and decreased RA susceptibility (OR = 0.55, 95% CI 0.35-0.87, p = 0.009; OR = 0.09, 95% CI 0.012-0.76, p = 0.011; OR = 0.49, 95% CI 0.32-0.76, p = 0.001, respectively). RA patients had higher mRNA expression and soluble levels of PRL than CS (p < 0.05). The PRL serum levels were similar in RA and CS according to genotypes. However, in CS, carriers of GT and TT genotypes showed lower PRL mRNA expression than GG genotype carriers (7.1-fold and 20-fold respectively, p = 0.006). This study demonstrated that the PRL -1149T allele is a genetic marker of decrease risk to RA in population from Southern Mexico, and it is associated with low PRL mRNA. • PRL -1149T allele is a marker of decreased RA susceptibility in population from southern Mexico. • PRL -1149TT genotype is associated with low PRL mRNA expression. • RA patients have higher mRNA expression and soluble levels of PRL than healthy subjects. • PRL serum levels are higher in those RA patients with < 2years of disease evolution.

Polymorphisms of the prolactin gene and their association with egg production traits in two Chinese domestic ducks

ABSTRACT1. Prolactin (PRL) as a polypeptide hormone which plays a crucial role in egg production traits.2. Polymorphisms of the PRL gene were analysed with DNA sequencing and polymerase chain reaction-single-strand conformation polymorphism methods in two Chinese domestic laying duck breeds (Jinding, n = 400, Youxian, n = 400, respectively).3. The results showed that one polymorphism was detected (A-412G) in intron 1 of the PRL gene, with three genotypes: AA, AG and GG. Association analysis showed that the ducks with the GG genotype had significantly greater egg production and egg weight than those with AG and AA genotype (p < 0.05). Hence, the 412A > G polymorphism of the PRL gene in intron 1 is a potentially valuable genetic marker for laying duck breeding programmes.

Potency of papua local chickens as egg producers: a molecular review

This research aims to evaluate potency of Papua Local Chickens as egg producers through allelic frequency estimation of 24-bp indel (insertion-deletion) on the prolactin gene promoter (cPRL) owned by the population. A total of 68 Papua Local Chickens (ALP) are used in this research, and blood samples are taken individually for DNA isolation. DNA fragment target size 130 or 154 bp contain the 24-bp indel locus is successfully amplified using Polymerase Chain Reaction (PCR) method with a pair of specific primer. Electrophoresis analysis on 1.5 % agarose gel indicates three genotypes: II, ID and DD with the frequency of 8.82 %, 41.18 % and 50.00 %, respectively. The I and D allele frequencies are 0.29 and 0.71. This result estimate shows that the I allele frequency is categorized medium in Papua Local Chickens population. Therefore, the Papua Local Chickens is potential in producing eggs. Their egg producer potency can be increased by eliminating the broodiness behaviour of the hens in the population or by increasing the I allele frequency through a controlled mating.

Prolactin gene polymorphism in indigenous and crossbred cattle of north east India

Bovine PRL gene is considered one of the important hereditary components of milk productivity, thus seems to be an excellent candidate gene for milk production traits. The study was conducted to study the distribution pattern of allelic variants at the prolactin-RsaI locus in 210 indigenous cattle of north east India (viz. Manipur, Mizoram, Nagaland, Tripura, Asom and Meghalaya,) and crossbred cattle of Mizoram. PCR-RFLP genotyping of a 156 bp fragment of prolactin (PRL) in exon 3 revealed two different allelic variants. The predominant genotype (s) were AB (0.40) and BB (0.40) in indigenous cattle of Manipur, BB in Mizoram (0.57), AB in indigenous cattle of Nagaland (0.50) and Meghalaya (0.63). Whereas in Tripura and Asom indigenous cattle, AA was the most common genotype (0.40 and 0.60). However, AB (0.80) was markedly higher in crossbred (HF × indigenous) cattle. All the six indigenous cattle populations conforming to equilibrium indicated lack of selection pressure for PRL gene in these cattle population. The most frequent allele was A allele in the indigenous cattle of Nagaland (0.55), Tripura (0.55) and Asom (0.70) and B allele in indigenous cattle of Manipur (0.60) and Mizoram (0.73). Whereas, in Meghalaya indigenous and crossbred cattle, both alleles were present in equal frequencies. The findings of differences in the distributions of allelic and genotype variants of PRL-RsaI locus among the indigenous cattle populations of six states of north east India, suggested the presence of scope for genetic selection for milk production traits.

Measuring Mobility in Chromatin by Intensity-Sorted FCS

The architectural organization of chromatin can play an important role in genome regulation by affecting the mobility of molecules within its surroundings via binding interactions and molecular crowding. The diffusion of molecules at specific locations in the nucleus can be studied by fluorescence correlation spectroscopy (FCS), a well-established technique based on the analysis of fluorescence intensity fluctuations detected in a confocal observation volume. However, detecting subtle variations of mobility between different chromatin regions remains challenging with currently available FCS methods. Here, we introduce a method that samples multiple positions by slowly scanning the FCS observation volume across the nucleus. Analyzing the data in short time segments, we preserve the high temporal resolution of single-point FCS while probing different nuclear regions in the same cell. Using the intensity level of the probe (or a DNA marker) as a reference, we efficiently sort the FCS segments into different populations and obtain average correlation functions that are associated to different chromatin regions. This sorting and averaging strategy renders the method statistically robust while preserving the observation of intranuclear variations of mobility. Using this approach, we quantified diffusion of monomeric GFP in high versus low chromatin density regions. We found that GFP mobility was reduced in heterochromatin, especially within perinucleolar heterochromatin. Moreover, we found that modulation of chromatin compaction by ATP depletion, or treatment with solutions of different osmolarity, differentially affected the ratio of diffusion in both regions. Then, we used the approach to probe the mobility of estrogen receptor-α in the vicinity of an integrated multicopy prolactin gene array. Finally, we discussed the coupling of this method with stimulated emission depletion FCS for performing FCS at subdiffraction spatial scales.

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.

Яичная продуктивность кур пушкинской и юрловской голосистой пород и ее взаимосвязь с полиморфными вариантами гена пролактина

Яичная продуктивность кур пушкинской и юрловской голосистой пород и ее взаимосвязь с полиморфными вариантами гена пролактина

A Prl/RsaI Polymorphism in Exon 3 and 4 of Prolactin Gene in Dairy Cattle

A Prl/RsaI Polymorphism in Exon 3 and 4 of Prolactin Gene in Dairy Cattle

Genetic variation of prolactin gene in Uttara fowl of Kumaon region and its association with growth traits

Genetic variation of prolactin gene in Uttara fowl of Kumaon region and its association with growth traits

Improvement of Phaseolus vulgaris on breastfeeding in female rats

Objective: To evaluate effect of Phaseolus vulgaris (P. vulgaris) on the breastfeeding in female rats.Methods: This experimental study was done from May 2018 to December 2018 in the Anatomical Department of Medical School in Kermanshah University of Medical Sciences in Iran. In this study, after one-week adaptation and fertilization by male, 40 female rats within 20 days of pregnancy (on average, every mother had 10 newborns) were equally separated into four groups (animals were administrated after delivery of offspring). Group 1 was control group receiving normal saline interaperitoneally, and groups 2, 3, 4 were treatment groups receiving the dose of 20, 50, 100 mg/kg of P. vulgaris interaperitoneally respectively once a day for 60 days. The prolactin hormone was measured by radio immune assay, number and diameter of alveoli via histological and morphometrical examinations, and receptor prolactin gene expression by real-time polymerase chain reaction method.Results: P. vulgaris significantly improved alveoli’s number and diameter, prolactin hormone and receptor prolactin expression when compared to the control group (P<0.001).Conclusions: P. vulgaris is helpful to improve the breastfeeding parameters of rats’ mammary glands.

Single nucleotide polymorphism analysis in meat-production related genes in broiler chickens

In broiler chickens, the intensive selection for growth rate, feed efficiency, body composition (breast muscle weight) traits in the last decades was successful. To improve economically important characteristics, it is possible to use molecular markers associated with meat production traits. The aim of this study was to examine genotype polymorphisms in ROSS 308 broilers for thyroid hormone responsive Spot14α, insulinlike growth factor 1 (IGF1), IGF-binding protein 2 (IGFBP2), somatostatin (SST) and prolactin (PRL) genes. A further goal of this investigation was to study the relationship between the polymorphisms and phenotypic characteristics.&#x0D; In the investigated broiler population, the frequency for CC homozygous genotype was 0.77 in Spot14α (AY568628), AA homozygous genotype was 0.80 in IGF1 (M74176), GG homozygous genotype was 0.85 in IGFBP2 (U15086), DD homozygous genotype was 0.60 in PRL (FJ663023 or FJ434669). Only the AA homozygous genotype was found in SST (X60191). Chickens with AC genotype in Spot14α, and with GG genotype in IGFBP2 had higher body weight (BW) and carcass weight (CW), compared to CC and GT genotypes. However, the differences were not significant (P&gt;0.05). There was significant association (P&lt;0.05) between PRL genotypes and body and carcass weight, where chicken with homozygous DD surpassed individuals with homozygous II genotypes.

ASSOCIATION OF PROLACTIN GENE POLYMORPHISM WITH EGG PRODUCTION TRAIT IN LIEN MINH CHICKEN

Lien Minh chicken is an indigenous breed with several favorable properties, such as good meat quality and associated with the economic development of the people in the Lien Minh village, Cat Hai, Hai Phong. Chicken prolactin is a candidate gene involved in egg production trait. The aim of this study was to investigate the association of single nucleotide polymorphism (SNP) of Prolactin gene (PRL) concerning with reproductive trait in Lien Minh chicken. All hens were kept in individual laying cage and phenotypic data for total 90 hens were recorded as following: age at first egg, first egg’s weight, number of eggs, eggs’ weight, and eggs’ shape index. Blood samples were used for DNA extraction and then genotyping by PCR-RFLP method. Two SNPs of PRL gene were analyzed including: 24 bp indel in promoter (PRL24) and C2402T in 5' flanking region (PRL5). The polymorphic sites at PRL24 and PRL5 showed two genotypes with the frequency of ID (0.27), DD (0.73) and CT (0.41), TT (0.59), respectively. Allele frequencies of PRL24 and PRL5 were: I (0.13), D (0.87) and C (0.21), T (0.79), respectively. Genotypes ID PRL24 and CT at PRL5 showed the highest values of mean weight of eggs 47.57 ± 3,11 g and 46.91 ± 4.29 g, respectively (P &lt; 0.05). In addition, the individual with genotype ID, CT also gave better reproductive characteristics such as number of eggs, first egg’s weight. These results suggested that the beneficial alleles/genotypes could be used to support the improvement of the egg production capacity in the breeding of Lien Minh chicken.

Identification of genetic marker for prolactin gene related to milk yield in pedigreed Sahiwal population

The study was conducted to identify the genetic marker for prolactin gene related to milk yield in pedigreed Sahiwal population. PCR-RFLP was carried out by using Rsa1 restriction endonuclease and revealed three genotypes GG, GA and AA in Sahiwal population. The frequency of G and A allele of Prolactin gene (exon 3) was estimated as 0.575 and 0.425, whereas the frequencies of GG, GA and AA genotypes for Prolactin gene (exon 3) were estimated as 0.45, 0.25 and 0.30, respectively. Regression analysis inferred that, SNP (G55A) of prolactin gene (exon 3) contributed to an increase in the average test day milk yield by 321.5g which was 3.92% of the average test day milk yield. Forty Sahiwal animals (20 families) under Daughter- Dam Design were screened and it was observed that one SNP (G55A) was found in all the nine families (45%) having heterozygous (GA) genotype. Thus, it could be considered as a potential genetic marker for high milk production in Sahiwal population.

Significant associations between prolactin gene polymorphisms and growth traits in the channel catfish (Ictalurus punctatus Rafinesque, 1818) core breeding population

The aim of this study was to analyze the effects of the prolactin gene (PRL) on growth traits in channel catfish (Ictalurus punctatus). We sequenced pooled DNA samples from channel catfish to identify single nucleotide polymorphisms (SNPs) in the PRL gene. The identified SNPs were then genotyped in 176 individuals from the G1 core breeding population of catfish using the multiplex SNaPshot method. SNP genotypes and diplotypes of the PRL gene and their associations with body mass and body length traits were also examined. Seven SNPs (g.951A>C, g.1293A>T, g.1359C>T, g.1374A>G, g.1620C>T, g.2675C>T, and g.3369A>G) were identified in introns and noncoding regions, and five of these were successfully genotyped. Seven haplotypes and ten diplotypes occurred in the group of 176 catfish. Association analyses showed that the g.1620 C>T SNP was significantly associated with growth traits in channel catfish, such that individuals with the TT genotype had significantly greater body weights and body lengths than those with CC or CT genotypes (P < 0.05). In addition, the g.3369 A>G SNP showed a significant negative association with growth traits, such that fish with the GG genotype had lower values for both growth traits compared with fish with genotype AA or AG (P < 0.05). Diplotypes H3/H3 and H4/H7 were associated with significantly higher body mass and body length than the other diplotypes (P < 0.01). The results of this study may provide basic data for marker-assisted selection and quantitative trait loci analysis of growth traits in channel catfish.

Role of kisspeptin and Kiss1R in the regulation of prolactin gene expression in rat somatolactotroph GH3 cells.

Hypothalamic kisspeptin is a known principal activator of gonadotropin-releasing hormone neurons and governs the hypothalamic-pituitary-gonadal axis. Previous reports have shown that kisspeptin is also released into the hypophyseal portal circulation and directly affects the anterior pituitary. In this study, we examined the direct effect of kisspeptin on pituitary prolactin-producing cells. The rat pituitary somatolactotroph cell line GH3 expresses the kisspeptin receptor (Kiss1R); however, in these cells, kisspeptin failed to stimulate prolactin-promoter activity. When GH3 cells overexpressed Kiss1R, kisspeptin clearly increased prolactin-promoter activity, with a concomitant increase in extracellular signal-regulated kinase (ERK) and cAMP/protein kinase A (PKA) signaling pathways. In the experiments using GH3 cells overexpressing Kiss1R, kisspeptin did not potentiate thyrotropin-releasing hormone (TRH)-induced prolactin-promoter activity, but it potentiated the pituitary adenylate cyclase-activating polypeptide-induced prolactin-promoter activity, with a concomitant enhancement of ERK and PKA signaling pathways. Although the basal and TRH-induced prolactin-promoter activities were not modulated by increasing amounts of Kiss1R expression in GH3 cells, kisspeptin-stimulated prolactin-promoter activity was increased by the amount of Kiss1R overexpression. Endogenous Kiss1r mRNA expression in GH3 cells was significantly increased by treatment with estradiol (E2) but not by TRH. In addition, kisspeptin's ability to stimulate prolactin-promoter activity was restored after E2 treatment in non-transfected GH3 cells. Our current observations suggest that kisspeptin might have a direct effect on prolactin expression in the anterior pituitary prolactin-producing cells under the influence of E2, which may regulate Kiss1R expression and function.