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Glucocorticoids (GCs) and adrenocorticotropic hormone (ACTH) are major components of the classic endocrine stress response. Free-living vertebrates are characterized by circannual changes in the baseline and/or stress-induced secretion of GCs and ACTH. In mammalian species, GC and ACTH levels vary seasonally but there is no consensus to the season in which animals have elevated GC and ACTH levels. The aim of our study was to determine, for the first time, the type and amount of glucocorticoids produced in free-living beaver (Castor fiber L.)--the largest rodent in Eurasia, and to find out whether stress-induced plasma GC and ACTH levels show seasonal variations. Blood samples were obtained from animals under general anesthesia in April (pregnancy in females), July (offspring rearing) and November (preparing for the winter). The adrenals of beavers produce both cortisol and corticosterone, and plasma cortisol levels were higher than corticosterone. In the current experiment, plasma cortisol concentrations in beavers were affected by the season. The highest stress-associated cortisol levels were noted in males in July during offspring rearing. Corticosterone and ACTH concentrations in beavers remained generally constant, regardless of the season and sex. In conclusion, seasonal changes were observed only in relation to stress-induced plasma cortisol levels in the beaver.

Orexin A and B are hypothalamic peptides derived from the prepro-orexin (PPO) precursor. Orexins stimulate food intake and arousal. Those peptides bind and activate two G protein-coupled receptors: orexin receptor 1 (OX1R) and orexin receptor 2 (OX2R). Numerous authors have suggested that orexins play an important role in the regulation of the reproductive functions. The objective of the present study was to analyse the presence of and changes in the gene and protein expression pattern of the orexin system in the porcine uterus, conceptus and trophoblast (chorioallantois) during early pregnancy. In the endometrium, the highest PPO and OX1R gene expression was detected on days 15 to 16 of gestation. The OX2R mRNA content in the endometrium was higher on days 10 to 11 and 15 to 16 than on days 12 to 13 and 27 to 28. In the trophoblasts, PPO gene expression was higher on days 30 to 32 than on days 27 to 28. The highest PPO protein content in the endometrium was noted on days 12 to 13. The highest OX1R protein content in the endometrium was detected on days 10 to 11, whereas OX2R protein on days 15 to 16. In the trophoblasts, PPO and OX1R protein levels were more pronounced on days 27 to 28 than on days 30 to 32, but OX2R expression was higher on days 30 to 32. The expression of PPO, OX1R and OX2R was different in the conceptuses and trophoblasts during early pregnancy. Local orexin production and the presence of the specific orexin receptors suggest that the orexin system may participate in the control of porcine reproductive functions by exerting endocrine and auto/paracrine effects on the uterus, conceptuses and trophoblasts during early pregnancy. This study provides the first evidence for the presence of orexins and their receptors in the uteri, conceptuses and trophoblasts in pigs during early pregnancy. The local orexin system is dependent on the stage of pregnancy.

The European beaver (Castor fiber L.) is an important free-living rodent that inhabits Eurasian temperate forests. Beavers are often referred to as ecosystem engineers because they create or change existing habitats, enhance biodiversity and prepare the environment for diverse plant and animal species. Beavers are protected in most European Union countries, but their genomic background remains unknown. In this study, gene expression patterns in beaver testes and the variations in genetic expression in breeding and non-breeding seasons were determined by high-throughput transcriptome sequencing. Paired-end sequencing in the Illumina HiSeq 2000 sequencer produced a total of 373.06 million of high-quality reads. De novo assembly of contigs yielded 130,741 unigenes with an average length of 1,369.3 nt, N50 value of 1,734, and average GC content of 46.51%. A comprehensive analysis of the testicular transcriptome revealed more than 26,000 highly expressed unigenes which exhibited the highest homology with Rattus norvegicus and Ictidomys tridecemlineatus genomes. More than 8,000 highly expressed genes were found to be involved in fundamental biological processes, cellular components or molecular pathways. The study also revealed 42 genes whose regulation differed between breeding and non-breeding seasons. During the non-breeding period, the expression of 37 genes was up-regulated, and the expression of 5 genes was down-regulated relative to the breeding season. The identified genes encode molecules which are involved in signaling transduction, DNA repair, stress responses, inflammatory processes, metabolism and steroidogenesis. Our results pave the way for further research into season-dependent variations in beaver testes.

Female reproductive success is closely associated with nutritional status and energy balance. In this context, adiponectin appears to be a key hormone connecting reproductive system function and metabolism regulation. It is hypothesized that adiponectin expression in the pituitary depends on the phase of the estrous cycle. The effect of adiponectin on luteinizing hormone (LH) and follicle-stimulating hormone (FSH) secretion is also postulated. Changes in the adiponectin gene and protein expression in the porcine anterior (AP) and posterior (NP) pituitaries as well as the effect of in vitro administration of adiponectin on basal and gonadotropin-releasing hormone (GnRH)- and/or insulin-stimulated LH and FSH secretion were investigated on days 2–3, 10–12, 14–16, and 17–19 of the estrous cycle. Adiponectin gene was more pronounced on days 2–3 in AP but on days 10–12 in NP. Protein concentration in AP was the highest on days 10–12 and in NP on days 10–12 and 17–19 of the cycle. In vitro, adiponectin did not affect basal LH secretion but increased FSH release by AP cells. Adiponectin administration affected GnRH- and/or insulin-induced LH and FSH output in a manner dependent on the phase of the estrous cycle. In this study we indicated for the first time adiponectin expression in the porcine AP and NP that was dependent on the phase of the estrous cycle. In vitro studies indicated that adiponectin may affect gonadotropin secretion. The above suggests that the studied adipokine may influence female reproductive functions via its effect on LH and FSH secretion by gonadotrophs, but the cellular mechanism of its action remains unknown.

Adiponectin, one of the several adipocytokines secreted mainly by the adipose tissue, plays an important role in regulating energy homeostasis and controls female fertility. Female reproductive functions are closely associated with nutritional status, and adiponectin seems to be an important factor linking the regulation of metabolic homeostasis with reproductive processes. The biological activity of adiponectin is mediated by two distinct receptors, adiponectin receptor 1 (AdipoR1) and adiponectin receptor 2 (AdipoR2). The objective of this study was to determine the presence of and changes in the gene and protein expression pattern of adiponectin and its receptors in the porcine uterus during early pregnancy and on Days 10 to 11 of the estrous cycle and in the conceptus and trophoblast. The highest level of adiponectin transcript was observed on Days 15 to 16 of gestation, Days 10 to 11 of the cycle in the endometrium, and Days 15 to 16 of gestation in the myometrium. The highest expression of AdipoR1 and AdipoR2 genes was detected on Days 10 to 11 of gestation in the endometrium, and Days 12 to 13 in the myometrium. The highest content of adiponectin protein was noted on Days 12 to 13 and 30 to 32 of gestation in the endometrium and Days 10 to 11 of the cycle in the myometrium. The expression of adiponectin protein was higher on Days 27 to 28 and 30 to 32 in the conceptuses. AdipoR1 protein content in the myometrium was highest on Days 12 to 13 and 30 to 32. In contrast, in the endometrium, it was more constant. The highest content of AdipoR2 protein was detected on Days 15 to 16 and 30 to 32 of gestation, Days 10 to 11 of the cycle in the endometrium, and Days 10 to 11 of gestation in the myometrium. In the conceptuses, the highest AdipoR1 protein content was observed on Days 15 to 16, and the highest AdipoR2 protein expression was determined on Days 15 to 16 and 27 to 28. In the trophoblasts, AdipoR1 protein content was higher on Days 27 to 28 than on Days 30 to 32, whereas the expression of AdipoR2 was higher on Days 30 to 32. This study demonstrated the presence of adiponectin and its receptors in the uteri, conceptuses, and trophoblasts of pregnant pigs and that the local adiponectin system is dependent on the stage of pregnancy.

The aim of this study was to compare the expression levels of adiponectin receptor 1 and adiponectin receptor 2 mRNAs and proteins in porcine ovaries during four stages (days 2 to 3, 10 to 12, 14 to 16, 17 to 19) of the oestrous cycle and to measure adiponectin plasma concentrations during the same phases of the cycle. Higher mRNA expression of adiponectin receptor 1 was detected in porcine granulosa cells than in corpora lutea and theca cells (P < 0.01). In contrast, higher gene expression of adiponectin receptor 2 occurred in newly developed and mature corpora lutea (P < 0.01). The adiponectin receptor 1 protein content was the highest in corpora lutea isolated on days 2 to 3 of the cycle and was the lowest in theca interna cells (P < 0.01). The profile of adiponectin receptor 2 protein was similar to that of adiponectin receptor 1. Adiponectin plasma concentrations were significantly higher throughout the luteal phase than in the follicular phase (P < 0.01). In conclusion, the presence of adiponectin receptor 1 and adiponectin receptor 2 mRNAs and proteins in the porcine ovary suggests that adiponectin may directly affect ovarian functions through its own specific receptors. The expression of both receptors and adiponectin plasma concentration were dependent on hormonal status related to the stage of the cycle.

Orexins A and B are hypothalamic neuropeptides associated with homeostasis and the reproductive system. The aim of the study was to compare the expression of the prepro-orexin gene and the intensity of orexins immunoreactivity in the porcine ovary (corpora lutea, granulosa and theca interna cells) during four different stages of the oestrous cycle (days: 2-3, 10-12, 14-16 and 17-19) and to examine the in vitro effect of orexins on the secretion of steroid hormones by porcine luteal, granulosa and theca interna cells. The highest expression of prepro-orexin mRNA was observed in theca interna cells on days 17-19 of the oestrous cycle. The highest content of immunoreactive orexin A was noted in corpora lutea on days 10-12 and the highest level of immunoreactive orexin B on days 14-16 of the cycle. Immunoreactive orexin A concentrations were higher in theca interna cells than in granulosa cells, whereas similar levels of immunoreactive orexin B were observed in both cell types. Under in vitro conditions, at the concentration of 10 nM, orexins A and B inhibited FSH-induced oestradiol secretion by granulosa cells. The obtained results suggest that the pattern of orexin peptide expression in the porcine ovary is related to the animals' hormonal status. Our findings imply that orexins can affect porcine reproductive functions through modulation of ovarian steroidogenesis.

Adiponectin is a hormonal link between obesity and reproduction, and its actions are mediated by two types of receptors: adiponectin receptor 1 (AdipoR1) and adiponectin receptor 2 (AdipoR2). This study compares the expression levels of adiponectin and adiponectin receptor mRNAs and proteins in selected areas of the porcine hypothalamus responsible for GnRH production and secretion: the mediobasal hypothalamus (MBH), pre-optic area (POA) and stalk median eminence (SME). The tissue samples were harvested on days 2-3, 10-12, 14-16 and 17-19 of the oestrous cycle. Adiponectin mRNA expression in MBH was significantly lower on days 14-16, whereas in SME, the most pronounced gene expression was found on days 2-3 of the cycle (p

Hypothalamic peptides orexin A (OXA) and orexin B (OXB) are derived from the proteolytic cleavage of a common precursor molecule, prepro-orexin (PPO). They act via two orexin receptors (OX1R and OX2R), which belong to the G-protein coupled receptor superfamily. Orexins are implicated in the regulation of arousal states, energy homeostasis and reproductive neuroendocrine function. The objective of this study was to investigate the presence and changes in orexin expression in the porcine pituitary during the estrous cycle. Adenohypophysis (AP) and neurohypophysis (NP) tissue samples were harvested on days 2 to 3, 10 to 12, 14 to 16, and 17 to 19 of the estrous cycle. The expression of the PPO gene increased in AP and NP during the estrous cycle. The highest PPO protein concentrations in AP were reported on days 2 to 3 (P

Chemerin (CHEM) may act as an important link integrating energy homeostasis and reproductive functions of females, and its actions are mediated by three receptors: chemokine-like receptor 1 (CMKLR1), G protein-coupled receptor 1 (GPR1), and C-C motif chemokine receptor-like 2 (CCRL2). The aim of the current study was to compare the expression of CHEM and its receptor (CHEM system) mRNAs (quantitative real-time PCR) and proteins (Western blotting and fluorescent immunohistochemistry) in the selected areas of the porcine hypothalamus responsible for gonadotropin-releasing hormone production and secretion: the mediobasal hypothalamus, preoptic area and stalk median eminence during the oestrous cycle and early pregnancy. Moreover, plasma CHEM concentrations were determined using ELISA. The expression of CHEM system has been demonstrated in the porcine hypothalamus throughout the luteal phase and follicular phase of the oestrous cycle, and during early pregnancy from days 10 to 28. Plasma CHEM levels and concentrations of transcripts and proteins of CHEM system components in the hypothalamus fluctuated throughout pregnancy and the oestrous cycle. Our study was the first experiment to demonstrate the presence of CHEM system mRNAs and proteins in the porcine hypothalamus and the correlations between the expression levels and physiological hormonal milieu related to the oestrous cycle and early pregnancy.

Comprehensive understanding of the regulatory mechanism of the implantation process in pigs is crucial for reproductive success. The endometrium plays an important role in regulating the establishment and maintenance of gestation. The goal of the current study was to determine the effect of adiponectin on the global expression pattern of genes and relationships among differentially expressed genes (DE-genes) in the porcine endometrium during implantation using microarrays. Diverse transcriptome analyses including gene ontology (GO), biological pathway, networks, and DE-gene analyses were performed. Adiponectin altered the expression of 1286 genes with fold-change (FC) values greater than 1.2 (p &lt, 0.05). The expression of 560 genes were upregulated and 726 downregulated in the endometrium treated with adiponectin. Thirteen genes were selected for real-time PCR validation of differential expression based on a known role in metabolism, steroid and prostaglandin synthesis, interleukin and growth factor action, and embryo implantation. Functional analysis of the relationship between DE-genes indicated that adiponectin interacts with genes that are involved in the processes of cell proliferation, programmed cell death, steroid and prostaglandin synthesis/metabolism, cytokine production, and cell adhesion that are critical for reproductive success. The presented results suggest that adiponectin signalling may play a key role in the implantation of pig.

Leptin is a polypeptide hormone produced primarily by adipocytes. It has been implicated in the regulation of satiety and energy homeostasis. Leptin has been suggested to play a role in reproduction based on its involvement in the regulation of the hypothalamic–pituitary–gonadal axis via endocrine, paracrine and/or autocrine pathways. The aim of the present study was to localize the cellular distribution of leptin and the long isoform of leptin receptor (OB-Rb) genes in porcine ovarian antral follicles and to compare the expression levels of leptin and OB-Rb mRNAs in porcine granulosa cells (GC), theca interna (TIC) and theca externa (TEC) cells during the luteal phase of the estrous cycle and in early pregnancy. The expression of leptin and OB-Rb genes was detected in GC, TIC and TEC. Significantly higher levels of leptin gene expression in GC were observed during the mid- and late-luteal phases of the cycle than on days 30 to 32 of pregnancy. On days 14 to 16 of pregnancy, leptin mRNA expression was higher than that on days 14 to 16 of the cycle. The expression of the OB-Rb gene in GC and TEC increased during pregnancy in comparison with the analyzed luteal phases of the cycle. Our results validate the hypothesis that locally produced leptin plays a role in the regulation of porcine reproduction at the ovarian level and exerts a direct effect on porcine follicles. The differences in OB-Rb gene expression in porcine GC and theca cells also suggest that their sensitivity to leptin varies in the ovaries of pregnant and cyclic pigs.

Orexin A (OXA) is a hypothalamic neuropeptide known for its role in the regulation of food intake and arousal. It is also considered as a link between energy homeostasis and reproduction. Nevertheless, very little is known on the role of this peptide in the uterus. The objective of this study was to investigate OXA effect on oestradiol (E 2 ), oestrone (E 1 ), and testosterone (T) secretion by porcine endometrial and myometrial explants and gene expression of key steroidogenic enzymes involved in steroid production, namely cytochrome P450c17 ( CYP17A1 ) and cytochrome P450 aromatase ( CYP19A3 ), on days 10–11, 12 to 13, 15 to 16 and 27 to 28 of pregnancy and on days 10–11 of the cycle. In endometrium, OXA increased E 1 secretion on days 10–11 and 15 to 27 of gestation, and T release on days 12–13. A decrease in E 2 , E 1 and T secretion was noted on days 27–28, 12 to 13 and 10 to 11 of gestation, respectively. OXA enhanced CYP17A1 and CYP19A3 expression on days 15–28 of pregnancy, whereas decreased their expression on days 10–13. In the myometrium, OXA increased E 1 secretion on days 10–16 of pregnancy, whereas inhibited the release of E 2 and T on days 10–11. CYP17A1 and CYP19A3 genes expression was enhanced on days 27–28 and 12 to 13 of pregnancy, respectively. The expression of both genes was suppressed on days 10–11 and 15 to 16 of pregnancy (P via its influence on steroidogenesis, may play a regulatory role in the uterus.

Studies have demonstrated that adiponectin could be a link between reproductive functions and energy metabolism in animals. The aim of the present study was to investigate the effects of prostaglandin (PG) E

Orexin A and B, also termed hypocretin 1 and 2, are associated with the stimulation of food intake and arousal. The biological actions of the hormones are mediated via two distinct G protein-coupled receptors, termed orexin receptor 1 (OX1R) and orexin receptor 2 (OX2R). OX1R is selective for orexin A and OX2R binds orexin A and orexin B with similar affinity. The present study analyzed mRNA and protein expressions of OX1R and OX2R in adenohypophysis (AP) and neurohypophysis (NP) of cycling pigs. The tissue samples were harvested on days 2-3, 10-12, 14-16, and 17-19 of the oestrous cycle. Using quantitative real-time PCR higher OX1R gene expression was detected in AP on days 2-3 relative to days 10-12, 14-16 and 17-19 (p0.05). In NP the OX1R mRNA level was elevated on days 10-12 compared to the remaining stages (p0.05). OX2R gene expression in AP was the lowest on days 10-12 (p0.05 compared to days 2-3 and 17-19) and the expression peak occurred on days 17-19 (p0.05 vs. the all studied stages). In NP the highest (p0.05) expression of OX2R mRNA was noted on days 17-19 in relation to the remaining periods. OX1R protein content in AP was greatest on days 10-12 (p0.05), whereas in NP it was greatest on days 2-3 and 14-16 (p0.05 vs. days 10-12 and 17-19). In both cases the lowest OX1R protein expression was observed during follicular phase (p0.05 in relation to three remaining studied stages). OX2R protein in AP was lower (p0.05) on days 2-3 and 14-16 compared to days 10-12 and 17-19. In NP the lowest (p0.05) expression of this protein was on days 17-19 and the highest on days 10-12 (p0.05 compared to days 2-3 and 17-19). In summary, the present findings provide the first evidence that OX1R and OX2R mRNAs and proteins occur in the pituitary of the pig and indicate the dependence of orexin receptor expression on the endocrine reproductive state.

Contents In this study, we examined the levels of leptin and OB-Rb protein expression in the discrete areas of the porcine hypothalamus (mediobasal hypothalamus – MBH, pre-optic area – POA, stalk median eminence – SME) during mid- and late-luteal phases of the oestrous cycle (days 10–12 and 14–16) as well as two stages of pregnancy (days 14–16 and 30–32). The analysis showed that during the cycle, leptin protein expression in MBH was higher in the mid-luteal phase than late-luteal phase. In the case of OB-Rb protein expression, a higher level was observed in MBH during the late-luteal phase in comparison to the mid-luteal phase, whereas in POA and SME the opposite dependence was noticed. In turn, during pregnancy, leptin protein expression in MBH and POA, and OB-Rb protein expression in POA were more pronounced on days 14–16 than on days 30–32. In contrast, leptin protein content in SME as well as OB-Rb protein in MBH and SME was higher on days 30–32 than during the earlier stage of pregnancy. Comparison of leptin and OB-Rb protein expression between the cycle (days 10–12) and pregnancy showed a higher level of leptin and OB-Rb protein contents in POA as well as in SME during pregnancy (on days 14–16 and 30–32, respectively). Yet, OB-Rb protein expression in POA on days 30–32 of pregnancy was lower in comparison to days 10–12 of the cycle. Furthermore, during pregnancy, leptin protein expression in MBH was lower (days 14–16 and 30–32), whereas OB-Rb protein expression in that area of hypothalamus was higher (days 30–32) in comparison to the mid-luteal phase. Our results indicate that both leptin and OB-Rb are synthesized in the porcine hypothalamus and suggest the participation of leptin in auto/paracrine regulation of these brain areas functions, including control of reproduction during the oestrous cycle and early pregnancy.

Leptin, the product of the OB gene, is a 16-kDa polypeptide of 146 amino acid residues produced mainly by adipocytes that regulates metabolism and reproduction. The actions of leptin are mediated mainly via the long form of the leptin receptor (OB-Rb). The identification of leptin and OB-Rb mRNAs and proteins in human and mouse endometrium, and placental trophoblast suggests that leptin may be involved in the implantation process. Thus, the aim of this study was to compare the expression levels of porcine OB-Rb mRNA and protein in the endometrium and myometrium during mid- and late-luteal phases of the oestrous cycle (days 10–12 and 14–16, respectively) as well as during two stages of pregnancy respondent to the beginning of the implantation process (days 14–16) and the post-implantation period (days 30–32), and in trophoblast during both periods of pregnancy. OB-Rb gene expression in endometrium during the examined stages of pregnancy and the mid- and late-luteal phases of the cycle was at the same level. In contrast, in myometrium leptin receptor gene expression decreased on days 14–16 of pregnancy compared to both phases of the cycle, and on days 30–32 of pregnancy in relation to late-luteal phase. OB-Rb protein expression in the tissues was lower during the examined stages of pregnancy in comparison to the mid- and late-luteal phases of the cycle. In trophoblast, OB-Rb mRNA and protein expression was higher on days 30–32 than during days 14–16 of pregnancy. In conclusion, our results might suggest that leptin can participate in the control of pig reproduction by exercising its action at the uterine and trophoblast level and have a direct effect on these organ during both the luteal phase of the cycle and early pregnancy. Moreover, changes in OB-Rb gene and protein expression in tissues of pig reproductive tract strongly suggest that their sensitivity to leptin varies throughout luteal phase of the cycle and early pregnancy.

Reproductive functions are closely related to nutritional status. Recent studies suggest that adiponectin may be a hormonal link between them. Adiponectin is an adipocytokine, abundantly expressed in adipose tissues. It plays a dominant role in lipid and carbohydrate metabolism by stimulating fatty acid oxidation, decreasing plasma triglycerides, and increasing cells' sensitivity to insulin and has direct antiatherosclerotic effects. The hormone is also postulated to play a modulatory role in the regulation of the reproductive system. The aim of this study was to identify differentially expressed genes (DE-genes) in response to adiponectin treatment of porcine luteal ovarian cells. The global expression of genes in the porcine ovary was investigated using the Porcine (V2) Two-color gene expression microarray, 4 × 44 (Agilent, USA). Analysis of the microarray data showed that 701 genes were differentially expressed and 389 genes showed a fold change greater than 1.2 (p

Studies have demonstrated that adiponectin could be a link between reproductive functions and energy metabolism in animals. The aim of the present study was to investigate the effects of prostaglandin (PG) E2 and PGF2α (10, 50, 100, 250 and 500 ng mL–1) on the expression and secretion of adiponectin and its receptor genes and proteins by cultured in vitro porcine endometrial and myometrial tissues on Days 10–28 of pregnancy and Days 10–11 of the oestrous cycle. The gene expression was analysed using the real-time PCR method. Adiponectin protein secretion was determined by ELISA, whereas the receptors proteins content was defined using Western Blot analysis. Both PGE2 and PGF2α modulated the expression of adiponectin system genes and proteins in the uterus during early pregnancy. PGE2 and PGF2α had similar effects on the adiponectin system, which differed between the stages of gestation and between pregnancy and the oestrous cycle. On Days 10–11 of gestation, PGE2 and PGF2α generally increased adiponectin secretion by endometrial and myometrial tissues. Both PGs decreased levels of endometrial adiponectin receptor type 1 (AdipoR1), whereas only PGF2α decreased myometrial levels of AdipoR1. Both PGs increased myometrial adiponectin receptor type 2 (AdipoR2) levels. On Days 12–13 of gestation, PGE2 decreased AdipoR1 concentrations in both tissues and AdipoR2 levels in the endometrium. PGF2α decreased myometrial concentrations of both receptors. On Days 15–16 of gestation, both PGE2 and PGF2α increased concentrations of AdipoR1 and AdipoR2 in the endometrium and myometrium. PGE2 stimulated the secretion of adiponectin in the endometrium, but not in the myometrium. On Days 27–28 of pregnancy, both PGE2 and PGF2α inhibited the expression of AdipoR1 and AdipoR2 in endometrial and myometrial tissues and decreased the secretion of endometrial adiponectin. Both PGE2 and PGF2α had tissue-specific and dose-dependent effects on the adiponectin system.

Adiponectin is an adipose-secreted hormone that regulates energy homeostasis and is also involved in the control of the reproductive system. The goal of the present study was to investigate changes in adiponectin gene and protein expression in porcine ovarian structures during the oestrous cycle and to examine the effects ofin vitroadministration of adiponectin on basal and gonadotrophin- and/or insulin-induced secretion of ovarian steroid hormones. Both gene and protein expression of adiponectin were enhanced during the luteal phase of the cycle. Adiponectin affected basal secretion of progesterone by luteal cells, oestradiol by granulosa cells, and testosterone by theca interna cells. The gonadotrophin/insulin-induced release of progesterone from granulosa and theca interna cells and the release of oestradiol and androstenedione from theca cells was also modified by adiponectin. In conclusion, the presence of adiponectin mRNA and protein in the porcine ovary coupled with our previous results indicating adiponectin receptors expression suggest that adiponectin may locally affect ovarian functions. The changes in adiponectin expression throughout the oestrous cycle seem to be dependent on the hormonal status of pigs related to the stage of the oestrous cycle. The effect of adiponectin on ovarian steroidogenesis suggests that this adipokine influences reproductive functions in pigs.

Adiponectin is a hormone secreted primarily by white adipose tissue. Recent studies have shown that adiponectin and its receptors (AdipoR1 and AdipoR2) are expressed in different reproductive tissues, including the ovary and uterus. This newly discovered endocrine system plays an important role in the regulation of reproductive processes. The expression of the adiponectin system in the porcine uterus during the oestrous cycle has not been researched to date. The aim of the present study was to investigate the presence and changes in adiponectin system expression in the porcine uterus on days 2–3, 10–12, 14–16, and 17–19 of the oestrous cycle. The expression of the adiponectin gene was highest on days 14–16 and 2–3 in the endometrium and myometrium, respectively. In the endometrium, the content of AdipoR1 and AdipoR2 mRNAs was highest on days 10–12, whereas significantly higher expression levels of both genes were noted in the myometrium on days 17–19. The highest content of adiponectin and AdipoR1 protein in the endometrium was reported on days 2–3. In the myometrium, the expression levels of both receptor proteins were significantly higher on days 17–19. Adiponectin system proteins were localized in endometrial epithelial glandular cells, luminal epithelial cells and stromal cells as well as in longitudinal and circular muscles of the myometrium. This study demonstrated the presence of adiponectin, AdipoR1 and AdipoR2 genes and proteins in the porcine uterus and the effect of the stage of the oestrous cycle on the expression of the adiponectin system. Our results suggest that locally synthesized adiponectin directly affects uterine functions.

Leptin is a polypeptide hormone produced predominantly in adipocytes. It has been found to be implicated in the regulation of satiety and energy homeostasis. A role for leptin in reproduction was later suggested by findings that this hormone may be involved in the regulation of the hypothalamic- pituitary-gonadal axis via endocrine, paracrine and/or autocrine pathways. The objective of the study was to investigate the ontogeny of the long isoform of leptin receptor (OB-Rb) gene in porcine ovarian follicles. The expression of OB-Rb gene was detected in porcine primordial, primary, secondary and antral follicles by in situ hybridization. In summary, our data suggest that leptin might have a direct effect on porcine follicles and plays an important role in the follicular development.

Background Adiponectin, protein secreted mainly by white adipose tissue, is an important factor linking the regulation of metabolic homeostasis and reproductive processes. The biological activity of the hormone is mediated via two distinct receptors, termed adiponectin receptor 1(AdipoR1) and adiponectin receptor 2 (AdipoR2). The present study analyzed mRNA and protein expression of AdipoR1 and AdipoR2 in the anterior (AP) and posterior (NP) pituitary of cyclic pigs. Methods The total of 20 animals was assigned to one of four experimental groups (n = 5 per group) as follows: days 2–3 (early-luteal phase), 10–12 (mid-luteal phase), 14–16 (late-luteal phase), 17–19 (follicular phase) of the oestrous cycle. mRNA and protein expression were analyzed using real-time PCR and Western Blot methods, respectively. Results The lowest AdipoR1 gene expression was detected in AP on days 10–12 relative to days 2–3 and 14–16 (p Conclusions Our study was the first experiment to demonstrate that AdipoR1 and AdipoR2 mRNAs and proteins are present in the porcine pituitary and that adiponectin receptors expression is dependent on endocrine status of the animals.

Orexin A (OXA) and orexin B (OXB) are neuropeptides synthesized mainly in the lateral hypothalamus, which are involved in the control of various physiological functions such as energy homeostasis, sleep, wakefulness and feeding behavior. The present study analyzes orexins A and B levels in the porcine plasma during the estrous cycle. The highest plasma concentrations of orexin A were observed on days 2-3 of the estrous cycle (p0.05 relative to days 10-12 and 14-16) and the lowest (p0.05) on days 14-16. The highest orexin B levels in the blood plasma were noted on days 17-19 (p0.05 vs. days 14-16). We demonstrated the presence of OXA and OXB in porcine blood plasma and the impact of the phase of the estrous cycle on the observed changes in plasma orexin levels.

SUMMARY The presence of orexins and their receptors in gonads indicate that these hormones participate in the control of reproductive functions. The aim of the study was to compare the expression of the prepro-orexin (PPO) gene in porcine endometrium and myometrium and the intensity of OXA-and OXB-immunoreactivity in the following uterine structures: endometrial glandular and luminal epithelium and stroma as well as the myometrial longitudinal and circular muscles during the four stages (days 2–3, 10–12, 14–16, 17–19) of the estrous cycle. The highest expression of PPO mRNA was observed in the endometrium and the myometrium on days 14–16 of the cycle. The expression of the PPO gene on days 2–3 was more pronounced in the myometrium than in the endometrium, whereas on days 17–19 the gene expression was markedly higher in the endometrium. The OXA signal intensity was highest on days 2–3 in the luminal epithelium and on days 2–3 and 10–12 in the stroma. In circular muscles of the myometrium, the highest immunoreactivity was found on days 2–3 and 10–12, while in longitudinal muscles on days 2–3. OXB-immunoreactivity was highest on days 10–12 in longitudinal muscles, on days 17–19 in glandular epithelium and stroma, and on days 10–12 and 14–16 in luminal epithelium. Our results suggest that orexin A and B are produced in the porcine uterus and that their release is dependent on the hormonal status of animals.

Orexin receptors type 1 (OX1R) and type 2 (OX2R) are G protein-coupled receptors whose structure is highly conserved in mammals. OX1R is selective for orexin A, and OX2R binds orexin A and orexin B with similar affinity. Orexin receptor expression was observed in human, rat, porcine, sheep as well as Xenopus laevis pituitaries, both in the adenohypophysis and in the neurohypophysis. The expression level is regulated by gonadal steroid hormones and GnRH. The majority of orexins reaching the pituitary originate from the lateral hypothalamus, but due to the presence of the receptors and the local production of orexins in the pituitary, orexins could deliver an auto/paracrine effect within the gland. Cumulative data indicate that orexins are involved in the regulation of LH, GH, PRL, ACTH, and TSH secretion by pituitary cells, pointing to orexins' effect on the functioning of the endocrine axes. Those hormones may also serve as a signal linking metabolic status with endocrine control of sleep, arousal, and reproduction processes.

Orexin receptors type 1 (OX1R) and type 2 (OX2R) are G protein-coupled receptors whose structure is highly conserved in mammals. OX1R is selective for orexin A, and OX2R binds orexin A and orexin B with similar affinity. Orexin receptor expression was observed in human, rat, porcine, sheep as well as Xenopus laevis pituitaries, both in the adenohypophysis and in the neurohypophysis. The expression level is regulated by gonadal steroid hormones and GnRH. The majority of orexins reaching the pituitary originate from the lateral hypothalamus, but due to the presence of the receptors and the local production of orexins in the pituitary, orexins could deliver an auto/paracrine effect within the gland. Cumulative data indicate that orexins are involved in the regulation of LH, GH, PRL, ACTH, and TSH secretion by pituitary cells, pointing to orexins’ effect on the functioning of the endocrine axes. Those hormones may also serve as a signal linking metabolic status with endocrine control of sleep, arousal, and reproduction processes.

Orexins A and B are neuropeptides which are synthesized mainly in the lateral hypothalamus and are associated with a variety of physiological functions such as energy homeostasis, sleep and wakefulness, feeding behavior, as well as the reproductive system. The orexins activate two G-protein-coupled receptors termed orexin receptor 1 (OX1R) and orexin receptor 2 (OX2R). The present study analyses OX1R and OX2R mRNAs and proteins' expression in the porcine ovary on days 2-3, 10-12, 14-16 and 17-19 of the oestrous cycle. Using real-time PCR, higher OX1R mRNA expression was detected in porcine CLs (p

Leptin, the product of the obese gene, is the hormone originally identified in adipocytes. It is involved in the control of satiety and energy metabolism. More recent observations suggest that leptin plays an important role in reproduction. Leptin mRNA and protein have been found in the human and the murine ovary. However, the expression of leptin in the porcine ovary has not been examined. Therefore, the aim of the present work was to compare the expression levels of porcine leptin mRNA by semiquantitative RT-PCR and in situ hybridization, as well as leptin protein by Western blotting in the corpus luteum (CL) and ovarian stroma (OS) during mid- and late-luteal phase of the oestrous cycle as well as during days 14-16 and 30-32 of pregnancy. Leptin gene and protein expression in CL was increased on days 14-16 of the cycle compared with pregnant animals. Leptin gene expression in OS was higher during the late-luteal phase of the cycle than on days 30-32 after conception. However, comparison of leptin protein expression in OS between days 14-16 of the cycle and days 30-32 of pregnancy indicates a higher protein expression during pregnancy. Moreover, leptin gene expression was higher in porcine CL and OS on days 14-16 of pregnancy in comparison to days 30-32. Contrary to leptin mRNA expression, a higher leptin protein expression was observed on days 30-32 compared with days 14-16 after conception. In summary, the present study provides the first evidence that leptin mRNA and protein occur in porcine ovary and vary during the oestrous cycle and pregnancy. Moreover, the obtained results indicate that also locally synthesized leptin may participate in the control of pig reproduction by exercising its action at the ovarian level.

Leptin, a hormone secreted by adipocytes, plays an important role in the regulation of metabolism and reproduction. The effect of leptin is mediated mainly via the long isoform of the leptin receptor (OB-Rb). Expression of leptin and its receptor has been identified in the central nervous system (hypothalamus, pituitary) and reproductive tract (uterus, ovary and placenta) in prepubertal gilts. Therefore, in the present study, the porcine OB-Rb expression was examined by a semiquantitative reverse transcription polymerase chain reaction (RT-PCR), in situ hybridization and Western blotting in the corpus luteum (CL) and ovarian stroma (OS) during the luteal phase of the estrous cycle (days 10-12 and 14-16) and two stages of early pregnancy (days 14-16 and 30-32). The OB-Rb gene expression in both ovarian structures was higher during early pregnancy in comparison to the mid- and late-luteal phase of the estrous cycle. Significant differences in OB-Rb gene expression between the two periods of the pregnancy were observed only in OS. The results of in situ hybridization are generally supported by data of semiquantitative RT-PCR. The CL expression of leptin receptor protein was significantly lower during early pregnancy compared to the cycle. In OS, the OB-Rb protein content was higher during pregnancy compared to the late-luteal phase of the estrous cycle. In summary, the obtained results indicate that leptin may participate in the control of pig reproduction at the ovarian level and have a direct effect on the ovary during both, the luteal phase of the cycle and early pregnancy. Moreover, changes in OB-Rb gene and protein expression in ovarian structures strongly suggest that their sensitivity to leptin varies throughout the luteal phase of the cycle and early pregnancy.

The aim of this study was the detection and location of long form leptin receptor (OB-Rb) in different area of hypothalamus and pituitary in the pig during early pregnancy.Expression of OB-Rb was examined by RT-PCR in the different area of hypothalamus: medial basal hypothalamus (MBH), preoptic area (POA), stalk median eminence (SME), as well as pituitary: the anterior (AP) and posterior (NP) lobe collected from gilts at days 14-16 (n=4) and 30-32 (n=4) of pregnancy.The results showed that OB-Rb mRNA was expressed in the hypothalamus (MBH, POA and SME), pituitary (AP, NP) and adipose tissue in the pig during early pregnancy (at days 14-16 and 30-32).These findings support the idea that leptin might play a role in the regulation of the hypothalamic-pituitary axis activity, and consequently in the control of pregnancy during critical period of embryo implantation in the pig.

Leptin, the 16-kDa peptide hormone product of the ob gene, is a regulatory hormone secreted mainly by adipose tissue. Recent studies have shown leptin production by other tissues, including rat hypothalamus, rat and human pituitary, rat skeletal muscle, kidney and stomach, human and porcine placenta, human mammary epithelial cells as well as endometrial tissues. This hormone is a central modulator of food intake, metabolism and neuroendocrine functions.The aim of the study was to detect and locate porcine leptin gene expression in the different areas of the hypothalamus and pituitary on days 14-16 and 30-32 of pregnancy in pigs.Leptin gene expression was analysed by RT-PCR method. PCR products were subjected to sequencing analysis.Leptin mRNA was expressed in the medial basal hypothalamus, preoptic area, stalk median eminence, anterior pituitary, posterior pituitary and adipose tissue on days 14-16 and 30-32 of pregnancy. Sequence analysis of the 258 bp product from the hypothalamus and pituitary confirmed 99% homology with the corresponding region of porcine leptin cDNA sequence.Leptin mRNA expression in the porcine hypothalamus and pituitary gland implies its paracrine and/or autocrine role in the regulation of hypothalamic-pituitary axis activity.

Background: Adiponectin, protein secreted mainly by white adipose tissue, is an important factor linking the regulation of metabolic homeostasis and reproductive processes. The biological activity of the hormone is mediated via two distinct receptors, termed adiponectin receptor 1(AdipoR1) and adiponectin receptor 2 (AdipoR2). The present study analyzed mRNA and protein expression of AdipoR1 and AdipoR2 in the anterior (AP) and posterior (NP) pituitary of cyclic pigs. Methods: The total of 20 animals was assigned to one of four experimental groups (n = 5 per group) as follows: days 2–3 (early-luteal phase), 10–12 (mid-luteal phase), 14–16 (late-luteal phase), 17–19 (follicular phase) of the oestrous cycle. mRNA and protein expression were analyzed using real-time PCR and Western Blot methods, respectively. Results: The lowest AdipoR1 gene expression was detected in AP on days 10–12 relative to days 2– 3a nd 14–16 (p< 0.05). In NP, AdipoR1 mRNA levels were elevated on days 10–12 and 14–16 (p< 0.05). AdipoR2 gene expression in AP was the lowest on days 10–12, and an expression peak occurred on days 2–3 (p