Role of AURKA in the hypothalamus-pituitary-testicular axis in Tibetan sheep from Tianzhu.

The mitosis-associated protein aurora kinase A (AURKA) regulates the maturation of germ cells. We have previously reported using transcriptome analysis that AURKA is expressed in yak testes. Although Tibetan sheep possess an immense economic value, their reproductive rate is low. Herein, the expression and functions of AURKA in the hypothalamus-pituitary-testicular (HPT) axis in Tibetan sheep from Tianzhu were investigated. The cDNA sequence of sheep AURKA was cloned and bioinformatics techniques were used to predict its structure. Tissue expression of AURKA was determined by qPCR, immunoblotting, immunostaining, and immunohistochemistry. The AURKA coding sequence was found to be 1218 bp in length, encoding a 405-amino acid polypeptide chain. Furthermore, the highest sequence similarity of AURKA with the corresponding sequence in other species was seen in goat and cattle; the least degree of similarity was seen in the domestic cat. In addition, AURKA expression was elevated in the testes compared to that in the hypothalamus and pituitary (p < 0.01). Moreover, AURKA was mainly localized in the hypothalamic paraventricular nucleus (magnocellular), chromophobe cells of the pituitary, and spermatogenic cells of the testis. These results indicated that AURKA might participate in sheep reproductive regulation, thus providing a reference for the study of AURKA function in the reproductive process of Tibetan sheep from Tianzhu.

MAPK pathway control of stem cell proliferation and differentiation in the embryonic pituitary provides insights into the pathogenesis of papillary craniopharyngioma.

Despite the importance of the RAS-RAF-MAPK pathway in normal physiology and disease of numerous organs, its role during pituitary development and tumourigenesis remains largely unknown. Here, we show that the over-activation of the MAPK pathway, through conditional expression of the gain-of-function alleles BrafV600E and KrasG12D in the developing mouse pituitary, results in severe hyperplasia and abnormal morphogenesis of the gland by the end of gestation. Cell-lineage commitment and terminal differentiation are disrupted, leading to a significant reduction in numbers of most of the hormone-producing cells before birth, with the exception of corticotrophs. Of note, Sox2+ stem cells and clonogenic potential are drastically increased in the mutant pituitaries. Finally, we reveal that papillary craniopharyngioma (PCP), a benign human pituitary tumour harbouring BRAF p.V600E also contains Sox2+ cells with sustained proliferative capacity and disrupted pituitary differentiation. Together, our data demonstrate a crucial function of the MAPK pathway in controlling the balance between proliferation and differentiation of Sox2+ cells and suggest that persistent proliferative capacity of Sox2+ cells may underlie the pathogenesis of PCP.

Seasonal variation of pituitary gonadotropin subunit, brain-type aromatase and sex steroid receptor mRNAs, and plasma steroids during gametogenesis in wild sablefish.

Pituitary-hormone signaling plays critical roles in the onset and progression of gametogenesis in vertebrates. This study characterized expression patterns of pituitary gonadotropin beta-subunits (fshb and lhb), brain-type aromatase (cyp19a1b), androgen (ar1, ar2) and estrogen receptors (esr1, esr2a, esr2b), and changes in plasma steroid levels by liquid chromatography/tandem mass spectrometry in wild sablefish (Anoplopoma fimbria, order Scorpaeniformes) during a complete reproductive cycle. Transcripts for fshb increased during early gametogenesis and peaked in late vitellogenic females and late recrudescent males, while expression of lhb reached maximum levels in periovulatory and spermiating fish. Pituitary levels of cyp19a1b and ar1 were strongly correlated with those of lhb in females and males, increasing during gametogenesis and reaching maximum levels prior to spawning. By contrast, expression of ar2, and the three estrogen receptors differed between female and male sablefish. 17ß-estradiol (E2) was the dominant steroid in females during vitellogenesis, while a range of at least 6 steroids (11ß-hydroxyandrostenedione, testosterone [T], E2, 11-ketotestosterone [11KT], 11-deoxycortisol, and 17α,20ß,21-trihydroxyprogesterone) were detected at similar levels in males during testicular development. Prior to spawning, a marked increase in 4-androstenedione, T, 11KT and E2 was found in both periovulatory females and spermiating males. In conclusion, the concomitant changes in plasma androgen levels and pituitary ar1 expression during gametogenesis suggest a specific role for androgens in pituitary hormone regulation of reproduction in sablefish. Further, our data highlight the importance of E2 during final stages of maturation in this species, which may regulate the transcription of pituitary lhb in a paracrine fashion.

Light-dark influence on enkephalinase activity in hypothalamus and pituitary.

OBJECTIVES: Enkephalins functions are partly modulated by enkephalinases especially membrane-bound alanyl aminopeptidase (EC-3.4.11.2) considered as the major enkephalin-degrading enzyme in brain. The analysis of its activity in standard and non-standard light/dark conditions may help the understanding of the regulatory mechanisms of enkephalins. METHODS: Enkephalinase activity was determined fluorometrically, using an arylamide derivative as substrate, in hypothalamus and pituitary of adult male rats, in a standard 12:12 h light/dark cycle; samples were collected at 10:00 h, 13:00 h and 16:00 h of the light period (light on from 7:00 to 19:00 h) and at 22:00 h, 01:00 h and 04:00 h of the dark one (light off from 19:00 h to 07:00 h). For comparison, the enzymatic activity was also measured in the same locations at the same time-points but under constant light conditions. RESULTS: In standard light/dark conditions, the results demonstrated an opposite daily rhythm in hypothalamus and pituitary. While the highest levels of AlaAP or enkephalinase activities were measured in hypothalamus during the dark period, they were the highest in the pituitary during the light one. In contrast, the lowest levels of activity were observed in the light period in the hypothalamus whereas they were in the dark one in the pituitary. A similar pattern was observed under constant light. The differences were however higher in hypothalamus and lower, but still significant, in pituitary. CONCLUSION: These results may reflect the behaviour of the endogenous substrates of enkephalinase and consequently be involved in their functions. This observation may affect the chronotherapeutic strategies.

Alkaline ceramidase 1 is essential for mammalian skin homeostasis and regulating whole-body energy expenditure.

The epidermis is the outermost layer of skin that acts as a barrier to protect the body from the external environment and to control water and heat loss. This barrier function is established through the multistage differentiation of keratinocytes and the presence of bioactive sphingolipids such as ceramides, the levels of which are tightly regulated by a balance of ceramide synthase and ceramidase activities. Here we reveal the essential role of alkaline ceramidase 1 (Acer1) in the skin. Acer1-deficient (Acer1(-/-) ) mice showed elevated levels of ceramide in the skin, aberrant hair shaft cuticle formation and cyclic alopecia. We demonstrate that Acer1 is specifically expressed in differentiated interfollicular epidermis, infundibulum and sebaceous glands and consequently Acer1(-/-) mice have significant alterations in infundibulum and sebaceous gland architecture. Acer1(-/-) skin also shows perturbed hair follicle stem cell compartments. These alterations result in Acer1(-/-) mice showing increased transepidermal water loss and a hypermetabolism phenotype with associated reduction of fat content with age. We conclude that Acer1 is indispensable for mammalian skin homeostasis and whole-body energy homeostasis. © 2016 The Authors. The Journal of Pathology published by John Wiley & Sons Ltd on behalf of Pathological Society of Great Britain and Ireland.

Development of an Assay for Measuring Proprotein-Conversion Activity on a Multiplex Magnetic Bead-Based Array Platform.

This chapter describes a protocol for measuring prolyl oligopeptidase (POP) activity using a biotinylated peptide substrate coupled to magnetic microspheres. The complex is incubated in the presence of a pituitary extract and activity can be detected by loss of the biotin label. The assay can be multiplexed for measuring multiple proprotein-cleaving enzymes simultaneously and can be used in analyses of neuropsychiatric diseases in which proteolytic cleavage of biologically active peptides is known to play a role.

Relation among Aromatase P450 and Tumoral Growth in Human Prolactinomas.

The pituitary gland is part of hypothalamic-pituitary-gonadal axis, which controls development, reproduction, and aging in humans and animals. In addition, the pituitary gland is regulated mainly by hormones and neurotransmitters released from the hypothalamus and by systemic hormones secreted by target glands. Aromatase P450, the enzyme responsible for the catabolization of aromatizable androgens to estrogens, is expressed in different parts of body, including the pituitary gland. Moreover, aromatase P450 is involved in sexual dimorphism where alteration in the level of aromatase can initiate a number of diseases in both genders. On the other hand, the direct actions of estrogens, mainly estradiol, are well known for stimulating prolactin release. Numerous studies have shown that changes in the levels of estrogens, among other factors, have been implicated in the genesis and development of prolactinoma. The pituitary gland can produce estradiol locally in several types of endocrine cells, and it is possible that aromatase could be responsible for the maintenance of the population of lactotroph cells and the modulation of the action of central or peripheral regulators. Aromatase overexpression due to inappropriate gene regulation has clinical effects such as the pathogenesis of prolactinomas. The present study reports on the synthesis of pituitary aromatase, its regulation by gonadal steroids, and the physiological roles of aromatase on pituitary endocrine cells. The involvement of aromatase in the pathogenesis of pituitary tumors, mainly prolactinomas, through the auto-paracrine production of estradiol is reviewed.

Pituitary Aromatase P450 May Be Involved in Maintenance of the Population of Luteinizing Hormone-Positive Pituitary Cells in Mice.

As aromatase P450 is located in several pituitary cells, testosterone can be transformed into 17ß-estradiol in the gland by the enzyme. The possible role of this transformation in pituitary function remains to be elucidated, but some evidence suggests a physiological and pathophysiological role for pituitary aromatase. To determine its relevance in the modulation of pituitary function, mainly associated with reproduction, luteinizing hormone (LH)-positive cells in the hypophysis of female and male aromatase knockout (ArKO) mice were studied. In all LH-positive cells, significant increases in the cellular (p < 0.01) and nuclear (p < 0.05) areas were found in the ArKO mice compared to the wild-type mice. In the ArKO mice, LH-positive cells were more abundant (p < 0.01); they were characterized by a stronger cytoplasmic reaction and the cells were more polygonal and exhibited more short, thick cytoplasmic prolongations than those in the wild-type mice. Moreover, LH-positive cells showed a greater proliferation rate in the ArKO mice compared to the wild-type mice (p < 0.01). These findings suggest that the local production of estradiol mediated by pituitary aromatase is necessary for the regulation of LH-positive gonadotropic cells, exerting an autoparacrine inhibitory regulation. These results could underlie the higher pituitary aromatase expression observed in male versus female mice. Similar effects were found in ArKO male and female mice, suggesting that in both sexes the effects of estrogens on maintenance of the LH-positive pituitary cell population could be related to the local aromatization of testosterone to estradiol inside the hypophysis. Therefore, aromatase could modulate pituitary LH-positive cells in males through local estradiol synthesis.

Biosynthesis of D-aspartate in mammals: the rat and human homologs of mouse aspartate racemase are not responsible for the biosynthesis of D-aspartate.

D-Aspartate (D-Asp) has important physiological functions, and recent studies have shown that substantial amounts of free D-Asp are present in a wide variety of mammalian tissues and cells. Biosynthesis of D-Asp has been observed in several cultured rat cell lines, and a murine gene (glutamate-oxaloacetate transaminase 1-like 1, Got1l1) that encodes Asp racemase, a synthetic enzyme that produces D-Asp from L-Asp, was proposed recently. The product of this gene is homologous to mammalian glutamate-oxaloacetate transaminase (GOT). Here, we tested the hypothesis that rat and human homologs of mouse GOT1L1 are involved in Asp synthesis. The following two approaches were applied, since the numbers of attempts were unsuccessful to prepare soluble GOT1L1 recombinant proteins. First, the relationship between the D-Asp content and the expression levels of the mRNAs encoding GOT1L1 and D-Asp oxidase, a primary degradative enzyme of D-Asp, was examined in several rat and human cell lines. Second, the effect of knockdown of the Got1l1 gene on D-Asp biosynthesis during culture of the cells was determined. The results presented here suggest that the rat and human homologs of mouse GOT1L1 are not involved in D-Asp biosynthesis. Therefore, D-Asp biosynthetic pathway in mammals is still an urgent issue to be resolved.

New potential targets for treatment of Cushing's disease: epithelial growth factor receptor and cyclin-dependent kinases.

BACKGROUND: Cushing's disease (CD) is caused by adrenocorticotropic hormone (ACTH)-producing pituitary adenomas (ACTHomas). Drug treatment for CD consists of three strategies: pituitary tumor-targeted therapy, steroidogenesis inhibitors, and glucocorticoid receptor antagonists. All of these strategies are under development, and several new drugs have recently been approved for clinical use or are being tested in clinical trials. Pituitary-targeted drugs are a particularly important method in the treatment of CD. Available pituitary tumor-targeted drugs include a dopamine receptor agonist and a somatostatin analog. Since disrupted cell cycle signaling is clearly associated with pathogenesis of ACTHomas which express active forms of epithelial growth factor receptor (EGFR), cyclins, and the catalytic subunit of cyclin-dependent kinases (CDKs), we focused on these molecules as therapeutic targets for ACTHomas. METHODS: In this review, a literature search were performed using PubMed with following terms; Cushing's disease, EGFR, CDKs, cell cycle, and targeted therapy. CONCLUSION: Accumulating evidence demonstrates that EGFR and cyclin E-CDK2 may be promising targets for treating ACTHomas.

Dynamic analysis of stochastic transcription cycles.

In individual mammalian cells the expression of some genes such as prolactin is highly variable over time and has been suggested to occur in stochastic pulses. To investigate the origins of this behavior and to understand its functional relevance, we quantitatively analyzed this variability using new mathematical tools that allowed us to reconstruct dynamic transcription rates of different reporter genes controlled by identical promoters in the same living cell. Quantitative microscopic analysis of two reporter genes, firefly luciferase and destabilized EGFP, was used to analyze the dynamics of prolactin promoter-directed gene expression in living individual clonal and primary pituitary cells over periods of up to 25 h. We quantified the time-dependence and cyclicity of the transcription pulses and estimated the length and variation of active and inactive transcription phases. We showed an average cycle period of approximately 11 h and demonstrated that while the measured time distribution of active phases agreed with commonly accepted models of transcription, the inactive phases were differently distributed and showed strong memory, with a refractory period of transcriptional inactivation close to 3 h. Cycles in transcription occurred at two distinct prolactin-promoter controlled reporter genes in the same individual clonal or primary cells. However, the timing of the cycles was independent and out-of-phase. For the first time, we have analyzed transcription dynamics from two equivalent loci in real-time in single cells. In unstimulated conditions, cells showed independent transcription dynamics at each locus. A key result from these analyses was the evidence for a minimum refractory period in the inactive-phase of transcription. The response to acute signals and the result of manipulation of histone acetylation was consistent with the hypothesis that this refractory period corresponded to a phase of chromatin remodeling which significantly increased the cyclicity. Stochastically timed bursts of transcription in an apparently random subset of cells in a tissue may thus produce an overall coordinated but heterogeneous phenotype capable of acute responses to stimuli.

Development of a novel assay for proprotein converting enzyme activity on a multiplex bead-based array system.

We describe the development of a novel, robust assay system for determining the changes in activity of proprotein converting enzymes. An assay for prolyl oligopeptidase (POP) activity was constructed using a peptide-streptavidin substrate coupled to magnetic microspheres and cleavage was detected by loss of streptavidin on the MAGPIX reader. Test analysis of postmortem pituitary extracts from schizophrenia patients showed an increase in POP activity compared to controls. The results were validated using both fluorometric and Western blot analyses for POP activity and immunoreactivity, respectively. The assays can be multiplexed for measuring the activity of multiple proprotein cleaving enzymes simultaneously in laboratory and clinical settings and should add valuable new information for conditions such as neuropsychiatric diseases, diabetes, endocrine dysfunction, and cancer, where effects on proteolysis of biologically active peptides play a key role.

Molecular cloning and expression of 17ß-hydroxysteroid dehydrogenase type 2 gene in Hu sheep.

17ß-Hydroxysteroid dehydrogenase type 2 (17ß-HSD2) catalyzes the NADP+-dependent oxidation of the most potent estrogen 17ß-estradiol into the weak estrogen estrone, and the conversion of testosterone to androstenedione. It has been reported that 17ß-HSD2 was expressed in many tissues in human, rats, however, the full-length sequence of 17ß-HSD2 gene and its expression in ewe were still unknown. In this study, we cloned the full-length cDNA sequence and investigated mRNA differential expression in 28 tissues of 12 adult Hu-Sheep which were fed with high- and low- dietary intake. The 1,317 bp full-length cDNA sequence was first cloned. The coding region was 1,167 bp in length, and the monomer was estimated to contain 389 amino acid residues. It shares high AA sequence identity with that of bos Taurus (96.13 %), sus scrofa (77.06 %), canis lupus familiaris (70.44 %), Callithrix jacchus (65.72 %), Nomascus leucogenys (65.46 %), pan troglodytes (65.21 %), human (64.69 %), mus musculus (58.35 %), and a comparatively lower identity to danio rerio (37.85 %). 17ß-HSD2 gene was high expressed in gastrointestinal (GI) tract, liver, but weakly expressed in other tissues. No detected expression was examined in lung. 17ß-HSD2 gene expression was significantly difference in rumen, omasum, duodenum, cecum, hypophysis after high- and low- dietary intake. Results from the present study suggested that 17ß-HSD2 plays a crucial role in almost all tissues protecting against excessive levels of active steroid hormone, and GI tract maybe an important steroid hormone metabolizing organ in Hu-Sheep. This present study is the first to provide the primary foundation for further insight into this ovine gene.

Resistin regulates pituitary lipid metabolism and inflammation in vivo and in vitro.

The adipokine resistin is an insulin-antagonizing factor that also plays a regulatory role in inflammation, immunity, food intake, and gonadal function and also regulates growth hormone (GH) secretion in rat adenopituitary cells cultures with the adipokine. Although adipose tissue is the primary source of resistin, it is also expressed in other tissues, including the pituitary. The aim of this study is to investigate the possible action of resistin on the lipid metabolism in the pituitary gland in vivo (rats in two different nutritional status, fed and fast, treated with resistin on acute and a chronic way) and in vitro (adenopituitary cell cultures treated with the adipokine). Here, by a combination of in vivo and in vitro experimental models, we demonstrated that central acute and chronic administration of resistin enhance mRNA levels of the lipid metabolic enzymes which participated on lipolysis and moreover inhibiting mRNA levels of the lipid metabolic enzymes involved in lipogenesis. Taken together, our results demonstrate for the first time that resistin has a regulatory role on lipid metabolism in the pituitary gland providing a novel insight in relation to the mechanism by which this adipokine can participate in the integrated control of lipid metabolism.

Hyperproinsulinaemia in obese fat/fat mice associated with a carboxypeptidase E mutation which reduces enzyme activity.

Mice homozygous for the fat mutation develop obesity and hyperglycaemia that can be suppressed by treatment with exogenous insulin. The fat mutation maps to mouse chromosome 8, very close to the gene for carboxypeptidase E (Cpe), which encodes an enzyme (CPE) that processes prohormone intermediates such as proinsulin. We now demonstrate a defect in proinsulin processing associated with the virtual absence of CPE activity in extracts of fat/fat pancreatic islets and pituitaries. A single Ser202Pro mutation distinguishes the mutant Cpe allele, and abolishes enzymatic activity in vitro. Thus, the fat mutation represents the first demonstration of an obesity-diabetes syndrome elicited by a genetic defect in a prohormone processing pathway.

Blunted response of pituitary type 1 and brown adipose tissue type 2 deiodinases to swimming training in ovariectomized rats.

Ovariectomy leads to significant increase in body weight, but the possible peripheral mechanisms involved in weight gain are still unknown. Since exercise and thyroid hormones modulate energy balance, we aimed to study the effect of swimming training on body weight gain and brown adipose tissue (BAT) type 2 iodothyronine deiodinase responses in ovariectomized (Ox) or sham-operated (Sh) rats. Rats were submitted to a period of 8-week training, 5 days per week with progressive higher duration of exercise protocol. Swimming training program did not totally prevent the higher body mass gain that follows ovariectomy in rats (16.5% decrease in body mass gain in Ox trained rats compared to 22% decrease in sham operated trained animals, in relation to the respective sedentary groups), but training of Ox animals impaired the accumulation of subcutaneous fat pads. Interestingly, swimming training upregulates pituitary type 1 (p<0.001 vs. all groups) and BAT type 2 iodothyronine deiodinases (p<0.05 vs. ShS and OxS) in sham operated but not in Ox rats, indicating an impaired pituitary and peripheral response to exercise in Ox rats. However, BAT mitochondrial O2 consumption significantly increased by swimming training in both sham and Ox groups, indicating that Ox BAT mitochondria responds normally to exercise stimulus, but does not result in a significant reduction of body weight. In conclusion, increased body mass gain produced by Ox is not completely impaired by 8 weeks of high intensity physical training, showing that these animals sustain higher rate of body mass gain independent of being submitted to higher energy expenditure.

Prolonged high iodine intake is associated with inhibition of type 2 deiodinase activity in pituitary and elevation of serum thyrotropin levels.

Our previous epidemiological study indicated that excessive intake of iodine could potentially lead to hypothyroidism. In the present study, we aimed to investigate the time and dose effect of iodine intake on serum thyrotropin (thyroid-stimulating hormone, TSH) levels and to explore the non-autoimmune regulation of serum TSH by pituitary type 2 deiodinase (D2). A total of 360 Wistar rats were randomly divided into five groups depending on administered iodine dosages (folds of physiological dose): normal iodine (NI), 3-fold iodine (3HI), 6-fold iodine (6HI), 10-fold iodine (10HI) and 50-fold iodine (50HI). At 4, 8, 12 and 24 weeks after administration of sodium iodide, blood was collected for serum TSH measurement by chemiluminescent immunoassay. Pituitaries were also excised for measurement of TSHß subunit expression, D2 expression and activity, monocarboxylate transporter 8 (MCT8) and thyroid hormone receptor ß2 isoform (TRß2) levels. The results showed that iodine intake of 10HI and 50HI significantly increased pituitary and serum TSH levels from 8 to 24 weeks (P < 0·05 v. NI). Excess iodine had no effect on D2 mRNA or protein expression; however, 10HI and 50HI administration significantly inhibited pituitary D2 activities from 8 to 24 weeks (P < 0·05 v. NI). Iodine had no effect on MCT8 or TRß2 protein levels. We conclude that prolonged high iodine intake inhibits pituitary D2 activity and induces elevation of serum TSH levels. These findings may provide a potential mechanism of iodine excess-induced overt and subclinical hypothyroidism.

Up-regulation of gonadotropin mRNA-expression at the onset of gametogenesis in the roach (Rutilus rutilus): evidence for an important role of brain-type aromatase (cyp19a1b) in the pituitary.

The present study characterized changes in key parameters of reproduction in adult roach (Rutilus rutilus) from Lake Grosser Mueggelsee (Berlin, Germany) during natural gametogenesis. Fish of both sexes were sampled in monthly intervals between April and August in order to cover the onset of gametogenesis. Investigated parameters included gonad histology, plasma levels of 17ß-oestradiol (E2), testosterone (T), 11-ketotestosterone (11-KT), and 17,20ß-dihydroxy-4-pregnen-3-one (17,20ß-P) as well as the expression of gonadotropin subunits in the pituitary. Furthermore, the mRNA-expression of brain-type aromatase (cyp19a1b), androgen receptor (ar), and estrogen receptor isoforms was studied at the pituitary level. The onset of gametogenesis - as indicated by follicles with cortical alveoli in females and first spermatogonia B in males - was observed in July, accompanied by a significant up-regulation of follicle-stimulating hormone ß (fshß) mRNA in the pituitary in both sexes. On the other hand, luteinizing hormone ß (lhß) mRNA increased later on in August. In males, the increase of fshß mRNA in July coincided with a rise in plasma 11-KT concentrations. In females, E2 in plasma increased later, not until August, shortly before true vitellogenesis (late cortical alveoli stage). Expression of sex steroid receptors in the pituitary revealed only minor seasonal fluctuations. Most pronounced, ar mRNA displayed the highest level pre-spawning in both sexes. Interestingly, cyp19a1b mRNA-expression in the pituitary increased in parallel with fshß already before any changes in plasma E2 or T occurred. These data suggest an important role of pituitary FSH and aromatase at the onset of gametogenesis in the roach.

Morphological and enzymatic changes caused by a long-term treatment of female rats with a low dose of gonadoliberin agonist and antagonist.

BACKGROUND: Long-term treatment with gonadoliberin analogs is used to block the hypothalamic-pituitary-gonadal axis. The use of these agents is generally considered to be safe; however, some observations suggest the possibility of adverse effects. MATERIAL/METHODS: We investigated whether a 3-months administration of a low dose (6 µg/kg b.w.) of dalarelin - a new agonist, and cetrorelix - a known antagonist of GnRH to female rats causes morphological changes in pituitary gland, ovaries, uterus and liver (HE and VG staining); effects on pituitary, hepatic and blood enzyme activities (histochemical and kinetic methods, respectively), and on the blood lipid profile (colorimetric methods); and to what extent these changes are reversible. RESULTS: Applying analogs effectively inhibited ovulation, affected the uterine endometrium and changed histological appearance of the liver (e.g., steatosis). They altered activities of marker enzymes of cellular respiration, gluconeogenesis and intracellular digestion in the liver and, partially in the pituitary gland, caused undesirable changes in the activities of aspartate aminotransferase, alanine aminotransferase, lactate dehydrogenase, and creatine kinase, and a concentration of cholesterol HDL fraction and triglycerides in the blood. Both morphological and enzymatic effects were more evident after antagonist administration; changes in the blood lipid profile were more evident after agonist administration. In both analogs histological and enzymatic changes persisted a relatively long time after the discontinuation of the treatment. CONCLUSIONS: The low dose of dalarelin and cetrorelix is sufficient to cause limited damage of hepatic cells and may modify the function of pituitary, ovaries, uterus and liver as well as other organs, even after discontinuation of the treatment.