No abstract

Gestagen is a collective term for endogenous and synthetic progesterone receptor (PR) ligands. In teleost fishes, 17α,20β-dihydroxy-4-pregnen-3-one (DHP) and 17α,20β,21-trihydroxy-4-pregnen-3-one (20β-S) are the predominant progestogens, whereas in other vertebrates the major progestogen is progesterone (P4). Progestins are components of human contraceptives and hormone replacement pharmaceuticals and, with P4, can enter the environment and alter fish and amphibian reproductive health. In this study, our primary objectives were to clone the fathead minnow (FHM) nuclear PR (nPR), to develop an in vitro assay for FHM nPR transactivation, and to screen eight gestagens for their ability to transactivate FHM nPR. We also investigated the ability of these gestagens to transactivate FHM androgen receptor (AR). Fish progestogens activated FHM nPR, with DHP being more potent than 20β-S. The progestin drospirenone and P4 transactivated the FHM nPR, whereas five progestins and P4 transactivated FHM AR, all at environmentally relevant concentrations. Progestins are designed to activate human PR, but older generation progestins have unwanted androgenic side effects in humans. In FHMs, several progestins proved to be strong agonists of AR. Here, we present the first mechanistic evidence that environmental gestagens can activate FHM nPR and AR, suggesting that gestagens may affect phenotype through nPR- and AR-mediated pathways.

Heat stress (HS) has been reported to alter fat deposition in broilers, however the underlying molecular mechanisms are not well-defined. The objectives of the current study were, therefore: (1) to determine the effects of acute (2 h) and chronic (3 weeks) HS on the expression of key molecular signatures involved in hepatic lipogenic and lipolytic programs, and (2) to assess if diet supplementation with dried Noni medicinal plant (0.2% of the diet) modulates these effects. Broilers (480 males, 1 d) were randomly assigned to 12 environmental chambers, subjected to two environmental conditions (heat stress, HS, 35°C vs. thermoneutral condition, TN, 24°C) and fed two diets (control vs. Noni) in a 2 × 2 factorial design. Feed intake and body weights were recorded, and blood and liver samples were collected at 2 h and 3 weeks post-heat exposure. HS depressed feed intake, reduced body weight, and up regulated the hepatic expression of heat shock protein HSP60, HSP70, HSP90 as well as key lipogenic proteins (fatty acid synthase, FASN; acetyl co-A carboxylase alpha, ACCα and ATP citrate lyase, ACLY). HS down regulated the hepatic expression of lipoprotein lipase (LPL) and hepatic triacylglycerol lipase (LIPC), but up-regulated ATGL. Although it did not affect growth performance, Noni supplementation regulated the hepatic expression of lipogenic proteins in a time- and gene-specific manner. Prior to HS, Noni increased ACLY and FASN in the acute and chronic experimental conditions, respectively. During acute HS, Noni increased ACCα, but reduced FASN and ACLY expression. Under chronic HS, Noni up regulated ACCα and FASN but it down regulated ACLY. In vitro studies, using chicken hepatocyte cell lines, showed that HS down-regulated the expression of ACCα, FASN, and ACLY. Treatment with quercetin, one bioactive ingredient in Noni, up-regulated the expression of ACCα, FASN, and ACLY under TN conditions, but it appeared to down-regulate ACCα and increase ACLY levels under HS exposure. In conclusion, our findings indicate that HS induces hepatic lipogenesis in chickens and this effect is probably mediated via HSPs. The modulation of hepatic HSP expression suggest also that Noni might be involved in modulating the stress response in chicken liver.

BackgroundThe hypothalamus plays a central role in regulating appetite and metabolism. However, the gene networks within the hypothalamus that regulate feed intake and metabolism, and the effects of fasting on those pathways are not completely understood in any species. The present experiment evaluated global hypothalamic gene expression in newly hatched chicks using microarray analysis to elucidate genes and pathways regulated by feeding, fasting, and delayed feeding. Ten groups of chicks were sampled over four days post-hatch, including fed, fasted, and 48 h fasted followed by access to feed for 4 h, 24 h, and 48 h. Hypothalamic samples were collected for microarray analysis (n = 4). Expression patterns of selected genes were confirmed by quantitative real-time PCR. Pathway analysis of the microarray results predicted a network of genes involved in neuropeptide or neurotransmitter signaling. To confirm the functionality of this predicted gene network, hypothalamic neurons from fed and fasted chicks were isolated and cultured in the presence of neuropeptide Y, somatostatin, α-melanocyte stimulating hormone, norepinephrine, and L-phospho-serine. Results confirmed functional relationships among members of the predicted gene network. Moreover, the effects observed were dependant upon the nutritional state of the animals (fed vs. fasted).ResultsDifferences in gene expression (≥ 1.6 fold) were detected in 1,272 genes between treatments, and of those, 119 genes were significantly (P < 0.05) different. Pathway Miner analysis revealed that six genes (SSTR5, NPY5R, POMC, ADRB2, GRM8, and RLN3) were associated within a gene network. In vitro experiments with primary hypothalamic neurons confirmed that receptor agonists involved in this network regulated expression of other genes in the predicted network, and this regulation within the network was influenced by the nutritional status and age of the chick.ConclusionsMicroarray analysis of the hypothalamus during different nutritional states revealed that many genes are differentially regulated. We found that functional interactions exist among six differentially regulated genes associated within a putative gene network from this experiment. Considering that POMC, an important gene in controlling metabolism, was central to this network, this gene network may play an important role in regulation of feeding and metabolism in birds.

No abstract

The anterior pituitary is comprised of five major hormone-secreting cell types that differentiate during embryonic development in a temporally distinct manner. Microarrays containing 5,128 unique cDNAs expressed in the chicken neuroendocrine system were produced and used to identify genes with potential involvement in the onset of thyroid-stimulating hormone beta-subunit (TSHbeta), growth hormone (GH), and prolactin (PRL) mRNA during embryonic development. We identified 352 cDNAs that were differentially expressed (P < or = 0.05) on embryonic day 10 (e10), e12, e14, or e17, the period of thyrotroph, somatotroph, and lactotroph differentiation. Self-organizing maps were used to identify genes that may function to initiate hormone gene transcription. Consistent with cellular ontogeny, TSHbeta mRNA increased steadily between e10 and e17, GH mRNA increased between e12 and e17, and PRL mRNA did not increase until e17. Expression of 141 genes increased in a manner similar to TSHbeta mRNA, and 64 genes decreased between e10 and e17. Although genes with these expression profiles are likely involved in development of the pituitary gland as a whole, some of these could be specifically associated with thyrotroph differentiation. Similarly, the expression profiles of 69 and 61 genes indicate a potential involvement in the induction of GH and PRL mRNA, respectively. Quantitative real-time RT-PCR was used to confirm microarray results for 31 genes. This is the first study to evaluate changes in anterior pituitary gene expression during embryonic development of any species using microarrays, and numerous transcription factors and signaling molecules not previously implicated in pituitary development were identified.

Heat stress (HS) adversely affects growth performance and inflicts heavy economic losses to the poultry industry. There is, therefore, a critical need to identify new alternative strategies to alleviate the negative effects induced by HS. The tropic medicinal plant, Morinda citrifolia (Noni), is being used in livestock nutrition, however the literature is limited and conflicting for its impact on growth performance. The present study aimed to determine the effect of Noni on feeding and drinking behavior as well as on the hypothalamic expression of stress- and metabolic-related genes in broiler chickens exposed to acute HS. A total of 480 1 day-old male broiler chicks were randomly assigned to 12 controlled environmental chambers. Birds were subjected to two environmental conditions (TN, 25°C vs. HS, 35°C for 2 h) and fed two diets (control vs. 0.2% Noni) in a 2 × 2 factorial design. Feed intake and core body temperature (BT) were recorded during HS period. Blood was collected and hypothalamic tissues were harvested for target gene and protein analyses. Acute HS-broilers exhibited higher BT (~1°C), spent less time eating with a significant decrease in feed intake, and spent more time drinking along with higher drinking frequency compared to those maintained under TN conditions. Although Noni supplementation did not improve feed intake, it significantly delayed (~30 min) and reduced the BT-induced by HS. At molecular levels and under HS conditions, Noni supplementation down regulated the hypothalamic expression of HSP90 and its related transcription factors HSF1, 2, and 4, increased orexin mRNA levels, and decreased the phosphorylation levels of AMPKα1/2Thr172 and mTORSer2481. Together, these data indicated that Noni supplementation might modulate HS response in broilers through central orexin-AMPK-mTOR pathways.

This study was conducted to evaluate potential hormonal mechanisms associated with the stress response, thermoregulation, and metabolic changes of broiler chickens exposed to high environmental temperature. Nine hundred 1-day-old male broiler chicks (Ross 708) were placed in floor pens and raised to 24 d. At 24 d, chicks were randomly assigned to 1 of 2 treatments, heat stress ( HS ) or no HS, and allocated into battery cages in 8 batteries (10 birds per cage, 2 cages per battery). On day 31, blood was collected prior to HS and analyzed using an iSTAT analyzer. Half of the batteries were then moved into 2 rooms with an elevated ambient temperature (35°C) for 8 h. The remaining batteries stayed in the thermoneutral rooms with an ambient temperature of 22°C. Beginning at 5 h after the initiation of HS, blood was collected and analyzed using an iSTAT analyzer, birds were euthanized, and hypothalamus and pituitary samples were collected (16 birds per treatment), flash frozen, and stored at −80°C until RNA extraction. Reverse transcription-quantitative PCR was used to compare mRNA levels of key corticotropic and thyrotrophic genes in the hypothalamus and pituitary. Levels of mRNA for each target gene were normalized to PGK1 (pituitary) and GAPDH (hypothalamus) mRNA. Differences were determined using mixed model ANOVA. HS decreased ( P < 0.05) feed intake, BW, bicarbonate, potassium, CO 2 , and triiodothyronine, while it increased mortality, glucose, pH, plasma thyroxine, and corticosterone. Expression of pituitary corticotropin-releasing hormone receptor 1 was downregulated ( P < 0.001), while corticotropin-releasing hormone receptor 2 mRNA levels were higher ( P = 0.001) in HS birds. HS increased expression of thyroid hormone receptor β ( P = 0.01) (2.8-fold) and thyroid stimulating hormone β ( P = 0.009) (1.4-fold). HS did not affect levels of mRNA of genes evaluated in the hypothalamus. Results showed that HS significantly affected both the thyrotropic and corticotropic axes. Understanding the role and regulation of these pathways during HS will allow researchers to better evaluate management strategies to combat HS.