Zinc glycinate alleviates LPS-induced inflammation and intestinal barrier disruption in chicken embryos by regulating zinc homeostasis and TLR4/NF-κB pathway.

The effect of an immune challenge induced by a lipopolysaccharide (LPS) exposure on systemic zinc homeostasis and the modulation of zinc glycinate (Zn-Gly) was investigated using a chicken embryo model. 160 Arbor Acres broiler fertilized eggs were randomly divided into 4 groups: CON (control group, injected with saline), LPS (LPS group, injected with 32 µg of LPS saline solution), Zn-Gly (zinc glycinate group, injected with 80 µg of zinc glycinate saline solution) and Zn-Gly+LPS (zinc glycinate and LPS group, injected with the same content of zinc glycinate and LPS saline solution). Each treatment consisted of eight replicates of five eggs each. An in ovo feeding procedure was performed at 17.5 embryonic day and samples were collected after 12 hours. The results showed that Zn-Gly attenuated the effects of LPS challenge-induced upregulation of pro-inflammatory factor interleukin 1ß (IL-1ß) level (P =0.003). The LPS challenge mediated zinc transporter proteins and metallothionein (MT) to regulate systemic zinc homeostasis, with increased expression of the jejunum zinc export gene zinc transporter protein 1 (ZnT-1) and elevated expression of the import genes divalent metal transporter 1 (DMT1), Zrt- and Irt-like protein 3 (Zip3), Zip8 and Zip14 (P < 0.05). A similar trend could be observed for the zinc transporter genes in the liver, which for ZnT-1 mitigated by Zn-Gly supplementation (P =0.01). Liver MT gene expression was downregulated in response to the LPS challenge (P =0.004). These alterations caused by LPS resulted in decreased serum and liver zinc levels and increased small intestinal, muscle and tibial zinc levels. Zn-Gly reversed the elevated expression of the liver zinc finger protein A20 induced by the LPS challenge (P =0.025), while Zn-Gly reduced the gene expression of the pro-inflammatory factors IL-1ß and IL-6, decreased toll-like receptor 4 (TLR4) and nuclear factor kappa-B p65 (NF-κB p65) (P < 0.05). Zn-Gly also alleviated the LPS-induced downregulation of the intestinal barrier gene Claudin-1. Thus, LPS exposure prompted the mobilization of zinc transporter proteins and MT to perform the remodeling of systemic zinc homeostasis, Zn-Gly participated in the regulation of zinc homeostasis and inhibited the production of pro-inflammatory factors through the TLR4/NF-κB pathway, attenuating the inflammatory response and intestinal barrier damage caused by an immune challenge.

Dietary 25-hydroxy-cholecalciferol and additional vitamin E improve bone development and antioxidant capacity in high-density stocking broilers.

This study aimed to investigate the effects of diets supplemented with 25-hydroxycholecalciferol [25-(OH)D3] and additional vitamin E on growth performance, antioxidant capacity, bone development, and carcass characteristics at different stocking densities on commercial broiler farms. A total of 118,800 one-day-old Arbor Acres broilers were assigned to a 2 × 2 factorial treatment consisting of two dietary vitamin levels (5,500 IU vitamin D3 and 60 IU vitamin E: normal diet, using half 25-(OH)D3 as a source of vitamin D3 and an additional 60 IU of vitamin E: 25-(OH)D3+VE diet) and two stocking densities (high density of 20 chickens/m2: HD and 16 chickens/m2: LD). The experiment lasted for 42 d. The results showed that high-density stocking negatively affected the growth performance of broilers during the first four weeks, whereas the vitamin diet treatment significantly improved the feed conversion ratios (FCR) during the last 2 wk. Vitamin diets increased catalase at 14 and 42 d, and the glutathione peroxidase (GSH-px) levels at 42 d in high-density-stocked broilers. The interaction showed that serum vitamin E levels were significantly improved at 28 d of age in high-density-stocked broilers as a result of the vitamin diets. Stocking density and dietary treatments were found to significantly affect bone development, with the vitamin diet significantly increasing metatarsal length and femoral bone strength in broilers from high-density stocking density at 28 d of age. High stocking density increased the proportion of leg muscles and meat yield per square meter. In general, 25-(OH)D3 and additional vitamin E suppressed oxidative stress and ameliorated the negative effects of high-density stocking on bone development in a commercial chicken farm setting. Vitamin diets improved the FCR of broilers, while high-density stocking resulted in better economic outcomes.

High-density stocking is often associated with animal welfare risks in broilers, mainly in terms of oxidative stress and bone development. Nevertheless, farming at too low a density remains for the most part economically unviable. Modulation of antioxidant capacity and bone development by nutritional strategies in high-density-farmed broilers has proven an effective tool in developing countries. Therefore, the present study investigated the effects of applying diets with a higher biological potency of vitamin D3 25-hydroxycholecalciferol [25-(OH)D3] and a higher concentration of vitamin E on broiler production performance, antioxidant capacity and meat production performance at different densities of stocking under commercial farming conditions. The results indicated that the vitamin dietary treatments suppressed oxidative stress and ameliorated the negative effects of high-density farming on bone development.

Glycerol monolaurate improves intestinal morphology and antioxidant status by suppressing inflammatory responses and nuclear factor kappa-B signaling in lipopolysaccharide-exposed chicken embryos.

Medium-chain fatty acids and their derivatives are natural ingredients that support immunological functions in animals. The effects of glycerol monolaurate (GML) on intestinal innate immunity and associated molecular mechanisms were investigated using a chicken embryo model. Sixty-four Arbor Acres broiler embryos were randomly allocated into four groups. On embryonic day 17.5, the broiler embryos were administered with 9 mg of GML, which was followed by a 12-h incubation period and a 12-h challenge with 32 µg of lipopolysaccharide (LPS). On embryonic day 18.5, the jejunum and ileum were harvested. Results indicated that GML reversed the LPS-induced decline in villus height and upregulated the expression of mucin 2 (P < 0.05). GML decreased LPS-induced malondialdehyde production and boosted antioxidant enzyme activity (P < 0.05). GML alleviated LPS-stimulated intestinal secretion of interleukin (IL)-1ß, IL-6, and tumor necrosis factor-α (TNF-α) (P < 0.05). GML also normalized LPS-induced changes in the gene expression of Toll-like receptor 4, nuclear factor kappa-B p65 (NF-κB p65), cyclooxygenase-2, NOD-like receptor protein 3, IL-18, zonula occludens 1, and occludin (P < 0.05). GML enhanced as well the expression of AMP-activated protein kinase α1 and claudin 1 (P < 0.05). In conclusion, GML improved intestinal morphology and antioxidant status by alleviating inflammatory responses and modulating NF-κB signaling in LPS-challenged broiler embryos.

Dietary Macleaya cordata extract supplementation improves the growth performance and gut health of broiler chickens with necrotic enteritis.

BACKGROUND: The poultry industry needs effective antibiotic alternatives to control outbreaks of necrotic enteritis (NE) caused by Clostridium perfringens. METHODS: The aim of this study was to investigate the effects of dietary supplementation with Macleaya cordata extract (MCE) on the immune function and gut microbiota of broilers with NE. A total of 288 1-day-old broiler chicks were randomly assigned to a 2 × 2 factorial arrangement with two concentrations of dietary MCE supplementation (0 or 350 mg/kg of diet) and two disease challenge statuses (control or NE). RESULTS: The results revealed that NE significantly increased the feed conversion rate (FCR), mortality, intestinal lesion score, the levels of IL-1ß, IL-17 and IFN-γ/IL-4 in serum and IL-17/IL-10 in the jejunal mucosa, mRNA levels of TLR2, IFN-γ and pIgR in the jejunum, and Clostridium perfringens concentrations in the cecum. NE significantly decreased the body weight (BW), body weight gain (BWG), jejunal villus height, V/C, mRNA level of AMPK-α1 in jejunum, IL-4 level in the jejunal mucosa and lactic acid bacteria abundance in the cecum. MCE significantly increased BW, BWG, jejunal villus height, V/C, mRNA levels of occludin, ZO-1 and AMPK-α1 in the jejunum, the levels of IgA and IgG in serum and IL-10 in the jejunal mucosa and mRNA levels of NF-κB, IL-10 and MHC-II in the jejunum. Additionally, MCE significantly decreased the FCR, mortality, intestinal lesion score, jejunal crypt depth, the levels of IFN-γ and IL-17 in serum and IL-17/IL-10 in the jejunal mucosa, Clostridium perfringens concentrations in the cecum, and mRNA levels of IL-17/IL-10 in the jejunum. Moreover, NE significantly increased the abundance of bacteria that are associated with inflammation, obesity and depression (Alistipes, Barnesiella, Intestinimonas, RF39 and UCG-005) and significantly decreased the abundance of short-chain fatty acid (SCFA)-producing bacteria (Anaerotruncus, Butyricicoccus and Bacteroides) in the cecum. MCE significantly increased the abundance of SCFA-producing bacteria (Streptococcus, Ruminococcus_torques_group and Lachnospiraceae_NK4A136_group) and significantly reduced the abundance of bacteria that are associated with inflammation and obesity (Alistipes, Barnesiella and UCG-010) in the cecum. In the cecum of broilers with NE, the relative abundance of Barnesiella and Alistipes was higher and that of Lachnoclostridium and Shuttleworthia was lower. Interestingly, these trends were reversed by the addition of MCE to the diet. Spearman correlation analysis showed that Barnesiella and Alistipes were associated with enhanced intestinal inflammation and inhibited growth performance, whereas Lachnoclostridium and Shuttleworthia were associated with anti-inflammatory effects. CONCLUSIONS: MCE ameliorated the loss of growth performance in broiler chickens with NE, probably by regulating the intestinal barrier, immune function, and gut microbiota.

RNA sequencing transcriptomics and metabolomics in three poultry breeds.

Chickens are remarkably versatile animals that are used as model organisms for biomedical research. Here, we performed metabolomic and RNA sequencing (RNA-Seq) transcriptomic analyses of the hypothalamus, liver tissue and serum of poultry with different genetic backgrounds, providing detailed information for hypothalamus and liver tissue at the transcriptional level and for liver tissue and serum at the metabolite level. We present two datasets generated from 36 samples from three poultry breeds using high-throughput RNA-Seq and liquid chromatography coupled with mass spectrometry acquisition (LC/MS). The transcriptomic and metabolomic data obtained for poultry of different genetic backgrounds will be a valuable resource for further studies on this model organism.

Dietary sodium butyrate improves female broiler breeder performance and offspring immune function by enhancing maternal intestinal barrier and microbiota.

This study aimed to investigate the effects of dietary sodium butyrate (SB) supplementation on the reproductive performance of female broiler breeders under intensive rearing conditions and to analyze antioxidant capacity, immune function, and intestinal barrier function of the female breeders and their offspring. A total of 96,000 40-wk-old Ross308 female broiler breeders were divided into the control (CON) and SB groups, each with 6 replicates of 8,000 birds. Each house with similar production performance characteristics was considered a replicate. The experiment lasted for 20 wk, whereupon sampling took place. Results showed that SB improved the egg production performance, egg quality of broiler breeders, and hatchability (P < 0.05). Maternal supplementation with SB substantially increased serum immunoglobulin A levels in broiler breeders and offspring (both P = 0.04) and offspring immunoglobulin G (P < 0.001). The levels of interleukin-1ß (P < 0.001) and interleukin-4 (P = 0.03) in the offspring were downregulated, while the total superoxide dismutase in the offspring and the eggs increased (P < 0.05). The serum biochemical components in breeders and offspring were altered by SB, as evidenced by the reduction in triglycerides, total cholesterol, and high- and low-density lipoproteins (P < 0.05). The intestinal morphology of broiler breeders and offspring also improved by the SB with the decreasing the jejunal crypt depth (P = 0.04) and increasing villus height in offspring (P = 0.03). Maternal jejunal and ileal intestinal barrier-related genes were also shown to be significantly affected by SB. Furthermore, SB altered the microbial diversity in maternal cecal contents, thus increasing the abundance of Lachnospiraceae (P = 0.004) and Ruminococcaceae (P = 0.03). Dietary SB enhanced the reproductive performance and egg quality of broiler breeders and improved the antioxidant capacity and immune function of broiler breeders and offspring, with the benefits potentially arising from the regulation of the maternal intestinal barrier and gut microbiota by SB.

Dietary Qi-Weng-Huangbo powder enhances growth performance, diarrhoea and immune function of weaned piglets by modulating gut health and microbial profiles.

Introduction: The evolution of nutritional strategies to improve the gut health and microbiota profiles of early-weaned piglets is essential to reduce diarrhoea caused by weaning stress. Therefore, the aim of this study was to determine the effects of dietary supplementation of Qi-Weng-Huangbo powder, a traditional herbal medicine consisting of a mixture of Pulsatilla chinensis, Chinese Schneid and Astragalus extracts (PCE), on the growth performance, diarrhoea rate, immune function and intestinal health of weaned piglets. Methods: 162 piglets were randomly assigned to the CON group (no PCE added), the PCEL group (300 mg/kg PCE) and the PCEH group (500 mg/kg PCE) at the end of the third week post farrowing. There were 9 replicates of each group with 6 pigs per replicate. The experiment lasted for 28 days and sampling was performed on the final day. Results: The results showed that the PCE diet increased the average daily gain (ADG) and final body weight (BW) compared to the CON group. Both supplemented doses of PCE reduced the faecal scores of piglets, and the diarrhoea rate in the PCEL group was significantly lower than that in the CON group. The application of PCE diets promoted the development of the spleen in piglets and up-regulated serum immunoglobulin concentrations to enhance immune function, which was also reflected in the down-regulated gene expression of the colonic TLR/MyD88/NF-κB pathway. Supplementation with PCE improved intestinal morphology, and all doses of PCE significantly increased villus height (VH) in the ileum, whereas colonic crypt depth (CD) was significantly lower in the PCEH group than in the CON group. The PCEH diet significantly increased the levels of valeric and isovaleric acid in the colon content. Dietary PCEH also improved the colonic microbial community profile, reflected by a significant increase in Shannon's index compared with CON group. The abundance of Veillonellaceae and Rhodospirillales was significantly increased in the PCEH group at the family level. Discussion: In conclusion, dietary PCE reduced diarrhoea rates, improved growth performance and enhanced immune function in weaned piglets. These improvements were potentially supported by altered ileum and colonic morphology, elevated colonic VFA levels, and modulation of colonic microbial profiles.

Glycerol monolaurate attenuated immunological stress and intestinal mucosal injury by regulating the gut microbiota and activating AMPK/Nrf2 signaling pathway in lipopolysaccharide-challenged broilers.

This study was conducted to investigate the effects of glycerol monolaurate (GML) on lipopolysaccharide (LPS)-induced immunological stress and intestinal mucosal injury in broilers and its underlying mechanisms. A total of 144 one-d-old Arbor Acres broilers were allocated to a 2 × 2 factorial arrangement involving dietary treatment (0 or 1,200 mg/kg dietary GML) and LPS challenge (injected with saline or Escherichia coli LPS on d 16, 18, and 20). Samples were collected on d 21. The results revealed that dietary GML augmented serum immunoglobulin A (P = 0.009) and immunoglobulin G (P < 0.001) levels in challenged birds. Dietary GML normalized LPS-induced variations in serum interleukin-6, interferon-gamma, and LPS levels (P < 0.05), jejunal villus height (P = 0.030), and gene expression of interleukin-6, macrophage inflammatory protein-3 alpha, Toll-like receptor 4, nuclear factor kappa-B, caspase-1, tight junction proteins, adenosine monophosphate-activated protein kinase alpha 1 (AMPKα1), nuclear factor-erythroid 2-related factor 2 (Nrf2), and superoxide dismutase-1 (P < 0.05). GML supplementation ameliorated LPS-induced peroxidation by reducing malondialdehyde content and increasing antioxidant enzyme activity (P < 0.05). Dietary GML enhanced the abundances of Anaerostipes, Pseudoflavonifractor, and Gordonibacter and reduced the proportion of Phascolarctobacterium in challenged birds. Dietary GML was positively correlated with alterations in antioxidant enzyme activities and AMPKα1, Nrf2, and zonula occludens-1 expressions. The genera Anaerostipes, Lachnospira, Gordonibacter, Lachnospira, Marvinbryantia, Peptococcus, and Pseudoflavonifractor were linked to attenuated inflammation and improved antioxidant capacity of challenged birds. In conclusion, dietary GML alleviated LPS-induced immunological stress and intestinal injury of broilers by suppressing inflammation and oxidative stress. Dietary GML regulated cecal microbiota and activated the AMPK/Nrf2 pathway in LPS-challenged broilers.

Probiotic Effects of Bacillus licheniformis DSM5749 on Growth Performance and Intestinal Microecological Balance of Laying Hens.

This study was conducted to investigate the effects of Bacillus licheniformis DSM5749 on the production performance and intestinal health in laying hens. A total of 32-week-old laying hens (Hyline Brown) were randomly assigned to two dietary groups (10 replicates of 27 laying hens), namely, basal diet and basal diet complemented with 200 g/t B. licheniformis (3.2 × 109 CFU/kg). The trial lasted for 8 weeks, and samples were collected at the last week. Results revealed that B. licheniformis DSM5749 significantly improved laying performance, including an increase in egg production rate and average daily egg yield, and a decrease in the feed-to-egg ratio during the entire 8-week experimental period (P < 0.05). B. licheniformis DSM5749 increased the levels of superoxide dismutase and glutathione peroxidase in the liver and decreased the IL-1 level in the serum (P < 0.05). In addition, the integrity of intestinal morphology (villus height, crypt depth, and villus height/crypt depth), tight junctions (ZO-1, Claudin-1, and Occludin), and lipase vitality in the intestine were potentiated by B. licheniformis DSM5749 in laying hens (P < 0.05). B. licheniformis DSM5749 decreased the Firmicutes/Bacteroidetes ratio (P < 0.05) in the cecum. Furthermore, B. licheniformis DSM5749 modulated the microbiota in the cecum of the laying hens, increased the relative abundance of beneficial bacteria (e.g., Prevotella) at the genus level and decreased the relative abundance of potential pathogens (e.g., Desulfovibrio). In conclusion, B. licheniformis DSM5749 can improve laying performance, promote intestinal health, affect the composition of cecal microorganisms, and regulate the intestinal micro-ecological balance, making B. licheniformis a good probiotic candidate for application in the laying hens industry.

High Temperature-Induced Oxidative Stress Affects Systemic Zinc Homeostasis in Broilers by Regulating Zinc Transporters and Metallothionein in the Liver and Jejunum.

To investigate the change in zinc homeostasis of broilers under heat stress, 512 broiler chickens were raised to the age of 28 days. The broilers were then assigned to heat stress and normal temperature (36.0°C vs. 26.0°C) groups for 7 days. The results showed that oxidative stress induced by high temperature had a negative effect on the growth performance of broilers. Heat stress altered zinc homeostasis and led to a redistribution of zinc in broilers, which was reflected in increased zinc concentrations in the jejunum, liver, and tibia. Upregulation of the expression of the zinc exporter ZnT1 and importers ZIP8 and ZIP14 in the jejunum indicated that more zinc was absorbed and transported from the jejunum into the blood, while the liver increased its capacity to hold zinc through upregulation of metallothionein (MT) expression, which was achieved by reducing ZnT1 expression and upregulating the expression of the importer ZIP3. The pathway was mediated by zinc transporters, but the capacity of MT to chelate and release zinc ions also played a crucial role. The mechanism of alterations in zinc homeostasis under heat stress was revealed by the changes in zinc transporters and MT levels in the intestine and liver. Heat stress also altered cecal microbial diversity and reduced the relative abundances of Bilophila and Dialister. In conclusion, broilers altered systemic zinc homeostasis through the regulation of zinc transporters and MT in the liver and jejunum to resist oxidative stress induced by high temperature.

Progress on Gut Health Maintenance and Antibiotic Alternatives in Broiler Chicken Production.

The perturbation of gut health is a common yet unresolved problem in broiler chicken production. Antibiotics used as growth promoters have remarkably improved the broiler production industry with high feed conversion efficiency and reduced intestinal problems. However, the misuse of antibiotics has also led to the increase in the development of antibiotic resistance and antibiotic residues in the meat. Many countries have enacted laws prohibiting the use of antibiotics in livestock production because of the increasing concerns from the consumers and the public. Consequently, one of the most significant discussions in the poultry industry is currently antibiotic-free livestock production. However, the biggest challenge in animal husbandry globally is the complete removal of antibiotics. The necessity to venture into antibiotic-free production has led researchers to look for alternatives to antibiotics in broiler chicken production. Many strategies can be used to replace the use of antibiotics in broiler farming. In recent years, many studies have been conducted to identify functional feed additives with similar beneficial effects as antibiotic growth promoters. Attention has been focused on prebiotics, probiotics, organic acids, emulsifiers, enzymes, essential oils, tributyrin, and medium-chain fatty acids. In this review, we focused on recent discoveries on gut health maintenance through the use of these functional feed additives as alternatives to antibiotics in the past 10 years to provide novel insights into the design of antibiotic-free feeds.

Glycerol Monolaurate Ameliorated Intestinal Barrier and Immunity in Broilers by Regulating Intestinal Inflammation, Antioxidant Balance, and Intestinal Microbiota.

This study was conducted to investigate the impact of glycerol monolaurate (GML) on performance, immunity, intestinal barrier, and cecal microbiota in broiler chicks. A total of 360 one-day-old broilers (Arbor Acres) with an average weight of 45.7 g were randomly allocated to five dietary groups as follows: basal diet and basal diets complemented with 300, 600, 900, or 1200 mg/kg GML. Samples were collected at 7 and 14 days of age. Results revealed that feed intake increased (P < 0.05) after 900 and 1200 mg/kg GML were administered during the entire 14-day experiment period. Dietary GML decreased (P < 0.05) crypt depth and increased the villus height-to-crypt depth ratio of the jejunum. In the serum and jejunum, supplementation with more than 600 mg/kg GML reduced (P < 0.05) interleukin-1ß, tumor necrosis factor-α, and malondialdehyde levels and increased (P < 0.05) the levels of immunoglobulin G, jejunal mucin 2, total antioxidant capacity, and total superoxide dismutase. GML down-regulate (P < 0.05) jejunal interleukin-1ß and interferon-γ expression and increased (P < 0.05) the mRNA level of zonula occludens 1 and occludin. A reduced (P < 0.05) expression of toll-like receptor 4 and nuclear factor kappa-B was shown in GML-treated groups. In addition, GML modulated the composition of the cecal microbiota of the broilers, improved (P < 0.05) microbial diversity, and increased (P < 0.05) the abundance of butyrate-producing bacteria. Spearman's correlation analysis revealed that the genera Barnesiella, Coprobacter, Lachnospiraceae, Faecalibacterium, Bacteroides, Odoriacter, and Parabacteroides were related to inflammation and intestinal integrity. In conclusion, GML ameliorated intestinal morphology and barrier function in broiler chicks probably by regulating intestinal immune and antioxidant balance, as well as intestinal microbiota.

Effects of Dietary Energy Level on Performance, Plasma Parameters, and Central AMPK Levels in Stressed Broilers.

This study aimed to characterize the effects of diets with different energy levels on the growth performance, plasma parameters, and central AMPK signaling pathway in broilers under dexamethasone (DEX)-induced stress. A total of 216 1-day-old male broiler chickens were allocated to groups fed with high (HED), National Research Council-recommended (control), or low (LED) energy diets. At 10 days old, chickens were treated with or without dexamethasone (DEX, 2 mg/kg body weight) for 3 consecutive days. HED increased broiler average daily gain (ADG) at 10 days old, compared with the LED (P < 0.05), while average daily feed intake (ADFI) and feed conversion rate (FCR) decreased as the dietary energy level increased (P < 0.05). Chickens fed a HED had higher total protein (TP) content, albumin (ALB), glucose (GLU), total cholesterol (TCHO), high-density lipoprotein (HDL) cholesterol, and low-density lipoprotein (LDL) cholesterol, compared with the control group (P < 0.05). At 13 days old, DEX decreased ADG and increased FCR in broilers fed with different energy diets (P < 0.05). The DEX-HED group had a higher ADFI than non-DEX treated HED group chickens. In addition, TP, ALB, triglycerides (TG), TCHO, HDL, and LDL content levels in the DEX group were higher than those in the control group (P < 0.05). The uric acid (UA) content of the LED group was higher than that of the HED group (P < 0.05). Further, gene expression levels of liver kinase B1, AMP-activated protein kinase α1, neuropeptide Y, and GC receptor in the hypothalamus were increased in chickens treated with DEX (P < 0.05). There was a trend toward interaction between plasma TCHO and hypothalamic LKB1 expression (0.05 < P < 0.1). In conclusion, this study suggests that HED improves growth performance, plasma glucose and total cholesterol at 10 days old broilers, but had no significant effect on performance, plasma parameters, and central AMPK in stressed broilers.

Effect of dexamethasone on gene expression of cannabinoid receptor type 1 and adenosine monophosphate-activated protein kinase in the hypothalamus of broilers (Gallus domesticus).

Hypothalamic neural circuits play a critical role in integrating peripheral signals and conveying information about energy and nutrient status. We detected cannabinoid receptor type 1 (CB1) distribution in the hypothalamus, liver, duodenum, jejunum, and ileum among 7- and 35-day-old broilers. The effects of dexamethasone (DEX) on CB1 gene expression were evaluated by in vitro and in vivo experiments on glucocorticoid receptor (GR) and adenosine monophosphate-activated protein kinase (AMPK) in the hypothalamus of broilers. In vitro, hypothalamic cells from 17-day-old broiler embryos were incubated with either 0.1% dimethyl sulfoxide or DEX (100 nmol/mL) for 1 h. In the in vivo study, 28-day-old broilers were injected with DEX for 24 h or 72 h. Results showed that CB1 was mainly expressed in the hypothalamus, and 72 h DEX treatment increased the expression. One-day treatment of broilers with DEX did not change the hypothalamic CB1 gene expression. Moreover, DEX treatment for 24 h and 72 h increased the mRNA level of hypothalamic AMPKα2 and GR. However, no differences were observed on the gene expression of CB1, GR, and AMPKα2 in hypothalamic cells with DEX-treated for 1 h. In conclusion, CB1 is mainly expressed in the hypothalamus of broilers; 72-h DEX exposure can regulate the CB1 system and AMPK signaling pathway of the broiler hypothalamus.

The AMPK-mTOR signaling pathway is involved in regulation of food intake in the hypothalamus of stressed chickens.

Glucocorticoids (GCs) can stimulate the appetite and AMPK in broilers. The activation of hypothalamic mTOR has been proposed as an important anorexigenic signal. However, inhibitory effect of AMPK activity on appetite and AMPK downstream signaling pathway under stress has not been reported. In this study, we performed an intracerebroventricular (icv) injection of compound C, an AMPK inhibitor, in GC-treated birds to explore the regulatory mechanism on appetite and AMPK downstream signaling pathway. A total of 48 7-day-old broilers, which had received an icv cannula, were randomly subjected to one of two treatments: subcutaneous injection of dexamethasone (DEX) or saline. After 3 days of continuous DEX injection, chicks of each group received an icv injection with either compound C (6 µg/2 µL) or vehicle (dimethyl sulfoxide, 2 µL). The results showed that body weight gain was reduced by the DEX treatment. Compared with the control, icv injection of compound C reduced feed intake at 0.5-1.5 h. In the DEX-treated group, the inhibitory effect of compound C on appetite remained apparent at 0.5-1 h. The DEX treatment increased the gene expression of liver kinase B1 (LKB1), neuropeptide Y (NPY), and decreased p-mTOR protein level. In stressed broilers, inhibition of AMPK relieved the decreased mTOR activity. A significant interaction was noted in DEX and compound C on protein expression of phospho-AMPK. Taken together, in stressed broilers, the central injection of compound C could inhibit central AMPK activity and reduce appetite, in which the AMPK/mTOR signaling pathway might be involved.