Response of Neuropeptides to Hunger Signals in Teleost.

INTRODUCTION: The perception of hunger is a complex physiological process that requires precise coordination between the central and peripheral tissues. METHODS: In this study, tilapia fasted for 24 h was chosen to establish a hunger model to study the mechanism of homeostasis recovery under the joint regulation of the central nervous system (CNS) and peripheral tissues. RESULTS: The gastric and intestinal contents of tilapia were predominantly depleted after a fasting period of 9 h and 24 h, respectively. The serum glucose level significantly decreased at the 9-h and 24-h fasting, respectively, and the glucokinase-dependent glucosensing mechanism in the liver was identified as well as the significant activation of phospho-AMPK. However, fasting for 24 h did not activate glucosensing mechanisms and AMPK signaling pathways in the hypothalamus. On the other hand, significant reductions were observed in the mRNA levels of the lipid synthesis-related genes fas and accα, and the serum triglyceride levels as well. The mRNA levels of npy, agrp, pomc, and cart in the hypothalamus fluctuated during the fasting period without significant differences. With in situ hybridization npy signals upregulated in the ventral zone of posterior periventricular nucleus after 24-h fasting, pomc signals enhanced in the lateral tuberal nucleus. Based on the serum metabolomic analysis, the levels of branched-chain amino acids, butyrate, and short-chain acylcarnitine decreased, while those of medium- and long-chain acylcarnitine increased. CONCLUSION: Fasting for 24 h resulted in changes in npy and pomc signals within the hypothalamus and triggered the glucosensing mechanism in the liver of tilapia. This study is beneficial for elucidating the response of neuropeptides in the CNS to the changes of nutritional factors when hungry.

Effects of nonantibiotic growth promoter combinations on growth performance, nutrient utilization, digestive enzymes, intestinal morphology, and cecal microflora of broilers.

Given the ban on antibiotic growth promoters, the effects of nonantibiotic alternative growth promoter combinations (NAGPCs) on the growth performance, nutrient utilization, digestive enzyme activity, intestinal morphology, and cecal microflora of broilers were evaluated. All birds were fed pellets of two basal diets-starter (0-21 d) and grower (22-42 d)-with either enramycin (ENR) or NAGPC supplemented. 1) control + ENR; 2) control diet (CON, basal diet); 3) control + mannose oligosaccharide (MOS) + mannanase (MAN) + sodium butyrate (SB) (MMS); 4) control + MOS + MAN + Bacillus subtilis (BS) (MMB); 5) control + MOS + fruit oligosaccharide (FOS) + SB (MFS); 6) control + FOS + BS (MFB); 7) control + MOS + FOS + MAN (MFM); 8) control + MOS + BS + phytase (PT) (MBP). ENR, MOS, FOS, SB, MAN, PT, and BS were added at 100, 2,000, 9,000, 1,500, 300, 37, and 500 mg/kg, respectively. The experiment used a completely random block design with six replicates per group: 2400 Ross 308 broilers in the starter phase and 768 in the grower phase. All NAGPCs significantly improved body weight gain (P < 0.01), utilization of dry matter, organic matter, and crude protein (P < 0.05), villus height and villus height/crypt depth in the jejunum and ileum (P < 0.01), and decreased the feed conversion ratio (P < 0.01) at d 21 and 42. MMS, MMB, MFB, and MFM duodenum trypsin, lipase, and amylase activities increased significantly (P < 0.05) at d 21 and 42. On d 21 and 42, MMS, MMB, and MBP increased the abundance of Firmicutes and Bacteroides whereas MMB, MFB, and MBP decreased the abundance of Proteobacteria, compared to ENR and CON. Overall, the NAGPCs were found to have some beneficial effects and may be used as effective antibiotic replacements in broilers.

Arginine infusion rescues ovarian follicular development in feed-restricted Hu sheep during the luteal phase.

The aim of this study was to investigate the molecular mechanisms of arginine (Arg) on follicular development of acute feed-restricted ewes during the luteal phase. From day 6 of the estrous cycle, 24 multiparous Hu sheep were randomly assigned into three groups: control group (a maintenance diet; n = 6), feed restriction group (0.5 maintenance diet, saline infusion; n = 9) and Arg treatment group (0.5 maintenance diet, infusion with 155 µmol of Arg-HCl/kg body weight; n = 9). The intravenous administrations were performed three times per day from day 6 to day 15 of the estrous cycle. At the end of treatment, the hypothalamus and pituitary were collected, as well as the follicular fluid (FF) and granulose cells (GCs) in the ≥2.5 mm follicles. The transcription level of NPVF was significantly increased, and the expression level of GNRH was significantly decreased in the hypothalamus with feed restriction. In addition, feed restriction significantly decreased the number of ≥2.5 mm follicles in the ovaries. In the ≥2.5 mm follicles, feed restriction significantly increased estradiol (E2) level in FF and the expression levels of steroidogenesis related genes (STAR, 3BHSD and CYP19A1) in GCs, while significantly decreased the expressions of FSHR and cell proliferation related genes (YAP1, CCND1 and PCNA) in GCs. Moreover, the activities of glucose metabolism enzymes (PFKP and G6PDH) were significantly decreased in GCs of the ≥2.5 mm follicles with feed restriction. Interestingly, as a precursor of nitric oxide, Arg supplementation can rescue the effects of feed restriction on follicular development by enhancing glucose metabolism and cell proliferation of GCs, and alleviating the abnormal E2 secretion in the ≥2.5 mm follicles, accompanied with recovering the expressions of NPVF and GNRH in the hypothalamus. These findings will be helpful for understanding the role of nutrition and Arg in sheep follicular development.

Effects of l-arginine on endometrial estrogen receptor α/ß and progesterone receptor expression in nutrient-restricted sheep.

This study aimed to determine the effects of l-arginine (L-Arg) supplementation on steroid hormone receptors in non-pregnant ovine endometrium. All experimental ewes were randomly assigned to either a control group (n = 6), a nutrient-restricted group (n = 6), or an L-Arg supplemented nutrient-restricted group (n = 6). The effects of L-Arg on estrogen receptor α/ß (ERα/ß) and progesterone receptor (PGR) expression in the ovine endometrium were assessed. Our results showed that levels of ERß and PGR expression were significantly increased by nutrient restriction, but L-Arg counteracted the effect of nutrient restriction on ERß and PGR expression (p < 0.05). Also, expression of endometrial ERα was substantially increased (p < 0.05) by L-Arg supplementation. Furthermore, ERα/ß and PGR were mainly detected in the endometrial luminal epithelium and glandular epithelium. Therefore, we isolated and identified endometrial epithelial cells (EECs) from sheep. Different concentrations of L-Arg were added to investigate the effects on ERα/ß and PGR in EECs. The expression levels of endothelial nitric oxide synthase, ERß, and PGR were significantly increased in response to low-concentration (200 µmol) L-Arg supplementation, which subsequently decreased with a high concentration (800 µmol) (p < 0.05). Otherwise, ERα expression was remarkably increased at both L-Arg concentrations in EECs (p < 0.05). Overall, the results indicated that L-Arg performed crucial roles in the regulation of ovine steroid hormone receptor expression in the endometrium. The results of this study provide a theoretical basis and technical means for the normal function of endometrium in response to low nutrient levels.

Effects of star anise (Illicium verum Hook.f.) essential oil on nutrient and energy utilization of laying hens.

An experiment using 96 Hy-Line brown laying hens at 29 weeks of age that were randomly allocated to four treatments with eight replicates was conducted to assess the effects of star anise (Illicium verum Hook.f.) oil (SAO) on nutrient and energy utilization. Dietary treatments were corn-soybean meal-based diets supplemented with 0, 200, 400, and 600 mg/kg SAO. The birds were fed the diets for 14 days. The conventional nutrients and amino acids of feed and excreta samples were measured for calculating apparent metabolic efficiency, and gross energy (GE) was analyzed for computation of apparent metabolic energy (AME) and true metabolic energy (TME). Endogenous losses were obtained from another 16 birds for calculating true metabolic efficiency. Birds supplemented with SAO had higher (p < 0.05) metabolic efficiency of crude protein (CP), organic matter (OM), Lys, Met, Arg, and Thr than those of control group. Quadratic effects on utilization of Lys, Met, and Thr were observed (p < 0.05). Inclusion of SAO to diet improved nutrient utilization, and the efficacy is best at 400 mg/kg.

Effects of nitric oxide on steroidogenesis and apoptosis in goat luteinized granulosa cells.

The aim of this study was to investigate effects of nitric oxide (NO) on steroidogenesis and apoptosis in goat luteinized granulosa cells (LGCs). We cultured goat LGCs from healthy follicles in culture medium supplemented with the NO donor sodium nitroprusside (SNP) or the NO synthase inhibitor Nω-Nitro-l-arginine methyl ester hydrochloride (l-NAME), then examined steroid synthesis, oxidative stress and apoptosis in vitro. The results showed that SNP treatment significantly increased the cGMP concentration in the LGCs (P < 0.05), whereas the l-NAME treatment significantly decreased cGMP concentration (P < 0.05). Then Inhibition of NO production significantly inhibited the expression of CYP19A1, a key gene that is involved in sex steroid hormones synthesis and is responsible for the decrease of E2. Inhibition of NO production resulted in an increased percentage of apoptosis, which was accompanied by upregulating expression levels of apoptosis-related markers BAX, CASP3 and CASP9. These data indicate that NO is required for goat LGCs steroidogenesis and cell survival. Furthermore, Inhibition of NO production decreased the expression of mitochondrial biogenesis related genes and proteins (PPARGC1A, NRF-1 and TFAM) and the mtDNA copy number. Simultaneously, inhibition of NO production suppressed the transcription and translation of SOD, GPX1, and CAT, and decreased the glutathione level and increased the 8-OHdG level. However, SNP treatment increased the expression of genes involved in mitochondrial function and biogenesis, and elevated the anti-oxidant stress system and steroid synthesis. Together, our results indicate that NO may up-regulate the expression of PPARGC1A and its downstream factors through the cGMP pathway, thereby decreasing granulosa cell apoptosis, and may participate in the regulation of granulocyte steroid production through the mitochondrial-dependent pathway.

Effects of l-arginine on endometrial microvessel density in nutrient-restricted Hu sheep.

Nutrient deficiency in ruminants can lead to estrus cycle disorders, a decreased pregnancy rate, and reduce birth weight. l-arginine (L-Arg), an important amino acid, can improve uterine homeostasis in pregnant sheep and prevent intrauterine growth restriction (IUGR). However, most studies of L-Arg have been conducted on pregnant sheep and few have reported the effects of L-Arg on microvessel density (MVD) in the non-pregnant ovine endometrium. The processes of normal uterine cyclical development and implantation are dependent on a balanced of endometrial MVD. In this study, female Hu sheep were randomly assigned to either a control group (n = 6), a nutrient-restricted group (n = 6), or an L-Arg supplemented nutrient-restricted group (n = 6). The effects of L-Arg on MVD in ovine endometrium were then studied. Our results showed that ovine endometrial MVD was significantly increased by nutrient restriction, but L-Arg counteracted the effect of nutrient restriction on MVD (P < 0.05). Levels of angiogenic growth factors (including VEGFA, VEGFR2, and FGF2) had significant increases (P < 0.05) in endometrium of nutrient restriction on sheep, but that L-Arg supplementation substantially decreased (P < 0.05) their expressions in nutrient restriction sheep. Furthermore, oxidative stress caused by nutrient restriction was also alleviated by L-Arg supplementation in the ovine endometrium. Overall, the results suggested that L-Arg has crucial roles in maintaining the balance of endometrial MVD and angiogenic growth factors, and increasing anti-oxidation capability in the endometrium of nutrient-restricted sheep. This study will provide a theoretical basis and technical means for the normal development of endometrial microvessels in low nutrition level.

Effects of nutrient restriction and arginine treatment on oxidative stress in the ovarian tissue of ewes during the luteal phase.

The aim of this study was to determine whether nutrient restriction and arginine treatment affect energy metabolism changes and oxidative stress through the mitochondrial pathway in the ovarian tissue of ewes during the luteal phase. On days 6-15 of the estrous cycle, 24 multiparous Hu sheep (BW = 43.56 ±â€¯1.53 kg) were randomly assigned to three groups: control group (CG; n = 6), restriction group (RG; n = 9), and l-arginine group (AG; n = 9) administered Arg treatment (or vehicle) three times per day. The ewes were slaughtered at the end of treatment, and blood samples and ovaries were collected for analysis. In this study, the expression levels of antioxidase enzymes (SOD2, CAT and GPX1) and mitochondrial biogenesis-related genes (ESRRA and TFAM), as well as antioxidase activity and mitochondrial function were examined in ovarian tissue. Nutrient restriction resulted in activation of ESRRA and TFAM and an increase in relative mtDNA copy number, whereas arginine treatment led to a pronounced recovery of ovarian tissue. In addition, we observed increased AMPK phosphorylation at Thr172 and SIRT3 levels in nutrient restricted ewes, and these effects decreased with arginine treatment. In conclusion, the present results indicated that short-term nutritional restriction led to changes in energy metabolism and oxidative stress. These changes disrupted the redox balance, thus leading to apoptosis through the mitochondria-dependent apoptosis pathway. Arginine treatment altered gene expression in ovarian tissue and increased the resistance to oxidative stress and the anti-apoptosis capacity. The results presented here suggest a potential method to increase agricultural productivity and economic benefits in the sheep industry by using dietary supplementation with arginine to decrease temporary undernutrition of ewes.

Dietary rumen-protected arginine and N-carbamylglutamate supplementation enhances fetal growth in underfed ewes.

The present study was conducted with an ovine intrauterine growth restriction (IUGR) model to test the hypothesis that dietary rumen-protected l-arginine (RP-Arg) or N-carbamylglutamate (NCG) supplementation in underfed ewes is effective in enhancing fetal growth. Between Days 35 and 110 of pregnancy, 32 multiparous ewes carrying two fetuses were randomly assigned to one of four groups: a control (CG) group (n=8; 100% National Research Council (NRC) requirements for pregnant sheep), a nutrient-restricted (RG) group (n=8; fed 50% NRC requirements, and two treatment (ARG and NCG) groups (n=8 in each group; fed 50% NRC requirements supplemented with 20gday-1 RP-Arg or 5gday-1 NCG. All ewes were killed on Day 110 of pregnancy to determine fetal weight and fetal organ weights, and metabolites and hormones in fetal plasma, amino acid concentrations in the fetal liver and longissimus dorsi muscle, and expression of mRNAs in the somatotropic axis. Maternal and fetal bodyweight and the weight of most fetal organs expressed as a percentage of bodyweight increased in response to ARG and NCG compared with values for fetuses from RG ewes. Fetal plasma concentrations of insulin, insulin-like growth factor 1, total amino acids, lactate, thyroxine, and the thyroxine/tri-iodothyronine ratio were lower in fetuses from RG ewes compared with the other treatment groups, but concentrations of growth hormone, non-esterified fatty acids, and total cholesterol were greater in fetuses from RG ewes. Maternal RP-Arg or NCG supplementation increased concentrations of amino acids in fetal tissues and expression of mRNAs for somatotropic axis proteins in fetuses from RG ewes. These findings suggest that maternal RP-Arg and NCG supplementation of underfed ewes decreases fetal IUGR by improving metabolic homeostasis of fetal endocrinology, increasing the availability of amino acids in the fetal liver and longissimus dorsi muscle and affecting the expression of somatotropic axis genes.

Vitamin D receptor expression and potential role of vitamin D on cell proliferation and steroidogenesis in goat ovarian granulosa cells.

This study aimed to investigate the expression of the vitamin D receptor (VDR) in goat follicles and to determine the effects of Vit D3 supplementation on goat granulosa cells (GCs) function linked to follicular development. The results demonstrated that VDR was prominently localized in GCs, with expression increasing with follicle diameter. Addition of Vit D3 (1α,25-(OH)2VD3; 10 nM) to GCs caused an increase in VDR and in steroidogenic acute regulator (StAR) and 3ß-hydroxysteroid dehydrogenase (3ß-HSD) mRNA expression. Additionally, Vit D3 increased the cyclic adenosine monophosphate (cAMP), estradiol (E2), and progesterone (P4) levels, while it decreased anti-müllerian hormone receptor (AMHR) and follicle-stimulating hormone receptor (FSHR) mRNA expression (P < 0.05). Addition of FSH remarkably increased E2, P4, and cAMP levels (P < 0.05), and Vit D3 further enhanced the E2 and cAMP levels in the presence of FSH (P < 0.05). Vit D3 significantly induced the mRNA expression of CDK4 and CyclinD1, and downregulated P21 gene expression (P < 0.05). In addition, Vit D3 significantly decreased reactive oxygen species (ROS) production and increased the mRNA and protein expression of superoxide dismutase 2 (SOD2) and catalase (CAT) (P < 0.05). In conclusion, VDR is expressed in GCs of the goat ovaries and Vit D3 might play an important role in GCs proliferation by regulating cellular oxidative stress and cell cycle-related genes. Meanwhile, Vit D3 enhances the E2 and P4 output of GCs by regulating the expression of 3ß-HSD and StAR and the level of cAMP, which regulate steroidogenesis, supporting a potential role for Vit D3 in follicular development.

Effects of diet and arginine treatment during the luteal phase on ovarian NO/PGC-1α signaling in ewes.

The aim of this study was to determine whether arginine (Arg) supplementation of malnourished ewes affects the expression of key NO/PGC-1α signaling pathway genes in the ovary. On Day 6-15 of the estrous cycle, 24 multiparous Hu sheep (BW = 43.56 ± 1.53 kg) were randomly assigned to three groups: control group (CG; n = 6), restriction group (RG; n = 9) and l-arginine group (AG; n = 9), and administered Arg treatment (or vehicle) three times per day. The ewes were slaughtered at the end of treatment, and blood samples and ovaries were collected for analysis. The results of our analyses showed that both short-term feed-restriction and/or supplementation with L-Arg-HCl affected the number of different size follicles observed in the ovary, and the relative day of estrus behavior initiation of ewes. Specifically, the relative day of estrus behavior initiation was significantly advanced in AG compared with that in RG ewes (P < 0.05). Both the number of ≤2 mm-ovarian follicles (P < 0.05) and the total number of ovarian follicles (P < 0.05) were significantly increased in the RG and AG compared with that in the CG ewes. RG ewes exhibited a higher proportion of ≤2 mm (P < 0.05), but a lower proportion of >5 mm follicles than did CG ewes (P < 0.05). The mean number of corpus lutea ≥5 mm was significantly increased in AG as compared to that in either CG or RG ewes. Furthermore, the expression of eNOS, nNOS, iNOS, PDE5A, PDE9A, PRKG2, and PPARGC1A varied significantly among the treatment groups (P < 0.05). GUCY1A3 mRNA levels were significantly increased in RG and AG as compared to those in CG ewes (P < 0.05), whereas conversely, GUCY1B3 mRNA levels were significantly decreased in CG and RG as compared to those in AG ewes (P < 0.05). P53 mRNA levels were found to vary significantly among the three experimental treatment groups (P < 0.05), and similarly, the relative expression levels of P53 were greater in AG and RG than in CG ewes (P < 0.05). The levels of eNOS protein were significantly higher in RG than in either CG or AG ewes (P < 0.05). The relative expression levels of PGC-1α were significantly higher in RG (P < 0.05) and significantly lower in AG ewes (P < 0.05) than in CG ewes. In conclusion, the results of the present study indicate that feed-restriction negatively affects follicular development, and that Arg-supplementation may modulate the expression of key NO/PGC-1α signaling pathway genes in the ovary and thereby accelerate ovulatory processes and the estrous rate. Elucidation of mechanisms underlying these effects of Arg on gene expression in the ewe ovary requires further investigation.

Effects of dietary star anise (Illicium verum†Hook f) supplementation during gestation and lactation on the performance of lactating multiparous sows and nursing piglets.

Thirty-two sows were allocated to four treatments to evaluate the effect of dietary star anise (SA) supplementation during gestation and lactation on the lactational performance of sows. At 85 days of gestation, sows were randomly allotted to one of two diets supplemented with 0.5% SA or basal diet. After farrowing, sows were further allotted to one of two lactation diets supplemented with 0.5% SA or basal diet. On a weekly basis, body weight (BW) of sows and piglets was measured. Blood and milk samples were obtained from the sows and piglets. Number of days from weaning to estrus, milk yield and feed intake were also recorded. Weight gain of piglets from sows fed the SA-supplemented diet during lactation was greater between days 7 and 14, days 14 and 21 and the overall experimental period compared with control groups. Supplementation of SA during lactation improved weaning weight of piglets, milk yield and average daily feed intake (ADFI) of sows. The SA diet increased concentrations of insulin-like growth factor-1 (IGF-1) in ordinary milk and prolactin (PRL) in serum of sows. In conclusion, this study has indicated the beneficial effects of dietary SA addition in improving the lactation performance of sows.