太田 能之

For the conservation of endangered avian species, developing gamete preservation technologies is essential. However, studies in oocytes have not been widely conducted. In this study, assuming that the ovaries are transported to a research facility after death, we investigated the effect of ovary storage on oocytes for the purpose of cryopreserving avian female gametes by using a chicken as a model of endangered avian species. After excision, the ovaries were stored at either a low temperature (4 °C) or room temperature for 1–3 days. Ovarian follicles stored under different conditions for each period were examined by neutral red staining, histology, and gene and protein expression analysis. In addition, the pH of the storage medium after preserving the ovaries was measured. Then, ovarian tissues were vitrified to determine the cryopreservation competence. Storing the ovarian tissues at 4 °C kept the follicles viable and morphologically normal for 3 days with slow decline. In contrast, although different storage temperature did not influence follicle viability and morphology after only 1 day of storage, ovarian tissues stored at room temperature rapidly declined in structurally normal follicles, and viable follicles were rarely seen after 3 days of storage. Gene and protein expression analysis showed that apoptosis had already started on the first day, as shown by the higher expression of CASP9 under room temperature conditions. Furthermore, high expression of SOD1 and a rapid decline of pH in the storage medium under room temperature storage suggested the influence of oxidative stress associated with low pH in this condition on the follicle survivability in hen ovarian tissues. Our cryopreservation study also showed that ovarian tissues stored at 4 °C could recover after cryopreservation even after 3 days of storage. The described storage conditions and cryopreservation methods, which preserve chicken follicle survival, will lay the foundation of ovarian tissue preservation to preserve the fertility of wild female birds.

To explore metabolic characteristics during the post-hatch developmental period, metabolomic analyses of breast muscle and plasma were performed in chickens. The most significant growth-related changes in metabolite levels were observed between seven and 28 days of age. Some of these metabolites are essential nutrients or reported as growth-promoting metabolites. In the muscle, two imidazole dipeptides—carnosine and its methylated metabolite, anserine—increased with the development. These dipeptide levels may be, in part, regulated transcriptionally because in the muscle mRNA levels of carnosine synthase and carnosine methylation enzyme increased. In contrast, taurine levels in the muscle decreased. This would be substrate availability-dependent because some upstream metabolites decreased in the muscle or plasma. In branched-chain amino acid metabolism, valine, leucine, and isoleucine decreased in the muscle, while some of their downstream metabolites decreased in the plasma. The polyamines, putrescine and spermidine, decreased in the muscle. Furthermore, mRNA levels associated with insulin/insulin-like growth factor 1 signaling, which play important roles in muscle growth, increased in the muscle. These results indicate that some metabolic pathways would be important to clarify metabolic characteristics and/or growth of breast muscle during the post-hatch developmental period in chickens.

Feeding behavior and energy metabolism are precisely regulated by humoral and/or neural factors in the central nervous system. In particular, nuclei, such as the arcuate nucleus, ventromedial hypothalamic nucleus, and lateral hypothalamic area located near the third ventricle of the hypothalamus are the centers of feeding and energy metabolism in various vertebrate species, including chickens. In this study, we evaluated the effects of cannulation of the third ventricle on chick growth and feeding behavior in the neonatal stage, to develop a method for local and chronic central nervous system-mediated energy metabolism. Referring to the chick brain atlas, a guide cannula was inserted into the third ventricle of the chick under anesthesia immediately after hatching using a stereotaxic instrument. The chicks that recovered from anesthesia were bred for 11 days under normal feeding management conditions, and then feed intake amount, body weight gain, and metabolic tissue weight were measured. The effects of direct stimulation of the third ventricle with 2-deoxy-D-glucose on the expression level of the immediate-early gene, cFOS, and feed intake in 5-day-old chicks were also evaluated. There were no differences in feed intake, body weight gain, and metabolic tissue weight between 11-day-old cannulated and control chicks. The expression of cFOS mRNA in the ventromedial hypothalamic nucleus was higher than that in the amygdala after the third ventricular administration of 2-deoxy-D-glucose. Additionally, direct third ventricular injection of 2-deoxy-D-glucose attenuated thefeeding behavior of chicks for a while. Overall, wespeculatethat thetechniqueis effectivefor local and/or chronic stimulation of the nucleus near the third ventricle of the chick hypothalamus, which is important for feed and energy metabolism regulation.

Low protein diets (LPs) constitute a reportedly effective form of nutritional therapy for canine chronic kidney disease and cirrhosis. These diets have long been feared to result in reduced muscle mass due to protein catabolism. This adverse effect, however, remains largely unrecognized in veterinary medicine as there are no easily applicable catabolism indicators. Therefore, we focused on urinary creatinine, a metabolite of protein in the urine, and examined whether its ratio to urinary urea nitrogen (UCrn/UN) can be used to assess protein catabolism. In Experiment 1, we first consecutively fed seven healthy beagles an LP, standard protein (SP), and high protein (HP) diet for 1 week each and then measured the UCrn/UN ratio at 2-h intervals from fasting to 16 h post-prandially. We consequently found that the UCrn/UN ratio was significantly elevated in the LP pre-prandially and at all post-prandial measurement points (P < 0.01). No significant differences were observed between the SP and HP. Analysis of fasting plasma amino-acid concentrations revealed that the concentration of methionine was significantly lower in the LP than in the other diets (P < 0.05). Although the effects of this change in amino-acid concentration were unclear, the UCrn/UN ratio was considered having increased due to a deficiency in protein and/or amino acids during LP feeding. In Experiment 2, we continuously fed five healthy beagles an LP for 18 weeks and then measured the UCrn/UN ratio as described above. We also measured changes in body composition with computed tomography. At weeks 10 and 18, the fasting UCrn/UN ratio was significantly higher than it was prior to the start of the LP; however, post-prandially, the UCrn/UN ratio decreased to the point that the significant difference disappeared. Muscle mass decreased at weeks 10 and 18. These results suggest that the fasting UCrn/UN ratio could be used as an indicator of protein catabolism in LP feeding. Our experiments thus indicate that examination of potential increases in the UCrn/UN ratio 1 week after introduction of LP feeding to healthy dogs could enable detection of body protein catabolism in long-term feeding of LP before muscle breakdown occurs.

Nociceptin/orphanin FQ (N/OFQ) was identified in 1995 as the endogenous ligand for the orphan Gi/G coupled opioid receptor-like 1 receptor (NOPI). Exogenous N/OFQ increases food intake in mammals, but its effect and mode of action in chicks are not fully known. We report herein that N/OFQ(5.0 nmol) has a stimulatory effect on food intake in layer-type chicks over a 2-h period after intracerebroventricular (icv) injection. Thirty minutes after central injection of N/CFQ (5.0 nmol) the concentration of agouti-related protein (AGRP) mRNA in the diencephalon increased, while cocaine- and amphetamine-regulated transcript (CART) mRNA decreased. However, concentrations of neuropeptide Y, proopiomelanocortin and glutamate decarboxylase mRNAs, and of catecholamines and excitatory amino acids were not affected. Simultaneous administration of alpha-melanocyte stimulating hormone (alpha-MSH: 1.0 pmol), a competitor of ACRP, completely blocked the orexigenic effect of N/OFQ (5.0 nmol). These data suggest that N/OFQ functions in layer chicks as an orexigenic peptide in the central nervous system, and that the AGRP and the CART neurons may mediate this function, as in mammals. (C) 2009 Elsevier Inc. All rights reserved.

Germline chimeric chickens were produced by the transfer of primordial germ cells (PGCs) or blastoderm cells. The hatchability of eggs produced by transfer of exogenous PGCs is usually low. The purpose of the present study was investigated to express (3-hydroxyacyl CoA dehydrogenase) 3HADH which is a limiting enzyme in the beta-oxidation of fatty acids for hatching energy. Manipulations of both donor and recipient eggshells were as follows. A window approximately 10 mm in diameter was opened at the pointed end of the eggs at stage 12-15 days incubation. Donor PGCs, taken from the blood vessels of donor embryos from fertilized eggs at the same stage of development, were injected into the blood vessels of recipient embryos. The muscles of chicks in the eggs with transferred PGCs were removed after 20 days of incubation. A cDNA was prepared from the total RNA. The expression of 3HADH in the manipulated embryos was investigated using real-time PCR analysis. Real-time PCR analysis showed that expression of 3HADH was reduced in the muscles of manipulated embryos.

Synthetic growth hormone secretagogues stimulate growth hormone secretion by binding to a specific receptor, growth hormone secretagogue receptor (GHS-R). In this study, we investigated the cDNA and the genomic structure of chicken GHS-R. Chicken GHS-R gene is composed of two exons separated by an intron. Two GHS-R mRNA species, cGHS-R1a and cGHS-R1a-variant (cGHS-R1aV) are generated by alternative splicing of a primary transcript. cGHS-R1a protein is predicted to have seven transmembrane domains by a high degree of amino acid sequence identity with mammalian and teleost homologs. cGHS-R1aV lacks the transmembrane-6 domain due to a 48 bp deletion. RT-PCR analysis showed widespread tissue distributions of cGHS-R1a and cGHS-R1aV mRNAs with much higher amounts of cGHS-R1a in all the tissues. (C) 2003 Elsevier Inc. All rights reserved.

The objective of this study was to determine if the bore diameter of needles used for in ovo injection of amino acids (AA) would affect subsequent chick hatchability. In Experiment 1, needles of different bore diameter were inserted into the egg. In Experiment 2, needles of different bore diameter were used to administer an AA solution that is identical to the AA pattern of the whole egg protein. In both experiments, 27, 24, 22, 20, and 18 gauge needles were tested. Needles were inserted to a depth of 13 mm in both experiments. Non-injected eggs were served as control in each experiment. Needle bore diameter did not affect (P&lt 0.05) hatchability or body weight (BW) of hatched chicks in either experiment. © 2002, Japan Poultry Science Association. All rights reserved.

Major objectives of researchers for poultry nutrition are to improve the performance and efficiency of gain and quality of products. Because feed is the major cost in poultry production, even a small improvement in feed efficiency is economically important. In addition, to produce enough for human consumption, the products of the poultry industry are required to be safe for human health, and the poultry industry has to be sustainable and benign for the ecosystem of the environment. Therefore, this review focuses on 1) the development of new feedstuffs and additives, especially on re-evaluation of by-products which are not evaluated highly because of some weak points, and biomass, organic acids, beta-agonist and organic and chelated minerals, 2) enzymes to improve the utilization of phytate P, oligosaccharide and cellulose and proteins, 3) quality of poultry products to enrich the contents of vitamins, n-3 polyunsaturated fatty acids, to reduce egg yolk cholesterol, and fortification eggshell strength, and 4) taste of meat and eggs.