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Abstracts from VI International Symposium on Avian EndocrinologyMarch 31 - April 5, 1996 Chateau Lake Louise, Alberta Continued The GH Receptor during Growth and DevelopmentL.a. Cogburn Department of Animal and Food Sciences, University of
Delaware, Newark, DE 19717, USA The chicken growth hormone receptor (cGHR) mediates multiple effects of GH which are essential for normal growth and development. The progressive developmental increase in expression of the cGHR gene is reflected by the presence of three transcripts (0.8, 3.2 and 4.3 kb) in target tissue, especially liver. Our previous studies have shown that there is an inverse relationship between plasma GH levels and hepatic expression of the cGHR and that exogenous cGH fails to stimulate growth rate or affect expression of the cGHR in young rapidly-growing broiler chickens. We have used other models to examine the relationship between cGH and the cGHR in growth and development of chickens. We have identified a mutation in the cGHR of a sex-linked dwarf (dwdw) broiler chicken that prohibits expression of a functional cGHR protein which is reflected by the absence of GH-binding activity in liver and serum. The absence of a functional cGHR protein in the dwdw chicken results in overexpression of a truncated transcript (0.8 kb) which shows a developmental increase of 5-fold between 1 and 7 weeks of age. There is a 2-fold developmental increase in expression of all three cGHR transcripts in normal (DwDw) chickens from this strain. At 7 weeks of age, the total cGHR gene expression of the dwdw chicken (0.8 kb) is about 7-times greater than that of the DwDw chicken (0.8, 3.2 and 4.3 kb transcripts). Our studies on growth-selected strains of broiler chickens show that slow-growing strains have higher plasma levels of GH and lower GH-binding activity in liver and serum than fast-growing strains. The expression of the cGHR gene in liver is also greater in fast-growing chickens than in the slower-growing strains. Thus, the developmental regulation of cGHR gene expression in the chicken is independent of GH action, although GH down-regulates expression of cGHR transcripts. Developmental and hormonal factors appear to regulate expression of the cGHR which could be a rate-limiting step in GH-dependent growth of the broiler chicken. The Role of Insulin and Insulin Receptors during DevelopmentF. de Pablo Department of Cell and Developmental Biology, CIB-CSIC,
Madrid, Spain In addition to the well defined metabolic function of insulin and its highly abundant expression in the pancreas, insulin and/or its precursor proinsulin appear to act as locally produced growth factors in development. The pattern of expression of insulin mRNA in two experimental systems, the neurulating chicken embryo (E) and the early embryonic chicken retina, is complementary in time to the expression of IGFI mRNA. Both (pro)insulin mRNA and insulin receptor mRNA are found in a widespread pattern from gastrulation (E 0.5) to late neurulation (E2) and in the developing retina from E3 to E6. At the protein level, the receptors found in the highly proliferating retina at E6 have high affinity but low discriminating capacity for [125I]insulin and [125I]IGFI, binding as well proinsulin. When insulin is added to cultured early whole embryos or retinas, proliferation is stimulated, as determined by DNA synthesis. To further characterize the role of this paracrine acting insulin we have added (pro)insulin mRNA and insulin receptor mRNA antisense oligonucleotides to embryos during neurulation.The percent of cells that undergo apoptosis is increased by these oligonucleotides, and its effect is reversed by coaddition of insulin. Therefore, insulin and/or proinsulin, in addition to IGFI, are required growth factors for cell survival and proliferation during early development in the chick embryo. Neuroendocrine Control of Early Sexual DevelopmentW.J. Kuenzel, K. M. Walsh, J.a. Proudman1 Poultry Science Department, University of Maryland, College
Park, MD 20742, USA; 1Germplasm Gamete Physiology Lab, USDA-ARS,
Beltsville, MD 20705, USA Neural pathways within the avian brain responsible for initiating gonadal development are not fully understood. A key environmental manipulation utilized to examine altered hypothalamo-pituitary function during sexual maturation is the photoperiod. Another manipulation that will help elucidate neuroendocrine systems involved in puberty is the use of sulfamethazine (SMZ). Both SMZ feeding and photostimulation appear to involve the central nervous system. Within two days of consuming SMZ mixed in a standard starter ration, broiler males show significantly elevated plasma luteinizing hormone levels (P _0.05) that remain higher than controls for several weeks. Plasma T3 levels, in contrast, are slightly but significantly lower only during the first six days following SMZ intake. Within two weeks of chronic feeding of SMZ, gonads are significantly larger compared to controls. However, the progonadal effect can be blocked by adding 0.5ppm T3 to the diet. Within the brain, two significant changes occur to thyroid releasing hormone (TRH) and neuropeptide Y (NPY) containing neurons. TRH-like ir within the median eminence (ME) is greater in experimentals. NPY-like neurons within the inferior hypothalamic nucleus (IHn), infundibular n. (IN) and internal zone of the ME are significantly more numerous and show a greater number of darker ir neurons in SMZ-fed chicks versus controls. Results suggest that early sexual maturation in chicks may be characterized by a transient reduction in plasma T3 and an activation of NPY-like neurons in the IH, IN and ME. (Research supported in part by U.S.D.A. - CGP #90-37240-5506 and MAES-CGP-POUL-96-40) Regulation of Somatotroph Differentiation during Chicken Embryonic DevelopmentT.E. Porter Department of Poultry Science, Texas AandM University, College
Station, TX 77843-2472, USA We have previously demonstrated that growth hormone (GH)-secreting cells differentiate between days 12 and 16 of chicken embryonic development. Furthermore, we have shown that GH cell differentiation can be induced in vitro by treatment with serum from day 16 (but not day 12) embryos, indicating that somatotroph ontogeny may be stimulated by a blood-borne signal. We have recently identified the GH cell-differentiating compound present in embryonic (day 16) serum. Pituitary cells from 12 d embryos were cultured for 2 days with untreated serum from day 16 embryos or the same serum that was first subjected to various biochemical treatments. Somatotroph differentiation was then assessed by reverse hemolytic plaque assays for chicken GH. We first tested whether the activity was sensitive to trypsin digestion. Interestingly, trypsin digestion increased the extent of GH cell differentiation in response to day 16 serum. Next we tested whether the activity was aqueous or ether soluble. All activity was contained in the ether phase. Together, these results indicated that the activity was an ether-soluble compound bound by a trypsin-sensitive protein. Evaluation of several candidate steroids revealed that glucocorticoids, progesterone and estradiol could induce GH cell differentiation. However, the glucocorticoid-receptor blocker, RU486, but not the estrogen-receptor blocker, tamoxifen, completely abolished embryonic (day 16) serum-induced somatotroph differentiation, indicating glucocorticoid receptor involvement. Finally, we found that removal of corticosterone, but not progesterone, by immunoprecipitation abolished the stimulatory effect of embryonic (day 16) serum on GH cell differentiation. Taken together, our findings indicate that corticosterone is the blood-borne signal that stimulates somatotroph differentiation during chicken embryonic development. Actions of Growth Hormone on Insulin-like Growth Factor-I Vary with AgeC.G. Scanes, S.V. Radecki1 Department of Animal Science, Iowa State University, Ames, IA
50011, USA; 1Rutgers-The State University of New Jersey, New
Brunswick, NJ 08903, USA. In mammals, there is a marked developmental change in the somatotropic response to growth hormone (GH), with growth being predominantly GHdependent. In young poultry, growth appears to be largely (but not completely) GH independent. Moreover, GH administration to intact young chicks fails to evoke the marked increases in plasma concentrations of insulinlike growth factorI (IGFI) or IGFbinding proteins (IGFBPs) observed in mammals. There is evidence from the literature that the intermediate stages of growth are somewhat responsive to GH. However, there is no information on effects of GH on the somatotropic axis. The ability of the chronic administration of GH to influence plasma concentrations of IGFI and IGFBPs was examined in adult chickens. Circulating concentrations of IGFI and the putative avian IGFBP3 were elevated markedly (35fold) in both adult male and female chickens. Increases were observed within 2 days of treatment (via continuous infusion employing osmotic pumps) and plateau high concentrations of both were achieved by 5 days of GH administration. There were, however, little changes in body or organ weights. Concurrent with the changes in circulating concentrations of IGFI and the putative IGFBP3, there were marked increases in the expression of GH dependent mRNA (e.g., GH responsive gene product1). It is concluded that the adult chicken with its low ambient circulating concentrations of GH is very sensitive to GH. Effect of Cysteamine on the Plasma Concentration of Several Hormones Related to Growth in Growing GeeseA. Xiaojie, H. Zhengkang Laboratory of Animal Physiology and Biochemistry, Nanjing
Agricultural University, Nanjing, 210095 P.R. China It had been found that cysteamine(CS) could promote the growth of chickens and geese, but the mechanism was not clear. Twenty-four two-month-old crossbred geese (Chuan White X Tai Lake) were used to evaluate the effect of CS on several hormones in this study. The birds were allocated randomly into control and treatment group. CS was added in the diet of treatment group at the dosage of 100 mg/kg·BW on the first day of the experiment. Blood samples were collected on day three from the wing vein. Plasma hormone concentrations were measured by radioimmunoassay. In the treatment group, the somatostatin (SS) concentrations was significantly lower than that of the control group (P<0.05), while growth hormones (GH) and ß-endorphin (ß-END) levels were increased by 76.92% (P<0.01) and 44.55%(P<0.05). The thyrotropic hormone (TSH) level was reduced by 32.32%(P<0.05), while T3 and T4 were increased by 76.86%(P<0.01) and 14.72%(P>0.05) respectively. Insulin concentration was also increased by 14.69%(P<0.05). The results indicate that CS can decrease the plasma SS and TSH concentrations, but increase the levels of b-END, GH, T3, T4 and insulin, thereby inducing fast growth in geese. Growth Hormone (GH) Infusion Decreases HepaticGH Binding in Adult Male Chickens, but has Little Effect on Organ Weights, Circulating Metabolite Concentrations or Reproductive Hormones.S.V. Radecki, L.M. McCann-Levorse, J.a. Proudman1, C.G. Scanes2 Department of Animal Science, Rutgers-The State University of
NJ, New Brunswick, NJ, USA; 1USDA, Beltsville, MD, USA; 2College
of Agriculture, Iowa State University Ames, IA, USA Previously, we have shown that infusion of adult male White Leghorn chickens with recombinant chicken GH (rcGH) increases circulating concentrations of insulinlike growth factor (IGF)I. To further investigate the effect of GH in the adult chicken (White Leghorn roosters), two studies were conducted in which chickens were implanted with osmotic minipumps (Model 2ml, Alzet Corp., Palo Alto, CA) to deliver 100 ug rcGH/kg body weight/day (n=6). Unimplanted (study 1) or chickens implanted with pumps containing vehicle (study 2) served as controls. In study 1, blood was collected from chickens at 2, 5 and 10, and analyzed for glucose. In study 2, chickens were killed after 2, 5, and 10 days, at which time blood was collected and organs removed. Plasma was analyzed for nonesterified fatty acids (NEFA), triiodothyronine (T3),thyroxine (T4), prolactin (PRL), luteinizing hormone (LH), and follicle stimulating hormone (FSH). Organs were weighed, then frozen in liquid nitrogen. Hepatic GH binding was measured by a radioreceptor assay. Body, breast muscle, liver, spleen, thymus and testis weights were not altered by GH infusion. Abdominal fat pad weight was decreased after 10 days of GH infusion (vehicle: 26 +/4 (SEM); GH: 10 +/3 g; P < .05). Glucose and NEFA were unaffected by GH infusion, as were circulating concentrations of PRL, LH, and FSH. Although circulating concentrations of T3 were not different between treatments, on day 10, T4 was greater (P < .05), and the T3 to T4 ratio lower (P < .05) in GH treated roosters. Specific hepatic GH binding was decreased 31, 67 and 55% by GH infusion after 2, 5, and 10 (P < .05), respectively, as compared to vehicle treatment. We conclude that the GH axis is responsive in the adult, but the affect this axis has on metabolism is unclear. A Heterologous Radioimmunoassay to Measure Growth Hormone in the Ostrich (Struthio camelus).M. Mathiu1, W.H. Burke Department of Poultry Science, The University of Georgia,
Athens, GA 30602-2772, USA; 1Permanent address: University of
Nairobi, Department of Animal Physiology, P.O. Box 30197,
Nairobi, Kenya Studies were conducted to determine if reagents developed for measurement of chicken growth hormone (cGH) could be used to measure GH in the ostrich (Struthio camelus). Ostrich GH (oGH) preparations SC52RC, SC55B, SC35GBR and SC15CR were purified according to Papkoff et al. (Gen. Comp. Endocr. 48:181-195) and kindly provided by Dr. H. Papkoff. All of the oGH preparations reacted with an anti-chicken GH antibody (Dean et al., Growth Dev. Aging 57:57-72), but not with an anti-turkey GH antibody (Proudman and Wentworth, Gen. Comp. Endocr. 36:194-200) in western blots. Double antibody radioimmunoassays were established using the anti-cGH antibody in combination with either radioiodinated rcGH or SC52RC. All oGH preparations displaced iodinated rcGH, but the dose response curves appeared nonparallel to the rcGH standard. The 50% bound point of rcGH was about 1ng, but it ranged from 1.6 to 2.1 for the oGH's and 160ng for oPrl. All GH preparations effectively displaced iodinated SC52RC from the anti-chicken GH antibody with the 50% bound point of SC52RC, SC35GBR, SC55B, SC15CR and rcGH being .85, 1.1, 1.5, 1.5 and 2.2ng, respectively; all dose response curves were parallel. The minimum detectable dose of SC52RC was .198ng. The assay system using iodinated oGH was capable of measuring GH in 25µl samples of plasma from ostriches at all ages tested (10-19 weeks). Plasma levels of oGH were increased by i.v. administration of TRH (10µg/kg). Metabolic Responses of the Chicken to an Intravenous Injection of Either Chicken (CIGF-I) or Human Insulin-like Growth Factor-I (hIGF-I)J. McMurtry, G. Francis, T. Caperna, R. Vasilatos-Younken, R. Rosebrough, D. Brocht USDA, ARS, Growth Biology Lab, Beltsville, MD, USA The purpose of this study was to compare the metabolic and endocrine responses of male broiler chickens to an intravenous injection of either cIGFI, hIGFI or cIGFILR3 (an IGFI analogue). At 5 weeks of age, chickens were surgically implanted with a jugular cannula. Following an 8 h fast, equimolar amounts of cIGFI, hIGFI and cIGFILR3 (100, 102 and 126 mg/kg bw, respectively) were infused. Control birds were infused with an equal volume of saline. Plasma samples were collected at selected intervals pre and postinfusion and analyzed for various hormonal and metabolite parameters as well as insulinlike growth factor binding protein (IGFBP) activity. Similar hypoglycemic responses (50%) were induced by cIGFI and hIGFI, and to a lesser extent (30%) by LR3. A concomitant decline in insulin concentrations was noted. GH levels were differentially affected following growth factor treatment in that cIGFI and hIGFI were similarly depressed, while only minor decreases in GH were observed following LR3 treatment. Regardless of treatment, plasma T4 and T3 declined during the sampling period. Significant increases in plasma glucagon and free fatty acid concentrations were observed, while triglyceride levels were unaffected. Changes in IGFBP activity were noted. Similar metabolic and endocrine responses were elicited by cIGFI and hIGFI, while less dramatic changes in these parameters were induced by cIGFI LR3. Higher Stimulation of Dna Synthesis By Igf-1 in Muscle Cells From Chickens Selected for GrowthM.J. Duclos, B. Chevalier, J. Simon Endocrinologie de la Croissance et du Métabolisme, Station de
Recherches Avicoles, INRA, 37380 Nouzilly, France Our purpose was to compare the growth and metabolic potentials of muscle satellite cells (SC) prepared from two strains of chickens selected over several generations for high or low growth rate (HG and LG). SC were prepared by enzymatic digestion of pectoralis muscle from 1 day old male chicks and maintained in culture. DNA synthesis by proliferating SC was estimated by thymidine incorporation for 2 h following a 12 h incubation period in the absence or presence of fetal calf serum (FCS) or IGF1. Myotubes (MT) obtained following terminal differentiation and fusion of the SC were used for metabolic studies. Aminoacid uptake was measured using the non metabolizable analog aminoisobutyric acid (AIB) and protein synthesis was estimated by the incorporation of tyrosine into proteins. DNA synthesis by SC from both strains was stimulated in a dose dependent manner by FCS and by IGF1. Maximal stimulation by FCS (2.5%) or IGF1 (50 ng/ml) was significantly higher for HG than for LG (557 ±61% of basal vs 359 ± 38% for FCS and 502 ± 17% vs 305 ± 15% for IGF1). IGF1 also stimulated AIB uptake and protein synthesis by MT prepared from chickens of both strains. Stimulation of both parameters was dose dependent but the efficacy of IGF1 did not differ between HG and LG cells. Our data show that proliferating SC from HG chicks exhibit a markedly greater stimulation of DNA synthesis by FCS and IGF1 than SC from LG chicks. In contrast, MT exhibit a similar response to IGF1 for metabolic parameters. Divergent Selection of Japanese Quail for 4 Week Body Weight Alters the Somatotropic AxisM.C. McGuinness, D.L. Long, K.E. Nestor, W.L. Bacon. Department of Animal Sciences, The Ohio State University, Ohio
Agricultural Research and Development Center, Wooster, OH 44691,
USA Divergent selection for body weight (BW) might be expected to alter the expression of growth-related hormones, particularly the relationship between insulin-like growth factor-I (IGF-I) and growth hormone (GH). This hypothesis was tested using an experimental model that consisted of male and female Japanese quail selected for high or low 4 week BW, at generation 29. At the age of selection, the divergence in body weight from that of the randombred control line was 2-fold for either selected line (P<.05). There were no gender differences in plasma hormone levels, but there were significant genetic (line) differences (P<0.05). Plasma GH was dramatically elevated in low BW birds at 3 wk, and decreased linearly with age. IGF-I in plasma was maintained at high and constant levels throughout the sampling period in low BW birds, but declined with age and the onset of sexual maturity in high BW birds. Plasma prolactin profiles followed a similar pattern. Hepatic expression of IGF-I mirrored the relationship among lines for plasma IGF-I levels. However, plasma GH levels and hepatic GH receptor expression were inversely related, with very low GH receptor expression in low BW birds and high levels of expression in high BW birds. Therefore, elevation of plasma GH and IGF-I levels and enhanced hepatic IGF-I gene expression are negatively associated with selection for BW. Examination of the quail IGF-I gene in the selected lines identified RFLP's in the peptide coding region, indicating alterations of gene structure. Effect of Anesthesia on GH Secretion and Gene Expression in Meat-Type and Laying-Type ChickensR.Q. Zhao1, E. Mühlbauer, E. Decuypere2, R. Grossman Institute for Small Animal Research, Federal Research Centre
of Agriculture, Celle, Germany; 1Laboratory for Animal Physiology
and Biochemistry, Nanjing Agricultural University, Nanjing, P.R.
China; 2Laboratory for Physiology of Domestic Animals, Catholic
University Leuven, Heverlee, Belgium The effect of anesthesia on plasma GH concentrations and pituitary GHmRNA content was compared between meattype (Broiler) and layingtype (White Leghorn) chickens at 5 days of age. Birds were catheterized via the jugular vein at day 3 and serial blood samples were taken at 20min intervals for 100 minutes both in conscious and anaesthetized (2.5 mg/g B.W., 10% urethane, i.P.) animals. GH concentrations in plasma were measured by RIA and GHmRNA content in pituitaries was quantified by Northern blot analysis. In the layer, urethane anesthesia significantly reduced (P<0.001) GH plasma concentrations. Peak amplitude and nadir were significantly decreased (P<0.001) under anesthesia. The pulsatile pattern of GH secretion was almost completely abolished. On the contrary, there were no significant changes in overall mean plasma GH concentrations in the broiler, although a slightly reduced GH peak amplitude was observed under anesthesia. An obvious upregulation (P<0.05) of GHmRNA content in the pituitary was found one hour and four hours after the beginning of anesthesia in male and female layers; the males responded faster (within one hour) compared to the females (within four hours). In the broiler upregulation of GH gene expression under anesthesia occurred only in females, but not in males. These observations indicate that GH secretion and GH gene expression is regulated differently between the two strains of chickens. |