Effect of Incubator Temperature and Oxygen Concentration at the Plateau Stage in Oxygen Consumption on Turkey Embryo Muscle Growth and Development 1

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1 Interntionl Journl of Poultry Science 6 (6): , 007 ISSN Asin Network for Scientific Informtion, 007 Effect of Incutor Temperture nd Oxygen Concentrtion t the Plteu Stge in Oxygen Consumption on Turkey Emryo Muscle Growth nd Development 1 V.L. Christensen, M.J. Winelnd, J.L. Grimes, E.O. Oviedo, P.S. Mozdzik, D.T. Ort nd K.M. Mnn Deprtment of Poultry Science, College of Agriculture nd Life Sciences, North Crolin Stte University, Rleigh, North Crolin , USA Astrct: It ws hypothesized tht incutor temperture nd oxygen concentrtion ffect emryo muscle development. Turkey eggs were incuted until the 4th dy of development. At the eginning of the 4th dy, the eggs contining vile emryos were rndomly divided into 4 groups immeditely prior to the plteu stge in oxygen consumption. Four experimentl cinets ccommodting pproximtely 100 eggs were used for the ctul htching process. Ech cinet operted t predetermined tempertures (TEM) nd oxygen concentrtions (O ) in TEM (36 nd 39 C) x O (17 nd 3%) fctoril rrngement. At 7 nd 8 dys of development, immeditely following the plteu stge, 10 emryos or poults were smpled from ech of the 4 cinets. Blood ws otined following decpittion. From ech crcss the pipping (musculus complexus), rest (pectorlis thorcicus) nd thigh muscles (gstrocnemus) were collected. Muscles were plced into n pproprite volume of 7% perchloric cid preprtory to ssying for glycogen nd lctte. Five irds were smpled for histologicl nlyses of muscle fiers. Plsm Cretine Kinse (CK) nd lctte dehydrogense (LDH) ctivities were mesured. TEM nd O ffected muscle growth differently. High TEM nd O ffected pipping nd thigh muscle weights ut not tht of the rest muscle. Only TEM ffected rest muscle weights. Muscle function ws ffected differently when emryos were exposed to TEM nd O. The CK nd LDH ctivities were lso ffected t 7 dys 39 C cusing elevted CK nd LDH ctivities compred to 36 C. At 8 dys, only CK ws ffected s 39 C elevted CK ctivity in the 3% oxygen environment ut not in the 17% environment. Thus, incutor conditions my ffect muscle development nd function in poult emryos Key words: incutor temperture, oxygen concentrtion, emryo muscle development Introduction Incutor temperture (TEM), oxygen (O ) concentrtion (Metclfe et l., 1981; Mlty nd Stricklnd, 004) nd sex (Vellemn nd Nestor, 004) my ffect poult emryo muscle development. Development of the tht overll, erly postntl muscle development is the most criticl time period for stellite cell mitotic ctivity. Little is known of the physiology nd development of muscles in emryos when incutor conditions re not idel. We proposed the hypothesis tht incutor emryo muscle egins with the fusion of temperture, oxygen concentrtions nd emryo sex t mononucleted myolsts to form myotues, which mture into myofiers (Shultz nd McCormick, 1994). Feldmn nd Stockmn (199) showed tht during the plteu stge in oxygen consumption my interct to ffect muscle physiology nd growth of thigh, pipping nd rest muscles of developing turkey emryos. vin development distinct popultion of stellite cells is present t the midfetl stges of development nd y Mterils nd Methods lte emryogenesis myolsts found in the chicken Approximtely 500 fertilized turkey eggs (Nichols) were emryo re predominntly dult myolsts (Hrtley et otined from commercil flock (Prestge Frms, l., 199). Mlty et l. (004) showed the numer of Clinton, NC) for ech of three experiments. The eggs fiers in turkey emryo my e incresed y using were incuted (Ntureform I14, Jcksonville, FL) for 4 incresing incutor temperture t mid emryonic dys using stndrd industry procedures. The cinet stges. Once the ird is htched nd myofiers re operted t set point of 37.5 C dry ul temperture formed, norml skeletl muscle growth occurs through nd RH of 53%. Eggs contining vile emryos on n increse in myofier size, rther thn n increse in the 4th dy of development were rndomly divided into myofier numer (Remignon et l., 1995; Moore et l., four groups immeditely prior to the plteu stge in 005). Recently Vellemn et l. (00; 003) reported oxygen consumption. Four experimentl cinets tht the inheritnce of rest muscle is sex-linked nd ccommodting pproximtely 100 eggs were used for the dm is more responsile t specific ges thn is the the htching process. Ech cinet contined one sire. Thus, sex my e n dditionl fctor in muscle incutor try nd ws regulted y thermistors development of emryos. Moore et l. (005) concluded connected to microprocessors with temperture 406

2 Christensen et l.: Emryo Muscles sensitivity of±0.05 C. Humidity ws controlled y clculted y dividing the totl glycogen mesured in similr system using reltive humidity sensors. Digitl muscle y the totl lctte mesured in the sme thermometers (Cox, Lexington, NC) were used with ech muscle. cinet to verify set point tempertures. Atmospheric In ech of the tretments descried ove, the pectorlis gses were mesured y using n infrred detector for thorcicus ws hrvested from n dditionl 5 poults per cron dioxide (Engelhrd Ventostt 001V, Iselin, NJ tretment nd fixed overnight with uffered neutrl 08830) nd electrochemicl cells for oxygen (Teledyne, formldehyde. The following morning the tissues were Los Angeles CA 90064), ech with sensitivity of±0.1%. dehydrted, wshed nd emedded in prffin. Eight Mnully operted vlves infused gses into ech of the micron thick cross sections were cut on microtome cinets nd the flow rte ws djusted sed on the nd dhered to glss slides. Tissues were dewxed sensors to crete the desired gs concentrtions. Three nd stined using stndrd hemtoxylin nd eosin experiments were conducted using the experimentl procedures. A Leic DMR (Leic Microsystems, cinets. Experiment 1 nd were preliminry Bnnockurn, IL) microscope ws used to oserve the experiments to determine the effects of temperture or tissue sections. A Spot-RT CCD (Dignostic oxygen on muscle development. Experiment 3 exmined Instruments Inc., Sterling Heights, MI) cmer ws used the interctive effects on muscle of the most effective to cpture imges of ech of the cross sections. A nd lest effective for ffecting muscle weights due to rndomly selected re on ech slide ws selected to temperture nd oxygen tretments determined in mesure the dimeter of 100 fiers/section. Experiments 1 nd. Oxygen: Four O concentrtions were the tretments in Temperture: The eggs were moved to the tretment th the second preliminry tril. All procedures to the 4 cinet t the eginning of the 4th dy of development. dy of development were the sme s in Experiment 1. The 4th dy for poult emryos is the eginning of the The O concentrtions within the cinets were 17, 19, plteu stge in oxygen consumption (Rhn, 1981). All 1 or 3% of the tmosphere. The frctionl infertile eggs nd nonvile emryos were removed concentrtion t se level (Rleigh, NC) corresponded prior to trnsfer to one of four experimentl cinets. At to oxygen prtil pressures of 19, 144, 160 nd 175 the eginning of the 4th dy the eggs were incuted mm Hg, respectively. Concentrtions lower thn mient t one of the tretment TEM (36, 37, 38 or 39 C) to oxygen concentrtions (0.9%) were mintined y expose the emryos to different tempertures t the infusing nitrogen gs into the cinet t rte tht plteu stge. resulted in the desired concentrtion of 17 or 19% O. Concentrtions were mesured with n oxygen meter Tissue smpling: Ten emryos or poults were selected nd flow rtes from oxygen or nitrogen storge tnks rndomly from ech incutor t the end of the tretment were djusted to mintin the desired oxygen level. period s emryos were externlly pipping during the Emryo smples were collected nd nlyzed s in th 7 dy or htching on 8 dys of development. The Experiment 1. poults were decpitted nd trunk lood ws collected into tue contining 10 mg EDTA. Poult ody weights Temperture nd oxygen: The most nd lest effective were recorded (nerest 0.1g) with nd without yolk nd TEM (36 nd 39 C) nd O tretments (17 nd 3%) for muscles were dissected nd weighed (nerest 0.01 ltered physiology nd the reduction of muscle weight in mg) s quickly s possile. The lood ws centrifuged the preliminry experiments were comined in fctoril (700 x g) for 15 min under refrigertion (4 C). The rrngement for the third experiment. All tretments plsm ws decnted nd frozen (- C) for nlysis of were s descried previously. Fertilized eggs were Cretine Kinse (CK) (Oliver, 1955) nd lctte gin incuted 4 dys when vile emryos were dehydrogense (LDH) (Wcker et l., 1956) ctivities ssigned rndomly to one of the four cinets. The tht my e indictive of muscle oxygen det (Wilson et conditions were TEM of 36 C with 17 or 3% O nd l., 1990; Vellemn nd Nestor, 004). Ech smple 39 C with 17 or 3% O in fctoril rrngement. Birds ws sexed y visul inspection following dissection. were smpled identiclly to the previous experiments. From ech smple the pipping (musculus complexus), rest (pectorlis thorcicus) nd thigh muscles Sttisticl nlysis: Dt for ll three experiments were (gstrocnemus) were collected. The three muscles were nlyzed using the generl liner models procedure weighed (nerest 0.01g) nd plced into n pproprite (SAS Inc., 1998). Experiments 1 nd were rrnged s volume (5 ml/g of muscle) of cold (4 C) 7% perchloric four levels of TEM or O tretments y two sexes (hens cid. The plsm smples were cpped in storge vils nd toms) y two dys of development (7 nd 8 d) nd kept t 4 C until ssyed for glycogen (Dreiling et fctoril. In Experiment 3, the dt were rrnged s two l., 1987) nd lctte (Henry, 1968; Trinder, 1969). TEM y two O concentrtions y two sexes fctoril. Rtios of glycogen to lctte in ech muscle were s differing significntly were seprted y the lest 407

3 Christensen et l.: Emryo Muscles Tle 1: Body nd yolk weights (g) of turkey emryos exposed to different oxygen concentrtions nd tempertures during the plteu stge of development Oxygen (%) Temperture ( C) 7 dy emryo 8 dy poult BW without yolk ±SEM 53.5± ±0.4 Proilities NS TEM = Yolk ±SEM 14.6±0. 9.8±0.3 Proilities O = O = TEM = s in column with different superscripts re significntly different (p<0.05) squre mens procedure. s in tles re lest squre mens. All possile min nd interction effects were tested. All proilities were sed on p<0.05 unless otherwise noted. Results Temperture nd oxygen trils: The 39 C TEM nd 17% O reduced pipping nd thigh muscle weights compred to 36 C nd 3% O respectively. However,, weights of rest muscles were not ffected y O s only TEM ffected only rest muscle weights. To prevent redundncy in the pper, no dt will e reported from the initil two trils ecuse their ojective ws to provide effective TEM nd O vlues for ffecting muscle growth nd physiology. Dt from the initil tril indicted tht poults muscle growth nd physiology responded mximlly to 39 nd 36 C. Thus, these two TEM were selected for the third tril. Dt from the second tril indicted tht 17 nd 3% O were the oxygen tretments tht elicited the mximl responses of muscle growth nd physiology. Thus, the 17 nd 3% O tretments were used in the third tril. Temperture with oxygen: There were no significnt effects due to sex s fctor or intercting with other fctors in the third tril so no sex effects will e reported. Only TEM ffected ody weights s 36 C incresed weights compred to 39 C t 8 d of development (Tle 1). Both O nd TEM ffected residul yolk mss s 17% O nd 39 C incresed residul yolk weight compred to 3% O nd 36 C, ut the two fctors did not interct (p>0.05). Muscle weights t externl pipping (7d) were decresed y 3% O compred to 17% nd TEM nd O intercted t htching to ffect the pipping muscle weight (Tle ). Using higher incutor TEM in comintion with 17% O incresed pipping muscle weights in the htched poult. Emryo rest muscles were hevier t 7 dys in 39 C thn in 36 C, ut no differences were noted in poults due to either O or TEM At htch. The thigh muscle weight ws ffected only t htching s 3% O incresed weights compred to 17% nd 39 C depressed weights compred to 36 C, ut the two fctors did not interct. The rtio of pipping muscle glycogen to lctte displyed n O y TEM interction t 7 dys s 39 C decresed the rtio in the 3% O compred to 36 C ut not in the 17% cinets (Tle 3). No effects were seen t dy 8. The rtio of rest muscle glycogen to lctte ws depressed y higher TEM t 7 dys nd y greter O concentrtion t dy 8. O nd TEM intercted to ffect glycogen to lctte rtios in poult thigh muscles. In 3% O environment, 36 C elevted glycogen to lctte rtios compred to 39 C, ut in 17% O environment, the reverse ws noted. At 7 dys 39 C elevted CK nd LDH ctivities compred to 36 C. At 8 dys, only CK ws ffected s 39 C elevted CK ctivity in the 3% O environment ut not in the 17% environment (Tle 4). TEM nd O intercted to ffect rest muscle fier dimeter in poults htching from the tretments. When exposed to 17% O concentrtion, 39 C suppressed muscle dimeter compred to 36 C. When exposed to 3% O, no differences were noted etween the TEM tretments (Tle 5). 408

4 Christensen et l.: Emryo Muscles Tle : Muscle weights (% of ody mss) of turkey emryos exposed to different oxygen concentrtions nd tempertures during the plteu stge of development Oxygen (%) Temperture ( C) 7 dy emryo 8 dy poult Pipping muscle ±SEM 1.49± ±0.03 Proilities O = O x TEM = Brest muscle ±SEM 5.70± ±0.03 Proilities TEM = NS Thigh muscle ±SEM 4.78± ±0.06 Proilities NS O = TEM = s in column with different superscripts re significntly different (p<0.05) Tle 3: Rtio of muscle glycogen (mg) to lctte (mg) of turkey emryos exposed to different oxygen concentrtions nd tempertures during the plteu stge of development Oxygen (%) Temperture ( C) 7 dy emryo 8 dy poult Pipping muscle c ±SEM 4.35± ±0.14 Proilities O x T = NS Brest muscle ±SEM.16± ±0.06 Proilities T = O = Thigh muscle c ±SEM.71± ±0.0 Proilities NS O x TEM = c s in column with different superscripts re significntly different (p<0.05) 409

5 Christensen et l.: Emryo Muscles Tle 4: Plsm cretine kinse (CK) nd lctte dehydrogense (LDH) ctivities of turkey emryos exposed to different oxygen concentrtions nd tempertures during the plteu stge of development Oxygen (%) Temperture ( C) 7 dy emryo 8 dy poult CK (U/L) ,56, ,411, ,155,59 39,133,84 ±SEM 1,488±11,579±100 Proilities TEM = O x TEM = , ,77 LDH (U/L) ±SEM 415±13 696±5 Proilities TEM = NS s in column with different superscripts re significntly different (p<0.05) Tle 5: Brest muscle fier dimeters of turkey emryos exposed to different oxygen concentrtions nd tempertures during the plteu stge of development Oxygen Temperture Brest dimeter (µm) c ±SEM.77±0.14 Proilities TEM O TEM x O c s in column with different superscripts re significntly different (p<0.05) Discussion In vin emryos development of muscle fiers egins with the fusion of mononucleted myolsts to form myotues, which mture into myofiers (Shultz nd McCormick, 1994). A distinct popultion of stellite cells is present t the midemryonic stges of development nd y lte emryogenesis myolsts found in the chicken emryo re primrily dult myolsts (Hrtley et l., 199). Once the ird is htched nd myofiers re formed, norml skeletl muscle growth occurs through n increse in myofier size, rther thn n increse in myofier numer (Remignon et l., 1995). Myofier numer does not increse during norml postntl growth ecuse myonuclei re post mitotic nd cnnot synthesize DNA (Stockdle nd Holtzer, 1961). Mlty nd Stricklnd (004) suggested tht incutor temperture t criticl times of turkey emryo development my ffect the numer of muscle fiers. Thus, the numer nd size of fiers tht poult hs when it htches my e importnt in the overll muscle development of the ird t mrket ge (Moore et l., 005). Inheritnce of rest muscle is sex-linked in turkeys so the ird sex my e determinnt of muscle mturtion t htching s well (Vellemn et l., 00, 003). The hypothesis tested in the current reserch ws tht incution temperture nd oxygen concentrtions nd emryo sex t the plteu in oxygen consumption ffect the formtion of muscles nd their physiology. The hypothesis ws tested specificlly t the plteu stge in oxygen consumption, time when muscle tissue my lck sufficient nutrients (Dietz et l., 1998; Christensen et l., 1999). In this study, incutor temperture nd cinet oxygen concentrtions effects on the development of ech of the three muscles were followed. The fctors were primrily independent, so interctions of O nd TEM did not lter the growth nd physiology of rest muscle. Ech muscle rected differently to the TEM nd O tretments. Temperture is the mjor regultory fctor in egg incution nd emryo development ecuse of its effect on generl growth nd the coincidentl impct on tissue formtion (Romnoff nd Fer, 1933). Temperture mnipultion t vrious times throughout o incution (38.5 C during dys 5-8 nd dys 9-1 of development) cn lter turkey emryo muscle development (Mlty et l., 004). Mlty et l. (004) suggested tht the nuclei nd muscle fier numers in the posthtch turkey could e incresed y temperture nd Mlty nd Stricklnd (004) reported tht temperture mnipultions ffected the posthtch phenotype through the ltertion of muscle estlishment in the emryo, ut the pthwys seemed oscure. A lower temperture incresed gene expression of muscle t dys 16 nd 17 of development; however, if tempertures were higher (38.5 C) the expression ws delyed until following dy 1. In the current study we sw no increse in fier numers, ut 36 C compred to 39 C did increse rest muscle fier dimeter when the eggs were incuted in hypoxic environment. 410

6 Christensen et l.: Emryo Muscles Little is known of the effect of oxygen on muscle cell ech muscle is very different t the vrious emryonic expression, ut it hs een suggested tht oxygen my stges of development. The pipping muscle my e e the primry determinnt of ody nd orgn growth in contrcting wheres the rest muscle cn e less the emryo (Metclfe et l., 1981) nd hypoxi cn lter ctive t given stge of development. It is noteworthy the crohydrte metolism of growth-selected tht ll muscles continued to grow through the plteu turkeys differently thn tht of controls (Christensen et stge despite physiologicl perturtions. l., 1999). The impct of temperture nd oxygen on Gluconeogenesis requires recycling of lctte vi the emryo development is highly importnt ecuse of the proliferting pool of muscle cells ville in lte incution. In the current study TEM higher thn 38 C nd O concentrtions elow 1% t the plteu ffected muscle growth nd physiology. Further study will e required to determine the long term effects of TEM nd O on muscle growth nd physiology. Becuse of the decresed frctionl percentge of ir cell oxygen during the finl stge of emryonic development (Rhn, 1981), the emryo relies primrily on neroic metolism for energy. Blood glucose increses stedily during this stge nd tissue glycogen stores decrese (Freemn, 1965). The glucose from mternl investment in eggs is miniml y this time (Hzelwood nd Lorenz, 1959). Becuse the egg contins so little glucose, glycogen is synthesized nd stored in muscles erlier in incution s source of glucose in preprtion for the hypoxi of htching (Freemn, 1965). Gluconeogenic processes provide the necessry crohydrte for glycogen storge. The liver nd kidney re essentil to this process ecuse they convert lctte into glucose-6-phosphte when lood glucose levels decline (Wtford et l., 1981). Hert nd skeletl muscle in vin species lck the ility to recycle lctte (Wtford et l., 1981). Therefore, the liver nd kidney re supply orgns in this sense nd oth hert nd skeletl muscles re demnd orgns. Anything tht interferes with overll energy metolism my ffect skeletl muscle growth nd physiology s well. Previously it ws shown tht incutor temperture nd oxygen t the plteu stge ffect the crdiovsculr nd digestive systems (Christensen et l., 004). Our dt my e the first to show effects on emryo muscle development nd physiology t the plteu stge in oxygen consumption. We suggest tht ech muscle hs unique mechnism to respond to hypoxi or hyperoxi to control growth nd physiology. An exmple of uniqueness cn e seen y compring the results of the pipping nd rest muscles. Pipping muscle responded to oth incresed TEM nd decresed O y incresing its size nd incresing its rtio of glycogen to lctte. Conversely, the rest muscle weight decresed nd the rtio of glycogen to lctte declined. Severl possiilities exist to explin these oservtions. One possiility my e the muscle types. One is primrily fst-twitch wheres the other is slow-twitch muscle nd ech muscle type my hve different oxygen requirement. A second explntion my e the ctivity of Cori cycle or ctolism of existing tissues or dditionl ctolism of ville nutrients in residul yolk (Donldson nd Christensen, 1991) nd occurs differently in turkeys selected for incresed growth (Christensen et l., 1999). If greter mturtion could e ttined prior to htching, the poult in modern-type turkeys my e etter le to grow muscle nd hve full muscle function even in n oxygen deficient environment (Dietz et l., 1998). Eggs incuted t 36 C with greter thn 1% O hd more glycogen thn lctte in their muscles indicting positive energy lnce in those groups. The presence of greter mounts of glycogen my indicte metolic djustments to environmentl conditions tht could led to muscle dmge. Elevted CK nd LDH ctivities re chrcteristic of noxi nd hypercpni in nimls followed y concomitnt increse in gluconeogenesis (Oliver, 1955; Henry, 1968). Cretine phosphte crries high-energy phosphte, which cn e trnsferred to nd from ATP y CK in reversile rection. It is source of energy when other mens of supplying ATP to muscle re indequte (Lio et l., 1996). Simply stted, significnt decreses in the speed t which the phosphoryltion of ATP from phosphocretine were found in muscles from crdiomyopthy turkeys. LDH ctlyzes the reversile rection etween pyruvte nd lctte tht is essentil in gluconeogenesis. Elevted CK nd LDH hve een suggested to e correltes of skeletl muscle dmge (Wilson et l., 1990; Vellemn nd Nestor, 004). The 39 C TEM elevted oth CK nd LDH t pipping ut CK ws ffected only y TEM t htching. Both of these oservtions indicte tht oxygen det nd rpid lctte recycling were occurring due to elevted TEM. It is lso noteworthy tht incresed CK ctivity due to elevted TEM occurred t htching only in the presence of 3% O. In conclusion, we present the first evidence known to the uthors showing the criticl importnce temperture nd oxygen t the plteu stge of turkey emryo development for muscle development. Incutor tempertures nd oxygen concentrtions oth ffected growth nd function of muscles. It is recommended for optiml muscle growth nd function, turkey eggs should e incuted t less thn 37 C nd in greter thn 1% oxygen t the plteu stge in oxygen consumption. Acknowledgements We thnk US Poultry nd Egg Assocition for funding these studies. 411

7 Christensen et l.: Emryo Muscles References Christensen, V.L., W.E. Donldson nd K.E. Nestor, Effect of supplementl oxygen on lood plsm orgnic cids within emryos from selected lines of turkeys. Poult. Sci., 78: Christensen, V.L., M.J. Winelnd, I. Yildrum, D.T. Ort nd K.M. Mnn, 004. Incutor temperture nd oxygen concentrtion t the plteu stge ffects intestinl mturtion of turkey emryos. Int. J. Poult. Sci., 3: Christensen, V.L., M.J. Winelnd, I. Yildrum, D.T. Ort nd K.M. Mnn, 004. Incutor temperture nd oxygen concentrtion t the plteu stge ffect crdic helth of turkey emryos. J. Anim. And Vet. Advnces, 3: Dietz, M.W., M. vn Kmpen, M.J.M. vn Griensven nd S. vn Mourik, Dily energy udgets of vin emryos: The prdox of the plteu phse in egg metolic rte. Physiol. Zool., 71: Donldson, W.E. nd V.L. Christensen, Dietry crohydrte levels nd glucose metolism in turkey poults. Comp. Biochem. Physiol., 98A: Dreiling, D.E., D.E. Brown, L. Csle nd L. Kelly, Muscle glycogen: Comprison of iodine inding nd enzyme digestion ssys nd ppliction to met smples. Met Sci., 0: Feldmn, J.L. nd F.E. Stockmn, 199. Temporl ppernce of stellite cells during myogenesis. Dev. Biol., 153: Freemn, B.M., The importnce of glycogen t the termintion of the emryonic existence of Gllus domesticus. Comp. Biochem. Physiol., 8: Hrtley, R.S., E. Bndmn nd Z. Ylonk-Reuveni, 199. Skeletl muscle stellite cells pper during lte chicken emryogenesis. Dev. Biol., 153: Hzelwood, R.L. nd F.W. Lorenz, Effects of fsting nd insulin on crohydrte metolism of the domestic fowl. Am. J. Physiol., 197: Henry, R.J., Pges in: Clincl Chemistry- Principles nd Technics. Hrper nd Row, New York, NY. Lio, R., L. Nscimen, J. Friederich, J. K. Gwthmey, nd J. S. Ingwll, Decresed energy reserve in n niml model of dilted crdiomyopthy. Circultion Res. 78: Mlty, V., A. Somiy, N.A. French nd N.C. Stricklnd, 004. In ovo temperture mnipultion influences post-htch muscle growth in the turkey. Br. Poult. Sci., 45: Mlty, V. nd N.C. Stricklnd, 004. Mnipulting incution temperture lters muscle development in the turkey. Pges 1-13 in: Astrcts from Incution nd Fertility Res. Group, Univ. of Lincoln, UK, Septemer 004. Metclfe, J., I.E. McCutcheon, D.L. Frncisco, A.B. Metzenerg nd J.E. Welsh, Oxygen vilility nd growth of the chick emryo. Resp. Physiol., 46: Moore, D.T., P.R. Ferket nd P.E. Mozdzik, 005. Muscle development in the lte emryonic nd erly posthtch poult. Int. J. Poult. Sci., 4: Oliver, I.T., A spectrophotometric method for the determintion of cretine phosphokinse nd myokinse. Biochem. J., 61: Rhn, H., Gs exchnge of vin eggs with specil reference to turkey eggs. Poult. Sci., 60: Remignon, J., M.F. Grdhut, G. Mrche nd F.H. Ricrd, Selection for rpid growth increses the numer nd the size of muscle fires without chnging their typing in chickens. J. Musc. Res. Cell. Mot., 16: Romnoff, A.L. nd H.A. Fer, Effect of temperture on the growth, ft nd clcium metolism nd mortlity of the chick emryo during the ltter prt of incution. J. Cell. Comp. Physiol., : SAS Institute, SAS/STAT Guide for Personl Computers. Version 6 Edition. SAS Institute, Cry, NC. Shultz, E. nd K.M. McCormick, Skeletl muscle stellite cells. Re. Phys. Biochem. Phrm., 13: Stockdle, F.E. nd H. Holtzer, DNA synthesis nd myogenesis. Exp. Cell Res., 4: Trinder, P., Determintion of glucose in lood using glucose oxidse with n lterntive oxygen cceptor. Ann. Clin. Biochem., 6: 4. Vellemn, S.G., C.S. Coy, J.W. Anderson, R.A. Ptterson nd K.E. Nestor, 00. Effect of selection for growth rte on emryonic rest muscle development in turkeys. Poult. Sci., 81: Vellemn, S.G., J.W. Anderson nd K.E. Nestor, 003. Possile mternl inheritnce of rest muscle morphology in turkeys t sixteen weeks of ge. Poult. Sci., 8: Vellemn, S.G. nd K.E. Nestor, 004. Inheritnce of rest muscle morphology in turkeys t sixteen weeks of ge. Poult. Sci., 83: Wcker, W.E.C., D.D. Ulmer nd B.L. Vlee, Metlloenzymes nd myocrdil infrction. New Englnd J. Med., 55: Wtford, M.Y. Hod, Y. Chio, M. Utter nd R.W. Hnson, The unique role of the kidney in gluconeogenesis in the chicken. J. Biol. Chem., 56: Wilson, B.W., P.S. Nieerg nd R.J. Buhr, Turkey muscle growth nd focl myopthy. Poult. Sci., 69: The mention of trde nmes in this puliction does not imply endorsement of the products mentioned nor criticism of similr products not mentioned. 41