Effects of Post-Exercise Supplements on Glycogen Repletion in Skeletal Muscle*

Transcription

1 Veterinry Therpeutics Vol. 3, No. 3, Fll 2002 Effects of Post-Exercise Supplements on Glycogen Repletion in Skeletl Muscle* Joseph J. Wkshlg, MS, DVM Kimberly A. Snedden, MS Annette M. Otis, MS Christopher A. Kennedy, DVM Todd P. Kennett, BS Jnet M. Scrlett, DVM, PhD b Frncis A. Kllfelz, DVM, PhD Gry M. Dvenport, PhD b Arleigh J. Reynolds, DVM, PhD Gregory A. Reinhrt, PhD c Cornell University College of Veterinry Medicine Deprtment of Clinicl Sciences b Deprtment of Popultion Medicine Ithc, NY c The IAMS Compny Reserch nd Development Lewisburg, OH ABSTRACT Post-exercise crbohydrte supplementtion hs been routinely used to enhnce glycogen concentrtions in skeletl muscle, prticulrly during multiple-dy thletic events. Consumption of protein hydrolystes mixed with crbohydrte supplements during the post-exercise period my increse insulin response nd cuse glycogen repletion in skeletl muscle. A group of Alskn sled dogs were selected to exmine post-exercise supplementtion in pired crossover study design. The dogs were subjected to the sme exercise regimen *This study ws supported by generous grnt from the IAMS Compny, Lewisburg, OH. nd provided one of three tretments wter, glucose polymers, or glucose polymers with protein hydrolystes over 2-month period. Prmeters tested t vrious post-exercise time points included plsm insulin, glucgon nd glucose concentrtions, nd skeletl muscle glycogen content to gin better understnding of glucose metbolism nd glycogen repletion. The results showed n enhnced insulin, glucose, nd glucgon response immeditely fter supplementtion nd significntly incresed glycogen concentrtions in skeletl muscle within 24 hours when dogs received either of the glucose-contining supplements compred with wter lone. There were no 226

2 J. J. Wkshlg, K. A. Snedden, A. M. Otis, C. A. Kennedy, T. P. Kennett, J. M. Scrlett, F. A. Kllfelz, G. M. Dvenport, A. J. Reynolds, nd G. A. Reinhrt differences in the plsm prmeters or skeletl muscle glycogen stores in dogs provided the glucose polymers lone or the glucose polymers plus protein hydrolystes. Thus, post-exercise crbohydrte supplementtion incresed muscle glycogen repletion, but inclusion of protein hydrolystes in the supplements provided no dditionl benefits. INTRODUCTION Post-exercise crbohydrte supplementtion or vrible dietry crbohydrte intke hs long been suggested s strtegy for resolving muscle glycogen depletion in thletic dogs, prticulrly rcing Greyhounds nd Alskn sled dogs. 1 3 Findings in studies of glucose metbolism 4,5 verify the use of crbohydrte supplementtion for immedite oxidtion nd glycogen repletion during prolonged exercise. Results of field studies conducted in the mid- 1990s 6 suggested tht immedite post-exercise crbohydrte supplementtion in rcing sled dogs could substntilly increse muscle glycogen content within 4 hours. These nd other findings in humn thletes 7 9 suggest tht providing crbohydrte-contining supplements within 4 hours fter exercise confers n dvntge in multiple-dy events, bsed on incresed cpcity for insulin-independent glycogen repletion in skeletl muscle. More recently, it hs been suggested tht the ddition of protein to post-exercise crbohydrte supplementtion my enhnce muscle glycogen repletion, 10,11 possibly becuse enhnced insulin relese is induced by mino cid bsorption from the ingested protein. 12 However, results of studies using post-exercise crbohydrte nd protein supplementtion hve yielded mixed results, despite the direct correltion between enhnced insulin relese nd protein ingestion. 13 Previous studies 6 hve shown tht sled dogs cn benefit from postexercise crbohydrte supplementtion. Therefore, n dditionl controlled experiment ws conducted with tredmill-trined sled dogs to ssess the effects of crbohydrte supplementtion with or without dditionl protein on muscle glycogen repletion. The gol of this study ws to determine the hormonl (insulin nd glucgon) nd biochemicl (serum glucose nd muscle glycogen) responses using current feeding nd supplementtion methods. Functionl thletic dogs tend to be fed diets reltively high in protein once dily; therefore, it ws hypothesized tht the use of protein in post-exercise supplementtion my enhnce muscle glycogen repletion becuse of the incresed gluconeogenetic cpcity of crnivores. 14 The experimentl design nd model llowed controlled investigtion of the effects of vrious post-exercise supplements in optimizing muscle glycogen repletion nd hormonl responses when dogs re exercised. MATERIALS AND METHODS Dogs Ten tredmill-trined, ctive, purposely bred sled dogs were housed in US Deprtment of Agriculture pproved fcility nd were ccepted for use in this study by the Cornell University Institutionl Animl Cre nd Use Committee. The dogs were fed commercil performnce diet (Euknub Premium Performnce Diet, Ims) contining 32.2% protein, 21.5% ft, 35.5% crbohydrte, 3% fiber, nd 7.5% sh (dry-mtter bsis) during the 3-month period preceding the study nd throughout the study; this protocol exceeded Assocition of Americn Feed Control Officils stndrds for mintining n dult dog. The dogs were trined regulrly (n verge of 3.7 times week) on the tredmill nd in tril running (up to 8 miles per session t speeds rnging from 12 to 16 miles per hour) for 2 months before the study ws initited. Ech dog served s its own control in the 227

3 Veterinry Therpeutics Vol. 3, No. 3, Fll 2002 crossover design followed in the study. The tretment groups were rndomly chosen, s ws the order of tretments. To ensure tht no crossover effects occurred due to tretments, ech experiment ws scheduled t 21-dy intervls, with ech dog being evluted fter ingesting ech of the two nutritionl supplements or wter. In between experimentl sessions, the dogs were exercised normlly, ccording to the protocol lredy designted. To ensure mximl glycogen content in the muscle, the dogs were given 72 hours to recover from the most recent exercise session before the experiment ws conducted. Tretment nd Experimentl Exercise Dogs were exercised in tndem for 30 minutes on tredmill while in hrness designed to mimic the norml pulling ctions of tril running. The tredmill speed ws set t 6.5 m/sec t 1% slope. Within minutes fter completing the exercise protocol, ech dog ws provided one of three supplements: 1.5 g of glucose polymers/kg, with 100% of clories s glucose polymers (GP; Polycose, Abbott Lbortories; 99.7% crbohydrte s dry mtter, 100% clories s GP), in 400 ml of wter; 1.5 g of GP/kg plus 0.5 g of egg (55%)/kg nd liver (45%) protein hydrolystes (GP+P; Glycochrge, Annmet Petfoods; 74% crbohydrte, 24% protein s dry mtter, 75% clories s GPs, 25% of clories s protein), in 400 ml of wter; or 400 ml of wter lone. On verge, ech supplement provided 11% of the totl cloric intke in crbohydrte, nd the GP+P supplement provided 4% of the totl cloric intke s protein. This converts to pproximtely 27% of the totl crbohydrte intke for ech dog receiving supplements nd 15% of the totl protein for dogs receiving the GP+P tretment. Dogs tht refused the supplements or did not finish the supplements were tube-fed the remining solution to ensure consumption. Ech dog ws fed stndrd mount of the diet 2 hours fter being exercised. The mount of food offered to ech dog ws sufficient to mintin the specific body weight nd body condition of ech dog throughout the study (rnge = 140 to 310 g of food per dy). The verge consumption per dog ws 47.1 ± 13.7 kcl/kg/dy. Serum Collection nd Biopsies Immeditely preceding exercise, ech dog s posterior hind limbs were septiclly prepred for biopsy collection. A single muscle biopsy (<26 mg muscle tissue) from the right hindlimb semimembrnosis ws obtined under locl nesthetic (2% lidocine) using the percutneous biopsy pproch published by Reynolds nd collegues, 15 which took pproximtely 3 to 5 minutes per dog. The muscle biopsy smple ws immeditely frozen for subsequent glycogen nlysis. A blood smple ws lso obtined in n EDTA-treted tube nd immeditely centrifuged t 3500 rpm to collect plsm. A 1-ml plsm liquot ws trnsferred to tube contining 10 µl protinin (500 Kllikrein Inctivtor Units) nd 10 µl leupeptin (10 mg/ml) nd stored t 70 C for future insulin nd glucgon nlysis. The remining plsm ws stored t 70 C for future glucose nlysis. Using the sme procedures, second blood smple nd muscle biopsy smple were obtined (from the left hind limb) immeditely fter the dogs received the post-exercise supplement. Blood smples were lso obtined t 0.5, 1, 2, 3, nd 4 hours fter exercise; nd the plsm ws removed nd stored s previously described. Biopsy smples of skeletl muscle were lso obtined 4 nd 24 hours fter exercise. The 4-hour biopsy ws obtined from the right hind limb bout 2 inches distl to the originl biopsy. The 24-hour biopsy ws ob- 228

4 J. J. Wkshlg, K. A. Snedden, A. M. Otis, C. A. Kennedy, T. P. Kennett, J. M. Scrlett, F. A. Kllfelz, G. M. Dvenport, A. J. Reynolds, nd G. A. Reinhrt tined from the left hind limb t lest 2 inches distl to the originl biopsy. This procedure ws followed three times t 21- dy intervls, with ech dog being evluted fter ingesting one of the two nutritionl supplements or wter. The dogs were given 72 hours to recover from their most recent exercise bout before the experiment ws conducted. Glycogen content in skeletl muscle ws ssessed using the method described by Lo nd collegues 16 bse-ctlyzed glycogen hydrolysis nlysis, 17 with ech smple run in triplicte. Plsm glucose concentrtion ws mesured using utomted procedures stndrdized for clinicl screening on the Muscle Glycogen Concentrtions (g glycogen/100 g tissue) Hitchi 911 Automted Biochemicl Anlyzer (Hitchi High Technologies). Plsm insulin nd glucgon concentrtions were ssessed by stndrdized rdioimmunossys for cnine plsm, with 6.6% nd 5.4% intrssy coefficients of vrition for insulin nd glucgons, respectively, nd interssy coefficients of vritions of 5.1% nd 4.8%, respectively. Sttisticl Anlysis A previous evlution of the pired crossover design determined n α-vlue of.05 nd β-vlue of t lest.90, reveling the need to include t lest six dogs in the study to detect chnges in skeletl muscle glycogen. Thus, 10 dogs were chosen for evlution in this study. The study ws designed to evlute the response of vrious plsm indictors of glucose metbolism t individul time points in n effort to compre the control popultion with the two tretment groups; therefore, ech time Before Exercise After Exercise Figure 1. Glycogen depletion in skeletl muscle of Alskn sled dogs shown s the men skeletl muscle glycogen content (g of glycogen/100 g of muscle wet weight) before nd fter ech exercise session. The difference between the two times is significntly (P <.001) different. point ws exmined using one-wy nlysis of vrince (ANOVA) with Tukey s posthoc nlysis to determine whether there ws significnt difference mong the three different groups or ny combintion thereof t n α-vlue set t.05. Skeletl muscle glycogen content results were lso nlyzed similrly t 0, 4, nd 24 hours to look for significnt (P <.05) differences t ech time point mong the three tretments. In ddition, ll muscle glycogen vlues were verged before nd fter exercise nd compred using one-tiled pired t-test to ensure tht the exercise ws producing sttisticlly significnt decrese in skeletl muscle glycogen concentrtion. RESULTS Glycogen Concentrtion in Skeletl Muscle Comprison of glycogen concentrtions in skeletl muscle before nd fter exercise reveled significnt (P <.01) nd drmtic de- 229

5 Veterinry Therpeutics Vol. 3, No. 3, Fll 2002 Muscle Glycogen Increse (g glycogen/100 g tissue) Post-Exercise Supplementtion Control (wter) b Glucose polymer (GP) Glucose polymer + protein (GP+P) 4 Hours 24 Hours Time After Post-Exercise Supplementtion b 43%, nd 40%, respectively (Figure 2). However, there ws sttisticlly significnt (P <.011) increse in skeletl muscle glycogen content t 24 hours in both the GP nd GP+P groups compred with the control group, with n verge repletion of 39%, 85%, nd 81%, respectively. Glycogen repletion vlues were similr for the GP nd GP+P groups t ll time points. Figure 2. Glycogen repletion (men nd stndrd devition) in Alskn sled dogs 4 nd 24 hours fter exercise nd different post-exercise supplementtion.,b Groups with different superscript letters re significntly different (P <.011). Plsm Insulin (µlu/ml) crese (Figure 1). When evluting the post-exercise tretment responses, there were no significnt differences between the control group nd two tretment groups 4 hours fter exercise (P =.20) even though the verge repletion for control, GP, nd GP+P were 10%, Post-Exercise Supplementtion Control (wter) Glucose polymer (GP) Glucose polymer + protein (GP+P) Time (hr) Figure 3. Plsm insulin concentrtion (men nd stndrd devition) for Alskn sled dogs following exercise nd post-exercise supplementtion. Vlue for control group (wter) is significntly (P.001) different from the GP nd GP+P groups t the sme time. Plsm Insulin nd Glucgon Concentrtions Comprison of plsm insulin concentrtions mong the three groups showed significnt (P <.01) increse t 0.5 nd 1.0 hours for the GP nd GP+P groups compred with the control group. However, plsm insulin concentrtions rpidly returned to control levels t 2, 3, nd 4 hours fter tretment. There ws no observble difference in plsm insulin concentrtion between the GP nd GP+P groups throughout the study (Figure 3). Plsm glucgon concentrtions incresed modertely in the GP group compred with the control group t 0.5, 1, nd 2 hours fter tretment. (P.026) However, glucgon concentrtions were not different for these two groups t the 3- nd 4-hour time points (Figure 4). Furthermore, there were no differences in glucgon concentrtions for the GP nd GP+P tretment groups throughout the postexercise period. 230

6 J. J. Wkshlg, K. A. Snedden, A. M. Otis, C. A. Kennedy, T. P. Kennett, J. M. Scrlett, F. A. Kllfelz, G. M. Dvenport, A. J. Reynolds, nd G. A. Reinhrt Plsm Glucose Concentrtions When compring the three groups, there ws significnt (P.02) increse in plsm glucose concentrtion in the GP tretment group compred with the control group t 0.5 nd 1.0 hours fter exercise (Figure 5). In contrst, the GP+P group ws only significntly different from the control group t 1 hour fter tretment. There were no differences between the GP nd GP+P tretment groups throughout this period. Plsm Glucgon (pg/ml) Post-Exercise Supplementtion Control (wter) Glucose polymer (GP) Glucose polymer + protein (GP+P) Time After Supplementtion (hr) Figure 4. Plsm glucgon concentrtion (men nd stndrd devition) for Alskn sled dogs following exercise nd postexercise supplementtion. Vlue for the control group (wter) is significntly (P.026) different from the GP group t the sme time. There ws no sttisticl difference between the control nd GP+P groups. DISCUSSION Crbohydrte supplementtion fter exercise hs been used in thletic nimls for yers. 18,19 More recently, protein hs been dded to post-exercise supplementtion s mens of incresing the insulin response. 12 Some studies hve speculted tht post-exercise protein supplementtion, especilly the mino cid leucine, cn increse insulin concentrtions over longer period of time thn glucose polymers cn Plsm Glucose (mm/l) lone. 20 This supplementtion strtegy my enhnce the movement of glucose into muscle stores. To dte, there hs been little scientific investigtion of the puttive benefits of post-exercise protein supplementtion in the repletion of skeletl muscle glycogen in humns nd no reserch in thletic cnine breeds Time After Supplementtion (hr) Post-Exercise Supplementtion Control (wter) Glucose polymer (GP) Glucose polymer + protein (GP+P) Figure 5. Plsm glucose concentrtion (men nd stndrd devition) for Alskn sled dogs following exercise nd postexercise supplementtion. Vlue for the control group (wter) is significntly (P.02) different from the GP nd GP+P groups t the sme time. Previous studies using sled dogs showed drmticlly incresed skeletl muscle glycogen repletion (pproximtely 45%) with GP supplementtion within 4 hours fter 60-km strenuous exercise protocol. 6 Muscle glycogen 231

7 Veterinry Therpeutics Vol. 3, No. 3, Fll 2002 concentrtions in the study reported here did not rech sttisticl significnce t 4 hours, even though the verge glycogen repletion ws pproximtely 40%. Resons for this disprity could be twofold: First, the previously mentioned study ws conducted using three different groups of dogs. Biopsies were obtined in the initil group before nd fter exercise; wheres in the other two groups, biopsies were obtined before nd 4 hours fter exercise, with one group receiving crbohydrte supplementtion nd the other group receiving wter only. Therefore, the repletion vlues generted t 4 hours re compred with depletion vlues from seprte group of dogs. As result, it hd to be ssumed tht ll groups underwent the sme degree of depletion. This vrible ws removed from the current study becuse of the crossover pired design. Second, in the previous study, the dogs underwent more strenuous exercise (i.e., the dogs rn frther nd longer thn during preexperimentl trining), which my hve cused greter glycogen deficit thn in the study reported here. In the study reported here, the dogs were exercised t fster pce for shorter period of time, nd the exercise protocol ws no more strenuous thn their norml trining pttern. This difference is reflected by n verge of 80% depletion of skeletl muscle glycogen in the previous study, wheres glycogen depletion in the current study verged only 45%. Glycogen repletion pproched 100% by 24 hours fter supplementtion in the two crbohydrte tretment groups (P <.011), wheres the 4-hour biopsy smples showed only 40% increse in skeletl muscle glycogen (P <.20). It hs been suggested tht fter 4 hours of exercise, the insulin hypersensitivity nd/or insulin-independent glucose trnsporters become internlized in skeletl muscle, mking it more difficult to chieve mximl repletion However, in ll groups of dogs in the current study, there ws continul repletion of skeletl muscle glycogen fter the 4-hour time point. Becuse glycogen repletion fter 4 hours is primrily n insulin-driven response, it ws speculted tht feeding the dogs 2 hours fter exercise llowed continul glycogen repletion in skeletl muscle, regrdless of the tretment. This ssumption is bsed on the drmtic rise in plsm glucgon nd insulin concentrtions t 3 nd 4 hours in conjunction with the dditionl muscle glycogen repletion t 24 hours. The higher muscle glycogen levels t 24 hours in the tretment groups my be ttributble to glucose-spring effect of the tretments tht llowed these dogs to use diet-derived glucose to replenish muscle glycogen stores. In contrst, the control group my hve used diet-derived glucose to fulfill bsic metbolic needs while replenishing both heptic nd muscle glycogen stores. When ssessing the efficcy of the glucosecontining supplements used in this study, it is evident tht there is no difference between using GP or GP+P in post-exercise supplementtion. Both supplements provided pronounced benefits in muscle glycogen repletion within 24 hours. Similrly, the increses in plsm insulin nd glucgon concentrtions were not significntly different between the two tretment groups. Although it hs been postulted tht insulin response is higher in humns supplemented with both crbohydrte nd protein immeditely fter exercise, 12 there ws no evidence of n incresed insulin response with dded protein. Throughout the study, the GP group showed n immedite nd sustined increse in plsm glucgon concentrtions compred with controls until 4 hours fter exercise. This physiologic response suggested tht incresed plsm glucgon concentrtions would led to excessive ntgonistic ctivity ginst insulin, but pprently this did not trnslte into diminished skeletl muscle glycogen. Al- 232

8 J. J. Wkshlg, K. A. Snedden, A. M. Otis, C. A. Kennedy, T. P. Kennett, J. M. Scrlett, F. A. Kllfelz, G. M. Dvenport, A. J. Reynolds, nd G. A. Reinhrt though it my be specultive, incresed plsm glucgon concentrtion my result in more gluconeogenesis vi heptic sources, bsed on previous observtions. 24,25 Thus, the higher glucgon concentrtions in the GP tretment group my hve enhnced heptic glycogenolysis nd gluconeogenesis, leding to incresed glucose vilbility for skeletl muscle glycogen repletion during the post-exercise intervl; wheres the protein ingested by the GP+P group my ttenute glucgon secretion, preventing gluconeogenesis nd glycogenolysis. This my be the reson for the more drmtic erly rise in plsm glucose concentrtion tht occurred in the GP group compred with the control group t 0.5 hours post-exercise supplementtion. Therefore, in sled dogs, post-exercise supplementtion with protein my be reflected through diminished glucgon relese rther thn incresed insulin. Further investigtion of this phenomenon with lrger study popultions nd inclusion of protein-only supplements involving both heptic nd skeletl muscle glycogen nd prmeters of gluconeogenesis my be rewrding. In conclusion, there is profound increse in skeletl muscle glycogen concentrtion with the use of post-exercise crbohydrte supplementtion t dose of 1.5 g of supplement/kg of body weight. This supplementtion llows skeletl muscle glycogen concentrtions to return to preexercise levels within 24 hours. Although post-exercise supplementtion my hve definitive benefit, feeding norml mel within the initil 4 hours fter exercise my lso provide sufficient glucose for some muscle glycogen repletion. In ddition, the use of protein hydrolystes in post-exercise supplementtion to potentite n insulin response nd enhnced glycogen repletion could not be exhibited in this study. Therefore, it is unlikely tht protein in these crbohydrte-contining supplements ugments muscle glycogen repletion cpbilities. However, supplementl protein my ttenute plsm glucgon concentrtions during the post-exercise period, but the effect of this on cellulr ctivities during periods of exercise-induced metbolic stress hs not been determined. REFERENCES 1. Griffiths BC: Nutrition of the greyhound. Vet Rec 84(26): , Hill RC: The nutritionl requirements of exercising dogs. J Nutr 128(12 Suppl):2686S 2690S, Reynolds AJ, Fuhrer L, Dunlp HL, et l: Effect of diet nd trining on muscle glycogen storge nd utiliztion in sled dogs. J Appl Physiol 79(5): , Jeukendrup AE, Rben A, Gijsen A, et l: Glucose kinetics during prolonged exercise in highly trined humn subjects: Effect of glucose ingestion. J Physiol 515(Pt 2): , Wgenmkers AM, Beckers EJ, Brouns F, et l: Crbohydrte supplementtion, glycogen depletion, nd mino cid metbolism during exercise. Am J Physiol 260(Endocrinol Metb 23):E883 E890, Reynolds AJ, Crey DP, Reinhrt GA, et l: Effect of post-exercise crbohydrte supplementtion on muscle glycogen repletion in trined sled dogs. Am J Vet Res 58(11): , Coyle EF, Coggn AR, Hemmert MK, Ivy JL: Muscle glycogen utiliztion during prolonged strenuous exercise when fed crbohydrte. J Appl Physiol 61(1): , Ivy JL, Ktz AL, Cutler CL, et l: Muscle glycogen synthesis fter exercise: Effect of time of crbohydrte ingestion. J Appl Physiol 64(4): , Costill DL, Bowers R, Brnm G, Sprks K: Muscle glycogen utiliztion during prolonged exercise on successive dys. J Appl Physiol 31(6): , Zwdzki KM, Yspelkis III BB, Ivy JL: Crbohydrte-protein complex increses the rte of muscle glycogen storge fter exercise. J Appl Physiol 72(5): , Vn Loon LJ, Sris WH, Kruijshoop M, Wgenmkers AJ: Mximizing post-exercise muscle glycogen synthesis: Crbohydrte supplementtion nd the ppliction of mino cid or protein hydrolyste mixtures. Am J Clin Nutr 72(1): , Vn Loon LJ, Kruijshoop M, Verhgen H, et l: Ingestion of protein hydrolyste nd mino cid-crbohydrte mixtures increse post-exercise plsm insulin response in men. J Nutr 130(10): ,

9 Veterinry Therpeutics Vol. 3, No. 3, Fll Jentjens Rl, vn Loon LJ, Mnn CH, et l: Addition of protein nd mino cids to crbohydrtes does not enhnce post-exercise muscle glycogen synthesis. J Appl Physiol 91(2): , Wshizu T, Tnk A, Sko T, et l: Comprison of the ctivities of enzymes relted to glycolysis nd gluconeogenesis in the liver of dogs nd cts. Res Vet Sci 67(2): , Reynolds AJ, Fuhrer L, Vlentine BA, Kllfelz FA: New pproch to percutneous muscle biopsy in dogs. Am J Vet Res 56(8): , Lo S, Russell JC, Tylor AW: Determintion of glycogen in smll tissue smples. J Appl Physiol 28(2): , Pssonneu JV, Luderdle VR: A comprison of three methods of glycogen mesurement in tissues. Anl Biochem 60: , Bullimore SR, Pgn JD, Hrris PA, et l: Crbohydrte supplementtion of horses during endurnce exercise: Comprison of fructose nd glucose. J Nutr 130(7): , Reynolds AR: The effects of diet on sled dog performnce, oxidtive cpcity, skeletl muscle microstructure nd muscle glycogen metbolism. In: Crey DP, Norton SA, Bolser SM, eds: Recent Advnces in Cnine nd Feline Nutritionl Reserch: Proceedings of the 1996 Ims Interntionl Nutrition Symposium. Wilmington: Ornge Frzer Press; 1996: Mero A: Leucine supplementtion nd intensive trining. Sports Med 27(6): , Richter EA, Derve W, Wojtszewski JFP: Glucose, exercise, nd insulin: Emerging concepts. J Physiol 535(2): , Goodyer LJ, Khn BB: Exercise, glucose trnsport, nd insulin sensitivity. Annu Rev Med 49: , Ivy JL, Kuo CH: Regultion of GLUT4 protein nd glycogen synthse during muscle glycogen synthesis fter exercise. Act Physiol Scnd 162(3): , Wssermn DH, Splding JA, Lcy DB, et l: Glucgon is primry controller of heptic glycogenolysis nd gluconeogenesis during musculr work. Am J Physiol 257(Endocrinol Metb 20):E108 E117, Stevenson RW, Steiner KE, Dvis MA, et l: Similr dose responsiveness of heptic glycogenolysis nd gluconeogenesis to glucgon in vivo. Dibetes 36(3): ,