Effect of Cortisol on Growth, Food Intake, Dietary Preference and Plasma Amino Acid Pattern of Rats Fed Amino Acid Imbalanced Diets '

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1 Effect f Crtisl n Grwth, Fd Intake, Dietary Preference and Plasma Amin Acid Pattern f Rats Fed Amin Acid Imbalanced Diets ' PHILIP M.-B. LEUNG, QUINTON R. ROGERS ANDA. E. HARPER Department f Physilgical Sciences, Schl f Veterinary Medicine, University f Califrnia, Davis, Califrnia, and Department f Bichemistry, University f Wiscnsin, Madisn, Wiscnsin ABSTRACT Since crtisl is knwn t influence plasma amin acid cncentratins and amin acid metablism in general, rats fed amin acid unbalanced diets have been given crtisl, and grwth, fd intake, dietary chice and plasma amin acid cncentratins have been measured. Fd intake and grwth f rats fed a threniner a histidine-imbalanced diet were depressed. Hwever, fd intakes f rats injected with crtisl (1 mg/day) and fed the unbalanced diets were nt depressed. When rats were ffered a chice between the threnine-imbalanced diet and a prtein-free diet, they chse the prtein-free diet almst immediately, whereas crtisl-injected rats preferred the imbalanced diet. It has been shwn that the mst limiting amin acid â threnine â in the plasma f rats fed the threnine-imbalanced diet fell belw that f the cntrl and the fasting level. Crtisl caused an elevatin f the cncentra tin f threnine and f ther indispensable amin acids in the plasma f rats fed the imbalanced diet. The increase in the cncentratin f amin acids in the plasma f crtisl-injected rats, pssibly thrugh sme redistributin f amin acids within the bdy, may accunt fr the preventin, by crtisl, f the fd intake depressin and the change in fd preference f rats ingesting the imbalanced diet. Additin f an unbalancing amin acid mixture t a lw prtein diet causes a reductin in fd intake and retardatin f grwth f rats. One f the earliest and mst cnsistent bichemical changes b served in rats fed an imbalanced diet is a marked reductin in the cncentratin f the mst limiting amin acid in bld plasma (1, 2). The pssibility exists that alteratin in the fd intake f rats in gesting the imbalanced diet is caused directly by the change in plasma amin acid pattern (3). If the plasma amin acid pattern des, directly r indirectly, influence fd intake regulatin f rats, then mdificatin f the plasma amin acid pattern by sme means, such as the administratin f a gluccrticid, shuld affect the fd intake f rats ingesting an imbalanced diet. Since eleva tin f plasma amin acid cncentratins by crtisl has been reprted by Friedberg and Greenberg (4), Bndy (5), Ltspeich (6) and Kaplan and Shimizu (7), and since crtisl administratin usually causes n depressin f fd intake f rats fed a nutritinally adequate diet (8), the present studies were undertaken t examine the effect f crtisl n fd intake, grwth and fd preference f rats ingesting an imbalanced diet. T ascertain the effect f crtisl administratin n the changes in plasma amin acids, rats trained t eat a single meal daily were crtisl and the plasma-free injected amin with acids were determined in bld samples taken at varius intervals during a 24-hur perid after feeding a balanced r imbalanced diet. EXPERIMENTAL Weanling r yung male rats f Hltzman r Sprague-Dawley strains were used in all experiments. They were hused individual cages with screen bttms. in In the fd selectin studies, cages f duble the usual size were used and extra care was taken t prevent fd spillage by using Fisher cups inside a n. 303 tin can in which a ne and ne-half inch hle was placed abut 3 inches frm the bttm f Received fr publicatin April 8, l Supprted in part by Public Health Service Re search Grants ns. AM10615, AM10747 and AM11066 frm the Natinal Institute f Arthritis and Metablic Diseases. J. NUTRITION,96:

2 140 PHILIP M.-B. LEUNG, QUINTON R. ROGERS AND A. E. HARPER the can. The rats were fed as described belw and water was supplied ad libitum. Tw types f amin acid-imbalanced diets were used. The cntrl diet fr the threnine-imbalance studies cntained 6% casein supplemented with 0.3% DL-methinine, while that fr the histidine-imbalance studies cntained 5% casein supplemented with 0.3% L-methinine and 0.2% L- threnine. The threnine-imbalanced diet was prepared by adding 5.4% f a mixture f indispensable amin acids lacking threnine t the 6% casein diet.2 The histidineimbalanced diet was prepared by adding 3.6 r 5.4% f the amin acid mixture lacking histidine t the 5% casein cntrl diet.3 All diets cntained salt mixture,4 4% ; crn il,5 5% ; vitamin mixture,8 0.5% ; chline chlride, 0.2% and dextrinstarch (1:2) mixture as the carbhydrate t make 100%. The "crrected" diets were prepared by adding 0.45% DL-threnine and 0.225% L-histidine-HCl t the thre nine-imbalanced r histidine-imbalanced diets, respectively. The amin acids were added at the expense f the mixed carb hydrate. Crtisl7 (hydrcrtisne acetate), Img/ rat/day, was administered intraperitneally r subcutaneusly just prir t feeding the animals. Rats receiving the threnine-imbalanced diet were injected at 3 t 4 PM, whereas thse fed the histi dine-imbalanced diet were injected at 5 t 6 PM daily. In the plasma amin acid study, rats trained t accept a single 2-hur daily feeding were used. They were trained t eat a 15% casein diet fr 2 weeks and then were fed the 6% casein cntrl diet fr 3 weeks. They were then selected n the basis f unifrmity f bdy weight and fd cnsumptin. The animals were in jected with crtisl (1.5 mg/rat) intraperitneally befre being fed either the cntrl r unbalanced diet. The cntrl grup was pair-fed against the unbalanced grup. Rats fed each f the diets ate ap prximately 8 g f fd. Systemic bld (by cardiac puncture) was taken frm 5 rats fed each diet 2, 6, 9, 13, 18 and 24 hurs after the feeding perid. Bld samples frm animals in each grup were pled and the plasma btained after centrifugatin was added t 5 times its vlume f 1% picric acid fr preparatin f a prtein-free filtrate as described by Stein and Mre (9). After remving the picric acid with a clumn f Dwex 2-X8 resin (50 t 100 mesh chlride frm), the deprteinized filtrates were analyzed fr amin acids using a Technicn amin acid analyzer. RESULTS Fd intake and change in weight f yung rats (80-90 g) fed the unbalanced diet cntaining 6% casein and 5.4% f amin acid mixture lacking threnine and injected with either saline r crtisl intraperitneally are shwn in table 1 and figure 1. The usual depressin in grwth and fd intake was bserved in rats fed the imbalanced diet. The fd intake and grwth f rats fed the imbalanced diet and injected with crtisl were nt depressed even in the early part f the experiment. The ttal gain in weight f the crtisl-injected imbalanced grup was 80% f that f the saline-injected cntrl and 86% f that f the crtisl-injected cntrl. The saline-injected rats ingesting the im- «The mixture f indispensable amin acids devid f threnine prvided: (in percent) DL-tryptphan, O.32; l--icuclne, 1.12; r>i>lsleuclne, 1.6; nl.-valine, 1.68; L-histidine-HCl, 0.48; DL-phenylalanine, 0.72; and r-lysine-hcl, The 3.6% and 5.4% amin acid mixtures lacking histidine cntained, respectively: (in percent) L-me thinine, 0.2 and 0.3; 1,-phenylalanine, 0.6 and 0.9; L-leucine, 0.6 and 0.9; i.-isleucine, 0.4 and 0.6; L-valine, 0.4 and 0.6; L-lysine-HCl, 0.6 and 0.9; t-arginine-hcl, 0.4 and 0.6; i^threnine, 0.3 and 0.45; and L-trvtphan, 0.1 and â â The salt mixture cntained: (in percent) CaCOs, 29.29; CaHPO4-2H2O, 0.43; KH2PO4, 34.31; NaCl, 25.06; MgSO4-7H2O. 9.98: Fe (C6H.sO7)-6H2O, 0.623; CuSO4, 0.156; MnSO4-H2O, 0.121; ZnCU ; KI, ; (NH4)6 MO7OZ4-4H2O, ; and Na2SeO3-5H2O, The salt mix was prepared hv and purchased frm General Bichemicals, Inc., Chagrin Falls, Ohi. The salts at 5% f the diet prvided: (in percent f element) Ca, 0.592; P, 0.394; K, 0.493; Na 0.493; Cl, 0.760; Mg, 0.049; Fe, ; Cu, ; Mn, ; Zn ; I, ; M, ; and Se, Mazóla Oil, Crn Prducts, C., New Yrk. «0.5% f the vitamin mixture in the diet prvided the rats with 0.44% sucrse plus the fllwing vita mins: (in mg/kg diet) thiamine-hcl, 5; ribflavin, 5; niacinamide, 25.0; Ca D-pantthenate, 20; pyridxine- HC1, 5; flie acid, 0.5; menadine, 0.5; d-bitin, 0.2; vitamin Bi2 (0.1% in mannitl), 30; ascrbic acid. HO (added t prevent thiamine destructin); vitamin E acetate (25% in a mixture f gelatin, sugar and starchl. 400: vitamin A acetate and vitamin D2 (325,000 USP units f A per gram and 32,500 USP units f D2 per gram in a mixture f gelatin, sugar and starch), (The thiamine, niacinamide, flie acid, menadine and vitamin Bu were purchased frm Nutritinal Bichemicals Crpratin, Cleveland, and the thers were purchased frm Hffmann-La Rche, Inc., Nutley, New Jersey.) * Crtisl (Hydrcrtisne acetate) in aqueus sus pensin was purchased frm ABCO Dealers, Inc., New Yrk.

3 EFFECT OF CORTISOL ON AMINO ACID IMBALANCE 141 TABLE 1 Fd intake f rats fed the cntrl, threnine-imbalanced, r crrected diet, and injected intraperitneally with saline r crtisl Diet Treatment Avg daily fd intake ' Days Cntrl (6% casein plus 0.3% DL-methinine) Saline Cntrl Crtisl Imbalanced (cntrl plus 5.4% amin acid threnine)imbalancedcrrected mix minus )CrrectedSalineCrtislSalineCrtisl ( imbalanced plus 0.45% DL-threnine i Five rats per grup. e Crr. Imb.(sline) 40 Crr. Imb. (crtisl) 30 e UJ 20 IO Cntrl (saline) eri Cntrl (crtisl) * Imb. (crtisl) Imb. (saline) 14 Fig. 1 Effect f crtisl n grwth f rats fed ad libitum the cntrl, imbalanced r cr rected diet. Cntrl diet: 6% casein plus 0.3% DL-methinine; imbalanced diet: 6% casein plus 0.3% DL-methinine and 5.4% amin acid mixture lacking threnine; crrected diet: imbalanced diet plus 0.45% DL-threnine; saline: saline-injected (intraperitneally); crtisl: crtisl-injected (intraperitneally). balanced diet, n the ther hand, gained nly 6.7% and 7.2% f the respective saline-injected and crtisl-injected cn trls. The fd intake f the imbalanced grup injected with crtisl was 101% and 93%, respectively, f thse f the saline-injected and crtisl-injected cn trl grups. Althugh the fd intake f rats ingesting the crrected diet and in jected with either saline r crtisl was within the same range, the weight gain f the crtisl-injected grup was depressed 23% belw that f the saline-injected cr rected grup. The experiment was re-

4 142 PHILIP M.-B. LEUNG, QUINTON R. ROGERS AND A. E. HARPER peated using slightly yunger animals f Sprague-Dawley strain (10 rats/grup, 60â 80 g ) and similar results were btained except that the crtisl-injected rats in gesting the crrected diet grew at mre nearly the same rate as the saline-injected animals. When the histidine-imbalanced diet was fed t rats (5 rats/grup) injected with crtisl (subcutaneusly), a similar stimu- TABLE 2 Fd intake f rats fed the cntrl, histidine-imbalanced, r crrected diet, and injected subcutaneusly with saline r crtisl intakedietcntrl Avg daily fd (5% casein plus 0.3% L-methinine L-threnine)CntrlImbalanced and 0.2% (cntrl plus 5.4% amin acid histidine)imbalancedcrrected mixture minus (imbalanced diet plus 0.225% L-histidine-HClCrrectedTreatmentSalineCrtislSalineCrtislSalineCrtisl Days2-6B Crr.(S) ^^.^A Aâ """" Crr.(C) - IO Fig. 2 Effect f crtisl n grwth f rats fed ad libitum the cntrl, imbalanced r cr rected diet. Cntrl diet: 5% casein plus 0.3% L-methinine and 0.2% L-threnine; im balanced diet: cntrl diet plus 5.4% amin acid mixture lacking histidine; crrected diet: imbalanced diet plus 0.225% L-histidine-HCl; saline; saline injected (subcutaneusly); crtisl: crtisl injected (subcutaneusly).

5 EFFECT OF CORTISOL ON AMINO ACID IMBALANCE 143 latin f fd intake was bserved, even thugh the rats did nt gain mre weight than thse injected with saline. The crtisl-injected cntrl grup cnsumed mre fd than the saline-injected grup but again the gains in weight f bth grups were similar. The amunts f fd cn sumed by the crtisl- and saline-injected grups fed the crrected diet were similar, but the crtisl-injected grup suffered a 43% depressin in grwth as cmpared with the saline-injected crrected grup (table 2 and figure 2). Figures 3 and 4 shw the 24-hur fd intake pattern f rats (8 rats/grup) fed either the cntrl r the threnine-imbalanced diet and injected with either saline r crtisl just prir t feeding. All animals were held withut fd fr 18 hurs befre they were fed. The fd in take f the saline-injected rats ingesting the unbalanced diet was depressed within a few hurs, with the greatest depressin ccurring between 5 and 8 hurs. The fd cnsumptin f the unbalanced grup re mained belw that f the cntrl grup thrughut the 24-hur perid. Hwever, the fd intake f the crtisl-injected rats fed the unbalanced diet was nt depressed. Als, crtisl injectin had little, if any, effect n the fd intake pattern f rats fed the cntrl diet. The effect f crtisl n fd selectin f rats ffered a prtein-free diet and the threnine-imbalanced diet is shwn in figure 5. Crtisl was injected intraperitneally (1 mg/rat) daily. Tw ut f 5 rats selected the imbalanced diet n day 1. The ther 3 rats did nt make this selectin until day 3. At the end f 2 weeks, when all rats were cnsuming nly the im balanced diet, saline was substituted fr IO TIME(HOURS) TIME (HOURS) Fig. 3 Twenty-fur-hur fd intake pattern f saline-injected rats fed the cntrl r the threnine-imbalanced diet. cntrl diet; O O threnine-imbalanced diet. Fig. 4 Twenty-fur-hur fd intake pattern f crtisl-injected rats fed the cntrl r the threnine-imbalanced diet. Cntrl diet: 6% casein plus 0.3% DL-methinine; threnine-im balanced diet: cntrl diet plus 5.4% amin acid mixture lacking threnine. â cntrl diet; O O threnine-imbalanced diet.

6 144 PHILIP M.-B. LEUNG, QUINTON R. ROGERS AND A. E. HARPER ~ Fig. 5 Changes in bdy weight and fd preference f saline- and crtisl-injected (intraperitneally) rats fed a prtein-free diet and an unbalanced diet simultaneusly, unbalanced diet: 6% casein plus 0.3% DL-methinine and 5.4% amin acid mixture lacking threnine; prtein-free diet: same cmpsitin as the cntrl diet except that prtein and methinine were replaced by mixed carbhydrate (dextrinâ starch 1:2). crtisl. After this nly 1 rat cntinued t cnsume the unbalanced diet almst ex clusively thrughut the rest f the experimental perid and its ttal fd in take fell; the rest f the animals reduced their cnsumptin f the unbalanced diet gradually ver a perid f 1 week, and began t cnsume the prtein-free diet in increasing quantities until the end f the experiment. Saline-injected rats selected the prtein-free diet immediately when they were ffered the chice between the prtein-free and the unbalanced diet. T determine whether rats can discrimi nate between the threnine-imbalanced diet and the crrected diet (unbalanced supplemented with 0.45% DL-threnine), rats (5/grup) with and withut injectin f crtisl were ffered a chice between the 2 diets as shwn in figure 6. The salineinjected animals invariably chse the cr rected diet ver the unbalanced diet and made the chice very quickly, whereas the crtisl-injected animals cnsumed bth diets withut shwing any specific prefer ence during the 2 weeks. At the end f 2 weeks, crtisl was substituted fr the saune injectin, and the animals aban dned their preference fr the crrected diet after 3 days and started cnsuming apprximately equal quantities f the tw diets. T study the chrnic effect f crtisl administratin n rats ingesting the un balanced diet, 2 grups f rats, which had previusly been used in the threnine-imbalance experiment were fed the un balanced diet ad libitum fr an additinal 4 weeks after cmpletin f the 2-week grwth and fd intake study. One grup f animals cntinued t receive crtisl injectins (intraperitneally), while the ther was injected with saline. After this 6-week perid, they were ffered a chice between the prtein-free diet and the threnine-imbalanced diet fr 2 weeks. As

7 EFFECT OF CORTISOL ON AMINO ACID IMBALANCE 145 e 5 s â Saline «-Crtisl 0 40 O > -g 10 I -Crtisl 14 Days 27 S IO "â *5 lili 7 14 Days Fig. 6 Changes in bdy weight and fd preference f saline- r crtisl-injected (intraperitneally) rats fed the unbalanced diet and the crrected diet simultaneusly. Imbuà anced diet: 6% casein plus 0.3% DL-methinine and 5.4% amin acid mixture lacking threnine; crrected diet: unbalanced diet plus 0.45% Dt-threnine. shwn in figure 7, the saline-injected grup cnsumed mstly the prtein-free diet thrughut the experimental perid. Five f the 10 rats selected the prtein-free diet almst immediately, while the rest f the animals still cnsumed sme f the un balanced diet all thrugh the experimental perid. The crtisl-injected grup, hw ever, chse the unbalanced diet n the first day and cntinued t d s until the end f the experiment. The chrnic effect f crtisl administra tin was examined further in anther trial. The rats were first fed the histidine-imbalanced diet cntaining 5.4% amin acid mixture lacking histidine fr 2 weeks and were injected with crtisl r saline subcutaneusly. Then the amin acid mixture was reduced t 3.6% fr anther 10 days and crtisl and saline injectins were cntinued intraperitneally. The rats were then allwed t chse between a prteinfree diet and the unbalanced diet cn taining 3.6% amin acid mixture minus histidine. As shwn in figure 8, rats nt injected with crtisl invariably refused the unbalanced diet and cnsumed pri marily the prtein-free diet. Similar results were btained when a mre severly un balanced diet cntaining 5.4% f the un balancing amin acid mixture was sub stituted fr the less severely unbalanced ne n day 11 f the experiment. The cr tisl-injected rats immediately rejected the prtein-free diet and cnsumed mstly the unbalanced diets thrughut the 4-week experimental perid, even thugh the mre severely unbalanced diet was intrduced n day 11. The changes in bdy weight f rats fllwed clsely the changes in fd selec tin, in that the rats lst weight when they selected the prtein-free diet and gained weight when they selected the unbalanced diet. The rates f stmach emptying f mealfed rats fed the cntrl r the threnineimbalanced diet were similar. The changes in the plasma cncentra tins f amin acids ver a 24-hur perid in rats that had been injected with crtisl

8 146 PHILIP M.-B. LEUNG, QUINTON R. ROGERS AND A. E. HARPER Saline - injected 3O 20 IO Crtisl-lnjected i-i ~ i 5.4%AAmix - Thr %AAmix / -Thr Utili  5 u. ia. Th, m-i Days Fig. 7 Effect f lng-term saline r crtisl administratin (intraperitneally) n grwth and fd preferences f rats fed a prtein-free diet and the threnine-imbalanced diet simultaneusly and injected either with saline r crtisl (10 rats/grup). Threnine-im balanced diet: 6% casein plus 0.3% DL-methinine and 5.4% amin acid mixture lacking threnine; prtein-free diet: same cmpsitin as the cntrl diet except that prtein and methinine were replaced by mixed carbhydrate (dextrin-starch 1:2). 12 and fed abut 8 g f either the cntrl r the unbalanced diet are shwn in figure 9. Fllwing crtisl injectin, an elevatin f the mst limiting amin acid, threnine, was bserved in the plasma abve the fasting level. Althugh the cncentratin f threnine in the plasma f the un balanced grup was lwer than that f the cntrl until 18 hurs after the feeding perid, the value was well abve the fasting level at all times. The plasma threnine cncentratin f the unbalanced grup began t increase 6 hurs after the feeding perid reaching the highest pint in 18 hurs and declining tward the fasting level at the end f the 24-hur perid. The plasma threnine cncentratin f the cn trl grup als increased after 6 hurs, reaching a maximum in 13 hurs and falling steadily tward the fasting level thereafter. The cncentratin f methi nine was similar in the plasma f bth grups 9 hurs after feeding. The cncen tratins f methinine 2 and 6 hurs after feeding, hwever, were cnsiderably higher in rats fed the unbalanced diet. All f the amin acids added t cause the imbalance, except lysine, shwed an early peak 2 hurs after feeding the un balanced diet. The changes f the plasma valine and phenylalanine cncentratins fr the unbalanced grup were similar, in that, they increased sharply t the maxi mum value at 2 hurs and declined rapidly by 6 and 9 hurs, respectively. The plasma valine and phenylalanine cncentratins f the cntrl were relatively cnstant thrughut the 24-hur perid. Leucine cncentratin als rse rapidly in the plasma f rats fed the unbalanced diet and remained higher than the cntrl thrughut. Changes in the isleucine cn centratin in the plasma f the imbalanced grup were similar t thse fr leucine. The

9 EFFECT OF CORTISOL ON AMINO ACID IMBALANCE 147 > --i O -20 Saline - injected IO Crtisl-lnjec ted O CT a, ~ a> 15 IO â â «C IO - 3.6%AAmix -His â žn-i 5.4%AAmix / -His O IO O I 5 IO 5 Hln-m-n-irTà irarritrlth,3.6%aamix -His /5.4% A A mix / -His O IO Days Fig. 8 Effect f lng-term saline r crtisl administratin (intraperitneally) n grwth and fd preference f rats fed a prtein-free diet and the histidine-imbalanced diet simul taneusly (5 rats/grup). Histidine-imbalanced diet: 5% casein plus 0.3% L-methinine, 0.2% L-threnine and 3.6% r 5.4% amin acid mixture lacking histidine. lysine cncentratin in the plasma f the unbalanced grup increased less rapidly after feeding. The lysine cncentratin f the cntrl grup was belw that f the unbalanced grup at all times. The histi dine cncentratin f the grup fed the unbalanced diet increased shrtly after feeding, but fell belw that f the cntrl at 6 hurs. The cntrl histidine values were quite cnstant. The cncentratins f arginine and gly cine fr the unbalanced grups were higher than fr the cntrls at 13 hurs. N marked changes were bserved in the plasma cncentratins f arginine and glycine fr the cntrl grups. Serine, tyrsine, alanine and glutamic acid exhibited irregular patterns in grups fed either f the 2 diets. Aspartic acid in the plasma f the cntrl r unbalanced grup did nt shw prnunced peaks, and the values were either slightly abve r belw the fasting value thrughut the 24-hur perid. DISCUSSION The fd intake f rats fed the threnineimbalanced diet increased when they were injected with crtisl; the increase in fd intake was accmpanied by an increased grwth rate. Similar results were btained with rats fed the histidine-imbalanced diet and injected with crtisl but the stimulatins f fd intake and grwth were smaller. This indicates that the effect f crtisl in stimulating fd intake and grwth may depend upn the catablic characteristics f the amin acid that is mst limiting in the unbalanced diet. The slw catablic rate f threnine, as pinted ut by Charkey et al. (10) and demn strated by Ousterhut (11), as cmpared with that f histidine, may accunt fr the difference in magnitude f fd-intake sti mulatin f crtisl-injected rats ingesting the 2 different imbalanced diets. In sme instances in the present study, the expected benefit frm increased fd intake may be ffset by the catablic effect f crtisl, which is apparently dependent

10 148 PHILIP M.-B. LEUNG, QUINTON R. ROGERS AND A. E. HARPER Arginine Fasting I5 Tlevel t J I 6- spfbsting" level J *l i i Glycine Figure 9

11 EFFECT OF CORTISOL ON AMINO ACID IMBALANCE 149 Methinine Hurs Fig. 9 Effect f crtisl n amin acid cncentratins in plasma f rats trained t eat a single 2-hur meal daily and fed the cntrl r the unbalanced diet. C: Cntrl diet (6% casein plus 0.3% DL-methinine); Imb: unbalanced diet (cntrl diet plus 5.4% amin acid mixture lacking threnine). All rats were injected (intraperitneally) with 1.5 mg f crtisl just befre feeding.

12 150 PHILIP M.-B. LEUNG, QUINTON R. ROGERS AND A. E. HARPER n the dse and rute f administratin. As reprted by Wells and Kendall (12) crtisl (injected subcutaneusly) is knwn t inhibit grwth f animals. Bel lamy (8) als fund that crtisl (injected subcutaneusly) caused a rapid inhibitin f grwth, but that it had n significant effect n fd intake f rats fed a nutri tinally adequate diet. He suggested that crtisl may prduce bichemical changes leading t inhibitin f grwth which are independent f an effect n fd intake. Greenspan et al. (13) cmpared tw mdes f administratin (i.e., subcutaneus ver sus intraperitneal) and fund that intraperitneal injectins did nt cause weight lss and adrenal atrphy, whereas the same dse f crtisl injected subcutan eusly prduced bth weight lss and adrenal atrphy. This may accunt fr the fact that the rats fed the histidine-imbalanced diet and injected with crtisl subcutaneusly, suffered grwth depressin althugh the fd intake was increased. Since the present study agrees with earlier reprts by ther wrkers that crtisl treat ment elevates plasma amin acids (4-7), and since intraperitneally injected crtisl is rapidly absrbed and excreted (13, 14), it might be expected that the shrter dura tin f an effective crtisl level prduced by intraperitneal injectin wuld mdify the bld amin acid pattern, but might nt prvide a sustained stimulus lng enugh t prduce the maximum catablic effect. Certainly a significant generalized amin acid catablic effect culd nt have ccurred withut affecting grwth since the lw prtein cntent f the cntrl diet severely limits grwth. It has been suggested that sme un desirable physilgical r bichemical change may be assciated with the sharp drp in the cncentratin f the mst limiting amin acid in bld plasma f rats ingesting the imbalanced diet, and that these changes may be respnsible fr the depressin in fd intake (3). Hence, the bservatins that crtisl-injected rats selected the imbalanced diet ver the pr tein-free diet and that they failed t dis criminate between the crrected and the imbalanced diets suggest that the specific physilgical r bichemical effect pr duced by the ingestin f the imbalanced diet is ablished by crtisl treatment. The results f the fd selectin studies shw that rats injected with crtisl preferred the imbalanced diet ver the prtein-free diet as lng as crtisl was administered. Of interest in relatin t this is that the plasma cncentratin f the mst limiting amin acid did nt fall belw the fasting level when crtisl was injected. The variatin amng rats in adapting t the threnine-imbalanced diet can be seen frm the results f the fd selectin study. Half f the saline-injected animals fed the imbalanced diet ad libitum fr 4 weeks selected the prtein-free diet im mediately when they were ffered the chice, while the ther half did nt reject cmpletely the imbalanced diet thrughut the experimental perid. These results in dicate that, althugh the rat will adapt and grw when fed the imbalanced diet fr several weeks, the "metablic respnse" t the amin acid imbalance is still evident, thugh perhaps less severe. The 24-hur fd intake pattern f rats injected with crtisl shws that crtisl administratin (intraperitneally) was ef fective in stimulating the fd intake f rats ingesting the imbalanced diet within a shrt perid f time. This result, then, ex plains the selectin f the imbalanced diet by crtisl-injected rats n day 1 f the chice experiment. Crtisl had little, if any, effect n the fd intake pattern (24 hur) f rats cnsuming the cntrl diet, indicating that crtisl acted t enhance specifically the fd intake f rats fed the imbalanced diet and that it apparently in duces a higher intake f the different im balanced diets thrugh similar mechan ismsâ pssibly by raising the level f the mst limiting amin acid in the plasma. On the ther hand, crtisl is knwn t af fect amin acid metablism in many ways, especially by inducing sme f the en zymes in amin acid catablism; this culd als affect the plasma amin acid pattern f rats ingesting the imbalanced diet. Al thugh the results f the plasma amin acid studies are in accrd with the hyp thesis (3) that the plasma amin acid pat tern may influence the regulatin f fd intake, mre infrmatin is needed t establish with certainty whether crtisl altered the feeding behavir f rats in-

13 EFFECT OF CORTISOL ON AMINO ACID IMBALANCE 151 gesting the unbalanced diet thrugh this r sme ther mechanism. LITERATURE CITED 1. Kumta, U. S., and A. E. Harper 1962 Amin acid balance and imbalance. IX. Ef fect f amin acid imbalance n bld amin acid pattern. Prc. Sc. Exp. Bil. Med., 110: Sanahuja, J. C., and A. E. Harper 1963 Ef fect f dietary amin acid pattern n plasma amin acid pattern and fd intake. Amer. J. Physil., 204: Harper, A. E., P. Leung, A. Yshida and Q. R. Rgers 1964 Sme new thughts n amin acid imbalance. Federatin Prc., 23: Friedberg, F., and D. M. Greenberg 1947 Endcrine regulatin f amin acid level in bld and tissues. J. Bil. Chem., 168: Bndy, P. K The effect f the adrenal and thyrid gland upn the rise f plasma amin acids in the eviscerated rat. Endcrinlgy, 45: Ltspeich, W. D Relatins between insulin and pituitary hrmnes in amin acid metablism. J. Bil. Chem., 185: Kaplan, S. A., and C. S. Nagareda Shimizu 1963 Effect f crtisl n amin acids in skeletal muscle and plasma. Endcrinlgy, 72: Bellamy, D Effect f crtisl n grwth and fd intake in rats. J. Endcrinl., 31: Stein, W. H., and S. Mre 1954 The free amin acids f human bld plasma. J. Bil. Chem., 211: Charkey, C. W., A. K. Kan and J. A. Ander sn 1954 Effects f fasting n free amin acid level in chick bld as mdified by vitamin B12. J. Bil. Chem., 210: Ousterhut, L. E Cmparative threnine, valine, histidine and glucse xi datin in chicks. Prc. Sc. Exp. Bil. Med., 102: Wells, B. B., and E. C. Kendall 1940 The influence f crticsterne and C17-hydrxydehydrcrticsterne (cmpund E) n smatic grwth. Prc. Staff Meeting May Clin., 15: Greenspan, F. S., H. Giffrd and Q. B. Deming 1953 A cmparisn f the effects f crtisne administered intraperitneally and subcutaneusly in the rat. Endcrinlgy* 52: Prter, C. C., and R. H. Suba 1953 The absrptin and glycgenic activity f crti sne and hydrcrtisne after parenteral administratin t rats. Endcrinlgy, 53: 73.