effect of prandial glucose on brown fat

8/6/2019 Effect of Prandial Glucose on Brown Fat

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Effect of Prandial G lucose on Brown FatThermogenesis in R ats: PossibleIm plications for Dietary O besity

Z. CLICK, G. A. BRAY1 AND R . J. TEAGUE

D epartm ent of M edicine, H arbor-U CLA M edical C enter,Terranee, CA 90509

ABSTRACT We have previously shown that a single meal results in an increasedtherm ic activity of brown adipose tissue (BAT ). In the present study, we exam ine theimportance of the availability of glucose in the test meal and in the incubation

medium on the rate of in vitro respiration of BAT.We show that a high glucose testmeal results in a significant increase in BATweight and in its in vitro rate of respiration and th at th e effect o f th e h igh g lu co se meal on brown fat th ermogenesis is significantly greater than that of an equicaloric high fructose meal. T hese data suggest thata decreased postprandial BAT thermogenesis may contribute to the increasedmetabolic efficiency and to the obesity reported to be associated with sucroseconsumption in the rat. We also show that the reduced in vitro rate of respiration ofBAT from meal-deprived rats is largely corrected by the addition of glucose andinsulin to the incubation medium. J. Nutr. 114: 286-291, 1984.INDEXING KEY WORDS thermogenesis brown adipose tissue glucose. fructose in vitro respiration

When normal rats are maintained on ahigh fat diet (1, 2) or when they receivea sucrose drinking solution in addition to astandard stock diet (3), they develop hyper-phagia and obesity. Consumption of either ahigh fat (4) or a high sucrose diet (5, 6) isassociated with an increased metabolicefficiency, which in turn enhances thedevelopment of these dietary obesities.We have recently shown that brown

adipose tissue (BAT) plays a role in thetherm ic effect (specific dynam ic effect) ofsingle meals. This tissu e undergoes a hypertrophy (7) which is associated w ith signifi

cant compositional changes (8), and its invitro respiration rate (7), as well as the invivo rate of blood flow ing through it (9), arelargely increased following a single lowprotein, h igh carbohydrate test meal. In ratsconsuming a high fat test meal, however, thein vitro therm ic activity of BAT is less thanin rats consuming a low fat meal eventhough meal size in the former is much

larger (10). These latter data raise theposs ib ility that d ie ta ry obesity that developswhen rats consume a high fat diet mayresult from decreased therm ic effects thatoriginate in BAT.In the present study, we examine the

possibility that a second type of dietaryobesity, nam ely that which develops fromconsumption of a high sucrose diet, may alsobe associated w ith a decreased heat production from BAT.S ince sucrose is composed ofglucose and fructose and since in the ratglucose was shown to be utilized less efficiently than sucrose (5, 6), w e hypothesize

that a fructose m eal will result in a reducedin vitro therm ic activity in comparison w itha glucose meal. In the present study, we alsoexam ine the extent to which the increased

1984 Amer ic an Ins ti tu te o f Nut rit ion. Receiv ed for pub li ca tion24 October 1983.'Presen t address : Div is ion of Diabe tes and Nutr it ion, Department of

Hea lth Science , LAC-USC Medica l Cente r, 1200North Sta te S tree t, LosAngeles, CA 90033.

286


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GLUCOSE AND BROWN FAT THERMOGENESIS 287

thermic acti vi ty of BAT af ter a high carbohydrate (starch) meal may be accounted forsolely by avai labi li ty of glucose substrate to

the BAT.MATERIALS AND METHODS

Animais

All rats used were Wistar females, weighing about 200 g, purchased f rom Simonsen(Gilroy, CA). The rats were caged individually in rooms maintained at 22 2Cwith lights on from 0700 to 1800.

Exper iment 1. Type of mea l ca rbohydr atea nd BAT weight

Protocol and diet. Eighteen rats weremaintained on a scheduled feeding planas f ollows: They had access to f ood tw icedaily at 0900-1100 and at 1530-1730 forf ive consecut ive days. This was fol lowed by2 days of ad libitum food intake. Thisweekly schedule was carried out for 2 weeks,af ter which al l rats were put on thei r feedingplan of two meals per day for one addi tionalday. Duri ng the 2 weeks that preceded theexperiment and when the rats were receiving two meals per day, the morning meals(of all rats) al ternated daily among threediets: 1) stock diet (Purina Rodent Chow,

Ralston Pur ina, St. Louis, MO); 2) a glucosediet (9% protein, 72% glucose, 19% fat,table 1); and 3) a f ructose diet (9% protein,72% f ructose, 19% fat; tabl e 1). Duri ng therest of the feedings, the rats received a stockdiet.The rationale for adopting thi s prepara

tory f eeding protocol of schedul ed interrupted by ad l ibi tum feeding was threefold.a ) On their scheduled feeding the ra tstrained to eat l arge meal s so as to maximizemeal size on the experimental day. b)Al ternating meal composi tion al lowed therats familiarization with the test diets

of fered duri ng the experimental day. c) Adl ibi tum intakes al lowed the rats to catch upon some weight loss exper ienced dur ing thescheduled feeding.On the experimental day, the animals

were divided into three groups, closelymatched for body weight and for voluntaryfood intake duri ng the preceding 2 weeks.Group 1 (n = 6) was meal deprived; group

2 (n = 6) received the high fructose testmeal; and group 3 (n = 6) the high glucosetest meal . The test meal lasted 2 hours, af ter

which all rats were intubated intragastrical-ly, group 1 with 2-4 ml of tap water per ratand group 2 and 3 with respective liquiddiets, i.e., group 2 w ith the glucose mealand group 3 with the fructose meal (table1). Each fed rat received a total of 27 kcalf rom both voluntary intake of the sol id di etduring the 2 hours of access to food andfrom the supplemental gastr ic intubation.The volume intubated per rat ranged f rom2 to 4 ml. To minimize possible stressassociated with the gastric intubation on theexperimental day, rats were adapted to theintubati on procedure through dai ly i ntra-

gastri c administrations of the liquid testdiets (table 1) during the week that preceded the experiment. These intubationsal ternated dai ly between the glucose andthe f ructose liquid diets, and each rat wasintubated with a volume of about 2-3 mlimmediately af ter the morning meal .

TABLE 1Composition o f th e te st d ie ts

IngredientSolid

dietCasein,vitaminree1Rogers-Harper

saltix1Nonnutritionalfiber'Vitaminmix'di-Methionine'Cornstarch,'

glucose2 orructose3Crisco1TotalProteinCarbohydrateFatLiquid

dietCarnation(nonfat dryilk)WaterGlucose

orructoseLipomul5ProteinCarbohydrateFatg1908040202149617220

'Teklad Test Diet s, Madison, WI . "Baker analyzedreagent, ] . T. Baker Co., Phi l lipsburg, NJ. 3LindbergNutr ition, Los Angeles, CA. 'Vegetable shor tening,Proctor & Gamble, Cincinnati , OH. "Upjohn Corp.,Kalamazoo, MI.


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288 CL ICK , BRAYAND TEAGUE

The rats were killed within the secondhour after the end of the gastric intubations;the interscapular brown f at was removed

and trimmed from adhering muscle andwhi te fat and weighed.

E xper iment 2. Type of mea l ca rbohydr atea nd in vitr o respira tion ra te of BAT

P rotocol. The exper imen ta l pr otoco l a ndmeal composi tions were identi cal to thosedescribed above. In thi s experiment sevenrats were meal deprived, 11rats received thehigh f ructose test meal and 10 rats the highglucose test meal. Each f ed rat received atotal of 28 kcal. The BAT was put in oxygenated Krebs-Ringer bicarbonate (KRB)buffer for the measurement of respi rationrate as described below.Determina tion of BAT r espira tion ra te.

After blotting, the interscapular BAT wassliced in a tissue chopper adjusted toproduce 200-j im secti ons. Sl ici ng was repeated in perpendicular directi ons. Onehundred to 200 mg of the chopped tissuewas put in a respi rometer vessel containing2.9 ml oxygenated KRB buffer [finalconcentration: 136.2, 4.74, 1.27, 1.18 and1.18 mM for NaCl, KC1, CaCl2, KH2PO4and MgSO< respectively. This solution wasautoclaved, cooled and NaHCO3 (4.76mM) was added to produce a pH of 7.26].Oxygen uptake was determined in a Gi lsondifferential respirometer at 37.5Cincubation temperature; a 30-minute equil ibrat iontime was fol lowed by 10 minute recordingsof O2 uptake for 30 minutes.

Exper iment 3. Ava ila bility of glucose a ndin vitr o r espir ation r ate of BAT

P rotocol a nd diet. Thir ty-two ra ts wer emaintained for 2 weeks on a scheduled feeding pl an as described earlier. Duri ng thistime, all rats were fed a low protein, highcarbohydrate diet , the composition of which

is described in table 1, with cornstarchprov iding the carbohydrate On the experimental day, the rats were divided into twogroups: eight rats received the (low protein,hi gh carbohydrate) test meal for 2Vt hoursand 24 rats were meal deprived. A ll ratswere killed within the second hour afterfood was removed f rom the fed group, thei rinter scapular BAT removed and careful ly

trimmed f rom white f at and muscle.D etermina tion of BAT respir ation ra te.

The tissues removed f rom the 24 rats that

were meal deprived were divided into threegroups according to the avai labi li ty of substrate i n the incubati on medium during themeasurement of oxygen uptake. I n group 1(n = 8) no glucose was added. In group 2,glucose was added at a f inal concentrationof 130 mg/100 ml, and in group 3 (n = 8),both glucose (130 mg/100 ml) and insul in(0.1 U/ml) were added. BAT from the groupfed the test meal (group 4, n = 8) had noadded substrate or insulin in the incubationmedium. All four groups of rats were closelymatched for body weight before the testmeal was given to the fed group. Respirat ionrate of the tissues was determined as described earlier.

Statist ica l analysis

Data were anal yzed by analysis of variance, and comparisons between groupswere done by a i -test, using the Bonferroniapproach for mul ti compari sons betweentreatments (11).

RESULTS

Experiment 1. Mea l ca rbohydra te andBAT weight. Effect of dieta ry tr ea tment onBAT weight was stati sti cal ly signi fi cant [F(2,15) = 7.70; P < 0.01; table 2]. Theobserved hypertrophy of BAT was statistically significant after the glucose (P < 0.01)but not af ter the f ructose test meal .Experiment 2. Mea l ca rbohydra te and

BAT r espir ation r ate. Effect of the dieta rytreatment on rate of brown fat respi rati on

TABLE 2

Body weig ht, ca lo ric in ta ke a nd BAT we ig ht,by d ie ta ry t rea tmen t1 '1

Treatment Body wt Caloric intake BAT wt

Meal deprivedFructose mealGlucose mealg185

6179 3179 2kcal2727mg160

7174 9204 9b

'Data are means SE. sValues marked withsuperscripts are significantly different from each otherat P < 0.01.


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was statistically significant [F (2,25)= 8.74; P < 0.01]. As shown in figure 1,the glucose meal resulted in a more than

70% increase (P < 0.05) when expressedper 100 mg tissue and i n a more than 100%increase (P < 0.01) when expressed pertotal i nterscapul ar BAT, when comparedwith the respi rati on rate of BAT f rom meal -depri ved rats. Expressed per total ti ssue,and partly due to the greater hypertrophy ofthe interscapular brown fat af ter the glucosemeal, respiration rate af ter this test mealwas significantly greater than after thef ructose meal (P < 0.05).

Experiment 3. Ava ila bility of glucosea nd BAT respira tion ra te. Effect of ava ilabi li ty of substrate on rate of respi ration of

BAT was stati sti cal ly si gni fi cant [F (3,28)= 6.20; P < 0.05]. As shown in figure 2,addition of glucose and insulin to theincubation medium of BAT from meal-deprived rats resulted in a statisticallysignificant increase in the rate of respirat ionwhen expressed per 100 mg tissue (P< 0.05). The highest ra te of respira tion,however, was observed in the BAT from

80-

I 70-

36OHni55CHKZ40HoP30H

iaoHtoiOH

0J

n Vit?/ Deprived

Fructose

Glucose

PER 100 m gW ET TIS SUE

PER TOTAL TIS SUE

Fig. 1 Respiration rate in vitro of brown fatpreparat ions f rom meal -depr ived rats and f rom ratsthat received a glucose or a fructose test meal . Data aremeans SE; n = 7 for meal deprived; n = 11 forf ructose and n = 10for glucose meal . Di fferent superscripts denote that the means are significantly differentf rom each other: abP < 0.05; cbP < 0.01. Mean bodyweight of the rats on the exper imental day, pr ior to thetest meal were 212 15, 211 13, and 205 14 forthe meal -depri ved and f or the rats f ed a f ructose orglucose meal , respectively. The respect ive weights ofthe interscapular brown adipose tissues were 174 24,203 34 and 232 31 mg. The fed rats consumed atotal of 28 kcal f rom both voluntary intake of the sol idfood and through gastric intubat ion.

meal-fed rats, by utilizing endogenoussubstrate. Due in part to the significanthypertrophy of the BAT follow ing the testmeal , respi ration rate per total ti ssue in thefed group was almost three times that ofBAT with no added substrate when takenf rom meal -depri ved rats (P < 0.001).

DISCUSSIONObesi ty in affluent societi es has been on

the increase in recent decades (12). Thisincrease in prevalence of obesi ty i s parall el ed by an increase i n the consumpti on offat and sucrose and by a decrease in consumption of starch (13). Since a largeconsumption of either of these nutrients

resul ts in the spontaneous development ofobesi ty i n experimental animal s (1-3), anincreased consumpti on of sucrose and f atmay constitute an important etiologicalcause of human obesi ty.The mechani sm involved in the develop

ment of dietary obesity in animals is notclear. Increased palatabi li ty of diets ri ch infat or sucrose may consti tute one factor. Inthi s study, we examine the hypothesi s thatdietary obesity that develops when consuming a high sucrose diet may result in partf rom a reduced thermic response of BAT tothe f ructose moiety of a sucrose meal . Thi s

would l ead to an increased metabol ic ef fici ency and consequentl y to l arger gains i nbody fat.We have prev iously shown that rats that

were al lowed a chronic access to a glucosedri nking soluti on exhibi t a greater hypertrophy of the interscapul ar brown fat thanrats having access to a f ructose drinkingsolution of the same concentration (14).Consi stent w ith thi s f inding are resul ts wereport here that the hypertrophy induced bya single test meal is greater af ter the glucosethan af ter the f ructose meal (tabl e 2; f ig. 1,legend) . This increased hyper trophy af ter a

glucose meal i s associ ated with a greaterpostprandi al i ncrease i n the in vi tro rate ofBAT thermogenesis compared with anequicaloric fructose meal (fig. 1). In anearlier study that used a similar experimental paradigm i t was reported that a highfat meal induced a smal ler increment in thein vitro rate of BAT thermogenesis than alow fat (high starch, low protein) meal , even


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290 C L IC K , B RAYAND T EACUE

lOO-i| | MEAL DEPR IVED

I | MEAL DEPR IVED + GLUCOS E

K^ $3 M EA L D EP R IV ED + G LUCOS E + IN S ULIN^IM0_l3UJKccoroiRESPIRA'80-60-40-20-/\[vX V'1MELQpLFEDCr ;!IlPer

100 mgwet tis su e

a

P e r to ta lw e t tis s ue

Fig. 2 Ef fect of substrate avai labi lity on i n vi trorespi rati on rate of brown adi pose ti ssue. Data aremeans SE. Different superscr ipt s denote that themeans are signif icantly different from each other: abf< 0.05; acP < 0.01; and adP < 0.001. M ean bodyweights of the rats on the experimental day, prior to thetest meal, were 183 2 and 183 3 for the mealdeprived (n = 24) and for meal fed (n = 8), respect ively. Interscapular brown fat weights were 153 4and 221 9 mg for the meal depri ved and the mealfed, respectively (P < 0.001). Energy intake of the testmeal was 28 2 kcal .

though meal size was much greater (10).Put together, the data from these two studiesare consistent with and provide supportto the hypothesis that a reduced thermicresponse to meals that originates in BATcontri butes to the development of di etaryobesi ty i n the rat.I n vitro measurements of BAT thermo-

genesis can reflect a meaningful in vivothermic response as suggested by our recentfinding that mean blood flow into BAT ismore than doubled after a single lowprotein, hi gh carbohydrate (starch) testmeal (9). Blood flow has been known to behighly correlated with oxygen uptake inBAT (15, 16).Though apparently present in small

quantities in the adult human (17), theimportance of BAT in the regulation ofhuman energy balance i s at the present notclear. In contrast to data obtained in rats (5,6, 18) and baboons (19), suggesting thatsucrose is utilized more efficiently thanglucose, in men, a sucrose meal wasreported to induce a greater thermic effect

than an equicaloric glucose meal (20). Itwould therefore appear that our currentfindings in the rat , suggesting that a reduced

BAT thermic response to a meal contributesto the development of dietary obesi ty, neednot necessar ily apply to humans.That availability of glucose to BAT is

associated with an increased thermic activi ty of the ti ssue has been prev iously shownby others, both in vi tro (21) and in vivo (22).Though the main substrate for oxidati on inBAT isthought to be free fatty acids releasedthrough the action of norepinephrine (23,24), glucose may stimulate respi ration as aprecursor of oxaloacetate required formaintaining activity of the tricarboxylicacid cycl e (23). Glucose taken up into BAT

may also serve as a precursor for synthesis offatty acids that are readily utilized in thetissue. Accordingly, in the present study wef ind that the speci fi c respi ration rate of BATpreparations that contain glucose andinsulin in the incubation medium and aref rom meal-deprived rats is signif icantlygreater than that of BAT from meal-deprived rats w ith no substrate added. Further, the rate approaches the respirat ion rateof BAT from meal-fed rats that utilizeendogenous substrate (f ig. 2). These datasuggest that dietary substrate absorbed froma meal can exert a direct effect on BAT

thermogenesis.

LITERATURE CITED

1. M i ck el sen, O., Tak ahashi , S. & Crai g, C. (1955)Exper imental obesi ty. I . Production of obesi ty inrats by feeding high-fat diets. J. Nutr. 57, 541-554.

2. Schemmel, R., M ickelsen, O. & Gill, J. L .(1970) D ietary obesi ty i n rats: body weight andbody fat accret ion in seven strains of rats. J. Nutr.100, 1941-1948.

3. K anarek, R. B. & Hirsch, E. (1977) Dietaryinduced overeat ing in experimental animals. Fed.Proc. 36, 154-158.

4. Schemmel, R., M ickelsen, O. & M otawl, K .(1972) Conversi on of di etary to body energy i nrat s as affected by strain, sex and rat ion. J. Nut r.102, 1187-1198.

5. Feyder, S. (1935) Fat f ormati on f rom sucroseand glucose. J. Nutr. 9 , 457-468.

6. M acdonald, I. & Grenby, T. H. (1979) Somedifferences between dietary carbohydrates in theiref fects on weight l oss and body fat i n rats. Proc.Nutr. Soc. 38, 30A (abs.) .

7. Click, Z., Teague, R. J. & Bray, G. A . (1981)Brown adipose tissue: thermic response increased


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by a singl e l ow protei n high carbohydrate meal .Science 213, 1125-1127.

8. Click, Z., Teague, R. J., Bray, G. A . & L ee, M .(1983) Composi tional and metabol ic changes in

brown adipose t issue fol lowing a single test meal .Metabolism, in press.9. Wick ler, S. J., Cl ick , Z ., Stern, J. S. & Horw itz,B .A . (1983) B lood f low i nto brown f at and intoother ti ssues fol lowing a single meal . Fed. Proc.42, 1190 (abs .) .

10. Cl ick , Z . (1982) I nverse rel ati onshi p betweenbrown fat thermogenesis and meal size: thethermostati c control of f ood i ntake rev isi ted.Physiol. Behav. 29, 1137-1140.

11. M eter, J. & Wasserman, W. (1974) A ppliedL inear Stati stical Model s, pp. 480-482, R. D .Erwin, Homewood, IL .

12. Montoye, H . J., Epstei n, F. H . & Kjel sberg, M. O.(1965) The measurement of body fatness: a studyin a total community. A m. J. Clin. Nutr. 16,417-427.

13. Joint FAO/WHO A d Hoc Expert Committee(1973) Energy and Protein Requi rements, WHOTechnical Rep. Ser. No. 522, World Health Organization, Geneva, Switzerland.

14. Teague, R. J., Kanarek , R., B ray, G. A ., Cl ick , Z .& Orthen-Gambil l, N . (1981) Ef fect of diet onthe weight of brown adipose t issue in rodents. LifeSci. 29, 1531-1536.

15. Foster, D. O. & Frydman, M . L . (1978) Non-shiver ing thermogenesi s in the rat . I I . Measurements of bl ood f low w ith mi crospheres poi nt tobrown adipose ti ssue as the dominant si te of thecalorigenesi s induced by noradrenal ine. Can. J.Physiol. Pharmacol. 56, 110-122.

16. Foster, D. O., Depocas, F. & Frydman, M . L .

(1980) Noradrenal ine- induced calor igenesi s inwarm - and col d-accl imated rats: rel ati ons between concentrat ion of noradrenal ine in arter ialplasma, blood flow to differently located masses of

brown adipose t issue, and calorigenic response.Can. J. Physiol . Pharmacol. 58, 915-924.17. Heaton, J. M . (1972) The di stri buti on of brown

adipose t issue in the human. J. Anat . 112, 35-39.18. K anarek, R. & Orthen-Gambil l, N . (1982) D if

ferential effects of sucrose, fructose and glucose onthe development of carbohydrate-induced obesityin rat s. J. Nutr. 112, 1546-1554.

19. A l len, R. J. L ., B rook , M., L i ster, R. E., Sim, A . K .& Warwick, M . H. (1966) M etabolic differences between dietary liquid glucose and sucrose.[Lett .] Nature (London) 211, 1104.

20. Sharief, N. N. & M acdonald, I. (1982) Differences in dietary induced thermogenesi s wi thvarious carbohydrates in normal and overweightmen. Am. J. Cl in. Nutr. 35, 267-272.

21. Shackney, S. E. & Joel, C. D. (1966) Stimulation of glucose metabolism in brown adipose tissueby addi ti on of i nsul in i n v itro. J. B iol . Chem. 241,4004-4010.

22. Rothw ell , N . J. & Stock, M . J. (1981) A rol e f ori nsuli n i n the diet-i nduced thermogenesi s ofcafeteria-fed rats. Metabolism 30, 673-678.

23. Cannon, B., Nedergaard, J., Romert , L ., Sundin,V. & Svartengren, J. (1978) T he biochemi calmechani sm of thermogenesi s i n brown adiposetissue. In: Strategies in the Cold: Natural Turpidityand Thermogenesis (Wang, L . C. & Hudson,J. W, eds.), pp. 567-593, Academic Press, NewYork.

24. Nicholls, D. G. (1979) Brown adipose tissuemitochondria. Biochim. Biophys. Acta 549, 1-29.

effect of prandial glucose on brown fat

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