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Volwne 7 Number 3 Medical Journal of the Islamic: Republic of mm Fall 1372 1993 November CHA NGES IN BIOCHEMICAL PARAMETERS OF CARBOHYDRATE METABOLISM FOLLOWING Downloaded from mjiri.iums.ac.ir at 17:31 IRDT on Thursday August 3rd 2017 FENFLURAMINE ADMINIS TRATION IN RATS M. ANI, M. MESSRIPOUR AND N. NAGHSHINEH From the Dept. of Biochemistry, School of Pharmacy, Isfahan University of Medical Sciences, Isfahan, Islamic Republic of Iran. ABSTRACT Fenfluramine, an anorexigenic agent, is widely used in the treatment of obesity. Besides its anorectic effect, it may also have some effects on general metabolism with the consequence of weight loss. In this study, the effect of fenfluramine on the concentrations of some parameters related to carbohydrate metabolism was investigated. It was shown that the serum insulin level was reduced by 41 %, four hours after fenfluramine administration, which was accompanied by the elevation of serum glucose levels by 26%. The liver glycogen content showed a transient reduction, but reached the control level four hours post-treatment. Corticosterone levels were elevated immediately, followed by a 20% reduction after four hours. The short-term effects of fenfluramine on thyroid hormones were not statistically significant. Administration of fenfluramine for two weeks did not change the glycogen content of the liver significantly, but 64% and 103% increases were observed after four and six weeks of drug treatment, respectively. Insulin levels showed a gradual increase so that by the end of six weeks, 153% increase in insulin level was observed. No significant changes in serum glucose levels were seen during the period of treatment. Corticosterone concentration remained unchanged up to four weeks of treatment but a 32% reduction was seen after six weeks. The levels of T and T3 4 showed a transient increase, followed by a significant decrease after six weeks of treatment. It is concluded that fenfluramine has some important metabolic effect that is related to its action in decreasing food intake and weight loss. M.lIRI, Vol. 7, No.3, 183-186, 1993. INTRODUCTION decreasing food intake which would be of value in tile treatmentof obesity.' In this regard tile effect of fenflurarnine Fenfluramine, an amphetamine derivative, was on plasma and liver levels of lipid fractions and on plasma introduced origimdly as an anorectic drug and it is frequently and brain levels of tryptophan has already been investigated used in the treaunent of obesity.' Its action is thought to in our laboratory.'> There is also evidence that this agent involve direct effects on serotoninergic systems at the improves insulin action independent of its effects on food level of hypotllalamus.' leading to anorexia and weight loss. intake,' which in tum may affect carbohydrate metabolism. It is also evident tlmt tllis drug may have some addition,d The present study was undertaken to investigate changes peripheral actions that may be unrelated to its action in in the concentrations of some biochemical parameters 183 Carbohydrate Metabolism with Fenfluramine Table l. The shorl-term effect of fenfluramine nil some biochemical parameters in rals. Each value indicates the mean ± SEM of five experiments performed in duplicate. Values in parenthesis shuw percent increase or decrease from Ihe control level. For more details see text. � Downloaded from mjiri.iums.ac.ir at 17:31 IRDT on Thursday August 3rd 2017 T ime GJu!.:osc Glyt.:ogt::n Insulin Curticosll!ronc T, T, (Illg/dl) (Illg/g Prot.) �IU/dl (�g/dl) (J-lg/dl) (ng/dl) ConlIol 85.4 ±5.7 20fi±21.4 37.4±4.4 37.7± 3.2 1.9±0.2 65 ± 6.8 0.5 hr %5 ± 6.4 169.6 ± 7.6 33.4±4.7 47.2±3.0 1.6±0.2 5U.8±1.6 (13%) (-18%) (- JU%) (+25%) (-20%) 2 hr 109± 6.8 98.7±9.7 31.6±3.9 39.0±3.1 2.1±0.3 (+27%) (-52%) (+3%) (+5%) 4 hr 108 ± 4.6 201± 14.5 22 ± 3.5 2lJ.9 ± 1.2 (+26%) (-2%) (-41%) (-20%) (-15%) related to carbohydrate metabolism following fenOuramine 1.9± 0.4 --_. (-21%) 81 ±5.1 (+24%) 12.4± 1:1 (+11%) by decapitation, their blood collected and allowed to administmtion. The observed changes may be a secondary coagul:ue. Theirsl!ra were then separated by cenlrifugation effect due to the alteration in hormon,d b,d,mce of the and used for the an,dysis. To avoid tlle probable diurnal animal. changes in serum honnonc levels. care was taken to perform the sampling at the smne time each day. Livers were rt!moved immediately for the determination of MATERIAL glycogen content, which was performed according to the mcthod of Kemp. et al.1 Serum glucose was determined by Male Wistar flUS (100 -150 gr) (obtained from Pasteur orthotoluidine method as reported by Caraway.' The Serum Institute, Tehran) \Vcr!! used Cor the experiments. They cOllcentration were kept at st�Uldard conditions in our animal labs. corticosterone. thyroxine (T) and T) wefe determined by or hormones Fenfiurmnine hydrochloride (20 mg coated tablets) was iIIIIIIunorauiOJI1L:tric tcchniq ue. incluuing insulin, purchased from Loghman Pharmaceutical, Tehran. Radioactive kits for the detennination of hormones were RESULTS obtained from Diagnostic Products Company, (DPC), Los Angeles, USA. All other chemic,ds used in this study were of reagent grade �md obtained from Sigma Chemica.l Short term l!ffccts of fl!nfluramine on biochemical Company. parameters an: shown in Table I. It is shown that the concentration of glucose is elevated following fenOuflunine :uJministralion. which is accomprmied by a reduction in METHODS seruln insulin level. The liver glycogen content showed a reduction, hut it reached the control level four hours post Anirn,ds were chosen in groups of five 1md kept in lrealml!n!. The levcl of corticoslerone showed an increase separate c;:ages. In short-tenn study. animals werc injected (25 percenl) 0.5 hr after fcnlluraminc administration, intraperitone,dly witil the acute dose of i"enOuflunine (100 followed by a gradual decrease in this honnone level so that mg/kg body weight). Treated animals were killed either 20 percent reduction was observed after four hours. Changes 0.5, 2 and/or4 hours post-injection. Anim'ds used for long observed in the levels of thyroid honnones were not term study received daily intr.lperitone,d doses of 10 kg fenllununine for either 1,2,4, or 109! st;ltistic:dly significant. 6 weeks. Groups of Long-term effects of fenOuramine were also studied, animals werc injected with normal saline in parallel with the results of which are shown in Table II. According to l!<tch cxpl!riml!nt :U1d considered as controls. results, the concentration of insulin in serum tends to At the time of tlle experiment, the ,mim,ds were killed increase following the continous administration of 184 1 M. Ani, et al. Table II. The lung-term effect u f fen nu ramin c un the biochemical parameters in rats. Eal'il figur e shows the mean ± SEM of five experiments. In parallihesis the percent increase ur decrease afC shown. For details abuut the method uf the experiments, see text. Week � Downloaded from mjiri.iums.ac.ir at 17:31 IRDT on Thursday August 3rd 2017 Conlmi I 2 4 Glucose (m!<ldl) Insulin Cor1iL:oslcrunc T, TJ �IU"" (�g/dl) ()!g/dl) (ngldl) 85.4±5.7 2011±21 37.4 ± 4.4 37.7±3.2 1.9 ± 0.2 65 ±6.2 73.1±:! 176.6 ±3.1 40.6±10 30.4±2.6 1.2±0.1 73.4±2.7 (·14%) (.14%) (+9 %) (·19%) (-36%) flS.6 ±2.5 2U2 ±22 (·I�%) (.I.R%) 84J ±3.4 338± 41 .. .. [, Glycogen (mg/g Prot.) I 05 ± 4 (+ 23 %) I 45 ±10 38.9±1.7 4.0±0.5 80 ± 8 (+20%) (+3%) (+101%) (±23%) 44.5 ± 9 37 ±4.5 1.9 ±0.3 (+19%) ---- 89.8 ±2.8 ---- (+38%) 94.8±4 25.6 ± I 1.1±0.2 48± 6.4 (+153%) (-32%) (-41%) (-26%) (+64%) 474±40 (+103%) (+13%) fenfluramir'le, so that after six weeks of trt.!Htmenl, the honnone level reached 94.8 I1IU/mL compared wilh Ihe control level of37.4 (P<O.O I). The increase in insulin level was accomp;:mied by the accumulation of glycogen in the liver. Thus. 103 pcrcenl increase in glycogen conleOl of lhe liver was observed following six weeks of drug administralion (P<O.O I). Serum glucose level did nOI c1uUlge significantly afler four weeks of IrealmCnl. bUI Ihe continuation of the treatment for six weeks increased serum glucose level by 23 percen!. No signific'UlI c1uUlges were observed in corticoslerone level afler four weeks of fenfluramine administration,but a reduction of 32 percent. was seen after six weeks. Thyroid honnonl!s were first elevated due to the action of fenflununine, \VherC�L"i the continuation oflhe treatment led to a consequent reductil)n. Thus Ihe increase in T. level was 13,23 and38 percent .. .I . following trcaUnem for one Iwo and four weeks respecuvcly. bul levels decreased by 26 percem if Ihe treallnenl was continued for six weeks. Serum T-Ilcvel was also increased by aboul 100 percem afler Iwo weeks of fenflurrunine administration, whereas a reduction of 4 1 percent was observed after six weeks. We have already reporled Ihat fenfluramine affecls tile metabolism of lipids,Ihe body. The action of tilis drug on Ihe metabolism of biomolecules may be secondary 10 its effeclon Ihe honnonal slalUs oflhe body. In this sludy il was shown Ihal the levels of insulin. corticoslerone and Ihyroid honnones were a1Tected by fcnllu"unine. Ihe consequence of which are c1uUlges in cm'bohydrate melabolism. It has ,�so been reponed that fenlluramine treaUnenl improved glycemic control in obese glucose-intolerant humans and in non-insulin dependent diabeles mellilUs independenl of its effecls on food imake and body weiglll.OI Our resulls indicale Ihal following Ihe administration of acule doses offenfluramine, Ihe glycogen of the liver was immedialely reduced which may be due 10 eilher increased rale of glycogenolysis or decreased rale of glycogenesis. This reduction in liver glycogen conlent is accomp:mied by an increasein serum glucose and adecrease in serum insulin levels. II is probable Ihalthe reduclion in liver glycogen level is due to the inhibition of insulin symhesis or release from the pancreatic cells. There are reports indicating that fenflurrunine inhibiled IhereleHseofinsulin from the perfused rat pancrcasy,13 Alternatively fenflurrunine may affect the liver enzymes involved in glycogen melabolism direclly, theconsequenceof which is the reduction in the concentration of Ihis carbohydrale slOrage compound. DISCUSSION Fenfluramine. an 'Ulli-obesiIY drug,is struclu"�ly bUi nol funclionally relaled 10 amphetmnine and is reponed 10 have some effecls on biomolecular melabolism,·' 6 hormonal balance of obese subjecls.' The reduction in liver glycogenesis due 10 Ihe inhibition of liver phophoglucomulase by fenfl uramine has already been reporled." However Ihe reduclion in liver glycogen level is transienl, so Ihat il reaches the control level four 185 Carbohydrate Metabolism with Fenfluramine hours after fenfluramine injection. As the insulin level a t liver levels of lipid fTactions in the rat. Ind J Pharmnc 20: 175- this time is still below conuul, Ole increase in liver glycogen t77,t988. level may be mediated through some other mechanisms 5. Ani M. Boroumam.l A: Changes in plasma and brain tryptophan involving honnones such as corticosterone and thyroid and serotonin levels following fenfluramine administration in honnones. It has been reported Omt the administration of ral.lnd J Pharamoc 20, 171-t74, 1988. 6. Storlien LH, Thorburn A W. SmythcGA: Effectof fenflurnmine fcnflurmnine immediately increased the serum corticosterone on basal glucose turnover and fat-feeding-induced insulin Downloaded from mjiri.iums.ac.ir at 17:31 IRDT on Thursday August 3rd 2017 levell�.[6 which is in agreement with the results obtained in resistance in rals. Diabetes 38: 499-503,1989. this study. Fenfluramine did not have 'my significant effect 7. Kemp A, Kits AIM: Colorimetric micromethod for the on thyroid honnoncs in short-term, which is in agreement determination of glycogen in tissues. Biochem J 5b: 646-648, wiOl the report Olat this drug had no effect on TSH release." 1954. In long-tenn however, fenflurrunine had different effects 8. Caraway WT: Methods for the determination of glucose in body 011 serum insulin and liver glycogen levels. The fluids. In: Tietz NW. Fundamentals of Clinical Chrmistry. administration of fenfluramine for six weeks led to 153 W.B. Sounder> Company. Philadelphi" pp: 242-251. 1976. !O3 percent 9. Bernini OP. Argenio GF, Corso CD, Vivaldi MS: Seroto increase in liver glycogen level. This is consistent with the ninergic receptor activation by D-fenfluramine enhances the percent increase in insulin level, followed by blunted pituitary-adrenal responsiveness to corticotropin findin,gs that long term administration of fenfluramine increased insulin secretion III a n d also inhibits glycogenolysis.I' releasing honnone in obese subjects. Metabolism 41 (I) 17-21. 1992. 10. Ani M. Amini SA: The effect of fenfluraminc on mitochondrial energy metabolism. Abstructs of 9111 Int. Congress of Physiol Our result showed that long term administration of fenflurmnine led to a decrease in corticosterone level which and Pharmac, Shnheed Bchcshti Universtiy. Tehran, IRAN. may be partly responsible for the observed metabolic changes. Administration of fenfluramine for two weeks increased II. the serum concentration of T, and T, which in turn may 224, May 15-18, 1989. Verdy M, Charbonneuu L, Verdy I, Blanger R, Bolte E, Chaissun IL: Fenfluramine in the treatment of non-insulin increase the rate of basaI metabolism leading to weight loss. dependent diabetics. Int J abc, 7: 289-297, 1983. However continuation of the treatment for up to six weeks 12. Barseghinn G, Lev-Ran A, Hwang D, Josefsberg Z. Tomkinson led to the reduction of these honnone levels which is in good C: Fenfluramine inhibits insulin secretion and potentiates agreement with the well-documented tnmsient effect of Olis glucagon release by the perfused rat pancreas. Eur J Pharmacal 96: 53-59,1983. ,tnoreclic agenl.:!o.:!J 13. Altamonte L.Zoli A. Manna R,Greco AV: Erfect offenfluramine The reduction in thyroid honnone levels after six weeks on insulin/growth honnoneratio in obcsesubjccts. Pharmacolog.y of treatment may be mediated through some unknown 36: t06-III,1988. mechanism responsible for setting up the metabolic balance 14. Macrae SM: Peripheral and metabolic effects of fennuraminc. in Ole body. Po,tgrad Med J (SuppJ. The slight increase in theconccnlIation of serum glucose I) 13-17, 1975. 15. McElroy IF, Miller 1M. Meyer JS: Fenfluramine. P despite Ole elevated level of insulin could be explained by chloroamphetamine and P-fluoro.unphetamine stimulation of the insulin resistance amI reduced glucose tolerancl:! which pituitary-adrenocortical activity in rat. J Pharmacol Exp Ther might have occuned in these conditions. 228(3) 593-599. 1984. It is concluded that although the mech,mism by which 16. Van ue Kar LO. Urben JH, Richardson KD, Belthea CL: fenflummine exerts its effects in short tenn lTIily be different Phannacologica1 studies on the serotoninergic .and nonserotonin from ilS long-tenn effeclS, Olis anorectic drug affects Ole mediated stimulation of prolactin imd corticosterone secretion by fenlluramine. Neuroencloc 41: 283-288. 1985. honnon'� stmus of the body, the outcome of which could be 17. Coccaro EF. Siever LJ, Kouridcs IA. Adan F. Davis KL: changes in the metabolism of fuels in the body. Central serotoninergic stimulation by fenlluramine does not affect plasma TSH level in man. Neuroendo. c 41.: 273-276. REFERENCES t988. 18. Lebovitz HE, Feldman JM: Pancreatic biogenic amine and insulin secretion in health and disease. Proc Fed Am Soc Exp 1. Rowland NE, Carlton J: Neurobiology of an anorectic Jrug: Bioi 32: 1797-1802, 1973. fcnnuraminc. Prog Ncurobiol 27: 13·62, 1986. 19. Brindley DN: Metabolicand honnonaleffectsof D·fenfluramine 2. Jespersen S, Scileel.K.rugcr J: Eviucnce for a difference in mechanism of action between fenfluramincand amphetamine. on stress situation. Clin Neuropharmac induced anorexia. J Pharm Phannac 25: 49-54, 1973. t988. 3. Al·Sieni AI, Plesled CP, Rolland Y, Brindley DN: Decreased 20. Brindley DN. Saxton J. Shahidullah H, Armstrong M: Possible incorporation of glucose into lipids and increased laclate relationship between changes in body weight, set·point and production by adipose tissue after long-tcnn treatment of rats stress metabolism after treating rats chronically with fcnflummine. Biochcm Pharmacal 34(8): 1265- I271. 1985. with D-fennuramine. Biochem Pharmacal 38: 3661·3667, 21. Turner P: Peripheral mechanism of action of fenOuramine. 1989. 4. Ani M, Boroumand A: The effect of fenOuraminc on plasma and \ I 1(suppl. 1) 586-589, CUrT Med Opin 6(Suppl. 186 t) 101-106. 1979.