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Comparison of the effects of fenfluramine and other anorectic agents in different feeding and drinking paradigms in rats
Life Sciences, Vol. 36, pp. 2295-2300 Printed in the U.S.A.
COMP~N~
~
AGENTS~D~FEED~
Pergamon Press
~S~FENFLURAMINE~OTHER~RECTIC ~ D ~ P A R A D I ~
~RATS
Neil E. Rowland, Seymour M. Antelman, Timothy J. Bartness Department of Psychology and Center for Neurobiological Sciences, University of Florida, Gainesville, Florida 32611 and Department of Psychiatry, University of Pittsburgh, Pittsburgh, Pennsylvania 15260 (Received in final form April 3, 1985)
S~mary Fenfluramine (2.5 and 5 mg/kg) significantly suppressed the food intake of rats following food deprivation, administration of 2-deoxy-D-glueose (2DG) , and during tail pressure. This suggests that fenfluramine has relatively general anorectic potency. Other "serotonergic" anorectics were studied for comparison. In a second experiment we determined that norfenfluramine and quipazine greatly suppressed food intake following food deprivation but, at the same doses, had relatively small effects on water intake following water deprivation. This was true for intraperitoneal and cerebroventricular routes of administration. The data have relevance for specificity of action of these agents and for the possible contribution of dopamine antagonist properties. Anorectic agents are of potential clinical use in cases of obesity. There can be little d i s a g ~ t that the anorectic potency of a potential clinical anorectic should be tested in multiple feeding paradigms (e.g., i), and that sc~e specificity for feeding relative to another behavior (e.g., water intake) should be demonstrated. Many laboratory studies have focussed on food deprivation paradigms, mostly in rats. However, it has been noted that feeding in starved rats is hardly likely to be a viable model of excessive eating in overweight h ~ a n s (1-8). Instead sc~e human overeating may be stress-related and so animal models such as tail pressure (1-8) and glucoprivation (9) may be more appropriate. The aim of the present work is twofold: first, to explicitly ccmpare the efficacy of several anorectics administered i.p. in three different feeding paradigms; second, to examine and conpaz~ the effects of i.p. vs intracerebroventricular (i.c.v.) adainistration of anorectics on food or water intake following acute deprivation. The main focus of the studies were the "serotonergic" anorectics quipazine, fenfluramine and its dealkylated metabolite, norfenfluramine. Experiment i. Effects of various anorectics, administered i.p., on eating during tail pressure, 2-deoxy-D-gluoose g lucoprivation, or after food deprivation. Three different feeding paradigms were used in this experiment: (i) Feeding after a 24 h fast, (ii) Eating in non-deprived rats during tail pressure stress (2,3) , (iii) Gluaoprivic feeding induced by the glucose antimetabolite 2-deoxy-D-glucose (2DG) (e.g., 9). In the present experiment 0024-3205/85 $3.00 + .00 Copyright (c) 1985 Pergamon Press Ltd.
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we have oDmpared the efficacy of several anorectics in these three feeding paradigms. Methods Male Sprague-Dawley rats weighing 230-310 g (Zivic Miller Laboratories) were used in this study. They were housed individually in stainless steel cages in a vivarium maintained at 23°C with lights on 8 a.m.-8 p.m. Chow pellets and tap w a t e r w e r e available unless stated. All tests were performed in the morning between 9 and 12 a.m. Each rat was used in only one test. The anorectic drugs were administered i.p. in a volume of 1 ml/kg of saline vehicle, 30 min prior to a feeding test. The latter consisted of measuring the const~ption of Purina chow pellets from the floor of the cage during the test period. Any spillage was included in the weight of the uneaten food. Deprivation Rats were deprived of food for 24 h and the amount of chow consumed in 1 h recorded in the home cage. Tail Pressure Rats were screened for tail pressure-induced eating (3) prior to the main experiment. These tests were conducted in circular stainless steel bowls and the pressure applied via a calibrated piston-inflated tail cuff. On the test day, the rats received five 120 sec tail pressure trials with a i0 min intertrial interval. The rats never were observed to eat in these intertrial intervals and so the total food constmption was exclusively during the pressure periods. 2-Deoxy-D-Glucose For several days prior to and on the day of the 2DG experiment the animals were presented a dish of wet chow mash at 8:30 evex~ morning for 30 min. This was to ensure that they would eat a meal and, therefore, be satiated at 9 a.m. On the test day, food was removed at 9 a.m. and the animals were injected with 2DG (350 mg/kg i.p.) . At 10:30 the drugs were administered i.p. and at ii a.m. food was presented for a 1 h feeding test. All of these manipulations occurred in the hc~e cage. This paradigm is a modified delayed feeding test designed so that all of the intake induced by the 2DG occurred in a 1 h period, i.e., the same total duration as the deprivation and tail pressure eating tests. Data Analysis All of the data were expressed as a percentage of the respective saline treated control group mean. Comparisons were made both within and between treatments by analyses of variance and post hoc conparisons (Duncan). Drugs The drugs used in the study were quipazine maleate (Miles Laboratories 1.5, 3 and 5 mg/kg), the hydrochloride salt of dl-fenfluramine (A. H. Robins, 2.5 and 5 r~/kg), and fluoxetine hydrochloride (Lilly, i0 ml/kg) . All drug doses refer to the weight of the salt. Fluoxetine solutions were warmed to assist dissolution prior to administration.
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Anorectics and Ingestion
2297
Results The percentage suppressions of food intake are shown in Table I along with the absolute intakes after vehicle treatment. All dose levels of all of the drugs produced reliable suppressions of food intake in 24 h food deprived animals, F(6,48)=18.9, p<0.001. TABLE I EFFECT OF VARIOUS ANORF/~ICS ON FOOD INTAKE, EXPRESSED AS M+SE P ~ OF SALINE CONTROL, IN FOOD DEPRIVATION, --2-DEOXY-D-GLUOOSE, AND TAIL PRESSURE PARADI6~4S Drug, Dose (mg/kg)
24 h Food Deprivation
2DG (350 mg/kg )
vehicle Quipazine, 1.5 Quipazine, 3.0 Quipazine, 5.0 Fenfluramine, 2.5 Fenfluramine, 5.0 Fluoxetine, i0.0
100-+6 (6.4 g) 73-+7* 45-+7* 50-+4* 24-+7* 19-+7* 33-+5*
100-+17 129-+14T 63-+17 76-+22 42-+9* 57-+IIT 78-+16
(2.7 g)
Tail Pressure 100-+20 --n.d. --n. d. 58-+26 --n.d. i-+i* 65-+33
(0.9 g)
Shown are M-+SE for groups of 5-22 (n.d.=not deteunined) *p43.05 less than respective saline vehicle condition %p<0.05 2DG effect different from food deprivation and tail pressure The anorectic drugs also had a statistically reliable effect upon 2DG food intake, F(6,54)=3.4, p<0.01. However, the post hoc comparisons showed that this suppression was reliable only for the fenfluramine (2.5 mg/k~ treatment. The main effect of drugs in the tail pressure paradigm did not reach customary levels of significance (F(3,31)=2.8, 0.06>p>0.05), but the post hoc ~mparison revealed that fenfluramine did have a statistically significant effect (p~0.05 vs. vehicle). Indeed, the animals ate virtually nothing after fenfluramine. When the main drug effects were cc~pared across the feeding paradigms it was found that quipazine (I. 5 mg/kg) had a significantly different effect on food deprivation (a decrease) and on 2DG (nonsignificant increase). The effect of fenfluramine (5 mg/kg) likewise showed a significant drug effect with its effect on 2DG induced feeding being significantly less than that in the tail pinch or food deprivation paradigms (p<0.01). No other crossparadigm cc~parisons were statistically significant. In the tail pinch paradigm the effects of the drugs on total food intake, reported in Table I, are a good reflection of the total amount of time spent eating or gnawing the food pellets and of the total percentage of trials on which tail pressure-induced behavior was initiated. Of particular note is the fact that the fenfluramine treated animals initiated tail pinchinduced eating in only 9% of the trials and food-directed oral behavior for less than 10% of the duration of saline-treated controls. These latter ate or gnawed for 201±28 sec out of the total maxim~n of 600 sec. Discussion The anorectic effect of these various agents in the food deprivation paradigm is expected and consistent with many previous reports (e.g., 2,4,10)
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Anorectics and Ingestion
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The effects in the 2DGparadi~nwere less consistent and, in general, indicated that these drugs were less effective in this paradigm than after food deprivation. We cannot presently state whether this is because the 2DG induced less food intake than did food deprivation, or whether the anorectics really act differently in these two paradigms. In the tail pressure paradigm, although only one dose of each drug was tested, there is a trend toward slightly less potency than in the food deprivation paradigmwith the notable exception of fenfluramine. This agent is particularly interesting because it totally abolished all oral behaviors in this acute testing paradigm. We have also shown that tail pressure administered to food deprived rats will overcome the anorectic effect of amphetamine, but not that of fenfluramine (4,6). These data are not inconsistentwith the general notion that fenfluramine interferes with the expression of stress-relatedeating. Insofar as tail pressure, and possibly the 2DG paradigms are models for stress-related eating inhumans, any differences frcm theunrealistic deprivation are of potential clinical interest. We have previously shown that fenfluramine-treated rats which are totally anorectic during tail pressure tests develop tolerance to the initial suppression of the behavior (2,4). This tolerance is similar to that observed in rats in other feeding paradigmswith fenfluramine. Experiment 2. Effects of qui~azine and norfenfluramine .... administered peripherally or centrally on food and water intake. The purpose of this experiment was twofold: (i) to ascertain whether the anorectic effect of quipazine and norfenfluramine was selective for food intake after food deprivation, con~oared with water intake after water deprivation; (ii) to determine whether that selectivity would also be evident after i.c.v, injection of these agents at doses which have been shown to produce anorexia. The reason for using norfenfluramine instead of fenfluramine is that this dealkylated metabolite is about twice as potent an anorectic both peripherally and i.c.v. (e.g., 4) , and thus less drug was
r~ui~. Methods Adult male Sprague-Dawley rats housed as before, were used for the i.p. experiments. Adult male Long-Evans rats (University of Florida, Psychology Colony) were used for the i.c.v, studies. The food intake experiments were performed after 24 h food deprivation as described in Experiment i. The water intake experiments w e r e p e r f o r m e d b y g i v i n g w a t e r in tubes, calibrated to the nearest 0.1 ml, following 24 h water deprivation. Food was available during this deprivation period but not during the drinking tests. I.p. injections in a volt,he of 1 ml/kg were given 30 min prior to the tests, while the i.c.v, injections in a voltm]e of i0 D1 were given immediately prior to the ingestion test. The animalswere stereotaxically fitted with indwelling lateral ventricular cannulas (flat skull coordinates: L 1.5 m m o n bregma, V - 3 . 5 m m from skull) under pentobarbital anesthesia at least 1 week prior to these experiments. Results The results are shown in Table II. As expected, both quipazine and norfenfluramine reduced food intake with i.p. and i.c.v, routes of injection.
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Anorectics and Ingestion
2299
T~RLV. II FOOD AND WATER INTAKES EXPRESSED AS % OF SALINE ~ I T I O N IN THE FIRST HOUR F O ~ N G 24 H FOOD OR WATER DEPRIVATION, RESPECYIVELY, AND INTRAPERITONEAL OR CEREBROVENIS~ICUIAR INJECTICN OF SALINE, QUIPAZINE, OR NOI~ENFI//RAMINE Drug, Dose Intraperitoneal Saline Quipazine (5 mg/kg) Norfenfluramine (2.5 mg/kg) Cerebroventricular Saline Quipazine (200 ~g) Norfenfluramine (I00 Dg)
Food Intake
Water Intake
100+6 47_+7*% 19±4*%
(6.4 g)
100±14 80±14 46±10"
(10.9 ml)
100_+10 27±II*T 48±6*%
(6.5 g)
100±9 76±13 86±11
(15.4 ml)
Shown are MiSE % for groups of 6-10 rats *p~.05 below saline vehicle condition %p<0.05 difference between food and water paradigms The effects on water intake were in each case significantly less than the effects on food intake. The only significant reduction in water intake from the vehicle condition was with i.p. norfenfluramine. Notice also with regard to food intake, quipazine was more effective i.c.v, than i.p. while the converse was true for norfenfluramine despite the same relative i.c.v./ i.p. dosage for each agent. Discussion This experiment demonstrates that the anorectic agents quipazine and norfenfluramine both have a relative specificity for food intake as cc~npared to water intake. Further, that specificity is also found following i.c.v, injection. We note, however, that at the doses tested the relative potency of norfenfluramine and quipazine on food intake seem to be reversed in the i.p. as opposed to the i.c.v, studies. That is, following the peripheral administration, norfenfluramJ_ne seems to be disportionately effective compared with that following central administration. This is circumstantial evidence that some of the actions of norfenfluramine may be of a peripheral origin. General Discussion It is widely-believed that fenfluramine exerts its anorectic properties via release of brain 5HT (i0). However, recent evidence also suggests that both the d- and l-enanticmers may have some dopamine antagonist properties (i). The present studies with the racemic mixture do not bear directly upon the enantiomeric specificity, but are of interest in conloarison with other studies using more specific dopamine antagonists. Many neuroleptics can inhibit both food and water intake. However, water intake is attenuated by butyrophenones at doses about one-half those needed to reduce food intake (12). This strongly suggests that fenfluramine, which preferentially blocks food rather than water intake, does not work via a neuroleptic-like mechanism. Also, haloperidol (0.I mg/kg) had no effect on food intake of 24 hr deprived rats (12), or tail-pressure-induced eating (3), but did partially attenuate 2DG-induced feeding (12). The profile of fenfluramine in the present experiments is clearly different: a smaller effect on 2DG than deprivation-
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Vol. 36, No. 24, 1985
induced feeding, and a large suppression of tail-pressure-induced feeding. The apparent dopamine-blocking properties of fenfluramine thus appear to contribute minimally to its anorectic action. Quipazine has relatively potent serotonin receptor agonist properties (13) and, together with fenfluramine, has been used to support a central serotonergic involvement in feeding (i0). However, recent studies from our laboratories have questioned the similarity of action of these agents as well as the issue of central 5HT involvement (4,14-16). We have emphasized that these agents may exert anorexia via quite different mechanisms (e.g., central vs. peripheral routes). The fact that central administrations can attenuate intake does not guarantee that this is their normal avenue of action on feeding; indeed, the fact that such high doses are needed i.c.v, argues against a CNS involvement and/or a locus r~mote from the cerebral ventricles. Acknowledgement We thank Donna Kocan for assistance. Supported in part by RSDA MH00738 to SMA. Address correspondence to N. Rowland, Department of Psychology, University of Florida, Gainesville, Florida 32611. References i. 2. 3. 4.
5. 6. 7. 8. 9.
i0. ii.
12. 13. 14. 15. 16.
A.S. LEVINE, J. KNEIP, M. GRACE AND J.E. MORLEY, Pharmac. Bioche~. Behav. 18: 19-23, (1983). N.---E.ROWIAND AND S.M. ~ , Science 191: 310-312, (1976). S.M. ~ , H. SZECHUIMAN, P. CHIN AND A.E. FISHER, Brain Res. 99: 219-227, (1975). S.M. ANTEI/4_AN, N. RC~K26ND AND D. KOCAN, In: Anorectic ~@ents: Mechanisms of Action and Tolerance edited by S. GRATTINI AND R. SAMANIN, 45-62, Raven Press, New York, (1981). J.E. MORLEY, A.S. LEVINE AND N.E. ROWIAND, Life Sci. 32: 2169-2187, (1983). S.M. ANTE[MAN, A.R. C A G G I ~ , A.J. EICHLER AND R.R. LUCIK, Curr. Med. Res. Opin. 6 (Suppl. i): 73-82, (1979). T.W. ROBBINS AND P.J. FRAY, Appetite i: 103-133, (1980). M.B. WALLACH, M. DAWBER, M. MCMAHON A ~ C. ROGERS, Pharmac. Biochem. Behav 6: 529-531, (1977). A.N. EPSTEIN, S. NICOLAIDIS AND R. MISELIS, In: Neural Integration of Physiological Mechanisms and Behaviour edited by G.J. ~ S O N AND F.R. CALARESU, 148-168, Toronto Univ. Press, (1975). R. SAMANIN, T. MI~Z~INI AND S. GARATTINI, Prog. Neuropsychopharmacol. 4: 363-369, (1980). T. MENNINI, A DEBIASI, E. BORRONI, C. BENDOTTI, F. BORSINI, R. SAMANIN AND S. GARATrINI, In: Anorectic Agents: Mechanisms of Action and Tolerance edited by S. GARATrINI AND R. SAMANIN, 87-100, Raven Press, New York, (1981). N. ROWLAND AND D.J. ENGLE, Pharmac. Biochem. Behav. 7: 295-301, (1977). R. SAMANIN, C. BENIX3T2I, F. MIRANDA AND S. GARATrINI. J. Pharm. pharmacol. 29: 53-54, (1977). N. ROWLAND, S.M. ~ AND D. KOCAN, Eur. J. Pharmacol. 81: 57-66, (1982). J. CARLTON AND N. ROWLAND, Pharmac. Biochem. Behav. 20: 739-745, (1984). N. ROWLAND AND J. CARLTON, Life Sci. 34: 2495-2499, (1984).
COMP~N~
~
AGENTS~D~FEED~
Pergamon Press
~S~FENFLURAMINE~OTHER~RECTIC ~ D ~ P A R A D I ~
~RATS
Neil E. Rowland, Seymour M. Antelman, Timothy J. Bartness Department of Psychology and Center for Neurobiological Sciences, University of Florida, Gainesville, Florida 32611 and Department of Psychiatry, University of Pittsburgh, Pittsburgh, Pennsylvania 15260 (Received in final form April 3, 1985)
S~mary Fenfluramine (2.5 and 5 mg/kg) significantly suppressed the food intake of rats following food deprivation, administration of 2-deoxy-D-glueose (2DG) , and during tail pressure. This suggests that fenfluramine has relatively general anorectic potency. Other "serotonergic" anorectics were studied for comparison. In a second experiment we determined that norfenfluramine and quipazine greatly suppressed food intake following food deprivation but, at the same doses, had relatively small effects on water intake following water deprivation. This was true for intraperitoneal and cerebroventricular routes of administration. The data have relevance for specificity of action of these agents and for the possible contribution of dopamine antagonist properties. Anorectic agents are of potential clinical use in cases of obesity. There can be little d i s a g ~ t that the anorectic potency of a potential clinical anorectic should be tested in multiple feeding paradigms (e.g., i), and that sc~e specificity for feeding relative to another behavior (e.g., water intake) should be demonstrated. Many laboratory studies have focussed on food deprivation paradigms, mostly in rats. However, it has been noted that feeding in starved rats is hardly likely to be a viable model of excessive eating in overweight h ~ a n s (1-8). Instead sc~e human overeating may be stress-related and so animal models such as tail pressure (1-8) and glucoprivation (9) may be more appropriate. The aim of the present work is twofold: first, to explicitly ccmpare the efficacy of several anorectics administered i.p. in three different feeding paradigms; second, to examine and conpaz~ the effects of i.p. vs intracerebroventricular (i.c.v.) adainistration of anorectics on food or water intake following acute deprivation. The main focus of the studies were the "serotonergic" anorectics quipazine, fenfluramine and its dealkylated metabolite, norfenfluramine. Experiment i. Effects of various anorectics, administered i.p., on eating during tail pressure, 2-deoxy-D-gluoose g lucoprivation, or after food deprivation. Three different feeding paradigms were used in this experiment: (i) Feeding after a 24 h fast, (ii) Eating in non-deprived rats during tail pressure stress (2,3) , (iii) Gluaoprivic feeding induced by the glucose antimetabolite 2-deoxy-D-glucose (2DG) (e.g., 9). In the present experiment 0024-3205/85 $3.00 + .00 Copyright (c) 1985 Pergamon Press Ltd.
2296
Anorectics and Ingestion
Vol. 36, No. 24, 1985
we have oDmpared the efficacy of several anorectics in these three feeding paradigms. Methods Male Sprague-Dawley rats weighing 230-310 g (Zivic Miller Laboratories) were used in this study. They were housed individually in stainless steel cages in a vivarium maintained at 23°C with lights on 8 a.m.-8 p.m. Chow pellets and tap w a t e r w e r e available unless stated. All tests were performed in the morning between 9 and 12 a.m. Each rat was used in only one test. The anorectic drugs were administered i.p. in a volume of 1 ml/kg of saline vehicle, 30 min prior to a feeding test. The latter consisted of measuring the const~ption of Purina chow pellets from the floor of the cage during the test period. Any spillage was included in the weight of the uneaten food. Deprivation Rats were deprived of food for 24 h and the amount of chow consumed in 1 h recorded in the home cage. Tail Pressure Rats were screened for tail pressure-induced eating (3) prior to the main experiment. These tests were conducted in circular stainless steel bowls and the pressure applied via a calibrated piston-inflated tail cuff. On the test day, the rats received five 120 sec tail pressure trials with a i0 min intertrial interval. The rats never were observed to eat in these intertrial intervals and so the total food constmption was exclusively during the pressure periods. 2-Deoxy-D-Glucose For several days prior to and on the day of the 2DG experiment the animals were presented a dish of wet chow mash at 8:30 evex~ morning for 30 min. This was to ensure that they would eat a meal and, therefore, be satiated at 9 a.m. On the test day, food was removed at 9 a.m. and the animals were injected with 2DG (350 mg/kg i.p.) . At 10:30 the drugs were administered i.p. and at ii a.m. food was presented for a 1 h feeding test. All of these manipulations occurred in the hc~e cage. This paradigm is a modified delayed feeding test designed so that all of the intake induced by the 2DG occurred in a 1 h period, i.e., the same total duration as the deprivation and tail pressure eating tests. Data Analysis All of the data were expressed as a percentage of the respective saline treated control group mean. Comparisons were made both within and between treatments by analyses of variance and post hoc conparisons (Duncan). Drugs The drugs used in the study were quipazine maleate (Miles Laboratories 1.5, 3 and 5 mg/kg), the hydrochloride salt of dl-fenfluramine (A. H. Robins, 2.5 and 5 r~/kg), and fluoxetine hydrochloride (Lilly, i0 ml/kg) . All drug doses refer to the weight of the salt. Fluoxetine solutions were warmed to assist dissolution prior to administration.
Vol. 36, No. 24, 1985
Anorectics and Ingestion
2297
Results The percentage suppressions of food intake are shown in Table I along with the absolute intakes after vehicle treatment. All dose levels of all of the drugs produced reliable suppressions of food intake in 24 h food deprived animals, F(6,48)=18.9, p<0.001. TABLE I EFFECT OF VARIOUS ANORF/~ICS ON FOOD INTAKE, EXPRESSED AS M+SE P ~ OF SALINE CONTROL, IN FOOD DEPRIVATION, --2-DEOXY-D-GLUOOSE, AND TAIL PRESSURE PARADI6~4S Drug, Dose (mg/kg)
24 h Food Deprivation
2DG (350 mg/kg )
vehicle Quipazine, 1.5 Quipazine, 3.0 Quipazine, 5.0 Fenfluramine, 2.5 Fenfluramine, 5.0 Fluoxetine, i0.0
100-+6 (6.4 g) 73-+7* 45-+7* 50-+4* 24-+7* 19-+7* 33-+5*
100-+17 129-+14T 63-+17 76-+22 42-+9* 57-+IIT 78-+16
(2.7 g)
Tail Pressure 100-+20 --n.d. --n. d. 58-+26 --n.d. i-+i* 65-+33
(0.9 g)
Shown are M-+SE for groups of 5-22 (n.d.=not deteunined) *p43.05 less than respective saline vehicle condition %p<0.05 2DG effect different from food deprivation and tail pressure The anorectic drugs also had a statistically reliable effect upon 2DG food intake, F(6,54)=3.4, p<0.01. However, the post hoc comparisons showed that this suppression was reliable only for the fenfluramine (2.5 mg/k~ treatment. The main effect of drugs in the tail pressure paradigm did not reach customary levels of significance (F(3,31)=2.8, 0.06>p>0.05), but the post hoc ~mparison revealed that fenfluramine did have a statistically significant effect (p~0.05 vs. vehicle). Indeed, the animals ate virtually nothing after fenfluramine. When the main drug effects were cc~pared across the feeding paradigms it was found that quipazine (I. 5 mg/kg) had a significantly different effect on food deprivation (a decrease) and on 2DG (nonsignificant increase). The effect of fenfluramine (5 mg/kg) likewise showed a significant drug effect with its effect on 2DG induced feeding being significantly less than that in the tail pinch or food deprivation paradigms (p<0.01). No other crossparadigm cc~parisons were statistically significant. In the tail pinch paradigm the effects of the drugs on total food intake, reported in Table I, are a good reflection of the total amount of time spent eating or gnawing the food pellets and of the total percentage of trials on which tail pressure-induced behavior was initiated. Of particular note is the fact that the fenfluramine treated animals initiated tail pinchinduced eating in only 9% of the trials and food-directed oral behavior for less than 10% of the duration of saline-treated controls. These latter ate or gnawed for 201±28 sec out of the total maxim~n of 600 sec. Discussion The anorectic effect of these various agents in the food deprivation paradigm is expected and consistent with many previous reports (e.g., 2,4,10)
2298
Anorectics and Ingestion
Vol. 36, No. 24, 1985
The effects in the 2DGparadi~nwere less consistent and, in general, indicated that these drugs were less effective in this paradigm than after food deprivation. We cannot presently state whether this is because the 2DG induced less food intake than did food deprivation, or whether the anorectics really act differently in these two paradigms. In the tail pressure paradigm, although only one dose of each drug was tested, there is a trend toward slightly less potency than in the food deprivation paradigmwith the notable exception of fenfluramine. This agent is particularly interesting because it totally abolished all oral behaviors in this acute testing paradigm. We have also shown that tail pressure administered to food deprived rats will overcome the anorectic effect of amphetamine, but not that of fenfluramine (4,6). These data are not inconsistentwith the general notion that fenfluramine interferes with the expression of stress-relatedeating. Insofar as tail pressure, and possibly the 2DG paradigms are models for stress-related eating inhumans, any differences frcm theunrealistic deprivation are of potential clinical interest. We have previously shown that fenfluramine-treated rats which are totally anorectic during tail pressure tests develop tolerance to the initial suppression of the behavior (2,4). This tolerance is similar to that observed in rats in other feeding paradigmswith fenfluramine. Experiment 2. Effects of qui~azine and norfenfluramine .... administered peripherally or centrally on food and water intake. The purpose of this experiment was twofold: (i) to ascertain whether the anorectic effect of quipazine and norfenfluramine was selective for food intake after food deprivation, con~oared with water intake after water deprivation; (ii) to determine whether that selectivity would also be evident after i.c.v, injection of these agents at doses which have been shown to produce anorexia. The reason for using norfenfluramine instead of fenfluramine is that this dealkylated metabolite is about twice as potent an anorectic both peripherally and i.c.v. (e.g., 4) , and thus less drug was
r~ui~. Methods Adult male Sprague-Dawley rats housed as before, were used for the i.p. experiments. Adult male Long-Evans rats (University of Florida, Psychology Colony) were used for the i.c.v, studies. The food intake experiments were performed after 24 h food deprivation as described in Experiment i. The water intake experiments w e r e p e r f o r m e d b y g i v i n g w a t e r in tubes, calibrated to the nearest 0.1 ml, following 24 h water deprivation. Food was available during this deprivation period but not during the drinking tests. I.p. injections in a volt,he of 1 ml/kg were given 30 min prior to the tests, while the i.c.v, injections in a voltm]e of i0 D1 were given immediately prior to the ingestion test. The animalswere stereotaxically fitted with indwelling lateral ventricular cannulas (flat skull coordinates: L 1.5 m m o n bregma, V - 3 . 5 m m from skull) under pentobarbital anesthesia at least 1 week prior to these experiments. Results The results are shown in Table II. As expected, both quipazine and norfenfluramine reduced food intake with i.p. and i.c.v, routes of injection.
Vol. 36, No. 24, 1985
Anorectics and Ingestion
2299
T~RLV. II FOOD AND WATER INTAKES EXPRESSED AS % OF SALINE ~ I T I O N IN THE FIRST HOUR F O ~ N G 24 H FOOD OR WATER DEPRIVATION, RESPECYIVELY, AND INTRAPERITONEAL OR CEREBROVENIS~ICUIAR INJECTICN OF SALINE, QUIPAZINE, OR NOI~ENFI//RAMINE Drug, Dose Intraperitoneal Saline Quipazine (5 mg/kg) Norfenfluramine (2.5 mg/kg) Cerebroventricular Saline Quipazine (200 ~g) Norfenfluramine (I00 Dg)
Food Intake
Water Intake
100+6 47_+7*% 19±4*%
(6.4 g)
100±14 80±14 46±10"
(10.9 ml)
100_+10 27±II*T 48±6*%
(6.5 g)
100±9 76±13 86±11
(15.4 ml)
Shown are MiSE % for groups of 6-10 rats *p~.05 below saline vehicle condition %p<0.05 difference between food and water paradigms The effects on water intake were in each case significantly less than the effects on food intake. The only significant reduction in water intake from the vehicle condition was with i.p. norfenfluramine. Notice also with regard to food intake, quipazine was more effective i.c.v, than i.p. while the converse was true for norfenfluramine despite the same relative i.c.v./ i.p. dosage for each agent. Discussion This experiment demonstrates that the anorectic agents quipazine and norfenfluramine both have a relative specificity for food intake as cc~npared to water intake. Further, that specificity is also found following i.c.v, injection. We note, however, that at the doses tested the relative potency of norfenfluramine and quipazine on food intake seem to be reversed in the i.p. as opposed to the i.c.v, studies. That is, following the peripheral administration, norfenfluramJ_ne seems to be disportionately effective compared with that following central administration. This is circumstantial evidence that some of the actions of norfenfluramine may be of a peripheral origin. General Discussion It is widely-believed that fenfluramine exerts its anorectic properties via release of brain 5HT (i0). However, recent evidence also suggests that both the d- and l-enanticmers may have some dopamine antagonist properties (i). The present studies with the racemic mixture do not bear directly upon the enantiomeric specificity, but are of interest in conloarison with other studies using more specific dopamine antagonists. Many neuroleptics can inhibit both food and water intake. However, water intake is attenuated by butyrophenones at doses about one-half those needed to reduce food intake (12). This strongly suggests that fenfluramine, which preferentially blocks food rather than water intake, does not work via a neuroleptic-like mechanism. Also, haloperidol (0.I mg/kg) had no effect on food intake of 24 hr deprived rats (12), or tail-pressure-induced eating (3), but did partially attenuate 2DG-induced feeding (12). The profile of fenfluramine in the present experiments is clearly different: a smaller effect on 2DG than deprivation-
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Anorectics and Ingestion
Vol. 36, No. 24, 1985
induced feeding, and a large suppression of tail-pressure-induced feeding. The apparent dopamine-blocking properties of fenfluramine thus appear to contribute minimally to its anorectic action. Quipazine has relatively potent serotonin receptor agonist properties (13) and, together with fenfluramine, has been used to support a central serotonergic involvement in feeding (i0). However, recent studies from our laboratories have questioned the similarity of action of these agents as well as the issue of central 5HT involvement (4,14-16). We have emphasized that these agents may exert anorexia via quite different mechanisms (e.g., central vs. peripheral routes). The fact that central administrations can attenuate intake does not guarantee that this is their normal avenue of action on feeding; indeed, the fact that such high doses are needed i.c.v, argues against a CNS involvement and/or a locus r~mote from the cerebral ventricles. Acknowledgement We thank Donna Kocan for assistance. Supported in part by RSDA MH00738 to SMA. Address correspondence to N. Rowland, Department of Psychology, University of Florida, Gainesville, Florida 32611. References i. 2. 3. 4.
5. 6. 7. 8. 9.
i0. ii.
12. 13. 14. 15. 16.
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