intake. The animals tolerant to the anorectic effect offenfluramine had mark- edly low basal hypothalamic serotonin (5-HT) levels. In this brain area the levels of ...
0022-3565/84/2282-0446$02.0O/0 THE
JOURNAL
Copyright
OF PHARMACOLOGY
AND
0 1984 by The American
Society
EXPERIMENTAL for
Vol. 228, No.2
THERAPEUTICS
Pharmacology
and Experimental
Therapeutics
PrinLed
Possible Involvement of Endogenous the Anorectic Effect of Fenfluramine’ ANTONIO
GROPPETTI,
Department
for publication
Accepted
MARCO
of Pharmacology,
PARENTI,
School of Medicine,
October
LAURA UnWersity
DELLAVEDOVA
Opiates
and
FELICE
in the Tolerance
in USA.
to
TIRONE
of Milan, Milan, ftaly
21 , 1983
ABSTRACT tolerant rats, morphine restored the efficacy of the anorectic agent. After morphine pretreatment, fenfluramine depleted 5-HT and reduced food intake in tolerant animals. These findings, while further substantiating the importance of 5-HT in mediating fenfluramine anorexia, also suggest that endogenous opiates may play an important role in the processes through which tolerance to this drug develops. Fenfluramine reduces food intake by releasing 5-HT and tolerance to its anorectic effect would be a consequence of an inability to further release 5-HT. However, because release of 5-HT by fenfluramine seems to be modulated by opiates, repeated administration of fenfluramine mIght alter such modulatory mechanisms and tolerance to the effects of the drug would develop.
Fenflunamine is probably one of the drugs most widely used for treatment of obesity. Structurally related to amphetamine, fenfluramine lacks the central stimulant activity and is devoid of many of the side effects elicited by this drug (Ganattini et aL, 1979). As with amphetamine, however, repeated administration of fenfluramine leads to progressive changes in its effectiveness (Le-Douarec and Neveu, 1970; Ghosh and Parvathy, 1976; Lewander, 1978). The mechanisms by which fenfluramine reduces food intake have been extensively investigated in the past. It appears that
Morley and Levine, 1980), the question arises as to whether endogenous opioids are also involved in fenfluramine-induced anorexia and/or, in particular, in the development of tolerance to this drug. Here we report that opiates participate in the mechanisms by which fenfluramine reduces hypothalamic 5-HT and food intake. Moreover, we suggest that mutual interaction between opioid and serotonergic neurons may be important in the development of tolerance to the pharmacological effects of fenflunamine.
administration of fenfluramine leads loss of its effectiveness in reducing
progressive
the
ability
to
release
5-HT
is probably
the
food
most
important
neurochemical event related to this behavioral effect of fenflunamine (Costa et aL, 1971; Samanin et at., 1972; Fuxe et at., 1975; Duhault et at., 1978). Information about the mechanisms by which other
tolerance
hand,
to fenfluramine
scarce
and
largely
anorexia
develops
is, on the
incomplete.
1 This
for publication work
was
supported
November by
grants
18,
Italian
Consiglio
Richerche and Ministero Pubblica Istruzione. Preliminary at 6th European Neuroscience Congress, Malaga, Spain,
ABBREVIATIONS:
446
5-HT, serotonin;
ME-IR,
studies.
albino rats (Charles MA) weighing 180
River
Breeding Laboraused in these
to 220 g were
determinations, animals were killed either tissue levels of [Met5]enkephalin were to be exposure to high energy microwave irradiation
For the biochemical or, when
decapitation,
by
measured,
(Guidotti
4-sec
et aL, 1974).
Brains were quickly removed, dissected as described by Glowinaki and Iversen (1966) and stored at -20C. Food intake. Animals were housed individually (cages 42 x 22 x
18 cm) and maintained to the usual dry pellet
1982. from
Male Sprague-Dawley tories, Inc., Wilmington, by
Recently, elevation of [Met5]enkephalin and -endorphin concentrations has been found in the hypothalamus of rats treated with fenflunamine (Harsing et al., 1982; Dellavedova et at, 1982, 1983). As opiates have been reported to affect food intake (Grandison and Guidotti, 1977; Margules et a!., 1978; Received
Methods
Nazionale
delle
reports were presented September 1982.
[MetJenkephalin-like
on a feeding schedule which allowed free access diet for 4 hr a day (10 A.M. to 2 P.M.). Water ad Ithitum. After 2 weeks on this regimen the total 4 hr
was available of food consumption
immunoreactive
Experiments
material.
were
approximated performed
that normally consumed in 24 hr. at the end of the training period. Food
Downloaded from jpet.aspetjournals.org at ASPET Journals on June 6, 2016
to a rapid and intake. The animals tolerant to the anorectic effect offenfluramine had markedly low basal hypothalamic serotonin (5-HT) levels. In this brain area the levels of [MetJenkephalin-like immunoreactive material were, on the contrary, significantly higher in fenfluramine-tolerant animals than in controls. In tolerant animals the drug failed to further decrease 5-HT concentrations unless it was given at doses also reducing food intake. On the other hand, in acute experiments, morphine pretreatment potentiated and naloxone antagonized fenfluramine-induced depletion of striatal and hypothalamic 5-HT stores. In addition, when given to fenfluramine-
Repeated
Opiates
1984 intake (grams presentation.
per
100
b.w.) was measured
g
4 hr after
Catalepsy.
The degree
et aL
Tolerance
Fenfluramine
447
food
A Food
intake
of catalepsy
was expressed as the mean of scores (0-6) were obtained by adding from different tests, as described by
at least five total scores. Total the individual scores resulting
Dunstan
at 2 and
and
(1980).
Tissue concentrations homogenized in 50 volumes by the spectrophotofluorometric
of 5-HT. Striatum or hypothalamus was of acidified butanol. 5-HT was determined method of Curzon and Green (1970). Tissue concentrations of ME-IR Frozen tissues were rapidly (10-15 eec) homogenized in 30 volumes of ice-cold 0.1 N acetic acid, immediately heated at 100#{176}C for 5 mm and then centrifuged at 40,000 x g for 15 mm. Aliquoth of the supernatant were assayed for [Met6]enkephalin by radioimmunoassay. The antiserum used has been found to be highly specific for [Met5]enkephalin; however, interference of presently anknown endogenous compounds cannot be completely ruled out. For this reason, in this report we will use the term ME-IR. Drugs and treatments. The following compounds were useth dl-
Fenfi.
mg/kg
dafly)
(test-dose)
10
-
4
10
lo.inections
7
10
-
lfl mq/k
-
B
A
Intake
1
-
-
Food
intake
0
Saline
D
enfluramine
0
Fenfluramine
2r) rig/kq
0
Fenfluramine
40 ng/kg
In
L
8
8
a
2
6
c
S
>i
-V 0 4
C-’
2
Oi
L
C)
C
Fenfluramine
mi.)
-
-
6
6
10
Fenfl.(2nd
inj.)
-
10
-
10
-
Food
10
10
20
20 mg/kg
-
10 mg/kg
B
Intake
0
7 days)
2. Effects
of fenfiuramine (Fenfi)(10, 20 or 40 mg/kg) on food intake and Fenfi-pretreated (10 mg/kg daily) rats. A, rats were treated daily for 1 , 4 or 7 consecutive days with 10 mg/kg i.p. of dl-Fenfl. A test dose of the drug (10 mg/kg) or saline was given 24 hr after the last treatment. B, increasing test doses of d!-fenfi (1 0, 20 and 40 mg/kg) or saline were given to controls or to rats that had been treated daIly for 7 consecutive days with d!-Fenfi (10 mg/kg i.p.). Chronic treatment was suspended 24 hr before the test doses of dl-Fenfi or saline was given. Fig.
Fenfl.(lst
(10 mg/kg
in saline-
Rats were allowed
to eat for 4 hr starting
30 mm after the ijection
of
the test dose of fenfluramine or saline. Food intake was measured at the end of the 4th hour. Each value represents the mean ± S.E. for at least
8
five determinations.
0)
fenfluramine HCI, Carlo
>,
