- Email: [email protected]
Effect of dopamine-β-hydroxylase inhibitors on thermoregulation in reserpine-treated rats
EFFECT OF DOPAMINE-P-HYDROXYLASE INHIBITORS ON THERMOREGULATION IN RESERPINETREATED RATS Y. ISHII, A. YOSHIKAWA and MASAKO H~MMA Research
Laboratories.
Nippon
Kayaku
(Acqt~il
Co.. Shimo.
Kita-ku.
Tokyo
115.Japan
30 October 1975)
SummaryPDopamine-/I-hydroxylase inhibitors, disulfiram, diethyldithiocarbamatc and FD-WX. caused marked hypothermia in rats pretreated with reserpine. The hypothermic action was more prominent in 24 hr than in 48 or 72 hr after treatment with reserpine. The hypothermic action of disulfiram or diethyldithiocarbamate was inhibited by L-dihydroxyphenylalamne. but was not inhibited by plmozide. a dopamine receptor blocker. Brain dopamine and norepinephrine were almost depleted in rats treated with reserpine, but a further significant reduction of norepincphrine or a significant increase of dopamine was not observed following administration of disulliram. L-Dihydroxyphenylalanine increased brain dopamine levels in rats treated with reserpine or rcserpine plus disulfiram but did not affect norepinephrine levels.
tory bulbs and cerebellum. was removed. The adrenal glands were also quickly excised. The tissues were frozen in liquid nitrogen and stored at -25 C before use. Norepinephrine (NE). cpinephrine and dopamine (DA) were determined using fluorometric procedures described by SHELL~NBIXWK and GORDON (1971) with a slight modification. Details of the procedures have been reported previously (ISHII, HOMMA. YOSHIKAWA and UMEZAWA. 197%). The rats were treated with reserpine injection (Pharmacopoeia of Japan VIII. Daiichi Seiyaku) at a subcutaneous dose of 2.5mgikg 24 hr prior to the experiment, unless otherwise noted. The calcium salt of FD-008 and L-DOPA were synthesized in our laboratories. Other drugs used were sodium diethyldithiocarbamate (Tokyo Kasei). disulfiram (extracted from commercial tablets and recrystallized), and pimozide (kindly supplied by Dr. P. Janssen). Diethyldithiocarbamate was dissolved in saline and other drugs were suspended in 0.5”,, carboxymethylcellulose solution.
5-(4’Chloro) butylpicolinic acid (FD-008) is one of the most potent dopamine-fi-hydroxylase inhibitors among the fusaric acid derivatives synthesized by UMEZAWA.
TAKEUCHI,
HAMANO (1973). During macology (1975b)
of FD-008, observed
MIYANO,
KOSHIGOI::
the study of the general
and phar-
ISHII. NATSUCXX and UMEZAWA
that FD-008
caused a marked fall in body temperature and an inhibition of the performance on a shuttle box in rats previously treated with reserpine at a dose at which FD-008 alone had no effect. Dopamine-/?-hydroxylase inhibitors are now attracting attention as a pharmacological tool for studying the physiological role of catecholamines and the function of sympathetic nerves. The purpose of the present study was to investigate the effect of another well-known dopamine-fl-hydroxylase inhibitor, disulfiram and its active metabolite, diethyldithiocarbamate, on body temperature and tissue catecholamine levels in reserpine-treated rats and to elucidate whether the hypothermic action in reserpine-treated rats is a common property of dopamine+hydroxylase inhibitors. In addition, the effect of L-dihydroxyphenylalanine (L-DOPA) and pimozide, a selective dopamine receptor blocker, on the hypothermic action of dopamine-/?-hydroxylase inhibitors was investigated.
RESULTS
The body temperature of rats treated with reserpine (2.5 mg,‘kg SC.) fell by about 2 C during 8 hr after the injection but the temperature virtually recovered in the following day (Fig. I ). Diethyldithiocarbamate (400 mgjkg). disulfiram (I g/kg) or FD-008 (100mg/kg) administered orally 24 hr after the injection of rcserpine caused a marked decrease in body temperature. The doses of dopaminc-/i-hydroxylasc inhibitors (except FD-008) used in this experiment scarcely affected body temperature in normal rats. The dose-dependent hypothermic action of disulfiram
MATERIALS AND METHODS
Male Wistar rats weighing 20&3OOg were used. The ambient temperature was maintained at 23 k 0S”C. The body temperature was measured by a heat thermistor probe inserted into the rectum to a depth of 4 cm and retained there until a constant temperature reading was obtained. The rats serving for study on tissue catecholamines were sacrificed by decapitation and the brain. separated from the olfac359
1’. I\IIII.
,I. YOSIIIKAWA and MASAKO HOMMA Disulfiram 1 h
Fig. I. Ell’ects of oral administrntlon 01‘ FD-00X ( 100 mg/kg). disulfiram (I &kg) and ciicthqldithiocarb~umntc (DDTC. 400 mg;kg) on hod) tcmpcraturc m reacrpinctt-cated rats (A) and normal rats (HI. In rtxcrpinc-trcnted t-ah. drugs were given ?J hr after injectIon of rcsel-pine (2.5 mg kg. S.C.I.
32 1
! 0
1
2
3
4
5
6
7
HOURS in
rcserpine-treated
dislllfiram-induced rats
depended
hppothcrmic
rats
is shown
hypothermia on
action
the
in Figure in
ambient
2. The
reserpinc-treated tcmperaturc.
01‘ disulliram
ruthor
n;is
The
Fig. 3, Influence of amblent temperature on the hypothermic action of disulliram (1 g/kg,p.o.) in reserpine-treated rats.
beaker
at 28 c‘ than at 23 C‘ (Fig. 3).
As shown in Figure 3. the administration firam 14 hr after rcscrpine had a stronger mic action
compared
72 hr afttx
rcscrpine
disulfiram
with
its administration
trentmcnt.
the
drugs
M;;IS ohservcd
4X or
rcserpine
When
were given simult~~n~ousl~.
between
of’ disulhypothcrund
the potentiation 24 hr
after
the
~tdtl7inistration.
oral
The
administration
of I.-DOPA
3 hr after that of disulfiram
markedly
hypot!xrmia
induced
disulfram
treated
(Fig.
40-
rats
h> 5).
In
(600 m&kg) attenuated in
the
rescrpinc-
rcwrpinc-trca1ed
rats.
Fig. 4. Infuencc of time after injection of reserpine on the h) pothcrmic action of disulfiram (I g,/kg,p.o.). Disulfiram was administered simultaneously with reserpine (2.5 rngl kg.s.c.) 24. 48 or 72 hr after reserpine treatment.
Reserplne J
39 -
&348
100 mgikg 300mglkg 1OOOmglkg
33.
32L”...;;0
2
4
6
8
24““’ 26 28 30 32
’ ‘481
l-‘lg. 7. The dose-dependent h>pothcrm~ action liram in rcscrpine-treated rats. Disulliram was tered orally 24 hr after injection of rcscrpine kg.w.).
%---HOURS
of disuladminis(2.5 mg
Fig. 5. ElTect of L-DOPA (600 mgjkg.p.0.) on body temperature in rats treated with reserpine or reserpine plus disulliram. Disulfiram (1 g/kg.p.o.) was administered 24 hr after reserpine (1.5 mg/kg.s.c.) and L-DOPA was given 3 hr after disulfiram
DBH Table
1. Effect of orally
administered
L-DOPA
rats previously
Tissue
Amine
Brain
DA NE Epinephrine
Adrenal medulla
Control X76 * 47 52-l * 17 23.9 _ + 1.x
inhibitors
on thermoregulation
(600 mg/kg) treated
and disulfiram
361
(I g/kg)
on tissue catecholamine
levels in
with reserpine (2.5 mg,‘kg.s.c.)
Rescrpine alone
Concentration Reserpine + L-DOPA
155 k ‘6 35 & 15 12.4 f I.1
573 f 5c)* 32 * 14 Il.4 & I.2
(ng;g; adrenal: Reserpine + Disulhram 131 * 26 13 i 6 7.3 * 1.2
/lg,‘pair) Rcserpinc + L-DOPA + Disulhram 667 + IO?*1 7t4 5.5 * 3*t
Time schedule of drug administration is given in Figure 5. The means and standard errors of 6 assays are given in the Table. Statistically significnnt at P < 0.01 (*) when compared with rats given rcscrpinc alone and at P < 0.01 (t) uhen compared with rats given rcserpine and dtsulfiram.
The effect of L-DOPA, disulfiram or L-DOPA plus disulfiram on catecholamine levels in the brain and adrenal gland of normal and reserpine-treated rats is shown in Table I. The rats were sacrificed 30 hr after treatment of reserpine; namely 6 or 3 hr after administration of disulfiram or L-DOPA, respectively. The time schedule of drug administration and doses were same as those in the experiment on measurement of body temperature. After reserpine, brain NE and DA levels were lowered to 7”,, and IX”,, of levels in normal rats, respectively. and the adrenal epinephrine level was lowered to about jl”,,. L-Dihydroxyphenylalanine significantly raised the brain DA level, but not brain NE and adrenal epinephrine levels. Disulfiram did not cause a further significant reduction of brain NE. but significantly reduced epinephrinc in the adrenal gland. In rats treated with L-DOPA plus disulfiram, the brain DA level was markedly raised. but brain NE and adrenal epinephrine levels were the lowest among all groups. although the differences were not statistically significant.
.
DDTC
o
Rcserpinc
alone + DDTC
A
Rcserpine
* Pimozide
A
Reserpine
+ Pimozide
s DDTC
HOURS
Fig. 6. Effect of pimozide (10 mg/kg.s.c.) on hypothermia induced by diethyldithiocarbamate (DDTC. 400 mg/ kg.p.0.) in reserpine-treated rats. Pimozide was given I.5 hr before diethyldithiocarbamate.
The effect of pimozide. a selective dopamine receptor blocker, on the hypothermic action of diethyldithiocarbamate (400 mg/kg,p.o.) in reserpine-treated rats is shown in Figure 6. Pimozide (IO mg!kg.s.c.) given 1.5 hr prior to diethyldithiocarbamatc did not show a significant inhibition of the hypothermia induced by diethyldithiocarbamate. Pimo7ide alone had no effect on body temperature in rcserpinc-treated rats.
In the present investigation. representative compounds of two groups of dopamine-/I-hydroxylase inhibitors. disulfram or its active metabolite. diethyldithiocarbamate. and FD-008 caused marked falls in body temperature in rats previously treated with rcserpine. It suggests that the hypothermic action in reserpine-treated rats is a common property of dopamine-/I-hydroxylase inhibitors. VI.TIILANI. REICHENUEK, WISZNI~WSKA and MAKHAJ (1973) reported that fusaric acid (50 or 100 mg!kg.i.p.) and dimethylphenylthiourea (100 or 200 mg,!kg.i.p.) depressed the body temperature in normal rats. ISHIt ct trl. (1975b) also observed that fusaric acid and FD-00X lowered the body temperature in normal rats by about 2.5C at an oral dose of 100 mgikg. However, the hypothermic action of FD-00X in reserpine-treated rats was far stronger than that in normal rats. BAKN~TT and TAI~I K (1968) observed that diethyldithiocarbamate raised body tcmpcrature in reserpine-treated mice at a dose at which diethyldithiocarbamate lowered temperature in normal mice, but diethyldithiocarbamate had no effect in reserpinetreated rats. Apart from the species difference. the present finding is contradictory to the results of BAKNETT and TAHEK (196X). who report that body temperature decreased an average of I5 -16 C after reserpine (5 mg/kg,i.p.). The body temperature of reserpine-treated rats in the present experiment recovered to a range of 37-38’ C when dopamine-[Ghydroxylase inhibitors wcrc administered. The conditions of our experiment seem to be more physiologically suitable for an investigation of the effect of drugs on thermoregulation.
362
Y. ISHU. A. YOSHIlCAM’4 and MASAICO
Dopaminc-/i-h)droxylase is an enzyme which convcrts DA to NE in sympathetic nerves. brain and adrenal medulla and the reduction of NE in these tissues may be involved in the pharmacological action of dopaminc+hydroxylase inhibitors. In rcserpinctreated rats. brain NE was practically depleted. and 21 further significant reduction of NE was not observed following administration of disulfiram. It shows that the mechanism of hypothermin induced by dopamine$-hydroxyiase inhibitors in reserpinetreated rats can not bc cxplaincd in terms of a static brain NE level. The hypothermic action of disulfiram was inhibited by L-DOPA which 21lone increased the body tempcr:tturc in rescrpinc-treated rats. nnd the brnin DA Ie\cl in rats tre>lted with reserpinc ulonc or rcserpine plus disulfiram was markedly increased by I.-DOPA but the brain NE level W:IS not affected by I.-DOPA. These results suggest that brain DA ma> play an important role in the restoration of body tempel-ature in rcserpinc-treated rats. However. the hypothermic action of diethyldithiocarbamate in rcserpine-treated rats W;IS not inhibited b! pimozidc and the decrease in body temperature in reserpinctreated ruts W;IS observed when y-methyldopa. ;I DOPA-decarboxyklse inhibitor. wx given orally at ;I dose of 500 m&kg. which did not atTcct the hod! temperature in normal rats (ISHII. YOSHIF;AWA and HOMMA. unpublished observations). BAKNI:TT and TAWK (1968) suggested that dicthyldithiocarbamate raised body temperature by elevating depleted DA stores in reserpine-treated mice and lowered temperature by causing an increase in DA levels above ;I critical point in normal mice. However. ;LSfar :IS the prcsent results :u-e concerned. the hypothermic action of dopamine-[Ghydroxylase inhibitors is not solely attributable to the change of brilin DA level. Cox and THA ( 1975) suggested that in normal mice dopamincrgic mechanisms mediate the hypothermia and noradrenergic mechanisms mediate the hyperthcrmia and in reserpine-treated mice both sjstcms are involved
k)hfMA
in the mechanism that restores body temperature to normal. The hypothermia induced by dopamine-/Ihydroxylasc inhibitors and the restorution l-11 L-DOPA in rcserpinc-treated rats might be attributable to the mechanism postulated by COX and TOW, (1975). There is. however. one other possibilit) that the decrease of sympathetic nerve activity caused by dopamine-/l-hydroxylase inhibition might be involved in hypothcrmi:t in rcserpinc-treated rats. although further study is needed to :lscertain the precise mechanism involved.
REFF:RENC‘ES BAKhl.TT. A. and TAI~R, R. 1. (1968).The effects of diethyldithiocnrbamate and 1-dopa on body temperature in mice. J. P/rcr~~i~.Plitrrwccc. 20: 600 604. Cox. B. nnd TIIA. S. J. (1975). The role 01‘ dopaminc and noradrenalinc in temperature control or normal and reserpine-pretrcatcd mix. .I. PhtrrIli. Plrtrr/,ltrc. 27 : 74’ 247. ISHU. Y.. HOM~IA.M.. YOSHIMWA..A. and Llhiiz.\u.~ H. (1975a). Pharmacological actions of FD-008. a IIL‘W dopamine-l~-hydrolIvlase inhibitor. III. Ffkcts on cndogcnous blogrnlc amine It’vcls in rats. .~r~~ic~i~~fif/~~/-~o~.sc~/~. 25: 3x3 385. ISHII. Y., NATS~ GOT. K. and I!MI ZAW.A. H. (197%). Phnrmacological actions of FD-00X. ;i new dopaminc-/i&droxylasc inhibitor. II Ekctn on central n~‘rvous rystcm 111m~cc’ and rats. .4f~rrrcf~~rfrrc~/-F~~r.~~~/1. 25: 2 I3 215. SINI 1I LHIKMI<,M. K. and GOKIXI~. H. H. (1971). A rapld, wiplificd procedure [or simultancoua assay of norcpinephrine. dopamine and 5-hvdroxvtrvptaminc from bisc&te brain &xs. 4ntrl~~t. Bk&w. 391 ‘3% 372. UMIZ.AWA.H.. TAU UCHI. T.. M~~Auo. K.. Kosnlc;oi T. and HAMANO.H. (1973). Fusaric acid dericativch: the effect on dopaminc /i-h! drosylase. J. lniihior. (7.01, ro) 26: IX’). VI -rt LANI. .I.. RI IC.III SN ICC;. K.. W~s~uro\vsa,~, G. :\nd MAIICHAJ. J. (1973). The clkct of dop~~mine-/~-h)drouqIax inhibitors. dimeth~lphen~lthiourra and l’usaric acid. on locomotor and c*plorntor> activity. bod\i tempcrature. and regional distribution 01‘ biogenic ammc\ in the rat brain. PO/. .I. Phtrrirwc~. P/W,ll. 25: 559 572.
Laboratories.
Nippon
Kayaku
(Acqt~il
Co.. Shimo.
Kita-ku.
Tokyo
115.Japan
30 October 1975)
SummaryPDopamine-/I-hydroxylase inhibitors, disulfiram, diethyldithiocarbamatc and FD-WX. caused marked hypothermia in rats pretreated with reserpine. The hypothermic action was more prominent in 24 hr than in 48 or 72 hr after treatment with reserpine. The hypothermic action of disulfiram or diethyldithiocarbamate was inhibited by L-dihydroxyphenylalamne. but was not inhibited by plmozide. a dopamine receptor blocker. Brain dopamine and norepinephrine were almost depleted in rats treated with reserpine, but a further significant reduction of norepincphrine or a significant increase of dopamine was not observed following administration of disulliram. L-Dihydroxyphenylalanine increased brain dopamine levels in rats treated with reserpine or rcserpine plus disulfiram but did not affect norepinephrine levels.
tory bulbs and cerebellum. was removed. The adrenal glands were also quickly excised. The tissues were frozen in liquid nitrogen and stored at -25 C before use. Norepinephrine (NE). cpinephrine and dopamine (DA) were determined using fluorometric procedures described by SHELL~NBIXWK and GORDON (1971) with a slight modification. Details of the procedures have been reported previously (ISHII, HOMMA. YOSHIKAWA and UMEZAWA. 197%). The rats were treated with reserpine injection (Pharmacopoeia of Japan VIII. Daiichi Seiyaku) at a subcutaneous dose of 2.5mgikg 24 hr prior to the experiment, unless otherwise noted. The calcium salt of FD-008 and L-DOPA were synthesized in our laboratories. Other drugs used were sodium diethyldithiocarbamate (Tokyo Kasei). disulfiram (extracted from commercial tablets and recrystallized), and pimozide (kindly supplied by Dr. P. Janssen). Diethyldithiocarbamate was dissolved in saline and other drugs were suspended in 0.5”,, carboxymethylcellulose solution.
5-(4’Chloro) butylpicolinic acid (FD-008) is one of the most potent dopamine-fi-hydroxylase inhibitors among the fusaric acid derivatives synthesized by UMEZAWA.
TAKEUCHI,
HAMANO (1973). During macology (1975b)
of FD-008, observed
MIYANO,
KOSHIGOI::
the study of the general
and phar-
ISHII. NATSUCXX and UMEZAWA
that FD-008
caused a marked fall in body temperature and an inhibition of the performance on a shuttle box in rats previously treated with reserpine at a dose at which FD-008 alone had no effect. Dopamine-/?-hydroxylase inhibitors are now attracting attention as a pharmacological tool for studying the physiological role of catecholamines and the function of sympathetic nerves. The purpose of the present study was to investigate the effect of another well-known dopamine-fl-hydroxylase inhibitor, disulfiram and its active metabolite, diethyldithiocarbamate, on body temperature and tissue catecholamine levels in reserpine-treated rats and to elucidate whether the hypothermic action in reserpine-treated rats is a common property of dopamine+hydroxylase inhibitors. In addition, the effect of L-dihydroxyphenylalanine (L-DOPA) and pimozide, a selective dopamine receptor blocker, on the hypothermic action of dopamine-/?-hydroxylase inhibitors was investigated.
RESULTS
The body temperature of rats treated with reserpine (2.5 mg,‘kg SC.) fell by about 2 C during 8 hr after the injection but the temperature virtually recovered in the following day (Fig. I ). Diethyldithiocarbamate (400 mgjkg). disulfiram (I g/kg) or FD-008 (100mg/kg) administered orally 24 hr after the injection of rcserpine caused a marked decrease in body temperature. The doses of dopaminc-/i-hydroxylasc inhibitors (except FD-008) used in this experiment scarcely affected body temperature in normal rats. The dose-dependent hypothermic action of disulfiram
MATERIALS AND METHODS
Male Wistar rats weighing 20&3OOg were used. The ambient temperature was maintained at 23 k 0S”C. The body temperature was measured by a heat thermistor probe inserted into the rectum to a depth of 4 cm and retained there until a constant temperature reading was obtained. The rats serving for study on tissue catecholamines were sacrificed by decapitation and the brain. separated from the olfac359
1’. I\IIII.
,I. YOSIIIKAWA and MASAKO HOMMA Disulfiram 1 h
Fig. I. Ell’ects of oral administrntlon 01‘ FD-00X ( 100 mg/kg). disulfiram (I &kg) and ciicthqldithiocarb~umntc (DDTC. 400 mg;kg) on hod) tcmpcraturc m reacrpinctt-cated rats (A) and normal rats (HI. In rtxcrpinc-trcnted t-ah. drugs were given ?J hr after injectIon of rcsel-pine (2.5 mg kg. S.C.I.
32 1
! 0
1
2
3
4
5
6
7
HOURS in
rcserpine-treated
dislllfiram-induced rats
depended
hppothcrmic
rats
is shown
hypothermia on
action
the
in Figure in
ambient
2. The
reserpinc-treated tcmperaturc.
01‘ disulliram
ruthor
n;is
The
Fig. 3, Influence of amblent temperature on the hypothermic action of disulliram (1 g/kg,p.o.) in reserpine-treated rats.
beaker
at 28 c‘ than at 23 C‘ (Fig. 3).
As shown in Figure 3. the administration firam 14 hr after rcscrpine had a stronger mic action
compared
72 hr afttx
rcscrpine
disulfiram
with
its administration
trentmcnt.
the
drugs
M;;IS ohservcd
4X or
rcserpine
When
were given simult~~n~ousl~.
between
of’ disulhypothcrund
the potentiation 24 hr
after
the
~tdtl7inistration.
oral
The
administration
of I.-DOPA
3 hr after that of disulfiram
markedly
hypot!xrmia
induced
disulfram
treated
(Fig.
40-
rats
h> 5).
In
(600 m&kg) attenuated in
the
rescrpinc-
rcwrpinc-trca1ed
rats.
Fig. 4. Infuencc of time after injection of reserpine on the h) pothcrmic action of disulfiram (I g,/kg,p.o.). Disulfiram was administered simultaneously with reserpine (2.5 rngl kg.s.c.) 24. 48 or 72 hr after reserpine treatment.
Reserplne J
39 -
&348
100 mgikg 300mglkg 1OOOmglkg
33.
32L”...;;0
2
4
6
8
24““’ 26 28 30 32
’ ‘481
l-‘lg. 7. The dose-dependent h>pothcrm~ action liram in rcscrpine-treated rats. Disulliram was tered orally 24 hr after injection of rcscrpine kg.w.).
%---HOURS
of disuladminis(2.5 mg
Fig. 5. ElTect of L-DOPA (600 mgjkg.p.0.) on body temperature in rats treated with reserpine or reserpine plus disulliram. Disulfiram (1 g/kg.p.o.) was administered 24 hr after reserpine (1.5 mg/kg.s.c.) and L-DOPA was given 3 hr after disulfiram
DBH Table
1. Effect of orally
administered
L-DOPA
rats previously
Tissue
Amine
Brain
DA NE Epinephrine
Adrenal medulla
Control X76 * 47 52-l * 17 23.9 _ + 1.x
inhibitors
on thermoregulation
(600 mg/kg) treated
and disulfiram
361
(I g/kg)
on tissue catecholamine
levels in
with reserpine (2.5 mg,‘kg.s.c.)
Rescrpine alone
Concentration Reserpine + L-DOPA
155 k ‘6 35 & 15 12.4 f I.1
573 f 5c)* 32 * 14 Il.4 & I.2
(ng;g; adrenal: Reserpine + Disulhram 131 * 26 13 i 6 7.3 * 1.2
/lg,‘pair) Rcserpinc + L-DOPA + Disulhram 667 + IO?*1 7t4 5.5 * 3*t
Time schedule of drug administration is given in Figure 5. The means and standard errors of 6 assays are given in the Table. Statistically significnnt at P < 0.01 (*) when compared with rats given rcscrpinc alone and at P < 0.01 (t) uhen compared with rats given rcserpine and dtsulfiram.
The effect of L-DOPA, disulfiram or L-DOPA plus disulfiram on catecholamine levels in the brain and adrenal gland of normal and reserpine-treated rats is shown in Table I. The rats were sacrificed 30 hr after treatment of reserpine; namely 6 or 3 hr after administration of disulfiram or L-DOPA, respectively. The time schedule of drug administration and doses were same as those in the experiment on measurement of body temperature. After reserpine, brain NE and DA levels were lowered to 7”,, and IX”,, of levels in normal rats, respectively. and the adrenal epinephrine level was lowered to about jl”,,. L-Dihydroxyphenylalanine significantly raised the brain DA level, but not brain NE and adrenal epinephrine levels. Disulfiram did not cause a further significant reduction of brain NE. but significantly reduced epinephrinc in the adrenal gland. In rats treated with L-DOPA plus disulfiram, the brain DA level was markedly raised. but brain NE and adrenal epinephrine levels were the lowest among all groups. although the differences were not statistically significant.
.
DDTC
o
Rcserpinc
alone + DDTC
A
Rcserpine
* Pimozide
A
Reserpine
+ Pimozide
s DDTC
HOURS
Fig. 6. Effect of pimozide (10 mg/kg.s.c.) on hypothermia induced by diethyldithiocarbamate (DDTC. 400 mg/ kg.p.0.) in reserpine-treated rats. Pimozide was given I.5 hr before diethyldithiocarbamate.
The effect of pimozide. a selective dopamine receptor blocker, on the hypothermic action of diethyldithiocarbamate (400 mg/kg,p.o.) in reserpine-treated rats is shown in Figure 6. Pimozide (IO mg!kg.s.c.) given 1.5 hr prior to diethyldithiocarbamatc did not show a significant inhibition of the hypothermia induced by diethyldithiocarbamate. Pimo7ide alone had no effect on body temperature in rcserpinc-treated rats.
In the present investigation. representative compounds of two groups of dopamine-/I-hydroxylase inhibitors. disulfram or its active metabolite. diethyldithiocarbamate. and FD-008 caused marked falls in body temperature in rats previously treated with rcserpine. It suggests that the hypothermic action in reserpine-treated rats is a common property of dopamine-/I-hydroxylase inhibitors. VI.TIILANI. REICHENUEK, WISZNI~WSKA and MAKHAJ (1973) reported that fusaric acid (50 or 100 mg!kg.i.p.) and dimethylphenylthiourea (100 or 200 mg,!kg.i.p.) depressed the body temperature in normal rats. ISHIt ct trl. (1975b) also observed that fusaric acid and FD-00X lowered the body temperature in normal rats by about 2.5C at an oral dose of 100 mgikg. However, the hypothermic action of FD-00X in reserpine-treated rats was far stronger than that in normal rats. BAKN~TT and TAI~I K (1968) observed that diethyldithiocarbamate raised body tcmpcrature in reserpine-treated mice at a dose at which diethyldithiocarbamate lowered temperature in normal mice, but diethyldithiocarbamate had no effect in reserpinetreated rats. Apart from the species difference. the present finding is contradictory to the results of BAKNETT and TAHEK (196X). who report that body temperature decreased an average of I5 -16 C after reserpine (5 mg/kg,i.p.). The body temperature of reserpine-treated rats in the present experiment recovered to a range of 37-38’ C when dopamine-[Ghydroxylase inhibitors wcrc administered. The conditions of our experiment seem to be more physiologically suitable for an investigation of the effect of drugs on thermoregulation.
362
Y. ISHU. A. YOSHIlCAM’4 and MASAICO
Dopaminc-/i-h)droxylase is an enzyme which convcrts DA to NE in sympathetic nerves. brain and adrenal medulla and the reduction of NE in these tissues may be involved in the pharmacological action of dopaminc+hydroxylase inhibitors. In rcserpinctreated rats. brain NE was practically depleted. and 21 further significant reduction of NE was not observed following administration of disulfiram. It shows that the mechanism of hypothermin induced by dopamine$-hydroxyiase inhibitors in reserpinetreated rats can not bc cxplaincd in terms of a static brain NE level. The hypothermic action of disulfiram was inhibited by L-DOPA which 21lone increased the body tempcr:tturc in rescrpinc-treated rats. nnd the brnin DA Ie\cl in rats tre>lted with reserpinc ulonc or rcserpine plus disulfiram was markedly increased by I.-DOPA but the brain NE level W:IS not affected by I.-DOPA. These results suggest that brain DA ma> play an important role in the restoration of body tempel-ature in rcserpinc-treated rats. However. the hypothermic action of diethyldithiocarbamate in rcserpine-treated rats W;IS not inhibited b! pimozidc and the decrease in body temperature in reserpinctreated ruts W;IS observed when y-methyldopa. ;I DOPA-decarboxyklse inhibitor. wx given orally at ;I dose of 500 m&kg. which did not atTcct the hod! temperature in normal rats (ISHII. YOSHIF;AWA and HOMMA. unpublished observations). BAKNI:TT and TAWK (1968) suggested that dicthyldithiocarbamate raised body temperature by elevating depleted DA stores in reserpine-treated mice and lowered temperature by causing an increase in DA levels above ;I critical point in normal mice. However. ;LSfar :IS the prcsent results :u-e concerned. the hypothermic action of dopamine-[Ghydroxylase inhibitors is not solely attributable to the change of brilin DA level. Cox and THA ( 1975) suggested that in normal mice dopamincrgic mechanisms mediate the hypothermia and noradrenergic mechanisms mediate the hyperthcrmia and in reserpine-treated mice both sjstcms are involved
k)hfMA
in the mechanism that restores body temperature to normal. The hypothermia induced by dopamine-/Ihydroxylasc inhibitors and the restorution l-11 L-DOPA in rcserpinc-treated rats might be attributable to the mechanism postulated by COX and TOW, (1975). There is. however. one other possibilit) that the decrease of sympathetic nerve activity caused by dopamine-/l-hydroxylase inhibition might be involved in hypothcrmi:t in rcserpinc-treated rats. although further study is needed to :lscertain the precise mechanism involved.
REFF:RENC‘ES BAKhl.TT. A. and TAI~R, R. 1. (1968).The effects of diethyldithiocnrbamate and 1-dopa on body temperature in mice. J. P/rcr~~i~.Plitrrwccc. 20: 600 604. Cox. B. nnd TIIA. S. J. (1975). The role 01‘ dopaminc and noradrenalinc in temperature control or normal and reserpine-pretrcatcd mix. .I. PhtrrIli. Plrtrr/,ltrc. 27 : 74’ 247. ISHU. Y.. HOM~IA.M.. YOSHIMWA..A. and Llhiiz.\u.~ H. (1975a). Pharmacological actions of FD-008. a IIL‘W dopamine-l~-hydrolIvlase inhibitor. III. Ffkcts on cndogcnous blogrnlc amine It’vcls in rats. .~r~~ic~i~~fif/~~/-~o~.sc~/~. 25: 3x3 385. ISHII. Y., NATS~ GOT. K. and I!MI ZAW.A. H. (197%). Phnrmacological actions of FD-00X. ;i new dopaminc-/i&droxylasc inhibitor. II Ekctn on central n~‘rvous rystcm 111m~cc’ and rats. .4f~rrrcf~~rfrrc~/-F~~r.~~~/1. 25: 2 I3 215. SINI 1I LHIKMI<,M. K. and GOKIXI~. H. H. (1971). A rapld, wiplificd procedure [or simultancoua assay of norcpinephrine. dopamine and 5-hvdroxvtrvptaminc from bisc&te brain &xs. 4ntrl~~t. Bk&w. 391 ‘3% 372. UMIZ.AWA.H.. TAU UCHI. T.. M~~Auo. K.. Kosnlc;oi T. and HAMANO.H. (1973). Fusaric acid dericativch: the effect on dopaminc /i-h! drosylase. J. lniihior. (7.01, ro) 26: IX’). VI -rt LANI. .I.. RI IC.III SN ICC;. K.. W~s~uro\vsa,~, G. :\nd MAIICHAJ. J. (1973). The clkct of dop~~mine-/~-h)drouqIax inhibitors. dimeth~lphen~lthiourra and l’usaric acid. on locomotor and c*plorntor> activity. bod\i tempcrature. and regional distribution 01‘ biogenic ammc\ in the rat brain. PO/. .I. Phtrrirwc~. P/W,ll. 25: 559 572.