- Email: [email protected]
Effects of chlordiazepoxide, amphetamine and their combinations on avoidance behaviour of reserpinized mice
Pharmacological Research Communications, VoL
9, No. 9, 1977
879
E F F E C T S OF C H L O R D I A Z E P O X I D E , AMPHETAMI NE AND THEIR COM/3INATIONS ON AVOIDANCE BEHAVIOUR OF R E S E R P I N I Z E D MICE M a r i o Sansone Laboratorio di Pstcobiologia e Psicofarmacologia, C.N.R. via Reno n. I - 0 0 1 9 8 R o m e , Italy
Receivedin final form 23 August 1977
SUMMARY C h l o r d i a z e p o x i d e and a m p h e t a m i n e , give n a lone or in c o m b i n a u o n , have been t e s t e d in s h u t l l e - b q x t r a i n e d m i c e , 24 h r s a f t e r a p r e t r e a t m e n t with r e s e r p i n e ( 1 .5 mg/kg ). When give n a l o n e , both c h o l o r d i a z e p o x i d e and a m p h e t a mine i m p r o v e d a v o i d a n c e b e h a v i o u r of r e s e r p i n i z e d m i c e . H o w e v e r , the a n t a g o n i s t i c effect t o w a r d s the a v o i d a n c e d e p r e s s i o n p r o d u c e d by r e s e r p i n e was much m o r e m a r k e d when the two d r u g s w e r e c o m b i n e d .
INTRODUCTION
It h a s been often o b s e r v e d that c h l o r d i a z e p o x i d e , when given a lone at low d o s e s , e x e r t e d s t i m u l a t i n g e f f e c t s on a v o i d a n c e b e h a v i o u r of r o d e n t s (. se e r e v i e w by B i g n a m i / 1976 ). A v o i d a n c e f a c i l i t a t i o n a p p e a r e d even more c l e a r , when c h l o r d i a z e p o x i d e and o t h e r b e n z o d i a z e p i n e s w e r e c ombine d with o t h e r centrall-y--actin_.q d r u g s , s u c h as a m p h e t a m i n e ( S a n s o n e , 1975 a, c ) a n d o t h e r a d r e n e r g i c stimulants-'(--Sansone, 1975 b ), i m i p r a m i n e ( S a n s o n e ,
19~77 b ), MAO - i n h i b i t o r s ( S a n s o n e , i 9 7 7 c ) and m o r p h i n e ( S a n s o n e and Castellano, 1977 ). S i m i l a r l y to what o b s e r v e d by Rushton and S t e i n b e r g ( 1967 ), who t e s t e d c h l o r d i a z e p o x i d e - a m p h e t a m i n e c o m b i n a t i o n s on the e x p l o r a t o r y a c t i v i t y of r a t s , t h e s e d r u g c o m b i n a t i o n s p r o d u c e d s t i m u l a t i n g eff e c t s on a v o i d a n c e b e h a v i o u r of mice to a d e g r e e not often r e a c h e d when the d r u g s w e r e given s e p a r a t e l y . T h e s e r e s u l t s s u g g e s t f u r t h e r s t u d i e s on the
Pharmacological Research Communications, VoL'9, No. 9, 1977
880
e f f e c t i v e n e s s of s u c h d r u g c o m b i n a t i o n s . T h e u s e of r e s e r p i n i z e d a n i m a l s , c o u l d be s u i t a b l e at this p u r p o s e 9 s i n c e it has been o b s e r v e d ( S a n s o n e ~
1977 a ) that c h l o r d i a z e p o x i d e w a s a b l e to i n c r e a s e the s p o n t a n e o u s l o c o m o t o r a c t i v i t y and to i m p r o v e the a v o i d a n c e b e h a v i o u r of r e s e r p i n i z e d m i c e , thus s h o w i n g e f f i c a c y in a t e s t u s u a l l y e m p l o y e d in the s t u d y of a n t i d e p r e s sant drugs
( S u l s e r and B a s s , 1968; van R o s s u m , 1970 ).
In the p r e s e n t r e s e a r c h
r e s e r p i n i z e d mice h a v e been u s e d to t e s t the
e f f e c t i v e n e s s of c o m b i n a t i o n s of c h l o r d i a z e p o x i d e a n d a m p h e t a m i n e . T h e a p p r o a c h a d o p t e d has been to a d m i n i s t e r the two d r u g s , e i t h e r s e p a r a t e l y o r c o m b i n e d , a f t e r r e s e r p i n e p r e t r e a t m e n t 9 to mice t r a i n e d in the s h u t t l e * box.
METHODS T h e s u b j e c t s w e r e male mice ( 2 4 - 2 8 g ) b e l o n g i n g to the i n b r e d strain
(Charles
River).
BALB/c
T h e a p p a r a t u s was the s a m e p r e v i o u s l y d e s c r i b e d
f o r a v o i d a n c e t r a i n i n g of mice ( B o v e t et a l . ,
1969 ). E i g h t a u t o m a t e d s h u t t l e -
- b o x e s w e r e u s e d , e a c h d i v i d e d into two 2 O x l O cm c o m p a r t m e n t s , c o n n e c t e d
by a 3x3 cm o p e n i n g . A l i g h t ( 1 0 W ) was s w i t c h e d on a l t e r n a t e l y in the two compartments and used
as a conditioned stimulus ( C S ) .
The CSpreceded
the o n s e t of the u n c o n d i t i o n e d s t i m u l u s ( US ) by 5 s e c and o v e r l a p p e d it f o r 25 s e e . T h e US was an
e l e c t r i c s h o c k a p p l i e d c o n t i n u o u s l y to the g r i d f l o o r
through a selenium rectifier ( 110 V delivered through a 500.000
ohms r e -
s i s t a n c e ). T h e m t e r t i ~ i a l i n t e r v a l was 3 0 s e c . An a v o i d a n c e r e s p o n s e w a s r e c o r d e d when the a n i m a l a v o i d e d the US by r u n n i n g m t o the d a r k c o m p a r t ment w i t h i n 5 s e e a f t e r the o n s e t of the C S . If a n i m a l s f a i l e d to a v o i d tile s h o c k t h e y c o u l d e s c a p e it by c r o s s i n g d u r i n g the U S . I n t e r t r i a l c r o s s i n g s w e r e p u n i s h e d with s h o c k . M i c e t r a i n e d with d a i l y 1 O O - t r i a l s e s s i o n s and s h o w i n g a l e v e l of a v o i d a n c e r e s p o n s e s o f at l e a s t 7 0 % w e r e s u b j e c t e d to an a v o i d a n c e s e s s i o n a f t e r d i f f e r e n t d r u g t r e a t m e n t s . T h e e f f e c t s of the d r u g s w e r e c a l c u l a t e d by c o m p a r i n g the p e r f o r m a n c e of e a c h mouse d u r i n g the " d r u g s e s s i o n " to its p e r f o r m a n c e d u r i n g the p r e v i o u s n o n d r u g s e s s i o n ( c o n t r o l s e s s i o n ).
Pharmacological Research Communications, Vol. 9, No. 9, 1977
881
All mice r e c e i v e d r e s e r p i n e ( S e r p a s i l - C i b a ampullae diluted with distilled w a t e r ) at the dose of 1.5 m g / k g / i . p . , 2/4 hrs before the drug s e s s i o n . The r e s e r p i n i z e d mice were
a11ocated to the following treatment groups : saline
( I O ml/kg ), chlordiazepoxide hydrochloride 2.5, 5 and iO mg/kg; d-l-amphetamine sulfate i and 2 mg/kg; mixtures of chlordiazepoxide hydrochloride and amphetamine sulfate ( 2.5 + I ) and ( 5 + I ) mg/kg. These drugs were administered intraperitoneally 15 rain before the avoidance session. All groups included eight subjects. Reserpine was administered at the dose of 1.5 mg/kg, since in a preliminarly experiment avoidance depression produced by the dose of I mg/kg was completely antagonized by chlordiazepoxide alone ( 2.5 and 5 mg/kg ). The dosage of the mixtures was chosen on the basis of the effects prodttced by the single drugs.
RESULTS Avoidance r e s p o n s e s exhibited by the different experimental g r o u p s , 2£ hrs after reserpine ( i .5 mg/kg ), have been reported in Fig. i as per cent of the avoidance responses exhibited" by the same mice in the previous nondrug session. An analysis of variance concerning these data gave significant differences between groups ( F :12.69; d.f. 7/56 ; P ~ O. O O 1 ). All the values of F obtained in the individual between-groups comparisons reported below were tested for their significance on tl~e basis of 1/56 d.f. Saline - Reserpine, given at the dose of 1.5 mg/kg, produced a strong avoidance depression in control mice. A,Joidance responses of the saline group, 2~ hrs after reserpine, were at a level of 9.22 % in comparison with the performances evident in the previous nondrug session. This group showed a 8.75% e s c a p e f a i l u r e . Chlordiazepoxide alone - At the dose o f ' 2 . 5 mg/kg, chlordiazepoxide sign/fic~antly i n c r e a s e d the number of avoidance r e s p o n s e s of r e s e r p i n i z e d mice, in comparison with saline ( F : 1 2 . 5 8 ; P < O . O O I ). The dose of 5 mg/kg p r o duced a slight and not significant avoidance improvement, while the dose of IO mg/kg impaired r e m a r k a b l y the p e r f o r m a n c e of r e s e r p i n i z e d inice as also proved by the mean level of e s c a p e f a i l u r e s ( 65.75% ).
Pharmacologica/ Research Communications, goL 9, No. 9, 1977
882
Amp..hetamine alone - The l o w e r dose of the drug ( 1 mg/kg ) p r o d u c e d a slight and not significant i n c r e a s e of the a v o i d a n c e r e s p o n s e s . A s i g n i f i c a n t avoidance improvement, in c o m p a r i s o n with saline ( F :12.~,2; P<~O.C)O1 ), followed the a d m i n i s t r a t i o n of 2 mg]kg of amphetamine.
P RE TR EATME RESERPINE U) tu Z
A
D
Z
_oo
> (L uJ CO ., ¢Z ¢L it'- U.
O
O P"
Z
Z
O O
;>
1.5 m g / K g
100 ~r~
O
uJ
NT
m a.
CO CO uJ ¢n
(3
50"
ee
¢1 Z 0 Z
o
J
¢7 S
2.5
s CDP
w0
1
2
AMPH
(2.5" I } ( 5 ' I )
mg/Kg
C D P*A M PH
F i g . 1 - Effects of s a l i n e ( S ) and d i f f e r e n t d o s e s of c h l o r d i a z e p o x i d e h y d r o c h l o r i d e ( CDP ) and amphetamine sulfate ( AMPH ), given a l o ne o r in combination, on the avoidance p e r f o r m a n c e of t r a i n e d mice, p r e t r e a t e d ~/,ith ~,eserpine 2Z h r s before the drug s e s s i o n ,
C h l o r d i a z e p o x i d e g a m p h e t a m i n e combinations - When the two d o s e s of c h l o r d i a z e p o x i d e , 2~5 and 5 'mg/kg, w e r e given in combination with the l o w e r dose ( 1 mg/kg ) 6f amphetamine, s i g n i f i c a n t a v o i d a n c e improvements r e s u l t e d not \
only in c o m p a r i s 6 n with saline ( F :fiB.83 and 2fi.f5 r e s p e c t i v e l y ; P(~ O.OO1 ), \
bu~ a l s o in rela~tion to amphetamine ~lone ( F : 2 5 . 1 6 ,
P ~ O . O O 1 and F : 9 . 7 2 ,
P~' O.O1, respectively )and to the corresponding doses of chlordiazepoxide, k given alone ( F:10.66; ~P(~ O.O1 and F:I~.20, P < O.OO1., respectively ). Advantages from the combination of the two drugs were especially obtained when amphetami~ne was Combined with the lower dose ( 2.5 mg/kg ) of chlordiazepoxide. In this case avoidance performance of reserpinized mice was even higher of that exhlbited by the same mice in the control session.
Pharmaco/ogica/ Research Communications, VoL 9, No, 9, 1977
883
DISCUSSION The p r e s e n t findings indicate that both chlordiazepoxide and amphetamine,given alone, a r e able to improve the avoidance behaviour of r e s e r p i nized mice. However a s t r o n g e r antagonistic effect towards the avoidance d e p r e s s i o n produced by r e s e r p i n e can be obtained by using combinations of the two d r u g s . Improvement of avoidance behaviour of r e s e r p i n i z e d mice by c h l o r d i a zepoxide has been a l r e a d y o b s e r v e d in a p r e v i o u s r e s e a r c h , c a r r i e d out with mice of the S E C / 1 R e s t r a i n . In that r e s e a r c h chlordiazepoxide improved avoidance responding of r e s e r p i n i z e d mice at doses higher of those effec= t i r e , in the p r e s e n t experiment, in B A L B / c mice p r e t r e a t e d with the same dose of r e s e r p i n e ( 1.5 mg/kg ). Such d i s c r e p a n c y can be a s c r i b e d to a diff e r e n t s e n s i t i v i t y of the two s t r a i n s of mice to d r u g ' s e f f e c t s . A d i f f e r e n t s e n s i t i v i t y to the effects of chlo~'diazepoxide on avoidance responding has been a l r e a d y o b s e / v e d in the above tWO mouse s t r a i n s ( Sansone and M e s s e r i , 1974 ). As c o n c e r n s amphetamine, its ability to antagonize the disrupting effect of r e s e r p i n e on avoidance behaviour has been well documentated ( Rech, 1964; Hanson, 1 9 6 T ) . However, in the p r e s e n t r e s e a r c h , the r e s u l t s of g r e a t e r i n t e r e s t have been obtained with the use of drug combinations. Chlordiazepoxide and amphetamine, when given combined, s t r o n g l y enhanced avoidance responding of r e s e r p i n i z e d mice, s i m i l a r l y to what happened in norma'l animals ( S a n sane, 1975 a, c ). The role played by brain catecholamines in the maintenance of conditioned avoidance behaviour ( Selden et a l . ,
1973 ), the r e s u l t s p r e v i o u s l y
obtained by combining chlordiazepoxide with a d r e n e r g i c stimulants ( Sans a n e , 1975 b ) and the antagonistic effects towards the reserpine-indaced avoidance d e p r e s s i o n now d e m o n s t r a t e d , allow to suppose that a d r e n e r g i c mechanisms may be involved in the facilitating effects e x e r t e d by combina tions of chlordiazepoxide and amphetamine on avoidance b e h a v i o u r . Howe-
Pharmacological Research Communications, Vol. 9~ No. 9, 1977
884 ver further researches
a r e n e e d e d to e x p l a i n s u c h s t i m u l a t i n g e f f e c t s a n d
t h o s e o b s e r v e d when b e n z o d i a z e p i n e d e r i v a t i v e s a r e c o m b i n e d with o t h e r centrally acting drugs.
Acknowledgments
- T h e author' w i ~
to t h a n k M a r i o B a t t a g l i a f o r hi,~
skilful t e c h n i c a l a s s i s t a n c e .
REFERENCES Bignami, G. : Ann. Rev. Pharmacol. 16, 329, 1976
Bovet, D., Bovet-Nitti, g . , Oliverio, A. : Science !63, 139, 1969 Hanson, L . C . F . : Psychopharmacologia i__OO,289, 1967 Rech, R.H. : J. Pharmacol. exp. The,-. 146, 369, 1964 Rushton, R., Steinberg, H. : Neuropsychopharmacology,
H. Brill, P.
Deniker, H.Hippius and P.B. Bradley, eds. p. 464, Amsterdam: Excerpta Medica Foundation,
1967
S a n s o n e , M . : P s y c h o p h a r m a c o l o g i a 41, 117, 1975 a
Sansone, M . : Arch. int. Pharmacodyn. 215, 190, 1975 b Sansone, M. : Arch. int. Pharmacodyn. 218, 125, 1975 c Sansone, M. : Psychopharmacology,
submitted for publication, 1977 a
Sansone, M . : Psychopharmacology, submitted for publication, 1977 b Sansone, M.: Pharmacol. Res. Comm., in press, 1977 c Sansone, M., CastelIano, C.: Arch. int. Pharmacodyn., submitted for publication, 1977 Sansone, M., Messeri, P.: Pharmacol. Res. C o m m . 6, 179, 1974 Seiden, L.S., Brown, R . M . Lewy, A.]. : Chemical Modulation of Brain Function,.H.C. Sabelli, ed., N e w Yc <: Raven Press, 1973 Sulser, F., Bass, A.D. : Psychopharmacology - A review of Progress 1957-1967, D.H. Efron, ed., Public Health Service Publication, No. 1836,p. 1065, Washington, 1968 van Rossum, J . M . : Intern. Rev. N e u r o b i o l . 1 2 ,
307, 1 9 7 0
9, No. 9, 1977
879
E F F E C T S OF C H L O R D I A Z E P O X I D E , AMPHETAMI NE AND THEIR COM/3INATIONS ON AVOIDANCE BEHAVIOUR OF R E S E R P I N I Z E D MICE M a r i o Sansone Laboratorio di Pstcobiologia e Psicofarmacologia, C.N.R. via Reno n. I - 0 0 1 9 8 R o m e , Italy
Receivedin final form 23 August 1977
SUMMARY C h l o r d i a z e p o x i d e and a m p h e t a m i n e , give n a lone or in c o m b i n a u o n , have been t e s t e d in s h u t l l e - b q x t r a i n e d m i c e , 24 h r s a f t e r a p r e t r e a t m e n t with r e s e r p i n e ( 1 .5 mg/kg ). When give n a l o n e , both c h o l o r d i a z e p o x i d e and a m p h e t a mine i m p r o v e d a v o i d a n c e b e h a v i o u r of r e s e r p i n i z e d m i c e . H o w e v e r , the a n t a g o n i s t i c effect t o w a r d s the a v o i d a n c e d e p r e s s i o n p r o d u c e d by r e s e r p i n e was much m o r e m a r k e d when the two d r u g s w e r e c o m b i n e d .
INTRODUCTION
It h a s been often o b s e r v e d that c h l o r d i a z e p o x i d e , when given a lone at low d o s e s , e x e r t e d s t i m u l a t i n g e f f e c t s on a v o i d a n c e b e h a v i o u r of r o d e n t s (. se e r e v i e w by B i g n a m i / 1976 ). A v o i d a n c e f a c i l i t a t i o n a p p e a r e d even more c l e a r , when c h l o r d i a z e p o x i d e and o t h e r b e n z o d i a z e p i n e s w e r e c ombine d with o t h e r centrall-y--actin_.q d r u g s , s u c h as a m p h e t a m i n e ( S a n s o n e , 1975 a, c ) a n d o t h e r a d r e n e r g i c stimulants-'(--Sansone, 1975 b ), i m i p r a m i n e ( S a n s o n e ,
19~77 b ), MAO - i n h i b i t o r s ( S a n s o n e , i 9 7 7 c ) and m o r p h i n e ( S a n s o n e and Castellano, 1977 ). S i m i l a r l y to what o b s e r v e d by Rushton and S t e i n b e r g ( 1967 ), who t e s t e d c h l o r d i a z e p o x i d e - a m p h e t a m i n e c o m b i n a t i o n s on the e x p l o r a t o r y a c t i v i t y of r a t s , t h e s e d r u g c o m b i n a t i o n s p r o d u c e d s t i m u l a t i n g eff e c t s on a v o i d a n c e b e h a v i o u r of mice to a d e g r e e not often r e a c h e d when the d r u g s w e r e given s e p a r a t e l y . T h e s e r e s u l t s s u g g e s t f u r t h e r s t u d i e s on the
Pharmacological Research Communications, VoL'9, No. 9, 1977
880
e f f e c t i v e n e s s of s u c h d r u g c o m b i n a t i o n s . T h e u s e of r e s e r p i n i z e d a n i m a l s , c o u l d be s u i t a b l e at this p u r p o s e 9 s i n c e it has been o b s e r v e d ( S a n s o n e ~
1977 a ) that c h l o r d i a z e p o x i d e w a s a b l e to i n c r e a s e the s p o n t a n e o u s l o c o m o t o r a c t i v i t y and to i m p r o v e the a v o i d a n c e b e h a v i o u r of r e s e r p i n i z e d m i c e , thus s h o w i n g e f f i c a c y in a t e s t u s u a l l y e m p l o y e d in the s t u d y of a n t i d e p r e s sant drugs
( S u l s e r and B a s s , 1968; van R o s s u m , 1970 ).
In the p r e s e n t r e s e a r c h
r e s e r p i n i z e d mice h a v e been u s e d to t e s t the
e f f e c t i v e n e s s of c o m b i n a t i o n s of c h l o r d i a z e p o x i d e a n d a m p h e t a m i n e . T h e a p p r o a c h a d o p t e d has been to a d m i n i s t e r the two d r u g s , e i t h e r s e p a r a t e l y o r c o m b i n e d , a f t e r r e s e r p i n e p r e t r e a t m e n t 9 to mice t r a i n e d in the s h u t t l e * box.
METHODS T h e s u b j e c t s w e r e male mice ( 2 4 - 2 8 g ) b e l o n g i n g to the i n b r e d strain
(Charles
River).
BALB/c
T h e a p p a r a t u s was the s a m e p r e v i o u s l y d e s c r i b e d
f o r a v o i d a n c e t r a i n i n g of mice ( B o v e t et a l . ,
1969 ). E i g h t a u t o m a t e d s h u t t l e -
- b o x e s w e r e u s e d , e a c h d i v i d e d into two 2 O x l O cm c o m p a r t m e n t s , c o n n e c t e d
by a 3x3 cm o p e n i n g . A l i g h t ( 1 0 W ) was s w i t c h e d on a l t e r n a t e l y in the two compartments and used
as a conditioned stimulus ( C S ) .
The CSpreceded
the o n s e t of the u n c o n d i t i o n e d s t i m u l u s ( US ) by 5 s e c and o v e r l a p p e d it f o r 25 s e e . T h e US was an
e l e c t r i c s h o c k a p p l i e d c o n t i n u o u s l y to the g r i d f l o o r
through a selenium rectifier ( 110 V delivered through a 500.000
ohms r e -
s i s t a n c e ). T h e m t e r t i ~ i a l i n t e r v a l was 3 0 s e c . An a v o i d a n c e r e s p o n s e w a s r e c o r d e d when the a n i m a l a v o i d e d the US by r u n n i n g m t o the d a r k c o m p a r t ment w i t h i n 5 s e e a f t e r the o n s e t of the C S . If a n i m a l s f a i l e d to a v o i d tile s h o c k t h e y c o u l d e s c a p e it by c r o s s i n g d u r i n g the U S . I n t e r t r i a l c r o s s i n g s w e r e p u n i s h e d with s h o c k . M i c e t r a i n e d with d a i l y 1 O O - t r i a l s e s s i o n s and s h o w i n g a l e v e l of a v o i d a n c e r e s p o n s e s o f at l e a s t 7 0 % w e r e s u b j e c t e d to an a v o i d a n c e s e s s i o n a f t e r d i f f e r e n t d r u g t r e a t m e n t s . T h e e f f e c t s of the d r u g s w e r e c a l c u l a t e d by c o m p a r i n g the p e r f o r m a n c e of e a c h mouse d u r i n g the " d r u g s e s s i o n " to its p e r f o r m a n c e d u r i n g the p r e v i o u s n o n d r u g s e s s i o n ( c o n t r o l s e s s i o n ).
Pharmacological Research Communications, Vol. 9, No. 9, 1977
881
All mice r e c e i v e d r e s e r p i n e ( S e r p a s i l - C i b a ampullae diluted with distilled w a t e r ) at the dose of 1.5 m g / k g / i . p . , 2/4 hrs before the drug s e s s i o n . The r e s e r p i n i z e d mice were
a11ocated to the following treatment groups : saline
( I O ml/kg ), chlordiazepoxide hydrochloride 2.5, 5 and iO mg/kg; d-l-amphetamine sulfate i and 2 mg/kg; mixtures of chlordiazepoxide hydrochloride and amphetamine sulfate ( 2.5 + I ) and ( 5 + I ) mg/kg. These drugs were administered intraperitoneally 15 rain before the avoidance session. All groups included eight subjects. Reserpine was administered at the dose of 1.5 mg/kg, since in a preliminarly experiment avoidance depression produced by the dose of I mg/kg was completely antagonized by chlordiazepoxide alone ( 2.5 and 5 mg/kg ). The dosage of the mixtures was chosen on the basis of the effects prodttced by the single drugs.
RESULTS Avoidance r e s p o n s e s exhibited by the different experimental g r o u p s , 2£ hrs after reserpine ( i .5 mg/kg ), have been reported in Fig. i as per cent of the avoidance responses exhibited" by the same mice in the previous nondrug session. An analysis of variance concerning these data gave significant differences between groups ( F :12.69; d.f. 7/56 ; P ~ O. O O 1 ). All the values of F obtained in the individual between-groups comparisons reported below were tested for their significance on tl~e basis of 1/56 d.f. Saline - Reserpine, given at the dose of 1.5 mg/kg, produced a strong avoidance depression in control mice. A,Joidance responses of the saline group, 2~ hrs after reserpine, were at a level of 9.22 % in comparison with the performances evident in the previous nondrug session. This group showed a 8.75% e s c a p e f a i l u r e . Chlordiazepoxide alone - At the dose o f ' 2 . 5 mg/kg, chlordiazepoxide sign/fic~antly i n c r e a s e d the number of avoidance r e s p o n s e s of r e s e r p i n i z e d mice, in comparison with saline ( F : 1 2 . 5 8 ; P < O . O O I ). The dose of 5 mg/kg p r o duced a slight and not significant avoidance improvement, while the dose of IO mg/kg impaired r e m a r k a b l y the p e r f o r m a n c e of r e s e r p i n i z e d inice as also proved by the mean level of e s c a p e f a i l u r e s ( 65.75% ).
Pharmacologica/ Research Communications, goL 9, No. 9, 1977
882
Amp..hetamine alone - The l o w e r dose of the drug ( 1 mg/kg ) p r o d u c e d a slight and not significant i n c r e a s e of the a v o i d a n c e r e s p o n s e s . A s i g n i f i c a n t avoidance improvement, in c o m p a r i s o n with saline ( F :12.~,2; P<~O.C)O1 ), followed the a d m i n i s t r a t i o n of 2 mg]kg of amphetamine.
P RE TR EATME RESERPINE U) tu Z
A
D
Z
_oo
> (L uJ CO ., ¢Z ¢L it'- U.
O
O P"
Z
Z
O O
;>
1.5 m g / K g
100 ~r~
O
uJ
NT
m a.
CO CO uJ ¢n
(3
50"
ee
¢1 Z 0 Z
o
J
¢7 S
2.5
s CDP
w0
1
2
AMPH
(2.5" I } ( 5 ' I )
mg/Kg
C D P*A M PH
F i g . 1 - Effects of s a l i n e ( S ) and d i f f e r e n t d o s e s of c h l o r d i a z e p o x i d e h y d r o c h l o r i d e ( CDP ) and amphetamine sulfate ( AMPH ), given a l o ne o r in combination, on the avoidance p e r f o r m a n c e of t r a i n e d mice, p r e t r e a t e d ~/,ith ~,eserpine 2Z h r s before the drug s e s s i o n ,
C h l o r d i a z e p o x i d e g a m p h e t a m i n e combinations - When the two d o s e s of c h l o r d i a z e p o x i d e , 2~5 and 5 'mg/kg, w e r e given in combination with the l o w e r dose ( 1 mg/kg ) 6f amphetamine, s i g n i f i c a n t a v o i d a n c e improvements r e s u l t e d not \
only in c o m p a r i s 6 n with saline ( F :fiB.83 and 2fi.f5 r e s p e c t i v e l y ; P(~ O.OO1 ), \
bu~ a l s o in rela~tion to amphetamine ~lone ( F : 2 5 . 1 6 ,
P ~ O . O O 1 and F : 9 . 7 2 ,
P~' O.O1, respectively )and to the corresponding doses of chlordiazepoxide, k given alone ( F:10.66; ~P(~ O.O1 and F:I~.20, P < O.OO1., respectively ). Advantages from the combination of the two drugs were especially obtained when amphetami~ne was Combined with the lower dose ( 2.5 mg/kg ) of chlordiazepoxide. In this case avoidance performance of reserpinized mice was even higher of that exhlbited by the same mice in the control session.
Pharmaco/ogica/ Research Communications, VoL 9, No, 9, 1977
883
DISCUSSION The p r e s e n t findings indicate that both chlordiazepoxide and amphetamine,given alone, a r e able to improve the avoidance behaviour of r e s e r p i nized mice. However a s t r o n g e r antagonistic effect towards the avoidance d e p r e s s i o n produced by r e s e r p i n e can be obtained by using combinations of the two d r u g s . Improvement of avoidance behaviour of r e s e r p i n i z e d mice by c h l o r d i a zepoxide has been a l r e a d y o b s e r v e d in a p r e v i o u s r e s e a r c h , c a r r i e d out with mice of the S E C / 1 R e s t r a i n . In that r e s e a r c h chlordiazepoxide improved avoidance responding of r e s e r p i n i z e d mice at doses higher of those effec= t i r e , in the p r e s e n t experiment, in B A L B / c mice p r e t r e a t e d with the same dose of r e s e r p i n e ( 1.5 mg/kg ). Such d i s c r e p a n c y can be a s c r i b e d to a diff e r e n t s e n s i t i v i t y of the two s t r a i n s of mice to d r u g ' s e f f e c t s . A d i f f e r e n t s e n s i t i v i t y to the effects of chlo~'diazepoxide on avoidance responding has been a l r e a d y o b s e / v e d in the above tWO mouse s t r a i n s ( Sansone and M e s s e r i , 1974 ). As c o n c e r n s amphetamine, its ability to antagonize the disrupting effect of r e s e r p i n e on avoidance behaviour has been well documentated ( Rech, 1964; Hanson, 1 9 6 T ) . However, in the p r e s e n t r e s e a r c h , the r e s u l t s of g r e a t e r i n t e r e s t have been obtained with the use of drug combinations. Chlordiazepoxide and amphetamine, when given combined, s t r o n g l y enhanced avoidance responding of r e s e r p i n i z e d mice, s i m i l a r l y to what happened in norma'l animals ( S a n sane, 1975 a, c ). The role played by brain catecholamines in the maintenance of conditioned avoidance behaviour ( Selden et a l . ,
1973 ), the r e s u l t s p r e v i o u s l y
obtained by combining chlordiazepoxide with a d r e n e r g i c stimulants ( Sans a n e , 1975 b ) and the antagonistic effects towards the reserpine-indaced avoidance d e p r e s s i o n now d e m o n s t r a t e d , allow to suppose that a d r e n e r g i c mechanisms may be involved in the facilitating effects e x e r t e d by combina tions of chlordiazepoxide and amphetamine on avoidance b e h a v i o u r . Howe-
Pharmacological Research Communications, Vol. 9~ No. 9, 1977
884 ver further researches
a r e n e e d e d to e x p l a i n s u c h s t i m u l a t i n g e f f e c t s a n d
t h o s e o b s e r v e d when b e n z o d i a z e p i n e d e r i v a t i v e s a r e c o m b i n e d with o t h e r centrally acting drugs.
Acknowledgments
- T h e author' w i ~
to t h a n k M a r i o B a t t a g l i a f o r hi,~
skilful t e c h n i c a l a s s i s t a n c e .
REFERENCES Bignami, G. : Ann. Rev. Pharmacol. 16, 329, 1976
Bovet, D., Bovet-Nitti, g . , Oliverio, A. : Science !63, 139, 1969 Hanson, L . C . F . : Psychopharmacologia i__OO,289, 1967 Rech, R.H. : J. Pharmacol. exp. The,-. 146, 369, 1964 Rushton, R., Steinberg, H. : Neuropsychopharmacology,
H. Brill, P.
Deniker, H.Hippius and P.B. Bradley, eds. p. 464, Amsterdam: Excerpta Medica Foundation,
1967
S a n s o n e , M . : P s y c h o p h a r m a c o l o g i a 41, 117, 1975 a
Sansone, M . : Arch. int. Pharmacodyn. 215, 190, 1975 b Sansone, M. : Arch. int. Pharmacodyn. 218, 125, 1975 c Sansone, M. : Psychopharmacology,
submitted for publication, 1977 a
Sansone, M . : Psychopharmacology, submitted for publication, 1977 b Sansone, M.: Pharmacol. Res. Comm., in press, 1977 c Sansone, M., CastelIano, C.: Arch. int. Pharmacodyn., submitted for publication, 1977 Sansone, M., Messeri, P.: Pharmacol. Res. C o m m . 6, 179, 1974 Seiden, L.S., Brown, R . M . Lewy, A.]. : Chemical Modulation of Brain Function,.H.C. Sabelli, ed., N e w Yc <: Raven Press, 1973 Sulser, F., Bass, A.D. : Psychopharmacology - A review of Progress 1957-1967, D.H. Efron, ed., Public Health Service Publication, No. 1836,p. 1065, Washington, 1968 van Rossum, J . M . : Intern. Rev. N e u r o b i o l . 1 2 ,
307, 1 9 7 0