Life Sciences, Vol. Printed in the USA
THE
EFFECT
51, pp.
1535-1544
OF A M P E R O Z I D E ° CORTICOSTERONE Agneta
Kabi
Pergamon
A N E W A N T I P S Y C H O T I C DRUG, C O N C E N T R A T I O N IN THE RAT
Albinsson
and G u n n a r
Press
ON P L A S M A
Andersson*
P h a r m a c i a AB, D e p a r t m e n t of C N S - R e s e a r c h , Box 839, S-201 80 Malm6, S w e d e n (Received
in final
form September
8, 1992)
Summary A m p e r o z i d e is an a t y p i c a l a n t i p s y c h o t i c d r u g w i t h high a f f i n i t y for the s e r o t o n i n 5-HT 2 r e c e p t o r but w i t h low a f f i n i t y for the d o p a m i n e D I and D 2 receptors. A m p e r o z i d e dose-dependently i n c r e a s e d the level of p l a s m a c o r t i c o s t e r o n e in the rat. The effect of a m p e r o z i d e on p l a s m a c o r t i c o s t e r o n e was not i n h i b i t e d by p r e t r e a t m e n t w i t h the 5-HTIA r e c e p t o r a n t a g o n i s t p i n d o l o l or the 5-HT 2 r e c e p t o r a n t a g o n i s t ritanserin. Nor was it i n h i b i t e d by the dopa m i n e D 2 r e c e p t o r a n t a g o n i s t h a l o p e r i d o l . In c o n t r a s t to ritanserin, a m p e r o z i d e did not a n t a g o n i z e p l a s m a c o r t i c o sterone elevation e l i c i t e d by the s e r o t o n i n receptor a g o n i s t MK-212. S i m i l a r to the s e r o t o n i n u p t a k e i n h i b i t o r fluoxetine, a m p e r o z i d e (0.5 mg/kg) s i g n i f i c a n t l y (p10 pM), 5-HTIc (Ki=9.4 ~M) (13) and 5-HT 3 (Ki>10 M) ( S v a r t e n g r e n et al., u n p u b l i s h e d data) and is a m o d e r a t e i n h i b i t o r of in v i t r o s e r o t o n i n u p t a k e (13). B a s e d on c l i n i c a l o b s e r v a t i o n s (14,15) and pK i v a l u e s for d o p a m i n e and s e r o t o n i n b i n d i n g sites (16) a m p e r o z i d e has been c l a s s i f i e d as an a t y p i c a l a n t i p s y c h o t i c drug. In p r e c l i n i c a l s t u d i e s an a n t i p s y c h o t i c e f f e c t of a m p e r o z i d e is i n d i c a t e d from its p o t e n t i n h i b i t i o n of a m p h e t a m i n e i n d u c e d hypermotility in m i c e and b l o c k i n g of c o n d i t i o n e d a v o i d a n c e res p o n s e in rats (]7,18). F u r t h e r m o r e , a m p e r o z i d e has been found to selectively attenuate neurotransmission in the m e s o l i m b i c p a r t of the d o p a m i n e r g i c n e r v e s y s t e m (]9), s t i m u l a t e the T I D A n e u r o n act i v i t y (20) and to d e c r e a s e the p l a s m a c o n c e n t r a t i o n of p r o l a c t i n (2]). T h e s e e f f e c t s of a m p e r o z i d e on d o p a m i n e r g i c n e r v e a c t i v i t y c o m b i n e d w i t h its h i g h a f f i n i t y for s e r o t o n i n 5-HT 2 r e c e p t o r s s u g g e s t an i n f l u e n c e on n e u r o e n d o c r i n e p a r a m e t e r s s i m i l a r to that of a t y p i c a l antipsychotic drugs. The p r e s e n t experiments were designed to p o r t r a y the e f f e c t s of a m p e r o z i d e on p l a s m a c o r t i c o s t e r o n e c o n c e n t r a t i o n in the rat. Material
and m e t h o d s
Animals. Sprague-Dawley rats, p u r c h a s e d from M ~ l l e g a a r d B r e e d i n g C e n t r e Ltd., Denmark, w e r e u s e d in the study. A d u l t m a l e rats (180 g) w e r e u s e d in one e x p e r i m e n t w h e r e b r a i n 5-HT c o n t e n t was d e p l e t e d by the t r y p t o p h a n h y d r o x y l a s e i n h i b i t o r p a r a c h l o r o p h e n y l alanine (PCPA), 24 hrs b e f o r e t e s t i n g . A d u l t m a l e rats w e r e a l s o u s e d in the e x p e r i m e n t w i t h h y p o p h y s e c t o m i z e d rats. The adrenal f u n c t i o n of t h e s e rats was m a i n t a i n e d by d a i l y i n j e c t i o n s (2-3 t i m e s per day) of A C T H (20 IU/kg, s.c.) d u r i n g the last 3 days p r i o r to the e x p e r i m e n t . All the o t h e r s t u d i e s w e r e c a r r i e d out on 1 6 - d a y - o l d rat pups 'born at the d e p a r t m e n t . The p r e g n a n t rats w e r e m o n i t o r e d d a i l y and the day the l i t t e r was found was c o n s i d e r e d to be d a y 0. This m o d e l is r o u t i n e l y u s e d at our d e p a r t m e n t in s c r e e n ing of d r u g s w i t h p o t e n t i a l e f f e c t on p l a s m a c o r t i c o s t e r o n e c o n c e n tration. At this age the rat pups r e a c t to both c h e m i c a l and e n v i r o n m e n t a l s t r e s s o r s (22). The a n i m a l s w e r e h o u s e d in a t e m p e r a t u r e c o n t r o l l e d r o o m (20-21°C) u n d e r 12 h l i g h t / d a r k c y c l e b e g i n n i n g at 6:00 a.m. F o o d (RMI(E), S c i e n t i f i c D i e t s Service, W i t h a m , U.K.) and tap w a t e r w e r e a l l o w e d ad libitum. Drugs. Amperozide HCl ( s y n t h e s i z e d by Kabi P h a r m a c i a AB, Malm~. Sweden) and P C P A (Sigma C h e m i c a l Co) w e r e d i s s o l v e d in p h y s i o l o g i cal saline. H a l o p e r i d o l ( i n j e c t i o n s o l u t i o n , J a n s s e n P h a r m a c e u t i c a , Beerse, B e l g i u m ) was d i l u t e d w i t h p h y s i o l o g i c a l saline. Pindolol (Research Biochemical Inc., Natick, MA) r ritanserin (Janssen P h a r m a c e u t i c a , Beerse, B e l g i u m ) and f l u o x e t i n e (Eli L i l l y and Co, I n d i a n a p o l i s , IN) w e r e d i s s o l v e d in a m i n i m a l a m o u n t of l a c t i c a c i d a n d t h e n d i l u t e d w i t h d i s t i l l e d water. PCA (Sigma C h e m i c a l Co), 8 O H - D P A T ( R e s e a r c h B i o c h e m i c a l s Inc., Natick, MA), q u i p a z i n e (Miles L a b o r a t o r i e s , N a p e r v i l l e , IL) and M K - 2 1 2 (Merck S h a r p and Dohme, Rahway, NJ) w e r e all d i s s o l v e d in d i s t i l l e d water. The f o l l o w i n g drugs were kindly supplied by the manufacturer: ritanserin, f l u o x e t i n e and M K - 2 1 2 . The a c u t e e f f e c t on c o r t i c o s t e r o n e . On day 16 a f t e r pups w e r e injected s.c. w i t h the drugs. Each drug
d e l i v e r y rat was g i v e n in
Vol.
51, No. 19, 1992
Amperozide
and Plasma Corticosterone
1537
different concentrations, calculated so t h a t the i n j e c t e d v o l u m e was 0.1 ml per 32 g b o d y weight, and e a c h c o n c e n t r a t i o n was g i v e n to at l e a s t five rat pups. One h o u r a f t e r the d r u g a d m i n i s t r a t i o n c e r v i c a l d i s l o c a t i o n and t r u n k b l o o d was tubes.
the pups collected
Blood sample treatment. The b l o o d s a m p l e s w e r e 1800xg for 10 m i n u t e s and the p l a s m a was s t o r e d analysis.
w e r e k i l l e d by in h e p a r i n i z e d
centrifuged at at -20 C until
A n a l y s i s of c o r t i c o s t e r o n e . C o r t i c o s t e r o n e was m e a s u r e d in d u p l i c a t e by a r a d i o i m m u n o a s s a y u s i n g a n t i s e r u m from R a d i o a s s a y S y s t e m s L a b o r a t o r i e s , Carson, CA, USA, on u n e x t r a c t e d p l a s m a in w h i c h b i n d ing p r o t e i n s had been d e n a t u r a t e d by d i l u t i o n w i t h a p e p s i n solution (23,24). The i n t r a - a s s a y v a r i a n c e was
THE
EFFECT
51, pp.
1535-1544
OF A M P E R O Z I D E ° CORTICOSTERONE Agneta
Kabi
Pergamon
A N E W A N T I P S Y C H O T I C DRUG, C O N C E N T R A T I O N IN THE RAT
Albinsson
and G u n n a r
Press
ON P L A S M A
Andersson*
P h a r m a c i a AB, D e p a r t m e n t of C N S - R e s e a r c h , Box 839, S-201 80 Malm6, S w e d e n (Received
in final
form September
8, 1992)
Summary A m p e r o z i d e is an a t y p i c a l a n t i p s y c h o t i c d r u g w i t h high a f f i n i t y for the s e r o t o n i n 5-HT 2 r e c e p t o r but w i t h low a f f i n i t y for the d o p a m i n e D I and D 2 receptors. A m p e r o z i d e dose-dependently i n c r e a s e d the level of p l a s m a c o r t i c o s t e r o n e in the rat. The effect of a m p e r o z i d e on p l a s m a c o r t i c o s t e r o n e was not i n h i b i t e d by p r e t r e a t m e n t w i t h the 5-HTIA r e c e p t o r a n t a g o n i s t p i n d o l o l or the 5-HT 2 r e c e p t o r a n t a g o n i s t ritanserin. Nor was it i n h i b i t e d by the dopa m i n e D 2 r e c e p t o r a n t a g o n i s t h a l o p e r i d o l . In c o n t r a s t to ritanserin, a m p e r o z i d e did not a n t a g o n i z e p l a s m a c o r t i c o sterone elevation e l i c i t e d by the s e r o t o n i n receptor a g o n i s t MK-212. S i m i l a r to the s e r o t o n i n u p t a k e i n h i b i t o r fluoxetine, a m p e r o z i d e (0.5 mg/kg) s i g n i f i c a n t l y (p10 pM), 5-HTIc (Ki=9.4 ~M) (13) and 5-HT 3 (Ki>10 M) ( S v a r t e n g r e n et al., u n p u b l i s h e d data) and is a m o d e r a t e i n h i b i t o r of in v i t r o s e r o t o n i n u p t a k e (13). B a s e d on c l i n i c a l o b s e r v a t i o n s (14,15) and pK i v a l u e s for d o p a m i n e and s e r o t o n i n b i n d i n g sites (16) a m p e r o z i d e has been c l a s s i f i e d as an a t y p i c a l a n t i p s y c h o t i c drug. In p r e c l i n i c a l s t u d i e s an a n t i p s y c h o t i c e f f e c t of a m p e r o z i d e is i n d i c a t e d from its p o t e n t i n h i b i t i o n of a m p h e t a m i n e i n d u c e d hypermotility in m i c e and b l o c k i n g of c o n d i t i o n e d a v o i d a n c e res p o n s e in rats (]7,18). F u r t h e r m o r e , a m p e r o z i d e has been found to selectively attenuate neurotransmission in the m e s o l i m b i c p a r t of the d o p a m i n e r g i c n e r v e s y s t e m (]9), s t i m u l a t e the T I D A n e u r o n act i v i t y (20) and to d e c r e a s e the p l a s m a c o n c e n t r a t i o n of p r o l a c t i n (2]). T h e s e e f f e c t s of a m p e r o z i d e on d o p a m i n e r g i c n e r v e a c t i v i t y c o m b i n e d w i t h its h i g h a f f i n i t y for s e r o t o n i n 5-HT 2 r e c e p t o r s s u g g e s t an i n f l u e n c e on n e u r o e n d o c r i n e p a r a m e t e r s s i m i l a r to that of a t y p i c a l antipsychotic drugs. The p r e s e n t experiments were designed to p o r t r a y the e f f e c t s of a m p e r o z i d e on p l a s m a c o r t i c o s t e r o n e c o n c e n t r a t i o n in the rat. Material
and m e t h o d s
Animals. Sprague-Dawley rats, p u r c h a s e d from M ~ l l e g a a r d B r e e d i n g C e n t r e Ltd., Denmark, w e r e u s e d in the study. A d u l t m a l e rats (180 g) w e r e u s e d in one e x p e r i m e n t w h e r e b r a i n 5-HT c o n t e n t was d e p l e t e d by the t r y p t o p h a n h y d r o x y l a s e i n h i b i t o r p a r a c h l o r o p h e n y l alanine (PCPA), 24 hrs b e f o r e t e s t i n g . A d u l t m a l e rats w e r e a l s o u s e d in the e x p e r i m e n t w i t h h y p o p h y s e c t o m i z e d rats. The adrenal f u n c t i o n of t h e s e rats was m a i n t a i n e d by d a i l y i n j e c t i o n s (2-3 t i m e s per day) of A C T H (20 IU/kg, s.c.) d u r i n g the last 3 days p r i o r to the e x p e r i m e n t . All the o t h e r s t u d i e s w e r e c a r r i e d out on 1 6 - d a y - o l d rat pups 'born at the d e p a r t m e n t . The p r e g n a n t rats w e r e m o n i t o r e d d a i l y and the day the l i t t e r was found was c o n s i d e r e d to be d a y 0. This m o d e l is r o u t i n e l y u s e d at our d e p a r t m e n t in s c r e e n ing of d r u g s w i t h p o t e n t i a l e f f e c t on p l a s m a c o r t i c o s t e r o n e c o n c e n tration. At this age the rat pups r e a c t to both c h e m i c a l and e n v i r o n m e n t a l s t r e s s o r s (22). The a n i m a l s w e r e h o u s e d in a t e m p e r a t u r e c o n t r o l l e d r o o m (20-21°C) u n d e r 12 h l i g h t / d a r k c y c l e b e g i n n i n g at 6:00 a.m. F o o d (RMI(E), S c i e n t i f i c D i e t s Service, W i t h a m , U.K.) and tap w a t e r w e r e a l l o w e d ad libitum. Drugs. Amperozide HCl ( s y n t h e s i z e d by Kabi P h a r m a c i a AB, Malm~. Sweden) and P C P A (Sigma C h e m i c a l Co) w e r e d i s s o l v e d in p h y s i o l o g i cal saline. H a l o p e r i d o l ( i n j e c t i o n s o l u t i o n , J a n s s e n P h a r m a c e u t i c a , Beerse, B e l g i u m ) was d i l u t e d w i t h p h y s i o l o g i c a l saline. Pindolol (Research Biochemical Inc., Natick, MA) r ritanserin (Janssen P h a r m a c e u t i c a , Beerse, B e l g i u m ) and f l u o x e t i n e (Eli L i l l y and Co, I n d i a n a p o l i s , IN) w e r e d i s s o l v e d in a m i n i m a l a m o u n t of l a c t i c a c i d a n d t h e n d i l u t e d w i t h d i s t i l l e d water. PCA (Sigma C h e m i c a l Co), 8 O H - D P A T ( R e s e a r c h B i o c h e m i c a l s Inc., Natick, MA), q u i p a z i n e (Miles L a b o r a t o r i e s , N a p e r v i l l e , IL) and M K - 2 1 2 (Merck S h a r p and Dohme, Rahway, NJ) w e r e all d i s s o l v e d in d i s t i l l e d water. The f o l l o w i n g drugs were kindly supplied by the manufacturer: ritanserin, f l u o x e t i n e and M K - 2 1 2 . The a c u t e e f f e c t on c o r t i c o s t e r o n e . On day 16 a f t e r pups w e r e injected s.c. w i t h the drugs. Each drug
d e l i v e r y rat was g i v e n in
Vol.
51, No. 19, 1992
Amperozide
and Plasma Corticosterone
1537
different concentrations, calculated so t h a t the i n j e c t e d v o l u m e was 0.1 ml per 32 g b o d y weight, and e a c h c o n c e n t r a t i o n was g i v e n to at l e a s t five rat pups. One h o u r a f t e r the d r u g a d m i n i s t r a t i o n c e r v i c a l d i s l o c a t i o n and t r u n k b l o o d was tubes.
the pups collected
Blood sample treatment. The b l o o d s a m p l e s w e r e 1800xg for 10 m i n u t e s and the p l a s m a was s t o r e d analysis.
w e r e k i l l e d by in h e p a r i n i z e d
centrifuged at at -20 C until
A n a l y s i s of c o r t i c o s t e r o n e . C o r t i c o s t e r o n e was m e a s u r e d in d u p l i c a t e by a r a d i o i m m u n o a s s a y u s i n g a n t i s e r u m from R a d i o a s s a y S y s t e m s L a b o r a t o r i e s , Carson, CA, USA, on u n e x t r a c t e d p l a s m a in w h i c h b i n d ing p r o t e i n s had been d e n a t u r a t e d by d i l u t i o n w i t h a p e p s i n solution (23,24). The i n t r a - a s s a y v a r i a n c e was
