- Email: [email protected]
Controlled demonstration of metrazol kindling
Pharmacology Biochemistry & Behavior, Vol. 6, pp. 599-600. Copyright © 1977 by ANKHO International Inc. All rights of reproduction in any form reserved. Printed in the U.S.A.
BRIEF COMMUNICATION Controlled Demonstration of Metrazol Kindling J O H N P. J. P I N E L A N D K. F. C H E U N G
Department o f Psychology, University o f British Columbia, Vancouver, British Columbia, V6T 1 W5 (Received 7 J a n u a r y 1977) PINEL, J. P. J. AND K. F. CHEUNG. Controlled demonstration of metrazol kindling. PHARMAC. BIOCHEM. BEHAV. 6(5) 5 9 9 - 6 0 0 , 1977. - Convulsive responses elicited in rats by intraperitoneal injections of metrazol were intensified following a series of metrazol injections administered once every three days. However, neither handling alone nor placebo injections increased the susceptibility of control rats to metrazol-induced convulsive symptoms. Thus, although increases in the susceptibility to metrazol seizures following periodic placebo injections have been reported by others, such increases do not appear to be a critical factor in metrazol kindling. Seizures
Rats
Metrazol
Kindling
P E R I O D I C i n j e c t i o n s of m e t r a z o l have b e e n s h o w n to p r o d u c e a progressive increase in seizure s u s c e p t i b i l i t y (kindling). F o r e x a m p l e , Mason a n d C o o p e r [2] f o u n d t h a t a dose o f m e t r a z o l w h i c h was initially s u b c o n v u l s i v e w o u l d e v e n t u a l l y elicit grand real seizures in rats i n j e c t e d w i t h it once every t h r e e days. However, r e c e n t l y I z q u i e r d o , Fern a n d e z , Oliveira, a n d S e t t i n e r i [1] r e p o r t e d t h a t increases in t h e s u s c e p t i b i l i t y to m e t r a z o l seizures f o l l o w e d a series of daily p l a c e b o injections. Since p l a c e b o c o n t r o l s were i n c l u d e d in n e i t h e r t h e M a s o n and C o o p e r s t u d y n o r in its r e p l i c a t i o n b y Pinel and V a n O o t [ 3 ] , the m e t r a z o l k i n d l i n g in these studies m a y have b e e n an a r t i f a c t of the i n j e c t i o n p r o c e d u r e r a t h e r t h a n being a c o n s e q u e n c e of t h e m e t r a z o l per se.
the p l a c e b o c o n t r o l g r o u p were injected at 3-day intervals with an e q u a l dose of i s o t o n i c saline; and rats in the second c o n t r o l g r o u p were h a n d l e d similarly b u t n o t injected. This t r e a t m e n t r e g i m e n t c o n t i n u e d until Day 127 w h e n all animals again received the s t a n d a r d 20 m g / k g i n j e c t i o n of metrazol. R e s p o n s e s to this p o s t t r e a t m e n t m e t r a z o l inject i o n were assessed b y an e x p e r i m e n t e r u n a w a r e of each animal's experimental history. RESULTS AND DISCUSSION The r e s p o n s e s of the animals in each of the t h r e e groups to t h e p r e t r e a t m e n t and p o s t t r e a t m e n t m e t r a z o l injections are s u m m a r i z e d in Fig. I. It is readily a p p a r e n t t h a t the i n c i d e n c e of each of the f o u r epileptic s y m p t o m s was increased a p p r e c i a b l y by the i n t e r v e n i n g series of m e t r a z o l i n j e c t i o n s b u t n o t b y e i t h e r of the t w o c o n t r o l p r o c e d u r e s ( p > 0 . 1 ) . Thus, a l t h o u g h animals were assigned to groups on the basis of t h e i r initial r e s p o n s e to m e t r a z o l , p l a n n e d c o m p a r i s o n s i n d i c a t e d t h a t b y t h e end of the e x p e r i m e n t each of the four s y m p t o m s was significantly m o r e prevalent in t h e e x p e r i m e n t a l group t h a n in the c o n t r o l s c o m b i n e d (ear t w i t c h e s , F ( 1 , 5 4 ) = 13.76, p < 0 . 0 0 0 5 ; jaw clonus, F ( 1 , 5 4 ) = 9.94, p < 0 . 0 0 3 ; facial t r e m o r F ( 1 . 5 4 ) = 7.29, p < 0 . 0 1 ; m y o c l o n u s F ( 1 , 5 4 ) = 11.52, p < 0 . 0 0 2 ) . The responses to the t r e a t m e n t i n j e c t i o n s of m e t r a z o l were assessed a f t e r every third i n j e c t i o n d u r i n g the course of the e x p e r i m e n t , and the results of these assessments are p r e s e n t e d in Fig. 2 to illustrate the progressive d e v e l o p m e n t of s y m p t o m s within the e x p e r i m e n t a l animals. A l t h o u g h increases in the s u s c e p t i b i l i t y to m e t r a z o l i n j e c t i o n s following periodic p l a c e b o injections have been r e p o r t e d by o t h e r s [ 1 ], the p r e s e n t data i n d i c a t e t h a t such increases are n o t a critical f a c t o r in m e t r a z o l kindling.
METHOD The a n i m a l s were 57 male, 2 8 5 - 3 8 5 g, h o o d e d rats p u r c h a s e d f r o m t h e C a n a d i a n Breeding F a r m and Laboratories, St. C o n s t a n t , Q u e b e c . Each rat was i n j e c t e d intrap e r i t o n e a l l y w i t h 20 m g / k g of m e t r a z o l a n d the i n c i d e n c e of e a c h of f o u r convulsive s y m p t o m s - (1) r h y t h m i c ear t w i t c h e s , (2) j a w clonus, (3) facial t r e m o r s , a n d (4) m y o c l o n i c b o d y jerks - was assessed d u r i n g the ensuing ten 1-min intervals. Each rat received a score for each s y m p t o m w h i c h was t h e n u m b e r of l-rain intervals in w h i c h t h a t s y m p t o m was o b s e r v e d at least o n c e ; thus the m a x i m u m score t h a t e a c h s u b j e c t could o b t a i n for each s y m p t o m was ten. The r e s p o n s e s to this p r e t r e a t m e n t i n j e c t i o n were used as a basis for assigning the a n i m a l s t o t h r e e e q u i v a l e n t g r o u p s (n = 19). T h r e e days later, and at 3-day intervals t h e r e a f t e r , the s u b j e c t s in each of these g r o u p s received one of t h r e e t r e a t m e n t s . Rats in the e x p e r i m e n t a l g r o u p were i n j e c t e d every t h r e e days w i t h m e t r a z o l ( 2 0 mg/kg); rats in 599
600
PINEL AND CHEUNG
4
Ear Twitches
c-<~
Jaw CI0nus
o~j
Facial Tremors .yoclonic Jerks ~
//'/~,~
L
r,4
3
4
oc u~ z
i2
.a
u,a
1
pre' 4' treatment
i pre
pos t
i pre po~st METRAZOL
' pre
po; t
' pre
po', t
;
10 '
;
1
16 '
19 '
h
,'~
INJECTIONS
2;
31.
.34 .
37.
40 .
pest lrealmeet
INJECTIONS
lqG. 1. Convulsive responses elicited by metrazol before and after a series o f periodic metrazol injections {circles), placebo injections (squares), or control handling {triangles). Each point represents the mean n u m b e r of ten l-rain postinjection periods in which a particular s y m p t o m was observed in animals of that group.
FIG. 2. Progressive increases in the incidence of epileptic s y m p t o m s elicited by a series of 43 metrazol injections. Each point represents the mean n u m b e r of l-rain intervals in the 10 min l'ollowing each injection during which a particular s y m p t o m was observed at least once.
REFERENCES
2.
lzquierdo, 1., J. Fernandez, R. Oliveira and F. Settineri. Effect o f daily saline, drug or blank injections on the susceptibility to the convulsant effect of drugs. Pharmac. Biochem. Beha~,. 3: 721 722, 1975. Mason, C. R. and R. M. Cooper. A p e r m a n e n t change in convulsive threshold in normal and brain-damaged rats, with repeated small doses of pentylenetetrazol. Epilepxia 13: 6 6 3 - 6 7 4 , 1972.
3.
Pinel, J. P. J. and P. H. Van Oot. Generality of the kindling p h e n o m e n o n : some clinical implications. Can. J. Neurol. Sci. 2: 4 6 7 - 4 7 5 , 1975.
BRIEF COMMUNICATION Controlled Demonstration of Metrazol Kindling J O H N P. J. P I N E L A N D K. F. C H E U N G
Department o f Psychology, University o f British Columbia, Vancouver, British Columbia, V6T 1 W5 (Received 7 J a n u a r y 1977) PINEL, J. P. J. AND K. F. CHEUNG. Controlled demonstration of metrazol kindling. PHARMAC. BIOCHEM. BEHAV. 6(5) 5 9 9 - 6 0 0 , 1977. - Convulsive responses elicited in rats by intraperitoneal injections of metrazol were intensified following a series of metrazol injections administered once every three days. However, neither handling alone nor placebo injections increased the susceptibility of control rats to metrazol-induced convulsive symptoms. Thus, although increases in the susceptibility to metrazol seizures following periodic placebo injections have been reported by others, such increases do not appear to be a critical factor in metrazol kindling. Seizures
Rats
Metrazol
Kindling
P E R I O D I C i n j e c t i o n s of m e t r a z o l have b e e n s h o w n to p r o d u c e a progressive increase in seizure s u s c e p t i b i l i t y (kindling). F o r e x a m p l e , Mason a n d C o o p e r [2] f o u n d t h a t a dose o f m e t r a z o l w h i c h was initially s u b c o n v u l s i v e w o u l d e v e n t u a l l y elicit grand real seizures in rats i n j e c t e d w i t h it once every t h r e e days. However, r e c e n t l y I z q u i e r d o , Fern a n d e z , Oliveira, a n d S e t t i n e r i [1] r e p o r t e d t h a t increases in t h e s u s c e p t i b i l i t y to m e t r a z o l seizures f o l l o w e d a series of daily p l a c e b o injections. Since p l a c e b o c o n t r o l s were i n c l u d e d in n e i t h e r t h e M a s o n and C o o p e r s t u d y n o r in its r e p l i c a t i o n b y Pinel and V a n O o t [ 3 ] , the m e t r a z o l k i n d l i n g in these studies m a y have b e e n an a r t i f a c t of the i n j e c t i o n p r o c e d u r e r a t h e r t h a n being a c o n s e q u e n c e of t h e m e t r a z o l per se.
the p l a c e b o c o n t r o l g r o u p were injected at 3-day intervals with an e q u a l dose of i s o t o n i c saline; and rats in the second c o n t r o l g r o u p were h a n d l e d similarly b u t n o t injected. This t r e a t m e n t r e g i m e n t c o n t i n u e d until Day 127 w h e n all animals again received the s t a n d a r d 20 m g / k g i n j e c t i o n of metrazol. R e s p o n s e s to this p o s t t r e a t m e n t m e t r a z o l inject i o n were assessed b y an e x p e r i m e n t e r u n a w a r e of each animal's experimental history. RESULTS AND DISCUSSION The r e s p o n s e s of the animals in each of the t h r e e groups to t h e p r e t r e a t m e n t and p o s t t r e a t m e n t m e t r a z o l injections are s u m m a r i z e d in Fig. I. It is readily a p p a r e n t t h a t the i n c i d e n c e of each of the f o u r epileptic s y m p t o m s was increased a p p r e c i a b l y by the i n t e r v e n i n g series of m e t r a z o l i n j e c t i o n s b u t n o t b y e i t h e r of the t w o c o n t r o l p r o c e d u r e s ( p > 0 . 1 ) . Thus, a l t h o u g h animals were assigned to groups on the basis of t h e i r initial r e s p o n s e to m e t r a z o l , p l a n n e d c o m p a r i s o n s i n d i c a t e d t h a t b y t h e end of the e x p e r i m e n t each of the four s y m p t o m s was significantly m o r e prevalent in t h e e x p e r i m e n t a l group t h a n in the c o n t r o l s c o m b i n e d (ear t w i t c h e s , F ( 1 , 5 4 ) = 13.76, p < 0 . 0 0 0 5 ; jaw clonus, F ( 1 , 5 4 ) = 9.94, p < 0 . 0 0 3 ; facial t r e m o r F ( 1 . 5 4 ) = 7.29, p < 0 . 0 1 ; m y o c l o n u s F ( 1 , 5 4 ) = 11.52, p < 0 . 0 0 2 ) . The responses to the t r e a t m e n t i n j e c t i o n s of m e t r a z o l were assessed a f t e r every third i n j e c t i o n d u r i n g the course of the e x p e r i m e n t , and the results of these assessments are p r e s e n t e d in Fig. 2 to illustrate the progressive d e v e l o p m e n t of s y m p t o m s within the e x p e r i m e n t a l animals. A l t h o u g h increases in the s u s c e p t i b i l i t y to m e t r a z o l i n j e c t i o n s following periodic p l a c e b o injections have been r e p o r t e d by o t h e r s [ 1 ], the p r e s e n t data i n d i c a t e t h a t such increases are n o t a critical f a c t o r in m e t r a z o l kindling.
METHOD The a n i m a l s were 57 male, 2 8 5 - 3 8 5 g, h o o d e d rats p u r c h a s e d f r o m t h e C a n a d i a n Breeding F a r m and Laboratories, St. C o n s t a n t , Q u e b e c . Each rat was i n j e c t e d intrap e r i t o n e a l l y w i t h 20 m g / k g of m e t r a z o l a n d the i n c i d e n c e of e a c h of f o u r convulsive s y m p t o m s - (1) r h y t h m i c ear t w i t c h e s , (2) j a w clonus, (3) facial t r e m o r s , a n d (4) m y o c l o n i c b o d y jerks - was assessed d u r i n g the ensuing ten 1-min intervals. Each rat received a score for each s y m p t o m w h i c h was t h e n u m b e r of l-rain intervals in w h i c h t h a t s y m p t o m was o b s e r v e d at least o n c e ; thus the m a x i m u m score t h a t e a c h s u b j e c t could o b t a i n for each s y m p t o m was ten. The r e s p o n s e s to this p r e t r e a t m e n t i n j e c t i o n were used as a basis for assigning the a n i m a l s t o t h r e e e q u i v a l e n t g r o u p s (n = 19). T h r e e days later, and at 3-day intervals t h e r e a f t e r , the s u b j e c t s in each of these g r o u p s received one of t h r e e t r e a t m e n t s . Rats in the e x p e r i m e n t a l g r o u p were i n j e c t e d every t h r e e days w i t h m e t r a z o l ( 2 0 mg/kg); rats in 599
600
PINEL AND CHEUNG
4
Ear Twitches
c-<~
Jaw CI0nus
o~j
Facial Tremors .yoclonic Jerks ~
//'/~,~
L
r,4
3
4
oc u~ z
i2
.a
u,a
1
pre' 4' treatment
i pre
pos t
i pre po~st METRAZOL
' pre
po; t
' pre
po', t
;
10 '
;
1
16 '
19 '
h
,'~
INJECTIONS
2;
31.
.34 .
37.
40 .
pest lrealmeet
INJECTIONS
lqG. 1. Convulsive responses elicited by metrazol before and after a series o f periodic metrazol injections {circles), placebo injections (squares), or control handling {triangles). Each point represents the mean n u m b e r of ten l-rain postinjection periods in which a particular s y m p t o m was observed in animals of that group.
FIG. 2. Progressive increases in the incidence of epileptic s y m p t o m s elicited by a series of 43 metrazol injections. Each point represents the mean n u m b e r of l-rain intervals in the 10 min l'ollowing each injection during which a particular s y m p t o m was observed at least once.
REFERENCES
2.
lzquierdo, 1., J. Fernandez, R. Oliveira and F. Settineri. Effect o f daily saline, drug or blank injections on the susceptibility to the convulsant effect of drugs. Pharmac. Biochem. Beha~,. 3: 721 722, 1975. Mason, C. R. and R. M. Cooper. A p e r m a n e n t change in convulsive threshold in normal and brain-damaged rats, with repeated small doses of pentylenetetrazol. Epilepxia 13: 6 6 3 - 6 7 4 , 1972.
3.
Pinel, J. P. J. and P. H. Van Oot. Generality of the kindling p h e n o m e n o n : some clinical implications. Can. J. Neurol. Sci. 2: 4 6 7 - 4 7 5 , 1975.