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Ethacrynic acid-induced convulsions and brain noradrenaline in mice
European Journal of Pharmacology, 179 (1990) 221-223
221
Elsevier EJP 20596 Short communication
Ethacrynic acid-induced convulsions and brain noradrenaline in mice M a s a f u m i Inoue, T a k u j i Hirose, Y o s h i m i t s u Fukai, X u n - t i n g Zeng, T o h r u Y a s u k u r a , S u m i o Ohnishi, T o s h i k o U r i u and C h i y o k o Inagaki Department of Pharmacology, Kansai Medical University, Fumizono-cho 1, Mortguchi-Ci{y, Osaka 570, Japan
Received 15 January 1990.accepted 13 Februa~ 1990
The intracerebroventricular injection of ethacrynic acid (a 50% convulsive dose; 50 ~g/mouse) accelerated brain noradrenaline turnover and decreased noradrenaline contents. The decrease in noradrenaline contents was antagonized by 2-amino-5-phosphonovalerate but not by diazepam. Both 2-amino-5-phosphonovalerate and diazepam suppressed the incidence of ethacrynic acid-induced convulsions while reserpine, ct-methyl-para-tyrosine or FLA-63 augmented it. The results suggest that stimulation by ethacrynic acid of excitatory amino acid neurons enhances-noradrenergic neuronal anticonvulsive activity. Ethacrynic acid; Noradrenaline; Convulsion; Epilepsy
I. Introduction
Intracerebroventricularly (i.c.v.) administered ethacrynic acid shows strong convulsive activity in mice, probably due to an impairment of CItransport processes resulting in the enhancement of excitatory amino acid neuron activity (Inoue et al., 1989). Glutamate (Beas-Zarate et al., 1985), kainic acid (Baran et al., 1987) and bicuculline (Calderirti et al., 1978) reportedly reduce noradrenaline (NA) contents. Further, depletion of brain NA in kindled rats (Kokaia et al., 1989) and those receiving kainic acid (Baran et al., 1989) potentiates seizure activity. Therefore, we examined the changes in brain noradrenergic functions in a new model of epilepsy, ethacrynic acidinduced status epilepticus,
Correspondence to: C. Inagaki. Department of Pharmacology, Kansai Medical University, Fumizono-cho 1, Moriguchi-City,
Osaka 570, Japan.
2. Materials and methods
Male D D Y K mice (Tokushima Experimental Animal Laboratory, Japan)weighing 20-30 g were given i.c.v, either a 50% convulsive dose (50 /~g in 10 /~1) of ethacrynic acid (Inoue et al., 1989) or physiological saline with or without pretreatment. To estimate N A turnover, the mice were given i.p. either a tyrosine hydroxylase (EC 1.14.16.2) inhibitor, a-methyl-DL-p-tyrosine methyl ester (aMT: 250 mg/kg), or a dopamine B-hydroxylase (EC 1.14.17.1) inhibitor, bis-(4-methyl-l-homopiperazinylthiocarbonyl)-disulfide (FLA-63: 40 mg/kg) 10 rain before the i.c.v, injection of ethacrynic acid. All animals were decapitated 1 h after the i.c.v, injection of ethacrynic acid or vehicle and the contents of NA in the whole brains were measured by HPLC with electrochemical detection as described previously (Inoue et al., 1987). The behavior of the animals was also monitored for the occurrence of full tonic-clonic convulsions for 1 h after the ethacrynic acid injection
0014-2999/90/$03.50 © 1990 ElsevierScience Publishers B.V. (Biomedical Division)
222 as described previously ( I n o u e et al., 1989). The drugs used were: ethacrynic acid, a - M T , FLA-63, D L - 2 - a m i n o - 5 - p h o s p h o n o v a l e r i c acid ( A P V ) (Sigma Chemical Co., U.S.A.) and diazepam analysis was d o n e by u n p a i r e d S t u d e n t ' s t-test for significance of changes in mean values a n d the
TABLE 2 Effects of various drugs on the incidence of ethacrynic acid-induced tonic-clonic convulsions. Ethacrynic acid (50 ~tg in 10 ~tl, i.c.v.) was given i.p. after the injection of the drugs listed, at the doses and time periods in parentheses. Observations for the incidence of the convulsions were made for 1 h. ~ P < 0.05 comparedto control mice; Chi-square test with Yates" correction was used.
Chi-square test with Yates' correction for the incidence of tonic-clonic convulsions.
Pretreatment
(Takeda Pharmaceutical Co., Japan). Statistical
No. of mice convulsing No. of mice used 15/29 0/7 ~' 0/7 ~
W i t h i n 1 h after the ethacrynic acid-injection, the contents of N A in whole b r a i n s decreased as compared with the control value (table 1). After
None (control) APV (50 mg/kg, 30 min) Diazepam (10 mg/kg. 30 rain) a-Methyl-para-tyrosine (250 mg/kg, 10 min) F L A - 6 3(40 mg/kg, 10 min) Reserpine(5 mg/kg, 24 h)
the i n h i b i t i o n of tyrosine hydroxylase and dopamine/3-hydroxylase by a - M T a n d FLA-63, respectively, the N A level in ethacrynic acid-treated mice decreased to a level less than that of the control group, i.e., ethacrynic acid e n h a n c e d N A turnover. T r e a t m e n t with APV or diazepam alone had n o effect on the catecholamine contents in the brain. Pretreatment with APV abolished the ethacrynic acid-elicited decrease in N A c o n t e n t s while p r e t r e a t m e n t with diazepam had no effect on it. No difference in d o p a m i n e c o n t e n t s was observed in ethacrynic acid-treated animals and
the c o r r e s p o n d i n g control, whether with or without pretreatment. T r e m o r , m y o c l o n u s jerk a n d / o r tonic-clonic c o n v u l s i o n s were produced by the i.c.v, injection of ethacrynic acid. In the mice pretreated with APV or diazepam, ethacrynic acid injection elicited tremor, but no tonic-clonic convulsions (table 2). P r e t r e a t m e n t with a - M T , F L A 63 or reserpine a u g m e n t e d the incidence of convulsions.
3. Results
9/9 " 9/9 a 8/8 ~
TABLE 1 Brain catecholamine contents 1 h after i.c.v, ethacrynic acid (EA) injection. Either ethacrynic acid (EA-treated; 50 ~tg in 10 #1) or the vehicle (control) was given i.c.v, without pretreatment or after the i.p. injection of the drugs listed~ at the doses and time periods shown in parentheses. All mice were killed 1 h after the i.c.v, injection of ethacrynic acid or the vehicle. Values (ng/g) are given as means_+S.E. for animals in the numbers shown in parentheses, a p < 0.05, unpaired Student's t-test, compared to the corresponding control mice. Treatment
Noradrenaline (ng/g) Control
Dopamine (ng/g) EA-treated
Control
EA-treated
Without pretreatment
503+20 (6) 380+21 ~ (11) a-Methyl-para-tyrosine (250 mg/kg, 10 rain) 324 5:9.0 (6) 211 + 8.6 ~(6) FLA-63 (40 mg/kg, 10 rain) 330_+31 (6) 207 ± 5.6 a(6) DL-2-Amino-5-phosphonovalerate (50 mg/kg, 30 rain) 490+ 7.0(4) 470±12 (4) Diazepam (10 mg/kg, 30 rain) 501+24 (4) 375_+ 18 a (4)
929-+34(6)
996+48(11)
657 :t: 21 (6)
669 + 11 (6)
1233_+40(6)
1173_+78(6)
1013_+71(4)
912+18(4)
983+24(4)
970±31(4)
223 4. Discussion The injection of ethacrynic acid decreased the level of b r a i n N A . E n h a n c e m e n t of N A turnover b u t not the inhibition of tyrosine h y d r o x y l a s e a n d d o p a m i n e f l - h y d r o x y l a s e a c c o u n t e d for this decrease in N A level. Similar changes in N A levels r e p o r t e d in other e x p e r i m e n t a l convulsions (BedsZ a r a t e et al., 1985; Baran et al., 1987; C a l d e r i n i et al., 1978) m a y also be e x p l a i n e d by an accelerated N A release. A n N - m e t h y l - D - a s p a r t a t e r e c e p t o r antagonist, APV, and d i a z e p a m c o m p l e t e l y suppressed the e t h a c r y n i c a c i d - i n d u c e d convulsions as r e p o r t e d previously ( I n o u e et al., 1989). A P V a n t a g o n i z e d the decrease in the N A level but d i a z e p a m had no effect on it. Direct effects of A P V on c a t e c h o l a m i n e m e t a b o l i s m in rat brain s y n a p t o s o m e s (Yee et al., 1989) are p r o b a b l y not involved in a n t a g o n i z i n g the decrease in N A levels, since APV, u n d e r the present e x p e r i m e n t a l conditions, had no effect on b r a i n N A levels in the control mice. Therefore, stimulation of g l u t a m a tergic neurons by ethacrynic acid is p r o b a b l y responsible for the accelerated N A release, directly or through other neural systems s y n a p t i c a l l y related to the a m i n e neurons. Indeed, N - m e t h y l - D a s p a r t a t e as an agonist of g l u t a m a t e receptors r e p o r t e d l y a u g m e n t s N A release from rat h i p p o c a m p a l slices ( S c h m i d t and Taylor, 1988). T h e anticonvulsive effects of d i a z e p a m a p p e a r to be i n d e p e n d e n t of the m e c h a n i s m affecting N A turnover in this model. Central N A neurons are a s s u m e d to have a n t i c o n v u l s a n t activity ( G e l l m a n et al., 1987; K o k a i a et al., 1989; Baran et al. 1989). T h e incidence of ethacrynic a c i d - i n d u c e d convulsions was increased by reserpine, a - M T or F L A - 6 3 , suggesting that decreased c a t e c h o l a m i n e r g i c activity in the b r a i n facilitates epileptogenesis with ethacrynic acid, as observed in k i n d l e d rats ( K o k a i a et al., 1989) a n d those receiving kainic acid (Baran et al., 1989). Thus, activation of N A neurons yields a c o m p e n s a t o r y anticonvulsive m e c h a n i s m in the present model of epilepsy. The lack of effects of ethacrynic acid on b r a i n d o p a mine contents and turnover indicates that d o p a m i n e r g i c neuron activity is unlikely to be involved in the anticonvulsive or convulsive mechanisms in ethacrynic a c i d - i n d u c e d convulsions,
In conclusion, in e t h a c r y n i c a c i d - i n d u c e d convulsive states, the release of b r a i n N A is facilitared, thus a n t a g o n i z i n g the convulsions through the s t i m u l a t i o n of N - m e t h y l - D - a s p a r t a t e t y p e exc i t a t o r y a m i n o acid receptors.
Acknowledgement This study was supported by research Grants from the Ministry of Fxtucation, Science and Culture, Japan, (01770145, 01570116)and from Kansai Medical University and by the Katano (}rant given by the Alumni Association of Kansai Medical University. The authors thank Kansai Medical University Laboratory Research Center and Laboratory Animal Center.
References Baran, H., H. Hortnagl and O. HornykiewicT, 1989, Kainic acid-induced seizures: potentiation by a-methyl-p-tyrosine, Brain Res. 495, 253. Baran, H., H. Lassmann. G. Sperk, F. Seitelberger and O. Hornykiewicz, 1987, Effect of mannitol treatment on brain neurotransminer markers in kainic acid-induced epilepsy, Neuroscience, 21,679. Beas-Z~rate, C., J. Arauz-Contreras, A. Velazquez and A. Ferio-Velasco, 1985, Monosodium I-ghitamate-induced convulsions-II. Changes in catecholamine concentrations in various areas of adult rats, Gen. Phaxmacol. 16, 489. Calderini, G., A. Carlsson and C.-H. NordstrOm, 1978, Monoamine metabolism during bicuculline-induced epileptic seizures in the rat, Brain Res. 157, 295. Gellman, R.L., J.A. Kallianos and J.O. McNamara, 1987, Alpha-2 receptors mediate an endogenous noradernergic suppression of kindling development, J. Pharmacol. Exp. Ther. 241,891. Inoue, M., K. Kurahashi and M. Fujiwara, 1987, Noradrenaline turnover after left coronary artery ligation in rat heart, European J. Pharmacol. 143, 251. lnoue, M., T. Uriu, H. Otani, M. Hard, K. Omori and C. lnagaki, 1989, Intracerebroventricular injection of ethacrrynic acid induces status epilepticus, European J. Pharmacol. 166. 101. Kokaia, M., J. Bengzon, P. Kalen and O. Lindvall, 1989, Noradrenergic mechanism in hippocampal kindling with rapidly recurring seizures, Brain Res. 491,398. Schmidt, C.J. and V.L. Taylor, 1988, Release of [3HInorepinephrine from rat hippocampal slices by N-methylD-aspartate: comparison of the inhibitory effects of Mg 2" and MK-801, European J. Pharmacol. 156, 111. Vee, D.K., A. Pastuszko, D. Nelson and D.F. Wilson, 1989, Effects of DL-2-amino-5-phosphonovalerate on metabolism of catecholamines in synaptosomes from rat brain, J. Neurochem. 52, 54.
221
Elsevier EJP 20596 Short communication
Ethacrynic acid-induced convulsions and brain noradrenaline in mice M a s a f u m i Inoue, T a k u j i Hirose, Y o s h i m i t s u Fukai, X u n - t i n g Zeng, T o h r u Y a s u k u r a , S u m i o Ohnishi, T o s h i k o U r i u and C h i y o k o Inagaki Department of Pharmacology, Kansai Medical University, Fumizono-cho 1, Mortguchi-Ci{y, Osaka 570, Japan
Received 15 January 1990.accepted 13 Februa~ 1990
The intracerebroventricular injection of ethacrynic acid (a 50% convulsive dose; 50 ~g/mouse) accelerated brain noradrenaline turnover and decreased noradrenaline contents. The decrease in noradrenaline contents was antagonized by 2-amino-5-phosphonovalerate but not by diazepam. Both 2-amino-5-phosphonovalerate and diazepam suppressed the incidence of ethacrynic acid-induced convulsions while reserpine, ct-methyl-para-tyrosine or FLA-63 augmented it. The results suggest that stimulation by ethacrynic acid of excitatory amino acid neurons enhances-noradrenergic neuronal anticonvulsive activity. Ethacrynic acid; Noradrenaline; Convulsion; Epilepsy
I. Introduction
Intracerebroventricularly (i.c.v.) administered ethacrynic acid shows strong convulsive activity in mice, probably due to an impairment of CItransport processes resulting in the enhancement of excitatory amino acid neuron activity (Inoue et al., 1989). Glutamate (Beas-Zarate et al., 1985), kainic acid (Baran et al., 1987) and bicuculline (Calderirti et al., 1978) reportedly reduce noradrenaline (NA) contents. Further, depletion of brain NA in kindled rats (Kokaia et al., 1989) and those receiving kainic acid (Baran et al., 1989) potentiates seizure activity. Therefore, we examined the changes in brain noradrenergic functions in a new model of epilepsy, ethacrynic acidinduced status epilepticus,
Correspondence to: C. Inagaki. Department of Pharmacology, Kansai Medical University, Fumizono-cho 1, Moriguchi-City,
Osaka 570, Japan.
2. Materials and methods
Male D D Y K mice (Tokushima Experimental Animal Laboratory, Japan)weighing 20-30 g were given i.c.v, either a 50% convulsive dose (50 /~g in 10 /~1) of ethacrynic acid (Inoue et al., 1989) or physiological saline with or without pretreatment. To estimate N A turnover, the mice were given i.p. either a tyrosine hydroxylase (EC 1.14.16.2) inhibitor, a-methyl-DL-p-tyrosine methyl ester (aMT: 250 mg/kg), or a dopamine B-hydroxylase (EC 1.14.17.1) inhibitor, bis-(4-methyl-l-homopiperazinylthiocarbonyl)-disulfide (FLA-63: 40 mg/kg) 10 rain before the i.c.v, injection of ethacrynic acid. All animals were decapitated 1 h after the i.c.v, injection of ethacrynic acid or vehicle and the contents of NA in the whole brains were measured by HPLC with electrochemical detection as described previously (Inoue et al., 1987). The behavior of the animals was also monitored for the occurrence of full tonic-clonic convulsions for 1 h after the ethacrynic acid injection
0014-2999/90/$03.50 © 1990 ElsevierScience Publishers B.V. (Biomedical Division)
222 as described previously ( I n o u e et al., 1989). The drugs used were: ethacrynic acid, a - M T , FLA-63, D L - 2 - a m i n o - 5 - p h o s p h o n o v a l e r i c acid ( A P V ) (Sigma Chemical Co., U.S.A.) and diazepam analysis was d o n e by u n p a i r e d S t u d e n t ' s t-test for significance of changes in mean values a n d the
TABLE 2 Effects of various drugs on the incidence of ethacrynic acid-induced tonic-clonic convulsions. Ethacrynic acid (50 ~tg in 10 ~tl, i.c.v.) was given i.p. after the injection of the drugs listed, at the doses and time periods in parentheses. Observations for the incidence of the convulsions were made for 1 h. ~ P < 0.05 comparedto control mice; Chi-square test with Yates" correction was used.
Chi-square test with Yates' correction for the incidence of tonic-clonic convulsions.
Pretreatment
(Takeda Pharmaceutical Co., Japan). Statistical
No. of mice convulsing No. of mice used 15/29 0/7 ~' 0/7 ~
W i t h i n 1 h after the ethacrynic acid-injection, the contents of N A in whole b r a i n s decreased as compared with the control value (table 1). After
None (control) APV (50 mg/kg, 30 min) Diazepam (10 mg/kg. 30 rain) a-Methyl-para-tyrosine (250 mg/kg, 10 min) F L A - 6 3(40 mg/kg, 10 min) Reserpine(5 mg/kg, 24 h)
the i n h i b i t i o n of tyrosine hydroxylase and dopamine/3-hydroxylase by a - M T a n d FLA-63, respectively, the N A level in ethacrynic acid-treated mice decreased to a level less than that of the control group, i.e., ethacrynic acid e n h a n c e d N A turnover. T r e a t m e n t with APV or diazepam alone had n o effect on the catecholamine contents in the brain. Pretreatment with APV abolished the ethacrynic acid-elicited decrease in N A c o n t e n t s while p r e t r e a t m e n t with diazepam had no effect on it. No difference in d o p a m i n e c o n t e n t s was observed in ethacrynic acid-treated animals and
the c o r r e s p o n d i n g control, whether with or without pretreatment. T r e m o r , m y o c l o n u s jerk a n d / o r tonic-clonic c o n v u l s i o n s were produced by the i.c.v, injection of ethacrynic acid. In the mice pretreated with APV or diazepam, ethacrynic acid injection elicited tremor, but no tonic-clonic convulsions (table 2). P r e t r e a t m e n t with a - M T , F L A 63 or reserpine a u g m e n t e d the incidence of convulsions.
3. Results
9/9 " 9/9 a 8/8 ~
TABLE 1 Brain catecholamine contents 1 h after i.c.v, ethacrynic acid (EA) injection. Either ethacrynic acid (EA-treated; 50 ~tg in 10 #1) or the vehicle (control) was given i.c.v, without pretreatment or after the i.p. injection of the drugs listed~ at the doses and time periods shown in parentheses. All mice were killed 1 h after the i.c.v, injection of ethacrynic acid or the vehicle. Values (ng/g) are given as means_+S.E. for animals in the numbers shown in parentheses, a p < 0.05, unpaired Student's t-test, compared to the corresponding control mice. Treatment
Noradrenaline (ng/g) Control
Dopamine (ng/g) EA-treated
Control
EA-treated
Without pretreatment
503+20 (6) 380+21 ~ (11) a-Methyl-para-tyrosine (250 mg/kg, 10 rain) 324 5:9.0 (6) 211 + 8.6 ~(6) FLA-63 (40 mg/kg, 10 rain) 330_+31 (6) 207 ± 5.6 a(6) DL-2-Amino-5-phosphonovalerate (50 mg/kg, 30 rain) 490+ 7.0(4) 470±12 (4) Diazepam (10 mg/kg, 30 rain) 501+24 (4) 375_+ 18 a (4)
929-+34(6)
996+48(11)
657 :t: 21 (6)
669 + 11 (6)
1233_+40(6)
1173_+78(6)
1013_+71(4)
912+18(4)
983+24(4)
970±31(4)
223 4. Discussion The injection of ethacrynic acid decreased the level of b r a i n N A . E n h a n c e m e n t of N A turnover b u t not the inhibition of tyrosine h y d r o x y l a s e a n d d o p a m i n e f l - h y d r o x y l a s e a c c o u n t e d for this decrease in N A level. Similar changes in N A levels r e p o r t e d in other e x p e r i m e n t a l convulsions (BedsZ a r a t e et al., 1985; Baran et al., 1987; C a l d e r i n i et al., 1978) m a y also be e x p l a i n e d by an accelerated N A release. A n N - m e t h y l - D - a s p a r t a t e r e c e p t o r antagonist, APV, and d i a z e p a m c o m p l e t e l y suppressed the e t h a c r y n i c a c i d - i n d u c e d convulsions as r e p o r t e d previously ( I n o u e et al., 1989). A P V a n t a g o n i z e d the decrease in the N A level but d i a z e p a m had no effect on it. Direct effects of A P V on c a t e c h o l a m i n e m e t a b o l i s m in rat brain s y n a p t o s o m e s (Yee et al., 1989) are p r o b a b l y not involved in a n t a g o n i z i n g the decrease in N A levels, since APV, u n d e r the present e x p e r i m e n t a l conditions, had no effect on b r a i n N A levels in the control mice. Therefore, stimulation of g l u t a m a tergic neurons by ethacrynic acid is p r o b a b l y responsible for the accelerated N A release, directly or through other neural systems s y n a p t i c a l l y related to the a m i n e neurons. Indeed, N - m e t h y l - D a s p a r t a t e as an agonist of g l u t a m a t e receptors r e p o r t e d l y a u g m e n t s N A release from rat h i p p o c a m p a l slices ( S c h m i d t and Taylor, 1988). T h e anticonvulsive effects of d i a z e p a m a p p e a r to be i n d e p e n d e n t of the m e c h a n i s m affecting N A turnover in this model. Central N A neurons are a s s u m e d to have a n t i c o n v u l s a n t activity ( G e l l m a n et al., 1987; K o k a i a et al., 1989; Baran et al. 1989). T h e incidence of ethacrynic a c i d - i n d u c e d convulsions was increased by reserpine, a - M T or F L A - 6 3 , suggesting that decreased c a t e c h o l a m i n e r g i c activity in the b r a i n facilitates epileptogenesis with ethacrynic acid, as observed in k i n d l e d rats ( K o k a i a et al., 1989) a n d those receiving kainic acid (Baran et al., 1989). Thus, activation of N A neurons yields a c o m p e n s a t o r y anticonvulsive m e c h a n i s m in the present model of epilepsy. The lack of effects of ethacrynic acid on b r a i n d o p a mine contents and turnover indicates that d o p a m i n e r g i c neuron activity is unlikely to be involved in the anticonvulsive or convulsive mechanisms in ethacrynic a c i d - i n d u c e d convulsions,
In conclusion, in e t h a c r y n i c a c i d - i n d u c e d convulsive states, the release of b r a i n N A is facilitared, thus a n t a g o n i z i n g the convulsions through the s t i m u l a t i o n of N - m e t h y l - D - a s p a r t a t e t y p e exc i t a t o r y a m i n o acid receptors.
Acknowledgement This study was supported by research Grants from the Ministry of Fxtucation, Science and Culture, Japan, (01770145, 01570116)and from Kansai Medical University and by the Katano (}rant given by the Alumni Association of Kansai Medical University. The authors thank Kansai Medical University Laboratory Research Center and Laboratory Animal Center.
References Baran, H., H. Hortnagl and O. HornykiewicT, 1989, Kainic acid-induced seizures: potentiation by a-methyl-p-tyrosine, Brain Res. 495, 253. Baran, H., H. Lassmann. G. Sperk, F. Seitelberger and O. Hornykiewicz, 1987, Effect of mannitol treatment on brain neurotransminer markers in kainic acid-induced epilepsy, Neuroscience, 21,679. Beas-Z~rate, C., J. Arauz-Contreras, A. Velazquez and A. Ferio-Velasco, 1985, Monosodium I-ghitamate-induced convulsions-II. Changes in catecholamine concentrations in various areas of adult rats, Gen. Phaxmacol. 16, 489. Calderini, G., A. Carlsson and C.-H. NordstrOm, 1978, Monoamine metabolism during bicuculline-induced epileptic seizures in the rat, Brain Res. 157, 295. Gellman, R.L., J.A. Kallianos and J.O. McNamara, 1987, Alpha-2 receptors mediate an endogenous noradernergic suppression of kindling development, J. Pharmacol. Exp. Ther. 241,891. Inoue, M., K. Kurahashi and M. Fujiwara, 1987, Noradrenaline turnover after left coronary artery ligation in rat heart, European J. Pharmacol. 143, 251. lnoue, M., T. Uriu, H. Otani, M. Hard, K. Omori and C. lnagaki, 1989, Intracerebroventricular injection of ethacrrynic acid induces status epilepticus, European J. Pharmacol. 166. 101. Kokaia, M., J. Bengzon, P. Kalen and O. Lindvall, 1989, Noradrenergic mechanism in hippocampal kindling with rapidly recurring seizures, Brain Res. 491,398. Schmidt, C.J. and V.L. Taylor, 1988, Release of [3HInorepinephrine from rat hippocampal slices by N-methylD-aspartate: comparison of the inhibitory effects of Mg 2" and MK-801, European J. Pharmacol. 156, 111. Vee, D.K., A. Pastuszko, D. Nelson and D.F. Wilson, 1989, Effects of DL-2-amino-5-phosphonovalerate on metabolism of catecholamines in synaptosomes from rat brain, J. Neurochem. 52, 54.