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K+ channel openers decrease seizures in genetically epileptic rats
European Journal of Pharmacology, 167 (1989) 181-183 Elsevier
181
EJP 0211R
Rapid communication
K ÷ channel openers decrease seizures in genetically epileptic rats Gabriel G a n d o l f o 1, Sylvie R o m e t t i n o 1, C l a u d e G o t t e s m a n n 1, Gilles v a n Luijtelaar 2, A n t o n C o e n e n 2, Jean-Noel Bidard 3, a n d Michel L a z d u n s k i 3 1 Laboratoire de Psychophysiologie, Facultd des Sciences et Techniques, 06034 Nice C~dex, France, 2 Department of Comparative and Physiological Psychology, University of Nijmegen, The Netherlands, and 3 Centre de Biochimie, Centre National de la Recherche Scientifique, Parc Valrose, Nice C~dex, France Received 6 July 1989, accepted 10 July 1989
The inbred strain of rats, W A G / R i j , shows paroxysmal discharges occurring spontaneously in the cortex and seems to correspond to a model of absence epilepsy (Van Luijtelaar and Coenen, 1986). Spike-wave complexes appear in bursts reaching several seconds, during quiet waking or during the transition between slow wave sleep and waking, predominantly in the dark (Van Luijtelaar and Coenen, 1986). (-)BRL34915 (Cromakalim), the active isomer of a K ÷ channel opener developed for hypertension therapy (Cook, 1988), has been shown to prevent the appearance of paroxysmal activities induced in normal rats by a K ÷ channel blocker, the mast cell-degranulating peptide ( M C D ) f r o m bee venom (Gandolfo et al., 1989). This prompted us to study the influence of (-)BRL34915 on the spike-wave complexes of this model of petit mal epilepsy. Five W A G / R i j rats were anaesthetized (55 m g / k g i.p. of penthiobarbital) and implanted with a bevel-edged cannula directed towards the lateral ventricle. Permanent silver balls were placed on the frontal, parietal and occipital cortex. The electromyogram was recorded from the dorsal neck muscles. Postoperative recovery with cables for habituation lasted at least one week in an inverted 12:12 lighting. The ambient temperature was
Correspondence to: C. Gottesmann, Laboratoire de Psychophysiologic, Faeult6 des Sciences et Techniques, 06034 Nice C6dex, France.
maintained constant (23°C). Recordings were made from each rat for several days as control, then as sham (7% v / v ethanol/saline solution) recording and finally for three days after administration of 10 nmol of (-)BRL34915. The injections, were intracerebroventricular (i.c.v. 10/~1 for 2 min, the injection needle remaining in the cannula 2 more min to avoid flowing back of the solution) at 11 a.m. during a darkness period (from 7 a.m. to 7 p.m.) during which seizures predominate. The spike-wave bursts were quantified each hour for number, amplitude and duration. Because of the circadian distribution of the spike-wave bursts (Van Luijtelaar and Coenen, 1986) we dissociated two darkness periods one from 11 a.m. to 2 p.m. (called 'morning') and one from 2 p.m. to 5 p.m. (called 'afternoon'). Statistical analyses were done with Student's t-test. Figure 1A shows that the mean number of spike-wave bursts per hour in the control rats was slightly greater in the 'morning' than in the 'afternoon' (means _+ S.E.M.: 12.30 +_ 1.52 versus 8.20 _+ 0.78). This non-significant difference was abolished on the 'sham' day (8.47 +_ 0.29 versus 9.08_ 0.75), probably because of a transitory effect due to the alcohol in the vehicle solution. The number of bursts after i.c.v, injection of ( - ) B R L 34915 showed a slight tendency to decrease in the 'morning' (7.60 _+ 0.55). However a significant (P < 0.05) decrease (at 3.74 _ 0.36) was found in the 'afternoon', i.e. from 3 h after the administration of the K ÷ channel opener. On the following day,
0014-2999/89/$03.50 © 1989 Elsevier Science Publishers B.V. (Biomedical Division)
182
A 15"
[]
MORNING
•
AFTERNOON
0 C
S
DAY 1
DAY 2
DAY
13 Control
(-)BRL 349 15
P O EMG Fig. 1. Effects of (-)BRL34915 upon spontaneous spike-wave bursts. (A) Decrease of the number of bursts: the mean ( + S.E.M.) of bursts per hour was counted in the 'morning' (from 11 a.m. to 2 p.m.) and in the 'afternoon' (from 2 p.m. to 5 p.m.) during the darkness period. Vehicle (sham day) or (-)BRL34915 (on day 1) was injected i.c.v, at 11 a.m. (-)BRL34915 decreased the number of bursts in the 'afternoon' (from 3 h after injection). This number returned to its earlier value in the 'afternoon' of the second day and in the 'morning' of the third day. C: control; S: sham; * P < 0 . 0 5 (Student's t-test); n = 5. (B) Absence of effect upon amplitude and duration of the bursts: the two recordings performed in the 'afternoon' did not show significant change of the spikewave burst, which occurred at the frontal level during a transition between slow wave sleep and waking. Abbreviations: F: frontal cortex; P: parietal cortex; O: occipital cortex; EMG: dorsal neck electromyogram. Calibration: 200/xV; 1 s.
afternoon of the 3rd day showed no significant difference from the control (10.67 versus 9.50). At no stage were the amplitude (550 + 30 /~V) and duration (5.61 + 0.06 s) of the spike-wave bursts significantly modified by the K ÷ channel opener treatment (fig. 1B). This observation means that seizures are fully developed when they start. This work showed clearly that opening of K ÷ channels by ( - ) B R L 3 4 9 1 5 (Cook, 1988) significantly decreased the number of spike-wave bursts in genetically epileptic rats. ( - ) B R L 3 4 9 1 5 has been shown to have a potent anticonvulsant effect in a model of epilepsy induced by MCD, a K + channel blocker (Gandolfo et al., 1989). This effect was highly selective since ( - ) B R L 3 4 9 1 5 was unable to prevent paroxysmal crises induced by two other K + channel blockers, dendrotoxin I and 4-aminopyridine, which have a wider spectrum of selectivity for other subtypes of K ÷ channels, (Gandolfo et al., in press). The data presented here suggest that the occurrence of spike-wave bursts in this inbred strain of rats showing spontaneous epilepsy may be associated with abnormal functioning of MCD-sensitive K ÷ channels or may be associated with an abnormal level of an endogeneous MCD-like peptide which has been identified in mamalian brain (Cherubini et al., 1987).
Acknowledgements This work was supported by the Centre National de la Recherche Scientifique, the Fondation pour la Recherche Mrdicale and the Ministrre de la Drfense Nationale (Grant DRET 85/054). We thank Dr. M.J. Soulal (Beecham) for very kindly supplying us with BRL34915. The skilful secretarial assistance of C. Roulinat-Bettelheim is gratefully acknowledged.
References the number of bursts remained lowered in the 'morning' (8.08 + 0.85) but there was a recovery to the control value in the 'afternoon' (9.18 + 1.48). On the third day, the number of bursts seen in the 'morning' increased back towards.the control values found for untreated mutants (11.33_ 2.12). One rat that was also recorded during the
Cherubini, E., Y. Ben Ari, M. Gho, J.-N. Bidard and M. Lazdunski, 1987, Long-term potentiation of synaptic transmission in the hippocampus induced by a bee venom peptide, Nature (London) 328, 70. Cook, N.S., 1988, The pharmacology of potassium channels and their therapeutic potential, Trends Pharmacol. Sci. 9, 21.
183 Gandolfo, G., C. Gottesmann, J.-N. Bidard and M. Lazdunski, 1989, K + channel openers prevent epilepsy induced by the bee venom peptide MCD, European J. Pharmacol. 159, 329. Gandolfo, G., C. Gottesmann, J.-N. Bidard and M. Lazdunski, Sub-types of K + channels differentiated by the effect of
K + channel openers upon K + channel blockerinduced seizures, Brain Res. (in press). Van Luijtelaar, E.L.J.M. and A.M.L. Coenen, 1986, Two types of electrocortical paroxysms in an inbred strain of rats, Neurosci. Lett. 70, 393.
181
EJP 0211R
Rapid communication
K ÷ channel openers decrease seizures in genetically epileptic rats Gabriel G a n d o l f o 1, Sylvie R o m e t t i n o 1, C l a u d e G o t t e s m a n n 1, Gilles v a n Luijtelaar 2, A n t o n C o e n e n 2, Jean-Noel Bidard 3, a n d Michel L a z d u n s k i 3 1 Laboratoire de Psychophysiologie, Facultd des Sciences et Techniques, 06034 Nice C~dex, France, 2 Department of Comparative and Physiological Psychology, University of Nijmegen, The Netherlands, and 3 Centre de Biochimie, Centre National de la Recherche Scientifique, Parc Valrose, Nice C~dex, France Received 6 July 1989, accepted 10 July 1989
The inbred strain of rats, W A G / R i j , shows paroxysmal discharges occurring spontaneously in the cortex and seems to correspond to a model of absence epilepsy (Van Luijtelaar and Coenen, 1986). Spike-wave complexes appear in bursts reaching several seconds, during quiet waking or during the transition between slow wave sleep and waking, predominantly in the dark (Van Luijtelaar and Coenen, 1986). (-)BRL34915 (Cromakalim), the active isomer of a K ÷ channel opener developed for hypertension therapy (Cook, 1988), has been shown to prevent the appearance of paroxysmal activities induced in normal rats by a K ÷ channel blocker, the mast cell-degranulating peptide ( M C D ) f r o m bee venom (Gandolfo et al., 1989). This prompted us to study the influence of (-)BRL34915 on the spike-wave complexes of this model of petit mal epilepsy. Five W A G / R i j rats were anaesthetized (55 m g / k g i.p. of penthiobarbital) and implanted with a bevel-edged cannula directed towards the lateral ventricle. Permanent silver balls were placed on the frontal, parietal and occipital cortex. The electromyogram was recorded from the dorsal neck muscles. Postoperative recovery with cables for habituation lasted at least one week in an inverted 12:12 lighting. The ambient temperature was
Correspondence to: C. Gottesmann, Laboratoire de Psychophysiologic, Faeult6 des Sciences et Techniques, 06034 Nice C6dex, France.
maintained constant (23°C). Recordings were made from each rat for several days as control, then as sham (7% v / v ethanol/saline solution) recording and finally for three days after administration of 10 nmol of (-)BRL34915. The injections, were intracerebroventricular (i.c.v. 10/~1 for 2 min, the injection needle remaining in the cannula 2 more min to avoid flowing back of the solution) at 11 a.m. during a darkness period (from 7 a.m. to 7 p.m.) during which seizures predominate. The spike-wave bursts were quantified each hour for number, amplitude and duration. Because of the circadian distribution of the spike-wave bursts (Van Luijtelaar and Coenen, 1986) we dissociated two darkness periods one from 11 a.m. to 2 p.m. (called 'morning') and one from 2 p.m. to 5 p.m. (called 'afternoon'). Statistical analyses were done with Student's t-test. Figure 1A shows that the mean number of spike-wave bursts per hour in the control rats was slightly greater in the 'morning' than in the 'afternoon' (means _+ S.E.M.: 12.30 +_ 1.52 versus 8.20 _+ 0.78). This non-significant difference was abolished on the 'sham' day (8.47 +_ 0.29 versus 9.08_ 0.75), probably because of a transitory effect due to the alcohol in the vehicle solution. The number of bursts after i.c.v, injection of ( - ) B R L 34915 showed a slight tendency to decrease in the 'morning' (7.60 _+ 0.55). However a significant (P < 0.05) decrease (at 3.74 _ 0.36) was found in the 'afternoon', i.e. from 3 h after the administration of the K ÷ channel opener. On the following day,
0014-2999/89/$03.50 © 1989 Elsevier Science Publishers B.V. (Biomedical Division)
182
A 15"
[]
MORNING
•
AFTERNOON
0 C
S
DAY 1
DAY 2
DAY
13 Control
(-)BRL 349 15
P O EMG Fig. 1. Effects of (-)BRL34915 upon spontaneous spike-wave bursts. (A) Decrease of the number of bursts: the mean ( + S.E.M.) of bursts per hour was counted in the 'morning' (from 11 a.m. to 2 p.m.) and in the 'afternoon' (from 2 p.m. to 5 p.m.) during the darkness period. Vehicle (sham day) or (-)BRL34915 (on day 1) was injected i.c.v, at 11 a.m. (-)BRL34915 decreased the number of bursts in the 'afternoon' (from 3 h after injection). This number returned to its earlier value in the 'afternoon' of the second day and in the 'morning' of the third day. C: control; S: sham; * P < 0 . 0 5 (Student's t-test); n = 5. (B) Absence of effect upon amplitude and duration of the bursts: the two recordings performed in the 'afternoon' did not show significant change of the spikewave burst, which occurred at the frontal level during a transition between slow wave sleep and waking. Abbreviations: F: frontal cortex; P: parietal cortex; O: occipital cortex; EMG: dorsal neck electromyogram. Calibration: 200/xV; 1 s.
afternoon of the 3rd day showed no significant difference from the control (10.67 versus 9.50). At no stage were the amplitude (550 + 30 /~V) and duration (5.61 + 0.06 s) of the spike-wave bursts significantly modified by the K ÷ channel opener treatment (fig. 1B). This observation means that seizures are fully developed when they start. This work showed clearly that opening of K ÷ channels by ( - ) B R L 3 4 9 1 5 (Cook, 1988) significantly decreased the number of spike-wave bursts in genetically epileptic rats. ( - ) B R L 3 4 9 1 5 has been shown to have a potent anticonvulsant effect in a model of epilepsy induced by MCD, a K + channel blocker (Gandolfo et al., 1989). This effect was highly selective since ( - ) B R L 3 4 9 1 5 was unable to prevent paroxysmal crises induced by two other K + channel blockers, dendrotoxin I and 4-aminopyridine, which have a wider spectrum of selectivity for other subtypes of K ÷ channels, (Gandolfo et al., in press). The data presented here suggest that the occurrence of spike-wave bursts in this inbred strain of rats showing spontaneous epilepsy may be associated with abnormal functioning of MCD-sensitive K ÷ channels or may be associated with an abnormal level of an endogeneous MCD-like peptide which has been identified in mamalian brain (Cherubini et al., 1987).
Acknowledgements This work was supported by the Centre National de la Recherche Scientifique, the Fondation pour la Recherche Mrdicale and the Ministrre de la Drfense Nationale (Grant DRET 85/054). We thank Dr. M.J. Soulal (Beecham) for very kindly supplying us with BRL34915. The skilful secretarial assistance of C. Roulinat-Bettelheim is gratefully acknowledged.
References the number of bursts remained lowered in the 'morning' (8.08 + 0.85) but there was a recovery to the control value in the 'afternoon' (9.18 + 1.48). On the third day, the number of bursts seen in the 'morning' increased back towards.the control values found for untreated mutants (11.33_ 2.12). One rat that was also recorded during the
Cherubini, E., Y. Ben Ari, M. Gho, J.-N. Bidard and M. Lazdunski, 1987, Long-term potentiation of synaptic transmission in the hippocampus induced by a bee venom peptide, Nature (London) 328, 70. Cook, N.S., 1988, The pharmacology of potassium channels and their therapeutic potential, Trends Pharmacol. Sci. 9, 21.
183 Gandolfo, G., C. Gottesmann, J.-N. Bidard and M. Lazdunski, 1989, K + channel openers prevent epilepsy induced by the bee venom peptide MCD, European J. Pharmacol. 159, 329. Gandolfo, G., C. Gottesmann, J.-N. Bidard and M. Lazdunski, Sub-types of K + channels differentiated by the effect of
K + channel openers upon K + channel blockerinduced seizures, Brain Res. (in press). Van Luijtelaar, E.L.J.M. and A.M.L. Coenen, 1986, Two types of electrocortical paroxysms in an inbred strain of rats, Neurosci. Lett. 70, 393.