Astrid G. Chapman

The anticonvulsant effect of the non-NMDA antagonists, NBQX and GYKI 52466, in mice

The excitatory amino acid antagonists, NBQX (2,3-dihydroxy-6-nitro-7-sulfamoylbenzo(F)quinoxaline) and GYKI 52466 (1-(4-aminophenyl)-4-methyl-7,8-methylenedioxy-5H-2,3-benzodiazepine) that act on non-NMDA receptors, provide potent anticonvulsant protection against AMPA [RS)-alpha-amino-3-hydroxy-5-methyl-4-isoxazolepropionic acid)-induced seizures in Swiss mice and against sound-induced seizures in seizure-susceptible DBA/2 mice. Maximal anticonvulsant protection is observed 5-30 min after the i.p. administration of NBQX and 5-15 min after the i.p. administration of GYKI 52466 in DBA/2 mice. The ED50 values for the protection against AMPA-induced seizures by NBQX (30 min, i.p.) and GYKI 52466 (15 min, i.p.) are 23.6 (11.6-48.0) and 18.5 (11.5-29.5) mumol/kg, respectively. The ED50 values at 15 min for the protection against sound-induced seizures in DBA/2 mice are 31.3 (24.9-39.4) mumol/kg (NBQX, i.p.), 37.8 (21.2-67.4) mumol/kg (NBQX, i.v.) and 13.7 (11.5-16.5) mumol/kg (GYKI 52466, i.p.). In DBA/2 mice the therapeutic index (ratio of ED50 values for impaired rotarod performance and anticonvulsant action) is 6.6 for NBQX (15 and 30 min, i.p.) and 2.0 for GYKI 52466 (15 min, i.p.).

Glutamate metabotropic receptors as targets for drug therapy in epilepsy

Metabotropic glutamate (mGlu) receptors have multiple actions on neuronal excitability through G-protein-linked modifications of enzymes and ion channels. They act presynaptically to modify glutamatergic and gamma-aminobutyric acid (GABA)-ergic transmission and can contribute to long-term changes in synaptic function. The recent identification of subtype-selective agonists and antagonists has permitted evaluation of mGlu receptors as potential targets in the treatment of epilepsy. Agonists acting on group I mGlu receptors (mGlu1 and mGlu5) are convulsant. Antagonists acting on mGlu1 or mGlu5 receptors are anticonvulsant against 3,5-dihydroxyphenylglycine (DHPG)-induced seizures and in mouse models of generalized motor seizures and absence seizures. The competitive, phenylglycine mGlu1/5 receptor antagonists generally require intracerebroventricular administration for potent anticonvulsant efficacy but noncompetitive antagonists, e.g., (3aS,6aS)-6a-naphthalen-2-ylmethyl-5-methyliden-hexahydrocyclopenta[c]furan-1-on (BAY36-7620), 2-methyl-6-(phenylethynyl)pyridine hydrochloride (MPEP), and 2-methyl-6-(2-phenylethenyl)pyridine (SIB-1893) block generalized seizures with systemic administration. Agonists acting on group II mGlu receptors (mGlu2, mGlu3) to reduce glutamate release are anticonvulsant, e.g., 2R,4R-aminopyrrolidine-2,4-dicarboxylate [(2R,4R)-APDC], (+)-2-aminobicyclo[3.1.0]hexane-2,6-dicarboxylic acid (LY354740), and (-)-2-oxa-4-aminobicyclo[3.1.0]hexane-4,6-dicarboxylate (LY379268). The classical agonists acting on group III mGlu receptors such as L-(+)-2-amino-4-phosphonobutyric acid, and L-serine-O-phosphate are acutely proconvulsant with some anticonvulsant activity. The more recently identified agonists (R,S)-4-phosphonophenylglycine [(R,S)-PPG] and (S)-3,4-dicarboxyphenylglycine [(S)-3,4-DCPG] and (1S,3R,4S)-1-aminocyclopentane-1,2,4-tricarboxylic acid [ACPT-1] are all anticonvulsant without proconvulsant effects. Studies in animal models of kindling reveal some efficacy of mGlu receptor ligands against fully kindled limbic seizures. In genetic mouse models, mGlu1/5 antagonists and mGlu2/3 agonists are effective against absence seizures. Thus, antagonists at group I mGlu receptors and agonists at groups II and III mGlu receptors are potential antiepileptic agents, but their clinical usefulness will depend on their acute and chronic side effects. Potential also exists for combining mGlu receptor ligands with other glutamatergic and non-glutamatergic agents to produce an enhanced anticonvulsant effect. This review also discusses what is known about mGlu receptor expression and function in rodent epilepsy models and human epileptic conditions.

Anticonvulsant activity of two metabotropic glutamate Group I antagonists selective for the mGlu5 receptor: 2-methyl-6-(phenylethynyl)-pyridine (MPEP), and (E)-6-methyl-2-styryl-pyridine (SIB 1893)

The selective mGlu5 antagonists, MPEP, 2-methyl-6-phenylethynyl-pyridine, and SIB1893, (E)-6-methyl-2-styryl-pyridine, have been evaluated as antiepileptic drugs in DBA/2 mice and lethargic mice. Clonic seizures induced by the selective mGlu5 agonist, (R,S)-2-chloro-5-hydroxyphenylglycine (CHPG), 3 micromol intracerebroventricularly (i.c.v.), are potently suppressed by both compounds (MPEP, ED(50)=0.42 [0.28-0.62] mg/kg intraperitoneally (i.p.); SIB 1893 ED(50)=0.19 [0.11-0.33] mg/kg i.p. ). Clonic seizures induced by the mGlu1,5 agonist, 3, 5-dihydroxyphenylglycine (DHPG), 1.5 micromol i.c.v., are less potently suppressed by both compounds (MPEP, ED(50)=22 [13-38] mg/kg i.p., 110 [67-180] nmol i.c.v.; SIB1893, ED(50)=31 [18-54] mg/kg i.p. , 95 [82-110] nmol i.c.v.). Sound-induced seizures in DBA/2 mice are suppressed at 15 min by MPEP and SIB 1893 (MPEP ED(50) clonic seizures=18 [10-32] mg/kg i.p., 93 [69-125] nmol i.c.v.; tonic seizures=6.1 [4.5-8.3] mg/kg i.p., 46 [26-80] nmol i.c.v.; SIB 1893 ED(50) clonic seizures=27 [17-44] mg/kg i.p., 825 [615-1108] nmol i. c.v., tonic seizures=5.4 [3.4-8.6] mg/kg i.p., 194 [113-332] nmol i. c.v.). The ED(50) for MPEP for impaired rotarod performance is 128 [83-193] mg/kg i.p., at 15 min, i.e. a therapeutic index for sound-induced seizures of 5-20. In lethargic mice (lh/lh), a genetic absence model, MPEP, 50 mg/kg i.p., caused a marked reduction in the incidence of spontaneous spike-and-wave discharges. These selective antagonists of mGlu5 block seizures due to activation of mGlu5 at very low systemic doses. At rather higher doses they block convulsive and non-convulsive primary generalised seizures.

Extracellular amino acid levels in hippocampus during pilocarpine-induced seizures.

Extracellular levels of aspartate, glutamate and glutamine were monitored by microdialysis in the dorsal hippocampus of freely moving rats following the administration of a convulsant dose of pilocarpine (400 mg/kg, i.p.). Rats were either pretreated with the glutamate uptake inhibitor, 1-trans-pyrrolidine-2,4-dicarboxylic acid (PDC, 1 mM in the perfusion medium, -25 min), or received pilocarpine directly. All rats injected with pilocarpine (with or without PDC pretreatment) developed limbic seizures (latency 15.4 +/- 2.4 min). Without PDC pretreatment there were no significant changes in extracellular levels of aspartate, glutamate and glutamine following pilocarpine administration until the onset of limbic seizures when glutamine levels fell by 35%. Following PDC pretreatment there were large and sustained increases in extracellular hippocampal aspartate (250%) and glutamate (55%) levels, but no significant change in the glutamine level. When pilocarpine was administered to this group of rats, there were further selective, significant, transient increases in the extracellular levels of aspartate (31%) and glutamate (18%) which preceded the onset of seizures. Aspartate and glutamate levels were not significantly increased (relative to PDC controls) during seizures. The conditions for pilocarpine-induced increases in aspartate and glutamate release were established in parallel groups of anaesthetised rats where pilocarpine was administered via a microdialysis probe in the dorsal hippocampus. Following the infusion of 10 mM pilocarpine there were large and rapid increases in the levels of aspartate (143%) and glutamate (179%), which were completely abolished by the absence of calcium in the perfusion medium, or by the presence of atropine (20 mM) or tetrodotoxin (1 microM).

Prolonged anticonvulsant action of glutamate metabotropic receptor agonists in inferior colliculus of genetically epilepsy-prone rats

The anticonvulsant activity of (S)-4-carboxy-3-hydroxyphenylglycine ((S)-4C3HPG) (an antagonist of Group I and an agonist of Group II metabotropic glutamate (mGlu) receptors), of (1S,3S)-1-aminocyclopentane-1,3-dicarboxylic acid ((1S,3S)-ACPD) (an agonist of Group II mGlu receptors), and of L-serine-O-phosphate (an agonist of Group III mGlu receptors) was studied against sound-induced seizures in genetically epilepsy-prone (GEP) rats following bilateral microinjection into the inferior colliculus. All 3 drugs produce dose-dependent suppression of all phases of sound-induced seizures (wild running, clonic and tonic). (S)-4C3HPG produces an immediate and short-lasting (< 2 h) protection against sound-induced seizures with an ED50 value of 4.3 (3.2-5.7) nmol, at 5 min. The preferential agonists of Group II and Group III mGlu receptors produce an immediate, transient (< 10 min) proconvulsant effect followed by a prolonged (> 1 day) anticonvulsant effect against sound-induced seizures. The anticonvulsant ED50 value for (1S,3S)-ACPD is 9 (5-18) nmol at 2 h, and for L-serine-O-phosphate is 36 (6.5-199) nmol at 2 days. It is concluded that mGlu receptor activation potently modifies seizure threshold.

Anticonvulsant activity of the NMDA antagonists, D(-)4-(3-phosphonopropyl) piperazine-2-carboxylic acid (D-CPP) and D(-)(E)-4-(3-phosphonoprop-2-enyl) piperazine-2-carboxylic acid (D-CPPene) in a rodent and a primate model of reflex epilepsy.

D-(-)4-(3-phosphonopropyl)piperazine-2-carboxylic acid (D-CPP) and its unsaturated analogue (D(-)(E)-4-(3-phosphonoprop-2-enyl) piperazine-2-carboxylic acid (D-CPPene) have been administered to DBA/2 mice (intracerebroventricularly, i.c.v., intraperitoneally, i.p., and orally, p.o.) and to photosensitive baboons, Papio papio (intravenously, i.v., and orally), and their effects on reflexly induced epileptic responses assessed. In DBA/2 mice the clonic phase of the seizure response to sound is suppressed by D-CPP with an ED50 of 5.5 micrograms/mouse, i.c.v.; 0.69 mg (2.75 mumol)/kg i.p. and 16.6 mg (65.8 mumol)/kg p.o. compared with, for D-CPPene, 2.2 micrograms/mouse i.c.v., 0.41 mg (1.54 mumol)/kg i.p. and 10.8 mg (40.2 mumol)/kg, p.o. In Papio papio myoclonic responses to stroboscopic stimulation are suppressed 24 and 48 h after D-CPP 32 mg (127 mumol)/kg p.o. Administration of D-CPPene 8-16 mg (30-60 mumol)/kg i.v. produces protection against myoclonic responses after 1-2 h, lasting for 48 h. Oral administration of D-CPPene 32-64 mg (119-239 mumol)/kg produces protection beginning after 4 h and sustained for 48 h. Measurements of plasma D-CPPene concentration show rapid clearance after i.v. injection and a low plasma concentration 1.5-5 h after oral administration. The prolonged anticonvulsant action of D-CPP and D-CPPene following oral administration suggests that these compounds merit evaluation as antiepileptic therapy in man.

Non-competitive N-methyl-D-aspartate antagonists protect against sound-induced seizures in DBA/2 mice

Non-competitive antagonists of the N-methyl-D-aspartate (NMDA) receptor have been evaluated as anticonvulsants against sound-induced seizures in DBA/2 mice. The ED50 values for protection against sound-induced clonic seizures 15 min following the intracerebroventricular (i.c.v.) and intraperitoneal (i.p.) administration are: MK-801, ED50 = 0.5 nmol (i.c.v.); 0.14 mumol/kg (i.p.); phencyclidine, ED50 = 14 nmol (i.c.v.); 1.9 mumol/kg (i.p.); dextrorphan, ED50 = 35 nmol (i.c.v.); 18.5 mumol/kg (i.p.); tiletamine, ED50 = 40 nmol (i.c.v.); 5.6 mumol/kg (i.p.); SKF-10047, ED50 = 50 nmol (i.c.v.); 23.5 mumol/kg (i.p.); dextromethorphan, ED50 = 70 nmol (i.c.v.); 28.0 mumol/kg (i.p.); ketamine, ED50 = 110 nmol (i.c.v.); 15.5 mumol/kg (i.p.). The anticonvulsant effects of ketamine and tiletamine are of short duration (10-30 min), whereas the anticonvulsant effects of MK-801 and dextromethorphan last for 45 min or longer. The effects of phencyclidine, SKF-10047 and dextrorphan are of intermediate duration. Mild to moderate behavioural excitation is associated with the anticonvulsant activity of all the non-competitive NMDA antagonists. For MK-801, phencyclidine, dextrorphan, SKF-10047 and ketamine there is a close correlation between their relative anticonvulsant potencies and their potencies for displacing [3H]MK-801. The anticonvulsant effect is likely to be primarily mediated via NMDA antagonism at the PCP/MK-801 site.

Acute anticonvulsant activity of structural analogues of valproic acid and changes in brain GABA and aspartate content

Ten analogues of valproic acid (substituted butyric, pentanoic and hexanoic acids) were tested for anticonvulsant activity against audiogenic seizures in DBA/2 mice. There is a consistent correlation between the structure of these branched-chain fatty acids and their anticonvulsant potency, the larger molecules being the more active. There is also a strong correlation between the anticonvulsant potency of these compounds and their ability to reduce cerebral aspartate levels. Cerebral GABA levels are elevated by most, but not all, of the actively anticonvulsant valproate analogues.

Anticonvulsant effects of 7-nitroindazole in rodents with reflex epilepsy may result from L-arginine accumulation or a reduction in nitric oxide or L-citrulline formation

1 To investigate the role of nitric oxide in epilepsy we have studied the effects of agents which affect nitric oxide synthesis in sound‐induced seizures in DBA/2 mice and in genetically epilepsy‐prone (GEP) rats. 2 The neuronal selective nitric oxide synthase inhibitor, 7‐nitroindazole (7‐NI) is anticonvulsant in these models with ED50 values against clonic seizures in mg kg−1 i.p. (times following injection) of: 74 (+0.25 h), 120 (+1 h) in DBA/2 mice, and 56 (+0.25 h), 42 (+0.5 h), 36 (+1 h), 28 (+2 h), 38 (+4 h), 93 (+8 h) in GEP rats. 3 Therapeutic indices (locomotor deficit ED50/anticonvulsant ED50) for 7‐NI are low, ranging from 0.6 to 1.1 at +0.25 h to +1 h after administration in GEP rats, but are more favourable at later times (1.6 at +2 h and 2.9 at +4 h). 4 The substrate for nitric oxide synthase, L‐arginine (500–5000 mg kg−1, i.p. or 100–300 μg, i.c.v.) but not D‐arginine (300 μg i.c.v.) is anticonvulsant in DBA/2 mice. L‐Arginine (500–5000 mg kg−1, i.p. or 1800–6000 μg, i.c.v.) is a more potent anticonvulsant than D‐arginine (1500–2500 mg kg−1, i.p. or 6000 μg, i.c.v.) in GEP rats. 5 In DBA/2 mice, L‐arginine (30 μg i.c.v.) reverses the anticonvulsant effect of 7‐NI (50 mg kg−1, i.p.). 6 In GEP rats, low dose L‐arginine (25–50 mg kg−1, i.p.) but not D‐arginine (50 mg kg−1, i.p.) reverses the anticonvulsant effect of low dose 7‐NI (25 mg kg−1, i.p.). A higher dose of L‐arginine (500 mg kg−1, i.p.) or 7‐NI (50 mg kg−1, i.p.) produces summation of anticonvulsant effect. 7 The product for nitric oxide synthase, L‐citrulline (250–831 μg i.c.v.), is convulsant in DBA/2 mice. 8 The anticonvulsant effect of the neuronal selective nitric oxide synthase inhibitor, 7‐nitroindazole, may therefore be mediated by L‐arginine accumulation, as well as by a reduction in nitric oxide and L‐citrulline formation in rodent models of reflex epilepsy.

Anticonvulsant actions of LY 367385 ((+)-2-methyl-4-carboxyphenylglycine) and AIDA ((RS)-1-aminoindan-1,5-dicarboxylic acid)

We have studied the effects in three rodent models of generalised convulsive or absence epilepsy of two antagonists of group I metabotropic glutamate receptors that are selective for the mGlu1 receptor. LY 367385 ((+)-2-methyl-4-carboxyphenylglycine) and AIDA ((RS)-1-aminoindan-1,5-dicarboxylic acid) have been administered intracerebroventricularly (i.c.v.) to DBA/2 mice and lethargic mice (lh/lh), and focally into the inferior colliculus of genetically epilepsy prone rats (GEPR). In DBA/2 mice both compounds produce a rapid, transient suppression of sound-induced clonic seizures (LY 367385: ED50 = 12 nmol, i.c.v., 5 min; AIDA: ED50 = 79 nmol, i.c.v., 15 min). In lethargic mice both compounds significantly reduce the incidence of spontaneous spike and wave discharges on the electroencephalogram, from <30 to >150 min after the administration of AIDA, 500 nmol, i.c.v., and from 30 to >150 min after the administration of LY 367385, 250 nmol, i.c.v. LY 367385, 50 nmol, suppresses spontaneous spike and wave discharges from 30 to 60 min. In genetically epilepsy prone rats both compounds reduce sound-induced clonic seizures. LY 367385, 160 nmol bilaterally, fully suppresses clonic seizures after 2-4 h. AIDA is fully effective 30 min after 100 nmol bilaterally. It is concluded that antagonists of mGlu1 receptors are potential anticonvulsant agents and that activation of mGlu1 receptors probably contributes to a variety of epileptic syndromes.