Ewart A. Swinyard

Laboratory Evaluation of Antiepileptic Drugs

The historical development of the laboratory evaluation of anticonvulsant drugs was briefly reviewed. Attention was directed to the fact that such evaluation can be approached from several biological levels. The numerous methods which have been employed for the study of antiepileptic drugs were summarized and mention made of their relative value in drug evaluation. The usual method for the determination of minimal neurotoxicity was described and six of the classical assay procedures were reviewed. It was pointed out that no one laboratory test is sufficient to delineate the anticonvulsant potential of a new drug and, therefore, a battery of 4 tests (Maximal Electroshock Seizure test, a.c. Electroshock Seizure Threshold test, Hyponatremic Electroshock Seizure Threshold test, and s.c. Metrazol test) has been employed in our laboratories. It was further emphasized that valuable confirmatory data and preliminary clinical information can be obtained by pre‐clinical testing of promising drugs in psychiatric patients undergoing electroshock seizure therapy. Finally, the predictive value of the battery of four tests was considered and the need for more selective assay procedures emphasized.

Comparative Anticonvulsant Activity and Neurotoxicity of Felbamate and Four Prototype Antiepileptic Drugs in Mice and Rats

Summary: Felbamate (2‐phenyl‐1, 3‐propanediol dicarbamate), phenytoin, phenobarbital, ethosuximide, and valproate were evaluated in mice and rats with a battery of well‐standardized anticonvulsant test procedures. The results obtained indicate that felbamate exhibits a wider range of experimental anticonvulsant activity than either phenytoin or ethosuximide and a somewhat more restricted range than either phenobarbital or valproate. Felbamate is effective in nontoxic intraperitoneal doses in mice by the maximal electroshock seizure (MES), pentylenetetrazol (s.c. PTZ), and picrotoxin (s.c. Pic) tests but ineffective against bicuculline‐ and strychnine‐induced seizures; it is effective after nontoxic oral doses in both mice and rats by the MES and s.c. PTZ tests. When compared on the basis of protective indices (PI = TD50/ED50) calculated from the intraperitoneal data in mice, the Pis for felbamate were from 1.05 to 2.37 times higher than those of the prototype antiepileptics. Overall, except for the s.c. PTZ test in mice and rats after oral administration, the Pis were equal to or higher than those of the prototype agents. The Pis for the s.c. PTZ test in mice and rats after oral administration were within the range of the prototype agents. These data indicate that felbamate is a relatively nontoxic agent with a unique profile of anticonvulsant action.

A Neuropharmacological Evaluation of Felbamate as a Novel Anticonvulsant

Summary: Felbamate (2‐phenyl‐1,3‐propanediol dicarbamate, FBM) was subjected to a series of carefully selected in vivo and in vitro tests to provide additional insight into mechanism of action, margin of safety, and clinical potential. FBM was effective against intracere broventricular (i.c.v.) N‐methyl‐D‐aspartate (NMDA)‐induced clonus and i.c.v. NMDA‐ and quisqualic acid (quis)‐induced forelimb tonic extension in mice and ineffective against i.c.v. quis‐induced clonus in mice. FBM was also effective in preventing the expression of Stage 5 kindled seizures in corneal‐kindled rats. The calculated protective indices (rotorod median toxic dose divided by anticonvulsant median effective dose) ranged from 28 to 146 for those tests in which FBM displayed activity. With the in vitro tests, FBM did not significantly displace [3H]MK‐801 from its binding site. In contrast, FBM was effective in blocking sustained repetitive firing in mouse spinal cord neurons grown in tissue culture (median inhibitory concentration 67 μg/ml). This effect on repetitive firing suggests indirectly that FBM modulates sodium channel conductance. The results, when compared to similar data for phenytoin, carbamazepine, valproate, and ethosuximide, support the concept that FBM is a relatively nontoxic agent with a unique profile of anticonvulsant action, a broad margin of safety, and a clinical potential that includes at least generalized tonic‐clonic and complex partial seizures.

Comparative Anticonvulsant Activity and Neurotoxicity of Clobazam, Diazepam, Phenobarbital, and Valproate in Mice and Rats

Summary: The 1,5‐benzodiazepine (clobazam), the 1,4‐benzodiazepine (diazepam), and two nonbenzodiazepine antiepileptic drugs (phenobarbital and valproate) were evaluated in mice and rats with a battery of well‐standardized anticonvulsant test procedures. The results obtained indicate that clobazam and valproate exhibit a wider range of experimental anticonvulsant activity than either diazepam or phenobarbital. Except for clobazam by the maximal electroshock seizure (MES) test in rats, clobazam and valproate are effective in nontoxic doses against MES and all four chemically induced seizures (Met‐razol®, bicuculline, picrotoxin, and strychnine). Clobazam is effective by the MES test in rats only in doses that exceed the median minimal toxic dose. Phenobarbital is effective against all of the above tests, but minimal toxic doses must be employed to prevent strychnine seizures. Diazepam, on the other hand, is effective in nontoxic doses against seizures induced by Metrazol, bicuculline, and picrotoxin, but protects animals from maximal electroshock and strychnine seizures only when given in toxic doses. When compared on the basis of protective indices (PI = TD50/ED50) calculated from intraperitoneal data, the Pis for clobazam were 1.6 to 13 times higher than those for diazepam. Overall, except for the MES test in rats, the Pis for clobazam were from 1.5 to 44 times higher than those for any of the other three substances. With respect to the MES test in rats, the PI for clobazam was 10.8 times higher than that for diazepam; however, the Pis for phenobarbital and valproate were 3.5 and 4.4 times higher, respectively, than that for clobazam. These data suggest that the spectrum of anticonvulsant activity for the 1,5‐benzodiazepine (clobazam) is superior to that for the 1,4‐benzodiazepine (diazepam). Also, the broad experimental profile of anticonvulsant activity of clobazam agrees well with its reported broad clinical efficacy.

Preclinical profile of remacemide: A novel anticonvulsant effective against maximal electroshock seizures in mice

Anticonvulsant tests in mice revealed specific, potent actions of remacemide for protection of mice against maximal electroshock seizures (MES). Comparisons of oral efficacy to reference compounds yielded the following ED50 values (expressed as mg/kg): remacemide = 33, phenytoin = 11, phenobarbital = 20, carbamazepine = 13 and valproate = 631. The duration for protection by remacemide was longer than carbamazepine or valproate, but shorter than phenytoin or phenobarbital. In neural impairment tests (inverted screen or rotorod) to determine the oral toxic dose 50 (TD50) the following therapeutic indices (TD50/ED50) were obtained: (1) inverted screen--remacemide = 17.6, phenytoin = 57.4, phenobarbital = 5.1, carbamazepine = 10.2, and valproate = greater than 3; and (2) rotorod--remacemide = 5.6, phenytoin = 9.6, phenobarbital 4.8, and valproate = 1.9. Remacemide was devoid of sedative actions and possessed a favorable 28.1 margin of safety value (median estimated lethal dose/ED50 for MES). An intermediate potency against either audiogenic- or N-methyl-D-aspartate-induced seizures was exhibited by remacemide. Tolerance to MES was not apparent after 5 days of oral daily dosing of remacemide. Remacemide was inactive in vitro against gamma-aminobutyrate or benzodiazepine receptors and adenosine uptake mechanisms. Therapeutic utility for generalized tonic/clonic seizures is predicted for remacemide.

The anticonvulsant properties of tridione: Laboratory and clinical investigations

T RIDIONE, 3,5,5-trimethyloxazolidine2, 4-dione, has recently been made available to physicians as a specific symptomatic therapy for petit ma1 epilepsy, and has been accepted by the Council on Pharmacy and Chemistry of the American Medical Association for inclusion in New and Nonofficial Remedies. The drug was synthesized by Spielman31 and initially studied and reported as an analgesic agent. 26,28 Its pharmacological and anticonvulsant properties were described by Richards and Everett8r26,27,28 and by LIS.~~~~~,-

Pentylenetetrazol May Discriminate Between Different Types of Benzodiazepine Receptors

Abstract: The effect of various concentrations of pentylenetetrazol (PTZ) on [3H]flunitrazepam binding to benzodiazepine receptors was investigated by Hofstee and Hill plot analyses. These analyses indicate the presence of two PTZ binding sites in forebrain, whereas a single PTZ binding site is present in cerebellum. The relative proportions of the two PTZ binding sites in forebrain are close to those of benzodiazepine Type II and Type I receptors, respectively. These results suggest that PTZ may actually discriminate between different types of benzodiazepine receptors.

Anticonvulsant Profiles of the Potent and Orally Active GABA Uptake Inhibitors SK&F 89976‐A and SK&F 100330‐A and Four Prototype Antiepileptic Drugs in Mice and Rats

Summary: The anticonvulsant profiles of two potent and orally active 7‐aminobutyric acid (GABA) uptake inhibitors, l‐(4,4‐diphenyl‐3‐butenyl)‐3‐piperidinecarboxylic acid hydrochloride (SK&F 89976‐A) and 1‐(4,4‐diphenyl‐3‐butenyl)‐l,2,5,6‐tetrahydro‐3‐pyridine‐carboxylic acid hydrochloride (SK&F 100330‐A), were determined with a battery of well‐standardized tests in mice and rats and compared with the profiles of pheny‐toin (PHT), carbamazepine (CBZ), valproate (VPA) and clonazepam (CZP) when subjected to the same tests. ED50 values were calculated and compared with TD50 values for minimal motor impairment to provide protective indexes (PI = TD50/ED50). The anticonvulsant profiles of SK&F 89976‐A and SK&F 100330‐A were similar and suggest that these compounds raise the threshold for seizure initiation rather than inhibit seizure spread. Like intraperitoneal (i.p.) PHT, CBZ, VPA, and CZP, SK&F 89976‐A and SK&F 100330‐A inhibited seizures in corne‐ally kindled rats. The profiles of SK&F 89976‐A and SK&F 100330‐A were most similar to that of CZP and virtually opposite to that of PHT. Intraperitoneal SK&F 100330‐A provided complete protection against pentyl‐enetetrazol‐induced seizures [subcutaneous (s.c.) PTZ] in mice but was ineffective against seizures induced by maximal electroshock (MES) at doses slightly greater than its TD50. SK&F 100330‐A provided complete protection against picrotoxin‐induced seizures (s.c. Pic) and against both clonus and forelimb tonic extension induced by NMDA Af‐methyl‐D‐aspartate [intracerebralventricular (i.c.v.) NMDA] in mice; however, SK&F 100330‐A was ineffective against seizures induced by bicuculline (s.c. Bic) and strychnine (s.c. Strych) at doses slightly greater than its TD50. SK&F 89976‐A was similar but provided partial protection against NMDA‐induced clonus. SK&F(R)‐89976‐A, which is more potent than the (5> enantiomer as a GABA uptake inhibitor in vitro, was also more potent in the PTZ test. SK&F 89976‐A and SK&F 100330‐A also inhibited PTZ‐induced seizures in mice and rats after oral (p.o.) administration but were more potent in mice. Both substances were adequately absorbed after p.o. administration. Neither compound inhibited MES in mice or rats after p.o. administration. The data support the concept that SK&F 89976‐A and SK&F 100330‐A may be useful in treatment of partial seizures.

Effect of GABA agonists on the neurotoxicity and anticonvulsant activity of benzodiazepines

Progabide (50 mg/kg, i.p.), a GABA receptor agonist, significantly decreases the median minimal neurotoxic dose (TD50) of clobazam, chlordiazepoxide, and diazepam; the receptor binding of these substances is highly enhanced by muscimol. Progabide has no significant effect on the TD50 of clonazepam and triazolam; the receptor bindings of these substances is either only slightly enhanced or not altered by muscimol. Progabide also significantly decreases the median antimaximal electroshock dose (MES ED50) of all the benzodiazepines tested. However, progabide has no effect on the median antipentylenetetrazol dose (PTZ ED50) of the benzodiazepines. Likewise, THIP (2.5 mg/kg, i.p.) significantly decreases the TD50 of chlordiazepoxide but not that of triazolam. THIP significantly decreases the MES ED50 of chlordiazepoxide and triazolam but has no effect on the PTZ ED50 of these two substances. The above data suggest that benzodiazepine receptors linked to GABA receptors contribute to the minimal neurotoxicity and anti-MES activity but not to the anti-PTZ activity of benzodiazepines.

Hypnotic action of benzodiazepines: a possible mechanism.

Abstract The objective of this investigation was to determine whether the effects of muscimol on benzodiazepine receptor binding relate to the hypnotic activity of nine benzodiazepines (clonazepam, triazolam, diazepam, flurazepam, nitrazepam, oxazepam, temazepam, clobazam, and chlordiazepoxide) and CL 218, 872. There was no correlation between the basal receptor binding affinities of the drugs tested and their hypnotic potencies, whereas the benzodiazepine receptor agonists whose receptor bindings are strongly modulated by muscimol possess potent hypnotic activity. These results indicate that benzodiazepine receptors that couple to GABA receptors are involved in the hypnotic activity of the benzodiazepines.