The role of technical, biological and pharmacological factors in the laboratory evaluation of anticonvulsant drugs. III. Pentylenetetrazole seizure models

Although seizure models using systemic administration of the chemoconvulsant pentylenetetrazol (PTZ) for induction of generalized clonic seizures in rodents are widely employed to identify potential anticonvulsants, the important role of diverse technical, biological and pharmacological factors in interpretation of results obtained with these models is often not recognized. The aim of this study was to delineate factors other than sex, age, diet, climate, and circadian rhythms, which are generally known. For this purpose, experiments with 8 clinically established antiepileptic drugs were undertaken in the following PTZ models: (1) the threshold for different types of PTZ seizures, i.e., initial myoclonic twitch, generalized clonus with loss of righting reflexes, and tonic backward extension of forelimbs (forelimb tonus), in mice; (2) the traditional PTZ seizure test with s.c. injection of the CD97 for generalized clonic seizures in mice; and (3) the s.c. PTZ seizure test in rats. In rats, in addition to evaluating drug effects on generalized clonic seizures, a ranking system was used to determine drug effects on other seizure types. When drugs were dissolved in vehicles which themselves did not exert effects on seizure susceptibility, the most important factors which influenced drug potencies were: (1) bishaped dose-response curves, i.e., a decline in anticonvulsant dose-response at high doses of some drugs, leading to misinterpretations of drug efficacy if only a single high drug dosage is tested; (2) effects of route of PTZ administration (i.v. infusion vs. s.c. injection) on estimation of anticonvulsant potency; (3) species differences in drug metabolism; (4) differences in drug potencies calculated on the basis of administered doses compared to potency calculations based on 'active' drug concentrations in plasma; (5) qualitative and quantitative species differences in drug actions; (6) endpoints used for PTZ tests; (7) misleading predictions from PTZ seizure models. Analysis of anticonvulsant drug actions indicated that myoclonic or clonic seizures induced by i.v. or s.c. PTZ might be suitable for predicting efficacy against myoclonic petit mal seizures in humans, but certainly not to predict efficacy against absence seizures. Tonic seizures induced by PTZ were blocked by drugs, such as ethosuximide, which exert no effect on tonic seizures in humans. In order to reduce the variability among estimates of anticonvulsant activity in PTZ seizure models, the various factors delineated in this study should be rigidly controlled in experimental situations involving assay of anticonvulsant agents.     

The effect of repeated seizures on anticonvulsant drug response in the kindling model

Clinical reports suggest that the occurrence of numerous seizures prior to the onset of anticonvulsant therapy is associated with a poor prognosis for drug control. This seems to imply that seizures become harder to suppress as they recur--a possibility which argues for the early initiation of anticonvulsant therapy. The present study attempted to determine whether this effect could be demonstrated under controlled laboratory conditions. The kindling preparation, an animal model involving the elicitation of repeated focal seizures, was used. Drug response (ED50) was measured in Royal Victoria hooded rats after either a small or a large number of pretreatment seizures, administered in the kindling paradigm. A variety of experiments were performed, involving: different drugs (phenobarbital, phenytoin, and carbamazepine), different seizure types (amygdala focal seizure, cortical focal seizure, generalized convulsion), and different stimulation parameters (0, 20, 40 or 100 pretreatment seizures: 'near-threshold' vs. 'standard' stimulation). In no case were seizures found to be harder to suppress following repeated pretreatment seizures. After large numbers of pretreatment seizures (40, 100), drug response was actually enhanced. These data indicate that the mere repetition of seizures does not automatically lead to a decrease in anticonvulsant effectiveness. They offer no particular rationale for the early initiation of anticonvulsant therapy in clinical situations.     

Classification criteria of epileptic seizures and syndromes

Care must be exercised not to intermingle with classification of seizures and classification of epilepsies in an inconsistent fashion. Criteria for each class must be defined as clearly as possible, and these criteria must be those which are necessary for classifying any given case. The international classification first proposed by the ILAE in 1970 was an attempt to distinguish seizures from epilepsies; the seizure types defined in the 2001 diagnostic scheme are conceptually akin to the syndromes in the 1989 classification in the sense that they imply etiological, therapeutic, and prognostic significance. However, there exists no room in the new diagnostic scheme to accommodate electro-clinical seizure types, which have been used for more than three decades. The concept of epileptic syndrome in the 1989 classifications seems to have been changed to epileptic seizure type in the 2001 diagnostic scheme, and seizure type of the latter virtually becomes synonymous with epileptic syndrome of the former. In the 2001 diagnostic scheme, seizure type can be used to supplement syndrome, and can stand alone when syndrome diagnosis cannot be made. In other words, seizure types may replace syndromes, or vice versa. We should not return to an era prior to 1970 where no distinction exists between epilepsies and seizures. In a cohort of patients with active epilepsy, to what extent is syndrome classification applicable? In 300 consecutive patients hospitalized in a tertiary center, syndromic diagnosis was applicable to only 61%. Similarly, another 100 consecutive patients, classification of epilepsy was possible but not defined as a syndrome in 32% patients, according to the 1989 classification. The 1989 syndrome classification assigned in each category "other epilepsies not defined as a syndrome." These epilepsies are diagnosed only dichotomously; idiopathic focal or generalized, symptomatic focal or generalized, or undetermined whether focal or generalized. In other words, even if we could complete a list to include all the new syndromes that may exist, it is very unlikely that it would cover all epilepsies.     

Complex partial seizures. Clinical features and differential diagnosis.

Complex partial seizures are the most common seizure type in patients with partial epilepsy. Most complex partial seizures emanate from the temporal lobe; however, the seizures also may be extratemporal in origin. The clinical phenomenology may distinguish complex partial seizures from nonepileptic paroxysmal disorders and other seizure types. Physiologic and psychological disorders need to be considered in the differential diagnosis of seizure activity. Long-term EEG monitoring may be necessary for select patients to confirm the diagnosis of epilepsy and to classify appropriately seizure type. Carbamazepine and phenytoin are the antiepileptic drugs of choice in the management of complex partial seizures. Polypharmacy and use of cognitively impairing antiepileptic drugs may reduce patient compliance and further impair the quality of life of the patient with epilepsy. Finally, epilepsy surgery is an important alternative for the patient with intractable partial seizures.     

Proof of efficacy trials: seizure types.

For proof of efficacy studies, ideally the seizure types included should also be the most common types of seizures so that an efficacy trial can recruit patients easily and in the shortest possible time frame, provide efficacy data that is valid for a large portion of the epilepsy population, include readily identifiable seizure types that are simple to count, and include seizures that occur frequently enough so that the trial can be short (3 months). Partial seizures are the most common seizure type in an adult population and therefore they represent the first choice for proof of efficacy studies, provided that data in experimental models support efficacy in this seizure type. Because there are already several antiepileptic drugs (AEDs) available which are effective for primary generalized seizure types, and patients with refractory primary generalized seizures are not so plentiful, and including these seizures as primary types to prove efficacy would be time consuming, expensive and therefore not feasible. It is concluded that, whereas proof of concept studies may include other seizure types, the first proof of efficacy studies should be undertaken in patients with frequent (>3/month) partial seizures.     

Clozapine-induced seizures: recognition and treatment.

OBJECTIVES: To inform clinicians about the types of seizures that can be induced by clozapine and to provide recommendations for treatment. METHODS: We identified articles on clozapine-induced seizures from a MEDLINE search of the English-language literature from 1978 to July 2006. The frequency of each type of seizure and the dosages of clozapine associated with seizures were compiled. In addition to this review, we report a new case illustrating the challenge of diagnosing subtle seizure activity. RESULTS: The tonic-clonic variety is the most frequently described clozapine-induced seizure. Myoclonic and atonic seizures together constitute about one-quarter of the reported seizures. The mean dosage of clozapine associated with seizures is not high (less than 600 mg daily). CONCLUSIONS: It may be difficult for clinicians to recognize subtle types of clozapine-induced seizures, such as myoclonic, atonic, or partial seizures. Clinicians should not place excessive reliance on the plasma level of clozapine or electroencephalogram findings to predict the occurrence of seizures. When a first seizure occurs, it is recommended that the dosage of clozapine be reduced or an alternative antipsychotic agent be employed. If a second seizure occurs, an anticonvulsant drug should be started. Special attention should be paid when commencing or discontinuing concurrent medication that may affect the plasma level of clozapine.     

Adult absence semiology misinterpreted as mesial temporal lobe epilepsy

Correct diagnosis of seizure type and epilepsy syndrome is the foundation for appropriate antiepileptic drug selection. Inappropriate medication choices occur in the treatment of generalized epilepsy and may aggravate some seizure types, including absence seizures, potentially leading to pseudo‐drug resistance. Fortunately, a correct diagnosis of absence seizures is usually not difficult, though rarely demonstrates electroclinical overlap with focal seizures. EEG can be especially misleading when secondary bilateral synchronous discharges occur in patients with focal seizures. However, the semiology of focal seizures associated with mesial temporal lobe epilepsy has a characteristic and consistent semiology that is the mark of this common epilepsy syndrome in adulthood. We recently encountered a 53‐year‐old female with refractory seizures and a semiology strongly suggesting mesial temporal lobe epilepsy. Instead of focal seizures, prolonged absence seizures were validated by video‐EEG monitoring and she became seizure‐free after a change to broad‐spectrum antiepileptic drugs. This case further expands our understanding of the complexity of semiology in electroclinical classification and the spectrum that may occur in adult absence seizures. It serves to underscore the need for ictal EEG recordings and the importance of concordance with the clinical course during the pre‐surgical evaluation of patients with lesions and drug‐resistant epilepsy. [Published with video sequences]     

Effect of anticonvulsant drugs on kainic acid-induced epileptiform activity

Five antiepileptic drugs were tested for their ability to block limbic seizures induced by systemic injection of kainic acid and to suppress kainic acid-induced epileptiform discharges in incubated hippocampal slices. Phenytoin, phenobarbital, ethosuximide, and valproic acid inhibited epileptiform discharges in hippocampal slices at concentrations approximating their respective clinically effective anticon-vulsant blood concentration in humans, and diazepam had a similar action at significantly higher concentrations. At these concentrations none of the drugs blocked evoked orthodromic responses of monosynaptic excitatory connections in the hippocampal slices. In contrast, none of the drugs, at therapeutic doses, prevented kainic acid-induced seizure discharges in the hippocampus, in situ. Phenobarbital and diazepam were effective at higher concentrations. These data demonstrate that antiepileptic drugs do not have identical effects on seizure discharges in one type of brain tissue in situ and in vitro even when both are elicited by the same convulsant agent. The results also indicate that limbic seizures induced by kainic acid in vivo, like many cases of complex partial seizures in humans, are highly resistant to conventional anticonvulsant drug therapy.     

Classification of seizures and epilepsy

The management of seizures and epilepsy begins with forming a differential diagnosis, making the diagnosis, and then classifying seizure type and epileptic syndrome. Classification guides treatment, including ancillary testing, management, prognosis, and if needed, selection of the appropriate antiepileptic drug (AED). Many AEDs are available, and certain seizure types or epilepsy syndromes respond to specific AEDs. The identification of the genetics, molecular basis, and pathophysiologic mechanisms of epilepsy has resulted from classification of specific epileptic syndromes. The classification system used by the International League Against Epilepsy is periodically revised. The proposed revision changes the classification emphasis from the anatomic origin of seizures (focal vs generalized) to seizure semiology (ie, the signs or clinical manifestations). Modified systems have been developed for specific circumstances (eg, neonatal seizures, infantile seizures, status epilepticus, and epilepsy surgery). This article reviews seizure and epilepsy classification, emphasizing new data.     

Utility of the Scalp-Recorded Ictal EEG in Childhood Epilepsy

PURPOSE: To evaluate the usefulness of the scalp-recorded ictal EEGs in diagnosing childhood epilepsy. METHODS: We analyzed the ictal EEGs of 259 seizures in 183 patients who visited the department of child neurology, Okayama University Medical School, during the past 6 years. RESULTS: We divided all seizures into the following four categories, according to the diagnostic usefulness of ictal EEGs in determining the seizure type: 1. (a) Ictal EEGs confirmed the diagnosis of the seizure type based on seizure symptoms (101 seizures); (b) Ictal EEGs aided in the classification of the seizure type based on the seizure symptoms (101 seizures); (c) Ictal EEGs corrected errors in the classification (37 seizures); and (d) Ictal EEGs revealed previously unreported/undocumented seizure type (20 seizures). 2. Of the 37 misdiagnosed seizures (group C), 11 were nonepileptic seizures misdiagnosed as epileptic seizures, eight were complex partial seizures (CPS) misdiagnosed as the other seizure types, and 10 were other seizure types misdiagnosed as CPSs. 3. Of the 20 previously unreported/undocumented seizures (group D), nine were myoclonic seizures, five were absence seizures, five were CPS, and one was tonic spasms. 4. Seventy-two patients had CPS. Among them, 11 patients showed no epileptic spikes in their interictal EEG recordings. Therefore, ictal recordings confirmed the diagnosis of epilepsy. CONCLUSIONS: Ictal EEG recording is a very useful diagnostic tool not only for determining seizure types, but also for uncovering the existence of the unsuspected seizure types. It supplies the physician with useful information for the classification and the treatment of epilepsy. In particular, ictal EEGs are useful in diagnosing patients with CPS.     

Increased secondary generalization of partial seizures after temporal lobectomy

OBJECTIVE: This study tests the primary hypothesis that secondary generalization of partial seizures is more likely after anterior temporal lobectomy (ATL) than before ATL, and the secondary hypothesis that antiepileptic drug withdrawal accounts for increased generalization of seizures postoperatively. BACKGROUND: The authors observed that some patients had generalized tonic-clonic (GTC) seizures after but not before ATL, by using a new classification of outcome that compares preoperative and postoperative seizure frequencies by seizure type. METHODS: Twenty patients with refractory temporal lobe epilepsy had postoperative GTC seizures or nongeneralizing complex partial (CP) seizures in a consecutive ATL series. All had reduced seizure frequency postoperatively and more than 2 years of follow-up on antiepileptic drugs. The authors calculated a generalization fraction, as (number of GTC seizures)/(number of CP and GTC seizures), for 2 years before and 2 years after surgery. RESULTS: Postoperative generalization fractions were greater than preoperative generalization fractions (Wilcoxon signed-rank test, p < 0.01). Most postoperative GTC seizures were not associated with antiepileptic drug withdrawal, and postoperative GTC seizures were not more associated with drug withdrawal than were postoperative CP seizures. Patients with more than two GTC seizures per year preoperatively were more likely than other patients to have postoperative GTC seizures. CONCLUSIONS: Patients with reduced seizure frequency after ATL have a greater tendency for partial seizures to secondarily generalize postoperatively. This phenomenon is not explained by antiepileptic drug withdrawal.     

Striking differences in proconvulsant-induced alterations of seizure threshold in two rat models

During drug development, seizure threshold tests are widely used to identify potential proconvulsant activity of investigational drugs. The most commonly used tests in this respect are the timed intravenous pentylenetetrazole (PTZ) infusion seizure test and the maximal electroshock seizure threshold (MEST) test in mice or rats. To our knowledge, no study is available in which proconvulsant drug activities in these models are directly compared, which prompted us to perform such experiments in male Wistar rats. Five drugs with reported proconvulsant activity were tested in the two models: d-amphetamine, chlorpromazine, caffeine, theophylline, and tramadol. Furthermore, the anticonvulsant drug phenobarbital was included in the experiments. While phenobarbital exerted anticonvulsant activity in both models, the five proconvulsant drugs markedly differed in their effects. In the dose range tested, d-amphetamine significantly lowered the PTZ seizure threshold but increased the MEST, caffeine and theophylline did not alter the PTZ seizure threshold but decreased the MEST, and tramadol reduced the PTZ threshold but increased the MEST. These marked differences between seizure threshold tests are most likely a consequence of the mechanisms underlying seizure induction in these tests. Our data indicate that using only one seizure threshold model during preclinical drug development may pose the risk that potential proconvulsant activity of an investigational drug is overseen. However, the label "proconvulsant" may be misleading if such activity only occurs at doses high above the therapeutic range, but the drug is not proconvulsant or even exerts anticonvulsant effects at lower, therapeutically relevant doses.     

Ictal single photon emission computed tomography in absence seizures: apparent implication of different neuronal mechanisms

Absence seizures represent a complex group of epilepsy, characterized by lapse of consciousness with staring. Bilateral, synchronous, and symmetric bursts of 3-Hz spike-and-wave discharges are observed on the electroencephalogram, whereas interictal background activity is normal. This kind of epilepsy has to be differentiated from other generalized epilepsies such as juvenile absence epilepsy and juvenile myoclonic epilepsy. Moreover, absence seizures, together with generalized spike-and-wave discharges, may coexist with other types of epilepsy such as frontal lobe epilepsy, temporal lobe epilepsy, benign epilepsy with centrotemporal spikes, and childhood epilepsy with occipital paroxysms. We have carried out ictal single photon emission computed tomography (SPECT) in 10 patients with clinical evidence of absence seizures with the aim to better understand and to distinguish this kind of seizure as primarily or secondarily generalized to a specific area and to obtain more information on the neuronal mechanisms involved in the different types of seizures, usually not identifiable at the first appearance. During the long follow-up period (9 months to 14 years), 7 of the 10 examined patients underwent interictal SPECT when they became seizure free. Our data permitted, in two patients, the diagnosis of childhood absence seizures; in three patients, they suggested the possibility of later appearance of other seizure types, on the basis of focal hyperperfusion indicating a possible focal firing. In three of the examined patients, the diagnosis of idiopathic localization-related epilepsies mimicking childhood absence seizures could be performed. In the last two patients, the hypothesis of a coexistence of absences with partial and generalized seizures was considered. From our results, it can be presumed that ictal SPECT findings may contribute to the physiopathologic classification of the different types of epilepsies. Moreover, anticonvulsant treatment more appropriate to the different forms of seizures can be used.     

Idiopathic generalized epilepsies: clinical and electroencephalogram diagnosis and treatment

This review concentrates on the principles of the clinical and electroencephalogram diagnosis of idiopathic generalized epilepsies and their treatment. The electroclinical variability of the main seizure types is detailed and particular emphasis is placed on the differential diagnosis from other seizures and nonepileptic conditions that is essential for the optimal management of these patients. The authors review the various idiopathic generalized epilepsy subsyndromes and conditions that are included in both the 1989 International League Against Epilepsy classification system and the recently proposed International League Against Epilepsy scheme, but also syndromes and forms that have not been formally recognized. Finally, the authors describe the principles of antiepileptic drug treatment with the old and newer drugs, and their specific indications and contraindications in the various syndromes and seizure types.     

Effects of carbamazepine, clonazepam, and phenytoin on seizure threshold in amygdala and cortex

This experiment was designed to determine whether or not the stronger effect of anticonvulsants on cortex than on amygdala focal seizures was due to a greater elevation of cortex seizure threshold. The effects of several doses of carbamazepine, clonazepam, and phenytoin were examined on the threshold for electrically induced afterdischarge in amygdala and cortex in 71 rats. All three drugs were found to be effective in increasing the seizure threshold with greater effects being produced in the cortex than in the amygdala. Carbamazepine produced the largest threshold increase in both foci, and clonazepam produced the weakest effects. These data are comparable to previous data on drug action against focal or partial seizures, and suggest that anticonvulsants may control partial attacks through their action on the local seizure threshold. This theory of anticonvulsant drug action adds to the common belief that carbamazepine and phenytoin act primarily by blocking seizure spread.     

Effects of Drug Withdrawal on Location of Seizure Onset

Fourteen patients with intractable epilepsy, candidates for surgical treatment, were investigated with intracerebral electrodes because of the presence of multifocal abnormalities in surface recordings. EEG and video monitoring was performed during a period of reduction and/or discontinuation of anticonvulsant medication performed to precipitate seizure occurrence. The clinical and electrical patterns of seizures recorded during the withdrawal of anticonvulsant drugs were compared to those of the patients' habitual seizures observed on full medication. For each patient, we determined a profile of the clinical and electrographic seizure activity observed before the withdrawal of medication or reported by the patient prior to hospitalization. Following the reduction or cessation of medication, an increase in partial seizure frequency was observed in all patients. All but one of these partial seizures had the same clinical pattern as the habitual attacks of the patients, and the EEG manifestations were compatible with the baseline data. Seven of the 14 patients had secondarily generalized seizures; for each patient, these seizures had the same clinical and electrographic onset as their partial seizures. Only one patient had a single partial seizure having an EEG onset different from the other seizures occurring around that time and having a clinical pattern never experienced before. In conclusion, the reduction of anticonvulsant medication extremely rarely causes the appearance of seizures having an electrical onset or a clinical pattern different from those observed on full medication.     

Clorazepate therapy for intractable epilepsy

Thirty-one epileptic patients with seizures refractory to conventional anticonvulsants were treated by adding clorazepate dipotassium to their regimen. Twelve cases showed improvement in seizure frequency, three of whom attained a seizure free state. Response to clorazepate was not related to the type of epilepsy, but patients with secondary generalized epilepsy tended to be less responsive than those with partial epilepsy. Among the various seizure types, generalized tonic-clonic seizures and simple partial seizures showed, although not significant, a tendency to be more responsive to clorazepate therapy than other seizure types, including complex partial seizures, atypical absence, atonic seizures, and tonic seizures. Drowsiness was the main adverse effect, of which 14 patients complained. Six patients were withdrawn from clorazepate because of drowsiness, but in the remaining 8 patients, this side effect disappeared within a week. The appearance of adverse effect was not related to the dose of clorazepate given. Clorazepate may be an effective secondary anticonvulsant in the treatment of intractable epilepsy.     

Patterns of seizure activation after withdrawal of antiepileptic medication

Effects of withdrawal of anticonvulsant drugs on the temporal profile of occurrence and the type of seizures were investigated in 40 intractable epileptic patients who were candidates for surgical treatment. EEG and behavior were monitored while drugs were reduced to allow localization of the epileptogenic region. The rapid withdrawal of drugs caused a rebound effect, triggering either generalized seizures during a brief period or a longer-lasting increase in partial seizures. These increases in seizure frequency appeared related to change in dosage rather than to dosage itself, since they remained largely confined to the early period following reduction of an anticonvulsant.     

Falsely localizing ictal onsets with depth EEG telemetry during anticonvulsant withdrawal

A patient with partial complex seizures evaluated for surgery with chronic depth electrode recordings demonstrated falsely localizing ictal onsets during anticonvulsant drug withdrawal. When phenytoin was being reduced, more seizures appeared to originate from the left temporal lobe than from the right. Right anterior temporal lobectomy, performed on the basis of other findings, revealed a small unsuspected tumor in the resected specimen, and the patient has remained seizure free for 3 years. The seizures that originated from the left temporal lobe were different from the patient's habitual attacks and appeared to be the result of anticonvulsant withdrawal and, perhaps, electrode irritation. Four other patients who received anterior temporal lobectomies at UCLA between 1977 and 1980 had at least one stereotaxic EEG (SEEG)-recorded contralateral seizure onset, and all have benefited from surgery. Although multifocal SEEG-recorded ictal onsets should be considered a poor prognostic sign, distant sites that give rise to atypical seizures during drug withdrawal may not generate spontaneous seizures postoperatively. Consequently, this finding should not be used as a sole criterion against the recommendation of surgical therapy.     

Is Adenosine an Endogenous Anticonvulsant?

The anticonvulsant properties of adenosine were tested pharmacologically on amygdala-kindled seizure activity in rats. The adenosine analogue 2-chloroadenosine and the adenosine uptake blocker papaverine both increased the latency to behavioral clonus as well as reduced the duration and severity of the clonic motor convulsion. Both drugs, however, failed to alter the postkindling afterdischarge (AD) threshold. Theophylline, an adenosine antagonist, had the opposite effects, prolonging the AD and motor seizure durations and facilitating partially kindled seizures, but again not altering the prekindling or postkindling AD thresholds of amygdala-elicited seizures. In contrast, carbamazepine raised AD thresholds, suggesting that it does not produce its anticonvulsant effects through adenosine systems. Since endogenous adenosine can impede seizure spread and seizure continuation, but does not affect seizure initiation from the amygdala, perhaps endogenous adenosine has the special property of being brought into play as an anticonvulsant only by the seizure itself.