Generalization and antagonism studies with convulsant,GABAergic and anticonvulsant drugs in rats trained to discriminate pentylenetetrazol from saline

Male hooded rats were trained on an FR 10 schedule of food reinforcement to discriminate the interoceptive stimulus produced by a subconvulsive dose of pentylenetetrazol from that produced by saline, by responding on a lever on one side of a food cup following a 20 mg/kg pentylenetetrazol injection and responding on a lever on the alternate side following a 1 ml/kg saline injection. All of the subjects learned this discrimination reliably. Picrotoxin and strychnine, but not bicuculline nor gamma-hydroxybutyrate, partially generalized to the pentylenetetrazol discriminative stimulus. Valproic acid, sodium pentobarbital and sodium phenobarbital antagonized the pentylenetetrazol discriminative stimulus but etomidate, gamma-hydroxybutyrate, gamma-acetylenic GABA, gamma-vinyl GABA, trimethadione, ethosuximide and diphenylhydantoin did not. These data suggest that the pentylenetetrazol discriminative stimulus is based upon neither a subconvulsant brain state produced by this drug nor a deficiency of GABA neuronal function. Drugs which antagonize the pentylenetetrazol stimulus are all anxiolytic drugs that show no other pharmacological property common to all of them.     

The role of bicuculline, aminooxyacetic acid and gabaculine in the modulation of ethanol-induced motor impairment

Ethanol's intoxicating effects may result from ethanol-induced changes in central gamma-aminobutyric acid (GABA) mechanisms. To further test this hypothesis, mice were pretreated with bicuculline (1 mg/kg s.c.), aminooxyacetic acid (15, 20, 25 or 30 mg/kg i.p.) or gabaculine (20 or 40 mg/kg i.p.). Following pretreatment, 20% ethanol (2.25 g/kg i.p.) was administered and rolling roller performance evaluated. All ethanol-treated control animals showed lack of rolling roller performance at 5 min post ethanol but regained rolling roller performance by 35 min. Only 42% of the bicuculline pretreated mice demonstrated lack of rolling roller performance at 5 min post ethanol and all regained rolling roller performance by 15 min. Impairment of rolling roller performance by ethanol was potentiated by aminooxyacetic acid in a dose-dependent manner. Aminooxyacetic acid (25 and 30 mg/kg doses) slowed blood ethanol disappearance although analysis of blood ethanol disappearance and motor impairment curves indicated that aminooxyacetic acid potentiation of ethanol-induced rolling roller performance impairment cannot be attributed solely to aminooxyacetic acid's effect on blood ethanol levels. Gabaculine also potentiated ethanol's impairment of rolling roller performance but was more effective than aminooxyacetic acid in slowing ethanol disappearance, suggesting that in comparison to aminooxyacetic acid, alteration of ethanol metabolism plays a greater role in gabaculine's potentiation of ethanol-induced motor impairment.     

Effects of anticonvulsant and convulsant drugs on the ATPase activities of synaptosomes and their components.

1. The effects of anticonvulsants, and other drugs on the Na+, K+-adenosine triphosphatase (ATPase) (ouabain-sensitive) and Mg++-ATPase activities of synaptosomes and their components have been determined. 2. The Mg++-ATPase activity of synaptosomes was not affected by the drugs but the Na+, K+-ATPase activity was inhibited by phenytoin (diphenylhydantoin), ethosuximide and diazepam. 3. Fractions containing mainly membranes, mitochondria or synaptic vesicles, were prepared from synaptosomes by osmotic shock and subsequent density gradient centrifugation. Inhibition of Na+, K+-ATPase activity by phenytoin, ethosuximide and diazepam was apparent only in the membrane fraction. 4. The fraction containing synaptic vesicles exhibited pronounced Md++-ATPase but no Na+, K+-ATPase activity. In contrast to the enzymes of the membranes and mitochondria, the Mg++-ATPase of the vesicles was inhibited by diazepam and all of the anticonvulsants tested.     

The effect of isoniazid and some anticonvulsant drugs on the gamma-aminobutyric acid content of mouse brain in insulin hypoglycaemia

Phenobarbitone, prominal and primidone protect mice from insulin convulsions and raise their lowered cerebral hemisphere GABA content. Phenobarbitone and primidone are superior to prominal and produce a significant increase in the insulin depleted GABA content. Isoniazid potentiates insulin convulsions and significantly lowers the insulin depleted GABA content. It is probable that insulin depletion of cerebral hemisphere GABA content is a rationale of hypolgycaemic convulsions.     

Comparison of the effects of benzodiazepines and other anticonvulsant drugs on synthesis and utilization of 5-HT in mouse brain

Acute administration of clonazepam (0.5-8.0 mg/kg, i.p.), diazepam (2-32 mg/kg, i.p.), chlordiazepoxide (1-40 mg/kg, i.p.) or diphenylhydantoin (5-320 mg/kg, i.p.), caused a dose-related elevation of the concentrations of, 5-hydroxytryptamine (5-HT), 5-hydroxyindoleacetic acid (5-HIAA) and tryptophan in whole mouse brain. Carbamazepine (5-100 mg/kg, i.p.), and phenobarbitone (10-80 mg/kg, i.p.) raised the concentrations of 5-HT and 5-HIAA in the brain, whereas flurazepam (5-80 mg/kg, i.p.) only elevated the level of 5-HIAA. After administration of L[G-3H]tryptophan (25 microCi, s.c.), clonazepam (4 mg/kg), diazepam (32 mg/kg, i.p.), chlordiazepoxide (40 mg/kg) or diphenylhydantoin (40 mg/kg), but not carbamazepine (50 mg/kg), flurazepam (40 mg/kg) or phenobarbitone (80 mg/kg), increased the content of labelled tryptophan in brain. However, administration of drugs did not alter the incorporation of the label into [3H]5-HT, suggesting that the synthesis of 5-HT was unaffected. When incorporation of [3H]tryptophan into [3H]5-HT was complete and the pool of labelled 5-HT was decreasing, clonazepam, diazepam, chlordiazepoxide and diphenylhydantoin elevated the content of [3H]5-HT in brain. Flurazepam, phenobarbitone and carbamazepine were without apparent effect. Calculation of the rate of utilization of 5-HT (Km) showed that all drugs, apart from flurazepam, reduced the utilization of 5-HT. Using the rate of disappearance of 5-HT after inhibition of tryptophan hydroxylase by p-chlorophenylalanine (PCPA), all drugs, except flurazepam, diphenylhydantoin and phenobarbitone, decreased the utilization of 5-HT. The major action of the anticonvulsant drugs on the function of 5-HT in brain appears to be a decrease in the utilization of 5-HT without altering synthesis.     

Inhibition in vitro of protein synthesis in brain subcellular fractions by the convulsant 3-mercaptopropionic acid.

The effect of the convulsant, 3-mercaptopropionic acid (MP) on [¹⁴C] leucine incorporation into proteins by brain subcellular fractions was studied. It was observed that this drug decreased protein synthesis by isolated nerve endings and mitochondria; on the other hand, microsomal fractions were not affected. The inhibition of mitochondrial protein synthesis produced by MP was not observed in the presence of an ATP-generating system. It was also found that mitochondrial ATPase was inhibited by MP. All these findings suggested that the inhibition of protein synthesis described in this paper could be an indication of an impairment of energy supply.     

The effect of baclofen and aminooxyacetic acid on the action of drugs stimulating the dopaminergic system

Baclofen and aminooxyacetic acid (AOAA) depressed the explorative and locomotor activity in rats. Baclofen, but not AOAA, decreased the locomotor stimulation, produced by apomorphine. Both compounds did not affect hyperactivity, induced by D, L-amphetamine, amantadine and methylphenidate. Neither baclofen nor AOAA influenced stereotypy induced by the above four substances.     

Short-term effects of administration of anticonvulsant drugs on free carnitine and acylcarnitine in mouse serum and tissues.

1. The short-term evolution of concentrations of free carnitine and acylcarnitine was studied in the serum, liver, kidney, heart and skeletal muscle of mice after administration of single therapeutic doses of the anticonvulsant drugs, valproic acid (VPA), carbamazepine (CBZ), phenytoin (PHT) and phenobarbitone (PHB). 2. The effects of the drugs were immediate but transitory, control levels of free carnitine and acylcarnitine having been recovered or almost recovered in serum and in all tissues 8 h post administration (p.a.). 3. VPA was the only drug that significantly reduced free carnitine concentration in serum, which recovered control levels by 4 h p.a. 4. All the drugs studied brought about marked deficits of serum acylcarnitine, which had disappeared 2 h p.a. in the case of VPA and not until 8 h p.a. for CBZ, PHT or PHB. 5. The minimum concentrations of free carnitine and acylcarnitine in serum were invariably associated with the maximum concentration of drug in serum. 6. Free carnitine concentration was not affected by VPA in any tissue, PHT and PHB brought about significant deficits in heart and kidney, and CBZ a significant deficit in muscle. 7. Acylcarnitine concentration was significantly reduced in heart, kidney and muscle by CBZ, PHT and PHB, but in liver the effects of all drugs were very small. 8. These results are compatible with the hypothesis that the primary cause of anticonvulsant-induced alteration of carnitine metabolism is interference with renal reabsorption of carnitine.     

Functional changes in cerebral 5-hydroxytryptamine metabolism in the mouse induced by anticonvulsant drugs.

1 Acute administration of clonazepam, diazepam, and diphenylhydantoin to mice elevated cerebral 5-hydroxytryptamine (5-HT) and 5-hydroxyindoleacetic acid (5-HIAA); chronic administration had less effect. 2 Acute administration of clonazepam and diazepam but not diphenylhydantoin raised cerebral trytophan levels; chronic administration of clonazepam caused a smaller elevation of cerebral tryptophan but chronic administration of diazepam still caused a large rise in cerebral tryptophan. 3 Neither clonazepam nor diazepam caused induction of drug metabolizing enzymes on chronic administration but diphenylhydantoin had a marked effect. 4 These data suggest that the altered 5-HT metabolism caused by these compounds is unrelated to a common action on tryptophan levels, and that the reduced effect of clonazepam and diazepam on chronic administration cannot be attributed to increased metabolism of these compounds. 5 Clonazepam induced abnormal head movements in mice in a dose-dependent manner. Pretreatment of animals with tranylcypromine increased the intensity of movement, although pargyline was without effect. Similar effects were observed with diazepam and diphenylhydantoin, suggesting that the increase in cerebral 5-HT caused by these compounds is of functional significance in stimulating 5-HT receptors.     

The effects of convulsant and anticonvulsant treatments on the behavioural effects of ultrasound presentation in Lister hooded rats

Lister hooded rats exhibit bursts of locomotion when exposed to a 20 kHz acoustic stimulus; this ultrasound-induced locomotion has been suggested as a potential model for panic attacks. The present studies determined the effects of treatment with the convulsant agents strychnine and pentylenetetrazole and the anticonvulsant agents pentobarbital and ethosuximide on locomotor behaviour elicited by experimenter-presented ultrasounds in Lister hooded rats. Behaviour in a circular arena was viewed live and tracked electronically. In Experiment 1, brief exposure to an ultrasound stimulus typically resulted in short intensity-related bursts of locomotion in control rats. Pentobarbital or ethosuximide treatment reduced this short-term ultrasound-induced locomotion in a dose-related manner, whereas pentylenetetrazole or strychnine treatment increased these locomotor bursts. In Experiment 2, exposure to the ultrasound stimulus for a longer period resulted in irregular cycles of bursts of locomotion followed by periods of relative inactivity in control rats. In addition, approximately 10% of control rats exhibited convulsions associated with this long-duration ultrasound exposure at 98 dB sound pressure level. Sub-convulsant doses of the convulsant treatments increased the frequency of occurrence of convulsions associated with the ultrasound stimulus; pentobarbital or ethosuximide pretreatment significantly reduced this effect. The present findings suggest that a relationship exists between ultrasound-induced locomotor bursts and convulsant activity.     

Protection from 1-cyano-3,4-epithiobutane nephrotoxicity by aminooxyacetic acid and effect on xenobiotic-metabolizing enzymes in male Fischer 344 rats.

1-Cyano-3,4-epithiobutane (CEB), a naturally occurring nitrile derived from cruciferous plants, causes nephrotoxicity and increased renal glutathione (GSH) concentration in male F-344 rats. This CEB-induced nephrotoxicity is dependent on GSH conjugation and bioactivation. The objectives of the present study were to investigate the effect of CEB on several xenobiotic-metabolizing enzymes and to evaluate the effect of modulators of GSH transport and metabolism on CEB-induced nephrotoxicity and GSH concentration. Animals received 125 mg kg-1 CEB alone or following pretreatment with one of three selective inhibitors of GSH metabolism: acivicin, probenecid or aminooxyacetic acid. There were no significant alterations in epoxide hydrolase (EH), P-450, ethoxyresorufin O-deethylase (EROD) or pentoxyresorufin O-depentylase (PROD) enzyme activity, but renal glutamyl cysteine synthetase (GCS) activity was decreased at 12 and 24 h, as was renal glutathione S-transferase 4 h after CEB administration. Renal ECOD activity was also diminished at 24 h and at 12 and 24 h in liver. Aminooxyacetic acid (AOAA) abrogated the nephrotoxicity, the renal GSH-enhancing effect, and decreased GCS of CEB alone. These findings provide further evidence for the importance of GSH conjugation as a significant pathway in CEB metabolism and the role of a reactive thiol in nephrotoxicity and altered renal GSH.     

Improved anticonvulsant activity of phenytoin by a redox brain delivery system. III: Brain uptake and pharmacological effects

Phenytoin (DPH) was delivered to the brain by a dihydropyridine in equilibrium pyridinium salt redox system, which was evaluated for anticonvulsant activity. Following iv injection of the lipophilic delivery system of DPH (2) to rats, concentrations of DPH were lower but sustained and, after 30 min, essentially the same as the levels after equimolar administration of DPH. While 2 delivered the same levels of DPH to the brain as DPH did, it was twice as potent as DPH in rats (ED50 was 7.5 mumol/kg for 2 and 14.2 mumol/kg for DPH) and mice (2: 10.5; DPH: 23.9) against maximal electroshock seizures (MES), and seven times more potent in mice (2: 10.0, DPH: 70.6) against maximal pentylenetetrazole seizures (MPS). Moreover, 2 was active against pentylenetetrazole threshold seizures (PTS) in mice and rats (ED50 = 44.1 and 40.5 mumol/kg, respectively), while DPH was ineffective (up to a dose of 79.2 mumol/kg). After evaluation of acute neurological toxicity in rats, 2 was found to possess 1.5 times higher a protective index (for MES) than DPH. It appeared also that while DPH was 2.9 times less sensitive to MPS than to MES, 2 was equally potent to both types of convulsions. Thus, the data indicate that 2 delivered DPH more efficiently to the brain. The better anticonvulsant activity (quantitatively as well as qualitatively) of 2 can be explained on the basis of an improved distribution in the brain due to its higher lipophilicity, and by favorable regional differences in the rates of conversion of 2 to DPH at the convulsing foci.     

Long-lasting anticonvulsant effects of diazepam in different mouse strains: Correlations with brain concentrations and receptor occupancy

There were marked strain differences in the duration of the protective effects of diazepam against the convulsant actions of penylenetetrazole and picrotoxin in mice. In no case was significant protection found at 12 h or longer, regardless of whether the incidence of or the latencies to myoclonus or tonic-clonic convulsions were considered. These behavioural differences could not be explained simply in terms of strain differences in benzodiazepine metabolism or in percent of receptor occupancy, as determined by the fractional displacement of ³H-flunitrazepam binding in vivo. It is suggested that there might be strain differences in the percent of receptor occupancy neeced in order to produce an anticonvulsant effect.     

Influence of ethacrynic acid on the anticonvulsant activity of conventional antiepileptic drugs in the mouse maximal electroshock seizure model

The aim of this study was to determine whether ethacrynic acid (EA), a loop diuretic with anticonvulsant activity, would affect the protective action of the conventional antiepileptics (AEDs) carbamazepine (CBZ), phenytoin (PHT), valproate (VPA) and phenobarbital (PB) in the mouse maximal electroshock seizure (MES) model. The effects of acute and chronic treatment with EA on these AEDs were examined. At a single dose of 100 mg/kg ip, EA enhanced the antielectroshock activity of VPA, decreasing its ED₅₀ value from 225.6 to 146.6 mg/kg (p < 0.05), but enhancement was not observed following continuous administration of EA (12.5 mg/kg) for seven days. Combined treatment of EA with other AEDs had no effect on their ED₅₀ values. The observed interaction between EA and VPA was pharmacodynamic in nature as EA did not alter free plasma (non-protein-bound) and total brain concentrations of VPA. Taking into consideration the clinical use of both drugs, this interaction between EA and VPA can be important for patients receiving these drugs.     

Discriminative stimulus properties of benzodiazepines, barbiturates and pharmacologically related drugs; relation to some intrinsic and anticonvulsant effects.

Using a food-reinforced two-lever operant method, rats (n = 12) were trained to discriminate chlordiazepoxide (5 mg/kg, p.o.) from solvent (p.o.). With rats trained thus as subjects, generalization experiments were done with various benzodiazepines, barbiturates and related compounds, and with two neuroleptic drugs. The ability of these drugs to induce a discriminative stimulus complex similar to that induced by chlordiazepoxide, was then compared with some intrinsic and anticonvulsant effects of the same drugs. It was found that the discriminative stimulus properties of benzodiazepines, barbiturates, and related compounds correlate with the ability of these drugs to induce ataxia, as well as with part of their anticonvulsant activity. However, the stimulus properties of these drugs, as defined by the procedure described, are based neither on their ataxia-inducing effect, nor on their general depressant or sedative action. It is concluded that these properties constitute a pharmacologically highly specific phenomenon.