Andrew Y. Chweh

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.

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.

Correlation of the hypnotic potency of benzodiazepines with inhibition of voltage-dependent calcium uptake into mouse brain synaptosomes

Nine benzodiazepines were tested for their ability to inhibit 45Ca2+ uptake into mouse whole brain synaptosomes and for hypnotic activity as indicated by their ability to produce loss of the righting reflex. Eight of the benzodiazepines significantly inhibited fast-phase voltage-dependent 45Ca2+ uptake and five exhibited hypnotic activity. There was a direct correlation between the hypnotic potency of these five benzodiazepines and their ability to inhibit 45Ca2+ uptake. There does not appear to be a correlation between the anticonvulsant potency of the benzodiazepines and their potency for inhibiting 45Ca2+ uptake. These results support previous findings with other sedative/hypnotic drugs and suggest that inhibition of presynaptic calcium uptake may be linked with hypnotic but not anticonvulsant actions of benzodiazepines.

Hypnotic action of pentobarbital in mice: A possible mechanism

The effect of GABA receptor agonists (THIP and baclofen) on the hypnotic activity (loss of righting reflex and sleep time) of pentobarbital in mice was investigated. Combinations of either THIP-pentobarbital or baclofen-pentobarbital interacted synergistically by increasing hypnotic activity. The GABAa receptor antagonist, bicuculline, decreased the hypnotic effect of THIP-pentobarbital combinations but not that of baclofen-pentobarbital combinations. These results suggest that the hypnotic activity of pentobarbital involves GABAa receptor function.

Involvement of a GABAergic mechanism in the pharmacologic action of phenytoin

In vivo interactions between phenytoin (PHT) and baclofen (a GABAb receptor agonist) or PHT and progabide (a GABAa receptor agonist) were investigated by the rotorod minimal neurotoxicity test and maximal electroshock seizure (MES) test. The combination of PHT and baclofen produced an additive effect by the rotorod test, whereas the combination of PHT and progabide elicited a supra-additive (synergistic) effect. The median minimal neurotoxic dose of baclofen augmented the anti-MES activity of PHT. The combination of PHT and progabide induced a supra-additive effect by the MES test. These results imply that GABAa receptors are involved in both the minimal neurotoxicity and anti-MES activity of PHT.

γ-Aminobutyric acid modulation of benzodiazepine receptor binding in vitro does not predict the pharmacologic activity of all benzodiazepine receptor ligands

gamma-Aminobutyric acid (GABA) modulation of triazolam and nicotinamide binding to benzodiazepine (BDZ) receptors in vitro was compared with the neurotoxicity and anticonvulsant activity of these two drugs in vivo. GABA had no significant effect on the inhibitory potency of triazolam in [3H]flunitrazepam receptor binding, whereas GABA decreased the inhibitory potency of nicotinamide. When administered to mice, both triazolam and nicotinamide exhibited neurotoxicity by the rotorod test and anticonvulsant activity by the pentylenetetrazol seizure threshold test. This suggests that GABA modulation of the receptor binding of a BDZ ligand in vitro is not a reliable predictor of the pharmacologic activity of the ligand.

γ-Aminobutyric Acid: Temperature Dependence in Benzodiazepine Receptor Binding

Abstract: The effects of muscimol and/or incubation temperature on the inhibition of [3H]flunitrazepam receptor binding by benzodiazepine receptor ligands were investigated. At 0°C muscimol decreased the Ki values for some ligands as displacers of [3H]flunitrazepam binding to brain‐specific sites while increasing or having no effect on the Ki values for other ligands. The Ki values for some ligands are higher at 37°C than at 0°C but are reduced by muscimol at both 0° and 37°C. In contrast, the ligands whose Kt values are increased by muscimol either decreased or did not alter the Ki values at 37°C as compared to those at 0°C. Incubation of membranes at 37°C for 30 min accelerated γ‐aminobutyric acid (GABA) release by 221% over that at 0°C. These results indicate that changes in incubation temperature alter benzodiazepine receptor affinity for ligands via GABA.

Benzodiazepine inhibition of [3H]flunitrazepam binding and caffeine-induced seizures in mice

The median inhibitory and anticonvulsant potencies of seven benzodiazepine (BDZ) agonists and one BDZ antagonist (Ro15-1788) were established by two tests: inhibition of [3H]flunitrazepam receptor binding and prevention of caffeine-induced seizures in mice. The effect of Ro15-1788 on the anticonvulsant potency of diazepam against caffeine-induced seizures was also investigated. The [3H]flunitrazepam receptor binding inhibitory potencies (IC50s) of the BDZ agonists correlate well with their anticonvulsant potencies (ED50s) against caffeine-induced seizures (r = 0.831; P greater than 0.01 and less than 0.05). Ro15-1788 and clonazepam are the most potent inhibitors (IC50s: 1.72 and 1.75 nM, respectively), but differ markedly in their ability to obtund caffeine-induced seizures (ED50s: 43.2 and 0.226 mg/kg, respectively). Although both Ro15-1788 and diazepam are effective against caffeine-induced seizures, when used in combination Ro15-1788 antagonizes the anti-caffeine effect of diazepam. These data indicate that Ro15-1788 is a BDZ partial agonist with low efficacy as well as a potent antagonist.

Effect of picrotoxinin on benzodiazepine receptor binding.

The effect of picrotoxinin on [3H]flunitrazepam binding to benzodiazepine receptors was investigated. In mouse forebrain membranes, picrotoxinin inhibited basal, GABA- and pentobarbital-stimulated [3H]flunitrazepam binding; this inhibitory activity was temperature- and chloride ion-dependent. Scatchard analysis of the data indicates that picrotoxinin decreases the number of binding sites without alterating binding affinity. In cerebellar membranes, picrotoxinin did not alter [3H]flunitrazepam receptor binding.