Cartazolate

Abstract: Benzodiazepine receptor binding heterogeneity evident from differential affinities of some ligands was compared with that suggested by differential interactions with γ-aminobutyric acid (GABA)/bicuculline and pyrazolopyridine/barbiturate receptor sites. The GABA receptor antagonist bicuculline only partially reverses pentobarbital enhancement of [3H] diazepam binding in rat brain membranes, while totally blocking both GABA and etazolate enhancement. The degree of bicuculline sensitivity varies with brain region (cortex > hippocampus > thalamus-midbrain = striatum > medulla-pons = cerebellum) in a manner which does not correlate with over-all barbiturate enhancement nor with baseline [3H]diazepam or [3H]GABA binding; it does correlate instead with the degree of barbiturate-enhanced [3H]GABA binding, which varies similarly with brain region. The bicuculline-insensitive barbiturate enhancement of [3H]diazepam binding in cortex is blocked by low concentrations of ethyl β-carboline-3-carboxylate sufficient to inhibit primarily its high-affinity sites. [3H]Diazepam binding remaining in the presence of nanomolar β-carbolines shows a similar degree of barbiturate enhancement, which is now totally blocked by bicuculline; an augmentation of the maximal enhancement by GABA is also observed. Furthermore, the residual [3H]diazepam binding unoccupied by nanomolar β-carbolines shows an increased relative enhancement by pyrazolopyridines, which appears to reflect a selective reversal of the inhibitory portion of the biphasic dose-response curve seen with these compounds. Etazolate, cartazolate, and certain barbiturates, which also show enhancement of [3H]diazepam binding at low concentrations but reversal of this effect at high concentrations, give essentially only the enhancement effect in the presence of low concentrations of β-carbolines. The independent nature of the enhancement and inhibition phases of the biphasic dose-response curves for these compounds was confirmed by selective blockade of only the enhancement phase by the GABA chloride channel antagonist picrotoxinin and by the GABA receptor antagonist bicuculline. The distinction of β-carbolines between bicuculline-sensitive and -insensitive barbiturate enhancement and pyrazolopyridine-enhanced and -inhibited [3H]diazepam binding sites defines two classes of benzodiazepine receptors. Discrimination between these subtypes was also observed with the benzodiazepine antagonist Ro 15-1788, but not with the pyrazoloquinoline antagonist CGS 8216. Neither were the two classes of sites differentiated by depressant benzodiazepines such as diazepam and lorazepam, nor by the anxiolytic triazolopyridazine CL 218,872. Since the two subpopulations differentiated by ethyl β-carboline-3-carboxylate do not correspond in toto with the two populations showing different affinities for CL 218,872, a third class of sites is defined. The results, taken together, suggest that the heterogeneous subpopulations of [3H]diazepam binding sites defined by the interactions with bicuculline/GABA and barbiturate/pyrazolopyridine receptors cannot yet be correlated in any simple way with agonist/antagonist conformational states of the benzodiazepine receptor nor subpopulations defined by heterogeneous binding affinities for some ligands. A speculative model suggests that multiple coupling states of a single type of GABA and benzodiazepine receptor might account for at least part of the heterogeneity. Ultimate resolution will await further biochemical evidence on subtypes and physiological studies on the functional relevance of this heterogeneity observed in vitro. (Less)