Certain GABA mimetics reduce the affinity of anions required for benzodiazepine binding in a GABA-reversible way

Binding of ³H-flunitrazepam to specific sites on rat-brain membranes is anion-dependent. In the absence of added ions, binding is reduced to less than 10% of maximum, and eight anions (Tris salts) were found to increase binding in a concentration-dependent and saturable manner with affinity constants (Kd) ranging from 0.75 mM (phosphate) to 4.5 mM (chloride). The GABA mimetic THIP (200 μM) increases these Kd values from three- to 10-fold, depending on the anion. The GABA mimetics isoguvacine and piperidine-4-sulfonic acid (P4S) increase the Kd values for chloride ion eight- to 10-fold, while amino-propanesulfonate and imidazole acetate produce smaller (2- to 4-fold) increases. These effects of THIP on anion affinities are reversed in a competitive manner by GABA and muscimol. Although there is no correlation between the ability of the anions to enhance ³H-flunitrazepam binding and their ability to substitute for chloride ion electrophysiologically, it seems possible that the ions bind to sites similar to the chloride channels associated with GABA receptors. All benzodiazepine receptors are probably coupled indirectly to GABA receptors through anion binding sites.