The organic anions picrate (2,4,6-trinitrophenol) and niflumate (2-[[3-(trifluoromethyl)phenyl]-amino]-3-pyridinecarboxylate) were examined for their effects on radioligand binding to the gamma-aminobutyric acid (GABA)/benzodiazepine receptor complex. Neither organic anion produced the enhancement of [35S] t-butylbicyclophosphorothionate (TBPS) binding characteristic of anions (such as Cl- and Br-) known to permeate GABA-gated chloride channels. However, both picrate and niflumate potently (IC50 values between 66 and 531 and 30 and 155 microM, respectively) inhibited the effect of 100-200 mM concentrations of anions (I-, Br-, Cl-, SCN-, and F-) to increase the binding of [35S]TBPS to GABA-gated chloride channels. This inhibition resulted from a decrease in both the maximum number of binding sites and the apparent affinity (increased Kd) of [35S]TBPS. Niflumate was consistently more potent than picrate, but both organic anions exhibited the same sequence of relative potencies against smaller anions (I- greater than Br- greater than Cl- greater than SCN- greater than F-). This sequence was similar to that described for the relative permeabilities of these anions through GABA-gated chloride channels. Niflumate and picrate were potent inhibitors of Cl-, but not GABA-modulated radioligand binding to benzodiazepine receptors. These findings suggest that picrate and niflumate bind with high affinity at or near an anion binding site that may regulate the movement of anions through GABA-gated chloride channels and radioligand binding at this "supramolecular complex."