The dissociation equilibrium constants (Kd values) of dequalinium (2) and the monoquinolinium compounds 1a and 1b have been determined from competition equilibrium radioligand binding with [125I]apamin on rat brain synaptic plasma membranes (SPMs). Dequalinium binds to the channel with 2 orders of magnitude higher affinity than 1a or 1b, suggesting that both quinolinium groups are needed for potent and selective SKCa channel blockade. The trisquinolinium compound 3 (1,1'-[5-[4-(4- aminoquinolinium-1-yl)but-1-yl]non-4-en-1,9-diyl]-bis-(4- aminoquinolinium)) has been synthesized and tested for inhibition of the afterhyperpolarization of rat sympathetic neurones and on the binding assay. Compound 3 shows approximately one order of magnitude higher potency than 2, being the most potent non-peptidic SKCa channel blocker reported so far (Kd approximately 30 nM). The higher affinity of 3 compared with 2 may be due to direct binding of the third quinolinium group to the channel or may arise from a reduction of the unfavorable entropy of binding via an increase of the "local concentration" of quinolinium groups.