Motor endplates of frog semitendinosus muscles were studied under voltage clamp. Current fluctuations by iontophoretic application of acetylcholine were analyzed to give the elementary conductance, γ, and mean open time, τ, of endplate channels. Total replacement of the external Na+ ion by several other metal ions and by many permanent organic cations changed both γ and τ. Except with NH4 + ions, the γ values with foreign test ions were all smaller than expected from the independence relation and their previously measured permeability ratios. The more hydrophobic ions gave the smallest γ values. Foreign permeant cations also depress γ when mixed with Na+ ions. These effects could be interpreted in terms of binding of ions to a saturable site within the endplate channel as they pass through. The site for organic ions would have a hydrophobic component. Similar evidence is given for a metal ion binding site on the cytoplasmic end of the channel accessible to internal ions. Most foreign cations also shortened τ when applied externally. The changes of gating did not seem to be correlated with changes in γ. Thus there is no evidence for control of τ by ions bound within the pore.