The specificity of bthe Na+ channel for monovalent cations has been analyzed in three different ways using neutrotoxins and excitable cells in culture. Influx experiments using radioactive monovalent cations have shown that all neurotoxins or combination of neurotoxins which change the gating system of the Na+ channel (veratridine, batrachotoxin, dihydrograyanotoxin, scorpion and sea‐anemone toxins) induce a considerable alteration of the permeability of the channel for monovalent cations. Theer is a much decreased ionic selectivity of the channel. Toxin‐induced changes in ionic selectivity are the same or very similar in neuroblastoma cells and in skeletal myotubes. Thefact that toxins which change the kinetics of the gating system also change channel selectivity indicates that the selectivity filter and the gating system may be coupled entities. Efflux experiments using neuroblastoma cells preloaded with 22N+, 42K+ or 86Rb+ indicate that the toxin stimulated release of these monovalent cations through the Na+ channel is not seen in the presence of cholinium in the external medium. It is seen only if the external medium contains permeant monovalent cations. The ‘chemical’ activation of the Na+ channel by neurotoxins necessitates the saturation at the external face of the channel of sites specific for permeant monovalent cations. Dose‐response curves for the effect of increasing external concentrations of Na+, K+, Rb+ or guanidinium on 22Na+ efflux indicate positive cooperativity in the stimulating effect of these cations. The best apparent affinity is observed for guanidinium, then for Na+, K+ and Rb+. Competition experiments between monovalent cations and a radiolabelled tritiated derivative of tetrodotoxin for binding to the tetrodotoxin receptor of the Na+ channel in neuroblastoma cells indicatae that monovalent cations bind cooperatively to external sites for Na+. The Hill coefficients and the dissociation constants for Na+ and guanidinium binding to the Na+ channel measured with this approach compare well to values found from 22Na+efflux experiments.