Tetraethylammonium blockade of calcium-activated potassium channels in clonal anterior pituitary cells

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Abstract
The effects of extracellular and intracellular tetraethylammonium (TEA) ions on single Ca2+-activated K+ channels were studied in excised membrane patches from the anterior pituitary clone AtT-20/D16-16 with the patch-clamp technique. The presence of TEA on either surface of the membrane resulted in a decrease in the single-channel current. Dissociation constants at zero voltage for the TEA-receptor complex were calculated to be 52.2 mM and 0.08 mM for external and internal TEA, respectively. The high sensitivity of AtT-20/D16-16 celsl to internal TEA is of considerable interest, since in other preparations, the greater TEA sensitivity for Ca2+-activated K+ channels has thus far been found to occur on the external membrane surface. Hill plot analysis of the dose-response data yielded a slope of 0.92, indicating a one-to-one stoichiometry for TEA-receptor binding. The blockade by TEA showed little voltage or current sensitivity over the membrane potential range studied, and could be fully reversed by washout in drug-free solution. The results suggest the presence of TEA receptors on both the external and internal membrane surfaces but with different binding affinities. Occupancy of either site by TEA leads to an apparent decrease in the single-channel conductance of Ca2+-activated K+ channels.