Summary
Ca2+-activated K+ channels were studied in cultured medullary thick ascending limb (MTAL) cells using the patch-clamp technique in the inside-out configuration. The Ca2+ activation site was modified using N-bromoacetamide (NBA). 1mm NBA in the bath solution, at 2.5 μm Ca2+ reduces the open probability,P o , of the channel to <0.01, without an effect on single-channel conductance. NBA-modified channels are still Ca2+-sensitive, requiring 25mm Ca2+ to raiseP o to 0.2. Both before and after NBA modification channel openings display at least two distributions, indicative of more than one open state. High Ca2+ (1mm) protects the channels from modification. Also presented is a second class of Ca2+-activated K+ channels which are normally present in MTAL cells which open infrequently at 10 μm Ca2+ (P o =0.01) but have aP o of 0.08 at 1mm Ca2+. We can conclude (i) that NBA modifies the channel by shifting Ca2+-sensitivity to very high Ca2+, (ii) that NBA acts on a site involved in Ca2+ gating, and (iii) that a low affinity channel is present in the apical cell membrane with characteristics similar to those of normal channels modified with NBA.
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Cornejo, M., Guggino, S.E. & Guggino, W.B. Modification of Ca2+-activated K+ channels in cultured medullary thick ascending limb cells by N-bromoacetamide. J. Membrain Biol. 99, 147–155 (1987). https://doi.org/10.1007/BF01871234
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DOI: https://doi.org/10.1007/BF01871234