DIVALENT-CATIONS INHIBIT ISK KVLQT1 CHANNELS IN EXCISED MEMBRANE PATCHES OF STRIAL MARGINAL CELLS/

The IsK/KvLQT1 K+ channel in the apical membrane of strial marginal ce lls and vestibular dark cells is an essential ion transport pathway fo r the secretion of K+ into the endolymph of the inner ear. Study of th is control point has been impeded by rundown of channel activity upon excision into commonly used cytosolic solutions. This paper describes conditions under which patches of apical membrane of strial marginal c ells and vestibular dark cells from gerbil containing this channel can be excised, retaining its characteristic voltage dependence, kinetic properties, ion permeability sequence and pharmacological sensitivity, similar to those found during on-cell and perforated-patch whole cell recordings (Shen et al., Audit. Neurosci. 3 (1997) 215-230). Those ex cised-patch conditions include removal of Mg2+ from the cytosolic solu tion and use of a K+-rich pipette electrolyte. The inhibition of chann el activity by Mg2+ was found to be a general feature of divalent cati ons; the channel was also inhibited by Ca2+, Ba2+ and Sr2+. The concen trations causing 50% inhibition of IsK/KvLQT1 channel current were 7x1 0(-5) M, 6x10(-6) M, 3x10(-4) M and 7x10(-5) M, respectively. It was a lso found that a chemical cross-linking agent, 3,3'-dithio-bis(sulfosu ccinimidyl propionate) (DTSSP), which was previously shown to persiste ntly activate IsK/KvLQT1 channels expressed in Xenopus oocytes, mainta ined in excised patches channel activity which retained voltage depend ence and pharmacological sensitivity. These data demonstrate that (1) the channel complex is inhibited by Ca2+, Mg2+ and other divalent cati ons, (2) the activation by Ca2+ observed previously in whole-cell prep arations was due to action via other cellular pathways. These findings must be taken into account when considering the action of receptors w hich alter the cytosolic Ca2+ activity. (C) 1998 Elsevier Science B.V. All rights reserved.