K(+)-induced stimulation of K+ secretion involves activation of the IsK channel in vestibular dark cells.
Vestibular dark cells in the inner ear secrete K+ from perilymph containing 4 mM K+ to endolymph containing 145 mM K+. Sensory transduction causes K+ to flow from endolymph to perilymph, thus threatening the homeostasis of the perilymphatic K+ concentration which is crucial for maintaining sensory transduction since the basolateral membranes of the sensory cells and adjacent neuronal elements need to be protected from K(+)-induced depolarization. The present study addresses the questions (1) whether increases in the perilymphatic K+ concentration by as little as 1 mM are sufficient to stimulate KCl uptake across the basolateral membrane of vestibular dark cells, (2) whether K(+)-induced stimulation of KCl uptake causes stimulation of the IsK channel in the apical membrane, and (3) whether the rate of transepithelial K+ secretion depends on the perilymphatic (basolateral) K+ concentration when the apical side of the epithelium is bathed with a solution containing 145 mM K+, as in vivo. Uptake of KCl was monitored by measuring cell height as an indicator for cell volume. The current (IIsK), conductance (gIsK) and inactivation time constant (tau IsK) of the IsK channel as well as the apparent reversal potential of the apical membrane (Vr) were obtained with the cell-attached macro-patch technique. Vr was corrected for the membrane voltage previously measured with microelectrodes. The rate of transepithelial K+ secretion JK was obtained as equivalent short circuit current from measurements of the transepithelial voltage (Vt) and resistance (Rt) measured in the micro-Ussing chamber. Cell height of vestibular dark cells was 7.2 microns (average). Elevations of the extracellular K+ concentration from 3.5 to 4.5 mM caused cell swelling with an initial rate of cell height change of 11 nm/s. With 3.6 mM K+ in the pipette IIsK was outwardly directed and elevation of the extracellular K+ concentration from 3.6 to 25 mM caused an increase of IIsK from 12 to 65 pA, gIsK from 152 to 950 pS and tau IsK from 278 to 583 ms as well as a hyperpolarization of Vr from -50 to -60 mV. With 150 mM K+ in the pipette IIsK was inwardly directed and the elevation of the extracellular K+ concentration caused an increase of IIsK from -1 to -143 pA, gIsK from 141 to 1833 pS and tau IsK from 248 to 729 ms. Vr remained within +/- 10 mV from zero. JK was 4.8 nmol x cm-2 x s-1 when the both the apical side and the basolateral side of the epithelium were perfused with a solution containing 3.5 mM K+. Elevation of the basolateral K+ concentration by 1 mM caused JK to increase by 1.1 nmol x cm-2 x s-1 or 23%. When the basolateral side of the epithelium was perfused with a solution containing 3.5 mM K+ and the apical side with a solution containing 145 mM K+, as in vivo, JK was 0.8 nmol x cm-2 x s-1 and elevation of the basolateral K+ concentration by 1 mM caused JK to increase by 0.8 nmol x cm-2 x s-1 or 100%. These data suggest that physiologically relevant increases in the perilymphatic K+ concentration increase JK by increasing KCl uptake across the basolateral membrane and activation of K+ release via the IsK channel in the apical membrane. Thus, the data demonstrate that vestibular dark cells adjust the rate of K+ secretion into endolymph according to the perilymphatic K+ concentration.