Hypo-osmotic challenge stimulates transepithelial K+ secretion and activates apical IsK channel in vestibular dark cells.

Volume regulation of vestibular dark cells from the gerbilline inner ear in response to a hypo-osmotic challenge depends on the presence of cytosolic K+ and Cl-. The present study addresses the questions: (i) whether and by what mechanism K+ is released during volume regulation, (ii) whether the osmolarity of the basolateral medium has an effect on the steady-state rate of transepithelial K+ transport and (iii) whether there is cross-talk between the basolateral membrane responsible for K+ uptake and the apical membrane responsible for K+ release. K+ secretion (JK+,probe) and current density (Isc,probe) were measured with vibrating probes in the vicinity of the apical membrane and the transepithelial potential (Vt) and resistance (Rt) were measured in a micro-Ussing chamber. The equivalent short-circuit current (Isc) was calculated. The current (IIsK), conductance (gIsK) and inactivation time constant (tau IsK) of the IsK channel and the apparent reversal potential of the apical membrane (Vr) were obtained with the cell-attached macropatch technique. Vr was corrected (Vrc) for the membrane voltage (Vm) measured separately with microelectrodes. A hypo-osmotic challenge (294 to 154 mosM by removal of 150 mM mannitol) on the basolateral side of the epithelium increased JK+,probe and Isc,probe by a factor of 2.7 and 1.6. When this hypo-osmotic challenge was applied to both sides of the epithelium Vt and Isc increased from 5 to 14 mV and from 189 to 824 microA/cm2 whereas Rt decreased from 27 to 19 omega-cm2. With 3.6 mM K+ in the pipette IIsK was outwardly directed, tau IsK was 267 msec and the hypo-osmotic challenge caused IIsK and gIsK to increase from 14 to 37 pA and from 292 to 732 pS. Vrc hyperpolarized from -44 to -76 mV. With 150 mM K+ in the pipette IIsK was inwardly directed, tau IsK was 208 msec and the hypo-osmotic challenge caused IIsK and gIsK to increase in magnitude from 0 to -21 pA and from 107 to 1101 pS. Vrc remained unchanged (-2 vs. 1 mV). These data demonstrate that a hypo-osmotic challenge stimulates transepithelial K+ secretion and activates the apical IsK channel. The hypo-osmotically-induced increase in K+ secretion exceeded the estimated amount of K+ release necessary for the maintenance of constant cell volume, suggesting that the rate of basolateral K+ uptake was upregulated in the presence of the hypo-osmotic challenge and that cross-talk exists between the apical membrane and the basolateral membrane.