Vestibular dark cells contain the Na+/H+ exchanger NHE-1 in the basolateral membrane

Vestibular dark cells and strial marginal cells transport K+ by similar mechanisms. We have shown that K+ transport in vestibular dark cells is sensitive to the cytosolic pH (pHi) (Wangemann et al. (1995a): J. Membrane Biol. 147: 255-262). The present study addresses pharmacologically the questions whether vestibular dark cells and strial marginal cells from the gerbil contain a Na+/H+ exchanger (NHE) and in which membrane, apical or basolateral, NHE is located. Further, the study addresses the question which NHE subtype is present in vestibular dark cells. pHi was measured micro-fluorometrically with the pH-sensitive dye 2',7'-bicarboxyethyl-5(6)-carboxyfluorescein (BCECF), cell volume which is a measure of the net balance between ion influx and efflux was monitored as cell height (CH) and the equivalent short circuit current (Isc) which is a measure of transepithelial K+ secretion was calculated from measurements of the transepithelial voltage (Vt) and the transepithelial resistance (Rt). Changes in pHi were induced by 20 or 40 mM propionate. In the presence of propionate a transient acidification of pHi was observed in vestibular dark cells as well as a subsequent alkalinization to pHi values exceeding those under control conditions. The alkalinization of pHi in the presence of propionate was inhibited by the NHE blockers amiloride and EIPA. Propionate-induced swelling of vestibular dark cells was inhibited by amiloride. The NHE blocker amiloride caused in vestibular dark cells an acidification of pHi and a decrease in CH. Amiloride caused in both vestibular dark cells and strial marginal cells a transient stimulation of Isc when added to the basolateral side but not added to the apical side. Similar effects and pHi were observed in vestibular dark cells with the amiloride analog ethyl-isopropyl-amiloride (EIPA) and similar effects on Isc were observed with EIPA and the NHE blocker HOE694 when applied to the basolateral side of vestibular dark cell epithelium. The IC50 for these basolateral effects of EIPA, HOE694 and amiloride on Isc in vestibular dark cells were 2 x 10(-7) M, 8 x 10(-7) M and 4 x 10(-5) M. These observation suggest that vestibular dark cells and strial marginal cells contain NHE in their basolateral membrane, that K+ transport in strial marginal cells in pHi sensitive similar to K+ transport in vestibular dark cells and that NHE in vestibular dark cells consists of the subtype NHE-1.