CHANGES IN THE VOLUME OF MARGINAL CELLS INDUCED BY ISOTONIC CL- DEPLETION RESTORATION - INVOLVEMENT OF THE CL- CHANNEL AND NA+-K+-CL- COTRANSPORTER/

Marginal cells constitute the endolymph-facing epithelium responsible for the secretion of endolymph by the stria vascularis in the inner ea r. We have studied the possible involvement of Cl- conductance and Na-K+-Cl- cotransport in the mechanism of changes in cell volume upon is otonic Cl- depletion/restoration. Changes in cell volume were estimate d from video-microscopic images with the aid of an image processor. Ma rginal cells shrank to similar to 80% of their original volume in 30 s and to 65-70% in 90 s upon total replacement of [Cl](0) (similar to 1 50 mM) by gluconate(-), and the original volume of the shrunken cells was restored within 2 min after restoration of Cl-. The order of poten cy of anions to induce isotonic shrinkage was gluconate(-) > I- > F- > Br-. The cell shrinkage caused by Cl- depletion was partially inhibit ed by 5-Nitro-2-(3-phenyl-propylamino)-benzoic acid (NPPB, 0.2 mM), bu t not by either cetamido-4'-isothiocyanato-stilbene-2,2'disulfonic aci d (SITS, 0.5 mM), bumetanide (10 mu M) or ouabain (1 mM). The cell shr inkage caused by a reduction of [Cl](0) from similar to 150 mM to 7.5 mM was not affected by [K](0) in the range of 3.6 mM to 72 mM. These r esults suggest that the main efflux pathway(s) responsible for the 'Cl removal'-induced shrinkage depends on volume-correlated Cl- conductan ce (Takeuchi and Irimajiri, J. Membrane Biol. 150, 47-62, 1996) and th at this pathway(s) is essentially independent of the Na+-K+-Cl- cotran sporter, the Na+,K+-ATPase, and the K+-Cl- cotransporter. With regard to volume recovery after isotonic shrinkage, its critical dependence o n the simultaneous presence of Na+, K+ and Cl- in the bath and its sub stantial inhibition by bumetanide (10 mu M) both indicate a major role for Na+-K+-Cl- cotransport. The strong influence on cell volume of so lute fluxes working through the Cl- channel and the Na+-K+-Cl- cotrans porter implies an essential role for these pathways in the ion transpo rt mechanism(s) of the marginal cell.