Early changes of Cl⁻ efflux and H⁺ extrusion induced by osmotic stress in <i>Arabidopsis thaliana</i> cells

Abstract

In various plant materials changes in turgor pressure, following hyper‐ or hypo‐osmotic stress, were associated with the activation or inactivation of the plasma membrane H + ‐ATPase, respectively. To see if the turgor changes might indirectly influence H + ‐ATPase activity by regulating ion fluxes through plasma membrane, we investigated, in cultured cells of Arabidopsis thaliana (L.) Heynh., the early effects of hyper‐ and hypo‐osmotic stress on Cl − fluxes in comparison, in the case of hyper‐osmotic treatment, with its effect on net H + extrusion. The results obtained showed that hyper‐osmotic stress (200 m M mannitol) quickly reduced Cl − efflux (−70%) from cells preloaded with 36 C1 − for 18 h. This inhibiting effect was independent of the simultaneous mannitol‐induced stimulation of Cl − influx and rapidly reversible after removal of the hyper‐osmotic treatment. The inhibition of Cl − efflux was associated with a stimulation of net H + extrusion, and these two effects showed the same dependence on the external mannitol concentration. Fusicoccin (FC, 20 µ M ), which stimulated H + extrusion to about the same extent as 200 m M mannitol, did not affect Cl − efflux. When cells preloaded with 36 C1 − for 18 h in the presence of mannitol (from 25 up to 200 m M ) were eluted in a mannitol‐free medium an early and strong increase in Cl − efflux was found. The increase of Cl‐ efflux was already detectable for a small hypo‐osmotic jump (25 m M ), and was reduced (−50%) by the anion channel inhibitor A9C (300 µM). These results lead to exclude a direct causal relationship mediated by E m changes between the effects of osmoticum on Cl − efflux and net H + extrusion, and favour the view that the changes in turgor pressure induced by hyper/hypo‐osmotic stress may respectively induce an early inactivation/activation of stretch‐sensitive anion channels.