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

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 mM mannitol) quickly reduced Cl− efflux (−70%) from cells preloaded with 36C1− 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 µM), which stimulated H+ extrusion to about the same extent as 200 mM mannitol, did not affect Cl− efflux. When cells preloaded with 36C1− for 18 h in the presence of mannitol (from 25 up to 200 mM) 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 mM), and was reduced (−50%) by the anion channel inhibitor A9C (300 µM). These results lead to exclude a direct causal relationship mediated by Em 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.