Cellular proton dynamics in Elodea canadensis leaves induced by cadmium

• A new method for protoplast isolation of Elodea canadensis leaves is reported.
• Cadmium treatment of protoplasts causes an immediate increase of cytosolic pH.
• The vacuolar pH, as well as cytosolic calcium concentration increases after cadmium exposure.
• Cadmium treatment decreases the apoplastic pH of intact leaves.
• Apoplastic acidosis increases cell-wall binding of cadmium and decreases cytosolic Cd uptake.

Our earlier investigations showed that Elodea canadensis shoots, grown in the presence of cadmium (Cd), caused basification of the surrounding medium. The present study was aimed to examine the proton dynamics of the apoplastic, cytosolic and vacuolar regions of E. canadensis leaves upon Cd exposure and to establish possible linkage between cellular pH changes and the medium basification. The changes in cytosolic calcium [Ca2+]cyt was also investigated as the [Ca2+]cyt and [pH]cyt homeostasis are closely linked. The cellular H+ and Ca2+ concentrations were monitored by fluorescence microscopy and ion-specific fluorescent dyes. Cadmium concentration of leaf-cell walls was measured after plant cultivation at different fixed levels of starting pH. The protoplasts from E. canadensis leaves were isolated by use of a newly developed enzymatic method. Upon Cd addition, both cytosolic and vacuolar pH of leaf protoplasts increased with a concomitant rise in the cytosolic Ca2+ concentration. Time course studies revealed that changes in [Ca2+]cyt and [pH]cyt followed similar dynamics. Cadmium (0.5 μM) exposure decreased the apoplastic pH by 0.85 units. The maximum cell wall bound Cd-contents were obtained in plants grown at low starting pH. It is concluded that Cd treatment causes apoplastic acidosis in E. canadensis leaves associated with enhanced Cd binding to the cell walls and, consequently, reduced Cd influx into the cytosol.