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Under NaCl-dominated salt stress, the key to plant survival is maintaining a low cytosolic Na(+) level or Na(+)/K(+) ratio. The OsHKT1, OsHKT2, and OsVHA transporter genes might play important roles in maintaining cytosolic Na(+) homeostasis in rice (Oryza sativa L. indica cvs Pokkali and BRRI Dhan29). Upon NaCl stress, the OsHKT1 transcript was significantly down-regulated in salt-tolerant cv. Pokkali, but not in salt-sensitive cv. BRRI Dhan29. NaCl stress induced the expression of OsHKT2 and OsVHA in both Pokkali and BRRI Dhan29. In cv. Pokkali, OsHKT2 and OsVHA transcripts were induced immediately after NaCl stress. However, in cv. BRRI Dhan29, the induction of OsHKT2 was quite low and of OsVHA was low and delayed, compared with that in cv. Pokkali. OsHKT2 and OsVHA induction mostly occurred in the phloem, in the transition from phloem to mesophyll cells, and in the mesophyll cells of the leaves. The vacuolar area in cv. Pokkali did not change under either short- (5-10 min) or long-term (24 h) salt stress, although it significantly increased 24 h after the stress in cv. BRRI Dhan29. When expressional constructs of VHA-c and VHA-a with YFP and CFP were introduced into isolated protoplasts of cvs Pokkali and BRRI Dhan29, the fluorescence resonance energy transfer (FRET) efficiency between VHA-c and VHA-a upon salt stress decreased slightly in cv. Pokkali, but increased significantly in cv. BRRI Dhan29. The results suggest that the salt-tolerant cv. Pokkali regulates the expression of OsHKT1, OsHKT2, and OsVHA differently from how the salt-sensitive cv. BRRI Dhan29 does. Together, these proteins might confer salt tolerance in Pokkali by maintaining a low cytosolic Na(+) level and a correct ratio of cytosolic Na(+)/K(+).

In this study, the uptake of Na+ into the cytosol of rice (Oryza sativa L. cvs Pokkali and BRRI Dhan29) protoplasts was measured using the acetoxy methyl ester of the fluorescent sodium-binding benzofuran isopthalate, SBFI-AM, and fluorescence microscopy. By means of inhibitor analyses the mechanisms for uptake and sequestration of Na+ in the salt-sensitive indica rice cv. BRRI Dhan29 and in the salt-tolerant indica rice cv. Pokkali were detected. Less Na+ was taken up into the cytosol of Pokkali than into BRRI Dhan29. The results indicate that K+-selective channels do not contribute to the Na+ uptake in Pokkali, whereas they are the major pathways for Na+ uptake in BRRI Dhan29 along with non-selective cation channels. However, non-selective cation channels seem to be the main pathways for Na+ uptake in Pokkali. Protoplasts from Pokkali leaves took up Na+ only transiently in the presence of extracellular Na+ at 5-100 mM. Therefore, it is likely that the protoplasts have a mechanism for fast extrusion of Na+ out of the cytoplasm. Experiments with protoplasts pretreated with NH4NO3 and NH4VO3 suggest that the salt-tolerant Pokkali extrudes Na+ mainly into the vacuole. After cultivation of both cultivars in the presence of 10 or 50 mM NaCl for 72 h, the isolated protoplasts from Pokkali took up less Na+ than the control protoplasts. The results suggest that the salt-tolerance in Pokkali depends on reduced uptake through K+-selective channels and a fast extrusion of Na+ into the vacuoles.

Addition of aluminium chloride (50 μM Al) caused different effects on the transmembrane electrical potential (PD) of root cells in Al‐tolerant wheat (Triticum aestivum) cv. Kadett and Al‐sensitive cv. WW 20299. As changes in PD of plant cells may depend on transient fluxes of protons, potassium and/or calcium through cell membranes, the effect of Al was investigated on the cytosolic concentrations of these ions in protoplasts isolated from root tips of the same cultivars. The tetra[acetoxymethyl] esters of the fluorescent dyes bis‐carboxyethyl‐carboxyfluorescein, BCECF, K+‐binding benzofuran isophthalate, PBFI, and the stilbene chromophore Fura 2‐AM were used to determine pH, K+ and Ca2+, respectively. Changes in fluorescence ratios, directly reflecting changes in [H+], [K+] and [Ca2+] in the cytosol, were determined by photometry fluorescence microscopy. Additions and removals of Al to and from both cultivars caused hyperpolarizations and depolarizations, respectively, but only in the sensitive cv. WW 20299 did the resting PD decrease gradually. Addition of Al to the protoplasts caused rapid changes in cytosolic pH, free [K+] and [Ca2+]. In both cultivars Al caused a transient oscillating increase in cytosolic [Ca2+] for 1 or 2 min and a rapid pH‐dependent change in cytosolic [K+]. At pH 5 the presence of K+ in the medium diminished the Al‐induced decrease in cytosolic [K+]. Aluminium (50 μM) induced a transient increase in cytosolic [H+] (pH decreased) in both cultivars, but the cytosolic pH returned to its initial value only in the Al‐tolerant cv. Kadett. In the Alsensitive cv. WW 20299, repeated additions of Al caused a gradual decline in pH. Moreover, in the presence of 1 mM KCl, pH recovered completely in both cultivars. Since only the effect on pH differed in the two cultivars, the more toxic effect of Al on the cv. WW 20299 should be related to the change in pH.

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