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The patch clamp measurements carried out on the patch of the cell membrane showed that amphotericin B (AmB) applied at a low concentration (0.05 and 0.1 µM) causes a decrease in seal resistance, indicating loss of membrane tightness. Application of AmB also decreased the number of active outward rectifying potassium channels TOK1, but did not change the unitary conductance of the channels. Interestingly, AmB did not significantly change the open probability of the single channel. Such results indicate a lack of direct interaction of AmB and the channels. Membrane permeability changes after AmB were also observed by fluorescent microscopy. This research allowed us to conclude that integrity of the membrane between the pores formed by AmB is maintained. Supported by NCN grant DEC-2012/05/BNZ1/00037
Liverworts are pioneer plants that colonized lands. They had to cope with frequent sea water flooding causing salt stress. The role of vacuoles and in particular slow-activating (SV) channels in the salt stress tolerance was addressed in the present study. A patch-clamp method was used to study sodium fluxes through the tonoplast of the liverwort Conocephalum conicum. The whole-vacuole measurements carried out in a symmetrical Na⁺ concentration allowed recording of slowly activated outward currents typical for SV channels. In a Na⁺ gradient promoting an efflux of Na⁺ from the vacuole, the outward rectifying properties of SV channels were reduced and inward Na⁺ currents with different inactivation rates were recorded. Single channel analysis proved that a decrease in cytoplasmic Na⁺ concentration evoked an increase in the open probability of the channels and shifted the activation voltages towards negative values. The number of SV channels recorded at negative voltages was dependent on the vacuolar calcium and decreased at the high concentration of this ion in the vacuole. In some of the tested patches, the channels exhibited a flickering type of activity and two different conductance levels. The role of SV channels in Na⁺ accumulation during salt stress and its removal after periods of flooding is discussed in the present paper.
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