Cation-permeable vacuolar ion channels in the moss Physcomitrella patens: a patch-clamp study

Patch-clamp studies carried out on the tonoplast of the moss Physcomitrella patens point to existence of two types of cation-selective ion channels: slowly activated (SV channels), and fast-activated potassium-selective channels. Slowly and instantaneously saturating currents were observed in the whole-vacuole recordings made in the symmetrical KCl concentration and in the presence of Ca2+ on both sides of the tonoplast. The reversal potential obtained at the KCl gradient (10 mM on the cytoplasmic side and 100 mM in the vacuole lumen) was close to the reversal potential for K+ (E (K)), indicating K+ selectivity. Recordings in cytoplasm-out patches revealed two distinct channel populations differing in conductance: 91.6 +/- A 0.9 pS (n = 14) at -80 mV and 44.7 +/- A 0.7 pS (n = 14) at +80 mV. When NaCl was used instead of KCl, clear slow vacuolar SV channel activity was observed both in whole-vacuole and cytoplasm-out membrane patches. There were no instantaneously saturating currents, which points to impermeability of fast-activated potassium channels to Na+ and K+ selectivity. In the symmetrical concentration of NaCl on both sides of the tonoplast, currents have been measured exclusively at positive voltages indicating Na+ influx to the vacuole. Recordings with different concentrations of cytoplasmic and vacuolar Ca2+ revealed that SV channel activity was regulated by both cytoplasmic and vacuolar calcium. While cytoplasmic Ca2+ activated SV channels, vacuolar Ca2+ inhibited their activity. Dependence of fast-activated potassium channels on the cytoplasmic Ca2+ was also determined. These channels were active even without Ca2+ (2 mM EGTA in the cytosol and the vacuole lumen), although their open probability significantly increased at 0.1 mu M Ca2+ on the cytoplasmic side. Apart from monovalent cations (K+ and Na+), SV channels were permeable to divalent cations (Ca2+ and Mg2+). Both monovalent and divalent cations passed through the channels in the same direction-from the cytoplasm to the vacuole. The identity of the vacuolar ion channels in Physcomitrella and ion channels already characterised in different plants is discussed.