Figure 1.

Fast-activating vacuolar currents inhibited by vacuolar Mg²⁺ in fava bean guard cells. A through C, Three representative whole-vacuole recordings are shown at Mg²⁺ concentrations of 0 mm (A), 0.5 mm (B), and 2 mm (C) in the pipette (luminal) solution. Membrane potential was stepped from −100 to +100 mV in 20-mV increments from a holding potential of 0 mV. In all traces, the vacuolar ion currents have been normalized to the whole-vacuolar capacitance (pA/pF). The solutions for FV current measurement contained 100 mm KCl, 4 mm EGTA, 10 mm HEPES-Tris, pH 7.5, in the bathing medium (cytosolic side), and 100 mm KCl, 5 mm CaCl₂, 5 mm MES-Tris, pH 5.5, with varied MgCl₂ of 0 to 2 mm in the pipette (vacuolar side). D, Average current-voltage relationships from experiments performed as in A through C at Mg²⁺ concentrations of 0, 0.1, 0.5, 1.0, and 2.0 mm in the pipette solution. FV currents were measured as the instantaneous component of whole-vacuole currents (n = 3–5 vacuoles per Mg²⁺ concentration; whole-vacuole capacitance = 9.7 ± 3.4 pF). Inset, Average percentage of inhibition of SV currents at +100 mV is plotted as a function of the concentrations of vacuolar Mg²⁺ and fitted to a Hill equation. E, Voltage dependence of vacuolar Mg²⁺ block. Average whole-vacuole currents in the presence of vacuolar Mg²⁺ as in D were normalized to currents in the absence of Mg²⁺ (Current_{0 mm}). Symbols are as in D. F, Mg²⁺ inhibition constant (K_i) plotted as a function of the applied membrane potentials. Inhibition constants at +40 to +100 mV were obtained using the Hill equation (see “Materials and Methods”).