Conduction of Monovalent and Divalent Cations in the Slow Vacuolar Channel

Abstract.

The conduction properties of individual physiologically important cations Na⁺, K⁺, Mg²⁺, and Ca²⁺ were determined in the slowly activating (SV) channel of sugar beet vacuoles. Current-voltage relationships of the open channel were measured on excised tonoplast patches in a continuous manner by applying a ±140 mV ramp-wave protocol. Applying KCl gradients of either direction across the patch we have determined that the relative Cl⁻ to K⁺ permeability was ≤1%. Symmetrical increase of the concentration of tested cation caused an increase of the single channel conductance followed by saturation. Fitting of binding isotherms at zero voltage to the Michaelis-Menten equation resulted in values of maximal conductance of 300, 385, 18, and 13 pS, and of apparent dissociation constants of 64, 103, 0.04, and 0.08 mm for Na⁺, K⁺, Mg²⁺, and Ca²⁺, respectively. Deviations from the single-ion occupancy mechanism are documented, and alternative models of permeation are discussed. The magnitude of currents carried by divalent cations at low concentrations can be explained by an unrealistically wide (∼140 Å) radius of the pore entrance. We propose instead a fixed negative charge in the pore vestibules, which concentrates the cations in their proximity. The conduction properties of the SV channel are compared with reported characteristics of voltage-dependent Ca²⁺-permeable channels, and consequences for a possible reduction of postulated multiplicity of Ca²⁺ pathways across the tonoplast are drawn.