Kinetics of Ca2+- and ATP-dependent, voltage-controlled anion conductance in the plasma membrane of mesophyll cells of Pisum sativum

Whole-cell patch-clamp techniques were used to measure anion currents through the plasma membrane of protoplasts of mesophyll cells of expanding pea (Pisum sativum L.) leaves. Voltage-induced changes of the currents could be modelled with single exponential activation and deactivation kinetics. The anion currents were activated at negative membrane potentials. The time constant of activation, tau(act), increased from 145 ms at -140 mV to 380 ms at -20 mV. A Boltzmann fit to the activation curve, n infinity (Delta G(Vm)/Delta G(max)), yielded a half-activation voltage of +27 mV. Opening and closing rate constants, alpha and beta respectively, were calculated from the values of tau and n(infinity). The currents depended on the presence of cytoplasmic Ca2+ concentrations higher than 10(-6) M. Including 3 mM MgATP in the intracellular solution resulted in a voltage-dependent inactivation of the anion current. The conductance-voltage relation resulting from the voltage-dependent activation and inactivation had a maximum at about -25 mV. The relations of the current in pea are discussed with respect to the anion currents in guard cells and suspension-cultured tobacco cells, and its possible role in growing leaf cells

Characterization of a Light-Controlled Anion Channel in the Plasma Membrane of Mesophyll Cells of Pea.

In leaf mesophyll cells of pea (Pisum sativum) light induces a transient depolarization that is at least partly due to an increased plasma membrane conductance for anions. Several channel types were identified in the plasma membrane of protoplasts from mesophyll cells using the patch-clamp technique. One of these was an anion channel with a single-channel conductance of 32 picasiemens in symmetrical 100/100 KCl solutions. In asymmetrical solutions the reversal potential indicates a high selectivity for Cl- over K+ at high cytoplasmic Cl-. At negative membrane voltages the channel openings were interrupted by very short closures. In the open channel conductance several substrates were identified. At a cytoplasmic negative logarithm of Ca concentration higher than 6.3, no channel openings were observed. When the protoplast was illuminated in the cell-attached configuration, at least one channel type had a higher opening probability. This channel can tentatively be identified as the above-described anion channel based on conductance and the characteristic short closures at negative membrane potentials. This light activation of the 32-picasiemen anion channel is a strong indication that this channel conducts the light-induced depolarizing current. Because channel activity is strongly Ca2+-dependent, a role of cytoplasmic Ca2+ concentration changes in the light activation of the conductance is discussed