SELECTIVE BLOCK BY ALPHA-DENDROTOXIN OF THE K-CELL PLASMA-MEMBRANE( INWARD RECTIFIER AT THE VICIA GUARD)

The efficacy and mechanism of alpha-dendrotoxin (DTX) block of K+ chan nel currents in Vicia stomatal guard cells was examined. Currents carr ied by inward- and outward-rectifying K+ channels were determined unde r voltage clamp in intact guard cells, and block was characterized as a function of DTX and external K+ (K-o(+)) concentrations. Added to th e bath, 0.1-30 nM DTX blocked the inward-rectifying K+ current (I-K,I- in), but was ineffective in blocking current through the outward-recti fying K+ channels (I-K,I-Out) even at concentrations of 30 nM. DTX blo ck was Independent of clamp voltage and had no significant effect on t he voltage-dependent kinetics for I-K,I-in, neither altering its activ ation at voltages negative of -120 mV nor its deactivation at more pos itive voltages. No evidence was found for a use dependence to DTX acti on. Block of I-K,I-in followed a simple titration function with an app arent K-1/2 for block of 2.2 nM in 3 mM K-o(+). However, DTX block was dependent on the external K+ concentration. Raising K-o(+) from 3 to 30 mM slowed block and resulted in a 60-70% reduction in its efficacy (apparent K-i = 10 mM in 10 nM DTX). The effect of K+ in protecting I- K,I-in was competitive with DTX and specific for permeant cations. A j oint analysis of IK,in block with DTX and K+ concentration was consist ent with a single class of binding sites with a K-d for DTX of 240 pM. A K-d of 410 mu M for extracellular K+ was also indicated. These resu lts complement previous studies implicating a binding site requiring e xtracellular K+ (K-1/2 similar to 1 mM) for I-K,I-in activation; they parallel features of K+ channel block by DTX and related peptide toxin s in many animal cells, demonstrating the sensitivity of plant plasma membrane K+ channels to nanomolar toxin concentrations under physiolog ical conditions; the data also highlight one main difference: in the g uard cells, DTX action appears specific to the K+ inward rectifier.