WHOLE-CELL K+ CURRENT ACROSS THE PLASMA-MEMBRANE OF GUARD-CELLS FROM A GRASS - ZEA-MAYS

Knowledge of ion fluxes in the dumbbell-shaped guard cells of grass species has been limited by the difficulty of obtaining isolated epidermes or guard-cell protoplasts for use in radioactive-tracer or electrophysiological studies. We describe here a method for isolating guard-cell protoplasts from Zea mays L. Whole-cell patch clamp has been used to measure K+-channel current across the plasma membrane surrounding these protoplasts. Two populations of K+-permeable channels have been identified. Hyperpolarization of the membrane to potentials (V(m)) more negative than -100 mV results in inward K+ current through one population of channels. Inward current activation is faster than in the dicotyledon, Vicia faba L. (mean activation half-time 26 ms (Z. mays) versus 123 ms ( V. faba) at V(m) = -180 mV). Steady-state current density is less than in V. faba (-22-mu-A . cm-2 (Z. mays) versus -40-mu-A . cm-2 (V. faba) at V(m) = -180 mV in 12 mM external K+). Depolarization of the membrane to potentials more positive than -20 mV results in outward K+ current through a second population of channels; these channels activate and (upon repolarization of the membrane) deactivate more slowly than in V. faba (mean activation half-time 375 ms (Z. mays) versus 187 ms (V. faba) at V(m) = +80 mV) but result in a similar steady-state current density (23.8-mu-A . cm-2 (Z. mays) versus 28.7-mu-A . cm-2 (V. faba) at V(m) = + 80 mV with 105 mM internal K+). Omission of K+ eliminates the current. The K+ current is sensitive to both internal and external Ca2+ concentration: increasing internal Ca2+ from 2 nM to 0.2-mu-M or increasing external Ca2+ from 1 mM to 8.5 mM reduces the magnitude of both inward and outward current.