Ca 2+ channels at the plasma membrane of stomatal guard cells are activated by hyperpolarization and abscisic acid

Abstract

In stomatal guard cells of higher-plant leaves, abscisic acid (ABA) evokes increases in cytosolic free Ca 2+ concentration ([Ca 2+ ] i ) by means of Ca 2+ entry from outside and release from intracellular stores. The mechanism(s) for Ca 2+ flux across the plasma membrane is poorly understood. Because [Ca 2+ ] i increases are voltage-sensitive, we suspected a Ca 2+ channel at the guard cell plasma membrane that activates on hyperpolarization and is regulated by ABA. We recorded single-channel currents across the Vicia guard cell plasma membrane using Ba 2+ as a charge-carrying ion. Both cell-attached and excised-patch measurements uncovered single-channel events with a maximum conductance of 12.8 ± 0.4 pS and a high selectivity for Ba 2+ (and Ca 2+ ) over K + and Cl − . Unlike other Ca 2+ channels characterized to date, these channels rectified strongly toward negative voltages with an open probability ( P o ) that increased with [Ba 2+ ] outside and decreased roughly 10-fold when [Ca 2+ ] i was raised from 200 nM to 2 μM. Adding 20 μM ABA increased P o , initially by 63- to 260-fold; in both cell-attached and excised patches, it shifted the voltage sensitivity for channel activation, and evoked damped oscillations in P o with periods near 50 s. A similar, but delayed response was observed in 0.1 μM ABA. These results identify a Ca 2+ -selective channel that can account for Ca 2+ influx and increases in [Ca 2+ ] i triggered by voltage and ABA, and they imply a close physical coupling at the plasma membrane between ABA perception and Ca 2+ channel control.