Extracellular Ba2+ and voltage interact to gate Ca2+ channels at the plasma membrane of stomatal guard cells.
Ca2+ channels at the plasma membrane of stomatal guard cells contribute to increases in cytosolic free [Ca2+] ([Ca2+](i)) that regulate K+ and Cl- channels for stomatal closure in higher-plant leaves. Under voltage clamp, the initial rate of increase in [Ca2+](i) in guard cells is sensitive to the extracellular divalent concentration, suggesting a close interaction between the permeant ion and channel gating. To test this idea, we recorded single-channel currents across the Vicia guard cell plasma membrane using Ba2+ as a charge carrying ion. Unlike other Ca2+ channels characterised to date, these channels activate at hyperpolarising voltages. We found that the open probability (P(o)) increased strongly with external Ba2+ concentration, consistent with a 4-fold cooperative action of Ba2+ in which its binding promoted channel opening in the steady state. Dwell time analyses indicated the presence of a single open state and at least three closed states of the channel, and showed that both hyperpolarising voltage and external Ba2+ concentration prolonged channel residence in the open state. Remarkably, increasing Ba2+ concentration also enhanced the sensitivity of the open channel to membrane voltage. We propose that Ba2+ binds at external sites distinct from the permeation pathway and that divalent binding directly influences the voltage gate.
Hamilton
D W
DW
Laboratory of Plant Physiology and Biophysics, Imperial College at Wye, Kent TN25 5AH, Wye, UK.
Hills
A
A
Blatt
M R
MR
D001464:Barium / Q000378:metabolism*; D015220:Calcium Channels / Q000502:physiology*; D002462:Cell Membrane / Q000378:metabolism / Q000502:physiology; D007887:Fabaceae / Q000378:metabolism / Q000502:physiology* / Q000648:ultrastructure; D015640:Ion Channel Gating; D018408:Patch-Clamp Techniques; D018515:Plant Leaves / Q000378:metabolism / Q000502:physiology / Q000648:ultrastructure; D010946:Plants, Medicinal; D011523:Protoplasts / Q000378:metabolism / Q000502:physiology / Q000648:ultrastructure
D015220:Calcium Channels; D001464:Barium