Increases in cytosolic Ca2+ are not required for abscisic acid-inhibition of inward K+ currents in guard cells of Vicia faba L.
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The inward K+ channels (IKin) of guard cells are inhibited upon application of abscisic acid (ABA). It has been postulated that IKin inhibition requires an elevation in cytosolic free Ca2+ levels ([Ca2+]c) because: (i) experimental increases in [Ca2+] c can mimic the ABA effect, and; (ii) ABA can trigger an elevation of [Ca2+]c in guard cells. However, not all guard cells respond to ABA with a [Ca2+]c increase, and the magnitude of the increases that do occur is variable. Therefore, an obligate role for Ca2+ in the regulation of downstream effectors of ABA response, such as the IKin channels, remains in question. In this study, we developed a methodology for simultaneous patch clamping and confocal ratiometric Ca2+ imaging of Vicia faba L. guard-cell protoplasts. This allowed us to directly assess the relationship between ABA-induced changes in [Ca2+]c and IKin inhibition. In the presence of extracellular Ca2+, the extent of [Ca2+]c elevation correlated with the extent of IKin inhibition. However, upon chelation of either extracellular Ca2+, [Ca2+]c, or both, extracellular Ca2+ and [Ca2+]c, [Ca2+]c elevation did not occur in response to ABA yet IKin currents were still strongly inhibited. These data illustrate that Ca2+-independent regulation is involved in ABA-inhibition of stomatal opening processes.
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