Changes in apoplastic pH and membrane potential in leaves in relation to stomatal responses to CO 2 , malate, abscisic acid or interruption of water supply

Abstract

Low CO2 concentrations open CO2-sensitive stomata whereas elevated CO2 levels close them. This CO2 response is maintained in the dark. To elucidate mechanisms underlying the dark CO2 response we introduced pH- and potential-sensitive dyes into the apoplast of leaves. After mounting excised leaves in a gas-exchange chamber, changes in extracellular proton concentration and transmembrane potential differences as well as transpiration and respiration were simultaneously monitored. Upon an increase in CO2 concentration transient changes in apoplastic pH (occasionally brief acidification, but always followed by alkalinization) and in membrane potential (brief hyperpolarization followed by depolarization) accompanied stomatal closure. Alkalinization and depolarization were also observed when leaves were challenged with abscisic acid or when water flow was interrupted. During stomatal opening in response to CO2-free air the apoplastic pH increased while the membrane potential initially depolarized before it transiently hyperpolarized. To examine whether changes in apoplastic malate concentrations represent a closing signal for stomata, malate was fed into the transpiration stream. Although malate caused apoplastic alkalinization and membrane depolarization reminiscent of the effects observed with CO2 and abscisic acid, this dicarboxylate closed the stomata only partially and less effectively than CO2. Apoplastic alkalinization was also observed and stomata closed partially when KCl was fed to the leaves. Respiration increased on feeding of malate or KCl, or while abscisic acid closed the stomate. From these results we conclude that CO2 signals modulate the activity of plasma-membrane ion channels and of plasmalemma H+-ATPases during changes in stomatal aperture. Responses to potassium malate and KCl are not restricted to guard cells and neighbouring cells.

Antisense Inhibition of the Iron-Sulphur Subunit of Succinate Dehydrogenase Enhances Photosynthesis and Growth in Tomato via an Organic Acid–Mediated Effect on Stomatal Aperture

“…Furthermore, malate induces stomata closure in epidermal strips of fava bean (Vicia faba) with a half maximal concentration of 0.3 mM. In good agreement with these results, feeding malate to excised leaves reduces the transpiration rate in a dose-dependent manner (Hedrich et al, 2001). However, even at the highest concentration of malate used (40 mM), stomata still responded to CO 2 in the atmosphere, indicating that guard cells must have at least one additional CO 2 sensing system.…”

CO 2 signaling in guard cells: Calcium sensitivity response modulation, a Ca 2+ -independent phase, and CO 2 insensitivity of the gca2 mutant

“…One such event could be a change in the membrane potential. Reports have shown that decreases in [CO 2 ] cause hyperpolarization, whereas increases in [CO 2 ] cause depolarization of the guard cell plasma membrane (11,14,15,53). Low CO 2 -induced hyperpolarization would activate voltage-gated K ϩ influx channels (54) and provide the driving force for K ϩ uptake, contributing to stomatal opening (55).…”

Rapid Structural Changes and Acidification of Guard Cell Vacuoles during Stomatal Closure Require Phosphatidylinositol 3,5-Bisphosphate

“…We used leaf fragments floating on buffer solution and found that stomata closed halfway at 60 min after ABA application. In gas exchange experiments using intact leaves, stomatal conductance reached halfway 10 to 15 min after ABA application (Hedrich et al, 2001), and guard cells of intact leaf and epidermal peels were found to differ in calcium dynamics (Levchenko et al, 2008). In addition, many other factors, including pH of the extracellular space, oxygen availability, and hydrodynamics, may vary, causing the difference in the speed of the change.…”