Availability of Chloride Affects the Balance between Potassium Chloride and Potassium Malate in Guard Cells of Vicia faba L.

Abstract

Electron probe microanalysis for K and Cl and enzymic determination of malate were performed on epidermal strips of Vicia faba L. which had been incubated with 0. Republic of Germany.We tested the effect of the availability of Cl-on the ionic composition of the guard cells of Viciafaba by incubating epidermal samples on solutions free of Cl-or containing Cl-and then determining the amounts of K+ and CF-absorbed as well as the amounts of malic acid synthesized. We used established methods: electron probe microanalysis for the determination of K+ and Clin the guard cells (6), and enzymic oxidation coupled to the reduction of NAD for the determination of malate in the epidermal samples (4,14).MATERIALS AND METHODS Plants. V.faba L. plants (cv. Long Pod; seeds from Lagomarsino Seeds Inc., Sacramento, Calif.) were grown for 3 to 4 weeks in a growth chamber in soil consisting of 2 parts Bacto potting soil (Michigan Peat, Houston, Texas) and I part Perlite (W. R. Grace and Co.) in pots with a soil depth of about 12 cm. The plants were watered once a week with Hoagland solution (full strength, pH 6.5), and twice a week with deionized H20. The day length was 16 hr light intensity 85 w m-2. The air temperature was 27 C during the light period and 23 C during the dark period. Relative humidity was about 85%, day and night.Preparation and Incubation of Epidermal Samples. The second, third and fourth fully expanded leaves were removed from the plants in the morning, immediately before the beginning of the light period. The leaves were washed in deionized H20. Sections measuring about 5 x 10 mm2 were cut from the intercostal areas of the leaves and floated, upper side down, on deionized H20 in the dark. Then the lower epidermis was peeled in room light (from fluorescent tubes), a process during which most of the epidermal cells are ruptured (3). The epidermal strips were rinsed in deionized H20, rubbed with a dissecting needle (bent at an angle of 1200) to remove adhering mesophyll cells, and collected on deionized H20. From there, the strips were transferred to the incubation media in small plastic beakers. The beakers were placed under an inverted crystallizing dish (150-mm diameter, 75-mm height). Humidified, C02-free air was passed through the inverted dish for 4 hr at 50 1 hr-1 and 21 C. The samples were illuminated by mercury vapor lamps (General Electric H 400 RDX 33-1); the irradiance below the water filter (5 cm deep) was 85 w m-2. The

Induction of Stomatal Closure by Vanadate or a Light/Dark Transition Involves Ca2+-Calmodulin-Dependent Protein Phosphorylations

“…Therefore, for the test of Na,VO,, the experimental solution contained the K+ salt of the impermeant anion IDA (Raschke and Schnabl, 1978) at a final Kf of 100 mM (50 mM K,IDA) instead of KCl. It was buffered at pH 5.8 with 10 mM Mes.…”

Potassium Chloride as Stomatal Osmoticum in Allium cepa L., a Species Devoid of Starch in Guard Cells

“…Iminodiacetate (J. T. Baker Chemical Co.) was used to balance K+ not balanced by CF-. Indirect evidence indicates that guard cells do not absorb iminodiacetate (16,17). The solutions were prepared with double distilled H20.…”

“…P-enolpyruvate is very likely derived from starch (4). In guard cells of Viciafaba, the balance between potassium chloride and potassium malate was determined by the availability of C1- (17). Guard cells of V. faba contain chloroplasts filled with starch, but guard cells of Allium species possess only poorly developed chloroplasts (1,10,22,23) which do not contain starch (1,2,8,22,24) or only traces ofit (11).…”

“…We set out to test this contention. We calibrated an electron probe microanalyzer for the simultaneous determination of K and Cl in epidermal cells, including guard cells (9,17). After K and Cl contents had been determined in epidermal samples, we measured the malate content of the same material.…”

Ion Transport at the Vacuole during Stomatal Movements

“…For example, Mal 22 is often listed as an important osmoticum. This is true for some species such as V. faba (Outlaw and Lowry, 1977;Outlaw and Kennedy, 1978; Raschke, 1978a, 1978b) and in certain growth conditions (Raschke and Schnabl, 1978;Van Kirk and Raschke, 1978a). But in Arabidopsis guard cells, on average, K + is charge balanced to 50% by Cl 2 and only to 5% by Mal 22 , with Mal 22 concentrations in the range of 1 to 2 mM…”