Sugar Concentrations in Guard Cells of Vicia faba Illuminated with Red or Blue Light : Analysis by High Performance Liquid Chromatography

Abstract

Concentrations of soluble sugars in guard cells in detached, sonicated epidermis from Vicia faba leaves were analyzed quantitatively by high performance liquid chromatography to determine the extent to which sugars could contribute to changes in the osmotic potentials of guard cells during stomatal opening. Stomata were illuminated over a period of 4 hours with saturating levels of red or blue light, or a combination of red and blue light. When stomata were irradiated for 3 hours with red light (50 micromoles per square meter per second) in a solution of 5 millimolar KCl and 0.1 millimolar CaCl(2), stomatal apertures increased a net maximum of 6.7 micrometers and the concentration of total soluble sugar was 289 femtomoles per guard cell (70% sucrose, 30% fructose). In an identical solution, 2.5 hours of irradiation with 25 micromoles per square meter per second of blue light caused a maximum net increase of 7.1 micrometers in stomatal aperture and the total soluble sugar concentration was 550 femtomoles per guard cell (91% sucrose, 9% fructose). Illumination with blue light at 25 micromoles per square meter per second in a solution lacking KCl caused a maximum net increase in stomatal aperture of 3.5 micrometers and the sugar concentration was 382 femtomoles per guard cell (82% sucrose, 18% fructose). In dual beam experiments, stomata irradiated with 50 micromoles per square meter per second of red light opened steadily with a concomitant increase in sugar production. Addition of 25 micromoles per square meter per second of blue light caused a further net gain of 3.7 micrometers in stomatal aperture and, after 2 hours, sugar concentrations had increased by an additional 138 femtomoles per guard cell. Experiments with 3-(3,4-dichlorophenyl)-1,1-dimethylurea (DCMU) were performed with epidermis illuminated with 50 micromoles per square meter per second of red light or with 25 micromoles per square meter per second of blue light in solutions containing or lacking KCl. DCMU completely inhibited sugar production under red light, had no effect on guard cell sugar production under blue light when KCl was present, and inhibited sugar production by about 50% when guard cells were illuminated with blue light in solutions lacking KCl. We conclude that soluble sugars can contribute significantly to the osmoregulation of guard cells in detached leaf epidermis of V. faba. These results are consistent with the operation of two different sugar-producing pathways in guard cells: a photosynthetic carbon reduction pathway and a pathway of blue light-induced starch degradation.