Light Saturation of Stomatal Opening on the Adaxial and Abaxial Epidermis of Commelina communis

The responses of adaxial and abaxial stomata to light were examinedon detached epidermis of Commelina communis. Stomata on theabaxial epidermis were considerably more sensitive to lightthan those on the adaxial epidermis, supporting the view thatthe differences in photosensitivity are inherent rather thanthe result of differences in the microenvironment within theleaf. The sensitivity of adaxial and abaxial stomata to bluelight was also examined. The quantum flux received by a pairof guard cells appears to be sufficient to support a directeffect of blue light on ion transport into the guard cells,but there is no evidence to suggest that blue light is essentialfor stomatal opening.     

Presence of Chloride Reduces Malate Production in Epidermis during Stomatal Opening

When stomata of isolated epidermis of Vicia faba are allowed to open in the presence of K⁺ and iminodiacetate (an impermeant zwitterion), malate is formed in the epidermis; the increases in malate content follow a nearly linear relationship with stomatal aperture. Stomata of leaf sections of V. faba floated on water during opening also exhibit this relationship. When isolated epidermis is offered KCI, this relationship is not observed and less malate is detected at comparable stomatal apertures. The data indicate that Cl⁻, if present at concentrations ≥ 10⁻⁵ eq liter⁻¹, can partially satisfy the anion requirement of guard cells of V. faba during stomatal opening. Discrepancies between earlier reports on the relative roles Cl⁻ and malate play as counterions for K⁺ in guard cells of V. faba could now be explained as resulting from variations in the availability of Cl⁻ to guard cells.     

Change in Levels of Starch and Sugars in Epidermis of Commelina benghalensis during Fusicoccin Stimulated Stomatal Opening

The concentrations of malate, starch and sugars were determinedin presonicated epidermal strips of Commelina benghalensis.During opening, the starch content of epidermis decreased whilethe level of sugars or malate increased. Fusicoccin (FC) stimulatedstomatal opening and elevated the levels of malate and sugars.However, the contribution from sugars was nearly 50% of theosmotic effect of malate and it increased to more than 60% inthe presence of FC. We conclude that FC stimulates stomatalopening by enhancing not only potassium influx into guard cellsbut also hydrolysis of starch into sugars (and malate). Significantcorrelations were noticed between the width of stomatal apertureand epidermal starch (negative), malate and sugars (both positive).The negative relationship between starch and malate or sugarswithin epidermis indicated that starch hydrolysis lead to formationof sugars as well as malate. Starch—sugar interconversioncan therefore play a significant role in modulating the solutepotential of guard cells. Key words: Commelina benghalensis, Stomatal opening, Fusicoccin, Epidermal starch and sugars     

冬小麦近轴和远轴叶面气孔对土壤水分胁迫反应的敏感性

当根层土壤水分含量不足,作物体内出现水分胁迫时,小麦叶片两面气孔的反应有明显差异。远轴叶面气孔对水分胁迫的反应比近轴叶面气孔敏感。当出现水分胁迫时,远轴叶面气孔首先收缩,且收缩的程度比近轴叶面气孔大。远轴与近轴叶面气孔阻力的比值(r_(ab)/r_(ab))与根层平均土壤水势(Ψ_s)有关,当Ψ_s大于-50 kPa时,r_(ab)/r_(ad)基本稳定在1.5左右,而当Ψ_s小于-50 kPa时,r_(ab )/r_(ab)随Ψ_s降低而明显增加。     

Abaxial and Adaxial Stomatal Responses to Light of Different Wavelengths and to Phenylacetic Acid on Isolated Epidermes of Commelina communis L.

Abaxial and adaxial stomatal responses to light of differentwavelengths and to phenylacetic acid (PAA), a molecule knownto form complexes with irradiated flavins, were examined onisolated epidermes of Commelina communis L. Blue light was superiorto red and green in promoting opening. Potassium accumulationand malate production were common to both abaxial and adaxialstomatal cells, but the photosensitivity was markedly higherin the former than in the latter. PAA suppressed opening andpotassium accumulation in guard cells, but hardly affected thelevel of epidermal malate; CO₂-free air failed to reverse thesesuppressions. The PAA-effect was more substantial in blue lightthan in red, green or darkness; thus, a flavin photoreceptoris indicated. Because of the overall effect of PAA under allconditions it is suggested that, in addition to its interactionwith blue light reception, PAA also has a more general effecton guard cells.     

Abaxial and Adaxial Stomatal Density, Stomatal Conductances and Water Status of Bean Primary Leaves as Affected by Paclobutrazol

The plant growth retardant, paclobutrazol at 8.5 or 17.0 μM concentrations effectively inhibited the stem elongation and primary leaf expansion of bean seedlings. Although the retardant reduced the relative water content in well-watered plants, the water and pressure potentials remained high in the primary leaves. K⁺, Na⁺, Mg²⁺ and Ca²⁺ contents in the primary leaves of the paclobutrazol-treated plants were not significantly different from those in the control. The stomatal density increased on both surfaces but the length of guard cells was not reduced significantly on the adaxial epidermes of the paclobutrazol-treated primary leaves. The inhibitory effect of paclobutrazol on the abaxial stomatal conductances became more pronounced with time during the light period but the adaxial surfaces displayed similar or slightly higher conductances than those of the control. The transpiration rate on a unit area basis did not change significantly or increased in the treated leaves thus the reduced water loss of paclobutrazol-treated plants was due to the reduced leaf area. Stomatal conductances of the adaxial surfaces responded more intensively to exogenous abscisic acid and the total leaf conductance decreased faster with increasing ABA concentration in the control than in the paclobutrazol-treated leaves. Paclobutrazol, an effective inhibitor of phytosterol biosynthesis, not only amplified the stomatal differentiation but increased the differences between the adaxial and abaxial stomatal conductances of the primary leaves.     

Light and Stomatal Metabolism : II. Effects of Sulfite and Arsenite on Stomatal Opening and Light Modulation of Enzymes in Epidermis

The effect of sulfite and arsenite on stomatal opening and light modulation of enzymes was examined in isolated epidermal strips of Pisum sativum L. var Little Marvel leaves. Sulfite or arsenite at 10 micromolar rapidly inhibited the stomatal opening process in light. Light activation of phosphoenolpyruvate carboxylase and NADP-malate dehydrogenase was completely diminished when the epidermal strips were incubated for 2 hours in light with either sulfite or arsenite at 10 micromolar. The data obtained suggest that the inhibition of stomatal opening by sulfite or arsenite in light might result from the inhibition of light modulation of key enzymes in guard cells.     

The Flux and Distribution of Xylem Sap Calcium to Adaxial and Abaxial Epidermal Tissue 8

Inherent differences in the responses of stomata on abaxialand adaxial epidermal surfaces of leaves of Commelina communishave previously been suggested to be due to differences in theconcentrations of apoplastic Ca. Adaxial stomata have also beenreported to be more sensitive than abaxial stomata to appliedabscisic acid (ABA). The aims of these experiments were to determinethe validity of these conclusions and to see if xylem sap Cahas a role in determining the response of stomata to ABA. It can be shown from measurements of relative stomatal resistance(determined with a viscous flow porometer) and stomatal conductancethat stomata were more open in plants grown on 8-0 mol m^–3Ca, than with those grown on 2-0 mol m^–3 Ca. When attachedleaves were fed with ABA via the transpiration stream neitherthe extent nor the rate with which conductance declined wasdependent on Ca nutrition. The extent of Ca accumulation within both epidermes was relatedto the concentration of Ca in the rhizosphere and in the xylemsap. It did not, however, appear to reflect the apparent differencesin the flux of the transpiration stream between the two epidermes.Plants growing at the lower Ca concentration accumulated proportionallymore epidermal Ca relative to Ca in xylem sap. The evidencepresented suggests that Ca movement from the xylem to the epidermiscannot be simply described by a mass flow model, and that thedistribution of Ca is not an adequate explanation of the differencesin the behaviour of adaxial and abaxial stomata. The potentialrole for changes in xylem sap Ca to act as a regulator of stomatalbehaviour are discussed. Key words: Abscisic acid, calcium, Commelina communis L., stomatal conductance     

Guard Cell Metabolism in Epidermis of Commelina communis L. During Stomatal Opening and Closing

The rates of CO² incorporation into the epidermis of C. communiswere linear and were similar during the completion of opening(2 h) and closing (1 h) movements of stomata. The kinetics of¹⁴C turnover between metabolites and the rates of ‘leakage’of metabolites were determined for opening and closing movements.When stomata were opening there was a slow turnover of ¹⁴C frommalate chiefly into sugars. Upon stomatal closure ¹⁴C was initiallymainly in sugars, malate, and sugar phosphates. Thereafter,there was a slight loss of label from sugar phosphates witha corresponding increase in malate. Starch became labelled duringopening and closing movements. Rates of incorporation of CO₂found in the ‘leakage’ fraction were greatest whenstomata were opening. Of the labelled compounds Most‘from the tissue, malate was the most highly labelled whetherstomata were opening or closing. Although interpretation of the turnover patterns is difficultwithout knowledge of pool sizes for the metabolites it is suggestedthat a pool of sugars exists within the guard cells, which havefairly direct and reversible access to carbon from starch andmalate. The implications of loss of malate from guard cellsduring stomatal opening and closing are discussed.     

Stimulation of Malate Synthesis by Glycolate in Tomato Leaves in Relation to CO2 Concentration

The mechanism by which malate synthesis from CO2 is increasedunder low concentrations of CO₂ was investigated in C₃ plants.A number of metabolites were administered to illuminated tomatoleaves, and their effects on the incorporation of ¹⁴CO₂ intomalate were determined. Compared with water as a control, glycolate,glyoxylate, D,L-glycerate, glycine, phosphoglycolate and L-serineincreased malate synthesis by factors of 6.8, 3.8, 3.3, 2.5,2.3 and 2.2, respectively. The effect of exogenous glycolateon malate synthesis from CO₂ was dependent on its concentrationup to 100 mu, but was independent of ambient CO₂ concentration.The feeding of l-¹⁴C-glycolate in the light indicated that glycolatestimulated the carbon flow from CO₂ to malate. The analysis of the products of ¹⁴CO₂ fixation in illuminatedleaves supplied with glycolate showed increases in malate andsugar and decreases in serine and phosphate esters. However,this stimulated malate synthesis ceased when malonate was suppliedsimultaneously with glycolate. Treatment with glycolate didnot affect the dark ¹⁴CO₂-fixation, but increased the ¹⁴C-malatesynthesis, with a corresponding decrease in ¹⁴C-aspartate and14^C-glutamate. These results suggest that exogenous glycolateactivates malate dehydrogenase in leaves, and that the increasedglycolate formation at low CO₂ concentrations is associatedwith the increased malate synthesis from CO₂. (Received January 12, 1981; Accepted May 20, 1981)     

Organic Acid Changes in the Epidermis of Vicia faba and Their Implication in Stomatal Movement

Considerable evidence indicates that the increase in guard cell turgor resulting in stomatal opening is brought about by active K⁺ uptake into guard cells. Only a small increase in inorganic anions appears to accompany the increase in K⁺. A plausible explanation is that organic acids are produced within guard cells and act as counterions, whereas the H⁺ produced are exchanged for K⁺.     

Stomatal Opening in Light of Different Wavelengths: Effects of Blue Light Independent of Carbon Dioxide Concentration

Stomatal opening in Xanthium pennsylvanicum was found to besignificantly greater in blue light than in red. Experimentsin which leaves were placed in a closed system and allowed toestablish their own steady-state carbon dioxide concentrationshowed that when the CO₂ concentration was about the same asthat in red, opening was much greater in blue light. Blue lightof low intensity could cause as great an opening as red of higherintensity, even though the CO₂ concentration was much higherin blue. Stomatal opening in light is considered as involvingat least two reactions: (1) a response to the removal of CO₂by photosynthesis; (2) a response to blue light not dependenton the removal of CO₂. Blue light became increasingly effective, relative to red, asthe length of night was increased over the range 2 to 14 hours.This might, in part, explain previously observed effects ofnight length on rate of opening in light. The initial very rapid phase of closure in darkness appearedto be independent of CO₂ accumulation, for it was not preventedby flushing the intercellular spaces with air free of CO₂. Itis suggested that closure in darkness, like opening in light,should be considered as involving components both dependentupon, and independent of, CO₂ concentration.     

Transport Processes and Corresponding Changes in Metabolite Levels in Relation to Starch Synthesis in Barley (Hordeum vulgare L.) Etioplasts

Intact etioplasts with an intactness of 85% and with a cytosolic and a mitochondrial contamination of less than 10% were isolated from 8-d-old dark-grown barley (Hordeum vulgare) leaves. These plastids contained starch equivalent to 21.5 μmol of glucose per mg protein. From various likely precursors applied to isolated etioplasts, only dihydroxyacetone phosphate (DHAP) had significant effects on metabolite levels and on the internal ATP/ADP ratio. The concentration dependence of DHAP uptake exhibited saturation characteristics with half saturation at 0.36 mm DHAP and a maximal velocity of 6.6 μmol mg⁻¹ of protein h⁻¹. The transport was significantly inhibited by inorganic phosphate, pyridoxal-5′-phosphate, and 4,4′-diisothiocyano-2,2′-stilbenedisulfonate. The rate of glucose-6-phosphate uptake was much lower and not saturable up to a concentration of 10 mm. Exogenously applied [¹⁴C]DHAP was incorporated into starch at a rate of 0.14 μmol of DHAP mg⁻¹ of protein h⁻¹. Enzyme activities required to convert DHAP into starch were found to be present in etioplasts. Furthermore, enzymes generating ATP from DHAP for ADPglucose synthesis were also detected. Finally, a scheme is presented suggesting DHAP uptake to serve both as carbon skeleton and as energy source for starch synthesis, mediated by a translocator with properties similar to those of the triose phosphate translocator from chloroplasts.     

Measurement of the Apoplastic Activity of K+ and Cl- in the Leaf Epidermis of Commelina communis in Relation to Stomatal Activity

Bowling, D. J. F. 1987. Measurement of the apoplastic activityof K⁺ and Cl^– in the leaf epidermis of Commelina communisin relation to stomatal activity.–J. exp. Bot. 38: 1351–1355. Ionic activity of K⁺ and Cl^– in the apoplast of the lowerepidermis of the leaf of Commelina communis was measured usingion selective micro-electrodes. Large gradients across the stomatalcomplex were observed which were related to stomatal aperture.On stomatal closure the activity of K⁺ and Cl^– in theapoplast of the guard cell rose from 3·0 mol m^–3to 100 mol m^–3 and 33 mol m^–3 respectively. It wasconcluded that the apoplast is an important pathway for iontransport between the cells. Key words: Stomata, ionic activity, leaves, apoplast     

Selection and Preparation of Leaf Epidermis for Experiments on Stomatal Physiology

A simple method for reproducibly peeling leaf epidermis tissueis described. Epidermis was obtained from eight species usingthe method, and its properties of value in research on stomatalphysiology evaluated. Effects of the method of peeling on cellviability and mesophyll contamination were quantified, and acomparison was made between the effects of peeling and subsequenttreatments on a plant with morphologically distinct subsidiarycells, Commelina communis, and one without, Vicia faba. Theresults indicate that artefacts in experiments on stomatal physiologyinvolving leaf epidermis could arise not only from the peelingmethod, but also the plant species chosen.     

Stomatal responses to light and CO2 are dependent on KCI concentration

Abstract. The responses of stomata on detached epidermis of Commelina communis to light and CO₂ have been shown to be strongly dependent on the concentration of KCI in the incubation medium. There was a high sensitivity to the two stimuli in 50 mM KCI, but there were much reduced responses at lower and higher concentrations. It is considered that an appropriate choice of medium is essential if useful physiological studies of stomata are to be made using epidermal strips. At lower KCI concentrations, the ability of the stomata to open is thought to be limited by the availability of K⁺ ions, and at higher concentrations their ability to close may be affected because of an inhibition of the net efflux of K⁺. The production of malate was related to KCI concentration, and was largest in the medium containing zero KCI which supported poor stomatal responses to light and CO₂It is concluded that malate metabolism is unlikely to play a central part in the changes in guard cell turgor that are brought about by light and CO₂.     

合意非洲铁羽片表皮及气孔器的发育研究

用光学显微镜和扫描电镜对合意非洲铁羽片表皮及气孔器的发育过程进行观察研究。结果显示,(1)在羽片发育过程中表皮细胞变化不明显,主要是细胞伸长和壁增厚的过程;表皮的角质层连续出现。(2)气孔器发育可大致分3个阶段:保卫细胞母细胞阶段、幼保卫细胞阶段和成熟保卫细胞阶段;气孔器发育方式为单唇型或周源型;成熟气孔器属单环型。(3)在羽片发育过程中,气孔密度起初很低,后急剧升高并达到峰值,之后逐渐下降并趋于稳定。(4)有早熟气孔器。(5)羽片表皮发育成熟的方式是由基部向顶部逐步推进。     

Properties of Proton Pumping in Response to Blue Light and Fusicoccin in Guard Cell Protoplasts Isolated from Adaxial Epidermis of Vicia Leaves

Guard cell protoplasts (GCPs) were isolated from the adaxial epidermis of Vicia leaves. The properties of isolated adaxial GCPs (ad GCPs) were compared with those of abaxial GCPs (ab GCPs) with respect to H+-pumping activity. A saturating pulse of blue light (200 [mu]mol m-2 s-1, 30 s) induced H+ pumping in both ad GCPs and ab GCPs under red light. The maximum rate of blue-light-dependent H+ pumping was slightly higher in ad GCPs than in ab GCPs, but the magnitude of H+ pumping in ad GCPs was 68% of that in ab GCPs. H+ pumping was responsive to the second pulse, and the rate and magnitude of the pumping increased with the time between two pulses. The periods required to achieve 50% of the maximum rate were 12 and 22 min for ad GCPs and ab GCPs, respectively. The rates of blue-light-dependent H+ pumping were saturable, with half-saturation at 630 [mu]mol m-2 (21 [mu]mol m-2 s-1, 30 s) for ad GCPs and 105 [mu]mol m-2 (3.5 [mu]mol m-2 s-1, 30s) for ab GCPs. In contrast, fusicoccin, an activator of the plasma membrane H+- ATPase, induced H+ pumping with a slightly higher rate in ad GCPs than in ab GCPs. Both types of protoplast swelled similarly in response to fusicoccin. These results suggest that ad GCPs have almost the same activity for H+ pumping as ab GCPs, whereas ad GCPs require a larger number of photons to activate the H+ pump than ab GCPs.     

Effect of Photoperiod on Stomatal Opening in Vicia faba

Stomatal apertures in darkness and subsequent average opening rates in light were measured in Vicia faba leaf discs throughout the nyctoperiods for plants grown on three light:dark cycles (8:16, 12:12, and 16:8). The time course of opening in darkness depended on the specific light:dark cycle with the maximum aperture always occurring at the time the lights normally went on. The light-induced opening rate was also maximum at the end of the nyctoperiod.     

Intra-Leaf and Intracellular Gradients in Chloroplast Ultrastructure of Dorsiventral Leaves Illuminated from the Adaxial or Abaxial Side during their Development

The number of thylakoids per granum, which is smaller in sun-typechloroplasts than in shade-type chloroplasts, was counted forthe chloroplasts at various positions within the single leavesof Spinacia oleracea L. and Glycine max (L.) Merrill. The thylakoidnumber increased with depth from the adaxial surface, but thistrend was not evident within respective cells. Therefore, photosyntheticproperties of chloroplasts should be similar within a cell butdifferent among cell layers. The similarity within a cell maybe due to the nuclear control of the chloroplast developmentand/or to chloroplast movement along the cell walls. Illumination of the leaves of G. max from the abaxial side duringtheir expansion resulted in the complete inversion of the intra-leafgradient in the thylakoid number, indicating that the formationof the intra-leaf gradient in chloroplast properties is influencedby the intra-leaf light environment during the later phase ofleaf development. ¹presennt address: Department of Environmental Biology, ResearchSchool of Biological Sciences, Australian National University,P.O. Box 475, Canberra, A.C.T. 2601, Australia. (Received March 13, 1986; Accepted May 30, 1986)