The Selection of an Infiltration Technique for Estimating the Degree of Stomatal Opening in Leaves of Field Crops in the Sudan and a Discussion of the Mechanism which Controls the Entry of Test Liquids

Estimation of the degree of stomatal opening by infiltrationof liquids provides a method whereby the crop plant may be usedas an indicator for timing of irrigation in field experimentswhere a simple routine procedure is Deeded and, as in some areasin the Sudan, fixed irrigation schedules or water balance estimatesare not feasible. Infiltration techniques used by previous workers,attempts by some of them at calibration by direct microscopicexamination of stomata and by stomatal resistance to air flowand investigations of the infiltration pattern in relation tochanges in environmental factors, such as are found in the diurnalcycle and the irrigation cycle, are discussed. Some techniques were considered unsuitable: the infiltrationof large areas of leaf becauseof difficulties in grading, thetiming of infiltration because of difficulty in determiningthe end point and in distinguishing between entry through stomataand subsequent spread through intercellular spaces and becauseof possible effects of the liquid on the stomata. The techniqueselected employs drops of graded mixtures of liquids allowingan observation time of 2 seconds. The mechanism of infiltration is not fully understood. The difficultiesin relating infiltration simply to pore diameter and to thesurface tension, viscosity, and contact angle of the liquidare examined. The possibility that the contact angle of thetest liquid and the slope of the wall within the stomatal poreor at the substomatal cavity may change in relation to themoisturestatus of the plant and the degree of stomatal opening is discussedin relation to Adam's treatment of the passage of a liquid througha pore with a constriction.     

Stomatal patchiness in Mediterranean evergreen sclerophylls

Midday depression of net photosynthesis and transpiration in the Mediterranean sclerophylls Arbutus unedo L. and Quercus suber L. occurs with a depression of mesophyll photosynthetic activity as indicated by calculated carboxylation efficiency (CE) and constant diurnal calculated leaf intercellular partial pressure of CO₂ (C_i). This work examines the hypothesis that this midday depression can be explained by the distribution of patches of either wide-open or closed stomata on the leaf surface, independent of a coupling mechanism between stomata and mesophyll that results in a midday depression of photosynthetic activity of the mesophyll. Pressure infiltration of four liquids differing in their surface tension was used as a method to show the occurrence of stomatal patchiness and to determine the status of stomatal aperture within the patches. Liquids were selected such that the threshold leaf conductance necessary for infiltration through the stomatal pores covered the expected diurnal range of calculated leaf conductance (g) for these species. Infiltration experiments were carried out with leaves of potted plants under simulated Mediterranean summer conditions in a growth chamber. For all four liquids, leaves of both species were found to be fully infiltratable in the morning and in the late afternoon while during the periods leading up to and away from midday the leaves showed a pronounced patchy distribution of infiltratable and non-infiltratable areas. Similar linear relationships between the amount of liquid infiltrated and g (measured by porometry prior to detachment and infiltration) for all liquids clearly revealed the existence of pneumatically isolated patches containing only wide-open or closed stomata. The good correspondence between the midday depression of CE, calculated under the assumption of no stomatal patchiness, and the diurnal changes in non-infiltratable leaf area strongly indicates that the apparent reduction in mesophyll activity results from assuming no stomatal patchiness. It is suggested that simultaneous responses of stomata and mesophyll activity reported for other species may also be attributed to the occurrence of stomatal patchiness. In Quercus coccifera L., where the lack of constant diurnal calculated C_i and major depression of measured CE at noontime indicates different stomatal behavior, non-linear and dissimilar relationships between g and the infiltratable quantities of the four liquids were found. This indicates a wide distribution of stomatal aperture on the leaf surface rather than only wide-open or closed stomata.Dedicated to Professor Otto L. Lange on the occasion of his 65th birthday     

Leaf diffusion resistance, illuminance, and transpiration

Stepwise increases in fluorescent illuminance, imposed as a single variable in a controlled environment, induced progressive stomatal opening in 8 plant species, as evidenced by a consistent decrease in leaf diffusion resistance (R_L), ranging from 15 to 70 sec cm⁻¹ in darkness to about 1 sec cm⁻¹ at approximately 40 kilolux. The minimum R_L values were the same for the upper and the lower epidermis, provided that stomatal density was adequate. Saturation illuminance was not achieved in any species; extrapolation indicates that 50 kilolux would bring about full stomatal opening (R_L ≤ 0.1 sec cm⁻¹).

In 4 species, reasonable agreement was obtained in a controlled environment between transpiration as measured by weight loss and that calculated from determination of (a) the difference in water vapor density from leaf to air, (b) the boundary layer resistance, and (c) the leaf diffusion resistance. This result confirms the physical validity of the resistance measurement procedure.

     

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.     

Stomatal opening quantitatively related to potassium transport: evidence from electron probe analysis

When stomata of Vicia faba opened (from a stomatal aperture of about 2 micrometers to one of 12 micrometers) the solute content of the guard cells increased by 4.8 × 10⁻¹² osmoles per stoma. During the same time an average of 4.0 × 10⁻¹² gram equivalents of K⁺ were transported into each pair of guard cells. This amount of K⁺, if associated with dibasic anions, is sufficient to produce the changes in guard cell volume and osmotic pressure associated with stomatal opening. Analysis of Cl, P, and S showed that these elements were not transported in significant amounts during stomatal opening. This finding suggests that the anions balancing K⁺ were predominantly organic. K⁺ was specifically required because no other elements, likely to be present as cations, were found to accumulate in appreciable quantities in guard cells of open stomata.     

Electrical potentials in stomatal complexes

Guard cells of several species, but predominantly Commelina communis, were impaled by micropipette electrodes and potential differences measured that occurred between cell compartments and the flowing bathing medium. The wall developed a Donnan potential that was between −60 and −70 millivolt in 30 millimolar KCl at pH 7. The density of the fixed charges ranged from 0.3 to 0.5 molar; its dependence on pH was almost identical with the titration curve of authentic polygalacturonic acid. The vacuolar potential of guard cells of Commelina communis L., Zea mays L., Nicotiana glauca Graham, Allium cepa L., and Vicia faba L. was between −40 and −50 millivolt in 30 millimolar KCl when stomata were open and about −30 millivolt when stomata were closed. The vacuolar potential of guard cells of C. communis was almost linearly related to stomatal aperture and responded to changes in the ionic strength in the bathing medium in a Nernstian manner. No specificity for any alkali ion (except Li⁺), ammonium, or choline appeared. Lithium caused hyperpolarization. Calcium in concentrations between 1 and 100 millimolar in the medium led to stomatal closure, also caused hyperpolarization, and triggered transient oscillations in the intracellular potential. Gradients in the electrical potential existed across stomatal complexes with open pores. When stomata closed, these gradients almost disappeared or slightly reverted; all epidermal cells were then at potentials near −30 millivolt in 30 millimolar KCl.     

Aftereffects of low and high temperature pretreatment on leaf resistance, transpiration, and leaf temperature in xanthium

Leaf resistance for water vapor (total diffusion resistance minus boundary layer resistance), transpiration, and leaf temperature were measured in attached leaves of greenhouse-grown Xanthium strumarium L. plants that had been pretreated for 72 hours with high (40 C day, 35 C night), or low (10 C day, 5 C night) air temperatures. Measurements were made in a wind tunnel at light intensity of 1.15 cal cm⁻² min⁻¹, air temperatures between 5 and 45 C, and wind speed of 65 cm sec⁻¹. Leaf resistances in low temperature pretreated plants were higher (8 to 27 sec cm⁻¹) than in controls or high temperature pretreated plants (0.5 to 3 sec cm⁻¹) at leaf temperatures between 5 and 25 C. Thus, the pretreatment influenced stomatal aperture.     

Effects of Phenylmercuric Acetate on Stomatal Movement and Transpiration of Excised Retula papyrifera Marsh. Leaves

Effects of 10⁻³m, 10⁻⁴m, and 10⁻⁵m phenylmercuric acetate (PMA) on stomatal movement and transpiration of excised Betula papyrifera leaves were investigated. Duco cement leaf prints and transpiration decline curves were used for the analysis of stomatal condition. PMA induced stomatal closure and decreased transpiration. Stomata of leaves treated with any of the 3 PMA concentrations closed earlier and at a higher relative water content than did stomata of untreated leaves. As determined from transpiration decline curves, PMA at 10⁻³m caused an increase in apparent “cuticular” transpiration. However, the increase appeared to result largely from some PMA-poisoned stomata which remained open for prolonged periods. Considerable PMA toxicity was observed, with 10⁻³m and 10⁻⁴m concentrations causing browning of leaves. PMA treatment caused a decrease in chlorophyll content, even at a low PMA concentration (10⁻⁵m) which influenced stomatal response only slightly and did not cause evident browning of leaves. The time and degree of stomatal opening varied with stomatal size. Large stomata tended to open earlier and close later than small stomata. Hence, in Betula papyrifera stomata of various size classes were considered as physiologically different populations.     

Effects of Light, Carbon Dioxide, and Temperature on Photosynthesis, Oxygen Inhibition of Photosynthesis, and Transpiration in Solanum tuberosum

Individual leaves of potato (Solanum tuberosum L. W729R), a C₃ plant, were subjected to various irradiances (400-700 nm), CO₂ levels, and temperatures in a controlled-environment chamber. As irradiance increased, stomatal and mesophyll resistance exerted a strong and some-what paralleled regulation of photosynthesis as both showed a similar decrease reaching a minimum at about 85 neinsteins·cm⁻²·sec⁻¹ (about ½ of full sunlight). Also, there was a proportional hyperbolic increase in transpiration and photosynthesis with increasing irradiance up to 85 neinsteins·cm⁻²·sec⁻¹. These results contrast with many C₃ plants that have a near full opening of stomata at much less light than is required for saturation of photosynthesis.     

Action of proline on stomata differs from that of abscisic Acid, g-substances, or methyl jasmonate

Methyl jasmonate (MJ) and a mixture of G₁, G₂, and G₃ (G-substances) inhibited stomatal opening in abaxial epidermis of Commelina benghalensis and complete closure occurred at 10⁻⁶ molar MJ, or 10⁻³ molar G-substances compared to 10⁻⁵ molar abscisic acid (ABA). Proline, even at 10⁻³ molar caused only a partial stomatal closure. Apart from ABA, other endogenous plant growth regulators do regulate stomata. Reduction in the stimulation by fusicoccin and complete stomatal closure, at 30 millimolar KCl or less, were affected by ABA, MJ, or G-substances, but not by proline. The action of MJ or G-substances was similar to ABA in decreasing proton efflux and the levels of potassium, malate, or reducing sugars. Proline, however, interfered with starch-sugar interconversion but had no effect on proton efflux or potassium content of epidermis.     

Stomatal response to abscisic Acid is a function of current plant water status

We investigated, under laboratory and field conditions, the possibility that increasing abscisic acid (ABA) concentrations and decreasing water potentials can interact in their effects on stomata. One experiment was carried out with epidermal pieces of Commelina communis incubated in media with a variety of ABA and polyethylene glycol concentrations. In the media without ABA, incubation in solutions with water potentials between −0.3 and −1.5 megapascals had no significant effect on stomatal aperture. Conversely, the sensitivity of stomatal aperture to ABA was trebled in solutions at −1.5 megapascals compared with sensitivity at −0.3 megapascals. The effect of the change in sensitivity was more important than the absolute effect of ABA at the highest water potential. In a field experiment, sensitivity of maize stomatal conductance to the concentration of ABA in the xylem sap varied strongly with the time of the day. We consider that the most likely explanation for this is the influence of a change in leaf or epidermal water potential that accompanies an increase in irradiance and saturation deficit as the day progresses. These observations suggest that epidermal water relations may act as a modulator of the responses of stomata to ABA. We argue that such changes must be taken into account in studies or modeling of plant responses to drought stress.     

Temperature-dependent stomatal movement in tulip petals controls water transpiration during flower opening and closing

Temperature‐dependent tulip petal opening and closing movement was previously suggested to be regulated by reversible phosphorylation of a plasma membrane aquaporin ( Azad et al., 2004a ). Stomatal apertures of petals were investigated during petal opening at 20°C and closing at 5°C. In completely open petals, the proportion of open stomata in outer and inner surfaces of the same petal was 27 ± 6% and 65 ± 3%, respectively. During the course of petal closing, stomatal apertures in both surfaces reversed, and in completely closed petals, the proportion of open stomata in outer and inner surfaces of the same petal was 74 ± 3% and 29 ± 6%, respectively, indicating an inverse relationship between stomatal aperture in outer and inner surfaces of the petal during petal opening and closing. Both petal opening and stomatal closure in the outer surface of the petal was inhibited by a Ca²⁺ channel blocker and a Ca²⁺ chelator, whereas the inner surface stomata remained unaffected. On the other hand, sodium nitroprusside, a nitric oxide donor, had no effect on stomatal aperture of the outer surface but influenced the inner surface stomatal aperture during petal opening and closing, suggesting different signalling pathways for regulation of temperature‐dependent stomatal changes in the two surfaces of tulip petals. Stomata were found to be differentially distributed in the bottom, middle and upper parts of tulip petals. During petal closing, water transpiration was observed by measuring the loss of ³H₂O. Transpiration of ³H₂O by petals was fivefold greater in the first 10 min than that found after 30 min, and the transpiration rate was shown to be associated with stomatal distribution and aperture. Thus, the stomata of outer and inner surfaces of the petal are involved in the accumulation and transpiration of water during petal opening.     

Water Relations of Cotton Plants under Nitrogen Deficiency: II. Environmental Interactions on Stomata

Nitrogen deficiency in cotton plants (Gossypium hirsutum L.) considerably increased the sensitivity of stomata to water stress. At air temperatures of 27, 35, and ≥40 C, threshold potentials for complete stomatal closure were −10, −15, and −26 bars in N-deficient plants and −20, −20, and −30 bars in high-N plants, respectively. This three-way interaction among N supply, water potential, and air temperature was similar to that exerted on leaf expansion. The effects of N supply on stomatal behavior could not be explained on the basis of either osmotic or structural considerations. Rather, effects of N deficiency on mesophyll and stomata were independent and divergent. Stomatal behavior may impart a stress avoidance type of drought resistance to N-deficient plants.     

STOMATAL PENETRATION: EFFECT OF SURFACTANTS AND ROLE IN FOLIAR ABSORPTION

Stomatal penetration of aqueous solutions into pear (Pyrus communis L. cv. Bartlett) leaves was induced with surfactants. The effectiveness of surfactants in promoting stomatal penetration was related to their effectiveness in decreasing surface tension. Vatsol OT (dioctyl ester of sodium sulfosuccinic acid) was most effective, X-77 (compounded product of alkyl aryl polyethoxy ethanol, free fatty acids and isopropanol) intermediate, and Tween 20 (polyoxyethylene sorbitan monolaurate) least effective. The percentage of stomata penetrated was limited, ranging from 0.5–4.5 %. There was no evidence of stomatal penetration of aqueous solutions with surface tensions of approximately 70 dyne cm⁻¹. The importance of stomatal penetration in foliar absorption under conditions of plant culture is discussed.     

Water Stress Reduces Ozone Injury via a Stomatal Mechanism

Various studies have shown that water-stressed plants are more tolerant of ozone exposures than are unstressed plants. Two probable explanations for this tolerance are (a) stomatal closure which reduces ozone uptake and (b) biochemical or anatomical changes within the leaves. Phaseolus vulgaris cv Pinto bean plants were established and transferred to membrane systems which controlled the osmotic potential around the roots at −35 or −80 kilopascals for 5 days prior to ozone treatment (0 or 1.0 microliters per liter for 2 hours). Both water-stressed and unstressed plants were sprayed with various concentrations of abscisic acid to close the stomata or with fusicoccin to induce stomata opening. The abaxial stomatal resistances of primary and trifoliate leaves were measured just prior to ozone exposure. Plant response to ozone was determined by stress ethylene production and chlorophyll loss. Both water stress and abscisic acid induced stomatal closure and reduced ozone injury. In water-stressed plants, fusicoccin induced stomatal opening and those plants were as sensitive to ozone as were the non-water-stressed plants. These data suggest that water stress protects plants from ozone injury mainly through its influence on stomatal aperture rather than through biochemical or anatomical changes.     

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

K⁺ and Cl⁻ contents of guard cells and of ordinary epidermal cells were determined in epidermal samples of Allium cepa L. by electron probe microanalysis; malate contents of the same samples were determined by enzymic oxidation. KCl was, in general, the major osmoticum in guard cells, irrespective of whether stomata had opened on leaves or in epidermal strips floating on solutions. The solute requirement varied between 50 and 110 femtomoles KCl per micrometer increase in aperture per pair of guard cells. Stomata did not open on solutions of K iminodiacetate, presumably because its anion could not be taken up. Stomata opened if KCl or KBr was provided. Taken together, the results indicate that the absence of starch from guard cells deprived them of the ability to produce malate in amounts of osmotic consequence and that the presence of absorbable Cl⁻ (or Br⁻) was necessary for stomatal opening.     

Light quality and osmoregulation in vicia guard cells : evidence for involvement of three metabolic pathways

Osmoregulation in opening stomata of epidermal peels from Vicia faba L. leaves was investigated under a variety of experimental conditions. The K⁺ content of stomatal guard cells and the starch content of guard cell chloroplasts were examined with cobaltinitrite and iodine-potassium iodide stains, respectively; stomatal apertures were measured microscopically. Red light (50 micromoles per square meter per second) irradiation caused a net increase of 3.1 micrometers in aperture and a decrease of −0.4 megapascals in guard cell osmotic potential over a 5 hour incubation, but histochemical observations showed no increase in guard cell K⁺ content or starch degradation in guard cell chloroplasts. At 10 micromoles per square meter per second, blue light caused a net 6.8 micrometer increase in aperture over 5 hours and there was a substantial decrease in starch content of chloroplasts but no increase in guard cell K⁺ content. At 25 micromoles per square meter per second of blue light, apertures increased faster (net gain of 5.7 micrometers after 1 hour) and starch content decreased. About 80% of guard cells had a higher K⁺ content after 1 hour of incubation but that fraction decreased to 10% after 5 hours. In the absence of KCl in the incubation medium, stomata opened slowly in response to 25 micomoles per square meter per second of blue light, without any K⁺ gain or starch loss. In dual beam experiments, stomata irradiated with 50 micomoles per square meter per second of red light for 3 hours opened without detectable starch loss or K⁺ gain; addition of 25 micomoles per square meter per second of blue light caused a further net gain of 4.4 micometers in aperture accompanied by substantial K⁺ uptake and starch loss. Comparison of K⁺ content in guard cells of opened stomata in epidermal peels with those induced to open in leaf discs showed a substantially higher K⁺ content in the intact tissue than in isolated peels. These results are not consistent with K⁺ (and its counterions) as the universal osmoticum in guard cells of open stomata under all conditions; rather, the data point to sugars arising from photosynthesis and from starch degradation as additional osmotica. Biochemical confirmation of these findings would indicate that osmoregulation during stomatal opening is the result of three key metabolic processes: ion transport, photosynthesis, and sugar metabolism.     

Water-Transport System and Its Components in Higher Plants: 5. Dynamic Model of the Water-Transport System in Transpiring Plants

The dynamic model of aperture behavior in leaf stomata of intact transpiring plants under constant environmental conditions is considered. The model is based on the supposed existence of feed-forward relations in the water-transport system: the decrease in root resistance – reduction in the capillary tension in the leaf apoplast – enlargement of the stomatal aperture. In parallel, there is a positive feedback relation: enlargement of the stomatal aperture – enhancement of transpiration – increase in the viscous flow through the root – intensified supply of dissolved oxygen to the respiratory centers of the root symplasm – elevation of turgor and the decrease in root resistance. Under the fulfillment of nonstringent physical requirementship, this system is capable of generating auto-oscillations with a period of 20–80 min. The model describes the relationship between the frequency–amplitude characteristics of auto-oscillations and the main parameters of the water-transport system in plants. In particular, the model presents a clue to one of the most intriguing phenomenon: the moistening of the root environment increases the frequency of auto-oscillations in some cases and reduces this frequency in others.     

Mutual Antagonism of Sulfur Dioxide and Abscisic Acid in Their Effect on Stomatal Aperture in Broad Bean (Vicia faba L.) Epidermal Strips

Abscisic acid (ABA) was found to counteract the stomatal opening in Vicia faba L. caused by SO₂. The antagonism between SO₂ and ABA was mutual, and their combined effect depended upon which compound was in the greatest concentration. Stomatal apertures were monitored in detached epidermal strips floated in the light on aqueous solutions of SO₂ (sulfurous acid) and/or ABA in 0.01 molar sodium citrate buffer (pH 5.8). Low concentrations of sulfurous acid (10⁻¹⁰ to 10⁻⁷ molar) increased stomatal aperture, but concentrations greater than 10⁻⁵ molar decreased it. A progressive decrease in aperture size occurred as ABA was increased from 10⁻¹⁰ to 10⁻⁵ molar.     

广州市常见行道树种叶片表面形态与滞尘能力

以广州市常见的18种行道树为对象,通过扫描电镜观察比较了行道树的叶表面形态结构、应用接触角测定仪测定了绿化树种叶片的接触角对滞尘能力的影响.结果表明:不同树种的滞尘量差异显著,18种植物叶片雨后第26天的最大滞尘量在0.066-1.831 g/m2,物种间相差达27倍以上.叶表面具有网状结构,气孔密度较大(20＜气孔密度＜60个)且气孔开口较大(如芒果)容易滞留粉尘;叶表面平滑具有蜡质层,气孔排列整齐,无明显起伏(如红花羊蹄甲、桃花心木、大叶紫薇、鹅掌藤),滞尘能力较弱.植物叶片接触角与滞尘量呈负相关(r=-0.614),接触角＜90°的表现为亲水性.易润湿的植物叶片雨后第26天最大滞尘量在1.0-1.831 g/m2,叶片表面的形态结构凹凸不平,具有钩状或脊状褶皱、突起等且20＜气孔密度＜60范围内,测得的接触角较小(芒果、重阳木、高山榕),使得粉尘与植物叶片接触面积较大,粉尘不易从叶面脱落,滞尘能力较强.而接触角较大的盆架树、麻楝、大叶紫薇、鹅掌藤和红花羊蹄甲的滞尘量均＜1.0g/m2,其特殊的表面结构和疏水的蜡质使颗粒物不易吸附在植物叶片上,因此滞尘能力较弱.由此可见,植物叶表面蜡质含量和气孔密度及其叶片接触角的大小是影响植物叶片滞尘能力的主要因素,在进行城市绿化时,适当考虑选择叶表面形态有利于滞尘的绿化树种,将可提高城市植被的环境效应.