Cell turgor, osmotic pressure and water potential in the upper epidermis of barley leaves in relation to cell location and in response to NaCl and air humidity

Previous single-cell studies on the upper epidermis of barleyleaves have shown that cells differ systematically in theirsolute concentrations depending on their location relative tostomatal pores and veins and that during NaCl stress, gradientsin osmotic pressure () develop (Fricke et al., 1995, 1996; Hinde,1994). The objective of the present study was to address thequestion to which degree these intercellular differences insolute concentrations and it are associated with intercellulardifferences in turgor or water potential (). Epidermal cellsanalysed were located at various positions within the ridgeregions overlying large lateral or intermediate veins, in thetrough regions between those veins or in between stomata (i.e.interstomatal cells). Turgor pressure of cells was measuredusing a cell pressure probe, and of extracted cell sap wasdetermined by picolitre osmometry. For both large and intermediatelateral veins, there were no systematic differences in turgorbetween cells located at the base, mid or top of ridges, regardlessof whether plants were analysed at low or high PAR (10 or 300–400µmol photons m^–2 s^–1). However, turgor withina ridge region was not necessarily uniform, but could vary byup to 0.14 MPa (1.4 bar) between adjacent cells. In 60 out of63 plants, turgor of ridge cells was either slightly or significantlyhigher than turgor of trough (lowest turgor) or interstomatalcells (intermediate turgor). The significance and magnitudeof turgor differences was higher in plants analysed under highPAR or local air flow than in plants analysed under low PAR.The largest (up to 0.41 MPa) and consistently significant differencesin turgor were found in plants treated for 3–9 d priorto analysis with 100 mM NaCl. For both NaCl-treated and non-treated(control) plants, differences in turgor between cell types weremainly due to differences in since differences in were negligible(0.01–0.04 MPa). Epidermal cell , in NaCl-treated plantswas about 0.38 MPa more negative than in control plants dueto higher . Turgor pressures were similar. Following a suddenchange in rooting-medium or air humidity, turgor of both ridgeand trough cells responded within seconds and followed the sametime-course of relaxation. The half time (T_1/2) of turgor relaxationwas not limited by the cell's T_1/2 for water exchange. Key words: Barley leaf epidermis, cell turgor, heterogeneity, NaCl stress, osmotic pressure, water potential     

Cell elongation, turgor and osmotic pressure in developing sunflower hypocotyls

The relationship between cell elongation, change in turgor andcell osmotic pressure was investigated in the sub-apical regionof hypocotyls of developing sunflower seedlings (Helianthusannuus L.) that were grown in continuous white light. Cell turgorwas measured with the pressure probe. The same hypocotyl sectionswere used for determination of osmotic pressure of the tissuesap. Acceleration of cell elongation during the early phaseof growth was accompanied by a 25% decrease in both turgor andosmotic pressure. During the linear phase of growth both pressuresremained largely constant. The difference between turgor andosmotic pressure (water potential) was –0.10 to –0.13MPa. Excision of one cotyledon had no effect on growth, turgorand osmotic pressure. However, after removal of both cotyledonscell elongation ceased and a substantial decrease in both pressureswas measured. In addition, we determined the longitudinal tissuepressure in seedlings from which one or both cotyledons hadbeen removed. Tissue pressure and turgor were very similar quantitiesunder all experimental conditions. Our results demonstrate thatturgor and cell osmotic pressure show a parallel change duringdevelopment of the stem. Cessation of cell elongation afterremoval of the cotyledons is attributable to a decrease in turgor(tissue) pressure, which provides the driving force for growthin the hypocotyl of the intact plant. Key words: Cell elongation, Helianthus annuus, osmotic pressure, tissue pressure, turgor     

Control of leaf cell elongation in barley. Generation rates of osmotic pressure and turgor, and growth-associated water potential gradients

In a previous study on the effects of N-supply on leaf cell elongation, the spatial distribution of relative cell elongation rates (RCER), epidermal cell turgor, osmotic pressure (OP) and water potential (Ψ) along the elongation zone of the third leaf of barley was determined (W. Fricke et al. 1997, Planta 202: 522–530). The results suggested that in plants receiving N at fixed relative addition rates (N-supply limitation of growth), cell elongation was rate-limited by the rate of solute provision, whereas in plants growing on complete nutrient solution containing excessive amounts of N (N-demand limitation), cell elongation was rate-limited by the rate of water supply or wall yielding. In the present paper, these suggestions were tested further. The generation rates of cell OP, turgor and Ψ along the elongation zone were calculated by applying the continuity equation of fluid dynamics to the previous data. To allow a more conclusive interpretation of results, anatomical data were collected and bulk solute concentrations determined. The rate of OP generation generally exceeded the rate of turgor generation. As a result, negative values of cell Ψ were created, particularly in demand-limited plants. These plants showed highest RCER along the elongation zone and a Ψ gradient of at least −0.15 MPa between water source (xylem) and expanding epidermal cells. The latter was similar to a theoretically predicted value (−0.18 MPa). Highest rates of OP generation were observed in demand-limited plants, with a maximum rate of 0.112 MPa · h⁻¹ at 16–20 mm from the leaf base. This was almost twice the rate in N-supply-limited plants and implied that the cells in the leaf elongation zone were capable of importing (or synthesising) every minute almost 1 mM of osmolytes. Potassium, Cl⁻ and NO₃ ⁻ were the main inorganic osmolytes (only determined for demand-limited plants). Their concentrations suggest that, unlike the situation in fully expanded epidermal cells, sugars are used to generate OP and turgor. Anatomical data revealed that the zone of lateral cell expansion extended distally beyond the zone of cell elongation. It is concluded that leaf cell expansion in barley relies on high rates of water and solute supply, rates that may not be sustainable during periods of sufficient N-supply (limitation by water supply: Ψ gradients) or limiting N-supply (limitation by solute provision: reduced OP-generation rates). To minimise the possibility of growth limitation by water and osmolyte provision, longitudinal and lateral cell expansion peak at different locations along the growth zone. Received: 15 October 1997 / Accepted: 12 March 1998     

Abscisic Acid accumulation in cotton leaves in response to dehydration at high pressure

Pressure-volume techniques were utilized to examine the control of abscisic acid (ABA) accumulation in dehydrated cotton (Gossypium hirsutum L. cv Tamcot SP 37) leaves. Leaves were rapidly dehydrated in a pressure chamber to a balance pressure coincident with the loss of cellular turgor, and then the pressure was either maintained at that level or released. Rapid accumulation of ABA began within two hours after the balance pressure was achieved, whether or not the high pressure potential of the cells was maintained by the externally imposed pressure. The results show that loss of pressure per se does not trigger ABA accumulation in dehydrated leaves. Rather, the stimulus may be related to cellular shrinkage and relaxation of the elastic wall.     

Stress-induced osmotic adjustment in growing regions of barley leaves

Young barley seedlings were stressed using nutrient solutions containing NaCl or polyethylene glycol and measurements were made of leaf growth, water potential, osmotic potential and turgor values of both growing (basal) and nongrowing (blade) tissues. Rapid growth responses similar to those noted for corn (Plant Physiology 48: 631-636) were obtained using either NaCl or polyethylene glycol treatments by which exposure of seedlings to solutions with water potential values of −3 to −11 bars effected an immediate cessation of leaf elongation with growth resumption after several minutes or hours. Latent periods were increased and growth resumption rates were decreased as water potential values of nutrient solutions were lowered. In unstressed transpiring seedlings, water potential and osmotic potential values of leaf basal tissues were usually −6 to −8 bars, and −12 to −14 bars, respectively. These tissues began to adjust osmotically when exposed to any of the osmotic solutions, and hourly reductions of 1 to 2 bars in both water potential and osmotic potential values usually occurred for the first 2 to 4 hours, but reduction rates thereafter were lower. When seedlings were exposed to solutions with water potential values lower than those of the leaf basal tissues, growth resumed about the time water potential values of those tissues fell to that of the nutrient solution. After 1 to 3 days of seedling exposure to solutions with different water potential values, cumulative leaf elongation was reduced as the water potential values of the root medium were lowered. Reductions in water potential and osmotic potential values of tissues in leaf basal regions paralleled growth reductions, but turgor value was largely unaffected by stress. In contrast, water potential, osmotic potential, and turgor values of leaf blades were usually changed slightly regardless of the degree and duration of stress, and blade water potential values were always higher than water potential values of the basally located cells. It is hypothesized that blades have high water potential values and are generally unresponsive to stress because water in most of the mesophyll cells in this area does not exchange readily with water present in the transpiration stream.     

Rapid and tissue-specific accumulation of solutes in the growth zone of barley leaves in response to salinity

The aim of the present study was to test whether rapid accumulation of solutes in response to salinity in leaf tissues of barley (Hordeum vulgare L.) contributes to recovery and maintenance of residual elongation growth. Addition of 100 mM NaCl to the root medium caused an immediate reduction close to zero in elongation velocity of the growing leaf 3. After 20–30 min, elongation velocity recovered suddenly, to 40–50% of the pre-stress level. Bulk osmolality increased first, after 60 min, significantly in the proximal half of the elongation zone. Over the following 3 days, osmolality increases became significant in the distal half of the elongation zone, the adjacent, enclosed non-elongation zone and finally in the emerged portion of the blade. The developmental gradient and time course in osmolality increase along the growing leaf was reflected in the pattern of solute (Cl, Na and K) accumulation in bulk tissue and epidermal cells. The partitioning of newly accumulated solutes between epidermis and bulk tissue changed with time. Even though solute accumulation does not contribute to the sudden and partial growth recovery 20–30 min after exposure to salt, it does facilitate residual growth from 1 h onwards. This is due to a high sink strength for solutes of the proximal part of the growth zone and its ability to accumulate solutes rapidly and at high rates.Abbreviations EDX analysis Energy-dispersive X-ray analysis - LEV Leaf elongation velocity - LVDT Linear variable differential transformer - REGR Relative elemental growth rate     

Proteomic response of barley leaves to salinity

Drought and salinity stresses are adverse environmental factors that affect crop growth and yield. Proteomic analysis offers a new approach to identify a broad spectrum of genes that are expressed in living system. We applied this technique to investigate protein changes that were induced by salinity in barley genotypes (Hordeum vulgare L.), Afzal, as a salt-tolerant genotype and L-527, as a salt-sensitive genotype. The seeds of two genotypes were sown in pot under controlled condition of greenhouse, using a factorial experiment based on a randomized complete block design with three replications. Salt stress was imposed at seedling stage and leaves were collected from control and salt-stressed plant. The Na⁺ and K⁺ concentrations in leaves changed significantly in response to short-term stress. About 850 spots were reproducibly detected and analyzed on 2-DE gels. Of these, 117 proteins showed significant change under salinity condition in at least one of the genotypes. Mass spectrometry analysis using MALDI-TOF/TOF led to the identification some proteins involved in several salt responsive mechanisms which may increase plant adaptation to salt stress including higher constitutive expression level and upregulation of antioxidant, upregulation of protein involved in signal transduction, protein biosynthesis, ATP generation and photosynthesis. These findings may enhance our understanding of plant molecular response to salinity.     

Biophysical characterization of changes in amounts and activity of Escherichia coli cell and compartment water and turgor pressure in response to osmotic stress

To obtain turgor pressure, intracellular osmolalities, and cytoplasmic water activity of Escherichia coli as a function of osmolality of growth, we have quantified and analyzed amounts of cell, cytoplasmic, and periplasmic water as functions of osmolality of growth and osmolality of plasmolysis of nongrowing cells with NaCl. The effects are large; NaCl (plasmolysis) titrations of cells grown in minimal medium at 0.03 Osm reduce cytoplasmic and cell water to approximately 20% and approximately 50% of their original values, and increase periplasmic water by approximately 300%. Independent analysis of amounts of cytoplasmic and cell water demonstrate that turgor pressure decreases with increasing osmolality of growth, from approximately 3.1 atm at 0.03 Osm to approximately 1.5 at 0.1 Osm and to less than 0.5 atm above 0.5 Osm. Analysis of periplasmic membrane-derived oligosaccharide (MDO) concentrations as a function of osmolality, calculated from literature analytical data and measured periplasmic volumes, provides independent evidence that turgor pressure decreases with increasing osmolality, and verifies that cytoplasmic and periplasmic osmolalities are equal. We propose that MDO play a key role in periplasmic volume regulation at low-to-moderate osmolality. At high growth osmolalities, where only a small amount of cytoplasmic water is observed, the small turgor pressure of E. coli demonstrates that cytoplasmic water activity is only slightly less than extracellular water activity. From these findings, we deduce that the activity of cytoplasmic water exceeds its mole fraction at high osmolality, and, therefore, conclude that the activity coefficient of cytoplasmic water increases with increasing growth osmolality and exceeds unity at high osmolality, presumably as a consequence of macromolecular crowding. These novel findings are significant for thermodynamic analyses of effects of changes in growth osmolality on biopolymer processes in general and osmoregulatory processes in particular in the E. coli cytoplasm.     

Correlation between loss of turgor and accumulation of abscisic acid in detached leaves

Mature leaves of Phaseolus vulgaris L. (red kidney bean), Xanthium strumarium L. (cocklebur), and Gossypium hirsutum L. (cotton) were used to study accumulation of abscisic acid (ABA) during water stress. The water status of individual, detached leaves was monitored while the leaves slowly wilted, and samples were cut from the leaves as they lost water. The leaf sections were incubated at their respecitive water contents to allow ABA to build up or not. At least 8 h were required for a new steady-state level of ABA to be established. The samples from any one leaf covered a range of known water potentials (), osmotic pressures (), and turgor pressures (p). The and p values were calculated from pressure-volume curves, using a pressure bomb to measure the water potentials. Decreasing water potential had little effect on ABA levels in leaves at high turgor. Sensitivity of the production of ABA to changes in progressively increased as turgor approached zero. At p=1 bar, ABA content averaged 4 times the level found in fully turgid samples. Below p=1 bar, ABA content increased sharply to as much as 40 times the level found in unstressed samples. ABA levels rose steeply at different water potentials for different leaves, according to the at which turgor became zero. These differences were caused by the different osmotic pressures of the leaves that were used; must cqual - for turgor to be zero. Leaves vary in , not only among species, but also between plants of one and the same species depending on the growing conditions. A difference of 6 bars (calculated at =0) was found between the osmotic pressures of leaves from two groups of G. hirsutum plants; one group had previously experienced periodic water stress, and the other group had never been stressed. When individual leaves were subsequently wilted, the leaves from stress-conditioned plants required a lower water potential in order to accumulate ABA than did leaves from previously unstressed plants. On the basis of these results we suggest that turgor is the critical parameter of plant water relations which controls ABA production in water-stressed leaves.Abbreviations ABA abscisic acid - me-ABA abscisic-acid methyl ester - leaf water potential - osmotic pressure - p volumeaveraged turgor - volumetric modulus of elasticity     

Changes in turgor pressure in response to increases in external NaCl concentration in the gas-vacuolate cyanobacterium Microcystis sp.

Changes in cell turgor pressure have been followed in cells of Microcystis sp. transferred to culture medium containing added NaCl at osmolalities of 30–1,500 mosmol kg^-1 ( 74–3,680 kPa). Upon upshock turgor decreased, due to osmotically-induced water loss from the cell. However, partial recovery of turgor was then observed in illuminated cells, with maximum turgor regain in media containing 30–500 mosmol kg^-1 NaCl. The lightdependent recovery of turgor pressure was completed within 60 min, with no evidence of further changes in cell turgor up to 24 h. This is the first direct evidence that turgor regulation may occur in a prokaryotic organism. Short-term increases in cell K⁺ content were also observed upon upshock in NaCl, indicating that turgor regain may involve a turgorsensitive K⁺ uptake system. Estimation of internal K⁺ concentration in cells transferred to 250 mosmol kg^-1 NaCl showed that changes in cell K⁺ may account for at least half of the observed turgor regain up to 60 min.     

Cell water potential,osmotic potential,and turgor in the epidermis and mesophyll of transpiring leaves

Water potential, osmotic potential and turgor measurements obtained by using a cell pressure probe together with a nanoliter osmometer were compared with measurements obtained with an isopiestic psychrometer. Both types of measurements were conducted in the mature region of Tradescantia virginiana L. leaves under non-transpiring conditions in the dark, and gave similar values of all potentials. This finding indicates that the pressure probe and the osmometer provide accurate measurements of turgor, osmotic potentials and water potentials. Because the pressure probe does not require long equilibration times and can measure turgor of single cells in intact plants, the pressure probe together with the osmometer was used to determine in-situ cell water potentials, osmotic potentials and turgor of epidermal and mesophyll cells of transpiring leaves as functions of stomatal aperture and xylem water potential. When the xylem water potential was-0.1 MPa, the stomatal aperture was at its maximum, but turgor of both epidermal and mesophyll cells was relatively low. As the xylem water potential decreased, the stomatal aperture became gradually smaller, whereas turgor of both epidermal and mesophyll cells first increased and afterward decreased. Water potentials of the mesophyll cells were always lower than those of the epidermal cells. These findings indicate that evaporation of water is mainly occurring from mesophyll cells and that peristomatal transpiration could be less important than it has been proposed previously, although peristomatal transpiration may be directly related to regulation of turgor in the guard cells.     

Changes in PS II heterogeneity in response to osmotic and ionic stress in wheat leaves (Triticum aestivum)

High salt stress involves ionic stress as well as osmotic stress. In this work we have tried to differentiate between the ionic and osmotic effects of salt stress on the basis of their ability to cause changes in PS II heterogeneity. PS II heterogeneity is found to vary with environmental conditions. Osmotic stress caused no change in the Q(B) reducing side heterogeneity and a reversible change in antenna heterogeneity. The number of Q(B) non-reducing centers increased under ionic stress but were unaffected by osmotic stress. On the other hand ionic stress led to a partially irreversible change in Q(B) reducing side heterogeneity and a reversible change in antenna heterogeneity. In response to both ionic and osmotic effect, there is conversion of active PS IIα centres to inactive PSIIβ and γ centres.     

Role of carbohydrates in proline accumulation in wilted barley leaves

The effect of wilting on proline synthesis, proline oxidation, and protein synthesis—all of which contribute to proline accumulation—was determined in nonstarved barley (Hordeum vulgare L.) leaves. Nonstarved leaves were from plants previously in the light for 24 hours and starved leaves were from plants previously in the dark for 48 hours. Wilted leaves from nonstarved plants accumulated proline at the rate of about 1 μmole per hour per gram of fresh weight whereas wilted leaves from starved plants accumulated very little proline. Wilting caused a 40-fold stimulation of proline synthesis from glutamate in nonstarved leaves but had very little effect in starved leaves. Proline oxidation and protein synthesis, on the other hand, were inhibited by wilting in both nonstarved and starved leaves. Thus, the role of carbohydrates in proline accumulation is to supply precursors for the stimulated proline synthesis. These results further indicate that the main metabolic response causing proline to accumulate in wilted barley leaves is the stimulation of proline synthesis from glutamate. The difference between these results and those obtained with beans is discussed.

Wilting caused an increased conversion of glutamate to other products. In nonstarved leaves, conversion to organic acids as well as to proline was increased. In starved leaves, wilting caused an increase in the conversion of glutamate to glutamine, aspartate, asparagine, and organic acids.

     

Changes in potassium fluxes in cells of carrot storage tissue related to turgor pressure

The effect of increasing external osmotic pressure on potassium fluxes in aged and fresh-cut discs of Daucus carota L. storage tissue was investigated. An increase of the external osmotic pressure by 5 bars of mannitol solution increased the rate of K⁺ net uptake of aged discs to 180% of their control rate. At 3°C and in 0.1 m M azide, in which a net efflux of potassium was observed, the mannitol treatment caused a reduction in the net efflux. In fresh-cut discs, in which the capability of net influx was rather low and a substantial net release of potassium was noted, the increase in the external osmotic pressure by mannitol caused a 70% inhibition in the net efflux. This effect was also observed at 3°C.
Measurements of separate fluxes confirmed the assumption that the mannitol treatment brought about two distinct effects on K⁺ fluxes: (a) raised the metabolically-dependent influx and (b) lowered the membrane permeability-dependent efflux. When a permeating solute (ethylene glycol) was used instead of mannitol, no effect on K⁺ flux was detectable. Reasons are given for relating the observed changes in K⁺ fluxes to the reduction in turgor pressure of the cells.     

Fruit ripening in Vitis vinifera: spatiotemporal relationships among turgor, sugar accumulation, and anthocyanin biosynthesis

This study reports the first observations indicating the spatiotemporal relationships among genetic and physiological aspects of ripening in the berry of Vitis vinifera. At the onset of ripening in the red flesh variety Alicante Bouschet, colour development began in the flesh at the stylar end of the fruit and progressed toward the pedicel end flesh and into the skin. Tissue solute potential and cell turgor also decreased first in the flesh. The decrease in flesh solute potential was due to accumulation of sugars, glucose and fructose, an accumulation that is integral to ripening. Expression of the anthocyanin biosynthesis-related genes VvMybA and VvUFGT was linearly related to the decrease in solute potential. Expression of VvMybA, and to a lesser extent VvUFGT, was correspondingly low in green tissue, higher in the red, stylar end flesh of berries beginning to ripen, and greatest in red berries. In contrast, expression of the abscisic acid biosynthesis-related genes VvNCED1 and VvNCED2 was not correlated with the other spatiotemporal aspects of the onset of ripening. These results, together with earlier work showing that sugar accumulation and acid loss also begin in the stylar flesh in other varieties, indicate that ripening in the grape berry originates in the stylar end flesh.     

The effect of blue light on stomatal oscillations and leaf turgor pressure in banana leaves

Stomatal oscillations are cyclic opening and closing of stomata, presumed to initiate from hydraulic mismatch between leaf water supply and transpiration rate. To test this assumption, mismatches between water supply and transpiration were induced using manipulations of vapour pressure deficit (VPD) and light spectrum in banana (Musa acuminata). Simultaneous measurements of gas exchange with changes in leaf turgor pressure were used to describe the hydraulic mismatches. An increase of VPD above a certain threshold caused stomatal oscillations with variable amplitudes. Oscillations in leaf turgor pressure were synchronized with stomatal oscillations and balanced only when transpiration equaled water supply. Surprisingly, changing the light spectrum from red and blue to red alone at constant VPD also induced stomatal oscillations – while the addition of blue (10%) to red light only ended oscillations. Blue light is known to induce stomatal opening and thus should increase the hydraulic mismatch, reduce the VPD threshold for oscillations and increase the oscillation amplitude. Unexpectedly, blue light reduced oscillation amplitude, increased VPD threshold and reduced turgor pressure loss. These results suggest that additionally, to the known effect of blue light on the hydroactive opening response of stomata, it can also effect stomatal movement by increased xylem–epidermis water supply.