Modulation of water stress effects on photosynthesis by altered leaf k

Wheat irrigated with nutrient solutions containing 0, 0.2, 0.5, 1, 2, or 6 millimolar K⁺ had maximum photosynthetic rates at 1 to 2 millimolar K⁺ concentrations. Rates in the 6 millimolar K⁺-grown plants were not higher than the 2 millimolar K⁺-grown wheat, and rates were inhibited below 0.5 millimolar K⁺. Photosynthesis was measured by both attached whole leaf CO₂ uptake and by ¹⁴CO₂ fixation of leaf slices in solution. Exposure of leaf slices from 0.2, 2, and 6 millimolar K⁺-grown wheat to various assay media water potentials showed that photosynthesis of the 0.2 millimolar K⁺-grown wheat decreased from control (high water potential) rates by 35%, that of the 2 millimolar K⁺-grown wheat by 20.4%, and that of the 6 millimolar K⁺-grown wheat by only 8.3% at −3.11 megapascals. Also, photosynthesis of the 6 millimolar K⁺-grown wheat was enhanced by 28% over that of the 2 millimolar K⁺ wheat at the most severe water stress (−3.11 megapascals), indicating that the excess leaf K⁺ in the 6 millimolar K⁺-grown wheat partially reversed dehydration effects on photosynthesis. Oligomycin eliminated the protective effects of high K⁺ on photosynthesis in dehydrated leaf slices. These results suggest that the protective effect of high K⁺ under water stress may involve the exchange of K⁺ in the cytoplasm for stroma H⁺, thus altering stromal pH and restoring photosynthesis. The protective effect of high K⁺ was also observed in attached whole leaf photosynthesis of in situ water-stressed wheat grown on 0.2, 2, and 6 millimolar K⁺. Under water stress, rates of the 6 millimolar K⁺-grown wheat were enhanced by 66.2% and 113.9% over that of 2 millimolar K⁺-grown wheat in two separate experiments. Internal CO₂ concentration of the 6 millimolar K⁺-grown wheat was lower than that of the 0.2 and 2 millimolar K⁺-grown wheat. These results suggest that the high K⁺ effects on chloroplast photosynthesis seen in leaf slices also occur at the whole plant level.     

Salinity effects on photosynthesis in isolated mesophyll cells of cowpea leaves

Mesophyll cells from leaves of cowpea (Vigna unquiculata [L.] Walp.) plants grown under saline conditions were isolated and used for the determination of photosynthetic CO₂ fixation. Maximal CO₂ fixation rate was obtained when the osmotic potential of both cell isolation and CO₂ fixation assay media were close to leaf osmotic potential, yielding a zero turgor pressure. Hypotonic and hypertonic media decreased the rate of photosynthesis regardless of the salinity level during plant growth. No decrease in photosynthesis was obtained for NaCl concentrations up to 87 moles per cubic meter in the plant growing media and only a 30% decrease was found at 130 moles per cubic meter when the osmotic potential of cell isolation and CO₂ fixation media were optimal. The inhibition was reversible when stress was relieved. At 173 moles per cubic meter NaCl, photosynthesis was severely and irreversibly inhibited. This inhibition was attributed to toxic effects caused by high Cl⁻ and Na⁺ accumulation in the leaves. Uptake of sorbitol by intact cells was insignificant, and therefore not associated with cell volume changes. The light response curve of cells from low salinity grown plants was similar to the controls. Cells from plants grown at 173 moles per cubic meter NaCl were light saturated at a lower radiant flux density than were cells from lower salinity levels.     

Nicotine Production by Tissue Cultures of Tobacco as Influenced by Various Culture Parameters

Assimilate efflux from vacuum-infiltrated leaf slices (spinach, barley) into a buffered solution was examined in relation to Ca^{+ +} -activity and osmotic conditions. Efflux from isolated mesophyll protoplasts and from a unicellular green alga (Eremosphaera viridis de Bary) was also measured.In the presence of Ca^{+ +}, assimilate efflux from leaf slices was small (1 to 5 % of the total carbon fixation rate, depending on osmotic conditions). Efflux was drastically stimulated by addition of Ca^{+ +} -chelators. If expressed as µmol carbon mg^-1 chlorophyll h^-1, it reached 50 % of the assimilation rate. Efflux from protoplasts or algae was slow and insensitive to Ca^{+ +} chelators at concentrations which caused fast efflux from leaf slices.Assimilate efflux from leaf slices was rather unspecific. Both in the tissue and the surrounding medium, sucrose was the most abundantly labelled compound (70 to 80 % of total soluble labelled material).A 50 % decrease of efflux was observed when turgor pressure was lowered by addition of sorbitol (200 to 300 mosmol kg^-1). At extremely high sorbitol concentrations (> 1500 mosmol kg^-1) efflux increased again and was relatively less stimulated by EDTA.It is suggested that assimilate efflux from leaf slices is mainly diffusion through open veins and/or plasmodesmata. When these symplastic connections are closed by addition of Ca^{+ +}, the remaining transmembrane flux into the apoplast is small. Thus, assimilate movement from the mesophyll to the phloem appears to be symplastic, not apoplastic as suggested in the literature.     

Stromal acidification mediates in vivo water stress inhibition of nonstomatal-controlled photosynthesis

Stromal acidification has been reported to mediate reduced osmotic potential (ψ_π) effects on photosynthesis in the isolated spinach chloroplast (Berkowitz, Gibbs 1983 Plant Physiol 72: 1100-1109). To determine if stromal acidification mediates osmotic dehydration inhibition of photosynthesis in vivo, the effects of a weak base (NH₄Cl), which raises stromal pH, on CO₂ fixation of vacuum-infiltrated spinach leaf slices, Chlamydomonas reinhardii cells and Aphanocapsa 6308 cells under isotonic and dehydrating conditions were investigated. Five millimolar NH₄Cl stimulated spinach leaf slice CO₂ fixation by 43% under stress (0.67 molar sorbitol) conditions, and had little effect on fixation under isotonic (0.33 molar sorbitol) conditions. Chlamydomonas cells were found to be more sensitive to reduced ψ_π than spinach leaf slices. CO₂ fixation in the cells of the green alga Chlamydomonas reinhardii was 99 and 17 micromoles per milligram chlorophyll per hour, respectively, at 0.1 molar mannitol and 0.28 molar mannitol. Five millimolar NH₄Cl stimulated CO₂ fixation of Chlamydomonas cells by 147% under stress (0.28 molar mannitol) conditions. Aphanocapsa 6308 cells (blue-green alga) were also found to be sensitive to reduced ψ_π, and inhibitions in photosynthesis were partially reversed by NH₄Cl. These data indicate that in vivo water stress inhibition of photosynthesis is facilitated by stromal acidification, and that this inhibition can be at least partially reversed in situ.     

Photosynthesis and ion content of leaves and isolated chloroplasts of salt-stressed spinach

Spinach (Spinacia oleracea) plants were subjected to salt stress by adding NaCl to the nutrient solution in increments of 25 millimolar per day to a final concentration of 200 millimolar. Plants were harvested 3 weeks after starting NaCl treatment. Fresh and dry weight of both shoots and roots was decreased more than 50% compared to control plants but the salt-stressed plants appeared healthy and were still actively growing. The salt-stressed plants had much thicker leaves. The salt-treated plants osmotically adjusted to maintain leaf turgor. Leaf K⁺ was decreased but Na⁺ and Cl⁻ were greatly increased.

The potential photosynthetic capacity of the leaves was measured at saturating CO₂ to overcome any stomatal limitation. Photosynthesis of salt-stressed plants varied only by about 10% from the controls when expressed on a leaf area or chlorophyll basis. The yield of variable chlorophyll a fluorescence from leaves was not affected by salt stress. Stomatal conductance decreased 70% in response to salt treatment.

Uncoupled rates of electron transport by isolated intact chloroplasts and by thylakoids were only 10 to 20% below those for control plants. CO₂-dependent O₂ evolution was decreased by 20% in chloroplasts isolated from salt-stressed plants. The concentration of K⁺ in the chloroplast decreased by 50% in the salt-stressed plants, Na⁺ increased by 70%, and Cl⁻ increased by less than 20% despite large increases in leaf Na⁺ and Cl⁻.

It is concluded that, for spinach, salt stress does not result in any major decrease in the photosynthetic potential of the leaf. Actual photosynthesis by the plant may be reduced by other factors such as decreased stomatal conductance and decreased leaf area. Effective compartmentation of ions within the cell may prevent the accumulation of inhibitory levels of Na⁺ and Cl⁻ in the chloroplast.

     

Osmoregulation in Cotton in Response to Water Stress : III. Effects of Phosphorus Fertility

Cotton (Gossypium hirsutum) (L.) was grown in a sand and nutrient solution system at two levels of phosphorus (0.5 and 5.0 millimolar). Within each phosphorus treatment, plants were either watered daily or acclimated to water stress by subjection to several water stress cycles.

Stress acclimation increased leaf starch at the low phosphorus level, but not at the high phosphorus level. High phosphorus increased leaf sucrose and glucose concentration in both acclimated and nonacclimated plants, but had little effect on osmotic adjustment or the relationship between turgor and water potential.

In nonacclimated plants, high phosphorus increased both leaf conductance and photosynthesis at high water potentials. In acclimated plants, high phosphorus increased photosynthesis but decreased conductance, thus increasing water use efficiency at the single leaf level.

     

Maintenance of photosynthesis at low leaf water potential in wheat : role of potassium status and irrigation history

The interaction of low water potential effects on photosynthesis, and leaf K⁺ levels in wheat (Triticum aestivum L.) plants was studied. Plants were grown at three K⁺ fertilization levels; 0.2, 2, and 6 millimolar. With well watered plants, 2 millimolar K⁺ supported maximal photosynthetic rates; 0.2 millimolar K⁺ was inhibitory, and 6 millimolar K⁺ was superoptimal (i.e. rates were no greater than at 2 millimolar K⁺). Photosynthesis was monitored at high (930 parts per million) and low (330 parts per million) external CO₂ throughout a series of water stress cycles. Plants subjected to one stress cycle were considered nonacclimated; plants subjected to two successive cycles were considered acclimated during the second cycle. Sensitivity of photosynthesis to declining leaf water potential was affected by K⁺ status; 6 millimolar K⁺ plants were less sensitive, and 0.2 millimolar K⁺ plants were more sensitive than 2 millimolar K⁺ plants to declining water potential. This occurred with nonacclimated and acclimated plants at both high and low assay CO₂. It was concluded that the K⁺ effect on photosynthesis under stress was not mediated by treatment effects on stomatal resistance. Differences between the K⁺ treatments were much less pronounced, however, when photosynthesis of nonacclimated and acclimated plants was plotted at a function of declining relative water content during the stress cycles. These results suggest that K⁺ effects on the relationship between relative water content and water potential in stressed plants was primarily responsible for the bulk of the K⁺-protective effect on photosynthesis in stressed plants. In vitro experiments with chloroplasts and protoplasts isolated from 2 millimolar K⁺ and 6 millimolar K⁺ plants indicated that upon dehydration, K⁺ efflux from the chloroplast stroma into the cytoplasm is less pronounced in 6 millimolar K⁺ protoplasts.     

Changes in the Water Balance and Photosynthesis of Onion, Bean and Cotton Plants under Saline Conditions

The osmotic concentration (osmotic potential) of onion leaf sap did not adjust to chloride salinity, and consequently water potential, turgor, stomatal aperture and transpiration were reduced. Although osmotic concentration of bean and cotton leaf sap did adjust to a saline root medium and turgor was no less in the salinized plants than in the controls, stomata of the salinized plants remained only partly open and transpiration was reduced. Net photosynthesis of onion plants was reduced by salinity (this effect being much enhanced in a hot dry atmosphere) but it could be rapidly raised to the level of the controls by inducing elevated leaf turgor. Stomatal closure was initially responsible for most of the ～30 % reduction in photosynthesis of salinized beans. This was due to interference with CO₂ diffusion and could be overcome by raising the CO₂ concentration in the air. At a later stage of growth, salinity affected the light reaction of bean photosynthesis, and elevation of the air CO₂ had little effect. Closure of stomata of salinized cotton plants had only a relatively small effect on net photosynthesis. Light intensity and CO₂ concentration experiments showed that salinity was reducing the photosynthesis of cotton leaves mainly by affecting the light reaction of photosynthesis. It is concluded that chloride salinity does affect the water balance and rate of photosynthesis of plants and that the nature and degree of the effects will depend upon climatic conditions and may be very different between plant species and in the same species at different periods of growth.     

Osmotic adjustment by intact isolated chloroplasts in response to osmotic stress and its effect on photosynthesis and chloroplast volume

Spinach leaf chloroplasts isolated in isotonic media (330 millimolar sorbitol, −1.0 megapascals osmotic potential) had optimum rates of photosynthesis when assayed at −1.0 megapascals. When chloroplasts were isolated in hypertonic media (720 millimolar sorbitol, −2.0 megapascals osmotic potential) the optimum osmotic potential for photosynthesis was shifted to −1.8 megapascals and the chloroplasts had higher rates of CO₂-dependent O₂ evolution than chloroplasts isolated in 330 millimolar sorbitol when both were assayed at high solute concentrations.

Transfer of chloroplasts isolated in 330 millimolar sorbitol to 720 millimolar sorbitol resulted in decreased chloroplast volume but this shrinkage was only transient and the chloroplasts subsequently swelled so that within 2 to 3 minutes at 20°C the chloroplast volume had returned to near the original value. Thus, actual steady state chloroplast volume was not decreased in hypertonic media. In isotonic media, there was a slow but significant uptake of sorbitol by chloroplasts (10 to 20 micromoles per milligram chlorophyll per hour at 20°C). Transfer of chloroplasts from 330 millimolar sorbitol to 720 millimolar sorbitol resulted in rapid uptake of sorbitol (up to 280 micromoles per milligram chlorophyll per hour at 20°C) and after 5 minutes the concentration of sorbitol inside the chloroplasts exceeded 500 millimolar. This uptake of sorbitol resulted in a significant underestimation of chloroplast volume unless [¹⁴C]sorbitol was added just prior to centrifuging the chloroplasts through silicone oil. Sudden exposure to osmotic stress apparently induced a transient change in the permeability of the chloroplast envelope since addition of [¹⁴C]sorbitol 3 minutes after transfer to hypertonic media (when chloroplast volume had returned to normal) did not result in rapid uptake of labeled sorbitol.

It is concluded that chloroplasts can osmotically adjust in vitro by uptake of solutes which do not normally penetrate the chloroplast envelope, resulting in a restoration of normal chloroplast volume and partially preventing the inhibition of photosynthesis by high solute concentrations. The results indicate the importance of matching the osmotic potential of isolation media to that of the tissue, particularly in studies of stress physiology.

     

Response of Photosynthesis and Dark-CO2-Fixation to Light, CO2 and Temperature in Leaf Slices under Osmotic Stress

Photosynthesis and dark-CO₂-fixation were measured in vacuum-infiltratedleaf slices from the mesophyte Spinacia oleracea and the Mediterraneanxerophyte Arbutus unedo under hypertonic stress as a functionof light-intensity, CO₂-concentration and temperature, in theabsence of stomatal control. Under hypertonic stress, photosynthesis and dark-CO₂-fixationwere inhibited in leaf tissue from both plants. 50% inhibitionof photosynthesis in spinach occurred at about –3.0 MPa,and of dark-CO₂-fixation at about –3.5 MPa. 50% inhibitionof photosynthesis in Arbutus unedo was reached at about –4.0MPa (sorbitol as osmoticum). In both plants, osmotic dehydration decreased the slope andthe maximum of the CO₂- and light-response curves. The slopeof the CO₂-response curve of dark-CO₂-fixation was also decreasedunder hypertonic stress, but the inhibition of the maximal fixationrate was less obvious than for photosynthesis. Photosynthesis and dark-CO₂-fixation differed significantlyin their response to high temperature: under light- and CO₂-saturation,photosynthesis of spinach leaf slices had a temperature optimumat about 37 °C, and it was nearly completely inhibited at45 °C. The rate of dark-CO₂-fixation, however, increasedcontinuously up to 45 °C. Osmotic dehydration increasedthe resistance of photosynthesis to high temperatures. Key words: CO₂ response, Heat stress, Light response, Photosynthesis, Water stress     

Reduced osmotic potential effects on photosynthesis : identification of stromal acidification as a mediating factor

Addition of sorbitol, which facilitated reductions in reaction medium osmotic potential from standard (0.33 molar sorbitol, −10 bars) isotonic conditions to a stress level of 0.67 molar sorbitol (−20 bars), inhibited the photosynthetic capacity of isolated spinach (Spinacia oleracea) chloroplasts. This inhibition, which ranged from 64 to 74% under otherwise standard reaction conditions, was dependent on reaction medium inorganic phosphate concentration, with the phosphate optimum for photosynthesis reduced to 0.05 millimolar at the low osmotic potential stress treatment from a value of 0.25 millimolar under control conditions.

Stromal alkalating agents such as NH₄Cl (0.75 millimolar) and KCl (35 millimolar) were also found to affect the degree of low osmotic potential inhibition of photosynthesis. Both agents doubled the rate of NaHCO₃-supported O₂ evolution under the stress treatment, while hardly affecting the control rate at optimal concentrations. These agents also reduced the length of the lag phase of photosynthetic O₂ evolution under the stress treatment to a much greater degree. The rate-enhancement effect of these agents under the stress treatment was reversed by sodium acetate, which is known to facilitate stromal acidification.

The reaction medium pH optimum for photosynthesis under the stress treatment was higher than under control conditions. In the presence of optimal NH₄Cl, this shift was no longer evident.

Internal pH measurements indicated that the stress treatment caused a 0.43 and 0.24 unit reduction in the stromal and intrathylakoid pH, respectively, under illumination. This osmotically induced acidification was not evident in the dark. The presence of 0.75 millimolar NH₄Cl partially reversed the osmotically induced reduction in the illuminated stromal pH. It was concluded that stromal acidification is a mediating mechanism of the most severe site of low osmotic potential inhibition of the photosynthetic process.

     

Stomatal and Nonstomatal Components to Inhibition of Photosynthesis in Leaves of Capsicum annuum during Progressive Exposure to NaCl Salinity

Young bell pepper (Capsicum annuum L.) plants grown in nutrient solution were gradually acclimated to 50, 100, or 150 moles per cubic meter NaCl, and photosynthetic rates of individual attached leaves were measured on several occasions during the salinization period at external CO₂ concentrations ranging from approximately 70 to 1900 micromoles per mole air. Net CO₂ assimilation (A) was plotted against computed leaf internal CO₂ concentration (C_i), and the initial slope of this A-C_i curve was used as a measure of photosynthetic ability. During the 10 to 14 days after salinization began, leaves from plants exposed to 50 moles per cubic meter NaCl showed little change in photosynthetic ability, whereas those treated to 100 or 150 moles per cubic meter NaCl had up to 85% inhibition, with increase in CO₂ compensation point. Leaves appeared healthy, and leaf chlorophyll content showed only a 14% reduction at the highest salinity levels. Partial stomatal closure occurred with salinization, but reductions in photosynthesis were primarily nonstomatal in origin. Photosynthetic ability was inversely related to the concentration of either Na⁺ or Cl⁻ in the leaf laminas sampled at the end of the experimental period. However, the concentration of Cl⁻ expressed on a tissue water basis was greater, exceeding 300 moles per cubic meter, and Cl⁻ was more closely associated (R² = 0.926) with the inhibition of photosynthetic ability. Leaf turgor was not reduced by salinization and leaf osmotic potential decreased to a slightly greater extent than the osmotic potential decreases of the nutrient solutions. Concentration of accumulated Na⁺ and Cl⁻ (on a tissue water basis) accounted quantitatively for maintenance of leaf osmotic balance, assuming that these ions were sequestered in the vacuoles.     

Potassium transport in two tomato species : lycopersicon esculentum and lycopersicon cheesmanii

The commercial tomato, lycopersicon esculentum Mill. cv Walter, and its wild relative, Lycopersicon cheesmanii ssp. minor (Hook.) C.H. Mull., were grown for 30 days under controlled conditions and in solution culture varying in its content of Na⁺ and K⁺. Subsequently, ⁸⁶Rb-labeled K⁺ uptake and distribution were studied. From all solutions, `Walter' consistently absorbed <sup>86</sup>Rb-labeled K<sup>+</sup> at a higher rate in micromoles per gram fresh weight per 30 minutes than L. cheesmanii. L. cheesmanii distributed a greater proportion of the absorbed K<sup>+</sup> from its root to its shoot. When 0.6 millimolar NaNO<sub>3</sub> replaced 0.6 millimolar KNO<sub>3</sub> in the pretreatment solution, intact plants of both genotypes followed a similar pattern as when they were pretreated with K<sup>+</sup> only, but a greater percentage of the absorbed K<sup>+</sup> remained in the roots. Leaf slices of L. cheesmanii plants deprived of K<sup>+</sup> for 6 days showed a greater rate of K<sup>+</sup> uptake than did slices from `Walter' plants pretreated the same way. Stem slices of L. cheesmanii, however, had a lower uptake rate than did those of `Walter'. Both leaf and stem slices of `Walter' plants, pretreated 6 days with 0.6 millimolar NaNO₃ substituting for 0.6 millimolar KNO₃ in their growth medium, had greater rates of ⁸⁶Rb-labeled K⁺ uptake from 0.5 and 20 millimolar KCl solutions than did slices of L. cheesmanii. These marked differences in patterns of ion uptake and translocation indicate that these genotypes of tomato have evolved different mechanisms to deal with K⁺ and Na⁺ in their environments.     

The chemiosmotic model of stomatal opening revisited

Stomata are light‐activated biological valves in the otherwise gas‐impermeable epidermis of aerial organs of higher plants. Stomata often regulate rates of photosynthesis and transpiration in ways that optimize whole‐plant carbon gain against water loss. Each stoma is flanked by a pair of opposing guard cells. Stomatal opening occurs by light‐activated increases in the turgor pressure of guard cells, which causes them to change shape so that the stomatal pore between them widens. These increases in turgor pressure oppose increases in cellular osmotic pressure that result from uptake of K⁺. K⁺ uptake occurs by a chemiosmotic mechanism in response to light‐activated extrusion of H⁺ outward across the plasma membrane of the guard cell. The initial changes in cellular membrane potential lead to the opening of inward‐rectifying K⁺ channels, after which K⁺ is taken up along its electrochemical gradient. Changes in membrane potential resulting from K⁺ uptake may be balanced by accumulation of Cl^?ions by guard cells and/or by synthesis of malic acid within each cell. Malic acid also acts to buffer increases in cytosolic pH caused by H⁺ extrusion. This review describes how the application of patch‐clamp technology to guard cell protoplasts has enabled investigators to elucidate the mechanisms by which H⁺ is extruded from guard cells, the types of ion channels present in the guard cell plasma membrane, how those ion channels are regulated, and the signal transduction processes that trigger stomatal opening and closing.     

Effects of Proline and Glycinebetaine on Vicia Faba Responses to Salt Stress

Plants of bean (Vicia faba L. cv. Calvor 103) were salt-stressed with NaCl and CaCl₂ in concentrations inducing soil osmotic potentials (ψ_soil) from 0 to -1.2 MPa and were sprayed with proline (8.7 μM) and glycinebetaine (8.5 μM) solutions. Bean plants respond to increasing soil salinity by decreased leaf relative water content and osmotic potential. Salinity decreased the contents of dry mass, chlorophyll, soluble and hydrolysable sugars, soluble proteins and enhanced content of total free amino acids, Na⁺, Ca²⁺ and Cl^-. The ratio of K⁺/Na⁺ was decreased on salinization. The membranes of leaf discs from salt-stressed plants appeared to be less stable under heat stress (51 °C) than that of unstressed plants. The reverse was true for discs placed under dehydration stress (40 % polyethylene glycol 6000). Proline and glycinebetaine application reduced membrane injury, improved K⁺ uptake and growth. Also both solutes increased chlorophyll contents. This revised version was published online in July 2006 with corrections to the Cover Date.     

Solute-dependent activation of cell motility in strongly hypertonic solutions in <Emphasis Type="Italic">Dictyostelium discoideum</Emphasis>, human melanoma HTB-140 cells and walker 256 carcinosarcoma cells

Published data concerning the effects of hypertonicity on cell motility have often been controversial. The interpretation of results often rests on the premise that cell responses result from cell dehydration, i.e. osmotic effects. The results of induced hypertonicity on cell movement of Dictyostelium discoideum amoebae and human melanoma HTB-140 cells reported here show that: i) hypertonic solutions of identical osmolarity will either inhibit or stimulate cell movement depending on specific solutes (Na⁺ or K⁺, sorbitol or saccharose); ii) inhibition of cell motility by hypertonic solutions containing Na⁺ ions or carbohydrates can be reversed by the addition of calcium ions; iii) various cell types react differently to the same solutions, and iv) cells can adapt to hypertonic solutions. Various hypertonic solutions are now broadly used in medicine and to study modulation of gene expression. The observations reported suggest the need to examine whether the other responses of cells to hypertonicity can also be based on the solute-dependent cell responses besides cell dehydration due to the osmotic effects.     

Phosphate cycles in energy crop systems with emphasis on the availability of different phosphate fractions in the soil

The growing cells of hydroponic maize roots expand at constant turgor pressure (0.48 MPa) both when grown in low-(0.5 mol m^-3 CaCl₂) or full-nutrient (Hoagland's) solution and also when seedlings are stressed osmotically (0.96 MPa mannitol). Cell osmotic pressure decreases by 0.1–0.2 MPa during expansion. Despite this, total solute influx largely matches the continuously-varying volume expansion-rate of each cell. K⁺ in the non-osmotically stressed roots is a significant exception-its concentration dropping by 50% regardless of the presence or absence of K⁺ in the nutrient medium. This corresponds to the drop in osmotic pressure. Nitrate appears to replace Cl^- in the Hoagland-grown cells.Analogous insensitivity of solute gradients to external solutes is observed in the radial distribution of water and solutes in the cortex 12 mm from the tip. Uniform turgor and osmotic pressures are accompanied by opposite gradients of K⁺ and Cl^-, outwards, and hexoses and amino acids, inwards, for plants grown in either 0.5 mol m^-3 CaCl₂ or Hoagland's solution (with negligible Cl^-). K⁺ and Cl^- levels within both gradients were slightly higher when the ions were available in the medium. The gradients themselves are independent of the direction of solute supply. In CaCl₂ solution all other nutrients must come from the stele, in Hoagland's solution inorganic solutes are available in the medium.24 h after osmotic stress, turgor pressure is recovered at all points in each gradient by osmotic adjustment using organic solutes. Remarkably, K⁺ and Cl^- levels hardly change, despite their ready availability. Hexoses are responsible for some 50% of the adjustment with mannitol for a further 30%. Some 20% of the final osmotic pressure remains to be accounted for. Proline and sucrose are not significantly involved. Under all conditions a standing water potential step of 0.2 MPa between the rhizodermis and its hydroponic medium was found. We suggest that this is due to solute leakage.Abbreviations EDX energy dispersive X-ray microanalysis - water potential - 11-1 cell osmotic pressure - P turgor pressure     

Role of K+ in leaf growth: K+ uptake is required for light-stimulated H+ efflux but not solute accumulation

The stimulation of dicotyledonous leaf growth by light depends on increased H⁺ efflux, to acidify and loosen the cell walls, and is enhanced by K⁺ uptake. The role of K⁺ is generally considered to be osmotic for turgor maintenance. In coleoptiles, auxin‐induced cell elongation and wall acidification depend on K⁺ uptake through tetraethylammonium (TEA)‐sensitive channels (Claussen et al., Planta 201, 227–234, 1997), and auxin stimulates the expression of inward‐rectifying K⁺ channels ( Philippar et al. 1999) . The role of K⁺ in growing, leaf mesophyll cells has been investigated in the present study by measuring the consequences of blocking K⁺ uptake on several growth‐related processes, including solute accumulation, apoplast acidification, and membrane polarization. The results show that light‐stimulated growth and wall acidification of young tobacco leaves is dependent on K⁺ uptake. Light‐stimulated growth is enhanced three‐fold over dark levels with increasing external K⁺, and this effect is blocked by the K⁺ channel blockers, TEA, Ba⁺⁺ and Cs⁺. Incubation in 10 mm TEA reduced light‐stimulated growth and K⁺ uptake by 85%, and completely inhibited light‐stimulated wall acidification and membrane polarization. Although K⁺ uptake is significantly reduced in the presence of TEA, solute accumulation is increased. We suggest that the primary role of K⁺ in light‐stimulated leaf growth is to provide electrical counterbalance to H⁺ efflux, rather than to contribute to solute accumulation and turgor maintenance.     

Effects of Alkalinization and ATPase Inhibition on Stromal Free Mg2+ Concentration in Spinach Chloroplasts

Our earlier studies indicate that stromal alkalinization is essential for light-induced increase in free Mg²⁺ concentration ([Mg²⁺]) in chloroplast. Stromal [Mg²⁺] was increased by dark incubation of chloroplasts in the K⁺-gluconate medium (pH 8.0), or by NH₄Cl. These results indicate that stromal alkalinization can induce an increase in stromal [Mg²⁺] without illumination. Some inhibitors of envelope proton-translocating ATPase activity involved in H⁺ efflux inhibited the alkalinization-induced increase in [Mg²⁺].     

Reduced osmotic potential inhibition of photosynthesis : site-specific effects of osmotically induced stromal acidification

The effects of reduced reaction medium osmotic potential (0.67 molar sorbitol as compared to a control treatment with 0.33 molar sorbitol) on the enzymic steps of the photosynthetic carbon reduction cycle were investigated using isolated spinach (Spinacia oleracea L. var Longstanding Bloomsdale) chloroplasts. Reversal of reduced osmotic potential inhibition of photosynthetic rates by a stromal alkalating agent (NH₄Cl) was associated with specific steps of the cycle. Low osmotic potential induced stromal acidification was found to be facilitated by osmotically induced chloroplast shrinkage. However, the action of the alkalating agent was found not to be associated with reversal of osmotically induced morphological changes of the stromal compartment.

Labeled metabolite analyses indicated that the osmotic stress treatment caused the substrate for fructose 1,6-bisphosphatase (FBPase) to build up in the absence of NH₄Cl, and the substrate for phosphoribulokinase to increase in the presence of NH₄Cl. These data were interpreted as indicating that the most severe effect of osmotic stress on photosynthesis is at the site of FBPase, and that this inhibition is mediated by osmotically induced stromal acidification. Phosphoribulokinase activity inhibition at the low osmotic potential treatment was apparently less severe and not mediated by stromal acidification. A third site of osmotic inhibition, which was reversed by NH₄Cl, and therefore was assumed to be mediated by stromal acidification, was at the step of ribulose 1,5-bisphosphate carboxylase.

Additions of NH₄Cl also enhanced the activity of the pH-insensitive phase of the photosynthetic carbon reduction cycle, 3-phosphoglyceric acid reduction, at the stress treatment. This effect was thought to be mediated by the removal of the block at FBPase. A model was proposed to outline the relative severity of osmotic stress effects at various sites of the photosynthetic carbon reduction cycle.