Compartment analysis of plant cells by means of turgor pressure relaxation: I. Theoretical considerations

Summary Turgor pressure relaxation curves for individual plant cells represent an important source of information for the plant physiologist. However, the accurate interpretation of these curves is strongly dependent on the model chosen to describe the plant cell. If the compartmentation of the cell into vacuole and cytoplasm is taken into account, a theoretical analysis shows that pressure relaxation curves can be represented by the sum of two exponential functions. Givena priori assumptions about the exchange area of the tonoplast and its reflection coefficient, the hydraulic conductivities of the plasmalemma and tonoplast can be determined and the proportion of the total cell volume occupied by the cytoplasm is also obtained. Numerical solutions to the flow equations have shown that the biphasic nature of pressure relaxations is maintained even when a permeable tonoplast is assumed. Depending on the magnitude of the reflection coefficient and the permeability of the vacuolar membrane, large errors can arise in the determination of the hydraulic conductivity of the tonoplast. However, under certain conditions, even a highly permeable tonoplast may behave like a nonpermeable membrane during pressure relaxation.     

Current-voltage relationships of a sodium-sensitive potassium channel in the tonoplast ofChara corallina

Summary The membrane of mechanically prepared vesicles ofChara corallina has been investigated by patch-clamp techniques. This membrane consists of tonoplast as demonstrated by the measurement of ATP-driven currents directed into the vesicles as well as by the ATP-dependent accumulation of neutral red. Addition of 1mm ATP to the bath medium induced a membrane current of about 3.2 mA·m^–2 creating a voltage across the tonoplast of about –7 mV (cytoplasmic side negative). On excised tonoplast patches, currents through single K⁺-selective channels have been investigated under various ionic conditions. The open-channel currents saturate at large voltage displacements from the equilibrium voltage for K⁺ with limiting currents of about +15 and –30 pA, respectively, as measured in symmetric 250mm KCl solutions. The channel is virtually impermeable to Na⁺ and Cl^–. However, addition of Na⁺ decreases the K⁺ currents. TheI–V relationships of the open channel as measured at various K⁺ concentrations with or without Na⁺ added are described by a 6-state model, the 12 parameters of which are determined to fit the experimental data.     

Hydrostatic pressure mimics gravitational pressure in characean cells

Summary Hydrostatic pressure applied to one end of a horizontalChara cell induces a polarity of cytoplasmic streaming, thus mimicking the effect of gravity. A positive hydrostatic pressure induces a more rapid streaming away from the applied pressure and a slower streaming toward the applied pressure. In contrast, a negative pressure induces a more rapid streaming toward and a slower streaming away from the applied pressure. Both the hydrostatic pressure-induced and gravity-induced polarity of cytoplasmic streaming respond identically to cell ligation, UV microbeam irradiation, external Ca²⁺ concentrations, osmotic pressure, neutral red, TEA Cl^–, and the Ca²⁺ channel blockers nifedipine and LaCl₃. In addition, hydrostatic pressure applied to the bottom of a vertically-oriented cell can abolish and even reverse the gravity-induced polarity of cytoplasmic streaming. These data indicate that both gravity and hydrostatic pressure act at the same point of the signal transduction chain leading to the induction of a polarity of cytoplasmic streaming and support the hypothesis that characean cells respond to gravity by sensing a gravity-induced pressure differential between the cell ends.     

Membrane orientation of droplets prepared fromChara corallina internodal cells

Summary It is generally accepted that the membrane surrounding droplets from characean cells originates from the tonoplast, but there is some uncertainty regarding droplet membrane sidedness. This issue was addressed directly by combining two different droplet isolation methods and the patch clamp technique. Neutral red accumulation was used to demonstrate the presence of H⁺-transport over the membrane and to predict membrane orientation. Two types of droplet populations with differently oriented membranes could be formed in an iso-osmotic bath solution. Cytoplasmic droplets (cytosolic side of the tonoplast inside) contained cytoplasm, while the second type of droplet population contained vacuolar sap (vacuolar droplets, vacuolar side of the tonoplast inside). Smaller vesicles also appeared inside the droplets, with an apparently inversely oriented membrane. Confocal laser scanning microscopy indirectly demonstrated that, at least with one of the droplet isolation methods, the plasma membrane entirely remains in the internodal cell after intracellular perfusion. Both types of droplet populations allowed the formation of excised patches and single-channel measurements by the patch clamp technique. Properties of anion channels in the tonoplast could be used to prove the predicted membrane orientation, knowing that Ca²⁺ can only activate these channels from the cytosolic side. These results provide useful data for studies addressing ligand-binding, block and modulation, organization and interaction of proteins within the membrane or with other regulatory factors, where it is important to control membrane orientation.     

Generation of action potentials in Chara corallina by turgor pressure changes

A depolarisation of the membrane potential difference (about-170 mV) of Chara corallina is observed in response to changes in cell turgor pressure using the pressure probe technique. The depolarisation occurs in phase with the pressure pulse (0.2 s duration) and is independent of the direction of the applied pressure gradient. This latter finding is in contradiction to results predicted on the basis of electro-kinetic phenomena. Pressure induced electrical leakages were ruled out by measuring the depolarisation in response to pressure in individual segments of the internode which were electrically isolated from one another. The changes in potential were recorded by external electrodes and an internal electrode which was positioned close to the micropipette of the pressure probe inserted through one of the electrically isolated nodes. The depolarisation in response to increasing positive or negative pressure gradients in the intact node region and in the intact middle segments was comparable to that monitored in the node region containing the pressure probe. Action potentials were initiated when the pressure gradients exceeded at least 2 bar. The action potentials were elicited at random in one of the two compartments adjacent to the node regions, but were never found to be initiated in the node regions themselves. The pressure-induced action potentials are explained in terms of an electro-mechanical compression (or expansion) of a local membrane area and discussed in their relevance to the propagation of pressure signals in response to water and salt stress in higher plants.Abbreviations PD potential difference     

Direct turgor pressure measurements in individual leaf cells of Tradescantia virginiana

The water relations of leaves of Tradescantia virginiana were studied using the miniaturized pressure probe (Hüsken, E. Steudle, Zimmermann, 1978 Plant Physiol. 61, 158–163). Under well-watered conditions cell turgor pressures, P _o, ranged from 2 to 8 bar in epidermal cells. In subsidiary cells P _o was about 1.5 to 4.5 bar and in mesophyll cells about 2 to 3.5 bar. From the turgor pressure, relaxation induced in individual cells by changing the turgor pressure directly by means of the pressure probe, the half-time of water exchange was measured to be between 3 and 100 s for the epidermal, subsidiary, and mesophyll cells. The volumetric elastic modulus, , of individual cells was determined by changing the cell volume by a defined amount and simultaneously measuring the corresponding change in cell turgor pressure. The values for the elastic modulus for epidermal, subsidiary, and mesophyll cells are in the range of 40 to 240 bar, 30 to 200 bar, and 6 to 14 bar, respectively. Using these values, the hydraulic conductivity, L _p, for the epidermal, subsidiary, and mesophyll cells is calculated from the turgor pressure relaxation process (on the basis of the thermodynamics of irreversible processes) to be between 1 and 55·10^-7 cm s^-1 bar^-1. The data for the volumetric elastic modulus of epidermal and subsidiary cells indicate that the corresponding elastic modulus for the guard cells should be considerably lower due to the large volume changes of these cells during opening or closing. Recalculation of experimental data obtained by K. Raschke (1979, Encycl. Plant Physiol. N.S., vol. 7, pp 383–441) on epidermal strips of Vicia faba indicates that the elastic modulus of guard cells of V. faba is in the order of 40–80 bar for closed stomata. However, with increasing stomatal opening, i.e., increasing guard cell volume, decreases. Therefore, in our opinion Raschke's results would indicate a relationship between guard cell volume and which would be inverse to that for plant cells known in the literature. assumes values between 20–40 bar when the guard cell colume is soubled.     

Flowers under pressure: ins and outs of turgor regulation in development

Background

Turgor pressure is an essential feature of plants; however, whereas its physiological importance is unequivocally recognized, its relevance to development is often reduced to a role in cell elongation.

Scope

This review surveys the roles of turgor in development, the molecular mechanisms of turgor regulation and the methods used to measure turgor and related quantities, while also covering the basic concepts associated with water potential and water flow in plants. Three key processes in flower development are then considered more specifically: flower opening, anther dehiscence and pollen tube growth.

Conclusions

Many molecular determinants of turgor and its regulation have been characterized, while a number of methods are now available to quantify water potential, turgor and hydraulic conductivity. Data on flower opening, anther dehiscence and lateral root emergence suggest that turgor needs to be finely tuned during development, both spatially and temporally. It is anticipated that a combination of biological experiments and physical measurements will reinforce the existing data and reveal unexpected roles of turgor in development.     

Responses of elongation growth rate,turgor pressure and cell wall extensibility of stem cells of Impatiens balsamina to lead,cadmium and zinc

Elongation growth rate of stem cells of Impatiens balsamina was inhibited by the heavy metals Pb²⁺, Cd²⁺ and Zn²⁺ due to their suppression on cell wall extensibility. Effective turgor was also inhibited by Pb²⁺ and Cd²⁺ but it played a secondary role in reducing the stem cell elongation growth rate. The major rate-limiting factor for cell elongation growth was the cell wall extensibility. Furthermore, Cd²⁺ was found to be more toxic than Pb²⁺, while Pb²⁺ was more toxic than Zn²⁺.     

Behavior of protoplasm for survival in injured cells ofValonia ventricosa: involvement of turgor pressure

Summary By injuring cells ofValonia ventricosa, one of two survival strategies — wound-healing and protoplast formation — is induced. The present study revealed that turgor pressure, as well as Ca²⁺ in bathing medium, is involved in the choice between these survival strategies. On the process of wound-healing, turgor pressure is recovered in the presence of both the wound plug, which closes the wound immediately after an injury, and the aggregation of protoplasm around the wound, which serves to protect the inflow of outer medium into the protoplasm layer and also to strengthen the wound plug. When the size of the wound is more than 150 m in diameter, the protoplasmic aggregate strengthen the wound plug incompletely and, as a result, wound-healing is unsuccessful. In this case, the ejection of vacuolar sap is repeated, due to partial restoration of turgor pressure. In each ejection, the wound plug is blown off, together with the aggregated protoplasm and, after several ejections are repeated, the cell is unable to heal the wound because of a lack of protoplasm around the wound. Continuous depression of turgor pressure, during the repeat of the unsuccessful wound-healing, induces disorganization of the protoplasm layer. Under these conditions, the centrifugal propagation of protoplasmic aggregation, followed by the protoplasts formation, takes place easily. Effects of turgor pressure and Ca²⁺ in the bathing medium upon the wound healing is discussed and the cytoplasmic behavior for survival of wounded cells is presented schematically.     

Transport properties of mobile charges in algal membranes: Influence of pH and turgor pressure

Summary Charge-pulse experiments were performed on giant algal cells ofValonia utricularis. If the tonoplast and plasmalemma in series are charged to voltages of the order of 10mV, the decay of the initial voltage with time can be described by the sum of two or three exponential relaxations. It is not possible to explain the exponential decay of the voltage by twoRC-circuits in series (e.g. tonoplast and plasmalemma), because this would lead to unreasonable values for the specific capacities of the two membranes. The exponential relaxations might be attributable to the transport of mobile negative charges present in both membranes, possibly as a part of a transport system. From an analysis of the experimental results in terms of the proposed model, the translocation rate constantk and the total surface densityN _t of the mobile charges in one membrane could be evaluated. On averagek is of the order of 600 sec^–1 andNt is about 5×10^–12 mol cm^–2 (average turgor pressure 1.6 bar). The transport properties of the mobile charges within the tonoplast and plasmalemma were studied as a function of different parameters such as external pH, glutardialdehyde, electrical breakdown and turgor pressure. When the pH is lowered from 8.2 to 4 or 5 the mobile charges disappear completely, presumably as the result of protonation of the anionic groups. This pH effect was found to be completely reversible. Electrical breakdown causes a reversible disappearance of the relaxation with the longer half-time due to the decrease in membrane resistance. The value of the electrical breakdown voltage determined by injection of charge pulses of 300-sec duration into the cell is pH-independent and therefore is consistent with the mobile charge model and with results previously reported (U. Zimmermann & R. Benz.J. Membrane Biol 53:33–43, 1980). Addition of glutardialdehyde leads also to a disappearance of the mobile charges probably due to cross-linkage. Increase of the turgor pressure from 0.05 bar to 2 bar results in an increase ink by a factor of 2 and inNt by about 30%. The increase ink is in reasonable agreement with that expected on the basis of the assumed compressibility of the membranes. The elastic compressive modulus perpendicular to the membrane plane calculated from the pressure dependence of the translocation rate constantk is in very good agreement with that derived from electrical breakdown experiments (14 and 13 bar, respectively). The presence of charges within the membranes as well as the compressibility of the membranes are discussed in terms of a possible turgor-pressure-sensing mechanism.     

The relationship between turgor pressure and titratable acidity in mesophyll cells of intact leaves of a Crassulacean-acid-metabolism plant,Kalanchoe daigremontiana Hamet et Perr

Day/night changes in turgor pressure (P) and titratable acidity content were investigated in the (Crassulacean-acid-metabolism (CAM) plant Kalanchoe daigremontiana. Measurements of P were made on individual mesophyll cells of intact attached leaves using the pressure-probe technique. Under conditions of high relative humidity, when transpiration rates were minimal, changes in P correlated well with changes in the level of titratable acidity. During the standard 12 h light/12 h dark cycle, maximum turgor pressure (0.15 MPa) occurred at the end of the dark period when the level of titratable acidity was highest (about 300 eq H⁺·g^-1 fresh weight). A close relationship between P and titratable acidity was also seen in leaves exposed to perturbations of the standard light/dark cycle. (The dark period was either prolonged, or else only CO₂-free air was supplied in this period). In plants deprived of irrigation for five weeks, diurnal changes in titratable acidity of the leaves were reduced (H=160 eq H⁺·g^-1 fresh weight) and P increased from essentially zero at the end of the light period to 0.02 MPa at the end of the dark period. Following more severe water stress (experiments were made on leaves which had been detached for five weeks), P was zero throughout day and night, yet small diurnal changes in titratable acidity were still measured. These findings are discussed in relation to a hypothesis by Lüttge et al. 1975 (Plant Physiol. 56,613-616) for the role of P in the regulation of acidification/de-acidification cycles of plants exhibiting CAM.Abbreviations CAM crassulacean acid metabolism - FW fresh weight - P turgor pressure     

Comparative measurements of the xylem pressure ofNicotiana plants by means of the pressure bomb and pressure probe

Determination of the pressure in the water-conducting vessels of intactNicotiana rustica L. plants showed that the pressure probe technique gave less-negative values than the Scholander-bomb method. Even though absolute values of the order of −0.1 MPa could be directly recorded in the xylem by means of the pressure probe, pressures between zero and atmospheric were also frequently found. The data obtained by the pressure probe for excised leaves showed that the Scholander bomb apparently did not read the actual tension in the xylem vessles ofNicotiana plants. The possibility that the pressure probe gave false readings was excluded by several experimental controls. In addition, cavitation and leaks either during the insertion of the microcapillary of the pressure probe, or else during the measurements were easily recognized when they occurred because of the sudden increase of the absolute xylem tension to that of water vapour or to atmospheric, respectively. Tension values of the same order could also be measured by means of the pressure probe in the xylem vessels of pieces of stem cut from leaves and roots under water and clamped at both ends. The magnitude of the absolute tension depended on the osmolarity of the bathing solution which was adjusted by addition of appropriate concentrations of polyethylene glycol. Partial and uniform pressurisation of plant tissues or organs, or of entire plants (by means of the Scholander bomb or of a hyperbaric chamber, respectively) and simultaneous recording of the xylem tension using the pressure probe showed that a 1∶1 response in xylem pressure only occurred under a few circumstances. A 1∶1 response required that the xylem vessels were in direct contact with an external water reservoir and/or that the tissue was (pre-)infiltrated with water. Corresponding pressure-probe measurements in isolated vascular bundles ofPlantago major L. orP. lanceolata L. plants attached to a Hepp-type osmometer indicated that the magnitude of the tension in the xylem vessels was determined by the external osmotic pressure of the reservoir. These and other experiments, as well as analysis of the data using classical thermodynamics, indicated that the turgor and the internal osmotic pressure of the accessory cells along the xylem vessels play an important role in the maintenance of a constant xylem tension. This conclusion is consistent with the cohesion theory. In agreement with the literature (P.E. Weatherley, 1976, Philos. Trans. R. Soc. London Ser. B23, 435–444; 1982, Encyclopedia of plant physiology, vol. 12B, 79-109), it was found that the tension in the xylem of intact plants under normal and elevated ambient pressure (as measured with the pressure probe) under quasi-stationary conditions was independent of the transpiration rate over a large range, indicating that the conductance of the flow path must be flow-dependent.     

Experimental evidence for negative turgor pressure in small leaf cells of Robinia pseudoacacia L versus large cells of Metasequoia glyptostroboides Hu et W.C.Cheng. 1. Evidence from pressure‐volume curve analysis of dead tissue.

This paper provides a mini‐review of evidence for negative turgor pressure in leaf cells starting with experimental evidence in the late 1950s and ending with biomechanical models published in 2014. In the present study, biomechanical models were used to predict how negative turgor pressure might be manifested in dead tissue, and experiments were conducted to test the predictions. The main findings were as follows: (i) Tissues killed by heating to 60 or 80 °C or by freezing in liquid nitrogen all became equally leaky to cell sap solutes and all seemed to pass freely through the cell walls. (ii) Once cell sap solutes could freely pass the cell walls, the shape of pressure‐volume curves was dramatically altered between living and dead cells. (iii) Pressure‐volume curves of dead tissue seem to measure negative turgor defined as negative when inside minus outside pressure is negative. (iv) Robinia pseudoacacia leaves with small palisade cells had more negative turgor than Metasequoia glyptostroboides with large cells. (v) The absolute difference in negative turgor between R. pseudoacacia and M. glyptostroboides approached as much as 1.0 MPa in some cases. The differences in the manifestation of negative turgor in living versus dead tissue are discussed.     

Experimental evidence for negative turgor pressure in small leaf cells of Robinia pseudoacacia L versus large cells of Metasequoia glyptostroboides Hu et W.C. Cheng. 2. Höfler diagrams below the volume of zero turgor and the theoretical implication for pressure‐volume curves of living cells

The physiological advantages of negative turgor pressure, P_t, in leaf cells are water saving and homeostasis of reactants. This paper advances methods for detecting the occurrence of negative P_t in leaves. Biomechanical models of pressure‐volume (PV) curves predict that negative P_t does not change the linearity of PV curve plots of inverse balance pressure, P_B, versus relative water loss, but it does predict changes in either the y‐intercept or the x‐intercept of the plots depending on where cell collapse occurs in the P_B domain because of negative P_t. PV curve analysis of Robinia leaves revealed a shift in the x‐intercept (x‐axis is relative water loss) of PV curves, caused by negative P_t of palisade cells. The low x‐intercept of the PV curve was explained by the non‐collapse of palisade cells in Robinia in the P_B domain. Non‐collapse means that P_t smoothly falls from positive to negative values with decreasing cell volume without a dramatic change in slope. The magnitude of negative turgor in non‐collapsing living cells was as low as ?1.3 MPa and the relative volume of the non‐collapsing cell equaled 58% of the total leaf cell volume. This study adds to the growing evidence for negative P_t.     

Electrical tolerance (breakdown) of theChara corallina plasmalemma: I. Necessity of Ca2+

Summary The relationship between the external Ca²⁺ concentrations [Ca²⁺]₀ and the electrical tolerance (breakdown) in theChara plasmalemma was investigated. When the membrane potential was negative beyond –350–400 mV (breakdown potential, BP), a marked inward current was observed, which corresponds to the so-called punch-through (H.G.L. Coster,Biophys. J. 5:669–686, 1965). The electrical tolerance of theChara plasmalemma depended highly on [Ca²⁺]₀. Increasing [Ca²⁺]₀ caused a more negative and decreasing it caused a more positive shift of BP. BP was at about –700 mV in 200 M La³⁺ solution. [Mg²⁺]₀ depressed the membrane electrical tolerance which was supposed to be due to competition with Ca²⁺ at the Ca²⁺ binding site of the membrane. Such a depressive effect of Mg²⁺ was almost masked when the [Ca²⁺]₀/[Mg²⁺]₀ ratio was roughly beyond 2.     

Long-term xylem pressure measurements in the lianaTetrastigma voinierianum by means of the xylem pressure probe

Diurnal changes of xylem pressure in the lianaTetrastigma voinierianum have been measured under greenhouse conditions by means of the recently developed xylem pressure probe. During the early morning hours, tensions in the vessels developed more or less rapidly with time, depending on light intensity. On sunny days, absolute negative pressures down to about -0.4 MPa (atmospheric = 0.1 MPa) were recorded around noon in petiolar or stem xylem vessels, whereas on rainy or cloudy days the xylem pressure remained in the positive sub-atmospheric or slightly negative pressure range. Towards the evening the tension in the vessels always decreased, i.e. the xylem pressure shifted to about atmospheric, or even above-atmospheric, values during the night. Simultaneous xylem pressure recordings at heights of 1 and 5 m frequently yielded either no gradient in tension at all, or far less than expected from the Cohesion Theory. Occasionally, tension gradients were even opposite to those predicted by this theory. Stem-toleaves pressure gradients in accord with the Cohesion Theory were recorded only when tension had been developed during sunny days in the upper branches of the liana, because increases in tension were not immediately propagated to the xylem of the leaves at ground level, as would be expected from a strictly coupled hydraulic system. Parallel recordings of the xylem tension using the pressure chamber yielded rather variable values ranging from 0.1 to 1 MPa; diurnal pressure changes could not be detected at all. The data are discussed on the basis of the equation for the chemical activity of water. They strongly suggest that the xylem tension induced by transpiration is not the sole force for water ascent. Other forces, such as osmotic pressure or convectional and interfacial forces, which to a remarkable extent have already been postulated for decades, seem to be equally important.Abbreviation R.H. relative humidity The authors are very grateful to Professor D. Fürnkranz, Institut für Botanik der Universität Salzburg, for his interest and help with the greenhouse facility, to Walter Gigerl for expert technical assistance, to Heike Schneider and Notburga Gierlinger for the petiolestaining experiments. This work was supported by a grant of the Deutsche Forschungsgemeinschaft to U.Z. (NMR-Graduiertenkolleg Ha 1232/8-1).     

A kinetic analysis of the electrogenic pump ofChara corallina: II. Dependence of the pump activity on external pH

Summary The current-voltage curve of theChara membrane was obtained by applying a slow ramp de- and hyperpolarization by use of voltage clamp. By inhibiting the electrogenic pump with 50m DCCD (dicyclohexylcarbodiimide), theI–V curve approached a steady state within 100 min, which gave thei _d -V curve of the passive diffusion channel. Thei _p -V curve of the electrogenic pump channel was obtained by subtracting the latter from the former. With the increase of external pH, thei _d -V curve showed only a slight change, while thei _p -V curve of the pump channel showed almost a parallel shift, in the hyperpolarizing direction, along the voltage axis in the pH range between 6.5 and 7.5. The sigmoidali _p -V curve in this pH range could be simulated satisfactorily with the five-state model reported previously (U. Kishimoto, N. Kami-ike, Y. Takeuchi & T. Ohkawa,J. Membrane Biol. 80:175–183, 1984) as well as with a lumped two-state model presented in this report. The analysis based on these models suggests that the electrogenic pump of theChara membrane is mainly a 2H⁺/1ATP pump. The forward rate constant in the voltage-dependent step increased with the increase of external pH, while the backward one decreased. On the other hand, the forward rate constant in the voltage-independent step remained almost unchanged with the increase of external pH, while the backward one increased markedly. The pump conductance at the resting membrane potential showed either a slight increase or a decrease with the increase of external pH, depending on the sample. Nevertheless, the pump current showed generally a slight increase with the increase of external pH.     

A kinetic analysis of the electrogenic pump ofChara corallina: I. Inhibition of the pump by DCCD

Summary The current-voltage curve of theChara membrane was obtained by applying a slow ramp depo- and hyperpolarization by use of voltage clamp. With the progress of poisoning by DCCD (dicyclohexylcarbodiimide) theI–V curve moved by about 50 mV (depolarization) along the voltage axis, reducing its slope, and finally converged to thei _d -V curve of the passive diffusion channel. Changes ofi _p -V curve of the electrogenic pump channel could be obtained by subtracting the latter from the former.The sigmoidali _p -V curve could be simulated satisfactorily by adopting a simple reaction kinetic model. Kinetic parameters of the successive changes of state of the H⁺ ATPase could be evaluated. Changes of these kinetic parameters during inhibition gave useful information about the molecular mechanism of the electrogenic pump.Depolarization of the membrane potential, decrease of membrane conductance, and decrease of pump current during inhibition of the pump with DCCD are caused mainly by the decrease of conductance of the pump channel. The decrease of this pump conductance is caused principally by a marked decrease of the rate constant for releasing H⁺ to the outside.     

The regulation of turgor pressure during sucrose mobilisation and salt accumulation by excised storage-root tissue of red beet

The changes in turgor pressure that accompany the mobilisation of sucrose and accumulation of salts by excised disks of storage-root tissue of red beet (Beta vulgaris L.) have been investigated. Disks were washed in solutions containing mannitol until all of their sucrose had disappeared and then were transferred to solutions containing 5 mol·m^-3 KCl+5 mol·m^-3 NaCl in addition to the mannitol. Changes in solute contents, osmotic pressure and turgor pressure (measured with a pressure probe) were followed. As sucrose disappeared from the tissue, reducing sugars were accumulated. For disks in 200 mol·m^-3 mannitol, the final reducing-sugar concentration equalled the initial sucrose concentration so there was no change in osmotic pressure or turgor pressure. At lower mannitol concentrations, there was a decrease in tissue osmotic pressure which was caused by a turgor-driven leakage of solutes. At concentrations of mannitol greater than 200 mol·m^-3, osmotic pressure and turgor pressure increased because reducing-sugar accumulation exceeded the initial sucrose concentration. When salts were provided they were absorbed by the tissue and reducing-sugar concentrations fell. This indicated that salts were replacing sugars in the vacuole and releasing them for metabolism. The changes in salf and sugar concentrations were not equal because there was an increase in osmotic pressure and turgor pressure. The amount of salt absorbed was not affected by the external mannitol concentration, indicating that turgor pressure did not affect this process. The implications of the results for the control of turgor pressure during the mobilisation of vacuolar sucrose are discussed.To whom correspondence should be addressed.     

Water relations of individual leaf cells ofMesembryanthemum crystallinum plants grown at low and high salinity

Summary The effects of saline conditions on the water relations of cells in intact leaf tissue of the facultative CAM plantMesembryanthemum crystallinum were studied using the pressure probe technique. During a 12-hr light/dark regime a maximum in turgor pressure was recorded for the mesophyll cells of salttreated (CAM) plants at the beginning of the light period followed 6 hr later by a pressure maximum in the bladder cells of the upper epidermis. In contrast, the turgor pressure in the bladder cells of the lower epidermis remained constant during light/dark regime. Turgor pressure maxima were not observed in untreated (C₃) plants.This finding strongly supports the assumption that water movement during malate accumulation and degradation in salttreated plants occurs predominantly between the mesophyll cells and the bladder cells of the upper epidermis. The necessary calculations take differences in the compartment volumes and in the elastic moduli of the cell walls () of the bladder cells of the lower and upper epidermis into account.Measurements of the kinetics of water transport showed that the half-time of water exchange for the two sorts of bladder cells were nearly identical in CAM plants and in C₃ plants. The absolute values of the half-times increased by about 45% in salttreated plants (about 113 sec) compared to the control plants (78 sec). Simultaneously, the half-time of water exchange of the mesophyll cells increased by about 60% from 14 sec (untreated plants) to 22 sec (salt-exposed plants). The leaves of this plant are apparently able to closely maintain the time of propagation of short-term osmotic pressure changes over a large salinity range.A cumulative plot of the data measured on both C₃ and CAM plants showed that the differences between the values of the elastic moduli of bladder cells from the lower and from the upper epidermis are due to differences in volume and suggested that the intrinsic elastic properties of the differently located bladder cells of C₃ and CAM plants were identical.A cumulative plot of the hydraulic conductivity of the membrane obtained both on mesophyll and on bladder cells of salttreated and of untreated plantsvs. the individual turgor pressure yielded a relationship well-known from giant algal cells and some higher plant cells: The hydraulic conductivity increased at very low pressure, indicating that the water permeability properties of the membrane of the various cell types of C₃ and CAM plants are pressure dependent, but otherwise identical.The results suggest that a few fundamental physical relationships control the adaptation of the tissue cells to salinity.