Salt Tolerance in the Halophyte Suaeda maritima L. Dum.: Intracellular Compartmentation of Ions

The time-course of exchange of sodium and potassium ions fromroot and leaf material of the halophyte Suaeda maritima hasbeen followed and the data analysed according to the phenomenologyof efflux, or compartmental, analysis. Sodium ions were exchangedmuch more slowly (c. 4 times) from the vacuoles of leaf cellsof plants grown in sodium chloride than were potassium ionsfrom the vacuoles of leaf cells of plants grown either in similarconcentrations of potassium chloride or in low concentrationsof potassium. In plants grown in sodium chloride, sodium ionswere exchanged 9 times more slowly from the vacuoles of leafcells than from the vacuoles of root cells. The concentration of sodium ions in the cytoplasm of leaf cellsof plants growing in 340 mol m^–3 sodium chloride was estimatedto be 165 mol m^–3 when the average concentration in theleaf tissue was about 600 mol m^–3. As measured by movement from mature to developing leaves inintact plants; there was less in vivo retranslocation of ²²Naand ³⁶CI in plants growing in sodium chloride than there wasof ⁸⁶Rb in plants growing either in potassium chloride or innon-saline conditions. The results are discussed in terms of the concept and energeticsof compartmentation of ions in the cells of halophytes.     

The Ultrastructure of Ion-Secreting and Non-Secreting Salt Glands of Limonium platyphyllum

The ultrastructure of salt glands in developing leaves of Limoniumplatyphyllum is described prior to exposure to 3% NaCl solution(with non-secreting glands) and after 4.5 and 18 h exposureto the salt solution. It is shown that in most glands, the transitionto active chloride transport was accompanied by the displacementof vacuoles toward the cell periphery and by the establishmentof plasmalemma contact sites with the tonoplast which appearedsimilar to gap junctions in animal epithelial cells. No evidencefor the exocytosis of vacuoles was found. It is suggested thatgland vacuoles may have a primary role in chloride secretionand that the tonoplast may be functionally asymmetrical, sothat the free part facing the hyaloplasm bears ion pumps, whereashighly permeable ion channels are active along the zone of contactwith the plasmalemma. It follows that the active step in chloridetransport in Limonium glands is the influx of ions into thevacuoles. Within the inner cup cells of the gland, vacuolescome into contact with the plasmalemma only at sites where thecell wall is adjacent to secretory and accessory cells. Suchan asymmetry appears to ensure the directed flux of ions intothis cell wall. Wall protuberances in the gland cells are rudimentaryand presumably not involved directly in NaCl secretion. Thenucleolus is activated during secretion and the frequency offree ribosomes is significantly increased, which is suggestiveof their involvement in the synthesis of membrane transportproteins. The ultrastructure of about one-third of the glandsremained unchanged in salt-treated leaves. Key words: Salt glands, ultrastructure, ion fluxes     

Changes in Activity of Malate Dehydrogenase, Catalase, Peroxidase and Superoxide Dismutase in Leaves of Halimione portulacoides (L.) Aellen Exposed to High Sodium Chloride Concentrations

Plants of Halimione portulacoides were grown in nutrient solutionscontaining NaCl at concentrations ranging from 0–2.0 MNaCl. They survived in this environment at least for 20 days.Malate dehydrogenase (MDH), catalase, peroxidase and superoxidedismutase (SOD) were extracted from the leaves of such plantsand enzyme activity was assayed in the absence of salt. Sodium chloride at low concentration (0–0.5 M) stimulatedthe activities of MDH and catalase but inhibited them at concentrationshigher than 0.5 M. Peroxidase and SOD were hardly affected byexposure to salinity in vivo. Salinity in vivo also affectedthe K_m and the V_max of the enzymes. The possibility that thethree enzymes (catalase, peroxidase and SOD) have a role inprotecting the leaf cells against oxygen toxicity caused byfree radicals, that may be formed in cells when growing undersaline and extreme climatic conditions, is discussed. Halimione portulacoides (L.) Aellen, salinity, catalase, peroxidase, superoxide dismutase     

An X-ray Microanalysis Study of the Salt Glands and Intracellular Calcium Crystals of Tamarix

X-ray microprobe analyses of the secreted salts from glandsof 20 Tamarix aphylla (L.) Karst trees growing in alluvial soilsin the north western district of Victoria, Australia, show thatthe glands secrete a variety of elements including calcium,magnesium and sulphur. However, sodium, potassium and chloride,common ions reported to be secreted predominantly in other studies,are present in relatively low amounts in the secreted salts.Calcium, the major cationic element in the secretory productof the gland, was not detected in the symplasm of the secretoryand collecting cells or in the vacuoles of the collecting cells.Some evidence indicates that silicon may be secreted by theglands but further corroboratory data are required. Our resultssupport previous suggestions that the glands of Tamarix havea low level of selectivity. Large, spherical aggregates of calcium crystals were observedin the vacuoles of the mesophyll cells and quantitative X-raymicroanalysis suggested that the chemical composition of thecrystalline aggregates was CaSO₄, 2H₂O. The predominant elementsin the vacuolar sap were magnesium and sulphur, probably presentas MgSO₄, whereas the major elements in the cytoplasm were potassium,sulphur and phosphorus. In Tamarix aphylla the calcium contentof the leaf appears to be regulated by both the secretion ofcalcium salts from the leaf and by intracellular calcificationin mesophyll cells.Copyright 1994, 1999 Academic Press X-ray microanalysis, Tamarix, salt gland, secretion, salts, ions, calcium crystals, compartmentation     

The Effect of Irradiance on the Recovery of Soybean Nodules from Sodium Chloride-Induced Senescence

Soybean (Glycine max L. Merr) cv. Clarke plants inoculated withBradyrhizobium japonicum strain RCR3407 were grown either ina greenhouse with a low irradiance (200–400)µmolm^–2 s^–1) or in a controlled-environment growth cabinetwith a higher irradiance (600 µimol m^–2 s^–1).At 42 d plants were given a nitrogen-free nutrient solutioncontaining 50 mol m^–3 sodium chloride for 2 weeks andthen allowed to recover from salt-stress for a further 2 weeks. Salt treatment reduced plant growth by at least half in bothgrowth regimes, however, the controlled environment-grown (CEG)plants were five times larger than the greenhouse-grown (GG)plants in terms of dry weight and number/weight of nodules perplant, regardless of treatment. The structure of nodules, from both growth regimes, harvestedat the end of the 2 week salt-stress was similar to unstressedcontrol nodules. However, nodules harvested 1 week later fromboth CEG and GG plants had structural changes including degradationof bacteria in vacuoles around host cell nuclei, particularlyin the outer cell layers of the infected tissue. In addition,meristematic activity was seen in the cortex of some nodulesfrom GG plants. Young cells here contained infection threadsand newly-released bacteria. Nodules harvested 2 weeks after removal of the salt-stress fromCEG plants showed an apparent recovery from the stress. However,there was a very marked increase in the amount of starch inthe cortex which was not seen in equivalent GG nodules. In contrast,nodules from GG plants contained many vacuolate infected cellsand, consequently, a lowered bacteroid population. Further,meristematic activity was seen in a zone concentric to the infectedzone, newly-formed cells contained many large infection threadsand were interspersed with intercellular bacteria. The meristematicactivity increased the relative volume of cortical to infectedcells in these nodules. Growth conditions did not affect control nodule specific nitrogenaseactivity or oxygen diffusion resistance (R) and these parameterswere also not altered in CEG nodules exposed to salt plus the14 d recovery period. However, nitrogenase activity was greatlyreduced, and R increased by more than eight times in equivalentGG nodules exposed to salt plus recovery. It is hypothesized that the gross morphological changes werean attempt to counter salt toxicity and/or oxygen damage underconditions of reduced photosynthate supply to the nodules dueto the poor light levels in the greenhouse. However, soybeannodules supplied with adequate photosynthate were able to withstandand recover from long-term salt-stress with little alterationto their structural integrity. Key words: Soybean, sodium chloride, nitrogen fixation, light intensity, oxygen diffusion resistance     

The Ultrastructure of Frankenia Salt Glands

The ultrastructure of the salt gland of Frankenia has been investigated.Glands were examined after treatment with distilled water andafter 4 h, 15 h, and 36 h of salt treatment. Characteristiccytoplasmic features can be correlated with each of these treatments.The results suggest that the acquisition and maintenance ofthe secreting state involves the vesiculation of endoplasmicreticulum.     

Salt Glands in the Zoysieae

Salt glands were found in two species of the genus Zoysia ofthe tribe Zoysieae, sub-family Chloridoideae (Poaceae). Glandsprotrude from and are recumbent to the leaf epidermis, and consistof two cells, a basal cell and upper cap cell. Glands were betterdeveloped on the adaxial surfaces. Those on the abaxial surfaceappear to be non-functional. Zoysia matrella, the more salt-tolerantspecies, had a higher density of larger glands, and secretedmore sodium per unit leaf mass, resulting in much lower leafsap osmolalities than those of the more salt-sensitive Z. japonica.The finding of salt glands in the tribe Zoysieae confirms itsrelation to the four other tribes within the sub-family Chloridoideaein which salt glands have previously been reported. Salt glands, Zoysieae, Poaceae, Japanese lawngrass, Zoysia japonica, Manilagrass, Zoysia matrella, sodium chloride, salt tolerance, secretion     

Anatomy, Ultrastructure and Assimilate Concentrations of Roots of Citrus Genotypes Differing in Ability for Salt Exclusion

Seedlings of Rangpur lime (Citrus reticulata var. austera hybrid?)and Etrog citron (C. medica) were treated in water culture with0, 25, 50 or 100 mol m^–3 NaCl, and in sand culture with0 or 100 mol m^–3 NaCl. Leaf chloride analyses indicatedthat Etrog citron accumulated the most chloride at all levelsof salinity. The structure, ultrastructure and concentrationsof chloride and reserve assimilates of the primary root up to50 mm back from the tip were compared between genotypes andbetween salt treatments. There were no differences in root anatomy in the absence ofsalt between the two genotypes. The hypodermal cells developedlignified and suberized walls which blocked the plasmodesmataand resulted in degeneration of the cell contents. Frequentthin-walled passage cells in the hypodermis had living contentsand may represent major sites of ion uptake into the symplasm,which was connected with the stele via plasmodesmatal connectionswith and between the cortical cells. The primary endodermalcells had lignified casparian strips and plasmodesmata in othercell wall areas. These connections were blocked by secondarysuberization of all except the endodermal passage cells oppositethe protoxylem arcs. Suberization of the hypodermis and endodermis and the appearanceof granular deposits in the vacuole occurred closer to the roottip of both genotypes after treatment with 100 mol m^–3NaCl. Levels of starch and triglyceride in 10 mm serial segmentswere similar between roots of control and salt-treated plantsand increased with distance back from the root tip. Chlorideconcentrations increased with salt treatment but values (ona tissue water basis) were similar between genotypes and betweenthe apical and proximal ends of the root. Reducing sugar concentrationsdecreased with salt treatment to a similar extent in roots ofboth genotypes. Key words: Citrus, Roots, Salinity     

The Movement of Ions to the Xylem Exudate of Maize Roots: II. A COMPARISON OF THE ELECTRICAL POTENTIAL AND ELECTROCHEMICAL POTENTIALS OF IONS IN THE EXUDATE AND IN THE ROOT CELLS

The electrical potential differences (E_ex) and the electrochemicalpotential differences for potassium and chloride in the xylemexudate of excised maize roots were determined and the valuesobtained indicated active transport of both ions to the xylemexudate. The potential differences were depolarized by increasesin the external KCl concentration ([KCl]_o), and E_ex was a linearfuction of log[KCl]_o. The corresponding data for the cell vacuolesof the same roots were also determined and trends similar tothose found for the exudate were obtained. The electrochemicalpotential differences between the cell vacuoles and the externalsolution for potassium (µ_rac.k) and chloride (µ_vac.Cl)indicated that both ions were actively transported to the vacuoles.Both µ_vac.k and µ_vac.cl decreased in a linear manneras [KCl]_o was increased. However, in spite of the similar trendsin both sets of data, the values of the electrical potentialand chloride electrochemical potential differences were alwaysmarkedly larger for the vacuoles than for the exudate. For potassium,there were no significant differences. It is concluded thatthe discrepancy between the data for the vacuoles and the wholeroots does not support the idea that the exuding root systembehaves like a single cell, as has been suggested by some workers.     

Drought effects on cellular and spatial parameters of leaf growth in tall fescue

The effect of drought and recovery on cellular and spatial parametersof the growth process in tall fescue leaves was studied in twoexperiments. In both experiments plants grown on vermiculiteand maintained in a controlled environment were submitted toa 7 d drought period generated by withholding water. Droughtwas followed by a 3 d recovery period in experiment II. As leafelongation rate (LER) decreased during developing drought boththe growth zone length (initially 40 mm) and the maximum relativeelemental growth rate (initially 0.09 mm mm^–1 h^–1during the dark period of diurnal cycles) within the growthzone declined. But the growth zone still exhibited a lengthof approximately 15 mm when LER approached 0 under severe drought(–2.0 MPa predawn leaf water potential). The growth potentialof the basal 15-mm-long portion of the leaf was conserved duringthe period when drought effected the complete arrest of leafelongation. A (retrospective) analysis of the position-timerelationships of epidermal cells identified on leaf replicas(experiment II) indicated that the cell flux out of the growthzone responded very sensitively to drought. Before drought theflux was maximum at approximately 3.2 cells (cell file h)^–1during the dark period. Flux decreased to 0 when leaf elongationstopped. Flux also varied diurnally both under well-wateredand droughted conditions. In well-watered conditions it wasabout 30% less during the light than the dark period. Cell elongationwas also sensitive to drought. Under well-watered conditionsepidermal cell elongation stopped when cells attained a lengthof approximately 480 µm. During developing drought cellsstopped elongating at progressively shorter lengths. When LERhad decreased to almost nil, cells stopped elongating at a lengthof approximately 250 µn. When drought was relieved followinga 2 d complete arrest of leaf elongation then cells shorterthan 250 µm were able to resume expansion. Following rewateringcell flux out of the growth zone increased rapidly to and abovethe pre-drought level, but there was only a slow increase overtime in the length at which cell elongation stopped. About 2d elapsed until the leaf growth zone produced cells of similarlength as before drought (i.e. approximately 480 µm). Key words: Epidermal cell length, cell flux, (leaf) growth zone, leaf elongation rate, relative elemental growth rate, position-time relationships (path line, growth trajectory), drought, water deficit     

Response of Melanthera biflora to Salinity and Water Stress

Melanthera biflora (Asteraceae) is a moderately salt-tolerantplant from the Indo-Pacific region. In laboratory studies itsgrowth was inhibited by salt above 50 mol m^–3, but itwas able to survive salinities approaching that of seawater,namely 400 mol m^–3. Shoot potassium concentrations weremaintained over a range of salinities up to 400 mol m^–3,while sodium and chloride accumulation followed closely theincrease in external osmotic pressure. In contrast, the increasein osmotic pressure of the leaf sap of Melanthera biflora, subjectedto water stress, was due mainly to a decrease in the ratio offresh weight/dry weight. 3-dimethylsulphoniopropionate (3-DMSP)and glycinebetaine were identified by fast atom bombardmentmass and ¹H -NMR spectroscopy, with 3-DMSP being the main oniumcompound and glycinebetaine absent in some accessions. Onium(quaternary ammonium and/or tertiary sulphonium) compounds andproline increased during salt and water stress due mainly toa decrease in the fresh weight/dry weight ratio of tissue, althoughpart of the increase in salt-stressed tissue was due to an increasein the accumulation of the onium compound. This salt-inducedincrease in 3-DMSP was inhibited in conditions of low sulphursupply and there was no compensatory increase in proline. Key words: Melanthera biflora, Asteraceae, salinity, glycinebetaine, 3-dimethylsulphonioproprionate     

Comparative Leaf Epidermal Anatomy of Mutants of Barley (Hordeum vulgare L. 'Himalaya') which Differ in Leaf Length

We wished to determine the nature of differences in epidermalcell numbers and dimensions between leaves of different lengthin mutants of barley (Hordeum vulgare L. ‘Himalaya’).Three comparisons were made: leaf one (L1)vs. leaf four (L4);wild typevs. nine dwarf mutants and wild typevs. a slender mutant.L1 was shorter than L4, and for most lines this was associatedwith a change in epidermal cell number for the blade, and inboth cell number and length for the sheath. Compared to wildtype, the smaller leaves of dwarf plants generally had shorterand fewer cells in both blade and sheath. The blade of slenderplants was the same length (L1) or longer (L4) than wild type,while the sheath was longer than that of wild type for bothL1 and L4. Slender plants had longer but fewer cells than thewild type along the blade of L1, and shorter but more cellsfor the blade of L4. In the sheath, slender plants had longerand more (L1) or fewer (L4) cells than did the wild type. ForL1, variation in blade width amongst the barley lines was associatedwith a change in file width and file number. For L4, blade widthvaried only with file number, except for slender plants wherenarrow blades were associated with reduced file width. Hencethere was no consistent correlation between changes in cellsize or cell (or file) number with changes in leaf length orwidth. Differences depended on the leaf (L1vs. L4), leaf part(bladevs. sheath), and the nature of the mutation (dwarfvs.slender). Barley (Hordeum vulgare L. ‘Himalaya’); leaf epidermis; dwarf mutant; slender mutant     

X-Ray Microanalysis and Ultrastructural Studies of Cell Compartments of Dunaliella parva

Hajibagheri, M. A., Gilmour, D. J., Collins, J. C. and Flowers,T. J. 1986. X-ray microanalysis and ultrastructural studiesof cell compartments of Dunaliella parva. -J. exp. Bot. 37:1725–1732. Ultrastructural studies of the unicellular green alga Dunaliellaparva showed the presence of cytoplasmic vacuoles. X-ray microanalysiswas performed on sections of cells which had been freeze substitutedin acetone. It was found that the concentrations of both Naand Cl were much higher in the vacuoles than in the cytoplasm.When cells were grown in 0·4 kmol m^–3 NaCl theNa and Cl concentrations in the vacuoles were 349 and 283 molm ^–3 respectively, whilst cytoplasmic Na and Cl concentrationswere 37 and 26 mol m^–3. Corresponding values for cellsgrown in 1·5 kmol m^–3 NaCl were 392 mol m^–3Na and 325 mol m^–3 Cl in the vacuoles and 36 mol m^–3Na and 30 mol m^–3 Cl in the cytoplasm. Immediately afterexposure to an increase in external salinity Na and Q concentrationsincreased in both vacuoles and cytoplasm. The results are discussedwith reference to compartmental models for the ionic relationsof Dunaiiella. Key words: X-ray microanalysis, ultrastructural studies, Dunaliella parva     

Salt Tolerance in Suaeda maritima (L.) Dum: THE EFFECT OF SODIUM CHLORIDE ON GROWTH, RESPIRATION, AND SOLUBLE ENZYMES IN A COMPARATIVE STUDY WITH PISUM SATIVUM L

The effect of sodium, chloride on the growth of a halophyte,Suaeda maritima (L.) Dum., was compared with its effect on Pisumsativum L. cv. Alaska under controlled environmental conditions.The salt stimulated the growth of Suaeda maximally at concentrationsof 170 to 340 mM while the growth of Pisum was inhibited evenby 100 mM. Both species accumulated ions in the tops and themaximum concentrations of Na⁺ and Cl^– rose in Suaeda to860 mM (based on the water content) and 730 mM and in Pisumto 170 mM and 300 mM respectively. Respiration in both specieswas inhibited as the NaCl level in the culture solution wasraised. Four supernatant enzymes (malic dehydrogenase, glucose-6-phosphatedehydrogenase, peroxidase, and acid phosphatase) prepared fromPisum and from Suaeda (grown either in the absence of addedNaCl or in the presence of 340 mM NaCl) were assayed in variouslevels of sodium chloride. The dehydrogenases were markedlyinhibited by increasing salt concentrations while there wasa smaller effect on the peroxidase and acid phosphatase. Therewas no difference in the effect of salt on the enzymes preparedfrom the two species although one is halophilic and the otherhalophobic.     

Salt Tolerance in the Triticeae: Growth and Solute Accumulation in Leaves of Thinopyrum bessarabicum

Gorham, J., McDonnell, E., Budrewicz, E. and Wyn Jones, R. G.1985. Salt tolerance in the Triticeae: growth and solute accumulationin leaves of Thinopyrum bessarabicum.—J. exp. Bot. 36:1021–1031. The diploid wheatgrass Thinopyrum bessarabicum was found towithstand prolonged exposure to 350 mol m^–3 NaCl in hydroponicculture. During the gradual addition of salt to the externalmedium, osmotic adjustment was rapidly achieved by the accumulationof Na and Cl. Following osmotic adjustment constant leaf Naand Cl concentrations were maintained, and K was retained ata high level. Thinopyrum bessarabicum may be described as anosmoconformer, adjusting its internal osmotic pressure to 400–500mOsmol kg^–1 above that of the external medium in hydroponicculture. Both slower shoot initiation and reduced leaf lengthcontributed to the reduced growth rates at higher salinities.Leaf width was not affected. Increasing salinity resulted inincreases in leaf concentrations of phosphate, glycinebetaine,sucrose and proline, and in decreases in the concentrationsof nitrate, sulphate, magnesium, calcium, total amino acidsand organic acids. Thinopyrum bessarabicum exhibits salt tolerancecharacters which may be useful in wheat breeding. Key words: Salt stress, solute accumulation, osmotic adjustment, Thinopyrum     

Effects of salinity levels and seed mass on germination in Australian species of Frankenia L. (Frankeniaceae)

Halophyte species demonstrate differing levels of salt tolerance. Understanding interspecific variation to salinity levels is of value from both the scientific perspective, which includes the identification of traits associated with salinity tolerance, as well as from an applied perspective, which includes identifying plant species for specific salinity restoration and remediation projects. This paper investigates the effects of salinity on germination of 12 Australian species of the plant genus Frankenia L. (Frankeniaceae). We use saline solutions that corresponded to the average soil–water salinity concentrations in the arid zones of inland Australia. These solutions consisted of 10 mM calcium chloride, 30 mM magnesium sulphate, and 450 mM sodium chloride. The aims of our study were: (1) to investigate the germination (germination rates, germination success) of Frankenia seeds to four salinity levels (0%, 10%, 20%, 30%), (2) to test for possible interaction effects between seed mass, germination, and salinity, and (3) to examine the effect of salinity levels on the inhibition of germination and/or seed damage. Species varied in their salt tolerance for germination rates and success. Species with larger seeds had higher germination rates and germination success for high salinity levels. Several species did not germinate well at any salinity level. Finally, no seeds were adversely affected by exposure to high salinity levels pre-germination. There is potential for including some Frankenia species in remediation and revegetation projects in areas affected by salinity, and also as garden plants in saline regions.     

Salt Retention by Bean Hypocotyl

The ultrastructure of hypocotyl, epicotyl, and petiole of bean(Phaseolus vulgaris, L.) was investigated in plants grown ina basic solution in the absence or presence of high levels ofchloride salts. The hypocotyl tissue of both control and salt-treatedplants showed frequent vesiculation similar to that previouslyobserved in the root and hypocotyl of the halophyte, Salicorniaeuropea, L. These vesicles were not previously observed in theroot and leaf of bean plants that were grown under identicalconditions in the absence of high levels of chloride salt. Experiments concerning the localization of chloride as electron-densesilver-complex showed that the vesicles contain a chloride concentrationas high as the cytoplasmic phase or the free space. These resultsare discussed in relation to the ionic retention property ofthe bean hypecotyl and the role of vesiculation in salt resistanceof plants.     

Sodium Chloride Compartmentation in Leaf Vacuoles of the Halophyte Suaeda maritima (L.) Dum. and its Relation to Tonoplast Permeability

Single channel properties, whole vacuole currents and protonpumping capacity were investigated in the intact vacuoles andmembrane patches of leaf tonoplast from the halophyte Suaedamaritima. ATP-dependent proton pumping capacity was similarto non-halophytes whether the plants were or were not grownwith added sodium chloride (200 mM). The most abundant ion channelwas inward rectifying and had a single channel conductance of58 pS in symmetrical KCl solutions (100 mM) to 170 pS usingphysiological conditions (50/150 mM KCl/NaCl cytoplasmic side,50/450 mM KCl/NaCl vacuolar side). The channel showed all thecharacteristics of the SV type channel described in many otherspecies. In the open state these channels caused tonoplast conductancesin excess of 0.5 nS m²– but conductances were much lowerusing physiological ion concentrations and membrane potentials.In spite of the poor selectivity and the potentially large tonoplastconductance it is calculated that compartmentation of NaCl inleaf vacuoles can be sustained by about 30% of ATP-dependentproton pumping capacity. The results do not indicate any specialadaptation of the tonoplast ion channels in the halophyte. Key words: Ion-channels, patch-clamp, salt-tolerance, vacuole     

Thermodynamics of Malate Transport across the Tonoplast of Leaf Cells of CAM Plants

The electrochemical potential difference for each dissociationstate of malic acid across the tonoplast of leaf cells was examinedin two CAM plants, Graptopetalum paraguayense and Kalanchoëdaigremontiana. The concentration of malic acid in each dissociationstate was estimated from an analysis of pH and concentrationsof ionic species that included calcium, malate and isocitrate.The vacuoles contained 30–40 mM isocitrate and 50–70mM calcium in G. paraguayense, and 20–30 mM isocitrateand 70–100 mM calcium in K. daigremontiana. For the calculationof the pattern of dissociation of malic acid, the formationof chelates of calcium with malate and isocitrate, which havedifferent stability constants depending on the dissociationof the acids, were also taken into consideration. The vacuolarconcentrations of the divalently dissociated form of malic acid(mal^2– were 4–7 mM and 1-3 mM in G. paraguayenseand in K. daigremontiana, respectively. To obtain informationabout the cytoplasmic concentration of malate, the apparentinhibition constant for malate of phosphoenolpyruvate carboxylasewas measured. It was about 330 µM in the dark period and60 µM in the light period. Considering an inside-positivemembrane potential, we conclude that mal^2– can be takenup passively into the vacuole during the dark period and canbe released passively from the vacuole during the light period.Two types of channel (the "SV-type" channel and a novel "MU-type"channel) which we found recently in G. paraguayense [Iwasakiet al. (1992) Plant Physiol. 98: 1494] are probably involvedin the uptake and the release of malate in the diurnal CAM rhythm.The existence of a large pH-buffering capacity due to isocitricacid in the vacuole allows the accumulation of a large amountof malic acid during the diurnal CAM rhythm. (Received February 12, 1992; Accepted July 10, 1992)     

Resting membrane potential and action potential of Nitella expansa protoplasts

Inside negative membrane potentials were observed for protoplastsobtained from Nitella expansa leaf internodal cells in mediacontaining 1 to 100 mM CaCl₂ using the microelectrode technique.The potential values were less negative than the membrane potentialof intact N. expansa leaf internodal cells. In addition, anaction potential consisting of two components—a fast componentand a slow component—was induced by electrical stimulationfor the protoplasts as well as the intact cells. (Received December 18, 1979; )