Aluminum-induced deposition of (1,3)-β-glucans (callose) inTriticum aestivum L.

Aluminum (Al)-induced damage to leaves and roots of two Al-resistant (cv. Atlas 66, experimental line PT741) and two Al-sensitive (cv. Scout 66, cv. Katepwa) lines ofTriticum aestivum L. was estimated using the deposition of (1, 3)--glucans (callose) as a marker for injury. Two-day-old seedlings were grown for forty hours in nutrient solutions with or without added Al, and callose deposition was quantified by spectrofluorometry (0–1000 µM Al) and localized by fluorescence microscopy (0 and 400 µM Al). Results suggested that Al caused little damage to leaves. No callose was observed in leaves with up to 400 µM Al treatment. In contrast, root callose concentration increased with Al treatment, especially in the Al-sensitive lines. At 400 µM Al, root callose concentration of Al-sensitive Scout 66 was nearly four-fold that of Al-resistant Atlas 66. After Al treatment, large callose deposits were observed in the root cap, epidermis and outer cortex of root tips of Scout 66, but not Atlas 66. The identity of callose was confirmed by a reduced fluorescence in Al-treated roots: firstly, after adding an inhibitor of callose synthesis (2-deoxy-D-glucose) to the nutrient solution, and secondly, after incubating root sections with the callosedegrading enzyme -D-glucoside glucohydrolase [EC 3.2.1.21]. Root callose deposition may be a good marker for Al-induced injury due to its early detection by spectrofluorometry and its close association with stress perception.Abbreviations DDG 2-deoxy-D-glucose - PAS periodic acid - Schiffs reagent - PE pachyman equivalents     

Solution pH modifies the response of Norway spruce seedlings to aluminium

Norway spruce (Picea abies) was exposed to nutrient solutions containing a range of aluminium (Al) concentrations at several pH levels (3.2, 4 and 5). Root growth was reduced by 100 µM and 400 µM Al at pH 4 and 5, but at pH 3.2 only by 400 µM Al. The Al content of the roots increased with increasing pH. The Al content of the roots was higher at the root tips than at the older root parts at all pH values. Using X-ray microanalysis it could be shown that higher levels of Al at increased pH were mainly due to increased Al contents in root cortex cell walls. In seedlings, mycorrhizal with Pisolithus tinctorius or Lactarius rufus, the Al concentration of cortex cell walls was higher when nitrate (NO₃) rather than ammonium (NH₄) was the nitrogen (N) source.     

Actions of aluminum on voltage-activated calcium channel currents

Summary 1. Extracellular and intracellular effects of aluminum (Al) on voltage-activated calcium channel currents (VACCCs) of cultured rat dorsal root ganglion (DRG) neurons were investigated. Al (0.54 to 5.4 µg/ml=20–200 µM) applied extracellularly reduces VACCCs in a concentration-dependent manner. The IC₅₀ was calculated to be 2.3 µg/ml (83 µM). All types of VACCCs were similarly reduced by Al treatment. A slight shift of the current-voltage relation to depolarized potentials was observed for higher Al concentrations (>2 µg/ml). The action of Al was found to be use dependent, with little recovery (max. 20%) after wash.2. The effect of Al was highly pH dependent in the investigated range (pH 6.4 to 7.8). We observed a rightward shift of the concentration-response curve at pH 7.7 (IC₅₀:3.1 µg/ml) and a leftward shift at pH 6.4 (IC₅₀:0.56 µg/ml) compared to the concentration-response curve at pH 7.3.3. The VACCC declined when 2.7 µg/ml Al was added to the internal solution. A steady state was reached within a few minutes. Additional extracellular application of the same concentration lead to an additional decrease of the current. These observations strongly suggest the existence of both intracellular and extracellular accessible binding sites for Al on voltage-activated calcium channels (VACCs).4. The special characteristics of the action of Al on VACCCs, i.e., the irreversibility, use dependence, and pH dependence, as well as the additional internal binding site may contribute to its neurotoxicity.     

Aluminium effects on transmembrane potential in cells of fibrous roots of sugar beet

The effect of aluminium (Al) on the electrical transmembrane potential of epidermal and outer cortical root cells of intact seedlings of sugar beet (Beta vulgaris L. cv. Monohill) was studied. The potential difference to the surrounding medium was recorded with microelectrodes inserted into the vacuoles (PD_v) and cytoplasm (PD_c) of intact roots. Both long-term effects of AlCl₃ (100, μM present during cultivation) and immediate effects of AlCl₃ (10, 50, or 100 μM present in the assay medium), were measured. The effect of Al was measured at pH 4.0, 5.0 and 6.5 in order to obtain information on the toxicity of different Al forms existing at different pH values. Low pH and/or the presence of AlCl₃ during cultivation caused large depolarizations of the PD_v. Since the immediate effect of 2,4-dinitrophenol (DNP) on the resting potential of cells from Al-cultivated plants was negligible, it is likely that Al affects the metabolic component of the transmembrane potential. Aluminium also had an immediate effect on the PD in root cells of plants cultivated without Al. Addition of 10 or 50 μM Al to the assay medium caused hyperpolarization of PD_v in the presence of 0.5 mM Ca²⁺ at all pH values studied, depolarization of PD_c at pH 6.5, and hyperpolarization of PD_c at lower pH. At 1 mM Ca²⁺, or in the presence of K⁺ (≥ 2 mM), however, the same Al concentrations had little effect on PD_c. The strongest depolarizing effects of 10 or 50 μM Al in short-term treatments were obtained at pH 6.5, and were probably due to the soluble species Al(OH)₃, which is more frequent at pH 6.5 than at a lower pH. Addition of 50 μM Al caused alkalinization of the root medium at pH 6.5, but not at pH 4.0. Therefore, it is possible that Al at pH 6.5 is bound to, or translocated across, the membrane without the accompanying hydroxide ions. It is likely that most of the Al is bound to the root cells, since removal of Al from the buffer surrounding the roots did not cause the changed PD values to return to the original values. Aluminium also interacts with effects of Ca²⁺ and K⁺ on the membrane potential, since changes in PD, induced by changes in concentrations of Ca²⁺ and K⁺ are different in the absence and presence of Al.     

Callose formation as parameter for assessing genotypical plant tolerance of aluminium and manganese

Callose ((1,3)--glucan) formation in plant tissues is induced by excess Al and Mn. In the present study callose was spectrophotometrically quantified in order to evaluate whether it could be used as a parameter to identify genotypical differences in Al and Mn tolerance. Mn leaf-tissue tolerance of cowpea and linseed genotypes was assessed using the technique of isolated leaf tissue floating on Mn solution. Genotypical differences in the density of brown speckles on the leaf tissue (Mn toxicity symptoms) correlated closely with the concentrations of callose for both plant species. In cell suspension cultures Mn excess also induced callose formation. However, differences in tolerance of cowpea genotypes using callose formation as a parameter could only be found in cultured cowpea cells if controls cultured at optimum Mn supply showed low background callose. As soon as after 1 h, Al supply (50 M) induced callose formation predominantly in the 5-mm root tip of soybean seedlings. Callose concentration in the 0–30 mm root tips was inversely related to the root elongation rate when roots were subjected to an increasing Al supply above 10 M. Three soybean genotypes differed in inhibition of root-elongation rate and induction of callose formation when treated with 50 M Al for 8 h. Relative callose concentrations and relative root-elongation rates for these genotypes were significantly negatively correlated.     

Effects of aluminium on tap-root elongation of soybean (Glycine max), cowpea (Vigna unguiculata) and green gram (Vigna radiata) grown in the presence of organic acids

The role of fulvic, malic, and oxalic acids in alleviating the toxic effects of aluminium (Al) on tap-root elongation of soybean cv. Fitzroy, cowpea cv. Vita 4, and green gram cv. Berken was studied. Treatments consisted of a factorial combination of four Al concentrations (0, 12.5, 25 and 50 µM as Al(NO₃)₃·9H₂O) and two concentrations either of malic or oxalic acid (0, 50 µM) or fulvic acid (0, 65 mg L^-1 of organic carbon). The free monomeric Al in solution was determined using a pyrocatechol violet procedure which distinguishes between monomeric and organically complexed Al. Fulvic acid completely alleviated the toxic effect of Al at all concentrations on soybean and cowpea and at concentrations <25 µM on green gram. The non-toxic Al-fulvate complex remained in solution. Both malic and oxalic acid, at the concentrations tested, failed to alleviate Al toxicity on any species; a much higher proportion of the added Al remained in monomeric form in the presence of these acids.     

The effects of low concentrations of aluminium on the growth and uptake of nitrate-N by white clover

Summary The effects of aluminium (Al³⁺) at concentrations of 0, 25, 50 and 100 μM on the growth of white clover, dependent upon N supplied as NO ₃ ⁻ , were examined in flowing solution culture. Plants were established with a normal nutrient supply for 7 weeks and then grown with carefully controlled pH (at 4.5) and P concentrations, and with 0, 25, 50 or 100 μM Al³⁺ for a further three weeks. There were rapid visual effects (i.e. symptoms of P deficiency and reduction in root extension) and the dry weights of shoots and roots were reduced at 50 and 100 μM. Less than 10% of Al absorbed from solution was transported to the shoots. The uptake of P, and its transport between roots and shoots, were reduced in plants grown with Al. The uptake of NO ₃ ⁻ stopped immediately after the introduction of 50 or 100 μM Al, and was significantly reduced at 25 μM after three weeks. During a second phase of the experiment, plants previously grown at 0, 25, 50 and 100 μM Al, were grown for a further 2 weeks either with NO ₃ ⁻ (with and without 50 μM Al³⁺) or without NO ₃ ⁻ but with inoculation by Rhizobia (and with or without 50 μM Al³⁺). The effects of the previous treatments with Al on N uptake were small during the second phase, but uptake by all plants was restricted when Al was present. Inoculation did not result in nodulation in the second phase when Al³⁺ was present in the solution, but Al already in the plant from the first phase did not prevent nodulation in the absence of Al during the second phase.     

Pectin methylesterase modulates aluminium sensitivity in Zea mays and Solanum tuberosum

Cell suspension cultures of Zeamays L. were adapted to grow under conditions of NaCl stress, which increased the cell‐wall pectin content of these cells by 31% compared with unadapted cells (controls). Both cultures were treated for 5 or 10 min with pectin methylesterase (PME) and afterwards incubated in the presence of Al for 2 h. The different capabilities of the cells to synthesise callose due to pre‐treatment were taken into account by calculating relative Al‐induced callose induction (digitonin=100%). Only in salt‐adapted cells with a degree of methylation of cell‐wall pectin (DM) decreasing from 34% (control) to 13%, did PME treatment enhance total and BaCl₂‐non‐exchangeable Al contents and Al sensitivity as indicated by increased callose formation. In a further step, a wider variation in DM was achieved by subculturing the NaCl‐adapted cells for up to 3 weeks without NaCl supply and adapting them to the cellulose‐synthesis inhibitor 2,6‐dichlorbenzonitrile (DCB). This reduced DM to 26%, while short‐term treatment with pectolyase resulted in the lowest DM (12%). After the 2 h Al treatment, there was a close negative relationship between DM and relative callose formation of Al contents, with the exception of pectolyase‐treated cells. In addition, intact plants of Solanumtuberosum L. genotypes were characterised for their Al sensitivity in hydroponics using root elongation, Al‐induced callose formation and Al contents of root tips as parameters. Based on all three parameters, the transgenic potato mutant overexpressing PME proved to be more Al‐sensitive than the wild type, the Al‐resistant and even the Al‐sensitive potato cultivar. Especially in the root tips (1 cm), Al treatment (2 h, 50 μM) increased the activity of PME more in the Al‐sensitive than in the Al‐resistant genotypes. The presented data emphasise the importance of the DM of the pectin matrix and the activity of PME for the expression of Al toxicity and Al resistance.     

Mechanisms involved in the antiplatelet activity of ketamine in human platelets

The aim of this study was to systematically examine the inhibitory mechanisms of ketamine in platelet aggregation. In this study, ketamine concentration-dependently (100–350 µM) inhibited platelet aggregation both in washed human platelet suspensions and platelet-rich plasma stimulated by agonists. Ketamine inhibited phosphoinositide breakdown and intracellular Ca²⁺ mobilization in human platelets stimulated by collagen. Ketamine (200 and 350 µM) significantly inhibited thromboxane (Tx) A₂ formation stimulated by collagen. Moreover, ketamine (200 and 350 µM) increased the fluorescence of platelet membranes tagged with diphenylhexatriene. Rapid phosphorylation of a platelet protein ofMr 47,000 (P47), a marker of protein kinase C activation, was triggered by phorbol-12,13-dibutyrate (100 nM). This phosphorylation was markedly inhibited by ketamine (350 µM). These results indicate that the antiplatelet activity of ketamine may be involved in the following pathways. Ketamine may change platelet membrane fluidity, with a resultant influence on activation of phospholipase C, and subsequent inhibition of phosphoinositide breakdown and phosphorylation of P47, thereby leading to inhibition of intracellular Ca²⁺ mobilization and TxA₂ formation, ultimately resulting in inhibition of platelet aggregation.     

The effects of soluble-Al on root growth and radicle elongation

Summary In order to determine the effects of concentration on plant growth, aluminium (Al) was extracted (10^–3 M CaCl₂) from 4 acid brown hill soils which had been treated with superphosphate at rates equivalent to 0 to 300 kg P ha^–1. The soils ranged in pH (CaCl₂) from 3.5 to 4.9, and Al concentration from 0 to 0.6 mM. The effects of Al on ryegrass growth in the 4 soils in a glasshouse was compared with its effect on radicle elongation of seeds germinated in contact with CaCl₂ extracts from the same soils.Ryegrass root growth in the glasshouse, and radicle elongation in the bioassay test were both unaffected by Al concentrations below 0.1 mM. Root growth was substantially reduced when Al concentration exceeded 0.1 mM and above 0.2 mM growth was almost completely inhibited. Radicle elongation rate was also reduced when the concentration of Al was greater than 0.2 mM agreeing well with the observation from the pot experiment.It is concluded that because of its speed and convenience the bioassay method offers a useful method of establishing critical levels of Al for crop plants.     

Growth and functional responses of different cultivars of Lotus corniculatus to aluminum and low pH stress

Aluminum toxicity is an important stress factor in acid soils. Growth, respiration and permeability properties of root cells were studied in five cultivars of Lotus corniculatus subjected to aluminum (Al) or low pH stress. The cultivars showed significant differences in root elongation under stress conditions, which correlated with changes in membrane potential (E_M) of root cortical cells. A pH drop from 5.5 to 4.0 resulted in significant membrane depolarization and root growth inhibition. The strongest inhibition was observed in cv. São Gabriel (33.6%) and least in cv. UFRGS (25.8%). Application of an extremely high Al concentration (2 mM) stopped the root growth in cv. INIA Draco, while inhibition in cv. UFRGS reached only 75%.The E_M values of cortical cells of Lotus roots varied between −115 and −144 mV. Treatment with 250 μM of AlCl₃ (pH 4) resulted in rapid membrane depolarization. The extent of the membrane depolarization ranged between 51 mV (cv. UFGRS) and 16 mV (cv. INIA Draco). The membrane depolarization was followed by a loss of K⁺ from Al-treated roots (2 mM Al) and resulted in a decrease of the diffusion potential (E_D). The total amount of K⁺ in Al-treated roots dropped from 31.4 to 16.8 μmol g⁻¹ FW in sensitive cv. INIA Draco, or from 26.1 to 22.7 μmol g⁻¹ FW in tolerant cv. UFGRS. The rate of root respiration under control conditions as well as under Al treatment was higher in cv. INIA Draco than in cv. UFRGS. Al-induced inhibition of root respiration was 21–34% of the control.     

Interactions between aluminium,phosphorus and pH in the response of barley to soil acidity

Summary Two acid soils showing different Al solubility as a function of pH were limed to a range of pH values (in 10^–2 M CaCl₂) between 4.1 and 5.6. The apparent critical pH for the growth of barley in pots was 0.25 lower in the soil showing lower Al solubility. The addition of phosphate reduced exchangeable and soluble Al in the soils, and lowered the apparent critical pH by 0.35 while maintaining the difference between the soils. The Al concentration at the critical pH, measured after cropping to take account of the treatment effects on soil Al, also varied with soil and with phosphate addition. These apparent critical values of both pH and soluble Al varied linearly with available phosphate, over the range 18 to 73 mg P/kg soil, as follows: pH from 4.9 to 4.3; soluble Al, from 0.010 mM to 0.056 mM; and the soluble Ca/Al mole ratio, from 1270 to 214.     

Effects of organic acid fractions extracted from Eucalyptus camaldulensis leaves on root elongation of maize (Zea mays) in the presence and absence of aluminium

Complexes of aluminium (Al) with organic ligands are believed to represent an important detoxification mechanism in acid soils. However, relatively little is known about the particular ligands produced by decomposing vegetation or about their effects on plant growth in the presence or absence of toxic Al. This paper reports an experiment on the effects of decomposition products of Eucalyptus camaldulensis leaves on the root elongation of maize (Zea mays) cv. DK687 in the presence or absence of Al. The static solution culture experiment used fulvic acid (FA) and humic acid (HA), extracted from E. camaldulensis leaves, at three nominal concentrations, viz. 40, 120 and 360 mg C L^-1, replicated 4 times in the presence and absence of 30 µM Al. In the absence of Al, root elongation was increased by 30% by HA at 40 mg C L^-1 and by 36% by FA at 120 mg C L^-1. In the presence of 30 µM Al, the effects of toxic Al on root elongation were negated by FA and HA at all concentrations. Aluminium was totally complexed in all treatments except FA at 40 mg C L^-1 in which treatment only 2.7 µM Al was present in the monomeric form. The E. camaldulensis FA and HA at concentrations of 40 and 120 mg C L^-1, either in the presence or absence of Al, stimulated maize root elongation. Aluminium was strongly complexed by the E. camaldulensis FA and HA. The present results, in which FA and HA alleviated Al toxicity limitations on root elongation of maize, are relevant to the protection afforded to plant growth in acid soils amended with organic materials. They highlight the need to focus more on the role of FA and HA.     

The effects of Al on the calcium currents inHelix neurons

Summary 1. The effects of aluminium (Al) on calcium (Ca) currents were investigated by using the conventional two-electrode voltage clamp technique inHelix pomatia neurons. The peak amplitude, kinetics, and voltage dependence of activation and inactivation of the Ca currents were studied in the presence of 10^–5–10^–3 M AlCl₃, at pH 6.2. Al prolonged the rising phase of the Ca currents and therefore increased the time to peak at each command voltage step used.3. There was no significant influence of Al on the peak amplitude of the Ca currents, but the voltage dependence of the time to peak, activation, and inactivation of the Ca currents shifted to more positive potentials as a consequence of Al treatment.4. The leak currents were not influenced by Al up to 1 mM, which was the maximal dose applied.5. The results support the suggestion that Al may modify the Ca homeostasis and that it exerts a neurotoxic effect, at least in part, by modulation of the Ca current of the neuronal membrane.     

The effect of aluminium concentration on root hairs in white clover (Trifolium repens L.)

The effect of aluminium (Al) on root hair length and number is quantified using solution culture techniques with genotypes from white clover cultivar Tamar, that had previously been selected for long and short root hairs. The population differences were maintained in control (0 Al) treatments, with the long-haired population having hairs three times longer than the short-haired population. At an activity of 2.2 µM Al³⁺, root hair length decreased in both populations, the magnitude of the decrease being greater for the long-haired population. Root hair numbers decreased in a similar manner for both populations. At an activity of 4.4 µM Al³⁺ or higher, root hairs virtually disappeared and root growth was very stunted. The effect of Al on root hair development has not been previously quantified, however other workers have observed reduced root hair development in other species at activities of Al greater than 2.5 µM Al³⁺.     

Muscarinic activation causes biphasic inotropic response and decreases cellular Na+ activity in canine cardiac purkinje fibers

In this study, the effects of carbachol (CCh) on twitch tension, intracellular Na⁺ activity (a _Na ⁱ ), and action potential were simultaneously measured in canine cardiac Purkinje fibers in order to examine the regulation of inotropy through muscarinic receptors and its relation to a _Na ⁱ . In fibers driven at 1 Hz, CCh (10 µM) initially and transiently decreased and then increased the twitch tension by 36±8%. The action potential showed a significant elevation of the plateau and a significant shortening of the duration at 90% repolarization (APD₉₀), from 403±7 to 389±7 ms. The a _Na ⁱ decreased from 7.4±0.4 to 6.7±0.3 mM (n=23, p<0.05). Atropine (1 µM) decreased the twitch tension by 21±6% (n=7, p<0.05) without significant effects on the action potential and a _Na ⁱ , and inhibited the effects of CCh. Cs⁺ (20 mM) increased the plateau height and APD₉₀, enhanced the twitch tension by 66±24%, but decreased a _Na ⁱ from 7.3±0.3 to 6.3±0.4 mM (n=6, p<0.05). In the presence of 20 mM Cs⁺, some fibers generated slow responses. The addition of 10 µM CCh further increased the twitch tension and APD₉₀, and decreased a _Na ⁱ from 6.3±0.4 to 5.3±0.3 mM. Ouabain (0.3 µM) increased the twitch tension and a _Na ⁱ , and inhibited the CCh-induced decrease of a _Na ⁱ . In the presence of ouabain, 20 mM Cs⁺ depolarized the fiber and generated slow responses with a decreased a _Na ⁱ . The addition of 10 µM CCh enhanced the slow action potential, and increased a _Na ⁱ although there was a transient decrease during early exposure. These results suggest that activation of muscarinic receptors in canine Purkinje fibers results in an enhancement of the Na⁺-K⁺ pump activity and a biphasic inotropic response, probably via different receptor subtypes. The inhibitory effect, most likely through M₂ receptors, is associated with the activation of K⁺ channels. The stimulatory effect, on the other hand, is probably due to the action on the M₁ receptors, resulting in increases in Ca²⁺ currents.     

Inhibition of the sodium channel by SK&F 96365, an inhibitor of the receptor-operated calcium channel,in mouse diaphragm

The effects of SK&F 96365, a blocker of the receptor-operated Ca²⁺ channel, on contractilities and the Na⁺ channel of mouse diaphragm were studied. SK&F 96365 (10–50 µM) reversibly inhibited twitches, tetanic contractions and muscle and nerve action potentials. The IC₅₀ was 17–24 µM. The inward Na⁺ current was suppressed and its recovery from inactivations delayed. Crotamine, a peptide toxin that binds to neurotoxin receptor site 3 of the muscle Na⁺ channel, enhanced the inhibitory effects of SK&F 96365 and reduced the IC₅₀ to about 4 µM. Veratridine had similar effects, although it was less effective than crotamine. On the other hand, the crotamine-induced membrane depolarizations and spontaneous discharges of muscle action potentials were inhibited by SK&F 96365 noncompetitively. The inhibitory effects of tetrodotoxin and tetracaine were additive with those of SK&F 96365 but were enhanced slightly by crotamine. The results suggested that SK&F 96365 acts on a distinct site and blocks the Na⁺ channel of excitable membranes at a concentration range that inhibits the receptor-operated calcium channel.     

A comparison of soil tests to predict the growth and nodulation of subterranean clover in aluminium-toxic topsoils

Total Al concentration or pH in 1∶5 10 mM CaCl₂ extracts and exchangeable Al in 100 mM BaCl₂ extracts cannot always distinguish between Al-toxic and Al-nontoxic topsoils. Our objectives were to compare the abilities of different measures of Al and pH in various extracts to predict the effects of acidity on growth and nodulation of subterranean clover. In a glasshouse experiment,Trifolium subterraneum L. cv. Mt Barker was grown in acidic soils from 3 sites in the Western Australian wheatbelt with different histories of phosphate fertilizer application. The pH was adjusted to give a range of 3.8–7 in the centrifuged soil solution (SS). Total (Al-tot), reactive Al (8-hydroxyquinoline-extractable, Al-HQ) and pH were measured in SS and 1∶5 extracts of KCl, CaCl₂ and LaCl₃. Another method of estimating reactive Al (Al which reacts with Chelex-100) was also measured in SS only. Other measurements included exchangeable Al and H, Ca in SS, and P in SS and the CaCl₂ extracts. Both plant growth and early nodulation decreased with increasing acidity. Plant growth in the acidified and unlimed treatments of all soils was best described by Al-HQ in SS, KCl or CaCl₂ (r²=0.68–0.70). Multiple regression of relative yield against Al or pH with the concentration of P in SS increased the percentage variation explained by 10% and 30%, respectively. Early nodulation was well correlated (r²=0.67–0.91) with pH or exch. H, Al-tot or exch. Al and Al-HQ. No improvement in the correlation was gained by including P using multiple regression. At constant ionic strength, increasing the valence of the extracting cation decreased the ability of soil tests to distinguish phytotoxic Al.     

铝胁迫下胡枝子根尖胼胝质形成规律及影响因素

采用水培试验,研究了铝胁迫下两个胡枝子品种根尖产生胼胝质的变化规律及影响因素。结果表明,两个品种的根尖铝吸收量与胼胝质形成量呈正比例关系。品种间差异主要是在根尖0—0.5 cm处。敏感品种胼胝质形成量同铝吸收量的变化趋势相一致,而耐性品种则在铝处理6 h时出现一个高峰值后下降。去除铝胁迫后,耐性品种胼胝质形成量并不显著减少。与单独铝处理相比,阴离子通道抑制剂苯甲酰甲醛加铝处理对两个品种胼胝质形成无影响;尼氟灭酸加铝处理抑制敏感品种胼胝质的形成,对耐性品种无影响;蒽-9-羧酸加铝处理显著抑制两个品种的胼胝质形成。另外,抑制剂2-去氧-D-葡萄糖加铝共同处理与单独铝处理相比,敏感品种的胼胝质形成量显著降低,耐性品种无影响。甘露醇对两个品种胼胝质形成的影响无显著差别。镧处理下胼胝质的形成量是耐性品种显著高于敏感品种,铝、镧同时处理胼胝质的形成量最高。敏感品种胼胝质形成处理间无差别。总之,耐性品种在铝胁迫下胼胝质形成与有机酸分泌可能存在一定的协调关系;铝胁迫下胼胝质形成是敏感指标;在一定条件下,特别是有机酸分泌前胼胝质的形成可能具有一定抗性意义;铝诱导胼胝质的形成受多种外界因素(浓度、时间、有机酸分泌,渗透压等)的影响。     

EGF Stimulates Growth by Enhancing Capacitative Calcium Entry in Corneal Epithelial Cells

In rabbit corneal epithelial cells (RCEC), we determined whether capacitative calcium entry (CCE) mediates the mitogenic response to epidermal growth factor, EGF. [Ca²⁺]_i was measured with single-cell fluorescence imaging of fura2-loaded RCEC. EGF (5 ng/ml) maximally increased [Ca²⁺]_i 4.4-fold. Following intracellular store (ICS) calcium depletion in calcium-free medium with 10 µM cyclopiazonic acid (CPA) (endoplasmic reticulum calcium ATPase inhibitor), calcium addback elicited plasma membrane Ca²⁺ influx as a result of activation of plasma membrane store operated channel (SOC) activity. Based on Mn²⁺ quench measurements of fura2 fluorescence, 5 ng/ml EGF enhanced such influx 2.3-fold, whereas with Rp-cAMPS (protein kinase A inhibitor) plus EGF it increased by 5.3-fold. In contrast, SOC activation was blocked with 100 µM 2-aminoethyldiphenylborate (2-APB, store-operated channel inhibitor). During exposure to either 50 µM UO126 (MEK-1/2 inhibitor) or 10 µM forskolin (adenylate cyclase activator), 5 ng/ml EGF failed to affect [Ca²⁺]_i. RT-PCR detected gene expression of: 1) transient receptor potential (TRP) protein isoforms 1, 3, 4, 6 and 7; 2) IP₃R isoforms 1–3. Immunocytochemistry, in conjunction with confocal and immunogold electron microscopy, detected plasma membrane localization of TRP4 expression. Inhibition of CCE with 2-APB and/or CPA, eliminated the 2.5-fold increase in intracellular [³H]-thymidine incorporation induced by EGF. Taken together, CCE in RCEC mediates the mitogenic response to EGF. EGF induces CCE through its stimulation of Erk1/2 activity, whereas PKA stimulation suppresses these effects of EGF. TRP4 may be a component of plasma membrane SOC activity, which is stimulated by ICS calcium depletion.