Effects of La3+ on the competence of Escherichia coli as studied by microcalorimetry

A series of microcalorimetric experiments were performed to investigate the effect of La³⁺ on the formation of the competent state of Escherichia coli HB101 by using a LKB-2277 BioActivity Monitor at 37°C. The thermogenic curves in the absence and in the presence of La³⁺ were obtained. Based on these curves, we calculated that the total heat effects (Q _T) and the maximal power (P _max) in the presence and absence of La³⁺. Our experiments indicate that the total heat effects in the presence of a low concentration of La³⁺ (≤ 100 μg/mL) are greater than those in the absence of La³⁺. Their trends are similar with respect to the increasing concentration of La³⁺. To the contrary, when the concentration of La³⁺ is greater than 100 μg/mL, the total heat effects decrease with the increasing concentration of La³⁺. Therefore, in the latter case, La³⁺ has inhibitory effects on the formation of competence. Our experimental results suggest that the La³⁺ ion in the environmental ecosystem can facilitate the formation of competence of E. coli HB101 and further stimulate the transfer of genetic materials between organisms.     

Study on the toxic mechanism of La3+ to Escherichia coli

The toxic mechanism of La³⁺ to Escherichia coli is investigated by detecting the concentration change of La³⁺ in E. coli cells growing in La³⁺-containing medium. Stimulatory action and inhibitory effect of La³⁺ in different concentrations can be attributed to the permeability alteration of the cell. Low concentration of La³⁺ increases the nutrition absorbability of the cells from the cultures as a result of increased cell permeability, and high concentration of La³⁺ causes the accumulation of La³⁺ in cells, resulting in the toxic effects on the E. coli cells.     

Properties of the K+ inward rectifier in the plasma membrane of xylem parenchyma cells from barley roots: Effects of TEA+, Ca2+, Ba2+ and La3+

Xylem parenchyma cells are situated around the (apoplastic) xylem vessels and are involved in the control of the composition of the xylem sap by exporting and resorbing solutes. We investigated properties of the K⁺ inward rectifier in the plasma membrane of these cells by performing patch clamp experiments on protoplasts in the whole-cell configuration. Inward currents were sensitive to the K⁺ channel blocker TEA⁺ at a high concentration (20 mm). Barium, another classical K⁺ channel blocker, inhibited K⁺ currents with a K _i of about 1.3 mm. In contrast to guard cells, the cytosolic Ca²⁺ level proved to be ineffective in regulating the K⁺ conductance at hyperpolarization. External Ca²⁺ blocked currents weakly in a voltage-dependent manner. From instantaneous current-voltage curves, we identified a binding site in the channel pore with an electrical distance of about 0.2 to 0.5. Lanthanum ions reduced the inward current in a voltage-dependent manner and simultaneously displaced the voltage at which half of the channels are in the open state to more positive values. This finding was interpreted as resulting from a sum of two molecular effects, an interaction with the mouth of the channel that causes a reduction of current, and a binding to the voltage sensor, leading to a shielding of surface charges and, subsequently, a modulation of channel gating.A comparison between the K⁺ inward rectifier in xylem parenchyma cells, guard cells and KAT1 from Arabidopsis leads to the conclusion that these rectifiers form subtypes within one class of ion channels. The ineffectiveness of Ca²⁺ to control K⁺ influx in xylem parenchyma cells is interpreted in physiological terms.     

Microcalorimetric studies of the action of Er3+ on Halobacterium halobium R1 growth

A microcalorimetric technique was used to evaluate the influence of Er³⁺ on Halobacterium halobium R1 growth. By means of a LKB-2277 Bioactivity Monitor ampoule method, we obtained the thermogenic curves of H. halobium R1 growth at 37°C. In order to analyze the results, the relationship between k and C was obtained. The addition of Er³⁺ in low concentration cause a decrease of the maximum heat production P _max and growth rate constants k; however, Er³⁺ in a high concentration might promote growth of H. halobium R1. When Er³⁺ is in a much higher concentration, the growth of H. halobium R1 is inhibited completely. For comparison, the shapes of H. halobium R1 cells were observed by means of transmission electron microscope (TEM). According to the thermogenic curves and TEM photos of H. halobium R1 under different conditions, it is clear that the metabolic mechanism of H. halobium R1 growth has been changed with the addition of Er³⁺.     

Research of the entry of rare earth elements Eu3+ and La3+ into plant cell

Whether rare earth elements can enter into plant cells remains controversial. This article discusses the ultracellular structural localization of lanthanum (La³⁺) and europium (Eu³⁺) in the intact plant cells fed by rare earth elements Eu³⁺ and La³⁺. Eu-TTA fluorescence analysis of the plasmalemma, cytoplast, and mitochondria showed that Eu³⁺ fluorescence intensities in such structures significantly increased. Eu³⁺ can directly enter or be carried by the artificial ion carrier A23187 into plant cells through the calcium ion (Ca²⁺) channel and then partially resume the synthesis of amaranthin in the Amaranthus caudatus growing in the dark. Locations of rare earth elements La³⁺ and Eu³⁺ in all kinds of components of cytoplasmatic organelles were determined with transmission electron microscope, scanning electron microscope, and energy-dispersive X-ray microanalysis. The results of energy-dispersive X-ray microanalysis indicated that Eu³⁺ and La³⁺ can be absorbed into plant cells and bind to the membranes of protoplasm, chloroplast, mitochondrion, cytoplast, and karyon. These results provide experimental evidence that rare earth elements can be absorbed into plant cells, which would be the basis for interpreting physiological and biochemical effects of rare earth elements on plant cells.     

稀土La3+跨PC12细胞膜行为研究

使用AR-CM-M1C阳离子测定系统,发展Fura-2荧光测定技术,将其应用于测定细胞内游离稀土离子La³⁺,并以此研究了La³⁺跨PC12细胞（大鼠嗜铬细胞瘤细胞）膜的行为.结果表明：在模拟细胞内离子组分,pH=7.05的溶液中,测得La³⁺-Fura-2的表观解离常数为3.27×10^-11 mol·L^-1.对于PC12细胞,静息条件下La³⁺不能跨越细胞膜进入胞内.与钙离子通道相关的KCl和去甲肾上腺素均不能刺激稀土La³⁺过膜.用哇巴因（ouabain）使胞内Na⁺超载后,La³⁺可过膜进入细胞内,且过膜量与胞外La³⁺浓度和胞内Na⁺超载程度有一定的浓度依赖关系,提示La³⁺可以经由Na⁺／La³⁺交换机制过膜而进入细胞内.     

Effects of La3+ on growth,transformation, and gene expression of Escherichia coli

Rare earth elements have been emitted into the environment largely as fertilizer components. This has caused much fear about whether they would influence our environment, especially on the metabolism and genetics of microorganisms. In this article, the trivalent ion of a rare earth element, lanthanum, was studied for the effects on growth, transformation, and gene expression of Escherichia coli. The results showed that La³⁺ at concentrations from 50 to 150 μg/mL stimulated the endogenic metabolism and ectogenic metabolism, but had few effects on gene expression. La³⁺ at lower concentrations from 0.5 to 30 μg/mL inhibit intensively E. coli-absorbing external DNA, decreasing the transformation efficiency. It is also supported by observations using transmission electron microscopy. Our results are significant in understanding the function of rare earth elements to microorganisms and assessing the risk of application of rare earth compounds.     

Effect of Sm3+ ion on growth of Bacillus thuringiensis by microcalorimetry

By using an LKB-2277 Bioactivity Monitor, cycle-flow method, the thermogenic curves of aerobic growth for Bacillus thuringiensis cry II strain at 28°C have been obtained. The metabolic thermogenic curves of B. thuringiensis cry II contained two distinct patterns: the first reflects the changes during the bacterial growth phase and the second corresponds to the sporulation phase. From these thermogenic curves in the absence and presence of Sm³⁺ ions, the thermokinetic parameters such as the growth rate constants k, the interval time τ_I, the maximum power P _{max 1} and heat-output Q _log for log phase, the maximum power P _{max 2} and heat-output Q _stat for stationary phase, the heat-output Q _spor for sporulation phase and total heat effects Q _T are calculated. Sm³⁺ ion has promoting action on the growth of B. thuringiensis cry II in its lower concentration range; on the other hand, this ion has inhibitory action on the sporulation of B. thuringiensis in its higher concentration range. We also found that the effects of Sm³⁺ ion on B. thuringiensis during the sporulation phase were far greater than that during the bacterial phase. It is concluded that the application of B. thruringiensis of controlling insecticides is not affected by the presence of the rare-earth elements in the environmental ecosystem.     

Ba2+ uptake and the inhibition by Ba2+ of K+ flux into rat liver mitochondria

Summary Rapid uptake of Ba²⁺ by respiring rat liver mitochondria is accompanied by a transient stimulation of respiration. Following accumulation of Ba²⁺, e.g. at a concentration of 120 nmol per mg protein, the mitochondria exhibit reduced rates of state 3 and uncoupler-stimulated respiration. ADP-stimulated respiration is inhibited at a lower concentration of Ba²⁺ than is required to affect uncoupler-stimulated respiration, suggesting a distinct effect of Ba²⁺ on mechanisms involved in synthesis of ATP. Ba²⁺, which has an ionic radius similar to that of K⁺, inhibits unidirectional K⁺ flux into respiring rat liver mitochondria. This effect on K⁺ influx is observable at concentrations of Ba²⁺, e.g. 23 to 37 nmol per mg protein, which cause no significant change in state 4 or uncoupler-stimulated respiration. The accumulated Ba²⁺ decreases the measuredV _max of K⁺ influx, while having little effect on the apparentK _m for K⁺. The inhibition of K⁺ influx by Ba²⁺ is seen in the presence and absence of mersalyl, an activator of K⁺ influx. In contrast, under the conditions studied, Ba²⁺ has no apparent effet on the rate of unidirectional K⁺ efflux. These data are consistent with the idea that K⁺ may enter and leave mitochondria via spearate mechanisms.     

H+-dependent efflux of Ca2+ from heart mitochondria

A rapid loss of accumulated Ca²⁺ is produced by addition of H⁺ to isolated heart mitochondria. The H⁺-dependent Ca⁺ efflux requires that either (a) the NAD(P)H pool of the mitochondrion be oxidized, or (b) the endogenous adenine nucleotides be depleted. The loss of Ca²⁺ is accompanied by swelling and loss of endogenous Mg^2–. The rate of H⁺-dependent Ca²⁺ efflux depends on the amount of Ca²⁺ and P_i taken up and the extent of the pH drop imposed. In the absence of ruthenium red the H⁺-induced Ca²⁺-efflux is partially offset by a spontaneous re-accumulation of released Ca²⁺. The H⁺-induced Ca²⁺ efflux is inhibited when the P_i transporter is blocked withN-ethylmaleimide, is strongly opposed by oligomycin and exogenous adenine nucleotides (particularly ADP), and inhibited by nupercaine. The H⁺-dependent Ca²⁺ efflux is decreased markedly when Na⁺ replaces the K⁺ of the suspending medium or when the exogenous K⁺/H⁺ exchanger nigericin is present. These results suggest that the H⁺-dependent loss of accumulated Ca²⁺ results from relatively nonspecific changes in membrane permeability and is not a reflection of a Ca²⁺/H⁺ exchange reaction.     

Ce^3+和La^3+对人工老化沙葱种子活力及生理特性的影响

以未老化和人工老化后的沙葱(Allium mongolicum Regel.)种子为材料,采用氯化铈(Ce3+)和氯化镧(La3+)浸种,测定种子萌发和生理指标,探讨Ce3+和La3+浸种对种子萌发、老化种子活力和生理特性的影响。结果显示:(1)在老化0~5 h时,Ce3+和La3+处理可显著促进沙葱种子萌发,提高种子活力;在老化5 h后,Ce3+和La3+处理对种子萌发无明显促进作用。(2)在老化0~15 h时,Ce3+和La3+处理的沙葱种子中抗氧化酶活性和抗坏血酸(AsA)含量提高,其超氧阴离子自由基(O2-·)产生速率、过氧化氢(H2O2)含量和丙二醛(MDA)含量显著降低;在老化15 h后,Ce3+和La3+处理的种子抗氧化酶活性提高、AsA含量降低,O2-·产生速率和MDA含量提高。(3)在老化5 h时,沙葱种子呼吸速率发生跃变达到最大,Ce3+和La3+处理显著降低了种子呼吸速率。(4)Ce3+和La3+处理在老化0~5 h时提高了沙葱种子超弱发光(UWL)强度,但在老化5 h后沙葱种子的UWL强度降低。研究认为,在沙葱种子人工老化初期,Ce3+和La3+浸种处理可以诱导增强种子抗氧化酶活性和提高AsA含量,有效清除因老化产生积累的过量活性氧(ROS),减轻过氧化伤害,提高种子活力;种子老化中后期,其内部ROS产生与清除系统发生紊乱,加剧了ROS对种子结构的损伤,Ce3+和La3+浸种处理的缓解效应丧失。     

Microcalorimetric study of the toxic effect of sodium selenite on the mitochondria metabolism of Carassius auratus liver

The fundamental thermogenesis curves of the metabolic process of liver mitochondria from Carassius auratus and the toxic effect of Na₂SeO₃ on it were studied by using an LKB-2277 bioactivity monitor, ampoule method, at 28°C. From the thermogenesis curves, the thermokinetic equations were established under different conditions. The kinetics show that a low concentration of Na₂SeO₃ (1–4 mg/L) had promoting action on the metabolism process of Carassius auratus liver mitochondria, but that a high concentration of Na₂SeO₃ (8–16 mg/L) inhibited the mitochondria metabolism.     

Rapid release of Mg(2+) from liver mitochondria by nonesterified long-chain fatty acids in alkaline media

Long-chain fatty acids induce a rapid release of Mg(2+) from both energized and nonenergized rat liver mitochondria suspended at pH 8 in isotonic saline but not sucrose media. The effect is observed only with fatty acids that possess protonophoric activity. The most active saturated fatty acids are myristic and palmitic, while the most active unsaturated acids are oleic, linolenic, and arachidonic. The rate of Mg(2+) release drastically decreases with decreasing medium pH to 7.2-7.6. However, at those pH values this rate is doubled by energization of mitochondria with respiratory substrates. Mg(2+) release is accompanied by cyclosporin A-insensitive large-amplitude swelling of mitochondria. This swelling is similar to that produced by the divalent metal ionophore A23187 and is interpreted as being due to activation of the inner membrane anion channel, the K(+) uniporter, and the K(+)/H(+) exchanger. In energized mitochondria, both swelling and Mg(2+) release are blocked by the exogenous K(+)/H(+) exchanger nigericin. It is proposed that fatty acids under conditions of alkaline mitochondrial matrix activate latent Mg(2+)-sensitive ion-conducting pathways in the inner mitochondrial membrane, which mediate swelling and Mg(2+) release. It is hypothesized that fatty acids activate an intrinsic Mg(2+)/H(+) exchanger that is related to, or identical with, the K(+)/H(+) exchanger.     

Ca2+ transport in mitochondria from yeast expressing recombinant aequorin

We have expressed aequorin in mitochondria of the yeast Saccharomyces cerevisiae and characterized the resulting strain with respect to mitochondrial Ca(2+) transport in vivo and in vitro. When intact cells are suspended in water containing 1.4 mM ethanol and 14 mM CaCl(2), the matrix free Ca(2+) concentration is 200 nM, similar to the values expected in cytoplasm. Addition of ionophore ETH 129 allows an active accumulation of Ca(2+) and promptly increases the value to 1.2 microM. Elevated Ca(2+) concentrations are maintained for periods of 6 min or longer under these conditions. Isolated yeast mitochondria oxidizing ethanol also accumulate Ca(2+) when ETH 129 is present, but the cation is not retained depending on the medium conditions. This finding confirms the presence of a Ca(2+) release mechanism that requires free fatty acids as previously described [P.C. Bradshaw et al. (2001) J. Biol. Chem. 276, 40502-40509]. When a respiratory substrate is not present, Ca(2+) enters and leaves yeast mitochondria slowly, at a specific activity near 0.2 nmol/min/mg protein. Transport under these conditions equilibrates the internal and external concentrations of Ca(2+) and is not affected by ruthenium red, uncouplers, or ionophores that perturb transmembrane gradients of charge and pH. This activity displays sigmoid kinetics and a K(1/2) value for Ca(2+) that is near to 900 nM, in the absence of ethanol or when it is present. It is furthermore shown that the activity coefficient of Ca(2+) in yeast mitochondria is a function of the matrix Ca(2+) content and is substantially larger than that in mammalian mitochondria. Characteristics of the aequorin-expressing strain appear suitable for its use in expression-based methods directed at cloning Ca(2+) transporters from mammalian mitochondria and for further examining the interrelationships between mitochondrial and cytoplasmic Ca(2+) in yeast.     

Effect of lanthanum ions (La3+) on the reactive oxygen species scavenging enzymes in wheat leaves

Physiological effects of lanthanum ions on the activities of the enzymes in the reactive oxygen species (ROS) scavenging system in leaves of wheat (Triticum aestivum L.) seedlings were studied. Wheat leaves treated in Hogland solution with 0.1 mM LaCl₃ for 48 h showed increased levels of superoxide dismutase (SOD), catalase (CAT), ascorbate-specific peroxidase (AsA-POD), and dehydroascorbate reductase (DHAR). However, a minor effect was observed on the levels of monodehydroascorbate reductase (MDAR) and glutathione reductase (GR), which regulate the release of energy required by the ROS scavenging system. The whole system was linked up by H⁺ transmission. Our results indicated that the activities of the enzymes that function directly to remove ROS were elevated by La³⁺ treatment, which is consistent with the observations that La³⁺-treated plants had increased tolerance to environmental stresses. The remaining levels of MDAR and GR suggested that these two enzymes might be regulated differently from that of the other four enzymes studied.     

NADH oxidation drives respiratory Na+ transport in mitochondria from Yarrowia lipolytica

It is generally assumed that respiratory complexes exclusively use protons to energize the inner mitochondrial membrane. Here we show that oxidation of NADH by submitochondrial particles (SMPs) from the yeast Yarrowia lipolytica is coupled to protonophore-resistant Na⁺ uptake, indicating that a redox-driven, primary Na⁺ pump is operative in the inner mitochondrial membrane. By purification and reconstitution into proteoliposomes, a respiratory NADH dehydrogenase was identified which coupled NADH-dependent reduction of ubiquinone (1.4 μmol min⁻¹ mg⁻¹) to Na⁺ translocation (2.0 μmol min⁻¹ mg⁻¹). NADH-driven Na⁺ transport was sensitive towards rotenone, a specific inhibitor of complex I. We conclude that mitochondria from Y. lipolytica contain a NADH-driven Na⁺ pump and propose that it represents the complex I of the respiratory chain. Our study indicates that energy conversion by mitochondria does not exclusively rely on the proton motive force but may benefit from the electrochemical Na⁺ gradient established by complex I. Electronic supplementary material  The online version of this article (doi:) contains supplementary material, which is available to authorized users.     

Interaction of Sr2+ with Ca2+-induced Ca2+ release in mitochondria

Respiring rat liver mitochondria are known to spontaneously release the Ca²⁺ taken up when they have accumulated Ca²⁺ over a certain threshold, while Sr²⁺ and Mn²⁺ are well tolerated and retained. We have studied the interaction of Sr²⁺ with Ca²⁺ release. When Sr²⁺ was added to respiring mitochondria simultaneously with or soon after the addition of Ca²⁺, the release was potently inhibited or reversed. On the other hand, when Sr²⁺ was added before Ca²⁺, the release was stimulated. Ca²⁺-induced mitochondrial damage and release of accumulated Ca²⁺ is generally believed to be due to activation of mitochondrial phospholipase A (EC 3.1.1.4.) by Ca²⁺. However, isolated mitochondrial phospholipase A activity was little if at all inhibited by Sr²⁺. The Ca²⁺ -release may thus be triggered by some Ca²⁺ -dependent function other than phospholipase.     

Influence of Ca2+ channel modulators on [3H]purine release from rat cultured glial cells

[³H]Purine release from rat striatum astrocyte cultures was studied at 14 days in vitro (DIV). Superfusion of cultures with a Ca²⁺-free medium +0.5 mM ethylene glycol-bis(-aminoethylether)N,N,N,N-tetracetic acid (EGTA) reduced the electrically evoked [³H]purine release. Nimodipine only at the concentration of 10 M modified [³H]purine outflow whereas 0.1 M -conotoxin and 0.03–0.1 M nitrendipine reduced the evoked one. Superfusion of cultures with 0.1 M -conotoxin +0.1 M nitrendipine antagonized the evoked [³H]purine release similarly to each drug given alone. Neither nitrendipine nor -conotoxin influenced the uptake of⁴⁵Ca²⁺ by the cultures. The treatment of cells with the Ca²⁺ agonist Bay K 8644 did not affect [³H]purine release or the⁴⁵Ca²⁺ uptake. The drug did not either alter [Ca²⁺]_i, evaluated by loading the cells with 3 M Fura-2/AM. 10–30 M 3,4,5-trimethoxybenzoic acid 8-(diethylamino)octyl ester (TMB-8), a blocker of intracellular Ca²⁺ discharge, significantly reduced the evoked [³H]purine release. On the other hand, 2 M thapsigargin, an inhibitor of the ion store Ca²⁺ ATPase, was able to increase either the culture [³H]purine release or the [Ca²⁺]_i. Together, the findings indicate that voltage-sensitive calcium channels (VSCC_s) of the neuronal N and L-types are not involved in the modulation of [³H]purine release from rat cultured astrocytes whereas Ca²⁺ coming from intracytoplasmic stores seems to play a prevailing role. Moreover, agents which block VSCC_s seem to be able to affect [³H]purine outflow with mechanisms other than VSCC gating.     

Mitochondrial free Ca2+ concentration in living cells

Evidence has accrued during the past two decades that mitochondrial Ca²⁺ plays an important role in the regulation of numerous cell functions such as energy metabolism. This implies that mitochondrial Ca²⁺ transport systems might be able to relay the changes of cytosolic Ca²⁺ concentration ([Ca²⁺]_c) into mitochondrial matrix for regulating biochemical activities. To substantiate this idea, measurements of intramitochondrial free Ca²⁺ concentration ([Ca²⁺]_m) become essential. In this article, we review the results from recent studies attempting to measure [Ca²⁺]_m in living cells. In addition, the significance of each study is discussed.     

Stimulation effect of lithium on the metabolic activity of liver tissue mitochondria measured by microcalorimetry

The effect of Li(I) on the metabolism of mitochondria isolated from Carassius auratus liver tissue was investigated by microcalorimetric method to provide evidence for mitochondria hypothesis of biporlar disorder (BPD) and to explore therapeutic mechanism of drug for treatment of BPD. Obvious stimulation induced by Li(I) on mitochondria metabolism was reflected by power-time (P-t) curves. The power-time curves of hepatic mitochondria metabolism without Li(I) could be divided into four parts: lag phase, active recovery phase, stationary phase, and decline phase. When Li(I) was added, the second heat peak occurred in a concentration-dependent sequence. Considering the first heat peak on the p-t curves, Li(I) in the range of therapeutic and lower concentration induced slight alterations in comparison with the characteristic heat peak observed in the control. However, Li(I) above the therapeutic concentration resulted in significant changes. Heat output increased with the concentration of Li(I), but the rate constant (k ₂) and the maximum heat power (P _max2) for the second heat peak reached maximum value in the range of therapeutic concentration. Mechanism of activation of mitoKatp was suggested and discussed.