Erythrocyte Lii-Nao countertransport system Inhibition by N-ethylmaleimide probes for a conformational change of the transport system

Human erythrocytes were treated by a series of SH-reagents, including maleimides, iodo compounds, mercurials and oxidizing agents. Rates of Li efflux into Na-rich medium, Li leak and Li_i-Na_o countertransport were then determined. Of the 13 different reagents studied, only N-ethylmaleimide, iodoacetamide and iodoacetate inhibited selectively the countertransport activity. The effect of the various reagents indicates that the sensitive SH-groups of the countertransport system are not externally exposed. N-Ethylmaleimide was used to probe for changes elicited by substrate cations in Li_i-Na_o countertransport. In Na- and Li-free medium, inhibition of Li_i-Na_o countertransport by N-ethylmaleimide of 35% was reached within 2 s. In Na or Li medium, maximal inhibition was twice as great, but was attained much more slowly, within 10 min. Kinetic data and Hill plot analysis indicate the involvement of two classes of SH-groups: one expressed in the various media with and without substrate cations, and an additional one, which becomes specifically available to N-ethylmaleimide in the presence of external Na or Li. The affinity of Na to the site promoting inhibition by N-ethylmaleimide (apparent K_m  12 mM) is higher than the affinity of Na to its external countertransport site (apparent K_m  25 mM), as reported by Sarakadi, B., Alifimoff, J.K., Gunn, R.B. and Tosteson, D.C. (1978) J. Gen. Physiol. 72, 249–265). Reactivity of $\text{N-}\text{ethyl}\text{[}{}^{14}\text{C}\text{]}\text{maleimide}$ was not modified by the media tested. It is concluded that external Na and Li cause a conformational change in the protein(s) of the countertransport system in human erythrocytes.     

Temperature dependence of the photosynthetic activities in the thylakoid membranes from the blue-green alga Anacystis nidulans

The photosynthetic electron transport and phosphorylation reactions were measured in the room temperature region in the thylakoid membranes prepared from the blue-green alga, Anacystis nidulans. The Arrhenius plot of the Hill reaction with 2,6-dichlorophenolindophenol showed a distinct break of straight lines at 21°C in the membranes from cells grown at 38°C, and at 12°C in those from cells grown at 28°C. The Arrhenius plot of the Hill reaction with ferricyanide showed a break at 13°C in the membranes from cells grown at 38°C, and at 7°C in those from cells grown at 28°C. On the other hand, the Arrhenius plot of the System I reaction with methylviologen as an electron acceptor and 2,6-dichlorophenolindophenol and ascorbate as an electron donor system was composed of a straight line in the membranes from cells grown at 28°C as well as at 38°C. The Arrhenius plot of the System II reaction measured by the ferricyanide reduction mediated by silicotungstate in the presence of 3-(3′,4′-dichlorophenyl)-1,1-dimethylurea also showed a break at 11°C in the membranes from cells grown at 38°C.The Arrhenius plot of the phosphorylation mediated by N-methylphenazonium methylsulfate showed a break at 21°C in the membranes from cells grown at 38°C and at 12°C in those from cells grown at 28°C. The Arrhenius plot of the phosphorylation mediated by the System I reaction showed a break at 24°C in the membranes from cells grown at 38°C.The characteristic features in the Arrhenius plots of the photosynthetic electron transport and phosphorylation reactions are discussed in terms of the transition of physical phase of the thylakoid membrane lipids.     

Phospholipase C digestion of rat liver plasma membrane stimulates adenylate cyclase

Brief treatment of rat liver plasma membranes with phospholipase C of Clostridium welchii increased both the ratio of saturated to unsaturated fatty acids and the ratio of cholesterol to phospholipids. Using 5-doxylstearic acid spin probes two breaks at 29 and 19.6 °C could be observed in the order parameter, S_A, vs temperature curve for untreated membranes. Upon phospholipase C digestion the lower phase transition temperature was shifted to 23 °C, while the higher phase transition temperature could not be detected up to 40 °C. The order parameter, S_A, was consistently higher at all temperatures in the phospholipase C-treated membranes. As phospholipase C is known to attack the outer lamella, these results can be interpreted as indicating an increase in ordering (i.e., decrease in fluidity) of the outer membrane lamella. On the other hand, an increase in basal activity of adenylate cyclase of the treated membranes was observed with an apparent reduction of the activation energies both below and above the break (at 20 °C) in the Arrhenius plot of enzyme activity. Phospholipase C treatment did not affect the temperature of the break in Arrhenius kinetics of the enzyme. The results are discussed in terms of the role of the ordering state of membrane lipids in adenylate cyclase activity.     

The erythrocyte membrane in essential hypertension: Modified temperature-dependence of Li+ efflux

The rate of ouabain-resistant Li⁺-efflux was studied in erythrocytes of normal controls and of patients with essential hypertension. Despite variability in rate, erythrocytes from normotensive persons revealed a uniform pattern of temperature dependence of the efflux, with two slopes (K_a = 9.4 and 19.1 kcal/mol, respectively) and a transition at about 25°C. Erythrocytes from the patients showed both a higher rate of Li⁺ efflux and significant changes in the temperature repsonse, with essentially a single slope (K_a = 14 kcal/mol). The data indicate localized changes in the membrane organization of hypertensive erythrocytes, involving lipid-protein interaction.     

Characteristic temperature dependences of respiratory and photosynthetic electron-transport activities in membrane preparations from Anacystis nidulans grown at different temperatures

Electron-transport activities supported by seven different electron donor/acceptor couples in the light and in the dark, respectively, were measured in particle preparations of the cyanobacterium (blue-green alga) Anacystis nidulans after growth at 40, 30 and 25°C. The Arrhenius plots of the photosynthetic electron-transport reactions between ascorbate (plus 2,6-dichlorophenolindophenol (DCIP)) and NADP⁺, diphenylcarbazide and DCIP, diaminodurene and benzyl viologen (O₂), and the plot of the photooxidation of reduced horse heart cytochrome c showed a single discontinuity at approx. 24–25, 15–17 and 10–13°C in membranes derived from cells grown at 40, 30 and 25°C, respectively. By contrast, the dark respiratory electron-transport reactions between NADPH, ascorbate (plus DCIP) or reduced horse heart cytochrome c and oxygen, and the reduction by horse heart cytochrome c of the aa₃-type terminal oxidase as followed directly by dual-wavelength spectrophotometry, all gave Arrhenius plots distinguished by two distinct breaks: The break at the higher temperature corresponded to the break also found in the Arrhenius plots of the photosynthetic reactions while an additional discontinuity was observed at 17–18, 8–9 and 5–6°C in membranes prepared from cells grown at 40, 30 and 25°C, respectively. The temperatures at which the discontinuities in the Arrhenius plots occurred depended on the temperature at which the cells had been grown; they were independent, however, of the specific electron donors and acceptors employed. The characteristic features in the Arrhenius plots of respiratory and photosynthetic electron-transport reactions are discussed in terms of lipid-phase transitions in the cytoplasmic and the intracytoplasmic (thylakoid) membranes of A. nidulans. Implications for possibly distinct sites of the respiratory and photosynthetic electron-transport systems in A. nidulans will be mentioned.     

Peripheral resistance and red cell LiNa countertransport in borderline hypertensives

We have studied ouabain-resistant, external sodium-stimulated, lithium efflux (LiNa countertransport) in red blood cells from 21 borderline hypertensives with at least one hypertensive first degree relative (BH-F), 19 borderline hypertensives without family history of essential hypertension (BH-NF), and 35 age-matched normotensive subjects. The data indicate the finding of an increased LiNa countertransport in all BH (F+NF), but with a significant overlap between BH values and control ones: LiNa countertransport is significantly higher only in BH-F but it is normal in BH-NF. Moreover, there is a significant correlation of LiNa countertransport to total peripheral resistance but not to mean blood pressure in all hypertensive patients. It is suggested that in BH the increase of erythrocyte Na flux is mediated by the NaNa exchange diffusion, and its abnormality may be associated to the hereditary trait of essential hypertension rather than the high blood pressure per se, probably resulting in the development of hypertension, through the increased vascular smooth muscle tone.     

Membrane phase transitions and succinate oxidase activity in an extremely thermophilic bacterium

ESR and succinate oxidase activity were used to investigate the membrane phase-transitions of an extreme thermophile, Thermus T351, over an 80°C temperature range in whole cells, membrane particles, and extracted lipid suspension. Three phase transitions were observed using both techniques. These occurred at about 19°C, 39°C and 66°C. The transition at 19°C is unusual in that the Arrhenius plot for succinate oxidase is concave upwards, implying an increase in activation energy (E_a) with increased temperature.     

Effect of different temperatures to remove reduction gas on the photoluminescence properties of Eu‐doped Li2(Ba1‐xSrx)SiO4 phosphors

In this study, Eu‐doped Li₂(Ba_1‐xSr_x)SiO₄ powders (x = 0, 0.2, 0.4, and 0.6) were synthesized at 850°C in a reduction atmosphere (5% H₂ + 95% N₂) for a duration of 1 h using a solid‐state reaction method. The reduction atmosphere was infused as the synthesis temperature reached 850°C, and was removed as the temperature dropped to 800–500°C. Li₂(Ba_1‐xSr_x)SiO₄ (or Li₂BaSiO₄), (Ba,Sr)₂SiO₄ (or BaSiO₄), and Li₄SiO₄ phases co‐existed in the synthesized Eu‐doped Li₂(Ba_1‐xSr_x)SiO₄ powders. A new finding was that the reduction atmosphere removing (RAR) temperature of the Li₂(Ba_1‐xSr_x)SiO₄ phosphors had a large effect on their photoluminescence excitation (PLE) and PL properties. Except for the 800°C‐RAR‐treated Li₂BaSiO₄ phosphor, PLE spectra of all other Li₂(Ba_1‐xSr_x)SiO₄ phosphors had one broad emission band with two emission peaks centred at ~242 and ~283 nm; these PL spectra had one broad emission band with one emission peak centred at 502–514 nm. We showed that the 800°C‐RAR‐treated Li₂BaSiO₄ phosphor emitted a red light and all other Li₂(Ba_1‐xSr_x)SiO₄ phosphors emitted a green light. Reasons for these results are discussed thoroughly.     

Passive potassium ion permeability of Halobacterium halobium cell envelope membranes

Cell envelope vesicles, prepared from Halobacterium halobium, were loaded with 3 M KCl suspended in 3 M NaCl, and the loss of K⁺ was followed at various temperatures. The Arrhenius plot of the K⁺-efflux rates shows a break at 30°C, with higher energy of activation above the break. This temperature dependence is consistent with earlier studies of chain motions in liposomes prepared from isolated lipids. The efflux of K⁺ is more rapid with increasing pH between pH 5 and 7. Since these vesicles do not respire under the experimental conditions it was expected that the K⁺-efflux data would be related to the passive permeability of the membranes to K⁺. The apparent K⁺ permeability at 30°C is 1–2· 10^?10 cm·^?1. This value corresponds to a 5-h half-life for retained K⁺ in the envelope vesicles and to a probably much longer half-life in whole cells. The previously observed ability of Halobacterium to retain K⁺ in the absence of metabolism can thus be explained solely by the permeability characteristics of the membranes.     

Effects of hypothermia on short latency somatosensory evoked potentials in humans

Short latency somatosensory evoked potentials (SSEPs) elicited by median nerve stimulation were monitored in 14 adult patients undergoing cardiac surgery under cardiopulmonary bypass and induced hypothermia. SSEPs were recorded at 1–2°C steps as the body temperature was lowered from 37°C to 20°C to determine temperature-dependent changes. Hypothermia produced increased latencies of the peaks of N₁₀, P₁₄ and N₁₉ components, the prolongation was more severe for the later components so that N₁₀−P₁₄ and P₁₄−N₁₉ interpeak latencies were also prolonged. The temperature-latency relationship had a linear correlation. The magnitude of latency prolongation (msec) with 1°C decline in temperature was 0.61, 1.15, 1.56 for N₁₀,P₄ and N₁₉ components, respectively, and 0.39 and 0.68 for interpeak latencies N₁₀−P₁₄ and P₁₄−N₁₉, respectively. The rise time and duration of the 3 SSEP components increased progressively with cooling. Cortically generated component, N₁₉ was consistently recordable at a temperature above 26°C, usually disappearing between 20°C and 25°C. On the other hand, more peripherally generated components, N₁₀ and P₁₄, were more resistant to the effect of hypothermia; P₁₄ was always elicitable at 21°C or above, whereas N₁₀ persisted even below 20°C. The amplitude of SSEP components had a poor correlation with temperature; there was a slight tendency for N₁₀ and P₁₄ to increase and for N₁₉ to decrease with declining temperature. Because incidental hypothermia is common in comatose and anesthetized patients, temperature-related changes must be taken into consideration during SSEP monitoring under these circumstances.     

Suppressing Li Dendrite Formation in Li2S‐P2S5 Solid Electrolyte by LiI Incorporation

Solid electrolytes have been considered as a promising approach for Li dendrite prevention because of their high mechanical strength and high Li transference number. However, recent reports indicate that Li dendrites also form in Li₂S‐P₂S₅ based sulfide electrolytes at current densities much lower than that in the conventional liquid electrolytes. The methods of suppressing dendrite formation in sulfide electrolytes have rarely been reported because the mechanism for the “unexpected” dendrite formation is unclear, limiting the successful utilization of high‐energy Li anode with these electrolytes. Herein, the authors demonstrate that the Li dendrite formation in Li₂S‐P₂S₅ glass can be effectively suppressed by tuning the composition of the solid electrolyte interphase (SEI) at the Li/electrolyte interface through incorporating LiI into the electrolyte. This approach introduces high ionic conductivity but electronic insulation of LiI in the SEI, and more importantly, improves the mobility of Li atoms, promoting the Li depositon at the interface and thus suppresses dendrite growth. It is shown that the critical current density is improved significantly after incorporating LiI into Li₂S‐P₂S₅ glass, reaching 3.90 mA cm^?2 at 100 °C after adding 30 mol% LiI. Stable cycling of the Li‐Li cells for 200 h is also achieved at 1.50 mA cm^?2 at 100 °C.     

Bovine adrenocortical microsomal hydroxylase and thermotropic transitions substrate-cytochrome P-450 binding reaction versus substrate hydroxylation

The effect of temperture on steroid C-21 hydroxylation and substrate-cytochrome P-450 binding reaction under turnover conditions (NADPH + O₂ are investigated. The Arrhenius activity plot exhibited a single break, while the van 't Hoff plot of the substrate dissociation constant (K_s) exhibited four breaks between 10 and 40°C which corresponded to the characteristic temperatures of the lipids' phase transitions. Unlike the case of the K_s value, the detergent Triton X-114 was without effect on the Arrhenius activity plot. This indicates that the single break in the case of the enzyme activity is distinct from but not necessarily independent of the multiple breaks in the case of the K_s. At physiologic temperature and concentration of the substrate, the free energy (?9.5 kcal/mol) of the substrate-cytochrome binding reaction is more than sufficient to account for the apparent activation energy (6.6 kcal/mol) of the overall hydroxylation. This suggests that the substrate-cytochrome P-450 binding reaction has the potential of being a source of energy for the overall reaction.     

Kinetics of water loss and the likelihood of intracellular freezing in mouse ova

To avoid intracellular freezing and its usually lethal consequences, cells must lose their freezable water before reaching their ice-nucleation temperature. One major factor determining the rate of water loss in the temperature dependence of the water permeability,L _p (hydraulic conductivity). Because of the paucity of water permeability measurements at subzero temperatures, that temperature dependence has usually been extrapolated from above-zero measurements. The extrapolation has often been based on an exponential dependence ofL _p on temperature. This paper compares the kinetics of water loss based on that extrapolation with that based on an Arrhenius relation betweenL _p and temperature, and finds substantial differences below ?20 to ?25°C. Since the ice-nucleation temperature of mouse ova in the cryoprotectants DMSO and glycerol is usually below ?30°C, the Arrhenius form of the water-loss equation was used to compute the extent of supercooling in ova cooled at rates between 1 and 8°C/min and the consequent likelihood of intracellular freezing. The predicted likelihood agrees well with that previously observed. The water-loss equation was also used to compute the volumes of ova as a function of cooling rate and temperature. The computed cell volumes agree qualitatively with previously observed volumes, but differ quantitatively.     

Discontinuous thermotropic response of Tetrahymena membrane lipids correlated with specific lipid compositional changes

Steady-state fluorescence polarization measurements of 1,6-diphenyl-1,3,5-hexatriene in microsomal lipids from Tetrahymena pyriformis cells grown at 39 or 15°C revealed discrete slope discontinuities in plots of polarization vs. temperature. Two well-defined ‘break points’ were present in the 0–40°C temperature range examined and their precise location was dependent upon the growth temperature of the cells. By mixing phospholipids from cells grown at different temperatures, the break points at 17.5 and 32°C in 39°C-lipid multilayer preparations were shown to correlate with the breaks at 12 and 27°C, respectively, in similar preparations from 15°C-grown cells. The discrete break points were also present, but at slightly different characteristic temperatures, in a phosphatidylcholine fraction and a phosphatidylethanolamine plus 2-aminoethylphosphonolipid fraction purified from the phospholipids and in total microsomal lipids (phospholipids plus the sterol-like triterpenoid, tetrahymanol). However, catalytic hydrogenation of the phospholipid fatty acids or mixing the non-hydrogenated phospholipids with increasing proportions of synthetic dipalmitoyl phosphatidylcholine eliminated the break points. We interpret this discontinuous thermotropic response in microsomal lipids as signalling a lipid phase separation of importance in regulating physiological events.     

Countering Voltage Decay and Capacity Fading of Lithium‐Rich Cathode Material at 60 °C by Hybrid Surface Protection Layers

Li‐rich layered metal oxides have attracted much attention for their high energy density but still endure severe capacity fading and voltage decay during cycling, especially at elevated temperature. Here, facile surface treatment of Li_1.17Ni_0.17Co_0.17Mn_0.5O₂ (0.4Li₂MnO₃·0.6LiNi_1/3Co_1/3Mn_1/3O₂) spherical cathode material is designed to address these drawbacks by hybrid surface protection layers composed of Mg²⁺ pillar and Li‐Mg‐PO₄ layer. As a result, the surface coated Li‐rich cathode material exhibits much enhanced cycling stability at 60 °C, maintaining 72.6% capacity retention (180 mAh g^?1) between 3.0 and 4.7 V after 250 cycles. More importantly, 88.7% average discharge voltage retention can be obtained after the rigorous cycle test. The strategy developed here with novel hydrid surface protection effect can provide a vital approach to inhibit the undesired side reactions and structural deterioration of Li‐rich cathode materials and may also be useful for other layered oxides to increase their cycling stability at elevated temperature.     

Potassium and Sodium Fluxes in the PhytopathogenicFungus Fusarium oxysporum var. lini

Rubidium uptake in potassium-starved cells followed biphasic kinetics in the micromolar and millimolar range and was independent of the temperature. In contrast, Rb⁺ uptake in normal-K⁺ cells followed a monophasic kinetics in the millimolar range and increased at temperatures higher than 30°C. Differences in the K _m values and in the Arrhenius plots of Rb⁺ uptake suggest different uptake systems in K⁺-starved and in normal-K⁺ cells. In addition, the substantial inhibition of Rb⁺ uptake caused by carbonyl cyanide-m-chlorophenyl hydrazone indicates that these systems are strongly dependent on membrane voltage. Lithium (sodium) tolerance, influx, and efflux were separately studied. F. oxysporum was shown to be very tolerant to sodium, while lithium caused a specific toxic effect. Li⁺ uptake in K⁺-starved cells exhibits a monophasic kinetics with low affinity. Li⁺ efflux was not affected by external pH or addition of potassium to the medium, suggesting that a Na⁺/cation antiporter is not involved in this process. Received: 14 March 2000 / Accepted: 5 June 2000     

Low Root Temperature Retardation of the Mineral Nitrogen Induced Decline in N2Fixation by a Northern Ecotype of White Clover

Nodulated white clover plants (Trifolium repensL.) of a Norwegian ecotype from Pasvik (70°N) were grown in flowing solution culture. Root temperature was 17°C until 51d after sowing, when it was lowered decrementally over 5d to 7°C in four of the eight plant culture units. After a further 24h, mineral N was supplied automatically at 20μMNH₄NO₃in three culture units at each root temperature (7 and 17°C) over 17d. The remaining two units provided control plants solely dependent on N₂fixation at 7 and 17°C.The supply of NH₄NO₃greatly reduced the nodule biomass per plant at 17°C over 17d compared with control plants, but had little effect at 7°C. The nodule decline at 17°C accompanied an acute and progressive decrease in specific rate of N₂fixation, from 9mmolN d^-1g^-1nodule d.wt on day 0 to zero by day 10. Whilst initial rates of N₂fixation were lower at 7°C, the mineral N-induced decrease in fixation rates was also less severe than at 17°C and specific fixation rates recovered after reaching a minimum on day 11. N₂fixation accounted for 36% of the total uptake of N by +min.N plants during the treatment period at 7°C as opposed to only 13% at 17°C. The total N₂fixed at 7°C was 86% of that fixed at 17°C, although the specific growth rate (d.wt) at 7°C was only 55% of that at 17°C. Addition of NH₄NO₃at 7°C had little effect on the gross amount of N₂fixed subsequently. In contrast, total N₂fixation by +min.N plants at 17°C was only 24% of that fixed by the corresponding controls. The possible mechanisms by which mineral N affects N₂fixation are discussed.     

Ultrathin,Flexible Polymer Electrolyte for Cost‐Effective Fabrication of All‐Solid‐State Lithium Metal Batteries

All‐solid‐state batteries are promising candidates for the next‐generation safer batteries. However, a number of obstacles have limited the practical application of all‐solid‐state Li batteries (ASSLBs), such as moderate ionic conductivity at room temperature. Here, unlike most of the previous approaches, superior performances of ASSLBs are achieved by greatly reducing the thickness of the solid‐state electrolyte (SSE), where ionic conductivity is no longer a limiting factor. The ultrathin SSE (7.5 µm) is developed by integrating the low‐cost polyethylene separator with polyethylene oxide (PEO)/Li‐salt (PPL). The ultrathin PPL shortens Li⁺ diffusion time and distance within the electrolyte, and provides sufficient Li⁺ conductance for batteries to operate at room temperature. The robust yet flexible polyethylene offers mechanical support for the soft PEO/Li‐salt, effectively preventing short‐circuits even under mechanical deformation. Various ASSLBs with PPL electrolyte show superior electrochemical performance. An initial capacity of 135 mAh g^?1 at room temperature and the high‐rate capacity up to 10 C at 60 °C can be achieved in LiFePO₄/PPL/Li batteries. The high‐energy‐density sulfur cathode and MoS₂ anode employing PPL electrolyte also realize remarkable performance. Moreover, the ASSLB can be assembled by a facile process, which can be easily scaled up to mass production.     

Effects of temperature on mitochondrial function in the Antarctic fish Trematomus bernacchii

Effects of temperature on O₂ consumption by mitochondria of the Antarctic fish Trematomus bernacchii were compared with effects obtained with mitochondria from tropical (Sarotheridon mossambica) and temperate zone fishes (Sebastes carnatus and Sebastes mystinus). Arrhenius plots of O₂ consumption versus temperature exhibited slope discontinuities (“breaks”) at temperatures (Arrhenius break temperatures: ABTs) reflective of the species' adaptation temperatures. The ABT for mitochondria of T. bernacchii is the lowest reported for any animal and is ∼12 °C below the value predicted by a regression equation based on ABT data for several invertebrates and fishes. The temperature at which the acceptor control ratio (ACR), an index of efficiency of coupling of electron transport to synthesis of ATP, began to decrease with rising temperature also reflected adaptation temperature. The decrease in ACR with rising temperature began at ∼18 °C for mitochondria of T. bernacchii, in contrast to ∼35 °C for mitochondria of Sarotheridon mossambica. Maintaining T. bernacchii at 4 °C for 2 weeks led to no changes in ABT or in the response of ACR to temperature. The thermal sensitivities of mitochondria of T. bernacchii reflect the high level of cold adaptation and stenothermy that is characteristic of Antarctic Notothenioid fishes. Accepted: 5 January 1998