Effects of Ammonia and Methylamine on Cl- Transport and on the pH Changes and Circulating Electric Currents Associated with HCO3- Assimilation in Chara corallina

Low concentrations of ammonia and methylamine greatly increaseCl^– influx into Chara corallina. Both amines have theirmaximum effect at pH 6.5–7.5. The amine stimulation ofCl^– influx is small below about pH 5.5. Above pH 8.5 theremay be inhibition of influx by amines. Concentrations of 10–25µM ammonia are sufficient to cause the maximum stimulationof Cl^– influx; the corresponding methylamine concentrationsare 0.1–0.2 mM. It is concluded that entry of amine cations(NH₄^$ and CH₃NH₃^$), rather than unionized bases (NH₃ and CH₃NH₂),causes Cl^– transport to be increased. Increases in rates of Cl^– transport are not necessarilyaccompanied by effects on HCO₃^$ assimilation and OH^– efflux.Measurements of localized pH differences at the cell surfaceand of circulating electric currents in the bathing solutionshow that these phenomena are only significantly affected byammonia at or above 50 µM and by methylamine at or above1.0 mM. The significance of the effects of amines is assessedin relation to current ideas about transport of Cl^–, HCO₃^–,and OH^–.     

Relation between Ultra-Violet Spectral Change and Nucleotide Binding of the Chloroplast Coupling Factor 1

Properties of the nucleotide binding sites on chloroplast couplingfactor 1 (CF₁) were studied by equilibrium dialysis and UV spectroscopy.From our direct binding studies, we identified at least fourkinds of ADP binding sites on CF₁; a barely dissociable ADPbinding site (site A), a slowly exchangeable high affinity sitewith dissociation constant (Kd) 0.021 µM (site B), anotherslowly exchangeable high affinity site with Kd 1.6 µM(site C) and several low affinity (Kd {small tilde}30 µM)sites. The Kd values for sites B and C of the other nucleotidestested were 0.5 µM and 16 µM (GDP), 8 µM and34 µM (CDP), 17 µM and 20 µM (UDP) and 1.4µM and 1.4 µM (PP₁). From a comparison of the observed UV spectral change and theamount of nucleotide bound to these sites, as calculated fromthe above Kd values, we concluded that the nucleotide bindingto site B or G induces UV spectral changes that are almost thesame in shape and magnitude. The estimated difference molarabsorption coefficient () was 3.4?10³M^–1_ADP cm^–1for ADP at 278 nm. Our conclusions were strengthened by thegood agreement between the observed spectra and the calculatedspectra (derived from the Kd and values of ADP and GDP) whenADP and GDP were added together to CF₁. The cause of the unusual behavior of GDP in the UV differencespectrum which was unexplained in our previous report was shownto be competition between the GDP added and previously boundADP at sites B and C; this distorted the real spectrum inducedby GDP. (Received October 3, 1983; Accepted February 13, 1984)     

Ascorbate enhancement of H1 histamine receptor sensitivity coincides with ascorbate oxidation inhibition by histamine receptors

Ascorbate has previously been shown to enhance both ₁- and ₂-adrenergic activity. This activity is mediated by ascorbate binding to the extracellular domain of the adrenergic receptor, which also decreases the oxidation rate of ascorbate. H1 histamine receptors have extracellular agonist or ascorbate binding sites with strong similarities to _1- and ₂-adrenergic receptors. Physiological concentrations of ascorbate (50 µM) significantly enhanced histamine contractions of rabbit aorta on the lower half of the histamine dose-response curve, increasing contractions of 0.1, 0.2, and 0.3 µM histamine by two- to threefold. Increases in ascorbate concentration significantly enhanced 0.2 µM histamine (5–500 µM ascorbate) and 0.3 µM histamine (15–500 µM ascorbate) in a dose-dependent manner. Histamine does not measurably oxidize over 20 h in oxygenated PSS at 37°C. Thus the ascorbate enhancement is independent of ascorbate's antioxidant effects. Ascorbate in solution oxidizes rapidly. Transfected histamine receptor membrane suspension with protein concentration at >3.1 µg/ml (56 nM maximum histamine receptor) decreases the oxidation rate of 392 µM ascorbate, and virtually no ascorbate oxidation occurs at >0.0004 mol histamine receptor/mol ascorbate. Histamine receptor membrane had an initial ascorbate oxidation inhibition rate of 0.094 min·µg protein^–1·ml^–1, compared with rates for transfected ANG II membrane (0.055 min·µg protein^–1·ml^–1), untransfected membrane (0.052 min·µg protein^–1·ml^–1), creatine kinase (0.0082 min·µg protein^–1·ml^–1), keyhole limpet hemocyanin (0.00092 min·µg protein^–1·ml^–1), and osmotically lysed aortic rings (0.00057 min·µg wet weight^–1·ml^–1). Ascorbate enhancement of seven-transmembrane-spanning membrane receptor activity occurs in both adrenergic and histaminergic receptors. These receptors may play a significant role in maintaining extracellular ascorbate in a reduced state. molecular complementarity; vitamin C; seven-transmembrane-spanning membrane receptors     

Changes in Outward K+ Currents in Response to Two Types of Sweeteners in Sweet Taste Transduction of Gerbil Taste Cells

Using the whole cell patch clamp technique, we measured changesin outward K⁺ currents of gerbil taste cells in response todifferent kinds of sweeteners. Outward K⁺ currents of the tastecell induced by depolarizing pulses were suppressed by sweetstimuli such as 10 mM Na-saccharin. The membrane-permeable analogof cAMP, cpt-cAMP, also decreased outward K⁺ currents. On theother hand, the K+ currents were enhanced by amino acid sweetenerssuch as 10 mM D-tryptophan. The outward K⁺ current was enhancedby external application of Ca²⁺-transporting ionophore, 5 µMionomycin, and intracellular application of 5 µM inositol-1,4,5-trisphosphate(IP₃). The outward K⁺ currents were no longer suppressed by10 mM Na-saccharin containing 20 µM gurmarin, but werestill enhanced by 10 mM D-tryptophan containing 20 µMgurmarin. These results suggest that sweet taste transductionfor one group of sweeteners such as Na-saccharin in gerbilsis concerned with an increase of the intracellular cAMP level,and that the transduction for the other group of sweetenerssuch as D-tryptophan is concerned with an increase of the intracellularIP₃ level which releases Ca²⁺ from the internal stores. Chem.Senses 22: 163–169, 1997.     

Oxidative Damages and Repair in Euglena gracilis Exposed to Ozone. I. SH Groups and Lipids

Malondialdehyde, a product of lipid oxidation, increased graduallywhen Euglena gracilis cells were bubbled with 240 µ1.liter^–1ozone (delivery rate of 1µmolO₃.min^–1) for 120 min.Simultaneously, the sulfhydryl group content decreased by 36%during the treatment, which was mainly due to oxidation of proteinsulfhydryl groups. The molar amount of SH groups oxidized was3 times higher than that of fatty acid oxidized, indicatingthat sulfhydryl groups were more accessible or more easily oxidizedby O₃ than fatty acids. When Euglena cells were allowed to recoverunder autotrophic growth conditions following O₃ treatment,viable cells were incapable of dividing during the first 5 hof the recovery period but regenerated SH groups nearly to thecontrol level. The increase of SH content during this periodpreceded the resumption of cell division and the restorationof normal growth. These results suggest that the regenerationof SH groups by Euglena cells is a part of a mechanism involvedin the repair of oxidative damage caused by ozone and is anessential step for the initiation of cell division. (Received July 20, 1987; Accepted December 14, 1987)     

Direct block of cloned hKv1.5 channel by cytochalasins, actin-disrupting agents

The action of cytochalasins, actin-disrupting agents on human Kv1.5 channel (hKv1.5) stably expressed in Ltk^– cells was investigated using the whole cell patch-clamp technique. Cytochalasin B inhibited hKv1.5 currents rapidly and reversibly at +60 mV in a concentration-dependent manner with an IC₅₀ of 4.2 µM. Cytochalasin A, which has a structure very similar to cytochalasin B, inhibited hKv1.5 (IC₅₀ of 1.4 µM at +60 mV). Pretreatment with other actin filament disruptors cytochalasin D and cytochalasin J, and an actin filament stabilizing agent phalloidin had no effect on the cytochalasin B-induced inhibition of hKv1.5 currents. Cytochalasin B accelerated the decay rate of inactivation for the hKv1.5 currents. Cytochalasin B-induced inhibition of the hKv1.5 channels was voltage dependent with a steep increase over the voltage range of the channel's opening. However, the inhibition exhibited voltage independence over the voltage range in which channels are fully activated. Cytochalasin B produced no significant effect on the steady-state activation or inactivation curves. The rate constants for association and dissociation of cytochalasin B were 3.7 µM/s and 7.5 s^–1, respectively. Cytochalasin B produced a use-dependent inhibition of hKv1.5 current that was consistent with the slow recovery from inactivation in the presence of the drug. Cytochalasin B (10 µM) also inhibited an ultrarapid delayed rectifier K⁺ current (I_K,ur) in human atrial myocytes. These results indicate that cytochalasin B primarily blocks activated hKv1.5 channels and endogenous I_K,ur in a cytoskeleton-independent manner as an open-channel blocker. voltage-gated K⁺ channel; heart; open channel block     

Effect of Methanol on the Nucleotide Binding to High-Affinity Sites on Chloroplast Coupling Factor 1

The effects of methanol on the nucleotide binding to isolatedchloroplast coupling factor 1 (CF₁) were investigated. IsolatedCF₁ has four kinds of nucleotide binding sites; a barely dissociableADP-binding site (site A), two slowly exchangeable high-affinitysites with different affinities for ADP (sites B and C) whichare not catalytic sites, and several low-affinity sites (Hisaboriand Sakurai 1984). Methanol at 20% (v/v) slightly acceleratedthe binding of ADP to CF₁ but did not influence the number ofbinding sites. Methanol at 10–24% (v/v) affected neitherthe total amounts of bound adenine nucleotides (2.5 mol/molCF₁) nor the incorporation of labeled ADP from the medium (1.5mol/mol CF₁ into the slowly exchangeable sites (sites A, B,C). These results indicate that no appreciable exchange of ADPoccurred at site A at 10–24% (v/v) methanol and excludethe possibility of direct participation of nucleotide bindingat this site in the regulation of ATPase. In 32% methanol, theamount of the labeled ADP bound increased, suggesting some exchangeat site A. Methanol at 20% (v/v) greatly increased the affinitiesof sites B and C for ADP, CDP, GDP, UDP and PP_i. Conformational change of CF₁ induced by the binding of nucleotidesto site(s) B (and C) increased the resistance of CF₁ to inactivationby methanol at high concentrations or by cold treatment. (Received August 16, 1984; Accepted January 23, 1985)     

Protease-activated receptor regulation of Cl- secretion in Calu-3 cells requires prostaglandin release and CFTR activation

Human lung epithelial (Calu-3) cells were used to investigate the effects of protease-activated receptor (PAR) stimulation on Cl^– secretion. Quantitative RT-PCR (QRT-PCR) showed that Calu-3 cells express PAR-1, -2, and -3 receptor mRNAs, with PAR-2 mRNA in greatest abundance. Addition of either thrombin or the PAR-2 agonist peptide SLIGRL to the basolateral solution of monolayers mounted in Ussing chambers produced a rapid increase in short-circuit current (I_sc: thrombin, 21 ± 2 µA; SLIGRL, 83 ± 22 µA), which returned to baseline within 5 min after stimulation. Pretreatment of monolayers with the cell-permeant Ca²⁺-chelating agent BAPTA-AM (50 µM) abolished the increase in I_sc produced by SLIGRL. When monolayers were treated with the cyclooxygenase inhibitor indomethacin (10 µM), nearly complete inhibition of both the thrombin- and SLIGRL-stimulated I_sc was observed. In addition, basolateral treatment with the PGE₂ receptor antagonist AH-6809 (25 µM) significantly inhibited the effects of SLIGRL on I_sc. QRT-PCR revealed that Calu-3 cells express mRNAs for CFTR, the Ca²⁺-activated KCNN4 K⁺ channel, and the KCNQ1 K⁺ channel subunit, which, in association with KCNE3, is known to be regulated by cAMP. Stimulation with SLIGRL produced an increase in apical Cl^– conductance that was blocked in cells expressing short hairpin RNAs designed to target CFTR. These results support the conclusion that PAR stimulation of Cl^– secretion occurs by an indirect mechanism involving the synthesis and release of prostaglandins. In addition, PAR-stimulated Cl^– secretion requires activation of CFTR and at least two distinct K⁺ channels located in the basolateral membrane. cystic fibrosis transmembrane conductance regulator; KCNQ1; calcium-activated potassium channels; KCNN4; cAMP     

Chloride Transport in Chara: I. KINETICS AND CURRENT-VOLTAGE CURVES FOR A PROBABLE PROTON SYMPORT

Chara cells show an inward positive electric current acrossthe plasmalemma when exposed to Cl^– under voltage-clampconditions. The rapid rise of this current suggests that itis directly associated with the inward transport of Cl^–.The dependence of the current on Cl^– concentration showssaturation, the data fitting the Michaelis-Menten equation withV_m up to 100 nmol m^–2 s^–1 (for Cl^–starvedcells) with K_M 10–20 µM, and with some allowancefor an unstirred layer of water adjacent to the membrane. Theeffects on the current of clamp potential, illumination, withdrawalof alkali metal cations, and addition of amine were also investigated.These results suggest that the mechanism is the symport of 2H⁺ with each Cl^–, and that the actions of light, externalK⁺, and amine in stimulating Cl^–, influx are indirect.     

Pacemaker activity in urethral interstitial cells is not dependent on capacitative calcium entry

The aim of the present study was to investigate the properties and role of capacitative Ca²⁺ entry (CCE) in interstitial cells (IC) isolated from the rabbit urethra. Ca²⁺ entry in IC was larger in cells with depleted intracellular Ca²⁺ stores compared with controls, consistent with influx via a CCE pathway. The nonselective Ca²⁺ entry blockers Gd³⁺ (10 µM), La³⁺ (10 µM), and Ni²⁺ (100 µM) reduced CCE by 67% (n = 14), 65% (n = 11), and 55% (n = 9), respectively. These agents did not inhibit Ca²⁺ entry when stores were not depleted. Conversely, CCE in IC was resistant to SKF-96365 (10 µM), wortmannin (10 µM), and nifedipine (1 µM). Spontaneous transient inward currents were recorded from IC voltage-clamped at –60 mV. These events were not significantly affected by Gd³⁺ (10 µM) or La³⁺ (10 µM) and were only slightly decreased in amplitude by 100 µM Ni²⁺. The results from this study demonstrate that freshly dispersed IC from the rabbit urethra possess a CCE pathway. However, influx via this pathway does not appear to contribute to spontaneous activity in these cells. smooth muscle; patch clamp; spontaneous transient inward currents     

Measurement of the Carbon Dioxide Compensation Point and the Rate of Loss of 14CO2 in the Light and Dark in Some Bryophytes

The CO₂ compensation point at 25 °C and 250 µEinsteinsm^–2 s^–1 wasmeasured for 27 bryo-phyte species, andwas found to be in the range of 45–160 µl CO₂ I^–1air. Under the same conditions Zea mays gave a value of 11 µlI^–1 and Horde um vulgare 76 µI^–1. The rate of loss of photosyntheticallyfixed ¹⁴CO₂ in the light and dark in six bryophytes (three mosses,two leafy liverworts, one thalloid liverwort) was determinedin CO₂-free air and 100% O₂. The rate of ¹⁴CO₂ evolution inthe light was less than that in the dark in CL₂-free air, butin 100% O₂ the rate in the light increased, so that in all butthe leafy liverworts it was greater than that in the dark. Raisingthe temperature tended to increase the rate of 14CO₂ evolutioninto CO₂-free air both in the light and dark, so that the light/dark(L/D) ratio did not greatly vary. The lower rate of loss of¹⁴CO₂ in the light compared tothe dark could be due to partialinhibition of ‘dark respiration’ reactions in thelight, a low rate of glycolate synthesis and oxidation, or partialreassimilation of the ¹⁴CO₂ produced, or a combination of someor all of these factors.     

Defoliation in White Clover: Nodule Metabolism, Nodule Growth and Maintenance, and Nitrogenase Functioning During Growth and Regrowth

In two experiments, the functioning and metabolism of nodulesof white clover, following a defoliation which removed abouthalf the shoot tissue, were compared with those of undefoliatedplants. In one experiment, the specific respiration rates of nodulesfrom undefoliated plants varied between 1160 and 1830 µmolCO₂ g^–1h^–1, of which nodule ‘growth and maintenance’accounted for 22 ± 2 per cent, or 27 ± 3.6 percent, according to method of calculation. Defoliation reducedspecific nodule respiration and nodule ‘growth and maintenance’respiration by 60–70 per cent, and rate of N₂ fixationby a similar proportion. The original rate of nodule metabolismwas re-established after about 5 d of regrowth; during regrowthnodule respiration was quantitatively related to rate of N₂,fixation: 9.1 µmol CO₂ µmol^–1N₂. With the possible exception of nodules examined 24 h after defoliation,the efficiency of energy utilization in nitrogenase functioningin both experiments was the same in defoliated and undefoliatedplants: 2.0±0.1 µmol CO₂ µmol^–1 C₂H₄;similarly, there was no change in the efficiency of nitrogenasefunctioning as rate of N₂ fixation increased with plant growthfrom 1 to 22 µmol N₂ per plant h^–1. Exposure of nodulated white clover root systems to a 10 percent acetylene gas mixture resulted in a sharp peak in rateof ethylene production after 1.5–2.5 min; subsequently,rate of ethylene production declined rapidly before stabilisingafter 0.5–1 h at a rate about 50 per cent of that initiallyobserved. Regression of ‘peak’ rate of ethyleneproduction on rate of N₂ fixation indicated a value of 2.9 µmolC₂H₄ µmol^–1 N₂, for rates of N₂ fixation between1 and 22 µmol N₂ per plant h^–1. The relationshipsbetween nitrogenase respiration, acetylene reduction rates andN₂ fixation rates are discussed. Trifolium repens, white clover, defoliation, nodule respiration, N₂, fixation, nitrogenase     

Nitrogenase Interrelationship with Nodule Ureide and Lipid Components at Anthesis and Seed Maturity of Jack Bean

Nodule lipid, glyoxylate and ureide transformations associatedwith nitrogenase activity of Jack bean (Canavalia ensiformis(L.) DC.) were determined at anthesis, 72 day age from emergence(DAE), and with mature seed pod development at 149 DAE. Totalnodule lipid content decreased about 86% during seed development.Acylglycerides were dominant, 10.9 g kg^–1 nodule, withtriacyl content decreasing from 55% at anthesis to about 13%at full seed set. Phosphatidylcholine composed about 39% oftotal nodule glycerphosphatides, 9.03 g kg^–1 nodule atanthesis, decreasing to 16% at full seed set. Oleic was theprincipal unsaturated fatty acid, 33.0% at anthesis decreasingto 25.8% at full seed set. Nitrogenase activity decreased from617.8 nmol C₂H₄ plant_–1s^–1 at anthesis to 499.5nmol C₂H₄ plant^–1s^–1 at full seed set. Key glyox-ylatecycle enzymes changed significantly with increase of isocitratasefrom 4.59 katal?10^-6 (µkat) kg^–1 to 18.36µkatkg^–1 nodule and decrease of malate synthetase from 32.47µkat kg^–1 to 4.25µkat kg^–1 nodule. Highlysignificant decreases occurred with purine catabolic enzymes;uricase decreased from 39.10µkat kg^–1 to 2.21 µkatkg^–1, allantoinase decreased from 4.08µkat kg^–1to 1.36µkat kg^–1 and allantoicase decreased from3.91 µkat kg^–1 to 0.85 µkat kg^–1 nodule.Ureide content decreased from 204.40 mMol kg^–1 to 33.03mMol kg^–1 nodule. (Received April 4, 1988; Accepted August 17, 1988)     

Effect of Inorganic Phosphate on the Biosynthesis of Purine and Pyrimidine Nucleotides in Suspension-Cultured Cells of Catharanthus roseus

The levels of purine and pyrimidine nucleotides in suspensioncultures of Catharanthus roseus were determined 24 h after stationary-phasecells were transferred to fresh complete (‘+Pi’)or phosphate-deficient (‘–Pi’) Murashige-Skoogmedium. The levels of ATP, GTP, UTP and CTP were from approx.3 to 5-fold greater in the cells grown in ‘+Pi’medium than in the cells grown in ‘–Pi’ medium.The levels of almost all other nucleotides were slightly higherin the cells in ‘+Pi’ medium. The rates of de novoand salvage biosynthesis of purine and pyrimidine nucleotideswere estimated from the rates of incorporation of radioactivityfrom [¹⁴C]formate, [2–¹⁴C]glycine, NaH¹⁴CO₃, [6–¹⁴C]orotate,[8–¹⁴C]adenine, [8–¹⁴C]adenosine, [2–¹⁴C]uraciland [2–¹⁴C]uridine. The results indicated that the activityof both the de novo and the salvage pathway was higher in thecells in ‘+Pi’ medium than in the cells in ‘–Pi’medium. The rate of degradation estimated from the rate of releaseof ¹⁴CO₂ from labelled purines and pyrimidines indicated thatdegradation of uridine was significantly reduced in the cellsin ‘+Pi’ medium, but no significant difference wasfound in the degradation of adenine, adenosine and uracil. Thepossible role of Pi in the control of the biosynthesis of nucleotidesand in the degradation of uridine is discussed. Catharanthus roseus, Madagascar periwinkle, suspension culture, inorganic phosphate, nucleotides, purines, pyrimidines, biosynthesis, degradation     

Enhancement of substrate-gated Cl- currents via rat glutamate transporter EAAT4 by PMA

Glutamate transporters (also called excitatory amino acid transporters, EAAT) are important in extracellular homeostasis of glutamate, a major excitatory neurotransmitter. EAAT4, a neuronally expressed EAAT in cerebellum, has a large portion (95% of the total L-aspartate-induced currents in human EAAT4) of substrate-gated Cl^– currents, a distinct feature of this EAAT. We cloned EAAT4 from rat cerebellum. This molecule was predicted to have eight putative transmembrane domains. L-Glutamate induced an inward current in oocytes expressing this EAAT4 at a holding potential –60 mV. Phorbol 12-myristate 13-acetate (PMA), a protein kinase C (PKC) activator, significantly increased the magnitude of L-glutamate-induced currents but did not affect the apparent affinity of EAAT4 for L-glutamate. This PMA-enhanced current had a reversal potential –17 mV at extracellular Cl^– concentration ([Cl^–]_o) 104 mM with an 60-mV shift per 10-fold change in [Cl^–]_o, properties consistent with Cl^–-selective conductance. However, PMA did not change EAAT4 transport activity as measured by [³H]-L-glutamate. Thus PMA-enhanced Cl^– currents via EAAT4 were not thermodynamically coupled to substrate transport. These PMA-enhanced Cl^– currents were partially blocked by staurosporine, chelerythrine, and calphostin C, the three PKC inhibitors. Ro-31-8425, a PKC inhibitor that inhibits conventional PKC isozymes at low concentrations (nM level), partially inhibited the PMA-enhanced Cl^– currents only at a high concentration (1 µM). Intracellular injection of BAPTA, a Ca²⁺-chelating agent, did not affect the PMA-enhanced Cl^– currents. 4-Phorbol-12,13-didecanoate, an inactive analog of PMA, did not enhance glutamate-induced currents. These data suggest that PKC, possibly isozymes other than conventional ones, modulates the substrate-gated Cl^– currents via rat EAAT4. Our results also suggest that substrate-gated ion channel activity and glutamate transport activity, two EAAT4 properties that could modulate neuronal excitability, can be regulated independently. oocytes; protein kinase C     

Effect of elevated CO2 on the utilization of light energy in Nothofagus fusca and Pinus radiata

Red beech (Nothofagus fusca (Hook. F.) Oerst.; Fagaceae) andradiata pine (Pinus radiata D. Don; Pinaceae) were grown for16 months in large open-top chambers at ambient (37 Pa) andelevated (66 Pa) atmospheric partial pressure of CO₂, and incontrol plots (no chamber). Summer-time measurements showedthat photosynthetic capacity was similar at elevated CO₂ (lightand CO₂-saturated value of 17.2 µmol m^–2 s^–1for beech, 13.5 µmol m^–2 s^–1 for pine), plantsgrown at ambient CO₂ (beech 21.0 µmol^–2 s^–1,pine 14.9 µmol m^–2s^–1) or control plants grownwithout chambers (beech 23.2 µmol m^–2 s^–1,pine 12.9 µmol m^–2 s^–1). However, the higherCO₂ partial pressure had a direct effect on photosynthetic rate,such that under their respective growth conditions, photosynthesisfor the elevated CO₂ treatment (measured at 70 Pa CO₂ partialpressure: beech 14.1 µmol m^–2 s^–1 pine 10.3)was greater than in ambient (measured at 35 Pa CO₂: beech 9.7µmol m^–2 s^–1, pine 7.0 µmol m^–2s^–1) or control plants (beech 10.8 µmol m^–2s^–1, pine 7.2 µmol m^–2 s^–1). Measurementsof chlorophyll fluorescence revealed no evidence of photodamagein any treatment for either species. The quantity of the photoprotectivexanthophyll cycle pigments and their degree of de-epoxidationat midday did not differ among treatments for either species.The photochemical efficiency of photosystem II (yield) was lowerin control plants than in chamber-grown plants, and was higherin chamber plants at ambient than at elevated CO₂. These resultssuggest that at lower (ambient) CO₂ partial pressure, beechplants may have dissipated excess energy by a mechanism thatdoes not involve the xanthophyll cycle pigments. Key words: Carotenoids, chlorophyll fluorescence, photosynthesis, photoinhibition, photoprotection, xanthophyll cycle     

Influence of Elevated CO2 and Temperature on the Photosynthesis and Respiration of White Clover Dependent on N2 Fixation

Single clonal plants of white clover (Trifolium repens L) grownfrom explants in a Perlite rooting medium, and dependent fornitrogen on N₂ fixation in root nodules, were grown for severalweeks in controlled environments which provided two regimesof CO₂, and temperature 23/18 °C day/night temperaturesat 680 µmol mol^–1 CO₂, (C680), and 20/15 °Cday/night temperatures at 340 µmol mol^–1 CO₂ (C340)After 3–4 weeks of growth, when the plants were acclimatedto the environmental regimes, leaf and whole-plant photosynthesisand respiration were measured using conventional infra-red gasanalysis techniques Elevated CO₂ and temperature increased ratesof photosynthesis of young, fully expanded leaves at the growthirradiance by 17–29%, despite decreased stomatal conductancesand transpiration rates Water use efficiency (mol CO₂ mol H₂O^–1)was also significantly increased Plants acclimated to elevatedCO₂, and temperature exhibited rates of leaf photosynthesisvery similar to those of C340 leaves ‘instantaneously’exposed to the C680 regime However, leaves developed in theC680 regime photosynthesised less rapidly than C340 leaves whenboth were exposed to a normal CO₂, and temperature environmentIn measurements where irradiance was varied, the enhancementof photosynthesis in elevated CO₂ at 23 °C increased graduallyfrom approx 10 % at 100 µmol m^–1 s^–1 to >27 % at 1170 µmol m^–2 s^–1 In parallel, wateruse efficiency increased by 20–40 % at 315 µmolm^–2 s^–1 In parallel, water use efficiency increasedby 20–40 % at 315 µmol m^–2 s^–1 In parallel,water use efficiency increased by 20–40 % at 315 µmolm^–2 s^–1 In parallel, water use efficiency increasedby 20–40 % at 315 µmol m^–2 s^–1 to approx100 % at the highest irradiance Elevated CO₂, and temperatureincreased whole-plant photosynthesis by > 40 %, when expressedin terms of shoot surface area or shoot weight No effects ofelevated CO₂ and temperature on rate of tissue respiration,either during growth or measurement, were established for singleleaves or for whole plants Dependence on N₂, fixation in rootnodules appeared to have no detrimental effect on photosyntheticperformance in elevated CO₂, and temperature Trifolium repens, white clover, photosynthesis, respiration, elevated CO₂, elevated temperature, water use efficiency, N₂ fixation     

Characterization of the rat Na+/nucleoside cotransporter 2 and transport of nucleoside-derived drugs using electrophysiological methods

The Na⁺-dependent nucleoside transporter 2 (CNT2) mediates active transport of purine nucleosides and uridine as well as therapeutic nucleoside analogs. We used the two-electrode voltage-clamp technique to investigate rat CNT2 (rCNT2) transport mechanism and study the interaction of nucleoside-derived drugs with the transporter expressed in Xenopus laevis oocytes. The kinetic parameters for sodium, natural nucleosides, and nucleoside derivatives were obtained as a function of membrane potential. For natural substrates, apparent affinity (K_0.5) was in the low micromolar range (12–34) and was voltage independent for hyperpolarizing membrane potentials, whereas maximal current (I_max) was voltage dependent. Uridine and 2'-deoxyuridine analogs modified at the 5-position were substrates of rCNT2. Lack of the 2'-hydroxyl group decreased affinity but increased I_max. Increase in the size and decrease in the electronegativity of the residue at the 5-position affected the interaction with the transporter by decreasing both affinity and I_max. Fludarabine and formycin B were also transported with higher I_max than uridine and moderate affinity (102 ± 10 and 66 ± 6 µM, respectively). Analysis of the pre-steady-state currents revealed a half-maximal activation voltage of about –39 mV and a valence of about –0.8. K_0.5 for Na⁺ was 2.3 mM at –50 mV and decreased at hyperpolarizing membrane potentials. The Hill coefficient was 1 at all voltages. Direct measurements of radiolabeled nucleoside fluxes with the charge associated showed a ratio of two positive inward charges per nucleoside, suggesting a stoichiometry of two Na⁺ per nucleoside. This discrepancy in the number of Na⁺ molecules that bind rCNT2 may indicate a low degree of cooperativity between the Na⁺ binding sites. two-electrode voltage clamp; concentrative nucleoside transport; presteady-state currents     

Rates of Photosynthesis in Characean Cells: II. PHOTOSYNTHETIC 14CO2 FIXATION AND 14C-BICARBONATE UPTAKE BY CHARACEAN CELLS

Photosynthetic ¹⁴C fixation by Characean cells in solutionsof high pH containing NaH¹⁴CO₃ gave a measure of the abilityof these cells to take up bicarbonate (H¹⁴CO₃^–). Whereascells of Nitella translucens from plants collected and thenstored in the laboratory absorbed bicarbonate at 1–1.5µµmoles cm^–2 sec^–1, rates of 3–8µµmoles cm^–2 sec^–1 were obtained withN. translucens cells from plants grown in the laboratory. Influxesof 5–6 µµmoles cm^–2 sec^–1 wereobtained with Chara australis, 3–8 µµmolescm^–2 sec^–1 with Nitellopsis obtusa, and 1–5µµmoles cm^–2 sec^–1 with Tolypella intricata.It is considered that these influxes represent the activityof a bicarbonate pump, which may be an electrogenic process. In solutions of lower pH, H¹⁴CO₃^– uptake would be maskedby rapid diffusion of ¹⁴CO₂ into the cells: the four Characeanspecies fixed ¹⁴CO₂ at maximum rates of 30–40 µµmolescm^–2 sec^–1 (at 21° C).     

Long Term Effects of High CO2 Concentration on Photosynthesis of Water Hyacinth (Eichhornia crassipes (Mart.) Solms)

The photosynthetic response to CO₂ concentration, light intensityand temperature was investigated in water hyacinth plants (Eichhorniacrassipes (Mart.) Solms) grown in summer at ambient CO₂ or at10000 µmol(CO₂) mol^–1 and in winter at 6000 µmol(CO₂)mol^–1 Plants grown and measured at ambient CO₂ had highphotosynthetic rate (35 µmo1(CO₂) m^–2 s^–1),high saturating photon flux density (1500–2000) µmolm^–2 s^–1 and low sensitivity to temperature in therange 20–40 °C. Maximum photosynthetic rate (63 µmol(CO₂)m^–2 s^–1) was reached at an internal CO₂ concentrationof 800 µmol mol^–1. Plants grown at high CO₂ in summerhad photosynthetic capacities at ambient CO₂ which were 15%less than for plants grown at ambient CO₂, but maximum photosyntheticrates were similar. Photosynthesis by plants grown at high CO₂and high light intensity had typical response curves to internalCO₂ concentration with saturation at high CO₂, but for plantsgrown under high CO₂ and low light and plants grown under lowCO₂ and high light intensity photosynthetic rates decreasedsharply at internal CO₂ concentrations above 1000 µmol^–1. Key words: Photosynthesis, CO₂, enrichment, Eichhornia crassipes