Chronic ozone exposure preferentially modifies root rather than foliar metabolism of date palm (Phoenix dactylifera) saplings.

In their natural environment, date palms are exposed to chronic atmospheric ozone (O3) concentrations from local and remote sources. In order to elucidate the consequences of this exposure, date palm saplings were treated with ambient, 1.5 and 2.0 times ambient O3 for three months in a free-air controlled exposure facility. Chronic O3 exposure reduced carbohydrate contents in leaves and roots, but this effect was much stronger in roots. Still, sucrose contents of both organs were maintained at elevated O3, though at different steady states. Reduced availability of carbohydrate for the Tricarboxylic acid cycle (TCA cycle) may be responsible for the observed reduced foliar contents of several amino acids, whereas malic acid accumulation in the roots indicates a reduced use of TCA cycle intermediates. Carbohydrate deficiency in roots, but not in leaves caused oxidative stress upon chronic O3 exposure, as indicated by enhanced malonedialdehyde, H2O2 and oxidized glutathione contents despite elevated glutathione reductase activity. Reduced levels of phenolics and flavonoids in the roots resulted from decreased production and, therefore, do not indicate oxidative stress compensation by secondary compounds. These results show that roots of date palms are highly susceptible to chronic O3 exposure as a consequence of carbohydrate deficiency.

Assessing social quality of sheltered independent housing: challenges of scale and group mix.

Forecasts for many cities and regions in Europe predict a growing share of 'elderly' people in the overall population. In addition to this general ageing process, the number of very old people is of specific importance for the issue under discussion. This article looks at sheltered independent housing and living. In particular, the article presents the results of a quantitative and qualitative multidisciplinary study of those facilities in the Netherlands. The research provides insight into the effect of physical scale and group mix on the social quality of sheltered independent housing. The results are based on a desk study of 265 projects and a detailed case study of 24 projects. The quantitative desk study provides reviews related to the time and location (and vice versa) and thus develops a picture of the variation in sheltered independent housing complexes for the period 1998-2010. The findings of the qualitative section in general are that ensuring security and belonging is an important function of sheltered independent housing for residents. Regarding the dimension of physical scale, the responses regarding the desired scale are surprising, with equal support for large as well as small scale. Preferences are strongly related to the location in towns or villages, as the scale surrounding the housing. Regarding group mix, the most important finding is a limit of tolerance between groups, particularly tolerance among vital elderly people towards groups of residents with a mental disability or dementia. This limit seems to be reached much sooner than commonly thought, or deployed on the basis of idealistic motives.

Potassium-dependent wood formation in poplar: seasonal aspects and environmental limitations.

Potassium availability and acquisition are pivotal for the generation of biomass and thus wood formation in growing poplar trees. Here, we focus on the role of potassium (K(+)) in wood production, transitions between dormancy and active growth, and limiting environmental conditions. Molecular mechanisms, such as expression and activity of K(+) transporters and channels controlling seasonal changes in wood formation, are discussed.

Guard cell anion channel SLAC1 is regulated by CDPK protein kinases with distinct Ca2+ affinities.

In response to drought stress, the phytohormone abscisic acid (ABA) induces stomatal closure. Thereby the stress hormone activates guard cell anion channels in a calcium-dependent, as well as -independent, manner. Open stomata 1 protein kinase (OST1) and ABI1 protein phosphatase (ABA insensitive 1) represent key components of calcium-independent ABA signaling. Recently, the guard cell anion channel SLAC1 was identified. When expressed heterologously SLAC1 remained electrically silent. Upon coexpression with Ca(2+)-independent OST1, however, SLAC1 anion channels appear activated in an ABI1-dependent manner. Mutants lacking distinct calcium-dependent protein kinases (CPKs) appeared impaired in ABA stimulation of guard cell ion channels, too. To study SLAC1 activation via the calcium-dependent ABA pathway, we studied the SLAC1 response to CPKs in the Xenopus laevis oocyte system. Split YFP-based protein-protein interaction assays, using SLAC1 as the bait, identified guard cell expressed CPK21 and 23 as major interacting partners. Upon coexpression of SLAC1 with CPK21 and 23, anion currents document SLAC1 stimulation by these guard cell protein kinases. Ca(2+)-sensitive activation of SLAC1, however, could be assigned to the CPK21 pathway only because CPK23 turned out to be rather Ca(2+)-insensitive. In line with activation by OST1, CPK activation of the guard cell anion channel was suppressed by ABI1. Thus the CPK and OST1 branch of ABA signal transduction in guard cells seem to converge on the level of SLAC1 under the control of the ABI1/ABA-receptor complex.

VOC emissions of Grey poplar leaves as affected by salt stress and different N sources.

Nitrogen nutrition and salt stress experiments were performed in a greenhouse with hydroponic-cultured, salt-sensitive Grey poplar (Populus x canescens) plants to study the combined influence of different N sources (either 1 mm NO(3) (-) or NH(4)(+)) and salt (up to 75 mm NaCl) on leaf gas exchange, isoprene biosynthesis and VOC emissions. Net assimilation and transpiration proved to be highly sensitive to salt stress and were reduced by approximately 90% at leaf sodium concentrations higher than 1,800 microg Na g dry weight (dw)(-1). In contrast, emissions of isoprene and oxygenated VOC (i.e. acetaldehyde, formaldehyde and acetone) were unaffected. There was no significant effect of combinations of salt stress and N source, and neither NO(3)(-) or NH(4)(+) influenced the salt stress response in the Grey poplar leaves. Also, transcript levels of 1-deoxy-d-xylulose 5-phosphate reductoisomerase (PcDXR) and isoprene synthase (PcISPS) did not respond to the different N sources and only responded slightly to salt application, although isoprene synthase (PcISPS) activity was negatively affected at least in one of two experiments, despite high isoprene emission rates. A significant salt effect was the strong reduction of leaf dimethylallyl diphosphate (DMADP) content, probably due to restricted availability of photosynthates for DMADP biosynthesis. Further consequences of reduced photosynthetic gas exchange and maintaining VOC emissions are a very high C loss, up to 50%, from VOC emissions related to net CO(2) uptake and a strong increase in leaf internal isoprene concentrations, with maximum mean values up to 6.6 microl x l(-1). Why poplar leaves maintain VOC biosynthesis and emission under salt stress conditions, despite impaired photosynthetic CO(2) fixation, is discussed.

Foliar water supply of tall trees: evidence for mucilage-facilitated moisture uptake from the atmosphere and the impact on pressure bomb measurements.

The water supply to leaves of 25 to 60 m tall trees (including high-salinity-tolerant ones) was studied. The filling status of the xylem vessels was determined by xylem sap extraction (using jet-discharge, gravity-discharge, and centrifugation) and by (1)H nuclear magnetic resonance imaging of wood pieces. Simultaneously, pressure bomb experiments were performed along the entire trunk of the trees up to a height of 57 m. Clear-cut evidence was found that the balancing pressure (P(b)) values of leafy twigs were dictated by the ambient relative humidity rather than by height. Refilling of xylem vessels of apical leaves (branches) obviously mainly occurred via moisture uptake from the atmosphere. These findings could be traced back to the hydration and rehydration of mucilage layers on the leaf surfaces and/or of epistomatal mucilage plugs. Xylem vessels also contained mucilage. Mucilage formation was apparently enforced by water stress. The observed mucilage-based foliar water uptake and humidity dependency of the P(b) values are at variance with the cohesion-tension theory and with the hypothesis that P(b) measurements yield information about the relationships between xylem pressure gradients and height.

Regulation of the ABA-sensitive Arabidopsis potassium channel gene GORK in response to water stress.

The phytohormone abscisic acid (ABA) regulates many stress-related processes in plants. In this context ABA mediates the responsiveness of plants to environmental stresses such as drought, cold or salt. In response to water stress, ABA induces stomatal closure by activating Ca2+, K+ and anion channels in guard cells. To understand the signalling pathways that regulate these turgor control elements, we studied the transcriptional control of the K+ release channel gene GORK that is expressed in guard cells, roots and vascular tissue. GORK transcription was up-regulated upon onset of drought, salt stress and cold. The wilting hormone ABA that integrates responses to these stimuli induced GORK expression in seedlings in a time- and concentration-dependent manner and this induction was dependent on extracellular Ca2+. ABA-responsive expression of GORK was impaired in the ABA-insensitive mutants abi1-1 and abi2-1, indicating that these protein phosphatases are regulators of GORK expression. Application of ABA to suspension-cultured cells for 2 min followed by a 4 h chase was sufficient to manifest transcriptional activation of the K+ channel gene. As predicted for a process involved in drought adaptation, only 12-24 h after the release of the stress hormone, GORK mRNA slowly decreased. In contrast to other tissues, GORK expression as well as K+(out) channel activity in guard cells is ABA insensitive, allowing the plant to adjust stomatal movement and water status control separately.

K(+) channel profile and electrical properties of Arabidopsis root hairs.

Ion channels and solute transporters in the plasma membrane of root hairs are proposed to control nutrient uptake, osmoregulation and polar growth. Here we analyzed the molecular components of potassium transport in Arabidopsis root hairs by combining K(+)-selective electrodes, reverse transcription-PCR, and patch-clamp measurements. The two inward rectifiers AKT1 and ATKC1 as well as the outward rectifier GORK dominated the root hair K(+) channel pool. Root hairs of AKT1 and ATKC1 loss-of-function plants completely lack the K(+) uptake channel or exhibited altered properties, respectively. Upon oligochitin-elicitor treatment of root hairs, transient changes in K(+) fluxes and membrane polarization were recorded in wild-type plants, while akt1-1 root hairs showed a reduced amplitude and pronounced delay in the potassium re-uptake process. This indicates that AKT1 and ATKC1 represent essential alpha-subunits of the inward rectifier. Green fluorescent protein (GFP) fluorescence following ballistic bombardment with GORK promoter-GFP constructs as well as analysis of promoter-GUS lines identified this K(+) outward rectifier as a novel ion channel expressed in root hairs. Based on the expression profile and the electrical properties of the root hair plasma membrane we conclude that AKT1-, ATKC- and GORK-mediated potassium transport is essential for osmoregulation and repolarization of the membrane potential in response to elicitors.

VFK1, a Vicia faba K(+) channel involved in phloem unloading.

In search of a K(+) channel involved in phloem transport we screened a Vicia faba cotyledon cDNA library taking advantage of a set of degenerated primers, flanking regions conserved among K(+) uptake channels. We cloned VFK1 (for Vicia faba K(+) channel 1) characterised by a structure known from the Shaker family of plant K(+) channels. When co-expressed with a KAT1 mutant in Xenopus oocytes, heteromers revealed the biophysical properties of a K(+) selective, proton-blocked channel. Northern blot analyses showed high levels of expression in cotyledons, flowers, stem and leaves. Using in situ PCR techniques we could localise the K(+) channel mRNA in the phloem. In the stem VFK1 expression levels were higher in the lower internodes. There channel transcripts increased in the light and thus under conditions of increased photosynthate allocation. VFK1 transcripts are elevated in sink leaves, and rise in source leaves during the experimental transition into sinks. Fructose- rather than sucrose- or glucose-feeding via the petiole induced VFK1 gene activity. We therefore monitored the fructose sensitivity of the sieve tube potential through cut aphid stylets. In response to an 1 h fructose treatment the sieve tube potential shift increased from 19 mV to 53 mV per 10-fold change in K(+) concentration. Under these conditions K(+) channels dominated the electrical properties of the plasma membrane. Based on the phloem localisation and expression patterns of VFK1 we conclude that this K(+) channel is involved in sugar unloading and K(+) retrieval.

KAT1 is not essential for stomatal opening.

It is generally accepted that K(+) uptake into guard cells via inward-rectifying K(+) channels is required for stomatal opening. To test whether the guard cell K(+) channel KAT1 is essential for stomatal opening, a knockout mutant, KAT1En-1, was isolated from an En-1 mutagenized Arabidopsis thaliana population. Stomatal action and K(+) uptake, however, were not impaired in KAT1-deficient plants. Reverse transcription-PCR experiments with isolated guard cell protoplasts showed that in addition to KAT1, the K(+) channels AKT1, AKT2/3, AtKC1, and KAT2 were expressed in this cell type. In impalement measurements, intact guard cells exhibited inward-rectifying K(+) currents across the plasma membrane of both wild-type and KAT1En-1 plants. This study demonstrates that multiple K(+) channel transcripts exist in guard cells and that KAT1 is not essential for stomatal action.

GORK, a delayed outward rectifier expressed in guard cells of Arabidopsis thaliana, is a K(+)-selective, K(+)-sensing ion channel.

Here we report on the molecular identification, guard cell expression and functional characterization of AtGORK, an Arabidopsis thaliana guard cell outward rectifying K(+) channel. GORK represents a new member of the plant Shaker K(+) channel superfamily. When heterologously expressed in Xenopus oocytes the gene product of GORK mediated depolarization-activated K(+) currents. In agreement with the delayed outward rectifier in intact guard cells and protoplasts thereof, GORK is activated in a voltage- and potassium-dependent manner. Furthermore, the single channel conductance and regulation of GORK in response to pH changes resembles the biophysical properties of the guard cell delayed outward rectifier. Thus GORK very likely represents the molecular entity for depolarization-induced potassium release from guard cells.

KDC1, a novel carrot root hair K+ channel. Cloning, characterization, and expression in mammalian cells.

Potassium is an essential nutrient which plays an important role in many aspects of plant growth and development. Plants have developed a number of highly specific mechanisms to take up potassium from the soil; these include the expression of K(+) transporters and potassium channels in root cells. Despite the fact that root epidermal and hair cells are in direct contact with the soil, the role of these tissues in K(+) uptake is not well understood. Here we report the molecular cloning and functional characterization of a novel potassium channel KDC1 which forms part of a new subfamily of plant K(in) channels. Kdc1 was isolated from carrot root RNA and in situ hybridization experiments show Kdc1 to be highly expressed in root hair cells. Expressing the KDC1 protein in Chinese hamster ovary cells identified it as a voltage and pH-dependent inwardly rectifying potassium channel. An electrophysiological analysis of carrot root hair protoplasts confirmed the biophysical properties of the Kdc1 gene product (KDC1) in the heterologous expression system. KDC1 thus represents a major K(+) uptake channel in carrot root hair cells.

Developmental and light-dependent regulation of a phloem-localised K+ channel of Arabidopsis thaliana.

K+ channels in plants can currently be classified into six families with individual members being involved in nutrient uptake, loading of the xylem and the physiology of stomatal movement. In this study we have focused on akt2/3. This K+ channel, as shown by GUS-expression analysis, is expressed in the phloem and xylem of the aerial parts of Arabidopsis thaliana. Northern blot analyses revealed the highest akt2/3-concentrations in the flower stalk, followed by the leaf, flower and stem. During the light period (8 am to 4 pm), transcripts reached a peak around noon (11 am), decayed to almost 50% in the afternoon and reached a low background level in the following dark period. In continuous darkness, however, the K+ channel mRNA content had already decreased beyond the background level by noon. In leaves and flower stalk, the light-induced transcription of akt2/3 was suppressed by CO2-free air, indicating that gene activity is under the control of photosynthates. Additionally, when rosette leaves were illuminated and flower stalks shaded, akt2/3-mRNA transcription was still inhibited in the shaded region. This indicates that channel gene activation is sensitive to photosynthesis-derived factors from neighboring cells rather than factors mobile in the phloem. We propose that the coupling between sugar production and allocation involves the photosynthate- and light-dependent phloem K+ channel AKT2/3.

AKT3, a phloem-localized K+ channel, is blocked by protons.

The potassium-channel gene, AKT3, has recently been isolated from an Arabidopsis thaliana cDNA library. By using the whole-mount and in situ hybridization techniques, we found AKT3 predominantly expressed in the phloem. To study the physiological role of this channel type, AKT3 was heterologously expressed in Xenopus oocytes, and the electrical properties were examined with voltage-clamp techniques. Unlike the plant inward-rectifying guard cell K+ channels KAT1 and KST1, the AKT3 channels were only weakly regulated by the membrane potential. Furthermore, AKT3 was blocked by physiological concentrations of external Ca2+ and showed an inverted pH regulation. Extracellular acidification decreased the macroscopic AKT3 currents by reducing the single-channel conductance. Because assimilate transport in the vascular tissue coincides with both H+ and K+ fluxes, AKT3 K+ channels may be involved in K+ transport accompanying phloem loading and unloading processes.

Measurement of carbamazepine and its main biotransformation products in plasma by HPLC.

We present a method that permits the simultaneous analysis of carbamazepine (CBZ) and its major biotransformation products, carbamazepine-10,11-epoxide (CBZ-E) and carbamazepine-10,11-dihydroxide (CBZ-diOH), in plasma samples. The method consists of plasma extraction in alkaline medium with NaCl added using chloroform-ethyl acetate (1:1, v/v) and later purification with n-hexane. The samples were submitted to reversed-phase chromatography (RP-18) using acetonitrile-water (3:7, v/v) as the mobile phase and detection at 220 nm. Recoveries of 62.0, 99.9, and 105.4% were obtained for CBZ-diOH, CBZ-E, and CBZ, respectively, with sensitivities of 0.32 micrograms/mL for CBZ-E and CBZ-dOH and of 0.64 micrograms/mL for CBZ. The method proved to be specific, thus permitting measurements in situations of drug combinations.