Search

1H NMR spectroscopic and pattern recognition-based methods (NMR-PR) were applied to the metabolic profiling studies on hemodialysis (HD). Plasma samples were collected from 37 patients before and after HD and measured by 600 MHz NMR spectroscopy. Each spectrum was data-processed and subjected to principal component analysis for pattern recognition. Spectral patterns of plasma between pre- and post-dialyses were clearly discriminated, together with significant fluctuations in the levels of creatinine, trimethylamine- N-oxide, glucose, lactate, and acetate, which were quantitated. We have first observed the significant elevation of lactate levels in post-dialysis plasma. The present study has demonstrated the high feasibility of NMR-PR method for monitoring the dialysis condition and comprehensive profiling of the change of low-molecular-weight metabolites in HD. [Figure not available: see fulltext.] [ABSTRACT FROM AUTHOR]

ATP and serotonin (5-HT) are neurotransmitters secreted from taste bud receptor (type II) and presynaptic (type III) cells, respectively. Norepinephrine (NE) has also been proposed to be a neurotransmitter or paracrine hormone in taste buds. Yet, to date, the specific stimulus for NE release in taste buds is not well understood, and the identity of the taste cells that secrete NE is not known. Chinese hamster ovary cells were transfected with α1A adrenoceptors and loaded with fura-2 ("biosensors") to detect NE secreted from isolated mouse taste buds and taste cells. Biosensors responded to low concentrations of NE (⩾10 nM) with a reliable fura-2 signal. NE biosensors did not respond to stimulation with KCl or taste compounds. However, we recorded robust responses from NE biosensors when they were positioned against mouse circumvallate taste buds and the taste buds were stimulated with KCl (50 mM) or a mixture of taste compounds (cycloheximide, 10 μM; saccharin, 2 mM; denatonium, 1 mM; SC45647, 100 μM). NE biosensor responses evoked by stimulating taste buds were reversibly blocked by prazosin, an α1A receptor antagonist. Together, these findings indicate that taste bud cells secrete NE when they are stimulated.Weisolated individual taste bud cells to identify the origin of NE release. NE was secreted only from presynaptic (type III) taste cells and not receptor (type II) cells. Stimulus-evoked NE release depended on Ca2+ in the bathing medium. Using dual biosensors (sensitive to 5-HT and NE), we found all presynaptic cells secrete 5-HT and 33% corelease NE with 5-HT. [ABSTRACT FROM AUTHOR]

We compared shoot and needle morphology and photosynthesis in Picea glehnii (Friedr. Schmidt) M.T. Mast. and Picea jezoensis (Sieb. et Zucc.) Carr. trees planted on immature volcanic ash and well-developed brown forest soils to investigate whether species differences in morphological and physiological plasticity affected tree growth on different soil types. Height growth of P. glehnii was reduced by about 10% on volcanic ash compared with forest soil, whereas that of P. jezoensis was reduced by more than 60%. Needle morphology of P. glehnii was unaffected by soil type. In contrast, needles of P. jezoensis trees growing on volcanic ash were shorter, narrower and thicker, with less dry mass and area, than those of trees growing on forest soil, and specific needle area was lower, indicating lower foliar light-interception efficiency. In both species, changes in needle morphology with increasing irradiance were similar in trees growing on both soil types, indicating that plasticity of needle morphology was unaffected by soil type. In both species, shoot mass and shoot silhouette area were lower and needle mass per unit shoot mass was higher in trees growing on volcanic ash than in trees growing on forest soil. Trees of both species had more needles per unit shoot length, lower shoot silhouette to projected needle area ratios and lower shoot silhouette areas per unit shoot mass (SAM) on volcanic ash than on forest soil, indicating lower shoot-level light-interception efficiency. For P. glehnii, the response of shoot morphology to increasing irradiance was similar on both soil types, with the exception of SAM, which showed lower plasticity in trees growing on volcanic ash. In contrast, shoot-level morphological plasticity of P. jezoensis was reduced in trees growing on volcanic ash. Light-saturated maximum photosynthetic rate (Pmax) of P. glehnii was unaffected by soil type, whereas mass-based Pmax of P. jezoensis was lower in trees growing on volcanic ash than in trees growing on forest soil. In P. jezoensis trees growing on forest soil, area-based Pmax increased with increasing irradiance, but this response was not observed in trees growing on volcanic ash. As a result, area-based Pmax at the top of the canopy was 39 to 54% lower in trees growing on volcanic ash than in trees growing on forest soil. Our results indicate that constraints on morphological acclimation to high irradiances may contribute to reduced height growth of P. jezoensis on volcanic ash. [ABSTRACT FROM AUTHOR]

The interaction of drought and elevated carbon dioxide concentration ([CO2]) on carboxylation capacity of Rubisco (Vcmax) and susceptibility to photoinhibition may be an important determinant of plant responses to seasonal fluctuations in precipitation in an anticipated elevated [CO2] environment. Japanese white birch (Betula platyphylla var. japonica) leaves that developed wholly during a period of drought showed an increase in leaf nitrogen and a decrease in leaf carbohydrates that could ameliorate photosynthetic down-regulation, defined as a decrease in Vcmax in response to elevated [CO2]. Photochemical quenching (qP) was decreased by elevated [CO2] but increased by drought when compared at a given intercellular [CO2] (Ci), indicating that elevated [CO2] could increase the risk of photoinhibition, whereas long-term drought could alleviate the risk of photoinhibition. However, only a small variation in qP was measured among seedlings in the various water availability x [CO2] treatment combinations, consistent with the small treatment differences in chronic photoinhibition among the seedlings, as indicated by the ratio of variable to maximum chlorophyll fluorescence after overnight dark-adaptation. Our results suggest that the offsetting responses--reduced Vcmax plus increased Ci at elevated [CO2] and increased Vcmax plus reduced Ci under drought conditions--resulted in a narrow range of susceptibility to photoinhibition at the growth [CO2] in Japanese white birch seedlings grownin variouswater availability x [CO2] treatment combinations. [ABSTRACT FROM AUTHOR]

ATP has been shown to be a taste bud afferent transmitter, but the cells responsible for, and the mechanism of, its release have not been identified. Using CHO cells expressing high-affinity neurotransmitter receptors as biosensors, we show that gustatory stimuli cause receptor cells to secrete ATP through pannexin 1 hemichannels in mouse taste buds. ATP further stimulates other taste cells to release a second transmitter, serotonin. These results provide a mechanism to link intracellular Ca2+ release during taste transduction to secretion of afferent transmitter, ATP, from receptor cells. They also indicate a route for cell-cell communication and signal processing within the taste bud. [ABSTRACT FROM AUTHOR]

A large number of complete microorganism genomes has been sequenced and submitted to the public database and then incorporated into our complete genome database, Genome Information Broker (GIB, ). However, when comparative genomics is carried out, researchers must be aware that there are protein-coding genes not confirmed by homology or motif search and that reliable protein-coding genes are missing. Therefore, we developed a protocol (Gene Trek in Prokaryote Space, GTPS) for finding possible protein-coding genes in bacterial genomes. GTPS assigns a degree of reliability to predicted protein-coding genes. We first systematically applied the protocol to the complete genomes of all 123 bacterial species and strains that were publicly available as of July 2003, and then to those of 183 species and strains available as of September 2004. We found a number of incorrect genes and several new ones in the genome data in question. We also found a way to estimate the total number of orthologous genes in the bacterial world. [ABSTRACT FROM PUBLISHER]

Taste buds are aggregates of 50 -100 cells, only a fraction of which express genes for taste receptors and intracellular signaling proteins. We combined functional calcium imaging with single-cell molecular profiling to demonstrate the existence of two distinct cell types in mouse taste buds. Calcium imaging revealed that isolated taste cells responded with a transient elevation of cytoplasmic Ca2+ to either tastants or depolarization with KCl, but never both. Using single-cell reverse transcription (RT)-PCR, we show that individual taste cells express either phospholipase C β2 (PLCβ2) (an essential taste transduction effector) or synaptosomal-associated protein 25 (SNAP25) (a key component of calcium-triggered transmitter exocytosis). The two functional classes revealed by calcium imaging mapped onto the two gene expression classes determined by single-cell RT-PCR. Specifically, cells responding to tastants expressed PLCβ2, whereas cells responding to KCl depolarization expressed SNAP25. We demonstrate this by two methods: first, through sequential calcium imaging and single-cell RT-PCR; second, by performing calcium imaging on taste buds in slices from transgenic mice in which PLCβ2-expressing taste cells are labeled with green fluorescent protein. To evaluate the significance of the SNAP25-expressing cells, we used RNA amplification from single cells, followed by RT-PCR. We show that SNAP25-positive cells also express typical presynaptic proteins, including a voltage-gated calcium channel (α1A), neural cell adhesion molecule, synapsin-II, and the neurotransmitter-synthesizing enzymes glutamic acid decarboxylase and aromatic amino acid decarboxylase. No synaptic markers were detected in PLCβ2 cells by either amplified RNA profiling or by immunocytochemistry. These data demonstrate the existence of at least two molecularly distinct functional classes of taste cells: receptor cells and synapse-forming cells. [ABSTRACT FROM AUTHOR]

A number of gustatory receptors have been proposed to underlie umami, the taste of L-glutamate, and certain other amino acids and nucleotides. However, the response profiles of these cloned receptors have not been validated against responses recorded from taste receptor cells that are the native detectors of umami taste. We investigated umami taste responses in mouse circumvallate taste buds in an intact slice preparation, using confocal calcium imaging. Approximately 5% of taste cells selectively responded to L-glutamate when it was focally applied to the apical chemosensitive tips of receptor cells. The concentration-response range for L-glutamate fell approximately within the physiologically relevant range for taste behavior in mice, namely 10mM and above. Inosine monophosphate enhanced taste cell responses to L-glutamate, a characteristic feature of umami taste. Using pharmacological agents, ion substitution, and immunostaining, we showed that intracellular pathways downstream of receptor activation involve phospholipase C β2. Each of the above features matches those predicted by studies of cloned and expressed receptors. However, the ligand specificity of each of the proposed umami receptors [taste metabotropic glutamate receptor 4, truncated metabotropic glutamate receptor 1, or taste receptor 1 (T1R1) and T1R3 dimers], taken alone, did not appear to explain the taste responses observed in mouse taste cells. Furthermore, umami responses were still observed in mutant mice lacking T1R3. A full explanation of umami taste transduction may involve novel combinations of the proposed receptors and/or as-yet-undiscovered taste receptors. [ABSTRACT FROM AUTHOR]

Mixed broad-leaved forests are abundantly populated by several kinds of herbivore species in northern Japan. The life of herbivores depends strongly on the quality of food leaves. Leaf quality is changing with increasing atmospheric CO2 and nitrogen deposition. Four seral species of deciduous broad-leaved tree seedlings (alder, birch, oak and maple) were raised in all four combinations of two levels of CO2 and two levels of nutrient, to examine foliar defense traits. To evaluate the relative defense capacity of the four tree species, we used wild silkworms (Erisan; a generalist herbivore) as a bioassay material. Except with alder, the survival rate and longevity (ML50) of the silkworms were least when they were fed with leaves of seedlings raised under high CO2 and infertile soil conditions, and longest on a diet of leaves grown in ambient CO2 and fertile soil, especially in birch and maple. The longevity of Erisan decreased in the order birch, oak, and maple. The longevity of Erisan fed with alder leaves was independent of CO2 levels and was longer on alder seedlings grown in infertile soil conditions. Alder is an actinorhizal plant that can fix atmospheric nitrogen in root nodules formed by the actinomycetes Frankia sp. The activity of symbiotic microbes would have been enhanced by the greater amount of photosynthates received from the host plants at high CO2, improving the food quality for the silkworms. It was concluded that in all but alder, leaf chemical traits, especially C/N ratio, affect the ML50 of Erisan larvae. [ABSTRACT FROM AUTHOR]

Synapses between gustatory receptor cells and primary sensory afferent fibers transmit the output signal from taste buds to the CNS. Several transmitter candidates have been proposed for these synapses, including serotonin (5-HT), glutamate, acetylcholine, ATP, peptides, and others, but, to date, none has been unambiguously identified. We used Chinese hamster ovary cells stably expressing 5-HT2c receptors as biodetectors to monitor 5-HT release from taste buds. When taste buds were depolarized with KCI or stimulated with bitter, sweet, or sour (acid) tastants, serotonin was released. KCI- and acid-induced 5-HT release, but not release attributable to sweet or bitter stimulation, required Ca2+ influx. In contrast, 5-HT release evoked by sweet and bitter stimulation seemed to be triggered by intracellular Ca2+ release. These experiments strongly implicate serotonin as a taste bud neurotransmitter and reveal unexpected transmitter release mechanisms. [ABSTRACT FROM AUTHOR]

Aspergillus oryzae is a fungus used extensively in the fermentation industry. We constructed cDNA microarrays comprising 2,070 highly expressed cDNAs selected from the ∼6,000 non-redundant expressed sequence tags (ESTs) in the A. oryzae EST database (http://www.aist.go.jp/RIODB/ffdb/index.html). Using the cDNA microarrays, we analyzed the gene expression profiles of A. oryzae cells grown under the glucose-rich (AC) and glucose-depleted (AN) liquid culture conditions used during the construction of the EST database. The sets of genes identified by the cDNA microarray as highly expressed under each culture condition agreed well with the highly redundant ESTs obtained under the same conditions. In particular, transcription levels of most catabolic genes of the glycolytic pathway (EMP) and tricarboxylic acid (TCA) cycle were higher under AC than AN conditions, suggesting that A. oryzae uses both EMP and TCA for glucose metabolism under AC conditions. We further studied the expression of genes encoding hydrolytic enzymes and enzymes involved in energy catabolism by using three industrial solid-phase biomass media, including wheat-bran. The wheat-bran culture gave the richest gene expression profile of hydrolytic enzymes and the lowest expression levels of catabolic genes (EMP, TCA) among the three media tested. The low expression levels of catabolic genes in the wheat-bran culture may release catabolite repression, consequently leading to the rich expression profiles of the hydrolytic enzymes. [ABSTRACT FROM AUTHOR]

A mammalian-like sugar moiety was created in glycoprotein by Saccharomyces cerevisiaein combination with bacterial α-mannosidase to produce a more economic enzyme replacement therapy for patients with Fabry disease. We introduced the human α-galactosidase A (α-GalA) gene into an S. cerevisiae mutant that was deficient in the outer chains of N-linked mannan. The recombinant α-GalA contained both neutral (Man8GlcNAc2) and acidic ([Man-P]1–2Man8GlcNAc2) sugar chains. Because an efficient incorporation of α-GalA into lysosomes of human cells requires mannose-6-phosphate (Man-6-P) residues that should be recognized by the specific receptor, we trimmed down the sugar chains of the α-GalA by a newly isolated bacterial α-mannosidase. Treatment of the α-GalA with the α-mannosidase resulted in the exposure of a Man-6-P residue on a nonreduced end of oligosaccharide chains after the removal of phosphodiester-linked nonreduced-end mannose. The treated α-GalA was efficiently incorporated into fibroblasts derived from patients with Fabry disease. The uptake was three to four times higher than that of the nontreated α-GalA and was inhibited by the addition of 5 mM Man-6-P. Incorporated α-GalA was targeted to the lysosome, and hydrolyzed ceramide trihexoside accumulated in the Fabry fibroblasts after 5 days. This method provides an effective and economic therapy for many lysosomal disorders, including Fabry disease. [ABSTRACT FROM PUBLISHER]

We compared effects of ambient (360 vpm) and elevated (720 vpm) carbon dioxide concentration ([CO2]) and high and low nutrient supply rates on stem growth, annual ring structure and tracheid anatomy of Siberian larch (Larix sibirica Ledeb.) seedlings over two growing seasons. Elevated [CO2] had no significant effect on either stem height or diameter growth; however, both stem height and diameter growth were enhanced by the high nutrient supply rate, and these increases were stimulated by elevated [CO2]. Elevated [CO2] tended to increase the width of the annual xylem ring, the number of cells in a radial file spanning the ring, and tracheid lumen diameter, whereas it tended to reduce cell wall thickness, although there were no statistically significant CO2 effects on tracheid anatomy. Changes in tracheid cell morphology seemed to be dependent on changes in shoot elongation rates. [ABSTRACT FROM PUBLISHER]

Photosynthetic acclimation of deciduous broad-leaved tree species was studied along a vertical gradient within the canopy of a multi-species deciduous forest in northern Japan. We investigated variations in (1) local light regime and CO2 concentration ([CO2]), and (2) morphological (area, thickness and area per mass), biochemical (nitrogen and chlorophyll concentrations) and physiological (light-saturated photosynthetic rate) attributes of leaves of seven major species on three occasions (June, August and October). We studied early successional species, alder (Alnus hirsuta (Spach) Rupr.) and birch (Betula platyphylla var. japonica(Miq.) Hara); gap phase species, walnut (Juglans ailanthifoliaCarrière) and ash (Fraxinus mandshurica var. japonica Rupr.); mid-successional species, basswood (Tilia japonica (Miq.) Simonk.) and elm (Ulmus davidiana var. japonica (Rehd.) Nakai); and the late-successional species, maple (Acer monoBunge). All but maple initiated leaf unfolding from the lower part of the crown. The [CO2] within the vertical profile ranged from 320–350 ppm in the upper canopy to 405–560 ppm near the ground. The lowest and highest ambient [CO2] occured during the day and during the night, respectively. This trend was observed consistently during the summer, but not when trees were leafless. Chlorophyll concentration was positively related to maximum photosynthetic rate within, but not among, species. Leaf senescence started from the inner part of the crown in alder and birch, but started either in the outer or top portion of the canopy of ash, basswood and maple. Chlorophyll (Chl) to nitrogen ratio in leaves increased with decreasing photon flux density. However, Chl b concentration in all species remained stable until the beginning of leaf senescence. Maximum photosynthetic rates observed in sun leaves of early successional species, gap phase or mid-successional species, and late successional species were 12.5–14.8 μmol m−2 s−1, 4.1–7.8 μmol m−2 s−1 and 3.1 μmol m−2 s−1, respectively. [ABSTRACT FROM AUTHOR]

Investigates calcium ion signaling in porcine duodenal glands by muscarinic receptor activation. Effects of acetylcholine (ACh) on calcium ion concentration; Membrane currents evoked by ACh; Concentration of messenger RNA expression in duodenal glands; Calcium ion inhibiting effect of thapsigargin.

Dryobalanops aromatica Gaertn. f. is a major tropical canopy species in lowland tropical rain forests in Peninsular Malaysia. Diurnal changes in net photosynthetic rate (A) and stomatal conductance to water vapor (gs) were measured in fully expanded young and old leaves in the uppermost canopy (35 m above ground). Maximum A was 12 and 10 μmol m−2 s−1 in young and old leaves, respectively; however, because of large variation in A among leaves, mean maximum A in young and old leaves was only 6.6 and 5.5 μmol m−2 s−1, respectively. Both gs and A declined in young leaves when Tleaf exceeded 34 °C and leaf-to-air vapor pressure deficit (ΔW) exceeded 0.025, whereas in old leaves, gs and A did not start to decline until Tleaf and ΔW exceeded 36 °C and 0.035, respectively. Under saturating light conditions, A was linearly related to gs. The coefficient of variation (CV) for the difference between the CO2 concentrations of ambient air and the leaf intercellular air space (Ca – Ci) was smaller than the CV for A or gs, suggesting that maximum gs was mainly controlled by mesophyll assimilation (A/Ci). Minimum Ci/Ca ratios were relatively high (0.72–0.73), indicating a small drought-induced stomatal limitation to A and non-conservative water use in the uppermost canopy leaves. [ABSTRACT FROM PUBLISHER]

An electronic structure calculation with full-potential and all-electron ab initio formalism is performed for hypothetical low-temperature simple cubic phases ( and space groups) of 60 molecular crystal, in which every molecule is orientationally freezed. The degeneracies at special k-points, the magnitudes of energy gaps, and the widths of bands have been found to be sensitive to the orientation of the fullerenes in the crystal. Our result suggests that the structure with space group with -22° rotation around [111] directions, which David et al. proposed, is more stable compared to the structure with space group with the same rotation angle. [ABSTRACT FROM AUTHOR]

A recent scanning tunneling microscopy (STM) experiment has revealed that 60-C70 mixture on the (111)-(1×1) substrate shows specific bias-voltage-dependent images which reflect intramolecular structure. In order to understand this observation, we perform a band-structure calculation. In this study, we adopt a simple model assuming a two-dimensional (2D) molecular crystal and a proper adsorption geometry. By comparing the calculated charge distributions of the lowest unoccupied molecular orbital (LUMO)-induced bands and the highest occupied molecular orbital (HOMO)-induced bands with the observed STM images, we find that the shapes of the intramolecular structures which change with the bias voltage strongly reflect the positions of pentagonal and hexagonal rings. [ABSTRACT FROM AUTHOR]