The Transport and Metabolism of Urea in Chara australis: III. TWO SPECIFIC TRANSPORT SYSTEMS

Reciprocal competitive inhibition studies were used to showthat N-methyl-urea (NMU), acetamide and urea all compete forbinding to a common transport system, designated system I andthat this system is one of two specific mechanisms transportingurea in Chara. System I binds urea with a Km of about 0–3mmol m-3 and is strongly influenced by metabolic controls. SystemI is active and electrogenic and may be energized by the couplingof urea uptake to an influx of protons. This is the first reportof an electrogenic urea transport system in an alga. The secondspecific mechanism for urea transport, designated system II,binds urea with a relatively low affinity (K_m c. 7–0 mmolm-3) and does not transport NMU to a significant extent. SystemII is less subject to metabolic control than system I and, thoughit may be active, is not electrogenic. Key words: Urea, methylurea, proton cotransport, metabolic control     

K+ current inhibition by amphiphilic fatty acid metabolites in rat ventricular myocytes

Fatty acid metabolites accumulate in the heart underpathophysiological conditions that affect -oxidation and can elicit marked electrophysiological changes that are arrhythmogenic. The purpose of the present study was to determine the impact of amphiphilic fatty acid metabolites on K⁺currents that control cardiac refractoriness and excitability. Transient outward(I_to) andinward rectifier(I_K1)K⁺ currents were recorded by thewhole cell voltage-clamp technique in rat ventricular myocytes, and theeffects of two major fatty acid metabolites were examined:palmitoylcarnitine and palmitoyl-coenzyme A (palmitoyl-CoA).Palmitoylcarnitine (0.5-10 µM) caused a concentration-dependent decrease in I_todensity in myocytes internally dialyzed with the amphiphile; 10 µMreduced mean I_todensity at +60 mV by 62% compared with control(P < 0.05). In contrast, externalpalmitoylcarnitine at the same concentrations had no effect, nor didinternal dialysis significantly alterI_K1. Dialysiswith palmitoyl-CoA (1-10 µM) produced a smaller decrease inI_to densitycompared with that produced by palmitoylcarnitine; 10 µM reduced meanI_to density at+60 mV by 37% compared with control(P < 0.05). Both metabolites delayedrecovery of I_tofrom inactivation but did not affect voltage-dependent properties.Moreover, the effects of palmitoylcarnitine were relatively specific,as neither palmitate (10 µM) nor carnitine (10 µM) alone significantly influencedI_to when added tothe pipette solution. These data therefore suggest that amphiphilicfatty acid metabolites downregulateI_to channels by amechanism confined to the cytoplasmic side of the membrane. Thisdecrease in cardiac K⁺ channelactivity may delay repolarization under pathophysiological conditionsin which amphiphile accumulation is postulated to occur, such asdiabetes mellitus or myocardial infarction.

     

Borate absorption in excised sugarcane leaves

Borate absorption in sugarcane consists of a rapid and reversibleinflux into the mesophyll cells of the leaf which is completedwithin 20 rains. (Phase I), followed by a slower and irreversibleaccumulatory phase (II). Phase II uptake represents the summationof 3 absorption mechanisms, each dependent upon the externalconcentration. Highly specific mechanisms 1 and 2 transportborate across the initial barrier into the cells, reaction 3carries the borate across the vacuolar membrane. Calcium isshown to be essential for maximum rates of borate absorption.All 3 reactions are inhibited by OH^– through a combinationof competitive inhibition and irreversible disruption of cellularfunction or structure. Temperature changes over the range of10–40 profoundly affect V_maz and K_m1, but have no effecton K_m2 and K_m3. Reactions 1 and 2 are unaffected by 50 mtl Cl^–,SO^–– or H2PO4^–, whereas each of these anionscompetes with H2BO3^– for site 3. Specific metabolic inhibitorswere used to delineate a linkage of mechanisms 1 and 2 to respiratoryelectron transport. Mechanism 3 is coupled to oxidative phosphorylation. ¹Published with the approval of the Director of the Hawaii AgriculturalExperiment Station as Technical Paper No. 954.     

Hormones in Plants Bearing Nitrogen-fixing Root Nodules: Metabolism of [8-14C]Zeatin in Root Nodules of Alnus glutinosa (L.) Gaertn

The metabolism of [8-¹⁴C]zeatin, supplied via micropipettesover a 24 h period to root nodules of Alnus gliutinosa (L.)Gaertn., was investigated. The major metabolites were tentativelyidentified by means of chromatographic, chemical, and enzymictreatments as adenine, adenosine, trans-zeatin riboside, dihydrozeatin,trans-zeatin-O-ß-D-glucoside, and the O-ß-D-glucosideof dihydrozeatin. In addition, a prominent water-soluble peakof radioactivity was present. This did not appear to be a ribosidebut was biologically active in the soybean callus test. The number and nature of the metabolites formed in the noduleswas similar in both dormant and non-dormant plants.     

EFFECT OF BODY SIZE ON THE RESPIRATION OF CERITHIDEA (CERITHIDEOPSILLA) CINGULATA (GMELIN, 1790) AND CERITHIUM CORALIUM KIENER, 1841

The effect of body size on the oxygen consumption and metabolicrate of Cerithidea (Cerithideopsilla) cingulata (Gmelin, 1790)andCerithium coralium Kiener, 1841 was studied at a constant temperatureof 25°C. An exponential relationship has been observed inboth animals. Oxygen consumption showed a positive linear correlationwith a ‘b’ value of 0.6518 in C. cingulata and 0.7667in C. coralium. A negative linear correlation was obtained forthe metabolic rate with a (b-1) value of –0.3482 in C.cingulata and –0.2333 in C. coralium.     

Mechanisms of Thermoregulation in Flying Bees

SYNOPSIS. Thermoregulation of elevated thorax temperatures isnecessary for bees to achieve the high rates of power productionrequired for flight, and is a key factor allowing them to occupywidely varying thermal environments. However, the mechanismsby which bees thermoregulate during flight are poorly understood.Thermoregulation is accomplished by balancing heat gain andheat loss via the following routes: convection, evaporation,and metabolic heat production. There appears to be a diversityof thermoregulatory mechanisms employed during flight amongbee species. Some species, particularly Bombus spp., activelyincrease the distribution of thoracic heat to the abdomen duringflight as air temperature (T_a) rises, and apparently thermoregulateby varying convective heat loss. However, thermal variationin convection has not been directly measured for any free-flyingbee. Above 33°C, flying Apis mellifera greatly increaseevaporative heat loss with T_a, and many other species "tongue-lash"during flight at high T_as or when artificially heated. Thus,evaporation seems to be important for preventing overheatingduring flight at very high T_as. Flying A. mellifera and Centrispallida strongly decrease metabolic rate as T_a increases, suggestingthat they are varying metabolic heat production for thermoregulationand not aerodynamic requirements. Variation in metabolic heatproduction appears to be mediated by changes in wingbeat kinematics,since wingbeat frequency decreases with T_a for A. melliferaand Centris spp. It is unknown if the decrease in flight metabolicrate at higher T_as occurs secondarily as a consequence of greaterefficiency or if it is truly an active response.     

A carrier-mediated mechanism for pyridoxine uptake by human intestinal epithelial Caco-2 cells: regulation by a PKA-mediated pathway

Vitamin B₆ is essential for cellular functions and growth due to its involvement in important metabolic reactions. Humans and other mammals cannot synthesize vitamin B₆ and thus must obtain this micronutrient from exogenous sources via intestinal absorption. The intestine, therefore, plays a central role in maintaining and regulating normal vitamin B₆ homeostasis. Due to the water-soluble nature of vitamin B₆ and the demonstration that transport of other water-soluble vitamins in intestinal epithelial cells involves specialized carrier-mediated mechanisms, we hypothesized that transport of vitamin B₆ in these cells is also carrier mediated in nature. To test this hypothesis, we examined pyridoxine transport in a model system for human enterocytes, the human-derived intestinal epithelial Caco-2 cells. The results showed pyridoxine uptake to be 1) linear with time for up to 10 min of incubation and to occur with minimal metabolic alteration in the transported substrate, 2) temperature and energy dependent but Na⁺ independent, 3) pH dependent with higher uptake at acidic compared with alkaline pHs, 4) saturable as a function of concentration (at buffer pH 5.5 but not 7.4) with an apparent Michaelis-Menten constant (K_m) of 11.99 ± 1.41 µM and a maximal velocity (V_max) of 67.63 ± 3.87 pmol · mg protein^-1 · 3 min^-1, 5) inhibited by pyridoxine structural analogs (at buffer pH 5.5 but not 7.4) but not by unrelated compounds, and 6) inhibited in a competitive manner by amiloride with an apparent inhibitor constant (K_i) of 0.39 mM. We also examined the possible regulation of pyridoxine uptake by specific intracellular regulatory pathways. The results showed that whereas modulators of PKC, Ca⁺²/calmodulin (CaM), and nitric oxide (NO)-mediated pathways had no effect on pyridoxine uptake, modulators of PKA-mediated pathway were found to cause significant reduction in pyridoxine uptake. This reduction was mediated via a significant inhibition in the V_max, but not the apparent K_m, of the pyridoxine uptake process. These results demonstrate, for the first time, the involvement of a specialized carrier-mediated mechanism for pyridoxine uptake by intestinal epithelial cells. This system is pH dependent and amiloride sensitive and appears to be under the regulation of an intracellular PKA-mediated pathway. vitamin B₆; intestinal transport; transport regulation; Caco-2 cell     

Relationship between Flower Induction by Benzoic Acid and its Metabolism in Lemna paucicostata 151

The flower-inducing activities in Lemna paucicostata 151 offour major metabolites of benzoic acid (N-benzoyl aspartate,benzyl 6-O-ß-D-apiofuranosyl-O-ß-D-glucopyranoside,O-benzoyl isocitrate and O-benzoyl malate) were measured, andthe effects on the uptake and metabolism of benzoic acid dueto change in the level of the benzoic acid concentration orto the addition of plant hormones were investigated. N-Benzoylaspartate had weak activity, and O-benzoyl isocitrate and malatehad fairly strong activities, while benzyl 6-O-ß-D-apiofuranosyl-ß-D-glucopyranosideshowed no activity. As the concentration of benzoic acid rose,the ratio of N-benzoyl aspartate increased and that of benzyl6-O-ß-D-apiofuranosyl-O-ß-D-glucopyranosidedecreased. GA₃ and IAA, inhibitors of flower induction by benzoicacid, seemed to promote conversion to N-benzoyl aspartate insteadof to benzyl 6-O-ß-D-apiofuranosyl-ß-D-glucopyranoside.The conversion to N-benzoyl aspartate was considered to be adetoxification process and that to benzyl 6-O-ß-D-apiofuranosyl-ß-D-glucopyranosidemay be directly related to flower induction in Lemna. (Received November 2, 1987; Accepted January 23, 1988)     

Kinetics of creatine kinase in an experimental model of low phosphocreatine and ATP in the normoxic heart

To study the dependence of the forward flux of creatine kinase(CK) on its substrates and products we designed an acute normoxic modelof steady-state depletion of phosphocreatine (PCr) and adenylate in theisovolumic acetate-perfused rat heart. Various concentrations of PCrand ATP were induced by prior perfusion with 2 deoxy-D-glucose in the presenceof insulin. The apparent rate constant(k_f) and theforward CK flux were measured under metabolic and contractile steadystate by progressive saturation-transfer³¹P nuclear magnetic resonance(NMR). At high adenylate content CK flux was constant for a twofoldreduction in PCr concentration ([PCr]); CK flux was 6.3 ± 0.6 mM/s (vs. 6.5 ± 0.2 mM/s in control) because of adoubling of k_f.Although, at the lowest ATP concentration and [PCr], CKflux was reduced by 50%, it nevertheless always remained higher thanATP synthesis estimated by parallel oxygen consumption measurement.NMR-measured flux was compared with the flux computed under thehypothesis of CK equilibrium. CK flux could not be fully predicted bythe concentrations of CK metabolites. This is discussed in terms ofmetabolite and CK isozyme compartmentation.

     

Possible physiological mechanisms for production of hydrogen peroxide by the ichthyotoxic flagellate Heterosigma akashiwo

Blooms of the toxic red tide phytoplankton Heterosigma akashiwo(Raphidophyceae) are responsible for substantial losses withinthe aquaculture industry. The toxicological mechanisms of H.akashiwoblooms are complex and to date, heavily debated. One putativetype of ichthyotoxin includes the production of reactive oxygenspecies (ROS) that could alter gill structure and function,resulting in asphyxiation. In this study, we investigated thepotential of H.akashiwo to produce extracellular hydrogen peroxide,and have investigated which cellular processes are responsiblefor this production. Within all experiments, H.akashiwo producedsubstantial amounts of hydrogen peroxide (up to 7.6 pmol min^–110⁴ cells^–1), resulting in extracellular concentrationsof ~0.5 µmol l^–1 H₂O₂. Measured rates of hydrogenperoxide production were directly proportional to cell density,but at higher cell densities, accuracy of H₂O₂ detection wasreduced. Whereas light intensity did not alter H₂O₂ production,rates of production were stimulated when temperature was elevated.Hydrogen peroxide production was not only dependent on growthphase, but also was regulated by the availability of iron inthe medium. Reduction of total iron to 1 nmol l^–1 enhancedthe production of H₂O₂ relative to iron replete conditions (10µmol l^–1 iron). From this, we collectively concludethat production of extracellular H₂O₂ by H.akashiwo occurs througha metabolic pathway that is not directly linked to photosynthesis.     

Cytokinins in Germinating Seeds of Phaseolus vulgaris L. III. Transport and Metabolism of 8[14C]t-Zeatin Applied to the Radicle

The application of 8[¹⁴C]t-zeatin to the cotyledons of germinatingbean seeds demonstrated that cytokinins are not readily exportedfrom the cotyledons to the embryonic axis during the early stagesof this process. In the cotyledons the applied zeatin is metabolizedextensively to metabolites which are polar and which occur atR_F 0·2–0·5 on paper chromatograms. Thesemetabolites are stable and are not readily exported from thecotyledons. In contrast the metabolites found at R_F 0–0·2are more readily exported. When exported to the radicles andplumules a large proportion of the translocated metaboliteswere converted to compounds which on paper co-chromatographedwith zeatin. This seems to suggest that the embryonic axis hasthe capacity to synthesize cytokinins and that some of the metabolitesformed during its catabolism can also be used for its synthesis. Phaseolus vulgaris, bean, germination, cytokinins, transport, cotyledons     

Junction Complexes between Sieve Tubes in the Secondary Phloem of Myrtaceae

Junction complexes of unusual structure form between neighbouringsieve tubes in the secondary phloem of Eucalyptus species. Thick-walledribs support thin-walled ‘sieve areas’. In longitudinalsections the structures have a ‘concertina’- likeappearance. They are relatively large, up to 0.2 mm in length.Electron micrographs confirmed that the structures consistedof thin-walled areas perforated with pores, supported by muchthicker ribs. The structures provide a vast surface area fortransfer of metabolites between sieve tubes compared with thatof lateral wall sieve areas of other plants. Hydrolysis of parenchymacell walls occurs during the development of the junction complexes.The structures are only found when sieve tubes are in closeproximity and it is the redifferentiation and partitioning ofintervening parenchyma cells which result in junction complexformation. A survey for the presence of the structures in thephloem of other genera in the family Myrtaceae was made andthey were found in Tristania and Angophora but were not observedin Acmena and Metrosideros. Eucalyptus, sieve tubes, lateral walls, ultrastructure     

Respiratory rates in the cladoceran Ceriodaphnia dubia in Lake Rotongaio, a monomictic lake

The experimentally measured oxygen consumption rate by the cladoceran,Ceriodaphnia dubia, showed a linear increase between 5 and 20°C.Oxygen consumption rates of C. dubia were estimated in situfrom respiratory electron transport system (ETS) activity inLake Rotongaio during summer stratification and winter mixing.Oxygen consumption was 0.002 µl O₂ animal^–1 h^–1in the hypolimnion and 0.076 µl O₂ animal^–1 h^–1in the epilimnion during stratification. Implications of respiredoxygen for metabolic carbon requirements are discussed.     

Are antarctic zooplankton metabolically more cold-adapted than arctic zooplankton? An intra-generic comparison of oxygen consumption rates

Because of the greater age of the antarctic ecosystem in relationto the arctic ecosystem, an elevated metabolic rate and moreobligate stenothermy of antarctic zooplankton compared withtheir arctic counterparts are predicted from the concept ofmetabolic cold adaptation. When oxygen consumption rates werestandardized to 1 mg body nitrogen and 0°C and comparedbetween the species belonging to the same genus, the antarcticrates were greater than arctic rates in Clione (pteropods) andCalanus (copepods), while the reverse was true in Parathemisto(amphipods), Thysanoessa (euphausiids) and Sagitta (chaetognaths).No significant differences were seen in Limacina (pteropods)and Metridia (copepods). These inconsistent results do not supportthe hypothesis of metabolic cold adaptation in the zooplanktonof polar seas.     

Effect of Salt Stress on Viability, Germination and Endogenous Levels of Some Metabolites and Ions in Maize (Zea mays L.) Pollen

Maize plants, subjected to 0, 80, 120 and 160 meq l^–1salinity using NaCl, showed adverse effects on viability, germinationand tube growth of pollen, besides enhancing the bursting ofpollen. The endogenous levels of various metabolites in pollenwere also affected. Pollen grains from salinized plants hadmore soluble carbohydrates, free amino acids, especially proline,phenols and DNA and less starch, protein and RNA compared tothe non-saline controls. Salinity also resulted in the accumulationof ions such as Na⁺, K⁺ and Cl^– while it caused a reductionin the boron content of pollen. These metabolic disturbancespossibly lead to decreased viability, germination and tube growthof pollen thereby resulting into a reduction in reproductivecapacity of the plants under salt stress. Zea mays L., maize, pollen, viability, germination, salt stress     

Murine renal organic anion transporters mOAT1 and mOAT3 facilitate the transport of neuroactive tryptophan metabolites

Tryptophan metabolites such as kynurenate (KYNA), xanthurenate (XA), and quinolinate are considered to have an important impact on many physiological processes, especially brain function. Many of these metabolites are secreted with the urine. Because organic anion transporters (OATs) facilitate the renal secretion of weak organic acids, we investigated whether the secretion of bioactive tryptophan metabolites is mediated by OAT1 and OAT3, two prominent members of the OAT family. Immunohistochemical analyses of the mouse kidneys revealed the expression of OAT1 to be restricted to the proximal convoluted tubule (representing S1 and S2 segments), whereas OAT3 was detected in almost all parts of the nephron, including macula densa cells. In the mouse brain, OAT1 was found to be expressed in neurons of the cortex cerebri and hippocampus as well as in the ependymal cell layer of the choroid plexus. Six tryptophan metabolites, including the bioactive substances KYNA, XA, and the serotonin metabolite 5-hydroxyindol acetate inhibited [³H]p-aminohippurate (PAH) or 6-carboxyfluorescein (6-CF) uptake by 50–85%, demonstrating that these compounds interact with OAT1 as well as with OAT3. Half-maximal inhibition of mOAT1 occurred at 34 µM KYNA and 15 µM XA, and it occurred at 8 µM KYNA and 11.5 µM XA for mOAT3. Quinolinate showed a slight but significant inhibition of [³H]PAH uptake by mOAT1 and no alteration of 6-CF uptake by mOAT3. [¹⁴C]-Glutarate (GA) uptake was examined for both transporters and demonstrated differences in the transport rate for this substrate by a factor of 4. Trans-stimulation experiments with GA revealed that KYNA and XA are substrates for mOAT1. Our results support the idea that OAT1 and OAT3 are involved in the secretion of bioactive tryptophan metabolites from the body. Consequently, they are crucial for the regulation of central nervous system tryptophan metabolite concentration. kidneys; brain; macula densa; transforming growth factor; N-methyl-D-aspartate receptor     

Silica Deposition in the Grass Leaf in Relation to Transpiration and the Effect of Dinitrophenol

Characteristic opal phytolith (‘silica body’) formationwas demonstrated in detached leaves of Sieglingia decumbens(Heath Grass), cultured in 100 ppm dissolved silicon (silicondioxide), previously, the leaves were free from intracellulardeposits as a result of silica-minimal tiller growth. The formertechnique allowed the study of the leaf deposition processesindependently of apical and root tissues, under growth-cabinetand glasshouse conditions. Deposition in excised leaves wascompletely suppressed by a surface, monomolecular coating, thusindicating that total net water loss was a limiting factor,however, evaporation from the recipient, epidermal tissues perse was not a requirement for this in situ deposition Generally,apart from an apparent, cell site shift in one treatment, phytolith-formationwas unaffected by the presence of the metabolic inhibitor 2,4-dinitrophenol. Also, some evidence was obtained of the influxof germanium dioxide into epidermal ldioblasts, which indicateda non-specificity of the host cell for silica. These results and those of earlier studies suggest that passive,non-metabolic mechanisms could account for the transport, influx,and cell lumen polymerization of silica in the grass leaf.     

Chemical Defence in Cephalaspidean Gastropods: Origin, Anatomical Location and Ecological Roles

Within the opisthobranchs, the cephalaspideans are traditionallyconsidered a transitional group between typical testacean prosobranchsand shell-less opisthobranchs. The cephalaspidean anatomy, includingthe presence of a cephalic shield, is related to burrowing throughsoft sediment. Recent studies have shown that some herbivorousand carnivorous cephalaspideans contain secondary metabolites.The micro-herbivorous bubble snails of the Bullidae and Haminoeidaefamilies are known to have secondary metabolites which have differentecological roles. The polypropionates isolated from Bulla gouldianaand B. striata were deterrent to fishes while the secondarymetabolites of Haminoea callidegenita, H. fusari, H. hydatis,H. navicula, H. orbignyana and H. orteai were alarm pheromonesemployed during cross copulation. In Bulla gouldiana and B.striata, the defensive secretion was located mainly in a whitegland along the margin of the mantle. In Haminoea species, alarmpheromones were located in external parts (cephalic shield,parapodial lobes and posterior pallial lobe). The carnivorous cephalaspideans Navanax inermis and Philinopsisdepicta employ chemotaxis to follow the slime trail of theirprey, which include other cephalaspideans or even congenericindividuals. N. inermis and P. depicta sequester alarm pheromonesand allomones from their cephalaspidean prey, which are ejectedwhen N. inermis and P. depicta are disturbed. The specific metabolic patterns of Mediterranean cephalaspideanssuggest that these patterns can be used as chemotaxonomic markers.We propose the use of a single Thin Layer Chromatography todifferentiate among Mediterranean Haminoea species. (Received 3 February 1997; accepted 22 May 1998)     

The Use of Organic and Inorganic Compounds to Increase the Accumulation of Indole Alkaloids in Catharanthus roseus (L.) G. Don Cell Suspension Cultures

Smith, J. I, Smart, N. J., Kurz, W. G. W. and Misawa, M. 1987.The use of organic and inorganic compounds to increase the accumulationof indole alkaloids in Catharanthus roseus (L.) G. Don cellsuspension cultures.—J. exp. Bot. 38: 1501–1506. The addition of sodium chloride, potassium chloride or sorbitolto 5–d–old cell suspension cultures of Catharanthusroseus stimulated an increase in the intracellular accumulationof catharanthine and other indole alkaloids within 48–72h. The magnitude of the response depended upon the concentrationof the compound added. The use of such inexpensive and readilyavailable compounds to increase the yields and reduce the requiredculture times has considerable potential for the productionof useful secondary metabolites from cell cultures of C. roseusand other plant species.