Dynamics of Cold Induced Inhibition of Phloem Transport

Inhibition of phloem transport caused by chilling has been studiedby in vivo measurement of temporal tracer profiles observedboth uptstream and downstream of the chilled region. The dynamicsof the profile changes indicate that blockage is immediate butdoes not cause flow along the pathway to cease for some considerabletime (50–100 min). This behaviour is anticipated for apressure driven mass flow system. These findings demonstratethe caution needed in interpretation of sink accumulation rates. Key words: Phloem transport, Chilling     

Multiple imputation when records used for imputation are not used or disseminated for analysis

When some of the records used to estimate the imputation modelsin multiple imputation are not used or available for analysis,the usual multiple imputation variance estimator has positivebias. We present an alternative approach that enables unbiasedestimation of variances and, hence, calibrated inferences insuch contexts. First, using all records, the imputer samplesm values of the parameters of the imputation model. Second,for each parameter draw, the imputer simulates the missing valuesfor all records n times. From these mn completed datasets, theimputer can analyse or disseminate the appropriate subset ofrecords. We develop methods for interval estimation and significancetesting for this approach. Methods are presented in the contextof multiple imputation for measurement error.     

The Adaptation of Enzymes to Pressure I. A Comparison of Trout Liver Fructose Diphosphatase with the Homologous Enzyme from an Off-Shore Benthic Fish

SYNOPSIS. At low temperature (3°C), in the absence of substrateand cofactor, trout liver fructose diphosphatase (FDPase) isinactivated by exposures to relatively low pressures. FDP andMg²⁺ protect against this inactivation; hence, maximum catalysisat pH 7.5 is pressure insensitive, while at more alkaline pH,it is markedly accelerated by pressure. The volume change ofactivation, V*, at saturating FDP and Mg²⁺ concentrations isabout –40 cm₃/mole. The apparent Km for FDP and the Kafor Mg²⁺ are markedly increased by pressure. At low FDP or Mg²⁺levels these kinetic properties outweigh V* in determining thereaction rate; hence, under these conditions, piessure retardscatalysis. Similarly, the K₄ for AMP is notably pressure sensitive.Comparable effects of pressure on the kinetic constants forliver FDPase from benthic Corypliaenoides are much less pronounced,suggesting that in these off-shore species enzyme-substrate,enzyme-cofactor, and enzyme-modulator interactions have beentailored through evolution for pressure independent catalyticfunction.     

A Reanalysis of the Kinetics of 14C Photosynthate Translocation in Morning Glory Vines

The spatial and temporal profiles of ¹⁴C photosynthate observedafter a 5 min pulse-labelling period in morning glory vinesare examined. In these experiments a peak of radioactivity appearsto travel down the stem, growing shorter and broader with timeand distance. It is possible to fit the known data reasonably well with aHorwitz reversible-loss model in which the observed changesin peak height and width are attributed to the reversible exchangeof tracer molecules between the translocating sieve tube anda stationary reservoir. The parameters that determine the spatialand temporal profiles are the input kinetics, the sap velocity,v, the ratio of the reservoir volume to sieve tube volume, ,and the permeability to radius ratio, P/r. Even when the inputkinetics are known it is possible to obtain several equallygood fits to the observed tracer profiles with different combinationsof v, , and P/r. With our present state of knowledge it is notpossible to characterize these parameters uniquely, and it isunlikely that tracer techniques alone will yield definite valuesfor v, , and P/r. Ipomoea oil Roth, morning glory, translocation profiles, Horwitz reversible-loss model, mathematical models     

Phloem Unloading within the Seed coat of Pisum sativum Observed using Surgically Modified Seeds

Phloem unloading in pea seed coats was observed by removingthe embryos from developing seeds and washing the attached coatswith a weakly buffered solution. The quantity of labelled photosynthateappearing in the washing solution varied immediately when thesolute concentration was changed, and is shown to be an osmoticresponse. This response is predicted by the Münch theoryof phloem transport with concentration dependent unloading.Respiratory inhibitors and the sulphydryl modifying reagentPCMBS had a slow effect upon the washout of tracer, which arrivedwithin the seed coat prior to inhibitor application, but completelystopped any washout of tracer arriving after its application.This time-course suggests that the inhibitors were not directlyinhibiting unloading, but preventing further tracer from enteringthe region of unloading within the seed coat. Phloem unloadingwithin the seed coats of Pisum appears to be passive and notdependent upon a PCMBS-sensitive carrier. Key words: Pisum sativum, seeds, phloem unloading     

Evaluation of bacterivory of Rotifera based on measurements of in situ ingestion of fluorescent particles, including some comparisons with Cladocera

Bacterivory of pelagic rotifers and cladocerans in eutrophicLake Loosdrecht (The Netherlands) was determined by microscopicobservation of in situ tracer particle uptake. Ingestion ratesof rotifer species using 0.51 µm microspheres or fluorescentlylabelled bacteria as tracers differed, with one exception. Theingestion rates depended on both the species and the tracertype. For cladocerans, fluorescently labelled bacteria seemedto underestimate grazing, presumably due to rapid digestionof tracer cells. Comparing results obtained with 0.51 µmmicrospheres, rotifers were much more important grazers on bacteriathan cladocerans in the study period (April-September). Basedon microspheres, the rotifer populations with the highest uptakeof bacteria were Filinia longiseta (May-July) andAnuraeopsisfissa (June-September). According to the uptake of fluorescentlylabelled bacteria, Conochilus unicornis had the highest uptakein June and A.fissa in July.     

Determining in situ bacterial filtration rates by zooplankton: improvements to the [methyl-3H]thymidine labelling technique

Bacterial feeding by macrozooplankton was studied in Lake Hartbeespoort,a hypertrophic reservoir with abundant coarse (>60 µm)organic particulates. principally Microcystis colonies. Feedingrates were measured in situ, using a twin-barrelled grazingchamber. Filtration rates determined for natural free-livingbacteria labelled with [methyl-³H]thymidine using publishedtechniques proved unacceptably imprecise, unreliable and insensitive.The nature and magnitude of contributory sources of error wereevaluated. Major shortcomings identified were: (i) inefficientconcentration of radiolabelled natural bacteria; (ii) inadequateradiolabel uptake by bacteria; (iii) inadequate removal of unincorporatedlabel and significant release of incorporated label; (iv) unacceptablyhigh and variable surface adsorption errors; (v) poor controlof isotope loss on preservation. New and modified experimentalprocedures, designed and tested to overcome these difficulties,are described. Efficient concentration of natural bacteria (10-foldincrease) using tangential ultrafiltration (TUF), and increasingspecific activity of the tracer by overnight incubation of bacteriawith [methyl-³H]thymidine improved measurement sensitivity.The removal of free (released) isotope from tracer suspensionsby TUF-rinsing shortly before in Situ exposure and the subsequentsaturation of uptake kinetics by the addition of unlabelledthymidine. along with the chilling of labelled bacteria duringtransport and pre-experimental manipulations, considerably reducedadsorption error. Adsorption of free radiolabelled compoundswas measured for each date- specific experimental series usinga modified ‘killed-control’ procedure. which reducedthis error to 11% on average. Estimates of isotope loss (averaging57.6%) associated with sample preservation were measured inparallel ship-board experiments on each occasion. These modificationsgave considerably more reliable and realistic measurements ofspecies-specific filtration rates.     

Radial Water and Solute Flows in Roots of Zea mays: IV. ELECTRICAL POTENTIAL PROFILES ACROSS THE ROOT

Techniques have been developed for continuous recording of electricalpotential difference in maize root preparations. In this waythe continuous p.d. profile along the lateral axis of the rootcould be obtained. The features of the profile are resolvedin terms of the root morphology. Analysis of the p.d. profile indicates (a) that the phases withinthe root are electrically negative with respect to the bathingmedium, (b) that the vacuoles are positive relative to the cytoplasm,i.e. a p.d. exists across the tonoplast, (c) that all cytoplasmicphases are equipotential, which indicates a continuity withinand between the cortical and stelar symplasms, (d) that theextracellular space of the root is presumably only slightlyinfluenced by the concentration of the external medium sincethe p.d. profile within the root does not react to changes ofconcentration in the external medium, (e) that the potentialof the stelar vacuoles is negative relative to the corticalone and this is consistent with the accumulation of surplusions in the stelar vacuoles. The implications of the present findings to the elucidationof ion transport mechanism across the root are briefly discussed.     

A Comparison of the Heat Pulse Method and Deuterium Tracing Method for Measuring Transpiration from Eucalyptus grandis Trees

Estimates of transpiration obtained with the heat pulse velocity(HPV) method and the deuterium tracing method were comparedin two Eucalyptus grandis Hill ex Maiden trees aged 18 monthsand 4 years, respectively. The HPV estimate of weighted meandaily flow rate differed from the deuterium mean by 7% in theolder tree, and 35% in the younger tree. An analysis of possiblesources of error revealed that the discrepancy in the youngertree may have arisen as a result of inadequate probe espacementradially through the sapwood, or from sapwood disruption causedwhen holes were drilled to release the tracer. We conclude thatthe two methods can provide very similar estimates of mean weightedtranspiration rate in E. grandis trees if due precautions aretaken to minimize known sources of error. The relative meritsof the two techniques, which in many respects are complementary,are discussed. Key words: Eucalyptus grandis, heat pulse velocity, deuterium, tracer, transpiration     

Two-stimulus versus one-stimulus procedure in the framework of functional measurement: a comparative investigation using quinine HCl/NaCl mixtures

When subjects are requested to judge single stimuli the observableresponses are the result of both sensory and judgmental processes.De Graaf et al. (1987) employed functional measurement in combinationwith a two-stimulus procedure in order to separate these twotypes of processes. This paper discusses the results of twoexperiments investigating taste interactions in quinine HCl/NaClmixtures. The first experiment employed a one-stimulus procedure,the second experiment employed De Graaf et al.'s two-stimulusprocedure. Comparing the two procedures, the main advantageof the one-stimulus procedure seems to lie in its simplicity.In addition, it enables the determination of a scale value forwater. However, the obtained ratings are context-dependent andare affected by end effects of the response scale. The mostimportant benefit of the two-stimulus procedure is that it allowsfor a post-experimental verification of the linearity of theresponse-output function. This check ensures that all scalevalues are assessed on an interval scale. If water can be assumedto be tasteless, ratio scale values can be obtained. It wasshown that quinine bitterness is largely suppressed by NaCl,whilst the saltiness intensity elicited by NaCl remains virtuallyunaffected. In both experiments, the perceived total taste intensityof a mixture could be well predicted by the weighted sum ofthe saltiness and bitterness sensations within the mixture percept.     

Ion Fluxes to the Vacuole of Nitella translucens

The time course of the appearance in the vacuole of Nitellatranslucens and of Tolypella intricata of tracer from the outsidesolution has been studied over short periods of uptake. Thereare two components of chloride transfer to the vacuole, a fastcomponent linear with time and a second component at longertimes whose behaviour is reasonably well described in termsof a single rate constant for exchange; a constant fractionof the total entry is in the fast component and the apparentrate constant for the second component is proportional to theinflux. In Nitella the path of rapid transfer involves chlorideand sodium, and may also involve a small but variable amountof potassium, but in Tolypella potassium has a significant componentof rapid transfer; these correspond to the cations for whichchloride-linked components of cation influx have been shownby another worker. Over both parts of the time course the level of activity inthe cytoplasm specifies, not the rate of transfer to the vacuoleas would be expected, but the rate as a fraction of the influx;the processes of influx to the cell and transfer to the vacuoleare intimately linked. It is difficult to explain the results in terms of static membranesand fixed compartments. An explanation in terms of the sequence,entry of salt by pino-cytotic vesicles at the plasmalemma, fusionof these vesicles with the endoplasmic reticulum after someloss of tracer to the surrounding cytoplasm, and transfer tothe vacuole in minivacuoles formed from the endoplasmic reticulum,is consistent with the time course found. A model of this kind,involving transport by a dynamic membrane system, seems necessaryto explain the results.     

Time-series measurement of grazing rates of zooplankton and bivalves

The current state of the art in automated measurement of grazingrates of zooplankton and bivalves is evaluated. Limitationsto the development of automated methods are discussed with referenceto theoretical considerations. Different approaches to time-seriesmeasurements are examined, and some inadequately investigatedtime scales for measurement of grazing rates are outlined. Levelsof automation of existing experimental systems are described,and a recently developed grazing system based on in vivo fluorescenceadvanced. The new system is suited to use with zooplankton orbivalves, but depends on precise calibration of in vivo fluorescence.It facilitates rapid measurement of functional response to foodconcentration, estimation of time-series rates at constant foodconcentrations, and measurement of rates while simulating patchyfood distribution. Results derived from experiments with krill(Nyctiphanes australis) and cockles (Chione stutchburyi) usingthis grazing system are presented, and some future directionsfor instrument development suggested.     

Spatially quantitative control of the number of cotyledons in a clonal population of somatic embryos of hybrid larch Larix x leptoeuropaea

• Background and Aims Many conifer embryos, both in naturalseeds and in clonal populations of somatic embryos, displayvariability in the number of cotyledons. In hybrid larch, Larixx leptoeuropaea (synonymous with L. x marschlinsii Coaz), suchvariability has previously been reported in somatic embryos,together with a decrease in the average cotyledon number whenbenzyladenine (BA) is applied exogenously. Described here isa spatially quantitative study with the aim of throwing somelight on the way cotyledon number is determined, and hence themechanism of cotyledon formation. • Methods Stock cultures of embryogenic tissue were maintainedand later made embryogenically active by standard methods. Developmentthrough cotyledon formation was followed by optical microscopywith quantitative measurement of embryo diameter and numberof cotyledons. SEMs of representative stages and cotyledon numberswere done for purposes of illustration in this account. Existingmathematics of waveforms on a disc were cast into a form suitableto compare with the quantitative data. • Key Results The number of cotyledons is linearly relatedto the diameter of the apical surface of the embryo (which approximatesa circular disc) at the time of first appearance of the cotyledonprimordia. This linearity is a constant-spacing phenomenon betweenadjacent primordia. Addition of BA to the medium restricts therange of apical diameters without changing inter-cotyledon spacing.Slope/intercept ratio of the linear plot matches expectationfor initiation of cotyledon pattern as a harmonic waveform ona circular disc. • Conclusions The entire pattern of cotyledon primordiaarises as a single entity coordinated by a mechanism with wave-formingproperties. This is explicable by diverse mechanisms, especiallyeither mechanical buckling (‘biophysical’) or reaction–diffusionkinetics (‘physicochemical’).     

Turnover of Carbohydrates in Relation to Growth in Apple Trees. II. Distribution of 14C Assimilates Labelled in Autumn, Spring and Summer

One-year-old plants of the apple rootstocks MM. 106 and M.7were allowed to assimilate ¹⁴CO₂ in autumn, spring and summerand the distribution of the tracer within the plant over a growingseason was followed. In MM.106 distribution of the tracer intwo fractions of extractable carbohydrate and the residues representingstructural material, was determined. The results of the radioactivityassay are discussed in relation to seasonal patterns of assimilatesupply and demand in the different regions of the plant. Malus sylvestris L, apple, ¹⁴C assimilates, distribution of carbohydrates     

Diffusion measurement of fluorescence-labeled amphiphilic molecules with a standard fluorescence microscope.

The lateral diffusion of fluorescence-labeled amphiphilic tracer molecules dissolved within a two-dimensional matrix of lipids was measured by continuous illumination of an elongated rectangular region. The resulting spatial concentration profile of unbleached tracer molecules was observed with a standard epifluorescence microscope and analyzed with digital image-processing techniques. These concentration profiles are governed by the mobility of the tracers, their rate of photolysis, and the geometry of the illuminated area. For the case of a long and narrow illuminated stripe, a mathematical analysis of the process is given. After prolonged exposure, the concentration profile can be approximated by a simple analytical function. This fact was used to measure the quotient of the rate of photolysis, and the diffusion constant of the fluorescent probe. With an additional measurement of the rate of photolysis, the mobility of the tracer was determined. As prototype experiments we studied the temperature dependence of the lateral diffusion of N-(7-nitrobenz-2-oxa-1,3-diazol-4-yl)-dipalmitoylphosphatidyl++ + ethanolamine in glass-supported bilayers of L-alpha-dimyristoylphosphatidylcholine. Because of its simple experimental setup, this technique represents a very useful method of determining the lateral diffusion of fluorescence-labeled membrane molecules.     

Rates of Axial Transport of 11C and 14C in Characean Cells: Faster than Visible Streaming?

Movements of ¹¹C and ¹⁴C in N. translucens occur at rates twoto five times greater than those of visually observed streaming.Like cytoplasmic streaming, longitudinal tracer movement isstopped reversibly by action potentials, irreversibly by N-ethylmaleimide(NEM) and is unaffected by colchicine (colch), suggesting thatboth processes share a common mechanism. Colchicine causes anincreased loading delay time. In both N. translucens and C. corallina the activity of ¹¹Cand ¹⁴C at the nodes shows a sharp discontinuity: it is lowon the feed side of the node and high at the node itself. Thissuggests that transport across plasmodesmata may be active. Key words: Axial-carbon transport, Cytoplasmic streaming, Carbon     

Bio-electric Potentials of Intact Green Plants: III. EFFECTS OF JACKETING A REGION OF THE PLANT TISSUE 3

When measuring the potential difference between two regionsof an oat seedling the P.D. may change if a region of the tissuebetween the measuring contacts is surrounded by a mineral saltsolution. This change is shown to be equal to (E₂—E₁)/(1+R₂/R₁)where E₁ is the E.M.F. generated by the shunted tissue and R₁its resistance, and E₂ is the net E.M.F. generated at the shuntingsolution and the tissue interfaces and R₂ the shunt resistance,expected to be proportional to the specific resistance of theshunting solution. However, in these experiments R₂/R₁ was closeto unity because of a large resistance from tissue to shuntingsolution and there was no relationship between the P.D. changeand the resistance of the shunt. Where the shunt was appliedbetween two dissimilar regions of the plant (coleoptile andprimary leaf) E₂ changed with the concentration of the shuntingsolution, and hence the measured potential difference was changedby varied amounts. When the shunt was applied to a primary leafalone, E₂ was independent of solution concentration, and thechange in P.D. was constant.     

Dispersion characteristics of two planktonic diatoms

Not all phyloplankton behave as ideal water tracers. Longitudinaldispersion experiments were conducted in a large flume withseawater of constant depth and velocity. The transport of thepassive, fluorescent tracer, Rhodamine B dye, was compared withunialgal cultures of a large, fast-sinking diatom (Odontellachinensis) and a small, slow-sinking diatom (Skeletonema costatum).Horizontal dispersion rates, proportional to variance per distance,were linear for the dye and Skeletonema but nonlinear for Odontella.Results are explained by the interaction of Odontella sinkingand advection. Longitudinal dispersion coefficients for thedye and Skeletonema were between 60–80 cm² s^–1 andas high as 280 cm² s^–1 for Odontella. This may have implicationsfor nutrient uptake in natural phytoplankton populations.     

Potassium Fluxes in Excised Barley Roots

The method of the modified compartmental analysis for excisedroots has been adopted for measuring K⁺-fluxes and compartmentationin barley (Hordeum distichon) roots. Efflux of ⁴²K and ⁸⁶Rbindicated that more than two intracellular compartments wereinvolved in the tracer exchange; the ⁴²K data clearly showedthe components. On the basis of the efflux behaviour of theapical and more basal tissues of the roots, the three componentsof efflux were attributed to the cytoplasm of differentiated(fast) and meristematic tissues (intermediate) and to the vacuoles(slow exchange) of the roots. A model is proposed on the basisof which, the fluxes corresponding to the meristematic and differentiatedtissues of the root can be estimated. Additionally, fluxes ofthe differentiated root tissues were determined by using effluxdata obtained with root segments without apical tissues. Thedata obtained in both ways compare reasonably well and agreeto independent chemical measurements. Comparison of the ⁴²K and ⁸⁶Rb efflux data show strong discriminationof K^– in favour of Rb⁺ and indicate that ⁸⁶Rb is not suitableas a tracer for K⁺ in efflux measurements, at least with barleyroots.     

Sulphate supply and its regulation of transport in roots of a tropical legume Macroptilium atropurpureum cv. Sirato

During a period of sulphate deprivation, roots of Macroptiliumatropurpureum responded by increasing their uptake capacityat the plasma membrane. This effect was apparent both in intactplants and in tissues excised prior to uptake. In experiments using excised root systems previousy labelledwith ³⁵SO₄^2- the rate of tracer transport to the xylem was muchgreater in roots subsequently deprived of external sulphatethan in those supplied with unlabelled sulphate. Removing theexternal sulphate to the external solution. Additionally, compartmentalanalysis of tracer exchange kinetics showed that the flux ofsulphate from the cytoplasm to the xylem(