Sample preconditioning for measurement of fluorescence induction of chlorophyll a in marine phytoplankton

Conditions for measuring fluorescence induction curves (time-scalems) of in vivo chlorophyll ^a were studied using cultures ofDunaliella tertiolecta Butcher (Chlorophyceae) and of Thalassiosirapseudonana Hustedt (3H) (Bacillariophyceae), and samples ofnatural phytoplankton populations from the Grand Banks. Thearea above the fluorescence induction curve (A_DCMU) and themaximum fluorescence intensity (F_max) measured in the presenceof 3-(3,4-dichlorophenyl)-1, 1-dimethylurea (DCMU) were computedby microcomputer. Cells must be ‘conditioned’ or‘adapted’ prior to obtaining a fluorescence inductioncurve; dark-adaptation resulted in a lower A_DCMU and F_max thandid adaptation in far-red (720 nm) light, and was the conditioningmethod chosen. A_DCMU and F_max increased linearly with increasingirradiance up to 32.8 W m^–2 the highest actinic irradianceavailable. Information on the light history of D. tertiolectawas obtained by following the time-course of change in A_DCMUand in F_max for cells exposed for 10 min to far-red or to bluelight. The rise-time of the fluorescence induction curve andvalues of F_max were greater for samples of D. tertiolecta concentratedonto glass-fiber filters than for liquid samples, however, valuesof A_DCMU for filtered and liquid samples were not significantlydifferent. Samples of Grand Banks phytoplankton collected ontoglass-fiber filters and frozen for 28 d exhibited a significantdecrease in F_max and in A_DCMU relative to the same freshly-filteredsamples. Filtration and freezing of samples is not recommended. ^*This paper is the result of a study made at the Group for AquaticPrimary Productivity (GAP). Second International Workshop heldat the National Oceanographic Institute. Haifa. Israel in April–May1984.     

The Steady State Chlorophyll a Fluorescence Exhibits in Vivo an Optimum as a Function of Light Intensity which Reflects the Physiological State of the Plant

Modulated (690 and 730 nm), as well as direct chlorophyll (Chl)a fluorescence and changes in the concentration of the oxidizedP700 were measured under steady state conditions in leaves ofhigher plants adapted to different light intensities. All theleaf samples exhibit an optimum curve of steady state fluorescenceyield (F_s) versus the light intensity but its position withrespect to light intensity varies considerably from one speciesto another or from one sample to other even in the same plantor within the same leaf sample. However, the optimum level ofF_s was always at a moderate light intensity. By using the modulatedfluorescence technique, the system with all closed (F^l_m) oropen reaction center (F^l_o) were measured in steady state conditions.Each experimentally measured fluorescence yield was separatedinto a fluorescence emission of open (F_open = F^l_o,(1—V_s))and closed (F_closed = (F^l_m . V_s)) reaction center (RC) of photosystemII where V_s=(F_s – F^l_o)/(F^l_m – F^l_o) is the functionof fraction of closed reaction centers. With increasing lightintensity, the fraction of open RC decreased while the fractionof closed RC increased. Maximum quantum efficiency (P_o) andactual quantum efficiency (P) decreased by increasing lightintensity. An optimum level of F_s was observed, when the fractionof closed reaction centers V_s of each sample was about 0.2 showinga common quenching mechanism which determines the fluorescenceproperties under steady state condition. This explains the apparentphenomenological contradiction that the fluorescence yield understeady state conditions can increase or decrease upon an increaseof actinic light. (Received December 31, 1994; Accepted May 1, 1995)     

Further Evidence for the Relationship between Light-Induced Changes of Plasmalemma Transport and Transthylakoid Proton Uptake

The simultaneous measurement of the induction curves of chlorophyllfluorescence, its responses to saturating flashes, light-scatteringat 532 nm, and plasmalemma voltage supports previous findings(Hansen, Kolbowski, and Dau, 1987), that light-induced uptakeof protons into the inner thylakoid space causes the rapid (5to 20 s) light-induced depolarization at the plasmalemma viasubstrate depletion of the electrogenic H⁺-pump. These conclusionsare based on kinetic studies which enable the separation ofindividual components in complex signals by means of their assignmentto different time-constants. In contrast to the previous investigation,binary noise was used for modulation of the actinic light. Thenew input signal not only increased the reliability of the previousresults obtained by sine-waves, but also led to the detectionof three additional time-constants. One of these is probablyrelated to the action of light on the potassium channel of theplasmalemma. The others are assigned to the quencher Q and toa still unknown process. Key words: Chlorophyll fluorescence, plasmalemma potential, proton fluxes, noise, scattering, spinach, state-transitions, thylakoid membrane     

Cerebral vasomotor reactivity at high altitude in humans

The purpose of this study was twofold:1) to determine whether at highaltitude cerebral blood flow (CBF) as assessed during CO₂ inhalation and duringhyperventilation in subjects with acute mountain sickness (AMS) wasdifferent from that in subjects without AMS and2) to compare the CBF as assessedunder similar conditions in Sherpas at high altitude and in subjects atsea level. Resting control values of blood flow velocity in themiddle cerebral artery (V_MCA), pulseoxygen saturation (Sa_O2), andtranscutaneous PCO₂ were measured at4,243 m in 43 subjects without AMS, 17 subjects with AMS, 20 Sherpas,and 13 subjects at sea level. Responses ofCO₂ inhalation andhyperventilation onV_MCA,Sa_O2, and transcutaneous PCO₂ were measured, and the cerebralvasomotor reactivity (VMR = V_MCA/PCO₂)was calculated as the fractional change ofV_MCA per Torrchange of PCO₂, yielding ahypercapnic VMR and a hypocapnic VMR. AMS subjects showeda significantly higher resting controlV_MCA than didno-AMS subjects (74 ± 22 and 56 ± 14 cm/s, respectively;P < 0.001), andSa_O2 was significantly lower (80 ± 8 and 88 ± 3%, respectively; P < 0.001). Resting control V_MCA values inthe sea-level group (60 ± 15 cm/s), in the no-AMS group, and inSherpas (59 ± 13 cm/s) were not different. Hypercapnic VMR valuesin AMS subjects were 4.0 ± 4.4, in no-AMS subjects were 5.5 ± 4.3, in Sherpas were 5.6 ± 4.1, and in sea-level subjects were 5.6 ± 2.5 (not significant). Hypocapnic VMR values were significantly higher in AMS subjects (5.9 ± 1.5) compared with no-AMS subjects (4.8 ± 1.4; P < 0.005) but werenot significantly different between Sherpas (3.8 ± 1.1) and thesea-level group (2.8 ± 0.7). We conclude that AMS subjects havegreater cerebral hemodynamic responses to hyperventilation, higherV_MCAresting control values, and lower Sa_O2 compared with no-AMSsubjects. Sherpas showed a cerebral hemodynamic patternsimilar to that of normal subjects at sea level.     

Membrane Depolarization by Hexacyanoferrate (III), Hexabromoiridate (IV) and Hexachloroiridate (IV)

Three artificial electron acceptors of different E_o and charge,hexacyanoferrate (III) (K₃Fe(CN)₆), hexachloroiridate (IV) (K₂IrCl₆),and hexabromoiridate (IV) (K₂IrBr₆), were compared with respectto their rate of reduction by roots of Zea mays L., the concomitantproton secretion, and to the effect on plasmalemma depolarization. It has been shown that these plasma membrane impermeable electronacceptors were reduced by a plasmalemma reductase activity.At low concentrations proton secretion was slightly inhibited,at higher concentrations, however, the rate of proton secretionwas stimulated. The root cell plasmalemma showed a transientdepolarization after addition of all three electron acceptors.The depolarization was concentration-dependent for the iridatecomplexes but not for hexacyanoferrate (III). For both iridatecomplexes maximum depolarization was reached at 50 µmoldm^–3. A hypothetical model as an explanation of the redox dependentproton secretion will be given. Key words: Hexachloroiridate (IV), hexabromoiridate (IV), hexacyanoferrate (III), plasmalemma redox, membrane potential, Zea mays     

The Influence of Temperature on a K+-Channel and on a Carrier-Type Transporter in Nitella

Fisahn, J. and Hansen, U-P. 1986. The influence of temperatureon a K⁺ -channel and on a carrier type transporter in Nilella—J.exp. Bot. 37. 440–460. In Nitella, the effects of temperature on membrane potentialand on resistance consist of several components. The evaluationof their associated time-constants measured in linear(ized)temperature responses at a resting potential of–120 mVprovides an approach to their identification. For changes slowerthan c. 1 s, the temperature effect on membrane potential andresistance does not originate from temperature action on theinvolved transporter, but is mediated by signals from temperaturesensitive metabolic processes. In the case of potential, theseprocesses seem to be identical to those which also mediate thelight effect: pH-regulation, and two direct signals from photosynthesis,as indicated by the similarities of the related time-constants( respectively). The temperature effect on resistance displays only one time-constant of 40 sinmost experiments. The related process is unknown. The non-coincidenceof the time-constants of the effect on potential and on resistanceimplies the involvement of a carrier-type transporter (H+-pumpor cotransporter) in the effect on potential, and of a K+channelin the effect on resistance. The K+-channel is identified bythe reversal potential of the effect on membrane potential measuredin cells depolarized or hyperpolarized by an injected electricalcurrent Under these conditions the temperature effect on resistancedominates the effect on potential. Key words: H⁺-pump, K⁺-channel, kinetic analysis, Nitella, oscillation, pH-regulation, reversal, potential, temperature, time-constants     

Effects of microtubules and microfilaments on [Ca(2+)](i) and contractility in a reconstituted fibroblast fiber

We used a reconstituted fiber formed when 3T3fibroblasts are grown in collagen to characterize nonmusclecontractility and Ca²⁺ signaling. Calf serum (CS) andthrombin elicited reversible contractures repeatable for >8 h. CSelicited dose-dependent increases in isometric force; 30% produced thelargest forces of 106 ± 12 µN (n = 30), whichis estimated to be 0.5 mN/mm² cell cross-sectionalarea. Half times for contraction and relaxation were 4.7 ± 0.3 and 3.1 ± 0.3 min at 37°C. With imposition of constant shortening velocities, force declined with time, yieldingtime-dependent force-velocity relations. Forces at 5 s fit thehyperbolic Hill equation; maximum velocity(V_max) was 0.035 ± 0.002 L_o/s.Compliance averaged 0.0076 ± 0.0006 L_o/F_o. Disruption of microtubules with nocodazole in a CS-contracted fiber had no net effects on force, V_max, or stiffness; force increased in 8, butdecreased in 13, fibers. Nocodazole did not affect baselineintracellular Ca²⁺ concentration([Ca²⁺]_i) but reduced (~30%) the[Ca²⁺]_i response to CS. The force afternocodazole treatment was the primary determinant of stiffness andV_max, suggesting that microtubules were not amajor component of fiber internal mechanical resistance. Cytochalasin Dhad major inhibitory effects on all contractile parameters measured butlittle effect on [Ca²⁺]_i.

     

Ionic Currents across the Plasmalemma of Chara inflata Cells: III. WATER-RELATIONS PARAMETERS AND THEIR CORRELATION WITH MEMBRANE ELECTRICAL PROPERTIES

The water-relations parameters of Chara inflata cells were determineddirectly using the micro pressure probe technique. The turgorpressure of cells in artificial pond water (₀ = 0.06 MPa) wasabout 0.65 MPa and the half-time (T_1/2) for water exchange wasabout 6.5 s. The calculated values of the hydraulic conductivity(L_P) were in the range 1–2 ? 10^–6m s^–1 (MPa)^–1.The volumetric elastic modulus () was 32.8 MPa for turgor rangingfrom 0.77 to 0.82 MPa. Large changes in the water-relations parameters and the electricalproperties of the membrane occurred when the turgor was decreasedto low values. These changes included: (i) a decrease in theT_1/2 for water exchange, (ii) an increase in L_P and (iii) depolarizationof the membrane potential difference (V_m). The micro pressure probe, which enabled the turgor pressureof the cell to be altered, was used in combination with thevoltage-clamp technique to determine the relationship betweenK⁺ and Cl^– conductances of the plasmalemma and the cellturgor. The K⁺ conductance increased reversibly as the turgorwas reduced in the range 0 to 0.6 MPa and the Cl^– -conductanceincreased as the turgor was reduced in the range 0.1 to 0.5MPa. It is suggested that these pressure-dependent K⁺ and Cl^–conductances may have a dual role in electrical events and thenon-electrical responses such as changes in the cell volume. Key words: Chara inflata, membrane conductances, ion channels, water-relations parameters     

Capillary and arteriolar responses to local vasodilators are impaired in a rat model of sepsis

Although sepsis isknown to affect vascular function, little is known about changes at thecapillary level. We hypothesized that sepsis attenuates the"upstream" arteriolar response to vasoactive agents appliedlocally to capillaries. Sepsis in rats was induced by cecal ligationand perforation. After 24 h, extensor digitorum longus muscle wasprepared for intravital microscopy. Phenylephrine (PE, 10 mM) andacetylcholine (ACh, 10 mM) were applied iontophoretically on terminalarterioles and on their downstream daughter capillaries (300 µm fromarteriole). There was no significant difference between control andseptic rats in baseline arteriolar diameters [8.0 ± 0.6 vs.9.8 ± 0.8 (SE) µm] or baseline red blood cellvelocity (V_RBC)in perfused daughter capillaries (255 ± 10 vs. 264 ± 13 µm/s). Application of PE onto arterioles resulted in comparable constrictions (i.e., 22% diameter change) andV_RBC reductions (100%) in control and septic rats. In contrast, arteriolardiameter and V_RBCincreases after application of ACh were attenuated in sepsis (diameter:from 41 to 14%;V_RBC: from 67 to24%). Application of PE onto the capillary reducedV_RBC to the samelevel (100%) in both groups, whereas application of AChincreased V_RBCless in septic than in control rats (20 vs. 73%). On the basis ofarteriolar-capillary pair stimulations, sepsis affectedV_RBC responses toACh more in the capillary than in the arteriole. When the adenosineanalog 5'-N-ethylcarboxamidoadenosine(0.1 mM) was used instead of ACh, similar effects of sepsis were seen.To test for a possible involvement of inducible NO synthase (iNOS) insepsis-induced attenuated ACh responses, arterioles and capillaries inseptic animals were locally pretreated with the iNOS blockeraminoguanidine (10 mM). In both microvessels, aminoguanidine restoredthe ACh response to the control level. We conclude that impairedcapillary V_RBCand arteriolar diameter responses to vasodilators applied tocapillaries in septic rat skeletal muscle were due to dysfunction atarteriolar and capillary levels. The study underscores the significantrole iNOS/NO may play in sepsis-induced alteration of vascularreactivity in vivo.

     

Rhizoid differentiation in Spirogyra II. Photoreversibility of rhizoid induction by red and far-red light

The optimum light conditions for rhizoid formation in Spirogyrawere determined. Red light was significantly effective on rhizoidformation while green, blue and violet lights had less effect.The dose-effect curve of red light was investigated and theminimum energy needed to saturate the effect was 8.1 Kergs.cm_–2.The effect of red light was completely reversed by subsequentirradiation with far-red light. The doseeffect curve of far-redlight was also obtained. The repeatedly reversible photoresponseswith red and far-red light strongly suggest that the photoreceptorof the rhizoid formation system is phytochrome. The existenceof phytochrome in the Spirogyra cell was also demonstrated spectrophotometrically.The half time for the escape reaction from the reversal effectof far-red light was 2hr. There may be no pigment other thanphytochrome mediating the photoreaction. (Received December 14, 1972; )     

The Location of the Transporting System for Inorganic Carbon and the Nature of the Form Translocated in Chlamydomonas reinhardtii

The permeability of the plasmalemma of Chlamydomonas reinhardtiicells was increased by treatment with poly-L-lysine or dimethylsulphoxideas indicated by 3-phosphoglyceric acid dependent O₂ evolution.These treatments decreased the ability of the cells to accumulateinorganic carbon internally and hence their photosynthetic affinityfor inorganic carbon in the medium. With saturating light andinorganic carbon, the photosynthetic rate was less affectedby the poly-L-lysine and dimethylsulphoxide treatments. Thusthe poly-L-lysine and dimethylsulphoxide did not alter the activityof the chloroplasts but rather made the intracellular inorganiccarbon pool more freely exchangeable with the medium. It isconcluded that the transporting system for inorganic carbonis located at the plasmalemma. Treatment with Diamox, an inhibitor of carbonic anhydrase, didnot affect photosynthetic rate and accumulation of inorganiccarbon when CO₂ was supplied but strongly inhibited both parameterswhen HCO^–₃ was supplied. In a mutant of Chlamydomonasreinhardtii lacking a cell wall, carbonic anhydrase leaks tothe medium and uptake of inorganic carbon is much faster whenCO₂ is supplied than when HCO^–₃ is supplied. These resultssuggest that CO₂ rather than HCO^–₃ is the inorganic carbonspecies that is actively translocated across the plasmalemma. Key words: Chlamydomonas, Inorganic carbon uptake     

The Permeability of Ammonia, Methylamine and Ethylamine in the Charophyte Chara corallina (C. australis)

Ritchie, R. J. 1987. The permeability of ammonia, methylamineand ethylamine in the charophyte Chara corallina (C. australis).—J.exp. Bot. 38: 67–76 The permeabilities of the amines, ammonia (NH₃), methylamine(CH₃NH₂) and ethylamine (CH₃CH₂NH₂) in the giant-celled charophyteChara corallina (C. australis) R.Br. have been measured andcompared. The permeabilities were corrected for uptake fluxesof the amine cations. Based on net uptake rates, the permeabilityof ammonia was 6?4?0?93 µm s^–1 (n = 38). The permeabilitiesof methylamine and ethylamine were measured in net and exchangeflux experiments. The permeabilities of methylamine were notsignificantly different in net and exchange experiments, norto that of ammonia (P_methylamine = 6?0?0?49 µm s^–1(n = 44)). In net flux experiments the apparent permeabilityof ethylamine was slightly greater than that of ammonia andmethylamine (P_{ethylamine, net} = 8?4?1?2 µm s^–1 (n= 40)) but the permeability of ethylamine based on exchangeflux data was significantly higher (P_{ethylamine, exchange} =14?1?2 µm s^–1 (n = 20)). Methylamine can be validlyused as an ammonium analogue in permeability studies in Chara. The plasmalemma of Chara has acid and alkaline bands; littlediffusion of uncharged amines would occur across the acid bands.The actual permeability of amines across the alkaline bandsis probably about twice the values quoted above on a whole cellbasis i.e. the permeability of ammonia across the permeablepart of the plasmalemma is probably about 12 µm s^–1. Key words: Chara, permeability, ammonia, methylamine     

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

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

The Diffusive Conductivity of the Stomata of Wheat Leaves

A leaf chamber (described in detail) was used alternately witha resistance porometer to measure resistance to viscous flowof air through the leaf, and with a diffusion porometer to measurethe differential diffusive flow of hydrogen and air (V_H–V_A)through the leaf and the component of hydrogen flow (V'_H) movingstraight across the leaf. The resistance of the mesophyll isneeded for interpretation: estimates by three different methodsfor viscous flow did not agree very well, but two differentmethods for diffusive flow gave good agreement. For wheat leaves,only very large errors are important. Formal analysis is in three appendixes: I. Interpretation ofviscous and diffusive flow in small pores involves some problemsin molecular physics, complicated by the particular geometryof the wheat stoma. With some uncertainty, formal expressionsare derived for the viscous resistance of a single stoma, rv,and for the resistances to diffusion of hydrogen and air, andof water vapour and carbon dioxide, all expressed as r_s persquare centimetre of leaf surface. The analysis for hydrogen/airis the most uncertain; that for water vapour and carbon dioxideis more reliable. II. An indication is given of the flow characteristicsof the leaf-chamber system, from which rv can be derived, andof the basis for estimating mesophyll resistance. III. The methodof converting estimates of r_s into estimates of V_H–V_Aand V'_H is given. The results presented are expressed as nearly as possible interms of the quantities which were measured. For five leavesthe dependence of V_H–V_A on V'_H agrees well with theoreticalpredictions; the dependence of V_H–V_A (and V'_H) on rv,on average, agrees well with prediction, but involves the assumptionthat the stomata get shorter as they close. The agreement isgood enough to suggest that the formal expressions for r_s interms of stomatal dimensions and molecular gas constants arereliable enough to be carried forward into future transpirationand assimilation studies. The minimum value of ra for watervapour (c. 3 sec cm⁺¹) is close to values found elsewhere bydifferent techniques. At very small stomatal openings there was a large deviationfrom predicted behaviour, such as would occur if the imposedexcess air pressure further closed the stomata during viscousflow experiments.     

Photomodulation of Stem Extension in Light-Grown Plants: Evidence for Two Reactions

Internode elongation was measured in plants of Phaseolus vulgarisand Glycine max grown under 8 h photoperiods at 25 W m^–2in white fluorescent light, followed by light-extensions varyingin quality, irradiance and duration. Two distinct responsesto light were observed under these conditions. A reduction in P_FR/P increased elongation, but elongation wasalso modified by a second reaction in which internode lengthincreased with increase in the duration and irradiance of theday-extension. This light-promoted response occurred in bothred and blue light. In the P_FR-inhibition response, light acteddirectly on the expanding internode. The light-promoted response,in contrast, required irradiation of the leaf. The response to a short end-of-day exposure to far-red lightprogressively diminished as successive internodes expanded underthe treatment, whereas the light-promoted response increased.The two processes appeared to interact and, in the later-expandinginternodes, the effect of a reduction in P_FR was greater underlong day-extensions with mixed red and far-red light than inthe end-of-day treatments. ¹ Present address: British Telecom, Brunel House, 2 FitzalanRoad, Cardiff, U.K.     

Nitrogen Utilization in N-limited Barley During Vegetative and Generative Growth: III. POST-ANTHESIS KINETICS OF NET NITRATE UPTAKE AND THE ROLE OF THE RELATIVE ROOT SIZE IN DETERMINING THE CAPACITY FOR NITRATE ACQUISITION

Barley (Hordeum vulgare L., cvs Golf, Mette, and Laevigatum)was grown under nitrogen limitation in solution culture untilnear maturity. Three different nitrogen addition regimes wereused: in the ‘HN’ culture the relative rate of nitrate-Naddition (RA) was 0·08 d^–1 until day 48 and thendecreased stepwise to, finally, 0·005 d^–1 duringgrain-filling; the ‘LN’ culture received 45% ofthe nitrogen added in HN; the ‘CN’ culture was maintainedat RA 0·0375 d^–1 throughout. Kinetics of net nitrateuptake were measured during ontogeny at 30 to 150 mmol m^–3external nitrate. V_max (which is argued to reflect the maximuminflux rate in these plants) declined with age in both HN andLN cultures. A pronounced transient drop was observed just beforeanthesis, which correlated in time with a peak in root nitrateconcentration. Similar, but less pronounced, trends were observedin CN. The relative V_max (unit nitrogen taken up per unit nitrogenin plants and day) in all three cultures declined from 1·3–2·3d^–1 during vegetative growth to 0·1–0·7d^–1 during generative growth. These values are in HN andLN cultures 15- to more than 100-fold in excess of the demandset by growth rates throughout ontogeny. Predicted balancingnitrate concentrations (defined as the nitrate concentrationrequired to support the observed rate of growth) were below6·0 mmol m^–3 in HN and LN cultures before anthesisand then decreased during ontogeny. In CN cultures the balancingnitrate concentration increased during grain-filling. Apartfrom the transient decline during anthesis, most of the effectof ageing on relative V_max can be explained in terms of reducedcontribution of roots to total biomass (R:T). The loss in uptakeper unit root weight is largely compensated for by the declinewith time in average tissue nitrogen concentrations. The quantitativerelationships between relative V_max and R:T in ageing plantsare similar to those observed for vegetative plants culturedat different RAs. The data support the contention that the capacity for nitrateacquisition in N-limited plants is under general growth control,rather than controlled by specific regulation of the biochemicalpathway of nitrate assimilation. Key words: Barley, nitrogen concentration, root: total plant biomass ratio, V_max     

An electrophysiological study of the membrane in aplanospore-like cell of Boergesenia forbesii

Membrane potential and resistance, each of which was the sumof those of the plasmalemma and tonoplast, measured in the coenocyticthallus of Boergesenia forbesii were 6.7 mv inside positiveand 2.8 k.cm², respectively. Protoplasm squeezed from the thallus into artificial sea water(ASW) formed numerous spherical bodies, which are termed aplanospore-likecells (simply "spores"). The following electrical propertiesof the "spores" 20–40 hr after squeezing were obtained:potential difference (p.d.) across plasmalemma (E_co) was –66mv (– means inside negative), plasmalemma resistance 665cm², p.d. across the tonoplast (E_vc) +73 mv, and tonoplast resistance2.6 k.cm². Tenfold increase in external [K⁺] caused +45 mv changein E_co and +17 mv in E_vc. The plasmalemma was entirely depolarizedin Ca⁺⁺-free ASW or ASW containing Triton X-100. When the "spore" was immersed in potassium-rich (277 mil) ASW,E_co was almost zero and the tonoplast showed two states (I andII, E_ve about +70 mv and +20 mv, respectively). E_vc went backand forth between the two states spontaneously or when a smallcurrent was applied. In most cases oscillatory changes in E^vcoccurred after the lapse of a long time in the K+-rich sea water.Membrane resistances in states I and II were 5 and 9 k.cm²,respectively. (Received July 11, 1977; )     

The Influence of Stress Conditions on Chlorophyll Content of Two Range Grasses with Contrasting Photosynthetic Pathways

The influence of various treatments and temperature regimeson total chlorophylls and on the chlorophyll a:b ratio of westernwheatgrass and blue grama plants was investigated at differenttime intervals during the 120-day growth period. Western wheatgrass,a C₃ species, accumulated greater amounts of chlorophyll thandid blue grama plants, a C₄ species. Maximum concentrations(mg gd wt^–1) of chlorophylls in western wheatgrass andin blue grama were recorded at the lower (13/7°C) and higher(30/18°C) temperature regimes. Nitrogen fertilizer alonedecreased the chlorophyll content in both species. The chlorophylla:b ratio in blue grama ranged from an average of 2·00under irrigated plus fertilized conditions to 3·00 undercontrol and fertilized conditions. On the other hand, the chlorophylla:b ratio in western wheatgrass remained constant at 3·00throughout the growing season under various treatments and temperatureregimes.     

Expression of the nuclear gene TaF(A)d is under mitochondrial retrograde regulation in anthers of male sterile wheat plants with timopheevii cytoplasm

Alterations of mitochondrial-encoded subunits of the F_oF₁-ATPsynthase are frequently associated with cytoplasmic male sterility(CMS) in plants; however, little is known about the relationshipof the nuclear encoded subunits of this enzyme with CMS. Inthe present study, the full cDNA of the gene TaF_Ad that encodesthe putative F_Ad subunit of the F_oF₁-ATP synthase was isolatedfrom the wheat (Triticum aestivum) fertility restorer ‘2114’for timopheevii cytoplasm-based CMS. The deduced 238 amino acidpolypeptide is highly similar to its counterparts in dicotsand other monocots but has low homology to its mammalian equivalents.TaF_Ad is a single copy gene in wheat and maps to the short armof the group 6 chromosomes. Transient expression of the TaF_Ad–GFPfusion in onion epidermal cells demonstrated TaF_Ad's mitochondriallocation. TaF_Ad was expressed abundantly in stem, leaf, anther,and ovary tissues of 2114. Nevertheless, its expression wasrepressed in anthers of CMS plants with timopheevii cytoplasm.Genic male sterility did not affect its expression in anthers.The expression of the nuclear gene encoding the 20 kDa subunitof F_o was down-regulated in a manner similar to TaF_Ad in theT-CMS anthers while that of genes encoding the 6 kDa subunitof F_o and the subunit of F₁ was unaffected. These observationsimplied that TaF_Ad is under mitochondrial retrograde regulationin the anthers of CMS plants with timopheevii cytoplasm. Key words: CMS, F_Ad subunit, F_oF₁-ATP synthase, retrograde regulation, wheat Received 8 October 2007; Revised 9 January 2008 Accepted 28 January 2008     

Perfusion of charophyte cells: a critical analysis of the method

The data obtained by different types of intracellular perfusionwere compared. As the ligated cells cannot be space-clamped,the efficiency of compartment-clamping was evaluated, showingthat the difference I/V (current-voltage) profile between space-clampedand compartment-clamped data could be approximated by a straightline. The time-dependence of the clamp currents was not affectedby the clamp technique. The comparison of different sets of data was quantified by fittingthe I/V curves with a mathematical model (Beilby and Walker,1996). The I/V curves of ligated cells perfused with 1 mM ATPshowed the closest similarity to intact cells with resting potentialsof –22010 mV (7 cells) and similar model parameter values.The cells under open-end perfusion with ATP showed less hyperpolarizedresting p.d.s (potential differences): –17512 mV (4 cells).For both preparations the —ATP data were similar withresting p.d.s at –80 12 mV (5 ligated cells, 7 open-endcells). The excited state was more pronounced in open-end cells(resting level: –5912 mV, 5 cells) than in ligated cells(resting level: –6512 mV, 7 cells). In open-end cellsthe pump responded faster to changes of ATP concentration thanthe excitation channels. The cells stabilized with Pb(NO₃)₂were strongly depolarized both with ATP: –8010 mV (6cells) and without: 010 mV (6 cells). Most model parametersdiffered from those in the intact cells. The excited state wasabolished. Key words: Intracellular perfusion, current-voltage characteristics, Chara, stabilization with Pb(NO₃)₂, ATP effects, voltage clamp techniques