Immunocytochemical Demonstration of Serotonin and Neuropeptides in the Nervous System of Gyrodactylus salaris (Monogenea)

A whole mount immunocytochemical (ICC) method has been used for the investigation of immunoreactivity (IR) to the molluscan cardioactive peptide FMRF-amide, to 9 vertebrate neuropeptides—leu-enkephalin, growth hormone-releasing factor (GRF), urotensin I, urotensin II, bovine pancreatic peptide (BPP), β-endorphin, substance P, secretin and insulin—and to the bioamine 5-HT in the nervous system (NS) of the trematode Gyrodactylus salaris belonging to the taxon Monogenea. Positive results were obtained using antisera to FMRF-amide, leu-enkephalin, urotensin I, GRF and to 5-HT. The present results are the first documentation of the presence of neuroregulatory peptides and a bioamine in the nervous system (NS) of Monogenea. Differences in the distribution pattern of the IR to the different antisera indicate that different subsets of neurons are revealed. In addition, details of the basic neuroanatomical pattern in monogeneans are confirmed by the whole mount ICC method used in this study. Negative results were obtained with antisera to urotensin II, substance P, β-endorphin, secretin, insulin and bovine pancreatic polypeptide (BPP).     

Efficiency of sunlight utilization: tubular versus flat photobioreactors

The light saturation effect imposes a serious limitation on the efficiency with which solar energy can be utilized in outdoor algal cultures. One solution proposed to reduce the intensity of incident solar radiation and overcome the light saturation effect is "spatial dilution of light" (i.e., distribution of the impinging photon flux on a greater photosynthetic surface area), but consistent experimental data supporting a significant positive influence of spatial light dilution on the productivity and the photosynthetic efficiency of outdoor algal cultures have never been reported. We used a coiled tubular reactor and compared a near-horizontal straight tubular reactor and a near-horizontal flat panel in outdoor cultivation of the cyanobacterium Arthrospira (Spirulina) platensis under defined operating conditions for optimum productivity. The photosynthetic efficiency achieved in the tubular systems was significantly higher because their curved surface "diluted" the impinging solar radiation and thus reduced the light saturation effect. This interpretation was supported by the results of experiments carried out in the laboratory under continuous artificial illumination using both a flat and a curved chamber reactor. The study also showed that, when the effect of light saturation is eliminated or reduced, productivity and solar irradiance are linearly correlated even at very high diurnal irradiance values, and supported findings that outdoor algal cultures are light-limited even during bright summer days. It was also observed that, besides improving the photosynthetic efficiency of the culture, spatial dilution of light also leads to higher growth rates and lowers the cellular content of accessory pigments; that is, it reduces mutual shading in the culture. The inadequacy of using volumetric productivity as the sole criterion for comparing reactors of different surface-to-volume ratio and of the areal productivity for evaluating the performance of elevated photobioreactors operated outdoors is stressed; it is furthermore suggested that the photosynthetic efficiency achieved by the culture also be calculated to provide a suitable parameter for comparison of different algal cultivation systems operated under similar climatic conditions. Copyright 1998 John Wiley & Sons, Inc.     

AQUEOUS POLYMER TWO-PHASE SYSTEMS AND THEIR USE IN FRAGMENTATION AND SEPARATION OF BIOLOGICAL MEMBRANES FOR THE PURPOSE OF MAPPING THE MEMBRANE STRUCTURE

When solutions of two different polymers are mixed, phase separation often occurs even at low concentrations of polymers. One polymer usually collects in one phase and the other polymer in the other phase. When water is used as solvent, two aqueous, immiscible, phases are obtained. The same holds for aqueous mixtures of a salt and a polymer. Such aqueous two-phase systems (ATPS) are very useful for separation of high-molecular-weight biomolecules such as proteins and nucleic acids and also for cells, cell organelles, and membrane vesicles. The phase systems can be made highly selective and they are also mild toward biomolecules and cell particles. In this review we describe how ATPS can be used for fragmentation and separation analyses of biological membranes and how this can be used for mapping of the photosynthetic membrane, the thylakoid, of green leaves.     

热带季雨林不同小生境大戟科植物幼树的叶片结构、耐旱性和光合能力之间的相关性

植物的叶片结构和功能性状受到自身、环境和系统发育的影响。该研究选取西双版纳20 hm²热带雨林动态监测大样地内18种分布格局不同的大戟科植物, 测量了幼树叶片的解剖结构、水分关系特征、最大光合能力和暗呼吸, 主要探讨了叶片结构对植物耐旱性和光合能力的影响, 耐旱性和光合能力之间的权衡关系, 以及环境水分条件对植物功能性状相关性的影响。结果表明: 1)生境内植物表现出一定的结构和功能的趋同性, 分布在山脊和山坡的种比沟谷种具有更强的耐失水能力; 2)去除了系统发育的影响后, 一些关键性状(特别是叶片密度和膨压丧失点时的水势、饱和渗透势等)之间存在跨生境尺度上的相关关系, 植物叶片结构同时影响了植物的耐失水能力和光合能力, 植物叶片自身的结构限制导致了植物的耐旱性(高的叶片密度、比叶质量)和光合能力(低的叶片密度、比叶质量)存在反向进化关系; 3)如果研究的植物类群亲缘关系较近, 传统的Pearson相关分析不能很好地揭示其性状间的相关关系, 因而必须采用系统发育独立对照差作相关分析。大戟科植物的结构和功能在水分梯度和光梯度上的生态位分化也从功能性状的角度为热带季雨林能维持高生物多样性, 保持植物物种长期共存提供了一个可能的解释。     

Biological applications of synchrotron radiation infrared spectromicroscopy

Extremely brilliant infrared (IR) beams provided by synchrotron radiation sources are now routinely used in many facilities with available commercial spectrometers coupled to IR microscopes. Using these intense non-thermal sources, a brilliance two or three order of magnitude higher than a conventional source is achievable through small pinholes (< 10 μm) with a high signal to-noise ratio. IR spectroscopy is a powerful technique to investigate biological systems and offers many new imaging opportunities. The field of infrared biological imaging covers a wide range of fundamental issues and applied researches such as cell imaging or tissue imaging. Molecular maps with a spatial resolution down to the diffraction limit may be now obtained with a synchrotron radiation IR source also on thick samples. Moreover, changes of the protein structure are detectable in an IR spectrum and cellular molecular markers can be identified and used to recognize a pathological status of a tissue. Molecular structure and functions are strongly correlated and this aspect is particularly relevant for imaging. We will show that the brilliance of synchrotron radiation IR sources may enhance the sensitivity of a molecular signal obtained from small biosamples, e.g., a single cell, containing extremely small amounts of organic matter. We will also show that SR IR sources allow to study chemical composition and to identify the distribution of organic molecules in cells at submicron resolution is possible with a high signal-to-noise ratio. Moreover, the recent availability of two-dimensional IR detectors promises to push forward imaging capabilities in the time domain. Indeed, with a high current synchrotron radiation facility and a Focal Plane Array the chemical imaging of individual cells can be obtained in a few minutes. Within this framework important results are expected in the next years using synchrotron radiation and Free Electron Laser (FEL) sources for spectro-microscopy and spectral-imaging, alone or in combination with Scanning Near-field Optical Microscopy methods to study the molecular composition and dynamic changes in samples of biomedical interest at micrometric and submicrometric scales, respectively.     

Microfluidics Approaches in Modern Developmental Biology

Modern automated microsystems based on microhydrodynamic (microfluidic) technologies— labs on chips—make it possible to solve various basic and applied research problems. In the last 15 years, the development of these approaches in application to the problems of modern quantitative (systems) development biology has been observed. In this field, high-throughput experiments aimed at accumulating ample quantitative data for their subsequent computer analysis are important. In this review, the main directions in the development and application of microfluidics approaches for solving problems of modern developmental biology using the classical model object, Drosophila embryo, as an example is discussed. Microfluidic systems provide an opportunity to perform experiments that can hardly be performed using other approaches. These systems allow automated, rapid, reliable, and proper placing of many live embryos on a substrate for their simultaneous confocal scanning, sorting them, or injecting them with various agents. Such systems make it possible, in particular, to create controlled gradients of microenvironmental parameters along a series of developing embryos or even to introduce discontinuity in parameters within the microenvironment of one embryo, so that the head half is under other conditions compared to the tail half (at continuous scanning). These approaches are used both in basic research of the functions of gene ensembles that control early development, including the problems of resistance of early patterns to disturbances, and in test systems for screening chemical agents on developing embryos. The problems of integration of microfluidic devices in systems for automated performance of experiments simultaneously on many developing embryos under conditions of their continuous scanning using modern fluorescence microscopy instruments will be discussed. The methods and approaches developed for Drosophila are also applicable to other model objects, even mammalian embryos.     

Comparative ecophysiology of leaf and canopy photosynthesis

Leaves and herbaceous leaf canopies photosynthesize efficiently although the distribution of light, the ultimate resource of photosynthesis, is very biased in these systems. As has been suggested in theoretical studies, if a photosynthetic system is organized such that every photosynthetic apparatus photosynthesizes in concert, the system as a whole has the sharpest light response curve and is most adaptive. This condition can be approached by (i) homogenization of the light environment and (ii) acclimation of the photosynthetic properties of leaves or chloroplasts to their local light environments. This review examines these two factors in the herbaceous leaf canopy and in the leaf. Changes in the inclination of leaves in the canopy and differentiation of mesophyll into palisade and spongy tissue contribute to the moderation of the light gradient. Leaf and chloroplast movements in the upper parts of these systems under high irradiances also moderate light gradients. Moreover, acclimation of leaves and chloroplasts to the local light environment is substantial. These factors increase the efficiency of photosynthesis considerably. However, the systems appear to be less efficient than the theoretical optimum. When the systems are optically dense, the light gradients may be too great for leaves or chloroplasts to acclimate. The loss of photosynthetic production attributed to the imperfect adjustment of photosynthetic apparatus to the local light environment is most apparent when the photosynthesis of the system is in the transition between the light-limited and light-saturated phases. Although acclimation of the photosynthetic apparatus and moderation of light gradients are imperfect, these markedly raise the efficiency of photosynthesis. Thus more mechanistic studies on these adaptive attributes are needed. The causes and consequences of imperfect adjustment should also be investigated.     

Comparative genomic analysis illustrates evolutionary dynamics of multisubunit tethering complexes across green algal diversity

The chlorophyte algae are a dominant group of photosynthetic eukaryotes. Although many are photoautotrophs, there are also mixotrophs, heterotrophs, and even parasites. The physical characteristics of green algae are also highly diverse, varying greatly in size, shape, and habitat. Given this morphological and trophic diversity, we postulated that diversity may also exist in the protein components controlling intracellular movement of material by vesicular transport. One such set is the multisubunit tethering complexes (MTCs)—components regulating cargo delivery. As they span endomembrane organelles and are well-conserved across eukaryotes, MTCs should be a good proxy for assessing the evolutionary dynamics across the diversity of Chlorophyta. Our results reveal that while green algae carry a generally conserved and unduplicated complement of MTCs, some intriguing variation exists. Notably, we identified incomplete sets of TRAPPII, exocyst, and HOPS/CORVET components in all Mamiellophyceae, and what is more, not a single subunit of Dsl1 was found in Cymbomonas tetramitiformis. As the absence of Dsl1 has been correlated with having unusual peroxisomes, we searched for peroxisome biogenesis machinery, finding very few components in Cymbomonas, suggestive of peroxisome degeneration. Overall, we demonstrate conservation of MTCs across green algae, but with notable taxon-specific losses suggestive of unusual endomembrane systems.     

Imaging of chlorophyll a fluorescence: theoretical and practical aspects of an emerging technique for the monitoring of photosynthetic performance

The development of chlorophyll (Chl) a fluorescence imaging systems has greatly increased the versatility of Chl a fluorometry as a non-invasive technique for the investigation of photosynthesis in plants and algae. For example, systems that image at the microscopic level have made it possible to measure PSII photochemical efficiencies from chloroplasts within intact leaves and from individual algal cells within mixed populations, while systems that image over much larger areas have been used to investigate heterogeneous patterns of photosynthetic performance across leaves and in screening programmes that image tens or even hundreds of plants simultaneously. In addition, it is now practical to use fluorescence imaging systems as real-time, multi-channel fluorometers, which can be used to record continuous fluorescence traces from multiple leaves, plants, or algal cells. This paper discusses some of the theoretical and practical issues associated with the imaging of Chl a fluorescence and with Chl a fluorometry in general. This discussion includes a review of the most commonly used Chl a fluorescence parameters.     

Direct and indirect climate change effects on photosynthesis and transpiration

Climate change affects plants in many different ways. Increasing CO(2) concentration can increase photosynthetic rates. This is especially pronounced for C(3) plants, at high temperatures and under water-limited conditions. Increasing temperature also affects photosynthesis, but plants have a considerable ability to adapt to their growth conditions and can function even at extremely high temperatures, provided adequate water is available. Temperature optima differ between species and growth conditions, and are higher in elevated atmospheric CO(2). With increasing temperature, vapour pressure deficits of the air may increase, with a concomitant increase in the transpiration rate from plant canopies. However, if stomata close in response to increasing CO(2) concentration, or if there is a reduction in the diurnal temperature range, then transpiration rates may even decrease. Soil organic matter decomposition rates are likely to be stimulated by higher temperatures, so that nutrients can be more readily mineralised and made available to plants. This is likely to increase photosynthetic carbon gain in nutrient-limited systems. All the factors listed above interact strongly so that, for different combinations of increases in temperature and CO(2) concentration, and for systems in different climatic regions and primarily affected by water or nutrient limitations, photosynthesis must be expected to respond differently to the same climatic changes.     

植物气孔扩散传导率的研究

本文对生长在可控环境温室中的花生气孔扩散传导率进行了实验研究,揭示了单个植株之间、上下表面之间、叶片不同部位以及冠层垂直方向上气孔扩散传导率的变异性。同时以气孔扩散传导率与环境条件的测定值为基础,对传导率对环境因子的反应进行了分析,植株顶部叶片气孔扩散传导率与太阳总辐射和空气饱和差有关系;冠层传导率与冠层截留辐射和空气饱和差有相关关系。     

Changes in photosynthetic activity of the diatom Phaeodactylum tricornutum in a culture of limited volume

Changes in photosynthetic activity of a marine diatom duringalgal growth were studied with a typical culture medium formarine algae, ASP-2 (l). As the algal specimen, Phaeodactylumtricornutum was used for the experiments. Nitzschia closteriumand Chaetoceros sp. were also supplementarily used. Photosynthetic and p-benzoquinone Hill activities remarkablychanged with time during algalgrowth; with maximum activityfound in cells at log phase. A rapid decrease occurred in theinterphase from the log to stationary phase. The activity changewas not accompanied by variation in photosynthetic pigment content. The low concentration of phosphorus source was suggested asthe main cause for the change. On supply of extra inorganicphosphate, the time length for holding high photosynthetic activitybecame longer; or, the activity of the cells at stationary phaserecovered at least partly even in the dark. Dark recovery wasnot accompanied by either algal growth or an increase in thecontent of photosynthetic pigments. Inactivation of photosynthesis in the stationary growth phaseand activation by added phosphate in the dark were inferredto be due to changes in activities of both the CO₂-fixing andelectron transfer systems. The observed activity change maynot be attributable to a deficiency in inorganic phosphate asthe substrate for photophosphorylation. Similar changes in photosynthetic activity were also observedwith Nitzschia closterium and Chaetoceros sp. (Received January 30, 1971; )     

The evaluation of photosynthetic parameters in maize inbred lines subjected to water deficiency: Can these parameters be used for the prediction of performance of hybrid progeny?

The response of selected photosynthetic and morphological parameters of plants to drought was examined in 5 inbred lines of maize (Zea mays L.) and their 10 F1 hybrids. The aim of the study was to establish whether the photosynthetic performance of parental genotypes under drought conditions correlates with the performance of their progeny and whether the net photosynthetic rate, the chlorophyll fluorescence parameters or the content of photosynthetic pigments could be used as reliable physiological markers for early breeding generations. The relative importance of the additive and the nonadditive (dominance, maternal) genetic effects in the inheritance of these parameters was also assessed by means of the quantitative genetics analysis. The results showed that the nonadditive genetic effects associated with a particular combination of genotypes or a particular direction of crossing are at least equally and often even more important as the additivity and that these genetic effects almost totally change with the exposure of plants to drought conditions. This was reflected in the inability to predict the response of F1 hybrids to drought on the basis of the photosynthetic performance of their parents, which indicates that the practical usability of such parameters in maize breeding programs is rather limited.     

Abschätzung von Turbulenzkorrekturen für die phytoplanktische O2-Produktion bei schwacher Turbulenz. Estimation of Turbulence Correction Terms for the Phytoplankton Production (O2) Under Conditions of Low Turbulence

The turbulence correction terms for the photosynthetic oxygen production of phytoplankton given by BAUMERT (1974) are used for the case that the variations in velocity are exponentially autocorrelated. In case of the P(I)-relation by STEEMANN NIELSEN one can show that the value of the relative correction term is — in the worst case — less than, or equal to 1% if the depth of epilimnion (mixing depth) and the light intensity at the water surface are high. The demand for weak turbulence and light extinction — necessary for validity of the equations — is met for some real objects, especially for oligotrophic seas and lakes.     

Monitoring the pH dependence of IR carboxylic acid signals upon Q(B)- formation in the Glu-L212 --> Asp/Asp-L213 --> Glu swap mutant reaction center from Rhodobacter sphaeroides

In the photosynthetic reaction center (RC) from the purple bacterium Rhodobacter sphaeroides, proton-coupled electron-transfer reactions occur at the secondary quinone (QB) site. Involved in the proton uptake steps are carboxylic acids, which have characteristic infrared vibrations in the 1770-1700 cm-1 spectral range that are sensitive to 1H/2H isotopic exchange. With respect to the native RC, a novel protonation pattern for carboxylic acids upon QB photoreduction has been identified in the Glu-L212 --> Asp/Asp-L213 --> Glu mutant RC using light-induced FTIR difference spectroscopy (Nabedryk, E., Breton, J., Okamura, M. Y., and Paddock, M. L. (2004) Biochemistry 43, 7236-7243). These carboxylic acids are structurally close and have been implicated in proton transfer to reduced QB. In this work, we extend previous studies by measuring the pH dependence of the QB-/QB FTIR difference spectra of the mutant in 1H2O and 2H2O. Large pH dependent changes were observed in the 1770-1700 cm-1 spectral range between pH 8 and pH 4. The IR fingerprints of the protonating carboxylic acids upon QB- formation were obtained from the calculated double-difference spectra 1H2O minus 2H2O. These IR fingerprints are specific for each pH, indicative of the contribution of different titrating groups. In particular, the 1752 cm-1 signal indicates that Glu-L213 protonates upon QB- formation at pH >or= 5, whereas the 1746 cm-1 signal indicates protonation of Asp-L212 even at pH 4. An unidentified carboxylic acid absorbing at approximately 1765 cm-1 could be the proton donor between pH 8 and 5. The observation that in the swap mutant there are several uniquely behaving carboxylic acids shows that electrostatic interactions occurring between them are sufficiently modified from the native RC to reveal their IR signatures.     

Translation of Rabbit Globin mRNA

IN certain conditions bacterial initiation factors can discriminate between different classes of mRNA, or the individual cistrons of bacteriophage RNA^1–5. Whether messenger specificity is a general phenomenon in eukaryotes as well is controversial, but several reports support the idea that mRNAs can be translated in heterologous systems without the addition of homologous tissue-specific factors, suggesting a lack of specificity^6,10,17. These results have been obtained by adding mRNA either to oocytes⁷ or to crude cell-free systems such as lysates or 30,000g supernatants (S-30). With more fractionated systems—separated ribosomes reconstituted with cell sap—the existence of messenger specific factors is supported^11–13. We have already demonstrated that an S-30 derived from either mouse or rat liver can faithfully translate rabbit globin mRNA without the addition of reticulocyte factors and now report that fractionation of this S-30 using methods similar to those used by previous workers^11–13 results in a system which will also translate globin mRNA. This fractionated system does not show a requirement for reticulocyte specific factors.     

Migration of electron excitation energy in mixed associates of chlorophyll and its derivatives]

The intracomplex migration is investigated of electron excitation energy between molecules which are a part of a mixed associate. Weakly fluorescent and nonfluorescent associates of chlorophyll and its analogues as well as of bacteriochlorophyll and neoxanthine have been used as donors and acceptors. The migration of excitation energy with variation of quantum yield of donor emission from 5-10(-3) to less than or equal to 10(-4) and also with very weak overlapping of luminescence and absorption spectra for the pair D leads to A is experimentally discovered by luminescence excitation spectra of mixed pigment aggregates. A specific disturbing action of Pd and Cu ions on chlorophyll a luminescence with formation of mixed complexes of chlorophyll with Pd-pheophytin a and Cu-pheophytin b is observed. The presence of Pd and Cu ions in the structure of the mixed complexes gives rise to a considerable reduction of chlorophyll apparently due to increasing intersystem crossing probability in the chlorophyll molecule. The probability of intracomplex energy migration in systems under investigation amounts to approximately 10(12) sec-1 according to the obtained estimations. The probable mechanism of energy migration in mixed associates is discussed. The obtained results may be used for elucidation of a possible role of the mixed aggregates and effective intracomplex migration of excitation energy in photosynthetic apparatus of green plants.     

Thermal acclimation of photosynthesis: on the importance of adjusting our definitions and accounting for thermal acclimation of respiration

While interest in photosynthetic thermal acclimation has been stimulated by climate warming, comparing results across studies requires consistent terminology. We identify five types of photosynthetic adjustments in warming experiments: photosynthesis as measured at the high growth temperature, the growth temperature, and the thermal optimum; the photosynthetic thermal optimum; and leaf-level photosynthetic capacity. Adjustments of any one of these variables need not mean a concurrent adjustment in others, which may resolve apparently contradictory results in papers using different indicators of photosynthetic acclimation. We argue that photosynthetic thermal acclimation (i.e., that benefits a plant in its new growth environment) should include adjustments of both the photosynthetic thermal optimum (T _opt) and photosynthetic rates at the growth temperature (A _growth), a combination termed constructive adjustment. However, many species show reduced photosynthesis when grown at elevated temperatures, despite adjustment of some photosynthetic variables, a phenomenon we term detractive adjustment. An analysis of 70 studies on 103 species shows that adjustment of T _opt and A _growth are more common than adjustment of other photosynthetic variables, but only half of the data demonstrate constructive adjustment. No systematic differences in these patterns were found between different plant functional groups. We also discuss the importance of thermal acclimation of respiration for net photosynthesis measurements, as respiratory temperature acclimation can generate apparent acclimation of photosynthetic processes, even if photosynthesis is unaltered. We show that while dark respiration is often used to estimate light respiration, the ratio of light to dark respiration shifts in a non-predictable manner with a change in leaf temperature.     

光合作用对光和二氧化碳响应的观测方法探讨

用便携式光合仪LI-6400观测自然条件下生长的盆栽蚕豆叶片光合作用对光和二氧化碳的响应发现：（1）用未经过光合诱导的叶片观测光合作用对光的响应会得到即使在全太阳光强下光合作用仍然不饱和的假象;（2）利用某些经验方程计算的饱和光强远低于实际观测值;（3）在观测光合作用对CO2的响应过程中,每一次CO2浓度变化都应当伴随一次光合仪的匹配步骤,否则所得结果偏差很大;（4）在不饱和光下观测光合作用对CO2的响应,会导致对叶片光合能力的低估。