Organellar Calcium Buffers

Ca²⁺ is an important intracellular messenger affecting many diverse processes. In eukaryotic cells, Ca²⁺ storage is achieved within specific intracellular organelles, especially the endoplasmic/sarcoplasmic reticulum, in which Ca²⁺ is buffered by specific proteins known as Ca²⁺ buffers. Ca²⁺ buffers are a diverse group of proteins, varying in their affinities and capacities for Ca²⁺, but they typically also carry out other functions within the cell. The wide range of organelles containing Ca²⁺ and the evidence supporting cross-talk between these organelles suggest the existence of a dynamic network of organellar Ca²⁺ signaling, mediated by a variety of organellar Ca²⁺ buffers.     

The occurrence of the freshwater isopod Heterias (Fritzianira) exul in the Lakes Region of Chile,with notes on the genus Heterias (Asellota: Janiridae)

Heterais exul, previously known from Brazil and Argentina, was collected from a number of lakes and streams in the Lakes Region of Chile (between Valdivia and Puerto Montt). It was found in a wide variety of habitats: on leaf litter, detritus, algae, rotting wood, emergent vegetation, and clean sandy bottoms. Heterias, Pseudasellus, and Fritzianira are placed in synonymy, but Fritzianira is maintained as a subgenus of Heterias because of its lack of an antenna) scale. The 3 Australian species of Heterias and H. exul are believed to have evolved from a common freshwater Gondwanaland ancestor.     

Light-Induced Polar pH Changes in Leaves of Elodea canadensis: I. Effects of Carbon Concentration and Light Intensity

Leaves of the submerged aquatic Elodea canadensis Michx. exhibit a light induced polar pH reaction. In this study, the effects of light intensity and dissolved inorganic carbon concentration on this polar reaction were examined. At a light intensity of 100 watts per square meter the leaf showed a polar pH response when the dissolved inorganic carbon concentration was less than about 1 millimolar. The polar reaction was suppressed at a higher dissolved inorganic carbon concentration. This suppression was not due to the buffering capacity of bicarbonate. Because another weak acid, acetate, did not inhibit the polarity, but even had a small stimulatory effect, the effect of bicarbonate is also not due to acidification of the cytoplasm. The suppression of the polar reaction by CO₂/HCO₃⁻ was relieved when the light intensity was increased. Apparently there is competition for product(s) of the photosynthetic light reactions between processes generating the polar reaction and the carbon fixation reactions. The possibility that the redox state of the cell regulates the generation of the polar reaction is discussed.     

Thermophiles: A life at elevated temperatures

Interest in the ecology, physiology and evolution of microorganisms adapted to growth at relatively high temperatures (up to 110°C) has increased enormously during the past two decades. This interest was stimulated by the discovery of marine hydrothermal vent ecosystems, and also by awareness of the potential of thermophilic microbes in biotechnological processes. Subsequent attempts to isolate new thermophilic organisms have been very successful. Moreover, these results, in combination with much-improved techniques for studying the phylogeny of microorganisms, have renewed interest in the evolution of microbes and the early development of life.     

Electrophysiological membrane characteristics of the salt tolerant Plantago maritima and the salt sensitive Plantago media

Plasmid pUCD607 was mobilized into the biocontrol agent Enterobacter cloacae strain E6 by conjugation and the resultant strain, E6(pUCD607), was bioluminescent. Biocontrol of Pythium ultimum by E6(pUCD607) was similar to that of the parent strain, E6. The location of E6(pUCD607) in the soil and in the rhizosphere of lettuce was readily determined by pressing agar medium against plant roots in a root box, allowing the bacteria to grow overnight on the medium, and detecting the presence of bioluminescence by autophotography. There was a positive, linear correlation between population sizes determined by dilution plating and the quantity of light emitted due to bioluminescence. However, both the intercept and slope of this line varied among experiments possibly due to the differing physiological states of cells recovered from soils. The amount of light emitted by the bioluminescent strain E6(pUCD607) was not quantitative. This technique is useful for qualitative determinations of populations and for photographically locating bacteria.     

Patch clamp studies on root cell vacuoles of a salt-tolerant and a salt-sensitive plantago species : regulation of channel activity by salt stress

Plantago media L. and Plantago maritima L. differ in their strategy toward salt stress, a major difference being the uptake and distribution of ions. Patch clamp techniques were applied to root cell vacuoles to study the tonoplast channel characteristics. In both species the major channel found was a 60 to 70 picosiemens channel with a low ion selectivity. The conductance of this channel for Na⁺ was the same as for K⁺, P_K⁺/P_Na⁺ = 1, whereas the cation/anion selectivity (P_K⁺/P_c1⁻) was about 5. Gating characteristics were voltage and calcium dependent. An additional smaller channel of 25 picosiemens was present in P. maritima. In the whole vacuole configuration, the summation of the single channel currents resulted in slowly activated inward currents (t^½ = 1.2 second). Inwardly directed, ATP-dependent currents could be measured against a ΔpH gradient of 1.5 units over the tonoplast. This observation strongly indicated the physiological intactness of the used vacuoles. The open probability of the tonoplast channels dramatically decreased when plants were grown on NaCl, although single channel conductance and selectivity were not altered.     

Utilization of Organic Nitrogen Sources by Two Phytoplankton Species and a Bacterial Isolate in Pure and Mixed Cultures

Algal production of dissolved organic carbon and the regeneration of nutrients from dissolved organic carbon by bacteria are important aspects of nutrient cycling in the sea, especially when inorganic nitrogen is limiting. Dissolved free amino acids are a major carbon source for bacteria and can be used by phytoplankton as a nitrogen source. We examined the interactions between the phytoplankton species Emiliania huxleyi and Thalassiosira pseudonana and a bacterial isolate from the North Sea. The organisms were cultured with eight different amino acids and a protein as the only nitrogen sources, in pure and mixed cultures. Of the two algae, only E. huxleyi was able to grow on amino acids. The bacterium MD1 used all substrates supplied, except serine. During growth of MD1 in pure culture, ammonium accumulated in the medium. Contrary to the expectation, the percentage of ammonium regenerated from the amino acids taken up showed no correlation with the substrate C/N ratio. In mixed culture, the algae grew well in those cultures in which the bacteria grew well. The bacterial yields (cell number) were also higher in mixed culture than in pure culture. In the cultures of MD1 and T. pseudonana, the increase in bacterial yield (number of cells) over that of the pure culture was comparable to the bacterial yield in mixed culture on a mineral medium. This result suggests that T. pseudonana excreted a more-or-less-constant amount of carbon. The bacterial yields in mixed cultures with E. huxleyi showed a smaller and less consistent difference than those of the pure cultures of MD1. It is possible that the ability of E. huxleyi to use amino acids influenced the bacterial yield. The results suggest that interactions between algae and bacteria influence the regeneration of nitrogen from organic carbon and that this influence differs from one species to another.     

Simulation of the light-induced oscillations of the membrane potential in Potamogeton leaf cells

Summary An attempt has been made to simulate the light-induced oscillations of the membrane potential of Potamogeton lucens leaf cells in relation to the apoplastic pH changes. Previously it was demonstrated that the membrane potential of these cells can be described in terms of proton movements only. It is hypothesized that the membrane potential is determined by an electrogenic H⁺-ATPase with a variable H⁺/ATP stoichiometry. The stoichiometry shifts from a value of two in the dark to a value of one in the light. Moreover, this H⁺ pump shows the characteristics of either a pump or a passive H⁺ conductance: the mode of operation of the H⁺ translocator is considered to be regulated by the external pH. The pump conductance is assumed to be dominant at low or neutral pH, while the passive H⁺ conductance becomes more significant at alkaline pH. The pH dependence of the transport characteristic is expressed by protonation reactions in the plasma membrane. The proposed model can account for most features of the light-induced oscillations but not for the absolute level of the membrane potential.This research was supported by the Foundation of Biophysics, part of the Dutch Organization for Scientific Research (NWO) ECOTRANS publication No. 34.     

Reflection contrast microscopy of ultrathin sections in immunocytochemical localization studies: a versatile technique bridging electron microscopy with light microscopy

Reflection contrast microscopy (RCM) of ultrathin sections was recently introduced as a sensitive technique for visualization with enhanced definition in immunogold histochemistry. Experience of using RCM as a major tool in immunocytochemical research in different fields is summarized, e.g. oncology, nephrology and embryology. The sensitive visualization of immunocytochemical labels, gold particles or peroxidase-diaminobenzidine deposits in or on ultrathin sections, by RCM instead of electron microscopy is demonstrated. RCM of ultrathin sections is an adequate light microscopical alternative for immunoelectron microscopy, since an overview of both label and tissue is obtained with a high image definition and high contrast of label. In the studies presented, RCM is shown to provide a better gradation in staining intensity and staining pattern than other light microscopical methods. Moreover, a precise localization of multiple labels is obtained with this method. Besides the applications shown, ultrathin section visualization by RCM is very useful for correlative light- and electron microscopical studies of fine structures. Commercially available fluorescence microscopes can be adapted for proper RCM functioning; an adaptation scheme and list of microscopes tested is provided.