X-RAY MICROANALYSIS IN CELL BIOLOGY AND CELL PATHOLOGY

Electron probe X-ray microanalysis has been used for the last 25 years by biologists to obtain information about the distribution of elements at the cell and tissue level. During this period, progress has mainly been made through the development of more adequate techniques for specimen preparation (mainly low temperature techniques) and quantitative analysis, so that accurate analysis of the physiologically important cellular ions can be carried out. Use ofin vitrosystems and cell cultures may further increase the number of problems to which X-ray microanalysis can be applied. Among the numerous applications of X-ray microanalysis in cell biology and cell pathology, applications in the areas of epithelial ion transport, the role of calcium in secretory and contractile cells, and the role of ions in cell proliferation and cancer are discussed in more detail.     

Influence of land use on stream ecosystem function in a Mediterranean catchment

1. Due to the hierarchical organization of stream networks, land use changes occurring at larger spatial scales (i.e. the catchment) can affect physical, chemical and biological characteristics at lower spatial scales, ultimately altering stream structure and function. Anthropogenic effects on streams have primarily been documented using structural metrics such as water chemistry, channel alteration and algal biomass. Functional parameters, including metrics of nutrient retention and metabolism, are now being widely used as indicators of stream condition. 2. Within this hierarchical context, we used a multivariate approach to examine how structural and functional (i.e. nutrient retention and metabolism) attributes of streams are related to catchment variables, including land use. The study was done in 13 streams located within a single Mediterranean catchment, but draining sub‐catchments with contrasting land use. 3. At the catchment scale, results showed two contrasting land use gradients: (i) from forested‐ to urban‐dominated catchments and (ii) from low to moderate agricultural‐dominated catchments. Variation in structural and functional parameters was strongly related to these land use gradients. Specifically, NH₄⁺ demand (measured as the uptake velocity, V_f) decreased along the gradient from forested‐ to urban‐dominated catchments primarily in response to increases in stream nutrient concentrations [NH₄⁺, dissolved organic nitrogen (DON) and carbon (DOC)]. Both primary production and respiration increased along the gradient of agricultural development in response to increases in algal biomass (chlorophyll a). Soluble reactive phosphorus demand was not related to any of the land use gradients. 4. Our results illustrate the connections among factors operating at different spatial scales (i.e. from catchments to streams) and their distinct influence on stream ecosystem function. Managers should take into consideration these connections when designing stream management and restoration plans. Because ecologically successful stream management and restoration is expected to restore function as well as structure to streams, the use of appropriate measures of functional processes is required. Nutrient retention and metabolism parameters are good candidates to fill this gap.     

Stomatal responses to CO2 during a diel Crassulacean acid metabolism cycle in Kalanchoe daigremontiana and Kalanchoe pinnata

To investigate the diurnal variation of stomatal sensitivity to CO₂, stomatal response to a 30 min pulse of low CO₂ was measured four times during a 24 h time-course in two Crassulacean acid metabolism (CAM) species Kalanchoe daigremontiana and Kalanchoe pinnata , which vary in the degree of succulence, and hence, expression and commitment to CAM. In both species, stomata opened in response to a reduction in p CO₂ in the dark and in the latter half of the light period, and thus in CAM species, chloroplast photosynthesis is not required for the stomatal response to low p CO₂. Stomata did not respond to a decreased p CO₂ in K. daigremontiana in the light when stomata were closed, even when the supply of internal CO₂ was experimentally reduced. We conclude that stomatal closure during phase III is not solely mediated by high internal p CO₂, and suggest that in CAM species the diurnal variability in the responsiveness of stomata to p CO₂ could be explained by hypothesizing the existence of a single CO₂ sensor which interacts with other signalling pathways. When not perturbed by low p CO₂, CO₂ assimilation rate and stomatal conductance were correlated both in the light and in the dark in both species.     

Effects of nutrients and light on periphyton biomass and nitrogen uptake in Mediterranean streams with contrasting land uses

1. Nutrient diffusing substrata (NDS) were used to determine the relative importance of nutrients and light as potential limiting factors of periphyton biomass and nitrogen (N) uptake in Mediterranean streams subjected to different human impacts. The nutrients examined were phosphorus (P) and N, and we also further differentiated between the response of periphyton communities to N species (i.e. NO₃‐N and NH₄‐N). To examine the effect of light and nutrients on periphyton biomass, chlorophyll a accrual rates on NDS located at open and closed canopy sites were compared. The effect of nutrient availability on periphyton uptake was measured by ¹⁵N changes on the NDS after NO₃‐¹⁵N short‐term nutrient additions. 2. Results show that light was the main factor affecting algal biomass in the study streams. Algal biomass was in general higher at open than at closed canopy sites. Nutrient availability, as simulated with the NDS experiments, did not enhance algal biomass accrual in either of the 2 light conditions. 3. In the control treatments (i.e. ambient concentrations), periphyton NO₃‐N uptake rates increased and C : N molar ratios decreased consistently with increases in N availability across streams. NO₃‐N uptake rates were altered when ambient N concentrations were increased artificially in the N amended NDS. Periphyton assemblages growing on N enriched substrata seemed to preferentially take up N diffusing from the substratum rather than N from the water column. This response differed among streams, and depended on ambient N availability. 4. Periphyton biomass was not significantly different between substrata exposed to the two forms of available N sources. Nonetheless, we found differences in the effects of both N sources on the uptake of N from the water column. NH₄‐N seemed to be the preferred source of N for periphyton growing on NDS. 5. Results suggest that the effect of riparian zones on light availability, although seldom considered by water managers, may be more important than nutrients in controlling eutrophication effects derived from human activities. Finally, our results confirm that not only increases in concentration, but also stoichiometric imbalances should be considered when examining N retention in human altered streams.     

Temperature response of carbon isotope discrimination and mesophyll conductance in tobacco

The partial pressure of CO₂ at the sites of carboxylation within chloroplasts depends on the conductance to CO₂ diffusion from intercellular airspace to the sites of carboxylation, termed mesophyll conductance (g_m). We investigated the temperature response of g_m in tobacco (Nicotiana tabacum) by combining gas exchange in high light, ambient CO₂ in either 2 or 21% O₂ with carbon isotope measurements using tuneable diode laser spectroscopy. The g_m increased linearly with temperature in 2 or 21% O₂. In 21% O₂, isotope discrimination associated with g_m decreased from 5.0 ± 0.2 to 1.8 ± 0.2‰ as temperature increased from 15 to 40 °C, but the photorespiratory contribution to the isotopic signal is significant. While the fractionation factor for photorespiration (f = 16.2 ± 0.7‰) was independent of temperature between 20 and 35 °C, discrimination associated with photorespiration increased from 1.1 ± 0.01 to 2.7 ± 0.02‰ from 15 to 40 °C. Other mitochondrial respiration contributed around 0.2 ± 0.03‰. The drawdown in CO₂ partial pressure from ambient air to intercellular airspaces was nearly independent of leaf temperature. By contrast, the increase in g_m with increasing leaf temperature resulted in the drawdown in CO₂ partial pressure between intercellular airspaces and the sites of carboxylation decreasing substantially at high temperature.     

Ontogeny and structure of the drupe of Ozoroa paniculosa (Anacardiaceae)

An exocarp sensu stricto develops from the outer epidermis of the ovary wall. At maturity it comprises extensively radially elongated palisade-like parenchyma cellS. Besides having an outer cuticle, the outer tangential and outer parts of the radial cell walls of these cells are strongly cutinized. Large, permanently open stomata and saucer-shaped depressions also characterize the exocarp. The mature mesocarp sensu stricto consists of secondarily thickened parenchyma and brachysclereidS. An abundance of tanniniferous deposits and crystals, as well as secretory ducts associated with the vascular bundles also form part of the mature mesocarp. Derivatives of the inner epidermis of the ovary wall differentiate into the stratified endocarp sensu stricto. At maturity this comprises consecutive layers of macrosclereids, osteosclereids (typified by a capitate part and cell wall flutes), brachysclereids, and crystalliferous sclereidS. Pericarp structure is related to its taxonomic significance and the possible role of micromorphological characters in the survival strategy of Ozoroa paniculosa. It is shown that ontogenetic studies contribute to the precise interpretation of previously described cell layers, ensuring that homologous tissues are compared in different taxa.