Differential and reversible responses of common fen meadow species to abandonment

Abstract. We studied the effects of abandonment on two common fen plant species. In mown and a chronosequence of abandoned fen meadows spanning 35 yr, we measured fitness traits of the sedge Carex davalliana and the forb Succisa pratensis. Cessation of mowing had little effect on fitness traits and seed production of C. davalliana, but seedling density decreased more than threefold. Population density of S. pratensis decreased with increasing community biomass, but was not affected by the cessation of mowing. However, flowering frequency increased threefold and seed production was 20% higher in fallow meadows. Consequently, seedling density of S. pratensis increased nearly threefold after abandonment. However, these changes were not dependent on the age of the fallow. In a common garden and germination experiment, we found no differences in either species between plants from fallows and mown fen meadows, except for the height of the flowering stalk of S. pratensis. The combined results from the common garden experiment and the field studies indicate that changes in fitness traits observed in fallows were mostly phenotypic and likely to be reversible. If other species react in similar ways, there is a high potential for re‐establishing traditional fen meadow communities from fallows by mowing.     

Equilibrium binding characteristics of [3H]thiomuscimol.

The equilibrium binding characteristics of the tritiated GABAA agonist, 5-aminomethyl-3-isothiazolol (thiomuscimol) are described. Using the filtration technique to separate bound- from free-ligand, [3H]thiomuscimol was shown to bind to the GABA(A) receptor site(s) in a saturable manner with a Kd value of 28+/-6.0 nM and a Bmax value of 50+/-4.0 fmol/mg original tissue. In parallel binding experiments, the Kd and Bmax values for [3H]muscimol were determined to be 5.4+/-2.8 nM and 82+/-11 fmol/mg original tissue, respectively. In binding assays using the centrifugation technique, Kd and Bmax values for [3H]thiomuscimol were found to be 116+/-22 nM and 154 13 fmol/mg original tissue, respectively, whereas a Kd value of 16+/-1.8 nM and a Bmax value of 155+/-8.0 fmol/mg original tissue were determined for [3H]muscimol. In comparative inhibition studies using the GABA(A) antagonist SR 95531 and a series of specific GABAA agonists, the binding sites for [3H]thiomuscimol and [3H]muscimol were shown to exhibit similar pharmacological profiles. Autoradiographic studies disclosed similar regional distribution of [3H]thiomuscimol and [3H]muscimol binding sites in rat brain. Highest densities of binding sites were detected in cortex, hippocampus, and cerebellum, whereas low densities were measured in the midbrain structures of rat cortex. In conclusion, the equilibrium GABA(A) receptor binding characteristics of [3H]thiomuscimol are very similar to those of [3H]muscimol.     

Mid-season gas exchange of an alpine grassland under elevated CO2

Ecosystem net CO₂ uptake, evapotranspiration (ET) and night-time CO₂ efflux were measured in an alpine grassland dominated by Carex curvula, treated with doubled ambient partial pressure of CO₂ via open-top chambers. One quarter of the plots were treated with mineral nutrients to simulate the effect of lowland nitrogen deposition rates. Depending upon fertilizer supply, ecosystem net CO₂ uptake per ground area in full sunlight (NCE_max) was 41–81% higher in open-top chambers supplied with doubled ambient partial pressure (p _a) of CO₂ than in plots receiving ambient CO₂. Short-term reversals of the CO₂ level suggest that the extent of downward adjustment of canopy photosynthesis under elevated CO₂ was 30–40%. ET tended to decline, while water use efficiency (WUE), expressed as the NCE_max:ET ratio, increased more than twofold under elevated CO₂. Night-time ecosystem CO₂ efflux did not respond to changes in CO₂ p _a. NCE_max and night-time CO₂ efflux were more responsive to mineral fertilizer than the doubling of CO₂. This suggests that in these alpine plant communities, atmospheric nutrient input may induce equal or greater effects on gas exchange than increased CO₂.     

Population dynamics and spatial arrangement of Ranunculus glacialis L., an alpine perennial herb,in permanent plots

In 1986 sixteen permanent plots (625 cm² each) were established in scree slopes dominated by Ranunculus glacialis at Mt. Glungezer, Austria (2600 m elevation) in order to document the population dynamics of herbaceous perennials near the upper altitudinal limits of plant existence. The abundance and sizes of individual R. glacialis shoots, their leaf numbers and reproductive status were evaluated over a 6-year period. On South-facing slopes the population sizes of adult and juvenile shoots remained constant over the years, while seedling numbers fluctuated significantly. Overall density of all developmental stages of R. glacialis was significantly lower on North-facing slopes and year-to-year fluctuations were greater, than on thermally-favorable Southern slopes. The spatial pattern of adult shoots and seedlings was clumped, while juvenile shoots had a random or clumped distribution. Fertilization had no effects on population dynamics. Proposed greenhouse effects, e.g. increases in CO₂ and temperature, should result in population growth on North-facing slopes and may increase mortality on South-facing sites.     

Poster Sessions CP11: Neurotoxicity and Neuroprotection. Secretory phospholipase A2 signalling partly overlaps glutamate‐mediated events

Here we explored the mechanisms of secretory phospholipase A2 (sPLA2) and glutamate (glu) in neuronal signalling and cell damage. Rats or primary neuronal cultures were treated with MK‐801 and injected with/exposed to sPLA2 or glu. MK‐801 partially inhibited sPLA2‐ and glu‐induced neuronal death as well as [3H]arachidonic acid release. The involvement of cytosolic PLA2 (cPLA2) and plateletactivating factor (PAF) in sPLA2 or glu signalling was explored by treating cells with the selective cPLA2 inhibitor, AACOCF3, PAF‐acetyl hydrolase (PAF‐AH) or the presynaptic PAF‐receptor antagonist, BN52021. AACOCF3 blocked sPLA2‐ and glu‐induced neuronal death by 26 and 77%, respectively. PAF‐AH ameliorated sPLA2 as well as glu neurotoxicity by 31 and 47%, whereas BN52021 inhibited sPLA2 induced neurotoxicity by 11% but did not significantly protect against glu‐induced neurotoxicity. Expression in neurons of early response genes in response to sPLA2 or glu was further examined. An up‐regulation of COX‐2, c‐fos, and c‐jun, but not COX‐1, was observed at earlier time points after rat striatal injection of glu as compared to sPLA2 injection. Moreover we treated neuronal cells with COX‐2 inhibitors and found that neuronal cell death after sPLA2 and glu exposure was inhibited by 35 and 33%, respectively. Thus sPLA2 activates a neuronal signalling cascade that includes activation of cPLA2, AA‐release, production of PAF and induction of COX‐2. Hence sPLA2 and glu signalling are overlapping, but not identical. Cytosolic PLA2 may primarily drive glutamatergic neurotransmission, whereas PAF plays a more crucial role in sPLA2 neuronal signalling. Acknowledgements: Supported by EPSCoR grant NSF/LEQSF(2001‐04)‐RII‐01 from the National Science Foundation.     

Activation of morphogenesis and proliferation by low specificity chemical effectors

Activation of highly specific biochemical processes by simple chemical agents is demonstrated for morphogenesis (anlage and development of female gametophyte in cereal) and mitosis (in cell cultures and animal and plant tissues). The effects of these agents are tissue-specific. Structure--activity relationship is analyzed in this group of compounds. Thus, the phenomenon reveals the exact pathways of the influence of allelopathic and anthropogenic chemical agents on evolution of plant biocenoses.     

Molecular Responses of Mouse Macrophages to Copper and Copper Oxide Nanoparticles Inferred from Proteomic Analyses

The molecular responses of macrophages to copper-based nanoparticles have been investigated via a combination of proteomic and biochemical approaches, using the RAW264.7 cell line as a model. Both metallic copper and copper oxide nanoparticles have been tested, with copper ion and zirconium oxide nanoparticles used as controls. Proteomic analysis highlighted changes in proteins implicated in oxidative stress responses (superoxide dismutases and peroxiredoxins), glutathione biosynthesis, the actomyosin cytoskeleton, and mitochondrial proteins (especially oxidative phosphorylation complex subunits). Validation studies employing functional analyses showed that the increases in glutathione biosynthesis and in mitochondrial complexes observed in the proteomic screen were critical to cell survival upon stress with copper-based nanoparticles; pharmacological inhibition of these two pathways enhanced cell vulnerability to copper-based nanoparticles, but not to copper ions. Furthermore, functional analyses using primary macrophages derived from bone marrow showed a decrease in reduced glutathione levels, a decrease in the mitochondrial transmembrane potential, and inhibition of phagocytosis and of lipopolysaccharide-induced nitric oxide production. However, only a fraction of these effects could be obtained with copper ions. In conclusion, this study showed that macrophage functions are significantly altered by copper-based nanoparticles. Also highlighted are the cellular pathways modulated by cells for survival and the exemplified cross-toxicities that can occur between copper-based nanoparticles and pharmacological agents.Manufactured nanoparticles are more and more widely used in more and more consumer products, ranging from personal care products to tires and concrete. Among the nanoparticles, metals and metal oxides represent an important part of the total production and are used in water treatment, as antibacterials, in antifouling paints, and in microelectronics. These varied uses in turn pose the problem of the toxicological evaluation of these nanoparticles (1, 2), and especially of the long-term effects that often come not from simple cell mortality but from altered cellular functions.Macrophages are one of the cell types that deserve special attention in toxicology, because of the variety of their functions. Altered cytokine production can lead to adverse long-term effects, as documented, for example, in the case of asbestos (3). Other dysfunctions of the innate immune system can lead to deregulation of the immune responses and to severe adverse effects, such as a higher incidence of tumors (4).It is therefore not surprising that the immunotoxicology of nanoparticles is a developing field (5–7), and several studies have been devoted to macrophages'' response to nanoparticles. However, most of these studies have been limited to the effect of nanoparticles on cell viability and on cytokine production (e.g. 8–11), although some also studied oxidative stress (12–14) and sometimes other functional parameters (15–17). Very few studies have used the analytical power of proteomics to go deeper into the mechanisms of the response to nanoparticles or metals (reviewed in Ref. 18). A few exceptions are studies on, for example, carbon-based nanoparticles (19) and titanium dioxide (20, 21).Most of the toxicological studies in this field have been focused on a few nanoparticles used either as health products, such as iron oxide (15, 17, 22), or in a variety of consumer products, such as silver (13, 14), silica (9, 12), and titanium dioxide (11, 16, 20, 21).However, many other nanoparticles are being used more and more in industrial applications without extensive toxicological testing. Good examples are indium-tin oxide, used in electronic screens, which appears to be toxic (23), and the copper-based nanoparticles used in high-performance batteries (24), in water depollution (25), and as bactericides as a replacement for nano-silver. Copper and copper oxide induce a strong toxicity (26, 27), coupled with inflammation (28), oxidative stress (29), and genotoxicity (30), at least in epithelial cells.In light of these various effects, we decided to use a combination of a proteomics approach and targeted approaches to address in more molecular detail the responses of macrophages to copper-based nanoparticles (i.e. both metallic copper and copper II oxide).     

Cellular Origin of Ischemia-Induced Glutamate Release from Brain Tissue In Vivo and In Vitro

The uptake and release of D-[3H]aspartate (used as a tracer for endogenous glutamate and aspartate) were studied in cultured glutamatergic neurons (cerebellar granule cells) and astrocytes at normal (5 mM) or high (55 mM) potassium and under conditions of hypoglycemia, anoxia or "ischemia" (combined hypoglycemia and anoxia). In glutamatergic neurons it was found that "ischemic" conditions led to a 2.4-fold increase in the potassium-induced release of D-[3H]aspartate as compared to normal conditions. Hypoglycemia or anoxia alone affected the release only marginally. The ischemia-induced induced increase in the evoked D-[3H]aspartate release was shown to be calcium-dependent. In astrocytes no difference was found in the potassium-induced release between the four conditions and the K+-induced release was not calcium-dependent. The uptake of D-[3H]aspartate was found to be stimulated at high potassium in both glutamatergic neurons (98%) and in astrocytes (70%). This stimulation of D-aspartate uptake, however, was significantly reduced under conditions of anoxia or "ischemia" in both cell types. In glutamatergic neurons (but not in astrocytes) hypoglycemia also decreased the potassium stimulation of D-aspartate uptake. In a previous report it was shown, using the microdialysis technique, that during transient cerebral ischemia in vivo the extracellular glutamate content in hippocampus was increased eightfold. In the present paper it is shown that essentially no increase in extracellular glutamate is seen under ischemia when the perfusion is performed using calcium-free, cobalt-containing perfusion media. The results from the in vitro and in vivo experiments indicate that the glutamate accumulated extracellularly under ischemia in vivo originates from transmitter pools in glutamatergic neurons.(ABSTRACT TRUNCATED AT 250 WORDS)