A Comprehensive Metabolic Profile of Cultured Astrocytes Using Isotopic Transient Metabolic Flux Analysis and 13C-Labeled Glucose

Metabolic models have been used to elucidate important aspects of brain metabolism in recent years. This work applies for the first time the concept of isotopic transient 13C metabolic flux analysis (MFA) to estimate intracellular fluxes in primary cultures of astrocytes. This methodology comprehensively explores the information provided by 13C labeling time-courses of intracellular metabolites after administration of a 13C-labeled substrate. Cells were incubated with medium containing [1-13C]glucose for 24 h and samples of cell supernatant and extracts collected at different time points were then analyzed by mass spectrometry and/or high performance liquid chromatography. Metabolic fluxes were estimated by fitting a carbon labeling network model to isotopomer profiles experimentally determined. Both the fast isotopic equilibrium of glycolytic metabolite pools and the slow labeling dynamics of TCA cycle intermediates are described well by the model. The large pools of glutamate and aspartate which are linked to the TCA cycle via reversible aminotransferase reactions are likely to be responsible for the observed delay in equilibration of TCA cycle intermediates. Furthermore, it was estimated that 11% of the glucose taken up by astrocytes was diverted to the pentose phosphate pathway. In addition, considerable fluxes through pyruvate carboxylase [PC; PC/pyruvate dehydrogenase (PDH) ratio = 0.5], malic enzyme (5% of the total pyruvate production), and catabolism of branched-chained amino acids (contributing with ∼40% to total acetyl-CoA produced) confirmed the significance of these pathways to astrocytic metabolism. Consistent with the need of maintaining cytosolic redox potential, the fluxes through the malate–aspartate shuttle and the PDH pathway were comparable. Finally, the estimated glutamate/α-ketoglutarate exchange rate (∼0.7 μmol mg prot−1 h−1) was similar to the TCA cycle flux. In conclusion, this work demonstrates the potential of isotopic transient MFA for a comprehensive analysis of energy metabolism

Influence of a breakwater on nearby rocky intertidal community structure

Copyright © 2009 Elsevier Ltd. All rights reserved.It is widely recognised that coastal-defence structures generally affect the structure of the assemblages they support, yet their impact on adjacent systems has been largely ignored. Breakwaters modify the nearby physical environment (e.g. wave action) suggesting a local impact on biological parameters. In the present study, an ACI (After-Control-Impact) design was used to test the general hypothesis that the artificial sheltering of an exposed coast has a strong effect on the structure and functioning of adjacent systems. The effects of a reduction in hydrodynamics were clear for a number of taxa and included the replacement of barnacles, limpets and frondose algae by an increasing cover of ephemeral algae. These effects were evident both at early and late successional stages. Results suggest that the artificial sheltering of naturally exposed coasts can have a strong impact promoting a shift from consumer- to producer-dominated communities, which has important ecological and energetic consequences for the ecosystem

Limpets (Patella and Haliotis) aquaculture in the Azores.

43rd European Marine Biology Symposium. Ponta Delgada, Açores, 8-12 de Setembro de 2008

Image analysis as a tool for viability and recombinant protein production assessment during E. coli fermentations

The development of monitoring methods for physiological state assessment during recombinant fermentation processes has been encouraged by the need to evaluate the influence of processing conditions in recombinant protein production. In this work, microscopy and image analysis techniques were used for the quantification of viability and protein production in two recombinant E. coli batch fermentations. Images obtained from light microscopy with phase contrast were used to assess the total number of cells in a given sample and, from epifluorescence microscopy, both producing and dead cells were counted using two different filters. This methodology allowed the extraction of information related to cell viability and recombinant protein production. This information, combined with standard fermentation data, allowed the derivation of interesting hypothesis that can be used afterwards for experimental design and further validation. Additionally, the ratios calculated in this work can be complemented with other parameters that can be extracted from image analysis

Aquaculture of the clam (Tapes decussatus) on a closed system.

43rd European Marine Biology Symposium. Ponta Delgada, Açores, 8-12 de Setembro de 2008

The use of digital photography for the definition of coastal biotopes in Azores.

Copyright © Springer Science+Business Media B.V. 2007.Sampling benthic communities usually requires intensive field and lab work which is generally performed by skilled staff. In algal dominated communities, like those on the shores of the Azores, biotope characterization studies focused on the more conspicuous algae categories, thus reducing the skills required for species identification. The present study compares in situ quadrat quantifications done by a skilled reader, with computer based quadrat quantifications using digital photographic records of the same areas read in situ, accomplished by skilled and non-skilled readers. The study was conducted inter- and subtidally at various shore heights/depths. Quantification of algal coverage, both in situ and computer based, used the point to point method with quadrats of 0.25 m × 0.25 m for the intertidal, and 0.50 m × 0.50 m for the subtidal surveys, both subdivided into 36 intersection points. Significant differences were found between in situ readings and computer based readings of photographic records conducted both by experienced and inexperienced readers. Biotopes identified using in situ data and image based data differ both for the subtidal and intertidal

Chemical strategies for dendritic magneto-plasmonic nanostructures applied to surface-enhanced Raman spectroscopy

Chemical analyses in the field using surface-enhanced Raman scattering (SERS) protocols are expected to be part of several analytical procedures applied to water quality monitoring. To date, these endeavors have been supported by developments in SERS substrate nanofabrication, instrumentation portability, and the internet of things. Here, we report distinct chemical strategies for preparing magneto-plasmonic (Fe3 O4  : Au) colloids, which are relevant in the context of trace-level detection of water contaminants due to their inherent multifunctionality. The main objective of this research is to investigate the role of poly(amidoamine) dendrimers (PAMAMs) in the preparation of SERS substrates integrating both functionalities into single nanostructures. Three chemical routes were investigated to design magneto-plasmonic nanostructures that translate into different ways for assessing SERS detection by using distinct interfaces. Hence, a series of magneto-plasmonic colloids have been characterized and then assessed for their SERS activity by using a model pesticide (thiram) dissolved in aqueous samples.publishe

Oligodendrocytes Do Not Export NAA-Derived Aspartate In Vitro.

Oligodendroglial cells are known to de-acetylate the N-acetylaspartate (NAA) synthesized and released by neurons and use it for lipid synthesis. However, the role of NAA regarding their intermediary metabolism remains poorly understood. Two hypotheses were proposed regarding the fate of aspartate after being released by de-acetylation: (1) aspartate is metabolized in the mitochondria of oligodendrocyte lineage cells; (2) aspartate is released to the medium. We report here that aspartoacylase mRNA expression increases when primary rat oligodendrocyte progenitor cells (OPCs) differentiate into mature cells in culture. Moreover, characterising metabolic functions of acetyl coenzyme A and aspartate from NAA catabolism in mature oligodendrocyte cultures after 5 days using isotope-labelled glucose after 5-days of differentiation we found evidence of extensive NAA metabolism. Incubation with [1,6-13C]glucose followed by gas chromatography-mass spectrometry and high performance liquid chromatography analyses of cell extracts and media in the presence and absence of NAA established that the acetate moiety produced by hydrolysis of NAA does not enter mitochondrial metabolism in the form of acetyl coenzyme A. We also resolved the controversy concerning the possible release of aspartate to the medium: aspartate is not released to the medium by oligodendrocytes in amounts detectable by our methods. Therefore we propose that: aspartate released from NAA joins the cytosolic aspartate pool rapidly and takes part in the malate-aspartate shuttle, which transports reducing equivalents from glycolysis into the mitochondria for ATP production and enters the tricarboxylic acid cycle at a slow rate.This work was supported by grants from the UK Multiple Sclerosis Society and from Qatar Foundation. The work was further supported by core funding from the Wellcome Trust and MRC to the Wellcome Trust-Medical Research Council Cambridge Stem Cell Institute. The authors acknowledge the excellent technical support in GC-MS and HPLC analysis from Lars Evje (NTNU, Norway).This is the final version of the article. It first appeared from Springer at http://dx.doi.org/10.1007/s11064-016-1985-y

Long-term TNT and DNT contamination: 1-D modeling of natural attenuation in the vadose zone: case study, Portugal

The vadose zone of a trinitrotoluene (TNT) and dinitrotoluene (DNT) contaminated site was investigated to assess the mobility of those explosives under natural conditions. Located in the left margin of the River Tejo Basin, Portugal, the site is located on unconsolidated sediments. Wastewaters associated with the 50-year explosives production were disposed in excavated ponds, from where water would infiltrate and pollute the unsaturated and saturated parts of the local aquifers. Two boreholes were drilled to 9 m depth in such a former waste pond to investigate the contaminant's fate in the vadose zone. Sediment samples were taken every 1-2 m for analysis of the polynitroaromatics (p-NACs) and organic volatile compounds, pH, organic carbon content, cation exchange capacity and grain size analysis. The main contaminant was TNT representing >70 % of the total p-NACs concentration that peaked approximately 7 mg/kg in one borehole, even if the median in both boreholes was of similar to 1 mg/kg. DNT was 4-30 % of the total p-NACs and nitrotoluene (NT), up to 5 %. No other (volatile) organic compound was detected. The predominance of TNT as the main contaminant implies that any natural mass reduction has been inefficient to clean the site. Several 1-D model simulations of p-NACs cleaning of the vadose zone under natural conditions indicated that the most probable scenario of combined advection and partitioning will only remove TNT after 10's of years, whereas DNT and NT will hardly be removed. Such low concentrations and long times for the p-NACs removal, suggest that by now those compounds have been washed-out to a level below standard limits

Consistent patterns of common species across tropical tree communities

D.L.M.C. was supported by the London Natural Environmental Research Council Doctoral Training Partnership grant (grant no. NE/L002485/1). This paper developed from analysing data from the African Tropical Rainforest Observatory Network (AfriTRON), curated at ForestPlots.net. AfriTRON has been supported by numerous people and grants since its inception. We sincerely thank the people of the many villages and local communities who welcomed our field teams and without whose support this work would not have been possible. Grants that have funded the AfriTRON network, including data in this paper, are a European Research Council Advanced Grant (T-FORCES; 291585; Tropical Forests in the Changing Earth System), a NERC standard grant (NER/A/S/2000/01002), a Royal Society University Research Fellowship to S.L.L., a NERC New Investigators Grant to S.L.L., a Philip Leverhulme Award to S.L.L., a European Union FP7 grant (GEOCARBON; 283080), Leverhulme Program grant (Valuing the Arc); a NERC Consortium Grant (TROBIT; NE/D005590/), NERC Large Grant (CongoPeat; NE/R016860/1) the Gordon and Betty Moore Foundation the David and Lucile Packard Foundation, the Centre for International Forestry Research (CIFOR), and Gabon’s National Parks Agency (ANPN). This paper was supported by ForestPlots.net approved Research Project 81, ‘Comparative Ecology of African Tropical Forests’. The development of ForestPlots.net and data curation has been funded by several grants, including NE/B503384/1, NE/N012542/1, ERC Advanced Grant 291585—‘T-FORCES’, NE/F005806/1, NERC New Investigators Awards, the Gordon and Betty Moore Foundation, a Royal Society University Research Fellowship and a Leverhulme Trust Research Fellowship. Fieldwork in the Democratic Republic of the Congo (Yangambi and Yoko sites) was funded by the Belgian Science Policy Office BELSPO (SD/AR/01A/COBIMFO, BR/132/A1/AFRIFORD, BR/143/A3/HERBAXYLAREDD, FED-tWIN2019-prf-075/CongoFORCE, EF/211/TREE4FLUX); by the Flemish Interuniversity Council VLIR-UOS (CD2018TEA459A103, FORMONCO II); by L’Académie de recherche et d’enseignement supérieur ARES (AFORCO project) and by the European Union through the FORETS project (Formation, Recherche, Environnement dans la TShopo) supported by the XIth European Development Fund. EMV was supported by fellowship from the CNPq (Grant 308543/2021-1). RAPELD plots in Brazil were supported by the Program for Biodiversity Research (PPBio) and the National Institute for Amazonian Biodiversity (INCT-CENBAM). BGL post-doc grant no. 2019/03379-4, São Paulo Research Foundation (FAPESP). D.A.C. was supported by the CCI Collaborative fund. Plots in Mato Grosso, Brazil, were supported by the National Council for Scientific and Technological Development (CNPq), PELD-TRAN 441244/2016-5 and 441572/2020-0, and Mato Grosso State Research Support Foundation (FAPEMAT)—0346321/2021. We thank E. Chezeaux, R. Condit, W. J. Eggeling, R. M. Ewers, O. J. Hardy, P. Jeanmart, K. L. Khoon, J. L. Lloyd, A. Marjokorpi, W. Marthy, H. Ntahobavuka, D. Paget, J. T. A. Proctor, R. P. Salomão, P. Saner, S. Tan, C. O. Webb, H. Woell and N. Zweifel for contributing forest inventory data. We thank numerous field assistants for their invaluable contributions to the collection of forest inventory data, including A. Nkwasibwe, ITFC field assistant.Peer reviewe