Ralf Dringen

Universität Bremen, Bremen, Bremen, Germany

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Publications (189)658.74 Total impact

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    ABSTRACT: Lead intoxication in humans is characterized by cognitive impairments, particularly in the domain of memory, where evidence indicates that glutamatergic neurotransmission may be impacted. Animal and cell culture studies have shown that lead decreases the expression and activity of glutamine synthetase (GS) in astrocytes, yet the basis of this effect is uncertain. To investigate the mechanism responsible, the present study exposed primary astrocyte cultures to a range of concentrations of lead acetate (0–330 μM) for up to 24 h. GS activity was significantly reduced in cells following 24 h incubation with 100 or 330 μM lead acetate. However, no reduction in GS activity was detected when astrocytic lysates were co-incubated with lead acetate, suggesting that the mechanism is not due to a direct interaction and involves intact cells. Since GS is highly sensitive to oxidative stress, the capacity of lead to inhibit the clearance of hydrogen peroxide (H2O2) was investigated. It was found that exposure to lead significantly diminished the capacity of astrocytes to degrade H2O2, and that this was due to a reduction in the effectiveness of the glutathione system, rather than to catalase. These results suggest that the inhibition of GS activity in lead poisoning is a consequence of slowed H2O2 clearance, and supports the glutathione pathway as a primary therapeutic target.
    Preview · Article · Dec 2015 · Frontiers in Integrative Neuroscience
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    ABSTRACT: In this study, we demonstrate how functional groups on the surface of mesoporous silica nanoparticles (MSNPs) can influence the encapsulation and release of the anticancer drug doxorubicin, as well as cancer cell response in the absence or presence of serum proteins. To this end, we synthesized four differently functionalized MSNPs with amine, sulfonate, polyethylene glycol or polyethylene imine functional surface groups, as well as one type of antibody-conjugated MSNP for specific cellular targeting and characterized these MSNPs regarding their physicochemical properties, colloidal stability in physiological media and uptake and release of doxorubicin in vitro. Then, the MSNPs were investigated for their cytotoxic potential on cancer cells. Cationic MSNPs could not be loaded with doxorubicin and did therefore not show any cytotoxic and antiproliferative potential on osteosarcoma cells, although they were efficiently taken up into the cells in the presence or absence of serum. In contrast, substantial amounts of doxorubicin were loaded into negatively charged and unfunctionalized MSNPs. Especially sulfonate functionalized doxorubicin-loaded MSNPs were efficiently taken up into the cells in the presence of serum and showed an accelerated toxic and antiproliferative potential compared to unfunctionalized MSNPs, antibody-conjugated MSNPs and even free doxorubicin. These findings stress the high importance of the surface charge as well as of the protein corona for designing and applying nanoparticles for targeted drug delivery.
    No preview · Article · Nov 2015 · ACS Applied Materials & Interfaces
  • Adrian Westhaus · Eva Maria Blumrich · Ralf Dringen
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    ABSTRACT: Metformin is the most frequently used drug for the treatment of type 2 diabetes in humans. However, only little is known about effects of metformin on brain metabolism. To investigate potential metabolic consequences of an exposure of brain cells to metformin, we incubated rat astrocyte-rich primary cultures with this compound. Metformin in concentrations of up to 30 mM did not acutely compromise the viability of astrocytes, but caused a time- and concentration-dependent increase in cellular glucose consumption and lactate production. For acute incubations in the hour range, the presence of 10 mM metformin doubled the glycolytic flux, while already 1 mM metformin doubled glycolytic flux during incubation for 24 h. In addition to metformin, also other guanidino compounds increased astrocytic lactate production. After 4 h of incubation, half-maximal stimulation of glycolysis was observed for metformin, guanidine and phenformin at concentrations of around 3 mM, 3 mM and 30 µM, respectively. The acute stimulation of glycolytic lactate production by metformin was persistent after removal of extracellular metformin and was also observed, if glucose was absent from the incubation medium or replaced by other hexoses. The metformin-induced stimulation of glycolytic flux was not prevented by compound C, an inhibitor of AMP-dependent protein kinase, nor was it additive to the stimulation of glycolytic flux caused by respiratory chain inhibitors. These data demonstrate that the antidiabetic drug metformin has the potential to strongly activate glycolytic lactate production in brain astrocytes.
    No preview · Article · Oct 2015 · Neurochemical Research
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    ABSTRACT: Copper is essential for several important cellular processes, but an excess of copper can also lead to oxidative damage. In brain, astrocytes are considered to play a pivotal role in the copper homeostasis and antioxidative defence. To investigate whether antioxidants and copper chelators can modulate the uptake and the toxicity of copper ions in brain astrocytes, we used primary astrocytes as cell culture model. These cells accumulated substantial amounts of copper during exposure to copper chloride. Copper accumulation was accompanied by a time- and concentration-dependent loss in cell viability, as demonstrated by a lowering in cellular MTT reduction capacity and by an increase in membrane permeability for propidium iodide. During incubations in the presence of the antioxidants ascorbate, trolox or ebselen, the specific cellular copper content and the toxicity in copper chloride-treated astrocyte cultures were strongly increased. In contrast, the presence of the copper chelators bathocuproine disulfonate or tetrathiomolybdate lowered the cellular copper accumulation and the copper-induced as well as the ascorbate-accelerated copper toxicity was fully prevented. These data suggest that predominantly the cellular content of copper determines copper-induced toxicity in brain astrocytes. Copyright © 2015 Elsevier GmbH. All rights reserved.
    No preview · Article · Oct 2015 · Journal of Trace Elements in Medicine and Biology
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    ABSTRACT: In this study we use a straightforward experimental method to probe the presence and activity of the proteolytic enzyme α-chymotrypsin adsorbed on titania colloidal particles. We show that the adsorption of α-chymotrypsin on the particles is irreversible and pH-dependent. At pH 8 the amount of adsorbed chymotrypsin is threefold higher compared to the adsorption at pH 5. However, we observe that the adsorption is accompanied by a substantial loss of enzymatic activity, and only around 6-9% of the initial enzyme activity is retained. A Michaelis-Menten kinetics analysis of both unbound and TiO2-bound chymotrypsin shows that the KM value is increased from ∼10μM for free chymotrypsin to ∼40μM for the particle bound enzyme. Such activity decrease could be related by the hindered accessibility of substrate to the active site of adsorbed chymotrypsin, or by adsorption-induced structural changes. Our simple experimental method does not require any complex technical equipment, can be applied to a broad range of hydrolytic enzymes and to various types of colloidal materials. Our approach allows an easy, fast and reliable determination of particle surface-bound enzyme activity and has high potential for development of future enzyme-based biotechnological and industrial processes. Copyright © 2015 Elsevier Inc. All rights reserved.
    No preview · Article · Oct 2015 · Journal of Colloid and Interface Science
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    ABSTRACT: A protein corona forms spontaneously around silica nanoparticles (SNPs) in serum-containing media. To test whether this protein corona can be utilized for the loading and release of anticancer drugs we incorporated the hydrophilic doxorubicin, the hydrophobic meloxicam as well as their combination in the corona around SNPs. The application of corona-covered SNPs to osteosarcoma cells revealed that drug-free particles did not affect the cell viability. In contrast, SNPs carrying a protein corona with doxorubicin or meloxicam lowered the cell proliferation in a concentration-dependent manner. In addition, these particles had an even greater antiproliferative potential than the respective concentrations of free drugs. The best antiproliferative effects were observed for SNPs containing both doxorubicin and meloxicam in their corona. Co-localization studies revealed the presence of doxorubicin fluorescence in the nucleus and lysosomes of cells exposed to doxorubicin-containing coated SNPs, suggesting that endocytotic uptake of the SNPs facilitates the cellular accumulation of the drug. Our data demonstrate that the protein corona, which spontaneously forms around nanoparticles, can be efficiently exploited for loading the particles with multiple drugs for therapeutic purposes. As drugs are efficiently released from such particles they may have a great potential for nanomedical applications.
    No preview · Article · Sep 2015 · Nanoscale
  • Charlotte Petters · Karsten Thiel · Ralf Dringen
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    ABSTRACT: Iron oxide nanoparticles (IONPs) are used for various biomedical and neurobiological applications. Thus, detailed knowledge on the accumulation and toxic potential of IONPs for the different types of brain cells is highly warranted. Literature data suggest that microglial cells are more vulnerable towards IONP exposure than other types of brain cells. To investigate the mechanisms involved in IONP-induced microglial toxicity, we applied fluorescent dimercaptosuccinate-coated IONPs to primary cultures of microglial cells. Exposure to IONPs for 6 h caused a strong concentration-dependent increase in the microglial iron content which was accompanied by a substantial generation of reactive oxygen species (ROS) and by cell toxicity. In contrast, hardly any ROS staining and no loss in cell viability were observed for cultured primary astrocytes and neurons although these cultures accumulated similar specific amounts of IONPs than microglia. Co-localization studies with lysotracker revealed that after 6 h of incubation in microglial cells, but not in astrocytes and neurons, most IONP fluorescence was localized in lysosomes. ROS formation and toxicity in IONP-treated microglial cultures were prevented by neutralizing lysosomal pH by the application of NH4Cl or Bafilomycin A1 and by the presence of the iron chelator 2,2′-bipyridyl. These data demonstrate that rapid iron liberation from IONPs at acidic pH and iron-catalyzed ROS generation are involved in the IONP-induced toxicity of microglia and suggest that the relative resistance of astrocytes and neurons against acute IONP toxicity is a consequence of a slow mobilization of iron from IONPs in the lysosomal degradation pathway.
    No preview · Article · Aug 2015 · Nanotoxicology
  • Felix Bulcke · Ralf Dringen
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    ABSTRACT: Copper is an essential trace element for many important cellular functions. However, excess of copper can impair cellular functions by copper-induced oxidative stress. In brain, astrocytes are considered to play a prominent role in the copper homeostasis. In this short review we summarise the current knowledge on the molecular mechanisms which are involved in the handling of copper by astrocytes. Cultured astrocytes efficiently take up copper ions predominantly by the copper transporter Ctr1 and the divalent metal transporter DMT1. In addition, copper oxide nanoparticles are rapidly accumulated by astrocytes via endocytosis. Cultured astrocytes tolerate moderate increases in intracellular copper contents very well. However, if a given threshold of cellular copper content is exceeded after exposure to copper, accelerated production of reactive oxygen species and compromised cell viability are observed. Upon exposure to sub-toxic concentrations of copper ions or copper oxide nanoparticles, astrocytes increase their copper storage capacity by upregulating the cellular contents of glutathione and metallothioneins. In addition, cultured astrocytes have the capacity to export copper ions which is likely to involve the copper ATPase 7A. The ability of astrocytes to efficiently accumulate, store and export copper ions suggests that astrocytes have a key role in the distribution of copper in brain. Impairment of this astrocytic function may be involved in diseases which are connected with disturbances in brain copper metabolism.
    No preview · Article · Aug 2015 · Neurochemical Research
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    ABSTRACT: To study the importance of the surface charge for cellular uptake of silica nanoparticles (NPs) we synthesised five different single- or multi-functionalised fluorescent silica NPs (FFSNPs) by introducing various ratios of amino and sulfonate groups into their surface. These FFSNPs were tailored in their zeta potential values from highly positive to highly negative, while other physicochemical properties remained almost constant. Irrespective of the original surface charge, serum proteins adsorbed onto the surface, neutralised the zeta potential values and prevented the aggregation of the tailor-made FFSNPs. Depending on the surface charge and on the absence or presence of serum, two opposite trends were found concerning the cellular uptake of FFSNPs. In the absence of serum, positively charged NPs were stronger accumulated by human osteoblast (HOB) cells than negatively charged NPs. In contrast, in serum-containing medium anionic FFSNPs were internalised by HOB cells more strongly, despite the similar size and surface charge of all types of protein-covered FFSNPs. Thus, at physiological condition, when the presence of proteins is inevitable sulfonate-functionalised silica NPs are the favourite choice to achieve a desired high rate of NP internalisation.
    Full-text · Article · Jun 2015 · ACS Applied Materials & Interfaces
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    Eva-Maria Blumrich · Reshma Kadam · Ralf Dringen

    Full-text · Article · Jun 2015 · SpringerPlus
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    Felix Bulcke · Ralf Dringen · Karsten Thiel

    Full-text · Article · Jun 2015 · SpringerPlus
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    Charlotte Petters · Ralf Dringen

    Preview · Article · Jun 2015 · SpringerPlus
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    ABSTRACT: The inorganic arsenic species arsenate and arsenite are common environmental toxins which contaminate the drinking water in many countries. Chronic intoxication with arsenicals has been connected with various diseases, but causes also neurological complications and impairs cognitive development, learning and memory. In brain, astrocytes have a pivotal role as partners of neurons in homeostatic and metabolic processes. In addition, astrocytes are the first parenchymal brain cell type which encounters substances which cross the blood-brain barrier and are considered as first line of defence against the toxic potential of xenobiotics. Therefore, astrocytes are likely to play a prominent role in the metabolism and potential detoxification of arsenicals in brain. This article summarizes the current knowledge on the uptake and toxicity of arsenate and arsenite in astrocytes and discusses the modulation of the astrocytic glucose and glutathione metabolism by arsenicals.
    No preview · Article · Apr 2015 · Neurochemical Research
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    ABSTRACT: The immobilization of enzymes on solid materials is a promising strategy in biotechnological applications and proteomics. It can improve the enzymes’ stability, and enables a more convenient handling, easy separation from the reaction solution, and cyclic reuse of the enzymes. In order to investigate the proteolytic properties of a particle-bound protease, chymotrypsin was covalently immobilized on silica and alumina colloidal particles. The enzymatic activity of the bound chymotrypsin at different times, in consecutive proteolytic cycles, and after storage up to several weeks was investigated by matrix-assisted laser desorption/ionization time-of-flight mass spectrometry (MALDI-ToF MS). Using this approach, the proteolysis products were identified without using artificial protease substrates or intermediate chemicals. Lysozyme was used as a model protein to perform enzymatic digestion using immobilized chymotrypsin and the peptides generated from the proteolytic digestion were determined. Compared to the activity of chymotrypsin applied for the immobilization reactions, more active chymotrypsin was bound to alumina (between 1 and 10% of the initial concentration) than to silica (below 1%) colloidal particles. Compared to an excess of unbound chymotrypsin, the digestion of lysozyme was slower with chymotrypsin immobilized on colloidal particles and only 60% of the maximal amounts of lysozyme peptides were detected. The proteolytic activity of chymotrypsin immobilized on colloidal particles was maintained during storage at room temperature for up to at least seven weeks, while it was lowered during consecutive digestions.
    No preview · Article · Feb 2015 · Analytical Letters
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    ABSTRACT: Based on the chemical structure and the known chemical synthesis of the marine sponge alkaloid ageladine A, we synthesized the ageladine A-derivative 4-(naphthalene-2-yl)-1H-imidazo[4,5-c]pyridine trifluoroacetate (LysoGlow84). The two-step synthesis started with the Pictet-Spengler reaction of histamine and naphthalene-2-carbaldehyde to a tetrahydropyridine intermediate, which was dehydrogenated with activated manganese (IV) oxide to LysoGlow84. Structure and purity of the synthesized LysoGlow84 were confirmed by NMR spectroscopy and mass spectrometry. The fluorescence intensity emitted by LysoGlow84 depended strongly on the pH of the solvent with highest fluorescence intensity recorded at pH 4. The fluorescence maximum (at 315 nm excitation) was 921 observed at 440 nm. Biocompatibility of LysoGlow84 was investigated using cultured rat brain astrocytes and the marine flatworm Macrostomum lignano. Exposure of the astrocytes for up to 6 h to micromolar concentrations of LysoGlow84 did not compromise cell viability, as demonstrated by several viability assays, but revealed a promising property of this compound for staining of cellular vesicles. Conventional fluorescence microscopy as well as confocal scanning microscopy of LysoGlow84-treated astrocytes revealed co-localization of LysoGlow84 fluorescence with that of LysoTracker ® Red DND-99. LysoGlow84 stained unclear structures in Macrostomum lignano, which were identified as lysosomes by co-staining with LysoTracker. Strong fluorescence staining by LysoGlow84 was further observed around the worms' anterior gut and the female genital pore which were not counterstained by LysoTracker Red. Thus, LysoGlow84 is a new promising dye that stains lysosomes and other acidic compartments in cultured cells and in worms.
    Full-text · Article · Feb 2015 · Marine Drugs
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    ABSTRACT: Colloidosomes are microcapsules consisting of nanoparticle shells. These microcarriers can be self-assembled from a wide range of colloidal particles with selective chemical, physical, and morphological properties and show promise for application in the field of theranostic nanomedicine. Previous studies have mainly focused on fairly large colloidosomes (>1 μm) based on a single kind of particle; however, the intrinsic building-block nature of this microcarrier has not been exploited so far for the introduction of tailored functionality at the nanoscale. We report a synthetic route based on interfacial shear rheology studies that allows the simultaneous incorporation of different nanoparticles with distinct physical properties, that is, superparamagnetic iron oxide and fluorescent silica nanoparticles, in a single submicron colloidosome. These tailor-made microcapsules can potentially be used in various biomedical applications, including magnetic hyperthermia, magnetic particle imaging, drug targeting, and bioimaging.
    Full-text · Article · Jan 2015 · Angewandte Chemie International Edition
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    ABSTRACT: Arsenate is an environmental pollutant which contaminates the drinking water of millions of people worldwide. Numerous in vitro studies have investigated the toxicity of arsenate for a large number of different cell types. However, despite the known neurotoxic potential of arsenicals, little is known so far about the consequences of an exposure of neurons to arsenate. To investigate acute effects of arsenate on the viability and the glutathione (GSH) metabolism of neurons, we have exposed primary rat cerebellar granule neuron cultures to arsenate. Incubation of neurons for up to 6 h with arsenate in concentrations of up to 10 mM did not acutely compromise the cell viability, although the cells accumulated substantial amounts of arsenate. However, exposure to arsenate caused a time- and concentration-dependent increase in the export of GSH from viable neurons with significant effects observed for arsenate in concentrations above 0.3 mM. The arsenate-induced stimulation of GSH export was abolished upon removal of arsenate and completely prevented by MK571, an inhibitor of the multidrug resistance protein 1. These results demonstrate that arsenate is not acutely toxic to neurons but can affect the neuronal GSH metabolism by stimulating GSH export.
    No preview · Article · Dec 2014 · Neurochemical Research
  • Charlotte Petters · Ralf Dringen
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    ABSTRACT: Magnetic iron oxide nanoparticles (IONPs) are frequently used for biomedical applications. Although nanoparticles can enter the brain, little is known so far on the uptake of IONPs in neurons and on their neurotoxic potential. Hence, we applied dimercaptosuccinate (DMSA)-coated IONPs to cultured primary rat cerebellar granule neurons. These IONPs had average hydrodynamic diameters of around 80 nm and 120 nm when dispersed in incubation medium in the absence and the presence of 10% fetal calf serum, respectively. Acute exposure of neurons with IONPs for up to 6 h did neither alter the cell morphology nor compromise cell viability, although neurons accumulated large amounts of IONPs in a time- and concentration-dependent manner which caused delayed toxicity. For the first 30 min of incubation of neurons at 37°C with IONPs the cellular iron content increased proportionally to the concentration of IONPs applied irrespective of the absence and the presence of serum. IONP-exposure in the absence of serum generated maximal cellular iron contents of around 3000 nmol iron/mg protein after 4 h of incubation, while the accumulation in presence of 10% serum was slower and reached already within 1 h maximal values of around 450 nmol iron/mg protein. For both incubation conditions was the increase in cellular iron contents significantly lowered by reducing the incubation temperature to 4°C. Application of inhibitors of endocytotic pathways did not affect neuronal IONP accumulation in the absence of serum, while inhibitors of clathrin-mediated endocytosis lowered significantly the IONP accumulation in the presence of serum. These data demonstrate that DMSA-coated IONPs are not acutely toxic to cultured neurons and that a protein corona around the particles strongly affects their interaction with neurons. Copyright © 2014. Published by Elsevier Ltd.
    No preview · Article · Dec 2014 · Neurochemistry International
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    ABSTRACT: Astrocytes have a pivotal role in brain as partners of neurons in homeostatic and metabolic processes. Astrocytes also protect other types of brain cells against the toxicity of reactive oxygen species and are considered as first line of defence against the toxic potential of xenobiotics. A key component in many of the astrocytic detoxification processes is the tripeptide glutathione (GSH) which serves as electron donor in the GSH peroxidase-catalyzed reduction of peroxides. In addition, GSH is substrate in the detoxification of xenobiotics and endogenous compounds by GSH-S-transferases which generate GSH conjugates that are efficiently exported from the cells by multidrug resistance proteins. Moreover, GSH reacts with the reactive endogenous carbonyls methylglyoxal and formaldehyde to intermediates which are substrates of detoxifying enzymes. In this article we will review the current knowledge on the GSH metabolism of astrocytes with a special emphasis on GSH-dependent detoxification processes.
    No preview · Article · Nov 2014 · Neurochemical Research
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    ABSTRACT: Fluorescently labeled nanoparticles (NPs) are used in a wide range of biomedical and nanotoxicological studies to elucidate their interactions with cellular components and their intracellular localization. As commonly used fluorescence microscopes are usually limited in their performance to a few channels which detect the emitted fluorescence light in the red, green and blue color range, the simultaneous colocalization of accumulated fluorescent NPs with cellular markers is often difficult and remains a challenge due to spectral overlay of NP-fluorescence and fluorescence of stained cellular components. To overcome this problem we have synthesized three different photostable dual-labeled fluorescent core/shell silica NPs with high fluorescence intensity (FI) and well defined shape, size and surface chemistry. The synthesis route of dual fluorophore doped silica (DFDS) NPs was based on a water in oil microemulsion method and includes the separate incorporation of two fluorophores in core or shell. The suitability of DFDS for colocalization studies was assessed and successfully demonstrated with human osteoblast (HOB) cells. Parallel visualization of DFDS NPs with two separate microscope channels allowed cellular NPs uptake and discrimination from fluorescently stained cellular components, even in triple stained cells that show fluorescence for the cytoskeleton protein actin (green), the nucleus (blue) and collagen (red). Our results demonstrate the feasibility and straightforwardness of the approach for colocalization studies at single-cell-level to discern clearly the accumulation of NPs from triple stained cellular components. Such NPs with multiple fluorescence characteristics have a great potential to replace single fluorescent NPs for in vitro studies, when multiple staining of cellular components is required. Copyright © 2014. Published by Elsevier Ltd.
    No preview · Article · Nov 2014 · Acta Biomaterialia

Publication Stats

9k Citations
658.74 Total Impact Points


  • 1998-2015
    • Universität Bremen
      • • Advanced Ceramics
      • • Center for Biomolecular Interactions CBIB
      Bremen, Bremen, Germany
  • 2012
    • Jacobs University
      Bremen, Bremen, Germany
  • 2006-2007
    • Monash University (Australia)
      • School of Psychology and Psychiatry
      Melbourne, Victoria, Australia
  • 1992-2005
    • University of Tuebingen
      • • Institute for Physiology
      • • Institute of Physical and Theoretical Chemistry
      Tübingen, Baden-Württemberg, Germany
  • 2004
    • University of Vic
      Vic, Catalonia, Spain