Softening of POPC membranes by magainin

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Microtubule-associated STOP protein deletion triggers restricted changes in dopaminergic neurotransmission.: Accumbic DA system in STOP KO mice

International audienceThe microtubule-associated stable tubule only polypeptide (STOP) protein plays a key-role in neuron architecture and synaptic plasticity. Recent studies suggest that schizophrenia is associated with alterations in the synaptic connectivity. Mice invalidated for the STOP gene display phenotype reminiscent of some schizophrenic-like symptoms, such as behavioral disturbances, dopamine (DA) hyper-reactivity, and possible hypoglutamatergia, partly improved by antipsychotic treatment. In the present work, we examined potential alterations in some DAergic key proteins and behaviors in STOP knockout mice. Whereas the densities of the DA transporter, the vesicular monoamine transporter and the D(1) receptor were not modified, the densities of the D(2) and D(3) receptors were decreased in some DAergic regions in mutant versus wild-type mice. Endogenous DA levels were selectively decreased in DAergic terminals areas, although the in vivo DA synthesis was diminished both in cell bodies and terminal areas. The DA uptake was decreased in accumbic synaptosomes, but not significantly altered in striatal synaptosomes. Finally, STOP knockout mice were hypersensitive to acute and subchronic locomotor effects of cocaine, although the drug equally inhibited DA uptake in mutant and wild-type mice. Altogether, these data showed that deletion of the ubiquitous STOP protein elicited restricted alterations in DAergic neurotransmission, preferentially in the meso-limbic pathway

Propriétés mécaniques de vésicules géantes fluctuantes

Les vésicules géantes sont un modèle des membranes biologiques souvent utilisées pour des études biophysiques, notamment pour étudier les propriétés mécaniques des bicouches lipidiques. Des développements ont été effectués sur le plan théorique avec la mise en place d'une analyse statistique des fluctuations membranaires permettant d'obtenir un module d'élasticité de courbure plus précis, et au niveau méthodologique avec l'amélioration de la méthode d'électroformation permettant de former des vésicules géantes à partir de différents types de liposomes ou protéoliposomes et de plus dans des conditions relevantes physiologiquement. Une première caractérisation des propriétés mécaniques de vésicules géantes dans des conditions physiologiques a été effectuée en étudiant la magainine. Sa présence conduit à un assouplissement de la bicouche par formation localement de régions à forte courbure à sa surface. Les mesures de l'élasticité de courbure peuvent aussi servir d'outil analytique capable de détecter des dégradations membranaires, comme illustré par l'étude de l'impact de différents fluorophores amphiphiles.L'étude des sels de sodium suggère une forte dépendance des propriétés mécaniques des bicouches de POPC avec la composition du solvant et démontre que la rigidité de courbure fait partie des paramètres qui sont perturbés par les solutions salines selon la série d'Hofmeister. Finalement les effets de l'activité de la pompe à sodium-potassium sur les fluctuations membranaires ont été étudiés. Une amplification des fluctuations de la membrane est observée, ce qui se reflète par une forte diminution de la rigidité de courbure. De plus, deux comportements différents des membranes actives semblent être discriminés : un état de pseudo-équilibre et un état très éloigné de l'équilibre .The giant vesicles are a model of biological membranes often used in biophysical studies, for instance to investigate the mechanical properties of lipid bilayers. Improvements have been performed theoretically by use of a statistical analysis of the membrane fluctuations allowing to obtain a bending rigidity which is more precise, and regarding the electroformation method that allows now to form giant vesicles from different types of liposomes and proteoliposomes and furthermore under physiologically relevant conditions. A first characterization of the mechanical properties of GUVs under physiological buffer conditions has been presented with the magainin study. Magainin molecules induce a softening of the lipid bilayer by formation of local regions of high curvature on the membrane surface. The measurements of the bending rigidity can also be considered as an analytical tool enabling to detect membranes degradation, as illustrated by the study of the impact of different amphiphilic fluorophores. The study of sodium salts effects suggest a strong dependence of the mechanical properties of POPC bilayers on the solvent composition and demonstrates that the bending rigidity falls into the category of phenomena, which are perturbed by salt solutions according to the Hofmeister series. Finally the activity effects of the sodium-potassium pump on the membrane fluctuations have been studied. An enhancement of the membrane fluctuations is observed, as suggested by the major decrease of the bending rigidity. Besides, two distinct behaviors of the active membrane seem to be distinguished: a pseudo-equilibrium state and a far-from-equilibrium state.RENNES1-BU Sciences Philo (352382102) / SudocSudocFranceF

Buffers Affect the Bending Rigidity of Model Lipid Membranes

International audienceIn biophysical and biochemical studies of lipid bilayers the influence of the used buffer is often ignored or assumed to be negligible on membrane structure, elasticity, or physical properties. However, we here present experimental evidence, through bending rigidity measurements performed on giant vesicles, of a more complex behavior, where the buffering molecules may considerably affect the bending rigidity of phosphatidylcholine bilayers. Furthermore, a synergistic effect on the bending modulus is observed in the presence of both salt and buffer molecules, which serves as a warning to experimentalists in the data interpretation of their studies, since typical lipid bilayer studies contain buffer and ion molecules

Advantages of statistical analysis of giant vesicle flickering for bending elasticity measurements

We show how to greatly improve precision when determining bending elasticity of giant unilamellar vesicles. Taking advantage of the well-known quasi-spherical model of liposome flickering, we analyze the full probability distributions of the configurational fluctuations instead of limiting the analysis to the second moment measurements only as usually done in previously published works. This leads to objective criteria to reject vesicles that do not behave according to the model. As a result, the confidence in the bending elasticity determination of individual vesicles that fit the model is improved and, consequently, the reproducibility of this measurement for a given membrane system. This approach uncovers new possibilities for bending elasticity studies like detection of minute influences by solutes in the buffer or into the membrane. In the same way, we are now able to detect the inhomogeneous behavior of giant vesicle systems such as the hazardous production of peroxide in bilayers containing fluorescent dyes

Analysis of the shape fluctuations of reconstituted membranes using GUVs made from lipid extracts of invertebrates

Summary Changes in the physical properties of the lipid matrix of cell membranes have repeatedly been proposed to underlie stresses associated with e.g. drought, cold and xenobiotics. Therefore, the ability to experimentally monitor such properties is central to the fundamental physiological understanding of adaptive changes. Here, we test the analysis of shape fluctuations in membranes composed of lipid extracts from two soil invertebrates, and show that theories and experimental approaches previously developed for simpler liposomes may be applied directly to reconstituted membrane lipids. Specifically, we show how the bending rigidity of giant unilamellar liposomes of lipid extracts can be determined precisely. We suggest that future measurements of this parameter could elucidate mechanisms of adaptive processes such as changes in lipid composition and accumulation of protective osmolytes

Impact of membrane-anchored fluorescent probes on the mechanical properties of lipid bilayers

International audienceFluorescent probes are used in membrane biophysics studies to provide information about physical properties such as lipid packing, polarity and lipid diffusion or to visualize membrane domains. However, our understanding of the effects the dyes themselves may induce on the membrane structure and properties are sparse. As mechanical properties like bending elasticity were already shown to be highly sensitive to the addition of "impurities" into the membranes, we have investigated the impact of six different commonly used fluorescent membrane probes (LAURDAN, TR-DPPE, Rh-DPPE, DiIC18, Bodipy-PC and NBD-PC) on the bending elasticity of dye containing POPC GUVs as compared to single component POPC GUVs. Small changes in the membrane bending elasticity compared to single POPC bilayers are observed when 2 mol% of Rh-DPPE, Bodipy-PC or NBD-PC are added in POPC membranes. These binary membranes are showing non reproducible mechanical properties attributed to a photo-induced peroxidation processes that may be controlled by a reduction of the fluorescent dye concentration. For TR-DPPE, a measurable decrease of the bending elasticity is detected with reproducible bending elasticity measurements. This is a direct indication that this dye, when exposed to illumination by a microscope lamp and contrary to Rh-DPPE, does not induce chemical degradation. At last, LAURDAN and DiIC18 probes mixed with POPC do not significantly affect the bending elasticity of pure POPC bilayers, even at 2 mol%, suggesting these latter probes do not induce major perturbations on the structure of POPC bilayers