Characterization of hormone and protein release from alpha-toxin- permeabilized chromaffin cells in primary culture

Addition of Staphylococcus aureus alpha-toxin to adult bovine chromaffin cells maintained in primary culture causes permeabilization of cell membrane as shown by the release of intracellular 86Rb+. The alpha-toxin does not provoke a spontaneous release of either catecholamines or chromogranin A, a protein marker of the secretory granule, showing the integrity of the secretory vesicle membrane. However the addition of micromolar free Ca2+ concentration induced the co-release of noradrenaline and chromogranin A. In alpha-toxin-treated cells, the released chromogranin A could not be sedimented and lactate dehydrogenase was still associated within cells, which provides direct evidence that secretory product is liberated by exocytosis. By contrast, permeabilization of cells with digitonin caused a Ca2+- dependent but also a Ca2+-independent release of secretory product, a dramatic loss of lactate dehydrogenase, as well as release of secretory product in a sedimentable form. Ca2+-dependent exocytosis from alpha- toxin-permeabilized cells required Mg2+-ATP and did not occur in the presence of other nucleotides. Thus alpha-toxin is a convenient tool to permeabilize chromaffin cells, and has the advantage of keeping intracellular structures, specifically the exocytotic machinery, intact

An ensemble approach to the analysis of weighted networks

We present a new approach to the calculation of measures in weighted networks, based on the translation of a weighted network into an ensemble of edges. This leads to a straightforward generalization of any measure defined on unweighted networks, such as the average degree of the nearest neighbours, the clustering coefficient, the `betweenness', the distance between two nodes and the diameter of a network. All these measures are well established for unweighted networks but have hitherto proven difficult to define for weighted networks. Further to introducing this approach we demonstrate its advantages by applying the clustering coefficient constructed in this way to two real-world weighted networks.Comment: 4 pages 3 figure

Evidence on the effect of the cocoa pulp flavour environment during fermentation on the flavour profile of chocolates

Empirical observations carried out by the first author during his many visits to cocoa producing countries suggested that a relationship might exist between cocoa pulp flavour attributes and sensory traits of cocoa liquor and chocolates. A first attempt to demonstrate such a relationship was made by evaluating the following flavour attributes: volatile aroma upon opening the pods, pulp sweetness, pulp acidity, pulp astringency, pulp flavour intensity and type, overall preference for pulp taste, and degree of bitterness of the cotyledon. Blind pulp tasting was carried out by a six-person panel of ripe fruits of nine cocoa clonal varieties, representing different genetic origins, several of which are well known for their contrasting sensory traits of their chocolates. Overall pulp preference was significantly related to the known fine flavour potential of the cocoa varieties tested, whereas specific pulp flavour attributes appeared related to specific types of fine flavour of the tested cocoa genotypes. One striking example was the EET 62 clone (Nacional x Trinitario), which displayed a "jasmine" flower odour upon opening the ripe fruit, sweet pulp with an intensive pleasant flavour that was identified as floral and fruity. These flavour attributes are apparently related to the known typical "Arriba" flavour of the EET62 clone. The typical fresh-fruit Trinitario cocoa flavour of the ICS 1 clone appeared to be related to a balanced combination of relatively high acidity and sweetness and a moderate citrus-like flavour of its pulp. On the other side, the pulp of the potentially low-flavour CCN51 clone was rated as very astringent and acid, with low flavour intensity and quality. The pulp of the Pandora 1 clone from Colombia was characterised by a sour sop (Guanabana) flavour and may therefore represent a new interesting type of fine-flavour cocoa. These results suggest that a significant part of fine-flavour attributes of cocoa products (excluding basic cocoa flavour) can be related to sensory traits identifiable in the pulp of ripe cocoa pods. The effect of the pulp flavour environment on the taste of chocolates was further experimentally demonstrated by adding equal quantities of aromatic pulp of two tropical fruit species (Theobroma grandiflorum and Anona muricata) with Amelonado beans during the fermentation process. The chocolates obtained by this method revealed the 2 presence of the typical flavour attributes of T. grandiflorum and A.muricata. This shows that the added aromatic compounds are absorbed by the cocoa cotyledon during the fermentation process. An amateur public in Montpellier, France, showed high levels of preference for the chocolates that had acquired flavours of T. grandiflorum and A. muricata in comparison to that of the pure Amelonado control treatment. The combined findings suggest that: 1) Rapid screening of germplasm and of breeding populations for pulp characteristics can be attempted to select cocoa varieties that display good potential for known as well as for new fine flavor traits, 2) Chocolate flavors can be modified substantially by adding aromatic substances to cocoa beans during the fermentation process. The further development of such an innovative way to produce cocoa liquors and chocolates with enhanced and/or modified flavour profiles is expected to have important commercial applications. It would also provide new opportunities for the use of aromatic tropical fruit pulps that are often under-utilized and under-valued. (Résumé d'auteur

Scaling of energy spreading in strongly nonlinear disordered lattices

To characterize a destruction of Anderson localization by nonlinearity, we study the spreading behavior of initially localized states in disordered, strongly nonlinear lattices. Due to chaotic nonlinear interaction of localized linear or nonlinear modes, energy spreads nearly subdiffusively. Based on a phenomenological description by virtue of a nonlinear diffusion equation we establish a one-parameter scaling relation between the velocity of spreading and the density, which is confirmed numerically. From this scaling it follows that for very low densities the spreading slows down compared to the pure power law.Comment: 4 pages, 4 figure

Applying weighted network measures to microarray distance matrices

In recent work we presented a new approach to the analysis of weighted networks, by providing a straightforward generalization of any network measure defined on unweighted networks. This approach is based on the translation of a weighted network into an ensemble of edges, and is particularly suited to the analysis of fully connected weighted networks. Here we apply our method to several such networks including distance matrices, and show that the clustering coefficient, constructed by using the ensemble approach, provides meaningful insights into the systems studied. In the particular case of two data sets from microarray experiments the clustering coefficient identifies a number of biologically significant genes, outperforming existing identification approaches.Comment: Accepted for publication in J. Phys.

Introduction of Macromolecules into Bovine Adrenal Medullary Chromaffin Cells and Rat Pheochromocytoma Cells (PC12) by Permeabilization with Streptolysin O: Inhibitory Effect of Tetanus Toxin on Catecholamine Secretion

Conditions are described for controlled plasma membrane permeabilization of rat pheochromocytoma cells (PC12) and cultured bovine adrenal chromaffin cells by Streptolysin O (SLO). The transmembrane pores created by SLO invoke rapid efflux of intracellular 86Rb+ and ATP, and also permit passive diffusion of proteins, including immunoglobulins, into the cells. SLO-permeabilized PC12 cells release [3H]dopamine in response to micromolar concentrations of free Ca2+. Permeabilized adrenal chromaffin cells present a similar exocytotic response to Ca2+ in the presence of Mg2+/ ATP. Permeabilized PC12 cells accumulate antibodies against synaptophysin and calmodulin, but neither antibody reduces the Ca2+-dependent secretory response. Reduced tetanus toxin, although ineffective when applied to intact chromaffin cells, inhibits Ca2+-induced exocytosis by both types of permeabilized cells studied. Omission of dithiothreitol, toxin inactivation by boiling, or preincubation with neutralizing antibodies abolishes the inhibitory effect. The data indicate that plasma membrane permeabilization by Streptolysin O is a useful tool to probe and define cellular components that are involved in the final steps of exocytosis

Eléments d'information sur les effets de l'environnement aromatique de la pulpe de cacao pendant la fermentation sur le profil d'arôme des chocolats

Des observations empiriques réalisées par le premier auteur au cours de ses nombreuses visites dans des pays producteurs de cacao ont laissé penser qn'il pour exister une relation entre les caractéristiques aromatiques de la pulpe de cacao et les caractéristiques sensorielles de la liqueur de cacao et des chocolats. Une première tentative visant à démontrer cette relation a été faite en évaluant les caractéristiques aromatiques suivantes: arôme volatil à l'ouverture des cabosses, douceur de la pulpe, acidité de la pulpe, astringence de la pulpe, intensité et type d'arôme de la pulpe, préférence globale pour le goût de la pulpe et degré d'amertume du cotylédon. Un test de dégustation à l'aveugle de la pulpe a été réalisé par un jury de six personnes sur des fruits mûrs de neuf variétés clonales de cacaoyers, représentant différentes origines génétiques, dont plusieurs sont bien connues pour les caractéristiques sensorielles contrastées de leurs chocolats. La préférence globale relative à la pulpe était significativement liée au potentiel d'arôme fin connu des diférentes variétés de cacao testés, alors que les caractéristiques aromatiques spécifiques de la pulpe sont apparues comme étant liées à des types spécifiques d'arôme fin des génotypes de cacaoyers testés. Une illustration frappante de ce phénomène a été donnée par le clone EET 63 (Nacional x Trinitario), qui présentait une odeur florale de "jasmin" à l'ouverture du fruit mûr, une pulpe sucrée avec un arôme agréable intense qui a été qualifié de floral et fruité. Ces caractéristiques aromatiques sont apparement liées à l'arôme "Arriba" typique reconnu du clone EET62. L'arôme typique de fruit frais du cacao Trinitario du clone ICS 1 semblait être associé à une combinaison équili"brée de teneur relativement élevée d'acidité et de douceur et un arôme d'agrumes modéré de sa pulpe. D'autre part, la pulpe du clone CCN51, potentiellement associée à un arôme léger, a été classée comme très astringente et acide, avec une faible intensité et qualité d'arôme. La pulpe du clone Pandora 1 de Colombie était caractérisée par un arôme de corossol (Guanabana) et pour donc représenter un nouveau type de cacao d'arôme fin intéressant Ces résultats suggèrent qu'une partie significative des caractéristiques aromatiques des produits cacaoyers (en dehors de l'arôme de base du cacao) peuvent être asés à des caractéristiques sensorielles identifiables dans la pulpe des cabosses de cacao mûres. L'effet de l'environnement aromatique de la pulpe sur le goût des chocolats a encore été démontré par des expériences, en ajoutant des quantités égales de pulpe aromatique de deux espèces de fruits tropicaux (Theobroma grandiflorum et Anona muricata) avec les fèves Amelonado pendant le processus de fermentation. Les chocolats obtenus par cette méthode ont révélé la présence des caractéristiques aromatiques typiques de T. grandiflorum et A. muricata. Ceci montre que les composés aromatiques ajoutés sont absorbés par le cotylédon de cacao pendant le processus de fermentation. Un panel d'amateurs à Montpellier, en France, a manifesté une préférence marquée pour les chocolats qui avaient acquis des arômes de T grandiflorum et A. muricata par rapport à ceux du traitement témoin Amelonado pur. Les conclusions combinées constituent: 1) des éléments démontrant clairement qu'une présélection de génotypes de cacaoyers de haute qualité spécifiques peut être faite efficacement par une évaluation sensorielle rapide des pulpes de cacao, 2) les premiers éléments scientifiques publiés démontrant que les arômes du chocolat peuvent être modifiés de façon significative en ajoutant des substances aromatiques aux fèves de cacao pendant le processus de fermentation; La poursuite du développement de cette méthode innovante de production de liqueurs de cacao et de chocolats avec des profils aromatiques améliorés et/ou modifiés devrait avoir des applications commerciales importantes. Elle offrirait aussi de nouvelles opportunités

Molecular Aspects of Secretory Granule Exocytosis by Neurons and Endocrine Cells

Neuronal communication and endocrine signaling are fundamental for integrating the function of tissues and cells in the body. Hormones released by endocrine cells are transported to the target cells through the circulation. By contrast, transmitter release from neurons occurs at specialized intercellular junctions, the synapses. Nevertheless, the mechanisms by which signal molecules are synthesized, stored, and eventually secreted by neurons and endocrine cells are very similar. Neurons and endocrine cells have in common two different types of secretory organelles, indicating the presence of two distinct secretory pathways. The synaptic vesicles of neurons contain excitatory or inhibitory neurotransmitters, whereas the secretory granules (also referred to as dense core vesicles, because of their electron dense content) are filled with neuropeptides and amines. In endocrine cells, peptide hormones and amines predominate in secretory granules. The function and content of vesicles, which share antigens with synaptic vesicles, are unknown for most endocrine cells. However, in B cells of the pancreatic islet, these vesicles contain GABA, which may be involved in intrainsular signaling.' Exocytosis of both synaptic vesicles and secretory granules is controlled by cytoplasmic calcium. However, the precise mechanisms of the subsequent steps, such as docking of vesicles and fusion of their membranes with the plasma membrane, are still incompletely understood. This contribution summarizes recent observations that elucidate components in neurons and endocrine cells involved in exocytosis. Emphasis is put on the intracellular aspects of the release of secretory granules that recently have been analyzed in detail

Additive Nonparametric Reconstruction of Dynamical Systems from Time Series

We present a nonparametric way to retrieve a system of differential equations in embedding space from a single time series. These equations can be treated with dynamical systems theory and allow for long term predictions. We demonstrate the potential of our approach for a modified chaotic Chua oscillator.Comment: accepted for Phys. Rev. E, Rapid Com