Safeguarding Organic Food: The Need for International Certification Standards

To certify an organic product, an accredited agent for the intended market must inspect each producer or manufacturer for compliance with that market’s standards. For example, the U.S. Department of Agriculture (“USDA”) has established a set of national standards that “organic” food must meet, whether it is grown in the United States or imported from abroad. Thus, a Brazilian farmer seeking to export his organic produce to the United States must obtain certification through one of the 95 Accredited Certifying Agents (“ACAs”) recognized by the USDA

G019 Cholesterol depletion enhances Kv1.5-encoded K+ current by increasing Rab11-mediated recycling

Membrane lipid composition is a major determinant of protein organisation in the cell membrane. In a previous study, we reported that depletion of membrane cholesterol by methyl-fÒ-cyclodextrin (MCD) causes a marked increase in Kv1.5-current (Ikur) in neonatal cardiac myocytes. Here, we examined the mechanisms of the cholesterol effects on potassium current in adult rat cardiomyocytes (ARC). GFP-tagged Kv1.5 channels were transduced in ARC using adenoviral vectors and patch clamp experiments were performed to record whole-cell currents and single channel activity. Fluorescence recovery after photobleaching (FRAP) technique was used to investigate GFP-Kv1.5 channels mobility; 3D-epifluorescence microscopy was conducted to follow Kv1.5 channels trafficking.In both freshly isolated and cultured ARC over-expressing GFP-Kv1.5 channels, MCD induced a rapid (< 7min) increase in Ikur but not Ito. On the contrary, incubation with the cholesterol donor LDL reduced Ikur. Single channel experiments revealed that MCD application caused a progressive and drastic increase of the number of active channels. Moreover, FRAP experiments showed that MCD reduced both mobility and recovery of GFP-Kv1.5. Several steps of the trafficking process of ion channels were studied. Blocking SNARE-mediated exocytosis with N-ethylmaleimide prevented the MCD-effect on Ikur. While disruption of Golgi complex/secretion pathway with brefeldine-A had no effect, manipulation of GTP-ases activity with GTP-f×-S suppressed the MCD effect. Transfection with a dominant negative (DN) form of Rab11 effect but not Rab4 DN prevented the MCD. Moreover, Kv1.5 channels co-immunoprecipitated with Rab11 which is stringly expressed in myocardium and ARC (qPCR and western blot). Finally, 3D-microscopy evidenced that Kv1.5 channels association with Rab11-positive recycling endosomes observed in control condition disappeared following cholesterol depletion.ConclusionLowering cholesterol rapidly induces the insertion of Kv1.5 channels by a process that involves vesicle fusion and trafficking processes, particularly the Rab11-associated slow recycling pathway. Given the role of Kv1.5 channel in normal and pathological atrial electrical properties, this study opens news perspectives for therapeutic modulation of cardiac myocytes excitability

Capacitação Tecnológica de Pequenos Agricultores para a Agroindustrialização de frutas e hortaliças

Trabalho apresentado no 31º SEURS - Seminário de Extensão Universitária da Região Sul, realizado em Florianópolis, SC, no período de 04 a 07 de agosto de 2013 - Universidade Federal de Santa Catarina.O emprego da matéria-prima vegetal por meio de técnicas adequadas para elaboração de produtos processados, proporciona alimentos mais seguros e de maior qualidade nutricional à população. Neste contexto, são abertas novas possibilidades para pequenos produtores rurais, dispostos a investir em novas estratégias de comercialização e a ampliar seus mercados. Fica evidente que existe um importante espaço a ser ocupado pelas universidades brasileiras junto a esses grupos. Um bom relacionamento entre universidades e agricultores permite reunir saberes diferentes e inovar em pesquisa e extensão. O objetivo principal deste projeto é capacitar produtores rurais do interior de Pelotas-RS, para produção de derivados de frutas e hortaliças, observando as boas práticas de fabricação, visando fundamentalmente a revitalização das atividades destes produtores. Como primeira etapa do trabalho foi realizado um levantamento, por meio de diálogo, dos anseios e necessidade de diferentes grupos que processam matéria-prima vegetal. A partir de suas necessidades foram ministrados cursos, dentre estes, cursos teóricos relativos a Boas Práticas de Fabricação (BPF), processamento de produtos de origem vegetal - minimamente processados, congelados e produtos desidratados, além de rotulagem/legislação de alimentos. No final das atividades os participantes foram convidados a responder um questionário para a avaliação dos cursos. Quando questionados sobre a avaliação geral dos cursos, 100% classificaram os cursos de bom a ótimo. Assim, acredita-se que as atividades do projeto estão produzindobenefícios relativos à questão da geração de emprego, e também colaborando para a manutenção de espaços socialmente reconhecidos pelo papel que desempenham

Intrahippocampal grafts containing cholinergic and serotonergic fetal neurons ameliorate spatial reference but not working memory in rats with fimbria-fornix/cingular bundle lesions

Three-month-old Long-Evans female rats sustained aspirative lesions of the dorsal septohippocampal pathways and, 2 weeks later, received intrahippocampal suspension grafts containing cells from the mesencephalic raphe, cells from the medial septum and the diagonal band of Broca, or a mixture of both. Lesion-only and sham-operated rats were used as controls. All rats were tested for locomotor activity 1 week, 3 and 5 months after lesion surgery, for spatial working memory in a radial maze from 5 to 9 months, and for reference and working memory in a water tank during the 9th month after lesioning. Determination of hippocampal concentration of acetylcholine, noradrenaline, and serotonin was made after completion of behavioral testing. Compared to sham-operated rats, all rats with lesions, whether grafted or not, exhibited increased levels of locomotor activity and made more errors in the radial maze. The lesioned rats were also impaired in the probe trial (30 first seconds) of the water-tank test made according to a protocol requiring intact reference memory capabilities. While rats with septal or raphe grafts were also impaired, the rats with co-grafts showed performances not significantly different from those of sham-operated rats. With a protocol requiring intact working memory capabilities, all lesioned rats, whether grafted or not, were impaired in the water-tank test. In the dorsal hippocampus of lesion-only rats, the concentration of acetylcholine and serotonin was significantly reduced. In rats with septal grafts or co-grafts, the concentration of acetylcholine was close to normal, as was that of serotonin in rats with raphe grafts or co-grafts. These results confirm previous findings showing that co-grafts enabled the neurochemical properties of single grafts to be combined. Data from the water-tank test suggest that cholinergic and serotonergic hippocampal reinnervations by fetal cell grafts may induce partial recovery of spatial reference, but not working memory capabilities in rats

Advanced Fluorescence Microscopy Techniques-FRAP, FLIP, FLAP, FRET and FLIM

Fluorescence microscopy provides an efficient and unique approach to study fixed and living cells because of its versatility, specificity, and high sensitivity. Fluorescence microscopes can both detect the fluorescence emitted from labeled molecules in biological samples as images or photometric data from which intensities and emission spectra can be deduced. By exploiting the characteristics of fluorescence, various techniques have been developed that enable the visualization and analysis of complex dynamic events in cells, organelles, and sub-organelle components within the biological specimen. The techniques described here are fluorescence recovery after photobleaching (FRAP), the related fluorescence loss in photobleaching (FLIP), fluorescence localization after photobleaching (FLAP), Forster or fluorescence resonance energy transfer (FRET) and the different ways how to measure FRET, such as acceptor bleaching, sensitized emission, polarization anisotropy, and fluorescence lifetime imaging microscopy (FLIM). First, a brief introduction into the mechanisms underlying fluorescence as a physical phenomenon and fluorescence, confocal, and multiphoton microscopy is given. Subsequently, these advanced microscopy techniques are introduced in more detail, with a description of how these techniques are performed, what needs to be considered, and what practical advantages they can bring to cell biological research

A Positive Feedback Synapse from Retinal Horizontal Cells to Cone Photoreceptors

Cone photoreceptors and horizontal cells (HCs) have a reciprocal synapse that underlies lateral inhibition and establishes the antagonistic center-surround organization of the visual system. Cones transmit to HCs through an excitatory synapse and HCs feed back to cones through an inhibitory synapse. Here we report that HCs also transmit to cone terminals a positive feedback signal that elevates intracellular Ca2+ and accelerates neurotransmitter release. Positive and negative feedback are both initiated by AMPA receptors on HCs, but positive feedback appears to be mediated by a change in HC Ca2+, whereas negative feedback is mediated by a change in HC membrane potential. Local uncaging of AMPA receptor agonists suggests that positive feedback is spatially constrained to active HC-cone synapses, whereas the negative feedback signal spreads through HCs to affect release from surrounding cones. By locally offsetting the effects of negative feedback, positive feedback may amplify photoreceptor synaptic release without sacrificing HC-mediated contrast enhancement