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Gonzalez-Gonzalez I.M.,University of Bristol | Jaskolski F.,University of Bristol | Jaskolski F.,Grenoble Institute des Neurosciences | Goldberg Y.,University of Bristol | And 2 more authors.
Methods in Enzymology

The use of genetically encoded fluorescent tags such as green fluorescent protein (GFP) as reporters to monitor processes in living cells has transformed cell biology. One major application for these tools has been to analyze protein dynamics in neurons. In particular, fluorescence recovery after photobleach (FRAP) of surface expressed fluorophore-tagged proteins has been instrumental to addressing outstanding questions about how neurons orchestrate the synaptic delivery of proteins. Here, we provide an overview of the methodology, equipment, and analysis required to perform, analyze, and interpret these experiments. © 2012 Elsevier Inc. Source

Chaari L.,French Institute for Research in Computer Science and Automation | Forbes F.,French Institute for Research in Computer Science and Automation | Ciuciu P.,IBM | Vincent T.,IBM | Doiat M.,Grenoble Institute des Neurosciences
IEEE Workshop on Statistical Signal Processing Proceedings

We address the issue of jointly detect brain activity and estimate brain hemodynamics from functional MRI data. To this end, we adopt the so-called JDE framework introduced in [1] and augmented in [2] with hidden Markov field models to account for spatial dependencies between voxels. This latter spatial addition is essential but also responsible for high computation costs. To face the intractability induced by Markov models, inference in [2] is based on intensive simulation methods (MCMC). In this work we propose an alternative to face this limitation by recasting the JDE framework into a missing data framework and to derive an EM algorithm for inference. We address the intractability issue by considering variational approximations. We show that the derived Variational EM algorithm outperforms the MCMC procedure on realistic artificial fMRI data. © 2011 IEEE. Source

Astolfo A.,Swiss Light Source | Astolfo A.,Australian Synchrotron | Qie F.,Beijing University of Chemical Technology | Qie F.,CSIRO | And 13 more authors.
Nanomedicine: Nanotechnology, Biology, and Medicine

The use of alginate based microcapsules to deliver drugs and cells with a minimal host interaction is increasingly being proposed. A proficient method to track the position of the microcapsules during such therapies, particularly if they are amenable to commonly used instrumentation, would greatly help the development of such treatments. Here we propose to label the microcapsules with gold nanoparticles to provide a bright contrast on small animal x-ray micro-CT systems enabling single microcapsule detection. The microcapsules preparation is based on a simple protocol using inexpensive compounds. This, combined with the widespread availability of micro-CT apparatus, renders our method more accessible compared with other methods. Our labeled microcapsules showed good mechanical stability and low cytotoxicity in-vitro. Our post-mortem rodent model data strongly suggest that the high signal intensity generated by the labeled microcapsules permits the use of a reduced radiation dose yielding a method fully compatible with longitudinal in-vivo studies. From the Clinical Editor: The authors of this study report the development of a micro-CT based tracking method of alginate-based microcapsules by incorporating gold nanoparticles in the microcapsules. They demonstrate the feasibility of this system in rodent models, where due to the high signal intensity, even reduced radiation dose is sufficient to track these particles, providing a simple and effective method to track these commonly used microcapsules and allowing longitudinal studies. © 2014. Source

Gottschalk M.,Lund University | Tropres I.,Grenoble University Hospital Center | Lamalle L.,Grenoble University Hospital Center | Grand S.,Joseph Fourier University | And 2 more authors.
NMR in Biomedicine

Short-TE 1H MRS has great potential for brain cancer diagnostics. A major difficulty in the analysis of the spectra is the contribution from short-T2 signal components, mainly coming from mobile lipids. This complicates the accurate estimation of the spectral parameters of the resonance lines from metabolites, so that a qualitative to semi-quantitative interpretation of the spectra dominates in practice. One solution to overcome this difficulty is to measure and estimate the short-T2 signal component and to subtract it from the total signal, thus leaving only the metabolite signals. The technique works well when applied to spectra obtained from healthy individuals, but requires some optimisation during data acquisition. In the clinical setting, time constraints hardly allow this. Here, we propose an iterative estimation of the short-T2 signal component, acquired in a single acquisition after measurement of the full spectrum. The method is based on QUEST (quantitation based on quantum estimation) and allows the refinement of the estimate of the short-T2 signal component after measurement. Thus, acquisition protocols used on healthy volunteers can also be used on patients without further optimisation. The aim is to improve metabolite detection and, ultimately, to enable the estimation of the glutamine and glutamate signals distinctly. These two metabolites are of great interest in the characterisation of brain cancer, gliomas in particular. When applied to spectra from healthy volunteers, the new algorithm yields similar results to QUEST and direct subtraction of the short-T2 signal component. With patients, up to 12 metabolites and, at least, seven can be quantified in each individual brain tumour spectrum, depending on the metabolic state of the tumour. The refinement of the short-T2 signal component significantly improves the fitting procedure and produces a separate short-T2 signal component that can be used for the analysis of mobile lipid resonances. Thus, in brain tumour spectra, distinct estimates of signals from glutamate and glutamine are possible. Copyright © 2016 John Wiley & Sons, Ltd. Copyright © 2016 John Wiley & Sons, Ltd. Source

Lopes C.,Grenoble Institute des Neurosciences | Lopes C.,French Institute of Health and Medical Research | Lopes C.,Grenoble Alpes University | Lopes C.,University of Coimbra | And 11 more authors.

Mutations of the huntingtin protein (HTT) gene underlie both adult-onset and juvenile forms of Huntington's disease (HD). HTT modulates mitotic spindle orientation and cell fate in mouse cortical progenitors from the ventricular zone. Using human embryonic stem cells (hESC) characterized as carrying mutations associated with adult-onset disease during pre-implantation genetic diagnosis, we investigated the influence of human HTT and of an adult-onset HD mutation on mitotic spindle orientation in human neural stem cells (NSCs) derived from hESCs. The RNAi-mediated silencing of both HTT alleles in neural stem cells derived from hESCs disrupted spindle orientation and led to the mislocalization of dynein, the p150Glued subunit of dynactin and the large nuclear mitotic apparatus (NuMA) protein. We also investigated the effect of the adult-onset HD mutation on the role of HTT during spindle orientation in NSCs derived from HD-hESCs. By combining SNP-targeting allelespecific silencing and gain-of-function approaches, we showed that a 46-glutamine expansion in human HTT was sufficient for a dominant-negative effect on spindle orientation and changes in the distribution within the spindle pole and the cell cortex of dynein, p150Glued and NuMA in neural cells. Thus, neural derivatives of disease-specific human pluripotent stem cells constitute a relevant biological resource for exploring the impact of adult-onset HD mutations of the HTT gene on the division of neural progenitors, with potential applications in HD drug discovery targeting HTT-dynein-p150Glued complex interactions. © 2016 Lopes et al. This is an open access article distributed under the terms of the Creative Commons Attribution License, which permits unrestricted use, distribution, and reproduction in any medium, provided the original author and source are credited. Source

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