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University of Lyon

4 ARTICLES PUBLISHED IN JoVE

Neuroscience

Use of pHluorin to Assess the Dynamics of Axon Guidance Receptors in Cell Culture and in the Chick Embryo

Céline Delloye-Bourgeois^*1, Arnaud Jacquier^*1, Julien Falk¹, Valérie Castellani¹

¹University Claude Bernard Lyon1, CGphiMC UMR CNRS 5534, University of Lyon

We describe here the use of a pH-sensitive green fluorescent protein variant, pHluorin, to study the spatio-temporal dynamics of axon guidance receptors trafficking at the cell surface. The pHluorin-tagged receptor is expressed both in cell culture and in vivo, using electroporation of the chick embryo.

Neuroscience

Culture of Isolated Floor Plate Tissue and Production of Conditioned Medium to Assess Functional Properties of Floor Plate-released Signals

Camille Charoy¹, Elise Arbeille¹, Karine Thoinet¹, Valérie Castellani¹

¹CGphiMC, UMR CNRS 5534, University of Lyon 1

The floor plate is a crucial structure of the developing spinal cord, providing diffusible signals to progenitors and differentiating neurons. We describe a method to produce floor-plate conditioned medium, to apply it to fresh spinal cord tissue, and to assess the consequences on proteins of interest by biochemistry.

JoVE Journal

Glucose Uptake Measurement and Response to Insulin Stimulation in In Vitro Cultured Human Primary Myotubes

Stephanie Chanon¹, Christine Durand¹, Aurelie Vieille-Marchiset¹, Maud Robert², Charna Dibner³, Chantal Simon¹, Etienne Lefai¹

¹CarMeN Laboratory, INSERM U1060, INRA 1397, University of Lyon, ²Department of digestive and bariatric surgery, Obesity Integrated Center, University Hospital of Edouard Herriot, Hospices Civils de Lyon, Lyon 1 University, ³Division of Endocrinology, Diabetes, Hypertension and Nutrition, Department of Clinical Medicine, Faculty of Medicine, University of Geneva

In this method, human primary muscle cells are cultured in vitro to obtain differentiated myotubes and glucose uptake rates are measured. We provide a detailed protocol to quantify rates in basal and insulin-stimulated states using radiolabeled [³H] 2-deoxy-D-Glucose.

Neuroscience

Modeling Charcot-Marie-Tooth Disease In Vitro by Transfecting Mouse Primary Motoneurons

Arnaud Jacquier^1,2, Valérie Risson¹, Laurent Schaeffer^1,3

¹Institut NeuroMyoGène, CNRS UMR5310, INSERM U1217, Université Lyon, ²Unité Fonctionnelle de Neurogénétique Moléculaire, Hospices Civils de Lyon, ³Centre de Biotechnologie Cellulaire, Hospices Civils de Lyon

The goal of this technique is to prepare a highly enriched culture of primary motoneurons (MNs) from murine spinal cord. To evaluate the consequences of mutations causing MN diseases, we describe here the isolation of these isolated MNs and their transfection by magnetofection.