Culturing of cerebellar granule cells to study neuronal migration: gradient and local perfusion assays

Curr Protoc Neurosci. 2012 Jul:Chapter 3:Unit 3.26. doi: 10.1002/0471142301.ns0326s60.

Abstract

Cultures of cerebellar granule cells are a suitable model to analyze the mechanisms governing neuronal migration. In this unit, we describe a protocol to obtain cultures of dissociated granule cells at a low density, where individual cells can be easily observed. In addition, we include a protocol for studying neuronal migration in these cultures, using single, actively migrating cerebellar granule cells. Following this protocol, a factor of interest can be applied either in a gradient concentration by means of a micropipet located near the neuron, or in a homogeneous concentration by locally perfusing a certain region of the neuron. Time-lapse images are taken to analyze changes in the speed and/or directionality of the observed neuron. Overall, the two protocols take more or less a day and a half to perform, and are a useful way to evaluate a certain factor/drug for its chemotactic activity or its capacity to alter migration speed.

MeSH terms

  • Animals
  • Cell Culture Techniques / methods*
  • Cell Movement / physiology*
  • Cerebellum / cytology*
  • Culture Media, Conditioned / pharmacology*
  • HEK293 Cells
  • Humans
  • Neurons / cytology*
  • Rats
  • Rats, Sprague-Dawley

Substances

  • Culture Media, Conditioned