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Assessment of Sphingosine-1-Phosphate Receptor Expression and Associated Intracellular Signaling Cascades in Primary Cells of the Human Central Nervous System

  • Veronique E. Miron
  • Bryce A. Durafourt
  • Jack P. Antel
  • Timothy E. KennedyEmail author
Protocol
Part of the Methods in Molecular Biology book series (MIMB, volume 874)

Abstract

Measuring the effects of sphingosine-1-phosphate (S1P) receptor modulators on human primary neural cells is of particular interest given the recent application of these central nervous system-accessible agents to the treatment of neurodegenerative diseases, such as multiple sclerosis. Issues to consider in experimental studies include the ability of some of these modulators to bind multiple S1P receptor subtypes simultaneously, the nonspecificity of commercially available S1P receptor antibodies, and activation of multiple intracellular signaling cascades by a given S1P receptor. Here, we discuss how to assay S1P receptor expression and activation using multiple agonists/antagonists, by linking the results of real-time reverse transcriptase polymerase chain reaction with the assessment of intracellular signaling derived from Western blot analyses.

Key words

Sphingosine-1-phosphate Sphingosine-1-phosphate receptor Intracellular signaling Central nervous system Human primary cells Real-time PCR Western blotting 

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Copyright information

© Springer Science+Business Media, LLC 2012

Authors and Affiliations

  • Veronique E. Miron
    • 1
  • Bryce A. Durafourt
    • 2
  • Jack P. Antel
    • 2
  • Timothy E. Kennedy
    • 3
    Email author
  1. 1.Centre for Multiple Sclerosis Research, Scottish Centre for Regenerative MedicineThe University of EdinburghEdinburghUK
  2. 2.Neuroimmunology Unit, The Montreal Neurological InstituteMcGill UniversityMontrealCanada
  3. 3.Centre for Neuronal Survival, The Montreal Neurological InstituteMcGill UniversityMontrealCanada

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