Expression and pharmacology of endogenous Cav channels in SH-SY5Y human neuroblastoma cells

PLoS One. 2013;8(3):e59293. doi: 10.1371/journal.pone.0059293. Epub 2013 Mar 25.

Abstract

SH-SY5Y human neuroblastoma cells provide a useful in vitro model to study the mechanisms underlying neurotransmission and nociception. These cells are derived from human sympathetic neuronal tissue and thus, express a number of the Cav channel subtypes essential for regulation of important physiological functions, such as heart contraction and nociception, including the clinically validated pain target Cav2.2. We have detected mRNA transcripts for a range of endogenous expressed subtypes Cav1.3, Cav2.2 (including two Cav1.3, and three Cav2.2 splice variant isoforms) and Cav3.1 in SH-SY5Y cells; as well as Cav auxiliary subunits α2δ1-3, β1, β3, β4, γ1, γ4-5, and γ7. Both high- and low-voltage activated Cav channels generated calcium signals in SH-SY5Y cells. Pharmacological characterisation using ω-conotoxins CVID and MVIIA revealed significantly (∼ 10-fold) higher affinity at human versus rat Cav2.2, while GVIA, which interacts with Cav2.2 through a distinct pharmacophore had similar affinity for both species. CVID, GVIA and MVIIA affinity was higher for SH-SY5Y membranes vs whole cells in the binding assays and functional assays, suggesting auxiliary subunits expressed endogenously in native systems can strongly influence Cav2.2 channels pharmacology. These results may have implications for strategies used to identify therapeutic leads at Cav2.2 channels.

Publication types

  • Research Support, Non-U.S. Gov't

MeSH terms

  • Binding, Competitive
  • Calcium Channel Blockers / metabolism
  • Calcium Channels / genetics*
  • Calcium Channels / metabolism*
  • Cell Line, Tumor
  • Cell Membrane / metabolism
  • Gene Expression*
  • Humans
  • Neuroblastoma / genetics*
  • Neuroblastoma / metabolism*
  • Protein Binding
  • Protein Isoforms
  • omega-Conotoxin GVIA / metabolism

Substances

  • Calcium Channel Blockers
  • Calcium Channels
  • Protein Isoforms
  • omega-Conotoxin GVIA

Grant support

This work was supported by an NHMRC Program Grant (RJL), an UQIRTA scholarship to SRS and NHMRC fellowships to RJL and IV. The FLIPR was supported by an ARC Linkage Grant. The funders had no role in study design, data collection and analysis, decision to publish, or preparation of the manuscript.