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

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 Ca-v channel subtypes essential for regulation of important physiological functions, such as heart contraction and nociception, including the clinically validated pain target Ca(v)2.2. We have detected mRNA transcripts for a range of endogenous expressed subtypes Ca(v)1.3, Ca(v)2.2 (including two Ca(v)1.3, and three Ca(v)2.2 splice variant isoforms) and Ca(v)3.1 in SH-SY5Y cells; as well as Ca-v auxiliary subunits alpha(2)delta(1-3), beta(1), beta(3), beta(4), gamma(1), gamma(4-5), and gamma(7). Both high-and low-voltage activated Ca-v channels generated calcium signals in SH-SY5Y cells. Pharmacological characterisation using omega-conotoxins CVID and MVIIA revealed significantly (similar to 10-fold) higher affinity at human versus rat Ca(v)2.2, while GVIA, which interacts with Ca(v)2.2 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 Ca(v)2.2 channels pharmacology. These results may have implications for strategies used to identify therapeutic leads at Ca(v)2.2 channels