Characterization of Endogenous Sodium Channels in the ND7-23 Neuroblastoma Cell Line: Implications for Use as a Heterologous Ion Channel Expression System Suitable for Automated Patch Clamp Screening
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The rodent neuroblastoma cell line, ND7-23, is used to express voltage-dependent sodium (Nav) and other neuronal ion channels resistant to heterologous expression in Chinese hamster ovary (CHO) or human embryonic kidney (HEK) cells. Their advantage is that they provide endogenous factors and signaling pathways to promote ion channel peptide folding, expression, and function at the cell surface and are also amenable to automated patch clamping. However, ND7-23 cells exhibit endogenous tetrodotoxin (TTX)-sensitive Nav currents, and molecular profiling has revealed the presence of Nav1.2, Nav1.3, Nav1.6, and Nav1.7 transcripts, but no study has determined which subtypes contribute to functional channels at the cell surface. We profiled the repertoire of functional Nav channels endogenously expressed in ND7-23 cells using the QPatch automated patch clamp platform and selective toxins and small molecules. The potency and subtype selectivity of the ligands (Icagen compound 68 from patent US-20060025415-A1-20060202, 4,9 anhydro TTX, and Protoxin-II) were established in human Nav1.3, Nav1.6, and Nav1.7 channel cell lines before application of selective concentrations to ND7-23 cells. Our data confirm previous studies that >97% of macroscopic Nav current in ND7-23 cells is carried by TTX-sensitive channels (300 nM TTX) and that Nav1.7 is the predominant channel contributing to this response (65% of peak inward current), followed by Nav1.6 (∼20%) and negligible Nav1.3 currents (∼2%). In addition, our data are the first to assess the Nav1.6 potency (50% inhibitory concentration [IC50] of 33 nM) and selectivity (50-fold over Nav1.7) of 4,9 anhydro TTX in human Nav channels expressed in mammalian cells, confirming previous studies of rodent Nav channels expressed in oocytes and HEK cells.