Voltage‐Sensitive Calcium Channels in Differentiated Neuroblastoma × Glioma Hybrid (NG108–15) Cells: Characterization by Quin 2 Fluorescence

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Abstract
Depolarization of differentiated neuroblastoma × glioma (NG108–15) cells with KC1 (50 mM) or veratridine (50 μM) stimulated Ca2+ accumulation, was detected by quin 2 fluorescence. Intracellular Ca2+ concentrations ([Ca2+]i) were elevated about threefold from 159 ± 7 to 595 ± 52 nM (n = 12). Ca2+ entry evoked by high extracellular K+ concentration ([K+]o) was voltage‐dependent and enhanced by the dihydropyridine agonists, BAY K 8644 and CGP 28 392, in a dose‐dependent manner. CGP 28 392 was less potent and less efficacious than BAY K 8644. The (+) and (‐) stereoisomers of 202–791 showed agonist and antagonist properties, respectively. (+)‐202–791 was less potent, but as efficacious as BAY K 8644. In the absence of KCl, BAY K 8644 had no effect on Ca2+ entry. Voltage‐sensitive calcium channel (VSCC) activity was blocked by organic Ca2+ channel antagonists (nanomolar range) both before and after KC1 treatment and also by divalent metal cations (micromolar range). High [K+]o‐in‐duced Ca2+ accumulation was dependent on external Ca2+, but not on external Na+ ions ([Na]o), and was insensitive to both tetrodotoxin (3 μM) and tetraethylammonium (10 μM). In contrast, veratridine‐induced Ca2+ accumulation required [Na+]o, and was blocked by tetrodotoxin, but not by nimodipine (1 μM). Veratridine‐induced Ca2+ accumulation was slower (∼45 s), smaller in magnitude (∼30% of [K+]o‐induced Ca2+ entry), and also enhanced by BAY K 8644 (∼50%). VSCC were identified in neuronal hybrid (NG108–15 and NCB‐20) cells, but not in glial (C6BU‐1), renal epithelial (MDCK), and human astrocytoma (1321N1) cells. NG108–15 cells differentiated with 1.0 mM dibutyryl cyclic AMP showed greater VSCC activity than undifferentiated cultures. These results suggest that cultured neural cells provide a useful system to study Ca2+ regulation via ion channels.