Ca currents in human neuroblastoma IMR32 cells: kinetics, permeability and pharmacology

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Abstract
We have investigated the kinetics, permeability and pharmacological properties of Ca channels in in vitro differentiated IMR32 human neuroblastoma cells. The lowthreshold (LVA, T) Ca current activated positive to −50 mV and inactivated fully within 100 ms in a voltage-dependent manner. This current persisted in the presence of 3.2 μM ω-conotoxin (ω-CgTx) or 40 μM Cd and showed a weaker sensitivity to Ni and amiloride than in other neurons. The high-threshold Ca currents (HVA,L and N) turned on positive to −30 mV, and inactivated slowly and incompletely during pulses of 200 ms duration. The amplitude of the HVA currents and the number of 125I-ω-CgTx binding sites increased markedly during cell differentiation. In agreement with recent reports, 6.4 μM ω-CgTx blocked only about 85% of the Ba currents through HVA channels in 50% of the cells. Residual ω-CgTx-resistant currents proved to be more sensitive to dihydropyridines (DHP) than total HVA currents. Bay K 8644 (1 μM) had a clear agonistic action on ω-CgTx-resistant currents and was preferred to other Ca antagonists for identifying HVA DHP-sensitive channels. Compared to the ω-CgTx-sensitive, the DHP-sensitive currents turned on at slightly more negative potentials and showed a weaker sensitivity to voltage. The two HVA currents were otherwise hardly distinguishable in terms of activation/inactivation kinetics, Ca/Ba permeability and sensitivity to holding potentials. This suggests that currently used criteria for identifying multiple types of neuronal Ca channels (T,L,N) may be widely misleading if not supported by pharmacological assays.