Action-potential-like depolarizations relieve opioid inhibition of N-type Ca²⁺ channels in NG108–15 cells

Abstract

 The ability of action-potential-like waveforms (APWs) to attenuate opioid-induced inhibition of N-type Ca²⁺ channels was investigated in the neuroblastoma × glioma cell line NG108–15 using whole-cell voltage clamp methods. In in vitro differentiated NG108–15 cells, the opioid agonist [d-ala²]-methionine-enkephalin (DAME) reversibly decreased ω-conotoxin-GVIA-sensitive Ba²⁺ currents (N-type currents). Agonist-mediated inhibition of N-type currents could be transiently relieved by strong unphysiological depolarizing prepulses to +80 mV (facilitation). Significant facilitation was also achieved by conditioning the cell with a train of 15 APWs, which roughly mimicked physiological action potentials (1- to 6-ms-long depolarizations to +30 mV from a holding potential of –40 mV). The APW-induced facilitation depended on both conditioning pulse frequency and duration. Summation of the disinhibition produced by each APW was possible because reinhibition following repolarization to –40 mV was a much slower process (τ=88 ms) than the onset of facilitation at +80 mV (τ=7 ms). These results provide evidence that N-type Ca²⁺ channel facilitation may be a physiologically relevant process, and suggest that neuronal firing may relieve agonist-induced inhibition of N-type currents to an extent depending on both the shape of action potentials and the frequency of firing.