On the Role of Ca 2+ - and Voltage-Dependent Inactivation in Ca v 1.2 Sensitivity for the Phenylalkylamine (-)Gallopamil

Abstract

Abstract— L-type calcium channels (Ca v 1.m) inactivate in response to elevation of intracellular Ca 2+ (Ca 2+ -dependent inactivation) and additionally by conformational changes induced by membrane depolarization (fast and slow voltage-dependent inactivation). Molecular determinants of inactivation play an essential role in channel inhibition by phenylalkylamines (PAAs). The relative impacts, however, of Ca 2+ -dependent and voltage-dependent inactivation in Ca v 1.2 sensitivity for PAAs remain unknown. In order to analyze the role of the different inactivation processes, we expressed Ca v 1.2 constructs composed of different β-subunits (β 1a -, β 2a -, or β 3 -subunit) in Xenopus oocytes and estimated their (-)gallopamil sensitivity by means of the two-microelectrode voltage clamp with either Ba 2+ or Ca 2+ as charge carrier. Ca v 1.2 consisting of the β 2a -subunit displayed the slowest inactivation and the lowest apparent sensitivity for the PAA (-)gallopamil. A significantly higher apparent (-)gallopamil-sensitivity with Ca 2+ as charge carrier was observed for all 3 β-subunit compositions. The kinetics of Ca 2+ -dependent inactivation and slow voltage-dependent inactivation were not affected by drug. The higher sensitivity of the Ca v 1.2 channels for (-)gallopamil with Ca 2+ as charge carrier results from slower recovery (τ rec,Ca ≈15 seconds versus τ rec,Ba ≈3 to 5 seconds) from a PAA-induced channel conformation. We propose a model where (-)gallopamil promotes a fast voltage-dependent component in Ca v 1.2 inactivation. The model reproduces the higher drug sensitivity in Ca 2+ as well as the lower sensitivity of slowly inactivating Ca v 1.2 composed of the β 2a -subunit.