Calmodulin inhibitors chlorpromazine and calmidazolium cause calcium overload and oxidative stress despite reducing peak density of CaV1.2 current in ventricular myocytes.

Calmodulin (CaM) plays crucial roles in the regulation of the voltage-dependent L-type calcium (Ca2+) channel α1C subunit (CaV1.2) channel opening and closing, which is closely associated with the occurrence and progression of cardiac diseases. However, the specific mechanisms by which CaM regulates CaV1.2 channel Ca2+ currents have not been well established experimentally, and whether CaM can be a therapeutic target for cardiovascular diseases deserves further study. In this study, we investigated the effects of CaM inhibitors chlorpromazine (CPZ) and calmidazolium (CMZ) on CaV1.2 channel function through a series of experiments. Molecular Operating Environment (MOE) docking analysis suggested that CPZ and CMZ alter the 3D conformation and binding sites between CaM and CaV1.2 channel. Furthermore, GST pull-down assays revealed that CPZ and CMZ significantly decreased the binding ability of CaM to CaV1.2 channel Ca2+-dependently. Patch clamp assays performed at the whole-cell level indicated that CPZ and CMZ induced marked electrical dysfunction in cardiomyocytes. including significantly reduced amplitude and delayed inactivation of the CaV1.2 currents. Additionally, confocal imaging analysis demonstrated that inhibition of CaM increased intracellular Ca2+ concentration ([Ca2+]) in response to KCl stimulation and increased basal reactive oxygen species (ROS), emphasizing CaM's critical role in maintaining intracellular Ca2+ homeostasis and redox balance. In conclusion, our data support the view that CaM has a significant regulatory effect on the electrophysiological properties of CaV1.2 channel, and CaM inhibitors can interfere with this regulatory effect.