The haemolytic effect of verapamil on erythrocytes exposed to varying osmolarity

The haemolytic effect of verapamil on red blood cells (RBCs) exposed to varying osmolarity was investigated. The experimental approach used a modified red cell haemolysis assay with concentrations of verapamil ranging from 50-1500 μM compared to drug free controls. The time-course of haemolytic effects was also investigated. We also briefly determined the haemolytic effects of verapamil in Ca2+-free conditions (with added EGTA). In conditions representing decreasing osmolarity (dilution from 140-0 mM NaCl) there was a significant increase in erythrocyte haemolysis that was also dependent on verapamil concentration (ANOVA, p < 0.05). The red cells also showed a significantly increased rate of haemolysis over 5 h with increasing verapamil concentration (ANOVA, p < 0.05). The degree of RBC hypotonic haemolysis was significantly increased in a Ca2+-free medium (+EGTA) compared to normal saline and this effect was exacerbated by additions of verapamil (ANOVA, p < 0.05). Overall the data suggested that verapamil can cause haemolysis of RBCs in a predictable time- and concentration-dependent manner, and that verapamil increases the fragility of the erythrocytes further during hypotonic osmotic stress and Ca2+-free conditions. The mechanism of verapamil-dependent haemolysis could be directly related to the observed biphasic concentration-effect and could consequently involve several ion transport pathways.