Application of Modulators of Ca²⁺-Activated Big-Conductance Potassium Channels Against Cd²⁺-Induced Cytotoxicity: A...

As we found earlier, paxilline (a Penicillium paxilli mycotoxin and blocker of Ca²⁺-activated big-conductance potassium channels, BK(Ca)s) attenuated Cd²⁺-induced cytotoxic effects, whereas BK(Ca) activators (NS004, NS1619) and Cd²⁺ were able to induce apoptosis, which was enhanced when used together. In this work, molecular mechanisms underlying the aforementioned effects were studied using two rat cell lines, PC12 and AS-30D, flow cytometry, and spectrofluorometric and polarographic techniques. Both NS004 and NS1619 were found to have time- and dose-dependent effects on cell viability, respiration, mitochondrial membrane potential, and intracellular reactive oxygen species (ROS) production. In PC12 cells, BK(Ca) openers exerted an uncoupling effect after 3 h, increasing the resting respiration, while they partially inhibited the maximal respiration after 5 and 24 h; in addition, after 3 h, a transient protection by NS004/NS1619 against Cd²⁺-induced decrease of cell viability was observed. In both cell types, NS004/NS1619 increased ROS production after 3 h and counteracted the mitigating effect of paxilline against Cd²⁺-induced necrosis. In turn, paxilline reduced NS004/NS1619-induced apoptosis in AS-30D cells and ROS increase produced by NS004/NS1619 and/or Cd²⁺ in PC12 cells. As a result, the involvement of the mitochondrial respiratory chain, ROS, and, very likely, BK(Ca)s, in the mechanisms of the modulatory effects of the BK(Ca) blocker/opener(s) used in the absence and presence of Cd²⁺ was revealed.