cytochrome-c-t and calmidazolium

The intracellular calcium concentration ([Ca](i)) regulates cell viability and contractility in myocardial cells. Elevation of the [Ca](i) level occurs by entry of calcium ions (Ca(2+)) through voltage-dependent Ca(2+) channels in the plasma membrane and release of Ca(2+) from the sarcoplasmic reticulum. Calmidazolium chloride (CMZ), a subgroup II calmodulin antagonist, blocks L-type calcium channels as well as voltage-dependent Na(+) and K(+) channel currents. This study elaborates on the events that contribute to the cytotoxic effects of CMZ on the heart. We hypothesized that apoptotic cell death occurs in the cardiac cells through calcium accumulation, production of reactive oxygen species, and the cytochrome c-mediated PARP activation pathway. CMZ significantly increased the production of superoxide (O(2)(*-)) and nitric oxide (NO) as detected by FACS and confocal microscopy. CMZ induced mitochondrial damage by increasing the levels of intracellular calcium, lowering the mitochondrial membrane potential, and thereby inducing cytochrome c release. Apoptotic cell death was observed in H9c2 cells exposed to 25 microM CMZ for 24 h. This is the first report that elaborates on the mechanism of CMZ-induced cardiotoxicity. CMZ causes apoptosis by decreasing mitochondrial activity and contractility indices and increasing oxidative and nitrosative stress, ultimately leading to cell death via an intrinsic apoptotic pathway.

Topics: Analgesics; Animals; Apoptosis; Calcium; Calmodulin; Cell Line; Cell Separation; Chlorides; Cytochromes c; Flow Cytometry; Imidazoles; Membrane Potential, Mitochondrial; Microscopy, Confocal; Mitochondria, Heart; Myocardial Contraction; Myocytes, Cardiac; Nitric Oxide; Oxidative Stress; Rats; Superoxides

Defects in proteasome function have been suggested to be involved in the pathogenesis of neurodegenerative diseases. We examined the effect of calmodulin antagonists on proteasome inhibitor-induced mitochondrial dysfunction and cell viability loss in undifferentiated PC12 cells. Caspase inhibitors (z-IETD.fmk, z-LEHD.fmk and z-DQMD.fmk) and antioxidants attenuated cell death and decrease in GSH contents in PC12 cells treated with 20 microM MG132, a proteasome inhibitor. Calmodulin antagonists (trifluoperazine, W-7 and calmidazolium) had a differential inhibitory effect on the MG132-induced cell death and GSH depletion depending on concentration with a maximal inhibitory effect at 0.5-1 microM. Addition of trifluoperazine and W-7 reduced the MG132-induced nuclear damage, loss of the mitochondrial transmembrane potential followed by cytochrome c release, formation of reactive oxygen species and elevation of intracellular Ca(2+) levels in PC12 cells. Calmodulin antagonists at 5 microM exhibited a cytotoxic effect on PC12 cells but attenuated the cytotoxicity of MG132. The results suggest that the toxicity of MG132 on PC12 cells is mediated by activation of caspase-8, -9 and -3. Trifluoperazine and W-7 at the concentrations of 0.5-1 microM may attenuate the MG132-induced viability loss in PC12 cells by suppressing change in the mitochondrial membrane permeability and by lowering of the intracellular Ca(2+) levels as well as calmodulin inhibition.

Topics: Animals; Calcium; Calmodulin; Caspase 3; Caspases; Cell Count; Cell Death; Cell Size; Cell Survival; Cytochromes c; Dose-Response Relationship, Drug; Drug Interactions; Enzyme Inhibitors; Glutathione; Imidazoles; Leupeptins; Mitochondrial Diseases; PC12 Cells; Rats; Reactive Oxygen Species; Sulfonamides; Trifluoperazine