sq-23377 and pyrvinium

Pharmacological modification of protein kinase CK1 (casein kinase 1) has previously been shown to influence suicidal erythrocyte death or eryptosis, which is triggered by activation of Cl(-)-sensitive Ca(2+)-permeable cation channels. Ca(2+) entering through those channels stimulates cell membrane scrambling and opens Ca(2+)-activated K(+)-channels resulting in KCl exit and thus cell shrinkage. The specific CK1-inhibitor D4476 (1 µM) blunted, whereas the specific CK1 αactivator pyrvinium pamoate (10 µM) enhanced cell membrane scrambling. The substances were at least partially effective through modification of cytosolic Ca(2+)-activity. The present study explored, whether pyrvinium pamoate indeed influences Cl(-)-sensitive cation-channels in erythrocytes. As a result, removal of Cl(-)increased Fluo3-fluorescence (reflecting cytosolic Ca(2+)-activity), triggered cell membrane scrambling (apparent from annexin-V-binding), and decreased forward scatter (pointing to cell shrinkage). Pyrvinium pamoate significantly augmented the effect of Cl(-)-removal on Fluo3 fluorescence and annexin-V-binding, but blunted the effect on forward scatter. According to whole cell patch clamp recording, Cl(-)removal activated a cation current, which was significantly enhanced by pyrvinium pamoate. Pyrvinium pamoate inhibited Ca(2+)-activated K(+)-channels. Ca(2+)-ionophore ionomycin (1 µM) decreased forward scatter, an effect significantly blunted by pyrvinium pamoate. In conclusion, pyrvinium pamoate activates Cl(-)-sensitive Ca(2+)-permeable cation channels with subsequent Ca(2+)-entry and inhibits Ca(2+)-activated K(+)-channels thus blunting the stimulating effect of Ca(2+) on those channels, K(+)-exit and thus cell shrinkage.

Topics: Aniline Compounds; Annexin A5; Calcium; Casein Kinase Ialpha; Cations; Cell Membrane; Cell Size; Electrophysiological Phenomena; Erythrocytes; Humans; Ion Channels; Ionomycin; Patch-Clamp Techniques; Potassium Channels, Calcium-Activated; Protein Binding; Pyrvinium Compounds; Xanthenes