The antifungal imidazole clotrimazole and its major in vivo metabolite are potent blockers of the calcium-activated potassium channel in murine erythroleukemia cells.
Clotrimazole (CLT), a member of the antifungal imidazole family of compounds, has been found to inhibit both calcium (Ca2+)-activated 86Rb and potassium (K) fluxes of human red cells and to inhibit red cell binding of 125I-charybdotoxin (ChTX) [11]. We have now used patch-clamp techniques to demonstrate reversible inhibition of whole cell KCa2+ currents in murine erythroleukemia (MEL) cells by submicromolar concentrations of CLT. Inhibition was equivalent whether currents were elicited by bath application of the Ca2+ ionophore A23187 or by dialyzing cells with a pipette solution containing micromolar concentrations of free Ca2+. The extent of inhibition of whole cell MEL KCa2+ currents was voltage-dependent, decreasing with increasing test potential. We also determined the single channel basis of the CLT inhibition in MEL cells by demonstrating the inhibition of a calcium-activated, ChTX-sensitive K channel by CLT in outside-out patches. The channel was also blocked by the des-imidazolyl metabolite of CLT, 2-chlorophenyl-bisphenyl-methanol (MET II) [15], thus demonstrating that the imidazole ring is not required for the inhibitory action of CLT. Single KCa2+ channels were also evident in inside-out patches of MEL cells. Block of K current by CLT was not unique to MEL cells. CLT also inhibited a component of the whole cell K current in PC12 cells. Channel specificity of block by CLT was determined by examining its effects on other types of voltage-sensitive currents. CLT block showed the following rank order of potency: K currents in PC12 cells > Ca2+ currents in PC12 cells >> Na currents in sympathetic neurons. These results demonstrate that direct inhibition of single KCa2+ by CLT can be dissociated from inhibition of cytochrome P-450 in MEL cells.
Rittenhouse
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Molecular Medicine & Renal Units, Beth Israel Hospital, 330 Brookline Ave., Boston, MA 02215, USA.
Vandorpe
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DH
Brugnara
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Alper
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D000818:Animals; D000831:Animals, Newborn; D000935:Antifungal Agents / Q000494:pharmacology*; D000001:Calcimycin / Q000494:pharmacology; D002118:Calcium / Q000378:metabolism*; D003022:Clotrimazole / Q000494:pharmacology*; D007476:Ionophores / Q000494:pharmacology; D004915:Leukemia, Erythroblastic, Acute / Q000378:metabolism*; D051379:Mice; D016716:PC12 Cells / Q000378:metabolism; D018408:Patch-Clamp Techniques; D015221:Potassium Channels / Q000187:drug effects* / Q000378:metabolism; D051381:Rats; D017208:Rats, Wistar; D017783:Superior Cervical Ganglion / Q000166:cytology / Q000187:drug effects; D014407:Tumor Cells, Cultured
D000935:Antifungal Agents; D007476:Ionophores; D015221:Potassium Channels; D000001:Calcimycin; D003022:Clotrimazole; D002118:Calcium