Cytochalasin D reduces Ca2+ currents via cofilin-activated depolymerization of F-actin in guinea-pig cardiomyocytes.
1. L-type Ca2+ channel currents (I(Ca)) were measured in guinea-pig ventricular myocytes (22 degrees C, 300 ms steps from -45 to +10 mV). Pulsing at 0.5 Hz reduced I(Ca) within 5 min to 92 +/- 3% (mean +/- S.E.M., n = 14) and within 10 min to 83 +/- 4 % ('run-down' with reference to I(Ca) after a 5 min equilibration period). 2. Bath-applied cytochalasin D (cytD, 10 microM) reduced I(Ca) to 75 +/- 4% within 5 min and to 61 +/- 4% within 10 min ('cytD reduction of I(Ca)') by reduction of maximal Ca2+ conductance (suggested by fits of time course and of current-potential (I-V) curves). 3. Preincubation with phalloidin (bath applied, 100 microM, 5 h) prevented the cytD reduction of I(Ca). Since phalloidin specifically blocks F-actin depolymerization, cytD reduction of I(Ca) is linked to depolymerization of F-actin. 4. CytD did not attenuate the beta-adrenergic stimulation of I(Ca) (30 nM isoproterenol), suggesting that A kinase anchoring proteins are unlikely to mediate the cytD reduction of I(Ca). The cytD reduction of I(Ca) was abolished by extra-/intracellular acidosis (pH(o) 6.9), by cell dialysis of 5 mM BAPTA, or by serine/threonine protein phosphatase inhibitors. 5. Actin-depolymerizing factor (ADF)/cofilin are proteins that bind to actin, mediate a pH-sensitive depolymerization of F-actin, and are activated by dephosphorylation. Western blots from hearts perfused with solutions containing zero or 10 microM cytD indicated that cytD reduces the ratio of phosphorylated to total ADF/cofilin content by 50%. 6. The data support the concept that cytD mediates dephosphorylation and activation of ADF/cofilin, leading to depolymerization of F-actin with a subsequent reduction of I(Ca).
Rueckschloss
U
U
Department of Physiology, Faculty of Medicine, Martin-Luther-University, 06097 Halle, Germany. uwe.rueckschloss@medizin.uni-halle.de
Isenberg
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G
D000138:Acidosis / Q000378:metabolism; D051339:Actin Depolymerizing Factors; D000199:Actins / Q000378:metabolism*; D000818:Animals; D002118:Calcium / Q000378:metabolism; D020746:Calcium Channels, L-Type / Q000187:drug effects* / Q000502:physiology*; D002352:Carrier Proteins / Q000502:physiology; D002614:Chelating Agents / Q000494:pharmacology; D017868:Cyclic AMP-Dependent Protein Kinases / Q000502:physiology; D015638:Cytochalasin D / Q000494:pharmacology*; D004533:Egtazic Acid / Q000031:analogs & derivatives* / Q000494:pharmacology; D004553:Electric Conductivity; D006168:Guinea Pigs; D008840:Microfilament Proteins / Q000494:pharmacology*; D009206:Myocardium / Q000166:cytology / Q000378:metabolism*; D010590:Phalloidine / Q000494:pharmacology; D010749:Phosphoprotein Phosphatases / Q000502:physiology; D011108:Polymers / Q000378:metabolism
D051339:Actin Depolymerizing Factors; D000199:Actins; D020746:Calcium Channels, L-Type; D002352:Carrier Proteins; D002614:Chelating Agents; D008840:Microfilament Proteins; D011108:Polymers; D010590:Phalloidine; D015638:Cytochalasin D; D004533:Egtazic Acid; D017868:Cyclic AMP-Dependent Protein Kinases; D010749:Phosphoprotein Phosphatases; C025603:1,2-bis(2-aminophenoxy)ethane-N,N,N',N'-tetraacetic acid; D002118:Calcium