Roscovitine, a cyclin-dependent kinase inhibitor, affects several gating mechanisms to inhibit cardiac L-type (Ca(V)1.2) calcium channels.
OBJECTIVE
L-type calcium channels (Ca((V))1.2) play an important role in cardiac contraction. Roscovitine, a cyclin-dependent kinase inhibitor and promising anticancer drug, has been shown to affect Ca((V))1.2 by inhibiting current amplitude and slowing activation. This research investigates the mechanism by which roscovitine inhibits Ca((V))1.2 channels.


METHODS
Ca((V))1.2 channels were transfected into HEK 293 cells, using the calcium phosphate precipitation method, and currents were measured using the whole-cell patch clamp technique.


RESULTS
Roscovitine slows activation at all voltages, which precludes one previously proposed mechanism. In addition, roscovitine enhances voltage-dependent, but not calcium-dependent inactivation. This enhancement resulted from both an acceleration of inactivation and a slowing of the recovery from inactivation. Internally applied roscovitine failed to affect Ca((V))1.2 currents, which supports a kinase-independent mechanism and extracellular binding site. Unlike the dihydropyridines, closed state inactivation was not affected by roscovitine. Inactivation was enhanced in a dose-dependent manner with an IC(50)=29.5+/-12 microM, which is close to that for slow activation and inhibition.


CONCLUSIONS
We conclude that roscovitine binds to an extracellular site on Ca((V))1.2 channels to inhibit current by both slowing activation and enhancing inactivation. Purine-based drugs could become a new option for treatment of diseases that benefit from L-channel inhibition such as cardiac arrhythmias and hypertension.
Yarotskyy
V
V
Department of Anesthesiology, Penn State College of Medicine, Penn State University, Hershey, PA 17033, USA.
Elmslie
K S
KS
D001665:Binding Sites; D020746:Calcium Channels, L-Type / Q000187:drug effects* / Q000378:metabolism; D002130:Calcium Phosphates / Q000378:metabolism; D002318:Cardiovascular Diseases / Q000188:drug therapy; D002460:Cell Line; D011232:Chemical Precipitation; D018844:Cyclin-Dependent Kinases / Q000037:antagonists & inhibitors*; D004594:Electrophysiology; D006801:Humans; D015640:Ion Channel Gating / Q000187:drug effects*; D018408:Patch-Clamp Techniques; D047428:Protein Kinase Inhibitors / Q000494:pharmacology*; D011687:Purines / Q000494:pharmacology*; D000077546:Roscovitine; D014162:Transfection
D020746:Calcium Channels, L-Type; D002130:Calcium Phosphates; C419385:L-type calcium channel alpha(1C); D047428:Protein Kinase Inhibitors; D011687:Purines; D000077546:Roscovitine; C020243:calcium phosphate; D018844:Cyclin-Dependent Kinases
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