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Roscovitine inhibits CaV3.1 (T-type) channels by preferentially affecting closed-state inactivation.

ABSTRACT: T-type calcium channels (Ca(V)3) play an important role in many physiological and pathological processes, including cancerogenesis. Ca(V)3 channel blockers have been proposed as potential cancer treatments. Roscovitine, a trisubstituted purine, is a cyclin-dependent kinase (CDK) inhibitor that is currently undergoing phase II clinical trials as an anticancer drug and has been shown to affect calcium and potassium channel activity. Here, we investigate the effect of roscovitine on Ca(V)3.1 channels. Ca(V)3.1 channels were transiently expressed in human embryonic kidney 293 cells, and currents were recorded by using the whole-cell patch-clamp technique. Roscovitine blocks Ca(V)3.1 channels with higher affinity for depolarized cells (EC?? of 10 ?M), which is associated with a negative shift in the voltage dependence of closed-state inactivation. Enhanced inactivation is mediated by roscovitine-induced acceleration of closed-state inactivation and slowed recovery from inactivation. Small effects of roscovitine were also observed on T-channel deactivation and open-state inactivation, but neither could explain the inhibitory effect. Roscovitine inhibits Ca(V)3.1 channels within the therapeutic range (10-50 ?M) in part by stabilizing the closed-inactivated state. The ability of roscovitine to block multiple mediators of proliferation, including CDKs and Ca(V)3.1 channels, may facilitate its anticancer properties.

SUBMITTER: Yarotskyy V

PROVIDER: S-EPMC3263959 | biostudies-literature | 2012 Feb

REPOSITORIES: biostudies-literature

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Roscovitine inhibits CaV3.1 (T-type) channels by preferentially affecting closed-state inactivation.

Yarotskyy Viktor V Elmslie Keith S KS

The Journal of pharmacology and experimental therapeutics 20111116 2

T-type calcium channels (Ca(V)3) play an important role in many physiological and pathological processes, including cancerogenesis. Ca(V)3 channel blockers have been proposed as potential cancer treatments. Roscovitine, a trisubstituted purine, is a cyclin-dependent kinase (CDK) inhibitor that is currently undergoing phase II clinical trials as an anticancer drug and has been shown to affect calcium and potassium channel activity. Here, we investigate the effect of roscovitine on Ca(V)3.1 channe ...[more]

PMID: 22088954

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Project description:Kv4 channels mediate the somatodendritic A-type K+ current (I(SA)) in neurons. The availability of functional Kv4 channels is dynamically regulated by the membrane potential such that subthreshold depolarizations render Kv4 channels unavailable. The underlying process involves inactivation from closed states along the main activation pathway. Although classical inactivation mechanisms such as N- and P/C-type inactivation have been excluded, a clear understanding of closed-state inactivation in Kv4 channels has remained elusive. This is in part due to the lack of crucial information about the interactions between gating charge (Q) movement, activation, and inactivation. To overcome this limitation, we engineered a charybdotoxin (CTX)-sensitive Kv4.2 channel, which enabled us to obtain the first measurements of Kv4.2 gating currents after blocking K+ conduction with CTX (Dougherty and Covarrubias. 2006J. Gen. Physiol. 128:745-753). Here, we exploited this approach further to investigate the mechanism that links closed-state inactivation to slow Q-immobilization in Kv4 channels. The main observations revealed profound Q-immobilization at steady-state over a range of hyperpolarized voltages (-110 to -75 mV). Depolarization in this range moves <5% of the observable Q associated with activation and is insufficient to open the channels significantly. The kinetics and voltage dependence of Q-immobilization and ionic current inactivation between -153 and -47 mV are similar and independent of the channel's proximal N-terminal region (residues 2-40). A coupled state diagram of closed-state inactivation with a quasi-absorbing inactivated state explained the results from ionic and gating current experiments globally. We conclude that Q-immobilization and closed-state inactivation at hyperpolarized voltages are two manifestations of the same process in Kv4.2 channels, and propose that inactivation in the absence of N- and P/C-type mechanisms involves desensitization to voltage resulting from a slow conformational change of the voltage sensors, which renders the channel's main activation gate reluctant to open.

Dataset Information

Roscovitine inhibits CaV3.1 (T-type) channels by preferentially affecting closed-state inactivation.

Publications

Roscovitine inhibits CaV3.1 (T-type) channels by preferentially affecting closed-state inactivation.

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