Inhibition of Kv4.3/KChIP2.2 channels by bupivacaine and its modulation by the pore mutation Kv4.3V401I.
BACKGROUND
The transient outward current Ito is an important repolarizing K current in human ventricular myocardium mediated by Kv4.3 and KChIP2.2 subunits. Inhibition of Ito by amino-amide local anesthetics may be involved in severe cardiotoxic side effects. This study elucidates the molecular mechanisms of bupivacaine interaction with complexes formed by Kv4.3 and KChIP2.2 as well as the modulatory effect of KChIP2.2. For this purpose, the pharmacologic effects of bupivacaine on Kv4.3wt/KChIP2.2 channels and on the pore mutant Kv4.3V401I were investigated.


METHODS
Kv4.3/KChIP2.2 cDNA was transiently expressed in Chinese hamster ovary cells. Site-directed mutagenesis and patch clamp experiments were performed to analyze the effects of bupivacaine on wild-type and mutant channels.


RESULTS
Inhibition of Kv4.3wt/KChIP2.2 channels by bupivacaine was concentration-dependent and reversible. The IC50s for inhibition of the charge conducted by Kv4.3wt/KChIP2.2 channels by bupivacaine and levobupivacaine were 55 +/- 8 and 50 +/- 5 microm, respectively. The local anesthetic accelerated macroscopic current decline of Kv4.3wt/KChIP2.2 and slowed recovery from inactivation without altering steady state inactivation. KChIP2.2 altered the response of Kv4.3wt channels to bupivacaine and bupivacaine modulated KChIP2.2 effects on Kv4.3wt channels. The pore mutation V401I slowed macroscopic current decline of Kv4.3 channels and recovery from inactivation, and it diminished modulation of gating by KChIP2.2. Bupivacaine inhibition of Kv4.3V401I resembled Kv4.3wt and was not changed by coexpression of KChIP2.2.


CONCLUSIONS
These results indicate that bupivacaine blocks Kv4.3/KChIP2.2 channels from the open state. They furthermore give structural evidence that amino-amide local anesthetics interfere with the effects of KChIP2.2 on Kv4.3 by an indirect mechanism.
Solth
Anna
A
Department of Anesthesiology and Institute for Neural Signal Transduction, University Medical Center Hamburg-Eppendorf, Germany.
Siebrands
Cornelia C
CC
Friederich
Patrick
P
D000779:Anesthetics, Local / Q000494:pharmacology*; D000818:Animals; D002045:Bupivacaine / Q000494:pharmacology*; D016466:CHO Cells; D006224:Cricetinae; D004305:Dose-Response Relationship, Drug; D051605:Kv Channel-Interacting Proteins / Q000037:antagonists & inhibitors*; D016297:Mutagenesis, Site-Directed; D051672:Shal Potassium Channels / Q000037:antagonists & inhibitors* / Q000235:genetics
D000779:Anesthetics, Local; D051605:Kv Channel-Interacting Proteins; D051672:Shal Potassium Channels; D002045:Bupivacaine