Thiol-dependent Redox Mechanisms in the Modification of ATP-Sensitive Potassium Channels in Rabbit Ventricular Myocytes.
- Author:
Jin HAN
1
;
Nari KIM
;
Dang Van CUONG
;
Chunghui KIM
;
Euiyong KIM
Author Information
1. Department of Physiology and Biophysics, College of Medicine, Inje University, Busan 614 735, Korea. phyhanj@ijnc.inje.ac.kr
- Publication Type:Original Article
- Keywords:
KATP channel;
Patch-clamp technique;
Rabbit ventricular myocytes;
Thiol redox
- MeSH:
Adenosine Triphosphate;
Collagenases;
Cytoplasm;
Dithionitrobenzoic Acid;
Dithiothreitol;
Ethylmaleimide;
Hand;
Ischemia;
KATP Channels*;
Membranes;
Muscle Cells*;
Oxidants;
Oxidation-Reduction*;
Patch-Clamp Techniques;
Perfusion
- From:The Korean Journal of Physiology and Pharmacology
2003;7(1):15-23
- CountryRepublic of Korea
- Language:English
-
Abstract:
Cellular redox state is known to be perturbed during ischemia and that Ca2+ and K+ channels have been shown to have functional thiol groups. In this study, the properties of thiol redox modulation of the ATP-sensitive K+ (KATP) channel were examined in rabbit ventricular myocytes. Rabbit ventricular myocytes were isolated using a Langendorff column for coronary perfusion and collagenase. Single-channel currents were measured in excised membrane patch configuration of patch-clamp technique. The thiol oxidizing agent 5, 5'-dithio-bis- (2-nitro-benzoic acid) (DTNB) inhibited the channel activity, and the inhibitory effect of DTNB was reversed by dithiothreitol (disulfide reducing agent; DTT). DTT itself did not have any effect on the channel activity. However, in the patches excised from the metabolically compromised cells, DTT increased the channel activity. DTT had no effect on the inhibitory action by ATP, showing that thiol oxidation was not involved in the blocking mechanism of ATP. There were no statistical difference in the single channel conductance for the oxidized and reduced states of the channel. Analysis of the open and closed time distributions showed that DTNB had no effect on open and closed time distributions shorter than 4 ms. On the other hand, DTNB decreased the life time of bursts and increased the interburst interval. N-ethylmaleimide (NEM), a substance that reacts with thiol groups of cystein residues in proteins, induced irreversible closure of the channel. The thiol oxidizing agents (DTNB, NEM) inhibited of the KATP channel only, when added to the cytoplasmic side. The results suggested that metabolism-induced changes in the thiol redox can also modulate KATP channel activity and that a modulatory site of thiol redox may be located on the cytoplasmic side of the KATP channel in rabbit ventricular myocytes.
