Hydrogen Peroxide Inhibits Chloride Channels of the Sarcoplasmic Reticulum of Skeletal Muscle |
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Authors: | JI Kourie |
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Institution: | (1) Membrane Transport Group, Department of Chemistry, The Faculties, The Australian National University, Canberra City, ACT, 0200 Australia, AU |
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Abstract: | Data obtained with the lipid bilayer technique indicate that cis (cytoplasmic) concentration of 4.4–22 mm hydrogen peroxide (H2O2), is a water-soluble oxidant. H2O2]
cis
(n= 26) reversibly inhibits the multisubconductance SCl channel of the sarcoplasmic reticulum vesicles from rabbit skeletal
muscle. At −40 mV, the mean values of the current amplitude (I) and the probability of the SCl channel being open (P
o
) were reduced significantly (n= 8) from −6.14 ± 0.42 pA and 0.69 ± 0.06 (for all conductance levels) in control 0.0 mm H2O2]
cis
to −1.10 ± 0.51 pA and 0.13 ± 0.04 (for the intermediate subconductance states) in 8.8 mm H2O2]
cis
, respectively. The H2O2]
cis
-induced decrease in P
o
is mainly due to a decrease in the mean open time T
o
. The mechanism of H2O2]
cis
effects on the multiconductance SCl channel is characterized by a mode shift in the channel state from the main conductance
state to the low subconductance states. The estimated concentration of the H2O2]
cis
for the half inhibitory constant, K
i
, was 11.78 mm, higher than the estimated 8.0 and 8.1 mm for the parameters P
o
and T
o
, respectively, indicating that the conductance of the SCl channel is less sensitive than the gating kinetics of the channel.
After a lag period of between 30 to 60 sec, the lipophilic SH-oxidizing agent 4,4′-dithiodipyridine (4,4′-DTDP) added to the
cis side at 1.0 mm removed the inhibitory effects of 8.8 mm H2O2]
cis
. The 4,4′-DTDP-enhanced SCl channel activity was blocked after the addition of 0.5 mm ATP to the cis side of the channel. The addition of 1.0 mm 4,4′-DTDP to the cis or trans solutions facing an SCl channel already subjected to 0.5 mm ATP]
cis
or ATP]
trans
failed to activate the ATP-inhibited SCl channel. These findings suggest that 4,4′-DTDP is not preventing the binding of
ATP to its binding site on the channel protein. The interaction of H2O2 with the SCl channel proteins is consistent with a thiol-disulfide redox state model for regulating ion transport, where
SH groups can directly modify the function of the channel and/or the availability of regulatory sites on the channel proteins.
The H2O2 effects on the Ca2+ countercurrent through the SCl channel are also consistent with H2O2-modification of the mechanisms involved in the Ca2+ regulation, which underlies excitation-contraction coupling in skeletal muscle.
Received: 27 April 1999/Revised: 1 July 1999 |
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Keywords: | : Reactive oxygen species — Sulfhydryl group (SH)-oxidizing and SH-reducing agents — ATP-sensitive channels — Bilayer technique — Calcium countercurrent |
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