Inhibitory effect of DIDS, NPPB, and phloretin on intracellular chloride channels.
We studied the effects of the chloride channel blockers, 5-nitro-2-(phenylpropylamino)-benzoate (NPPB), dihydro-4,4' diisothiocyanostilbene-2,2'-disulphonic acid (DIDS), and phloretin on H2O2-induced primary culture cardiomyocyte apoptosis and activity of intracellular chloride channels obtained from rat heart mitochondrial and lysosomal vesicles. The chloride channel blockers (100 micromol/l) inhibited the H2O2-induced cardiomyocytes apoptosis. We characterized the effect of the blockers on single channel properties of the chloride channels derived from the mitochondrial and lysosomal vesicles incorporated into a bilayer lipid membrane. The single chloride channel currents were measured in 250:50 mmol/l KCl cis/trans solutions. NPPB, DIDS, and phloretin inhibited the chloride channels by decreasing the channel open probability in a concentration-dependent manner with EC50 values of 42, 7, and 20 micromol/l, respectively. NPPB and phloretin inhibited the channel's conductance and open dwell time, indicating that they could affect the chloride selective filter, pore permeability, and gating mechanism of the chloride channels. DIDS and NPPB inhibited the channels from the other side than bongkrekic acid and carboxyatractyloside. The results may contribute to understand a possible involvement of intracellular chloride channels in apoptosis and cardioprotection.
Malekova
Lubica
L
Institute of Molecular Physiology and Genetics, Centre of Excellence for Cardiovascular Research, Slovak Academy of Sciences, Vlarska 5, 833 34 Bratislava, Slovak Republic.
Tomaskova
Jana
J
Novakova
Marie
M
Stefanik
Peter
P
Kopacek
Juraj
J
Lakatos
Boris
B
Pastorekova
Silvia
S
Krizanova
Olga
O
Breier
Albert
A
Ondrias
Karol
K
D017878:4,4'-Diisothiocyanostilbene-2,2'-Disulfonic Acid / Q000494:pharmacology*; D000818:Animals; D017209:Apoptosis / Q000187:drug effects; D002478:Cells, Cultured; D018118:Chloride Channels / Q000037:antagonists & inhibitors* / Q000187:drug effects / Q000378:metabolism; D004305:Dose-Response Relationship, Drug; D004594:Electrophysiology; D006861:Hydrogen Peroxide / Q000494:pharmacology; D008247:Lysosomes / Q000378:metabolism; D008297:Male; D008929:Mitochondria, Heart / Q000378:metabolism; D032383:Myocytes, Cardiac / Q000166:cytology / Q000187:drug effects*; D009579:Nitrobenzoates / Q000494:pharmacology*; D010693:Phloretin / Q000494:pharmacology*; D051381:Rats; D017208:Rats, Wistar
D018118:Chloride Channels; D009579:Nitrobenzoates; C058176:5-nitro-2-(3-phenylpropylamino)benzoic acid; D006861:Hydrogen Peroxide; D017878:4,4'-Diisothiocyanostilbene-2,2'-Disulfonic Acid; D010693:Phloretin