Direct inhibition of expressed cardiac L-type Ca2+ channels by S-nitrosothiol nitric oxide donors.
NO donors have complex effects on Ca2+ currents in native cardiac cells, with reports of direct stimulation and indirect cGMP-mediated inhibition or stimulation. To investigate the molecular basis of these effects, we tested the effects of one class of NO donors, S-nitrosothiols (RSNOs), on expressed cardiovascular L-type Ca2+ channels (alpha 1C +/- beta 1a +/- alpha 2 or alpha 1C +/- beta 2a +/- alpha 2) in human embryonic kidney (HEK293) cells. The RSNO compounds we used were S-nitroso-N-acetylpenicillamine (SNAP, 5 to 10 nmol/L or 100 to 800 mumol/L), S-nitrosocysteine (SNC, 100 mumol/L or 1 mmol/L), and S-nitrosoglutathione (GSNO, 1 mmol/L). Currents were measured using whole-cell patch recordings with 2 to 10 mmol/L Ba2+ as the charge carrier. SNAP reduced the amplitude of barium currents (IBa) through all the subunit combinations, with and EC50 of 360 mumol/L for alpha 1C + beta 1a channels. SNC or GSNO also inhibited IBa, albeit less potently. The inhibitory effect of SNAP was not affected by methylene blue (10 to 30 mumol/L) or 8-bromo-cGMP (200 to 400 mumol/L). The effects are relatively specific for Ca2+ channels, as expressed cardiac or skeletal muscle Na+ channels, which have a similar overall architecture, were barely affected by SNAP at concentrations as high as 1 mmol/L. We conclude that in the HEK293 expression system, the S-nitrosothiol NO donors inhibit L-type Ca2+ channels by a mechanism independent of cGMP.
Hu
H
H
Department of Medicine, Johns Hopkins University School of Medicine, Baltimore, MD 21205, USA.
Chiamvimonvat
N
N
Yamagishi
T
T
Marban
E
E
D000818:Animals; D002121:Calcium Channel Blockers / Q000494:pharmacology; D015220:Calcium Channels / Q000096:biosynthesis*; D002460:Cell Line; D004791:Enzyme Inhibitors / Q000494:pharmacology*; D006801:Humans; D008623:Mercaptoethanol; D009206:Myocardium / Q000378:metabolism*; D009569:Nitric Oxide / Q000378:metabolism*; D009603:Nitroso Compounds / Q000494:pharmacology*; D010396:Penicillamine / Q000031:analogs & derivatives* / Q000494:pharmacology; D026423:S-Nitroso-N-Acetylpenicillamine; D026403:S-Nitrosothiols; D014981:Xenopus / Q000378:metabolism
D002121:Calcium Channel Blockers; D015220:Calcium Channels; D004791:Enzyme Inhibitors; D009603:Nitroso Compounds; D026403:S-Nitrosothiols; D009569:Nitric Oxide; D008623:Mercaptoethanol; C032635:S-nitrosomercaptoethanol; D026423:S-Nitroso-N-Acetylpenicillamine; D010396:Penicillamine