Nitric oxide and thiol reagent modulation of Ca²⁺‐activated K⁺ (BK_Ca) channels in myocytes of the guinea‐pig taenia caeci

Abstract

The modulation of large conductance Ca2+‐activated K+ (BKCa) channels by the nitric oxide (NO) donors S‐nitroso‐L‐cysteine (NOCys) and sodium nitroprusside (SNP) and agents which oxidize or reduce reactive thiol groups were compared in excised inside‐out membrane patches of the guinea‐pig taenia caeci. When the cytosolic side of excised patches was bathed in a physiological salt solution (PSS) containing 130 mm K+ and 15 nm Ca2+, few BKCa channel openings were recorded at potentials negative to 0 mV. However, the current amplitude and open probability (NPo) of these BKCa channels increased with patch depolarization. A plot of ln(NPo) against the membrane potential (V) fitted with a straight line revealed a voltage at half‐maximal activation (V0.5) of 9.4 mV and a slope (K) indicating an e‐fold increase in NPo with 12.9 mV depolarization. As the cytosolic Ca2+ was raised to 150 nm, V0.5 shifted 11.5 mV in the negative direction, with little change in K (13.1 mV). NOCys (10 μm) and SNP (100 μm) transiently increased NPo 16‐ and 3.7‐fold, respectively, after a delay of 2–5 min. This increase in NPo was associated with an increase in the number of BKCa channel openings evoked at positive potentials by ramped depolarizations (between −60 and +60 mV). Moreover, this NOCys‐induced increase in NPo was still evident in the presence of 1H‐[1,2,4]oxadiazolo[4,3‐a]quinoxalin‐1‐one (ODQ; 10 μm), the specific blocker of soluble guanylyl cyclase. The sulfhydryl reducing agents dithiothreitol (DTT; 10 and 100 μm) and reduced glutathione (GSH; 1 mm) also significantly increased NPo (at 0 mV) 7‐ to 9‐fold, as well as increasing the number of BKCa channel openings evoked during ramped depolarizations. Sulfhydryl oxidizing agents thimerosal (10 μm) and 4,4′‐dithiodipyridine (4,4DTDP; 10 μm) and the thiol‐specific alkylating agent N‐ethylmaleimide (NEM; 1 mm) significantly decreased NPo (at 0 mV) to 40–50 % of control values after 5–10 min. Ramped depolarizations to +100 mV evoked relatively few BKCa channel openings. The effects of thimerosal on NPo were readily reversed by DTT, while the effects of NOCys were prevented by NEM. It was concluded that both redox modulation and nitrothiosylation of cysteine groups on the cytosolic surface of the α subunit of the BKCa channel protein can alter channel gating.

cGMP/Protein Kinase G-Dependent Inhibition of N-Type Ca2+Channels Induced by Nitric Oxide in Human Neuroblastoma IMR32 Cells

“…The SNP-induced inhibition was voltage independent, as shown by the similar magnitude of current reduction observed at the different voltages tested. The time course of the effects and the limited recovery observed during the first 2 min of washout are consistent with the reports of various investigators (Chen and Schofield, 1995;Chik et al, 1995;Tewari and Simard, 1997;Lang et al, 2000). At the single-channel level, the inhibitory action of cGMP consists of a reduction of the open probability of available channels and an increase in the mean closed time.…”

CO metabolism, sensing, and signaling

“…Conversely, the upregulation of the channel function in specific cell types may offer protection against some of the aforementioned disorders [107][108][109][110]. The BK Ca channel activity is subjected to modulation by a wide spectrum of biologically relevant factors such as Ser/Thr/Tyr phosphorylation, Cys/Met oxidation, steroid hormones, and diatomic gases (O 2 , NO, and CO) [111][112][113][114]. The PKG-dependent activation of the Na þ /Ca 2þ exchanger by CO has been proposed [12]: this may increase the submembrane Ca 2þ concentration in the vicinity of the BK Ca channels with their consequential opening [115].…”

Nitric oxide activates hypoglossal motoneurons by cGMP-dependent inhibition of TASK channels and cGMP-independent activation of HCN channels

“…This cGMP-independent signal is conferred by the covalent addition of an NO moiety to the sulfhydryl group of cysteine residues. This reversible posttranslational modification regulates function of several channels including NMDA receptors (NMDA-R) (Choi et al 2000), ATP-sensitive K ϩ (K ATP ) channels (Kawano et al 2009), Ca 2ϩ -dependent K ϩ channels (Lang et al 2000), and cyclic nucleotide-gated (CNG) channels (Broillet and Firestein 1996). However, despite compelling evidence that S-nitrosylation is an important component of NO signaling, physiological significance of this pathway remains uncertain.…”