Ca²⁺-activated K⁺ channel inhibition by reactive oxygen species

Abstract

We recorded potassium currents from single skeletal muscle channels incorporated into bilayers or using macropatches of Xenopus laevis oocytes membranes expressing the human Slowpoke (hSlo) ␣-subunit. Exposure of the intracellular side of KV,Ca channels to H2O2 (4-23 mM) leads to a time-dependent decrease of the open probability (Po) without affecting the unitary conductance. H2O2 did not affect channel activity when added to the extracellular side. These results provide evidence for an intracellular site(s) of H 2O2 action. Desferrioxamine (60 M) and cysteine (1 mM) completely inhibited the effect of H2O2, indicating that the decrease in Po was mediated by hydroxyl radicals. The reducing agent dithiothreitol (DTT) could not fully reverse the effect of H2O2. However, DTT did completely reverse the decrease in Po induced by the oxidizing agent 5,5Ј-dithio-bis-(2-nitrobenzoic acid). The incomplete recovery of KV,Ca channel activity promoted by DTT suggests that H2O2 treatment must be modifying other amino acid residues, e.g., as methionine or tryptophan, besides cysteine. Noise analysis of macroscopic currents in Xenopus oocytes expressing hSlo channels showed that H2O2 induced a decrease in current mediated by a decrease both in the number of active channels and Po.

A cytosolic source of calcium unveiled by hydrogen peroxide with relevance for epithelial cell death

“…47 Another example of a low-affinity peroxide effect is its inhibition of skeletal muscle Ca 2 þ -activated K þ channel, which in artificial bilayers showed a K i 410 mM. 48 A wellknown effect of peroxide in combination with transition metals is the production of highly reactive hydroxyl radicals in the Fenton or Haber-Weiss reactions; however, such mechanisms do not seem to be relevant as chelation of transition metals with desferroxamine failed to inhibit the calcium rise. A similar pattern of sensitivity to DTT/insensitivity to desferroxamine has been reported for peroxide-induced calcium rise in human platelets 49 and acinar pancreatic cells.…”

Activation of Vascular BK Channel by Tempol in DOCA-Salt Hypertensive Rats

“…39 This pathway is relevant to our data as H 2 O 2 relaxes porcine coronary arteries by stimulating BK channel activity via a phospholipase A2-coupled signaling cascade. 40 However, in MA, H 2 O 2 induced vasodilation was largely independent of BK channel activity 41,42 and exogenous H 2 O 2 inhibits BK channels in skeletal muscle 43 and hslo-tranfected HEK-293 cells. 44 Endogenous O 2 Ϫ in MA and mslo transfected HEK-293 cells could be converted to H 2 O 2 by tempol and H 2 O 2 could be the activator of BK channels as occurs in coronary arteries.…”

Opening of connexin 43 hemichannels is increased by lowering intracellular redox potential

“…The action of reducing agents suggests a role for redox potential in controlling hemichannel opening in cells under metabolic stress. Redox potential is known to modify the activity of different types of ion channels, including ryanodine receptors (12), P2X receptors (13), transient receptor potential (TRP) channels (14), potassium channels (15), and nonselective cation channels (16). At physiological cytosolic oxygen pressure, nitric oxide (NO) activates r yanodine receptors through Snitrosylation of free cysteines of the channel subunit, but at atmospheric oxygen pressure, cysteine S-nitrosylation does not occur because cysteine residues are oxidized (12).…”