ns-309 and 1-ethyl-2-benzimidazolinone

Small-conductance Ca

Topics: Benzimidazoles; Binding Sites; Crystallography, X-Ray; Humans; Indoles; Oximes; Small-Conductance Calcium-Activated Potassium Channels

This study was designed to determine whether putative openers of calcium-activated potassium channels of small and/or intermediate conductance (SK(Ca) and IK(Ca)) induce vascular smooth muscle hyperpolarizations and to identify the underlying mechanisms. The membrane potential of guinea pig carotid artery smooth muscle cells was recorded with intracellular microelectrodes in the presence of N(omega)-nitro-L-arginine and indomethacin. Acetylcholine and NS-309 produced endothelium-dependent hyperpolarizations. The effects of acetylcholine were partially and significantly inhibited by apamin. The combinations of charybdotoxin plus apamin and TRAM-34 plus apamin markedly and significantly reduced these hyperpolarizations. 1-ethyl-2-benzimidazolinone (1-EBIO) induced hyperpolarizations that were unaffected by TRAM-34 but partially inhibited by charybdotoxin, apamin, TRAM-34 plus apamin, and charybdotoxin plus apamin. Riluzole produced only marginal hyperpolarizations. Therefore, in the guinea pig carotid artery, endothelium-dependent hyperpolarization to acetylcholine involves the activation of both SK(Ca) and IK(Ca), with a predominant role for the former channel. 1-EBIO is a non-selective and weak opener of SK(Ca), while riluzole is virtually ineffective. By contrast, NS-309 is a reasonably potent and selective opener of both SK(Ca) and IK(Ca), and this compound mimics the endothelium-dependent hyperpolarizations to acetylcholine.

Topics: Acetylcholine; Animals; Benzimidazoles; Biological Factors; Carotid Arteries; Endothelium, Vascular; Guinea Pigs; Indoles; Male; Membrane Potentials; Oximes; Potassium Channels, Calcium-Activated; Pyrazoles; Riluzole

Positive modulators of small conductance Ca(2+)-activated K(+) channels (SK1, SK2, and SK3) exert hyperpolarizing effects that influence the activity of excitable and non-excitable cells. The prototype compound 1-EBIO or the more potent compound NS309, do not distinguish between the SK subtypes and they also activate the related intermediate conductance Ca(2+)-activated K(+) channel (IK). This paper demonstrates, for the first time, subtype-selective positive modulation of SK channels.. Using patch clamp and fluorescence techniques we studied the effect of the compound cyclohexyl-[2-(3,5-dimethyl-pyrazol-1-yl)-6-methyl-pyrimidin-4-yl]-amine (CyPPA) on recombinant hSK1-3 and hIK channels expressed in HEK293 cells. CyPPA was also tested on SK3 and IK channels endogenously expressed in TE671 and HeLa cells.. CyPPA was found to be a positive modulator of hSK3 (EC(50) = 5.6 +/- 1.6 microM, efficacy 90 +/- 1.8 %) and hSK2 (EC(50) = 14 +/- 4 microM, efficacy 71 +/- 1.8 %) when measured in inside-out patch clamp experiments. CyPPA was inactive on both hSK1 and hIK channels. At hSK3 channels, CyPPA induced a concentration-dependent increase in the apparent Ca(2+)-sensitivity of channel activation, changing the EC(50)(Ca(2+)) from 429 nM to 59 nM.. As a pharmacological tool, CyPPA may be used in parallel with the IK/SK openers 1-EBIO and NS309 to distinguish SK3/SK2- from SK1/IK-mediated pharmacological responses. This is important for the SK2 and SK1 subtypes, since they have overlapping expression patterns in the neocortical and hippocampal regions, and for SK3 and IK channels, since they co-express in certain peripheral tissues.

Topics: Algorithms; Benzimidazoles; Cell Line; Electrophysiology; Fluorescent Dyes; Humans; Indoles; Membrane Potentials; Oximes; Patch-Clamp Techniques; Small-Conductance Calcium-Activated Potassium Channels; Thallium

We have identified and characterized the compound NS309 (6,7-dichloro-1H-indole-2,3-dione 3-oxime) as a potent activator of human Ca2+ -activated K+ channels of SK and IK types, whereas it is devoid of effect on BK type channels. IK- and SK-channels have previously been reported to be activated by the benzimidazolinone, 1-EBIO and more potently by its dichloronated-analogue, DC-EBIO. NS309 is at least 1000 times more potent than 1-EBIO and at least 30 times more potent than DC-EBIO when the compounds are compared on the same cell.

Topics: Benzimidazoles; Calcium Channel Agonists; Cell Line; Dose-Response Relationship, Drug; Humans; Indoles; Intermediate-Conductance Calcium-Activated Potassium Channels; Kinetics; Oximes; Patch-Clamp Techniques; Potassium Channels, Calcium-Activated; Small-Conductance Calcium-Activated Potassium Channels