Abstract
The large-conductance Ca2+-activated K+ (BK) channel is activated by both the increase of intracellular Ca2+ concentration and membrane depolarization. The BK channel plays crucial roles as a key molecule in the negative feedback mechanism regulating membrane excitability and cellular Ca2+ in various cell types. Here, we report that a widely used slow-response voltage-sensitive fluorescent dye, bis(1,3-dibutylbarbituric acid)trimethine oxonol [DiBAC4(3)], is a potent BK channel activator. The application of DiBAC4(3) at concentrations of 10 nM and higher significantly increased whole-cell BK channel currents in human embryonic kidney 293 cells expressing rat BK channel α and β1 subunits (rBKαβ1). In the presence of 300 nM DiBAC4(3), the activation voltage of the BK channel current shifted to the negative direction by approximately 30 mV, but the single-channel conductance was not affected. DiBAC4(3) activated whole-cell rBKαβ1 and rBKαβ4 currents in the same concentration range but partially blocked rBKαβ2 currents. The BK channel α subunit alone and some other types of K+ channels examined were not markedly affected by 1 μM DiBAC4(3). Structure-activity relationship analyses revealed that a set of oxo- and oxoanion-moieties in two 1,3-dialkylbarbituric acids, which are conjugated by oligomethine, is the novel skeleton for the β-subunit-selective BK channel-opening property of DiBAC4(3) and related oxonol compounds. This conjugated structure may be located stereochemically in one plane. These findings provide a molecular and structural basis for understanding the regulatory mechanism of BK channel activity by an auxiliary β subunit and will be fundamental to the development of β-selective BK channel openers.
Footnotes
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This work was supported by a Grant-in-Aid for Scientific Research on Priority Areas (18059029) from the Ministry of Education, Culture, Sports, Science, and Technology, Japan, and by a Grant-in-Aid for Scientific Research (B) (17390045) from the Japan Society for the Promotion of Science (to Y.I.). This work was also supported by a Grant-in-Aid for Research on Health Sciences focusing on Drug Innovation (KH11001) from the Japan Health Sciences Foundation (to Y.I.).
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Article, publication date, and citation information can be found at http://molpharm.aspetjournals.org.
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doi:10.1124/mol.106.031146.
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ABBREVIATIONS: BK channel, large-conductance Ca2+-activated K+ channel; SK channel, small-conductance Ca2+-activated K+ channel, Kv channel, voltage-gated K+ channel; KATP channel, ATP-sensitive K+ channel; AE, anion exchanger; mUBSMC, mouse urinary bladder smooth muscle cell; HEK, human embryonic kidney; BKα, large-conductance Ca2+-activated K+ channel α subunit; BKβ, large-conductance Ca2+-activated K+ channel β subunit; BKαβ1, large-conductance Ca2+-activated K+ channel α plus β1 subunits; BKαβ2, large-conductance Ca2+-activated K+ channel α plus β2 subunits; BKαβ4, large-conductance Ca2+-activated K+ channel α plus β4 subunits; r, rat; m, mouse; DHS-I, dehydrosoyasaponin-I, BA, barbituric acid; DMBA, 1,3-dimethylbarbituric acid; DETBA, 1,3-diethyl-2-thiobarbituric acid; DiBAC, bis(1,3-dialkylbarbituric acid)oligomethine oxonol; DiSBAC, bis(1,3-dialkylthiobarbituric acid)oligomethine oxonol; DiBAC4(3), bis(1,3-dibutylbarbituric acid)trimethine oxonol; DiBAC4(5), bis(1,3-dibutylbarbituric acid)pentamethine oxonol; DiSBAC4(3), bis(1,3-dibutylthiobarbituric acid)trimethine oxonol; DiSBAC2(3), bis(1,3-diethylthiobarbituric acid)trimethine oxonol; DiSBAC2(5), bis(1,3-diethylthiobarbituric acid)pentamethine oxonol; DiSBAC2(1), bis(1,3-diethylthiobarbituric acid) methine oxonol; DiSBAC0(5), bis(barbituric acid)pentamethine oxonol; DiSBAC10(3), bis(1,3-didecylthiobarbituric acid)trimethine oxonol; Oxonol 595, bis(3-cyano-1-ethyl-4-methyl-2,6-dioxo-1,2,5,6-tetrahydropyridine)trimethine oxonol; Oxonol V, bis(3-phenyl-5-oxoisoxazol-4-yl)pentamethine oxonol; Oxonol VI, bis(3-propyl-5-oxoisoxazol-4-yl)pentamethine oxonol; DFT, density-functional theory; NS-1619, 1-(2′-hydroxy-5′-trifluoromethylphenyl)-5-trifluoromethyl-2(3H)-benzimidazolone; UCL 1684, 6,10-diaza-3(1,3)8,(1,4)-dibenzena-1,5(1,4)-diquinolinacy clodecaphane.
- Received September 23, 2006.
- Accepted January 3, 2007.
- The American Society for Pharmacology and Experimental Therapeutics
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