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Mechanism of inhibition of mouse Slo3 (KCa 5.1) potassium channels by quinine, quinidine and barium.

Wrighton DC, Muench SP, Lippiat JD - Br. J. Pharmacol. (2015)

Bottom Line: The F304Y mutation did not alter the effects of barium, but increased the potency of inhibition by both quinine and quinidine approximately 10-fold; this effect was not observed with the R196Q mutation.Barium inhibits mSlo3 outside the cell by interacting with the selectivity filter, whereas quinine and quinidine act from the inside, by binding in a hydrophobic pocket formed by the S6 segment of each subunit.Furthermore, we propose that the Slo3 channel activation gate lies deep within the pore between F304 in the S6 segment and the selectivity filter.

View Article: PubMed Central - PubMed

Affiliation: School of Biomedical Sciences, Faculty of Biological Sciences, University of Leeds, Leeds, UK.

No MeSH data available.


Related in: MedlinePlus

Concentration-dependent inhibition of WT, F304Y and R196Q mSlo3 currents by quinine and quinidine. Representative traces recorded before (0) and in the presence of quinine (A) and quinidine (B) (concentrations in μM as indicated). The dashed line represents the zero-current levels and scale bars represent equivalent current amplitudes and timescales. Mean (± SEM) concentration–inhibition plots for quinine (C) and quinidine (D) inhibition of WT mSlo3, R196Q mSlo3 and F304Y mSlo3 at +100 mV, fitted by the Hill equation provided in the Methods. Apparent voltage-dependence of the IC50 for quinine (E) and quinidine (F) of WT and F304Y mSlo3 currents. The data are described and analysed further in the main text. For key to symbols used see Figure 1.
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fig03: Concentration-dependent inhibition of WT, F304Y and R196Q mSlo3 currents by quinine and quinidine. Representative traces recorded before (0) and in the presence of quinine (A) and quinidine (B) (concentrations in μM as indicated). The dashed line represents the zero-current levels and scale bars represent equivalent current amplitudes and timescales. Mean (± SEM) concentration–inhibition plots for quinine (C) and quinidine (D) inhibition of WT mSlo3, R196Q mSlo3 and F304Y mSlo3 at +100 mV, fitted by the Hill equation provided in the Methods. Apparent voltage-dependence of the IC50 for quinine (E) and quinidine (F) of WT and F304Y mSlo3 currents. The data are described and analysed further in the main text. For key to symbols used see Figure 1.

Mentions: We studied the effects of quinine and quinidine in more detail. Quinine inhibited WT mSlo3 currents evoked by voltage pulses to +100 mV with an IC50 of 169 ± 40 μM and Hill slope of 1.0 ± 0.21 (n = 12; Figure 3A and C). F304Y mSlo3 was also blocked by quinine, but with a significantly lower IC50 of 15.9 ± 3.31 μM (n = 16, P < 0.0005; Hill slope of 0.69 ± 0.02). Potency was not significantly altered with the R196Q mSlo3 mutant, which was blocked by quinine with an IC50 of 166 ± 27.5 μM and Hill slope of 1.3 ± 0.04 (n = 4). The effects of quinidine, a stereoisomer of quinine, have previously been studied on the MC13 Slo1/Slo3 chimera (Tang et al., 2010b). Quinidine blocked WT mSlo3 channels with an IC50 of 19.9 ± 1.41 μM and Hill slope of 1.15 ± 0.15 (n = 7; Figure 3B and D). Again, the potency of inhibition by quinidine was higher for F304Y mSlo3 (IC50 of 2.42 ± 0.60 μM, n = 9, P < 0.005, anova; Hill slope of 0.98 ± 0.12), but lower with R196Q mSlo3 (IC50 of 38.4 ± 6.77 μM, n = 5, P < 0.001, anova; Hill slope of 1.05 ± 0.16). The inhibition of F304Y mSlo by quinidine was observed to have some time dependence (Figure 3B). To gain an insight into the mechanisms by which quinine and quinidine block WT and F304Y mSlo3 channels with altered potencies, we estimated the voltage dependence of the IC50 at different voltages. Exemplar currents evoked by voltage ramps and inhibition by quinidine are shown in Supporting Information Figure S2. We found that the F304Y mutation significantly increased the electrical distance, δ, sensed by the blockers from −0.12 ± 0.07 (n = 12) to −0.49 ± 0.22 (n = 12, P < 0.01) for block by quinine (Figure 3E), and from −0.12 ± 0.40 (n = 7) to −0.46 ± 0.15 (n = 9, P < 0.05) for block by quinidine (Figure 3F; two-way anova). The negative values for the electrical distance, δ, represent a positively charged molecule moving into the pore from the intracellular face of the channel.


Mechanism of inhibition of mouse Slo3 (KCa 5.1) potassium channels by quinine, quinidine and barium.

Wrighton DC, Muench SP, Lippiat JD - Br. J. Pharmacol. (2015)

Concentration-dependent inhibition of WT, F304Y and R196Q mSlo3 currents by quinine and quinidine. Representative traces recorded before (0) and in the presence of quinine (A) and quinidine (B) (concentrations in μM as indicated). The dashed line represents the zero-current levels and scale bars represent equivalent current amplitudes and timescales. Mean (± SEM) concentration–inhibition plots for quinine (C) and quinidine (D) inhibition of WT mSlo3, R196Q mSlo3 and F304Y mSlo3 at +100 mV, fitted by the Hill equation provided in the Methods. Apparent voltage-dependence of the IC50 for quinine (E) and quinidine (F) of WT and F304Y mSlo3 currents. The data are described and analysed further in the main text. For key to symbols used see Figure 1.
© Copyright Policy - open-access
Related In: Results  -  Collection

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Show All Figures
getmorefigures.php?uid=PMC4556473&req=5

fig03: Concentration-dependent inhibition of WT, F304Y and R196Q mSlo3 currents by quinine and quinidine. Representative traces recorded before (0) and in the presence of quinine (A) and quinidine (B) (concentrations in μM as indicated). The dashed line represents the zero-current levels and scale bars represent equivalent current amplitudes and timescales. Mean (± SEM) concentration–inhibition plots for quinine (C) and quinidine (D) inhibition of WT mSlo3, R196Q mSlo3 and F304Y mSlo3 at +100 mV, fitted by the Hill equation provided in the Methods. Apparent voltage-dependence of the IC50 for quinine (E) and quinidine (F) of WT and F304Y mSlo3 currents. The data are described and analysed further in the main text. For key to symbols used see Figure 1.
Mentions: We studied the effects of quinine and quinidine in more detail. Quinine inhibited WT mSlo3 currents evoked by voltage pulses to +100 mV with an IC50 of 169 ± 40 μM and Hill slope of 1.0 ± 0.21 (n = 12; Figure 3A and C). F304Y mSlo3 was also blocked by quinine, but with a significantly lower IC50 of 15.9 ± 3.31 μM (n = 16, P < 0.0005; Hill slope of 0.69 ± 0.02). Potency was not significantly altered with the R196Q mSlo3 mutant, which was blocked by quinine with an IC50 of 166 ± 27.5 μM and Hill slope of 1.3 ± 0.04 (n = 4). The effects of quinidine, a stereoisomer of quinine, have previously been studied on the MC13 Slo1/Slo3 chimera (Tang et al., 2010b). Quinidine blocked WT mSlo3 channels with an IC50 of 19.9 ± 1.41 μM and Hill slope of 1.15 ± 0.15 (n = 7; Figure 3B and D). Again, the potency of inhibition by quinidine was higher for F304Y mSlo3 (IC50 of 2.42 ± 0.60 μM, n = 9, P < 0.005, anova; Hill slope of 0.98 ± 0.12), but lower with R196Q mSlo3 (IC50 of 38.4 ± 6.77 μM, n = 5, P < 0.001, anova; Hill slope of 1.05 ± 0.16). The inhibition of F304Y mSlo by quinidine was observed to have some time dependence (Figure 3B). To gain an insight into the mechanisms by which quinine and quinidine block WT and F304Y mSlo3 channels with altered potencies, we estimated the voltage dependence of the IC50 at different voltages. Exemplar currents evoked by voltage ramps and inhibition by quinidine are shown in Supporting Information Figure S2. We found that the F304Y mutation significantly increased the electrical distance, δ, sensed by the blockers from −0.12 ± 0.07 (n = 12) to −0.49 ± 0.22 (n = 12, P < 0.01) for block by quinine (Figure 3E), and from −0.12 ± 0.40 (n = 7) to −0.46 ± 0.15 (n = 9, P < 0.05) for block by quinidine (Figure 3F; two-way anova). The negative values for the electrical distance, δ, represent a positively charged molecule moving into the pore from the intracellular face of the channel.

Bottom Line: The F304Y mutation did not alter the effects of barium, but increased the potency of inhibition by both quinine and quinidine approximately 10-fold; this effect was not observed with the R196Q mutation.Barium inhibits mSlo3 outside the cell by interacting with the selectivity filter, whereas quinine and quinidine act from the inside, by binding in a hydrophobic pocket formed by the S6 segment of each subunit.Furthermore, we propose that the Slo3 channel activation gate lies deep within the pore between F304 in the S6 segment and the selectivity filter.

View Article: PubMed Central - PubMed

Affiliation: School of Biomedical Sciences, Faculty of Biological Sciences, University of Leeds, Leeds, UK.

No MeSH data available.


Related in: MedlinePlus