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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

Expression of WT, R196Q and F304Y mSlo3 K+ channels in Xenopus oocytes. (A) Representative current families recorded by two electrode voltage clamp from oocytes injected with different mSlo3 RNA or with no RNA (control) as indicated. Oocytes were held at −80 mV and 100 ms pulses to potentials between −100 and +140 mV were applied. The dashed line represents the zero-current levels and scale bars represent equivalent current amplitudes and timescales. (B) Mean (± SEM) current–voltage relationships of oocytes expressing WT mSlo3 (WT, n = 29), R196Q mSlo3 (n = 8) F304Y mSlo3 (n = 26), and non-injected oocytes (control, n = 12). For symbols used see part (A). (C) Mean (± SEM) resting membrane potential of oocytes in standard Ringer's solution. *P < 0.0001 compared with control oocytes (Kruskal–Wallis test).
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fig01: Expression of WT, R196Q and F304Y mSlo3 K+ channels in Xenopus oocytes. (A) Representative current families recorded by two electrode voltage clamp from oocytes injected with different mSlo3 RNA or with no RNA (control) as indicated. Oocytes were held at −80 mV and 100 ms pulses to potentials between −100 and +140 mV were applied. The dashed line represents the zero-current levels and scale bars represent equivalent current amplitudes and timescales. (B) Mean (± SEM) current–voltage relationships of oocytes expressing WT mSlo3 (WT, n = 29), R196Q mSlo3 (n = 8) F304Y mSlo3 (n = 26), and non-injected oocytes (control, n = 12). For symbols used see part (A). (C) Mean (± SEM) resting membrane potential of oocytes in standard Ringer's solution. *P < 0.0001 compared with control oocytes (Kruskal–Wallis test).

Mentions: Full-length WT, R196Q and F304Y mSlo3 K+ channel subunits were expressed in Xenopus oocytes and currents recorded by two-electrode voltage clamp. Oocytes injected with WT mSlo3 gave rise to outwardly rectifying currents, which were absent in non-injected oocytes in the 0–100 mV range (Figure 1A and B). Oocytes injected with mRNA encoding the mSlo3 mutants R196Q and F304Y also yielded large outwardly rectifying currents, but exhibited channel activity at voltages below the threshold for WT mSlo3 activation (Figure 1A and B). Expressing R196Q and F304Y mSlo3 resulted in a significantly more negative oocyte resting membrane potential (Figure 1C) compared with control oocytes or those expressing WT mSlo3. We observed a reduction in the survival of oocytes expressing the gain-of-function mutants in normal Barth's medium, which was rectified by raising the K+ concentration (see Methods).


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)

Expression of WT, R196Q and F304Y mSlo3 K+ channels in Xenopus oocytes. (A) Representative current families recorded by two electrode voltage clamp from oocytes injected with different mSlo3 RNA or with no RNA (control) as indicated. Oocytes were held at −80 mV and 100 ms pulses to potentials between −100 and +140 mV were applied. The dashed line represents the zero-current levels and scale bars represent equivalent current amplitudes and timescales. (B) Mean (± SEM) current–voltage relationships of oocytes expressing WT mSlo3 (WT, n = 29), R196Q mSlo3 (n = 8) F304Y mSlo3 (n = 26), and non-injected oocytes (control, n = 12). For symbols used see part (A). (C) Mean (± SEM) resting membrane potential of oocytes in standard Ringer's solution. *P < 0.0001 compared with control oocytes (Kruskal–Wallis test).
© Copyright Policy - open-access
Related In: Results  -  Collection

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fig01: Expression of WT, R196Q and F304Y mSlo3 K+ channels in Xenopus oocytes. (A) Representative current families recorded by two electrode voltage clamp from oocytes injected with different mSlo3 RNA or with no RNA (control) as indicated. Oocytes were held at −80 mV and 100 ms pulses to potentials between −100 and +140 mV were applied. The dashed line represents the zero-current levels and scale bars represent equivalent current amplitudes and timescales. (B) Mean (± SEM) current–voltage relationships of oocytes expressing WT mSlo3 (WT, n = 29), R196Q mSlo3 (n = 8) F304Y mSlo3 (n = 26), and non-injected oocytes (control, n = 12). For symbols used see part (A). (C) Mean (± SEM) resting membrane potential of oocytes in standard Ringer's solution. *P < 0.0001 compared with control oocytes (Kruskal–Wallis test).
Mentions: Full-length WT, R196Q and F304Y mSlo3 K+ channel subunits were expressed in Xenopus oocytes and currents recorded by two-electrode voltage clamp. Oocytes injected with WT mSlo3 gave rise to outwardly rectifying currents, which were absent in non-injected oocytes in the 0–100 mV range (Figure 1A and B). Oocytes injected with mRNA encoding the mSlo3 mutants R196Q and F304Y also yielded large outwardly rectifying currents, but exhibited channel activity at voltages below the threshold for WT mSlo3 activation (Figure 1A and B). Expressing R196Q and F304Y mSlo3 resulted in a significantly more negative oocyte resting membrane potential (Figure 1C) compared with control oocytes or those expressing WT mSlo3. We observed a reduction in the survival of oocytes expressing the gain-of-function mutants in normal Barth's medium, which was rectified by raising the K+ concentration (see Methods).

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