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Alkanols inhibit voltage-gated K(+) channels via a distinct gating modifying mechanism that prevents gate opening.

Martínez-Morales E, Kopljar I, Snyders DJ, Labro AJ - Sci Rep (2015)

Bottom Line: Using the non-conducting Shaker-W434F mutant, we found that both alkanols immobilized approximately 10% of the gating charge and accelerated the deactivating gating currents simultaneously with ionic current inhibition.Thus, alkanols prevent the final VSD movement(s) that is associated with channel gate opening.Drug competition experiments showed that alkanols do not share the binding site of 4-aminopyridine, a drug that exerts a similar effect at the gating current level.

View Article: PubMed Central - PubMed

Affiliation: Laboratory for Molecular Biophysics, Physiology and Pharmacology, Department of Biomedical Sciences, University of Antwerp, Antwerp, 2610, Belgium.

ABSTRACT
Alkanols are small aliphatic compounds that inhibit voltage-gated K(+) (K(v)) channels through a yet unresolved gating mechanism. K(v) channels detect changes in the membrane potential with their voltage-sensing domains (VSDs) that reorient and generate a transient gating current. Both 1-Butanol (1-BuOH) and 1-Hexanol (1-HeOH) inhibited the ionic currents of the Shaker K(v) channel in a concentration dependent manner with an IC50 value of approximately 50 mM and 3 mM, respectively. Using the non-conducting Shaker-W434F mutant, we found that both alkanols immobilized approximately 10% of the gating charge and accelerated the deactivating gating currents simultaneously with ionic current inhibition. Thus, alkanols prevent the final VSD movement(s) that is associated with channel gate opening. Applying 1-BuOH and 1-HeOH to the Shaker-P475A mutant, in which the final gating transition is isolated from earlier VSD movements, strengthened that neither alkanol affected the early VSD movements. Drug competition experiments showed that alkanols do not share the binding site of 4-aminopyridine, a drug that exerts a similar effect at the gating current level. Thus, alkanols inhibit Shaker-type K(v) channels via a unique gating modifying mechanism that stabilizes the channel in its non-conducting activated state.

No MeSH data available.


Related in: MedlinePlus

Alkanols did not affect the IG behavior of Shaker-IR-W434F-P475A.(A) Representative IG recordings of Shaker-IR-W434F-P475A recorded in control condition (left recordings) and after approximately 10 minutes wash-in of 300 mM 1-BuOH (right recordings), elicited with the pulse protocols shown on top. Insets show scale up views of IGac and IGdeac respectively. (B) Representative IG recordings of Shaker-IR-W434F-P475A in presence of 30 mM 1-HeOH. (C) QV curves of Shaker-IR-W434F-P475A in control conditions (white circles) and presence of 300 mM 1-BuOH (black circles, n = 5) or 30 mM 1-HeOH (gray triangles, n = 4). (D) τIGac (gray circles) and τIGdeac (black circles) kinetics of Shaker-IR-W434F-P475A in presence of 300 mM 1-BuOH, which were similar to the kinetics in control condition (white circles). (E) Similar to 1-BuOH, 30 mM 1-HeOH did not affect the τIGac (gray triangles) or the τIGdeac (black triangles) kinetics compared to control condition (white triangles).
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f9: Alkanols did not affect the IG behavior of Shaker-IR-W434F-P475A.(A) Representative IG recordings of Shaker-IR-W434F-P475A recorded in control condition (left recordings) and after approximately 10 minutes wash-in of 300 mM 1-BuOH (right recordings), elicited with the pulse protocols shown on top. Insets show scale up views of IGac and IGdeac respectively. (B) Representative IG recordings of Shaker-IR-W434F-P475A in presence of 30 mM 1-HeOH. (C) QV curves of Shaker-IR-W434F-P475A in control conditions (white circles) and presence of 300 mM 1-BuOH (black circles, n = 5) or 30 mM 1-HeOH (gray triangles, n = 4). (D) τIGac (gray circles) and τIGdeac (black circles) kinetics of Shaker-IR-W434F-P475A in presence of 300 mM 1-BuOH, which were similar to the kinetics in control condition (white circles). (E) Similar to 1-BuOH, 30 mM 1-HeOH did not affect the τIGac (gray triangles) or the τIGdeac (black triangles) kinetics compared to control condition (white triangles).

Mentions: Since the Shaker-IR-P475A mutant did not affect the early VSD movements, the QV curve was split and displayed two gating charge components whereby the late one corresponded with the voltage dependence of channel gate opening29. Analyzing the gating currents of Shaker-IR-W434F-P475A in presence of 300 mM 1-BuOH or 30 mM 1-HeOH indicated that the voltage dependence of neither the early nor the late gating charge component was affected by 1-BuOH or 1-HeOH (Fig. 9 and Table 1). This was in agreement with the absence of an obvious shift in the threshold of channel opening (Fig. 8B). Also the IGac time constants, which in Shaker-IR-W434F-P475A report directly on the kinetics of the early VSD movements29, were unaffected by 1-BuOH or 1-HeOH. These IG data confirmed that 1-BuOH and 1-HeOH did not affect the voltage-dependent transitions of the Shaker-IR-P475A mutant but facilitated a late largely voltage-independent transition in the activation pathway, a transition that is compromised by the P475A mutation.


Alkanols inhibit voltage-gated K(+) channels via a distinct gating modifying mechanism that prevents gate opening.

Martínez-Morales E, Kopljar I, Snyders DJ, Labro AJ - Sci Rep (2015)

Alkanols did not affect the IG behavior of Shaker-IR-W434F-P475A.(A) Representative IG recordings of Shaker-IR-W434F-P475A recorded in control condition (left recordings) and after approximately 10 minutes wash-in of 300 mM 1-BuOH (right recordings), elicited with the pulse protocols shown on top. Insets show scale up views of IGac and IGdeac respectively. (B) Representative IG recordings of Shaker-IR-W434F-P475A in presence of 30 mM 1-HeOH. (C) QV curves of Shaker-IR-W434F-P475A in control conditions (white circles) and presence of 300 mM 1-BuOH (black circles, n = 5) or 30 mM 1-HeOH (gray triangles, n = 4). (D) τIGac (gray circles) and τIGdeac (black circles) kinetics of Shaker-IR-W434F-P475A in presence of 300 mM 1-BuOH, which were similar to the kinetics in control condition (white circles). (E) Similar to 1-BuOH, 30 mM 1-HeOH did not affect the τIGac (gray triangles) or the τIGdeac (black triangles) kinetics compared to control condition (white triangles).
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Related In: Results  -  Collection

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getmorefigures.php?uid=PMC4663795&req=5

f9: Alkanols did not affect the IG behavior of Shaker-IR-W434F-P475A.(A) Representative IG recordings of Shaker-IR-W434F-P475A recorded in control condition (left recordings) and after approximately 10 minutes wash-in of 300 mM 1-BuOH (right recordings), elicited with the pulse protocols shown on top. Insets show scale up views of IGac and IGdeac respectively. (B) Representative IG recordings of Shaker-IR-W434F-P475A in presence of 30 mM 1-HeOH. (C) QV curves of Shaker-IR-W434F-P475A in control conditions (white circles) and presence of 300 mM 1-BuOH (black circles, n = 5) or 30 mM 1-HeOH (gray triangles, n = 4). (D) τIGac (gray circles) and τIGdeac (black circles) kinetics of Shaker-IR-W434F-P475A in presence of 300 mM 1-BuOH, which were similar to the kinetics in control condition (white circles). (E) Similar to 1-BuOH, 30 mM 1-HeOH did not affect the τIGac (gray triangles) or the τIGdeac (black triangles) kinetics compared to control condition (white triangles).
Mentions: Since the Shaker-IR-P475A mutant did not affect the early VSD movements, the QV curve was split and displayed two gating charge components whereby the late one corresponded with the voltage dependence of channel gate opening29. Analyzing the gating currents of Shaker-IR-W434F-P475A in presence of 300 mM 1-BuOH or 30 mM 1-HeOH indicated that the voltage dependence of neither the early nor the late gating charge component was affected by 1-BuOH or 1-HeOH (Fig. 9 and Table 1). This was in agreement with the absence of an obvious shift in the threshold of channel opening (Fig. 8B). Also the IGac time constants, which in Shaker-IR-W434F-P475A report directly on the kinetics of the early VSD movements29, were unaffected by 1-BuOH or 1-HeOH. These IG data confirmed that 1-BuOH and 1-HeOH did not affect the voltage-dependent transitions of the Shaker-IR-P475A mutant but facilitated a late largely voltage-independent transition in the activation pathway, a transition that is compromised by the P475A mutation.

Bottom Line: Using the non-conducting Shaker-W434F mutant, we found that both alkanols immobilized approximately 10% of the gating charge and accelerated the deactivating gating currents simultaneously with ionic current inhibition.Thus, alkanols prevent the final VSD movement(s) that is associated with channel gate opening.Drug competition experiments showed that alkanols do not share the binding site of 4-aminopyridine, a drug that exerts a similar effect at the gating current level.

View Article: PubMed Central - PubMed

Affiliation: Laboratory for Molecular Biophysics, Physiology and Pharmacology, Department of Biomedical Sciences, University of Antwerp, Antwerp, 2610, Belgium.

ABSTRACT
Alkanols are small aliphatic compounds that inhibit voltage-gated K(+) (K(v)) channels through a yet unresolved gating mechanism. K(v) channels detect changes in the membrane potential with their voltage-sensing domains (VSDs) that reorient and generate a transient gating current. Both 1-Butanol (1-BuOH) and 1-Hexanol (1-HeOH) inhibited the ionic currents of the Shaker K(v) channel in a concentration dependent manner with an IC50 value of approximately 50 mM and 3 mM, respectively. Using the non-conducting Shaker-W434F mutant, we found that both alkanols immobilized approximately 10% of the gating charge and accelerated the deactivating gating currents simultaneously with ionic current inhibition. Thus, alkanols prevent the final VSD movement(s) that is associated with channel gate opening. Applying 1-BuOH and 1-HeOH to the Shaker-P475A mutant, in which the final gating transition is isolated from earlier VSD movements, strengthened that neither alkanol affected the early VSD movements. Drug competition experiments showed that alkanols do not share the binding site of 4-aminopyridine, a drug that exerts a similar effect at the gating current level. Thus, alkanols inhibit Shaker-type K(v) channels via a unique gating modifying mechanism that stabilizes the channel in its non-conducting activated state.

No MeSH data available.


Related in: MedlinePlus