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Direct Pore Binding as a Mechanism for Isoflurane Inhibition of the Pentameric Ligand-gated Ion Channel ELIC.

Chen Q, Kinde MN, Arjunan P, Wells MM, Cohen AE, Xu Y, Tang P - Sci Rep (2015)

Bottom Line: Electrophysiology measurements with a single-point mutation at position 6' or 13' support the notion that binding at these sites renders isoflurane inhibition.Molecular dynamics simulations suggested that isoflurane binding was more stable in the resting than in a desensitized pore conformation.This study presents compelling evidence for a direct pore-binding mechanism of isoflurane inhibition, which has a general implication for inhibitory action of general anesthetics on pLGICs.

View Article: PubMed Central - PubMed

Affiliation: Department of Anesthesiology, University of Pittsburgh School of Medicine, Pittsburgh, Pennsylvania, 15260, USA.

ABSTRACT
Pentameric ligand-gated ion channels (pLGICs) are targets of general anesthetics, but molecular mechanisms underlying anesthetic action remain debatable. We found that ELIC, a pLGIC from Erwinia chrysanthemi, can be functionally inhibited by isoflurane and other anesthetics. Structures of ELIC co-crystallized with isoflurane in the absence or presence of an agonist revealed double isoflurane occupancies inside the pore near T237(6') and A244(13'). A pore-radius contraction near the extracellular entrance was observed upon isoflurane binding. Electrophysiology measurements with a single-point mutation at position 6' or 13' support the notion that binding at these sites renders isoflurane inhibition. Molecular dynamics simulations suggested that isoflurane binding was more stable in the resting than in a desensitized pore conformation. This study presents compelling evidence for a direct pore-binding mechanism of isoflurane inhibition, which has a general implication for inhibitory action of general anesthetics on pLGICs.

No MeSH data available.


Related in: MedlinePlus

Isoflurane binding stabilized the closed channel.(a) A representative trace showing that pre-incubation with isoflurane (red arrows) slowed the onset rate and increased the maximum inhibition. The vertical and horizontal scale bars represent 20 μA and 30 seconds, representatively. (b) The onset rate of the channel current can be fitted to a single exponential decay function, I = Imax (1−e(−t/τ)). The fitting results for the onset rate τ and Imax for channels pre-incubated with isoflurane (red line) are 1.73 ± 0.04 s and −0.33 ± 0.02 μA, respectively; and 1.25 ± 0.05 s and −0.41 ± 0.03 μA, respectively, for channels without isoflurane pre-incubation (green line). The fitting parameters are expressed as mean ± SEM (n = 15).
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f4: Isoflurane binding stabilized the closed channel.(a) A representative trace showing that pre-incubation with isoflurane (red arrows) slowed the onset rate and increased the maximum inhibition. The vertical and horizontal scale bars represent 20 μA and 30 seconds, representatively. (b) The onset rate of the channel current can be fitted to a single exponential decay function, I = Imax (1−e(−t/τ)). The fitting results for the onset rate τ and Imax for channels pre-incubated with isoflurane (red line) are 1.73 ± 0.04 s and −0.33 ± 0.02 μA, respectively; and 1.25 ± 0.05 s and −0.41 ± 0.03 μA, respectively, for channels without isoflurane pre-incubation (green line). The fitting parameters are expressed as mean ± SEM (n = 15).

Mentions: Since isoflurane binds to the pore of the resting state of ELIC (Fig. 2a), pre-incubating the oocytes expressing ELIC with isoflurane for a short period of time is expected to enhance the inhibitory effect. As shown in Fig. 4, pre-incubation with isoflurane not only generated greater inhibition, but also decreased the apparent onset rate of agonist-induced current and slowed channel opening. Data indicate that isoflurane binding stabilizes the closed-channel conformation.


Direct Pore Binding as a Mechanism for Isoflurane Inhibition of the Pentameric Ligand-gated Ion Channel ELIC.

Chen Q, Kinde MN, Arjunan P, Wells MM, Cohen AE, Xu Y, Tang P - Sci Rep (2015)

Isoflurane binding stabilized the closed channel.(a) A representative trace showing that pre-incubation with isoflurane (red arrows) slowed the onset rate and increased the maximum inhibition. The vertical and horizontal scale bars represent 20 μA and 30 seconds, representatively. (b) The onset rate of the channel current can be fitted to a single exponential decay function, I = Imax (1−e(−t/τ)). The fitting results for the onset rate τ and Imax for channels pre-incubated with isoflurane (red line) are 1.73 ± 0.04 s and −0.33 ± 0.02 μA, respectively; and 1.25 ± 0.05 s and −0.41 ± 0.03 μA, respectively, for channels without isoflurane pre-incubation (green line). The fitting parameters are expressed as mean ± SEM (n = 15).
© Copyright Policy - open-access
Related In: Results  -  Collection

License
Show All Figures
getmorefigures.php?uid=PMC4561908&req=5

f4: Isoflurane binding stabilized the closed channel.(a) A representative trace showing that pre-incubation with isoflurane (red arrows) slowed the onset rate and increased the maximum inhibition. The vertical and horizontal scale bars represent 20 μA and 30 seconds, representatively. (b) The onset rate of the channel current can be fitted to a single exponential decay function, I = Imax (1−e(−t/τ)). The fitting results for the onset rate τ and Imax for channels pre-incubated with isoflurane (red line) are 1.73 ± 0.04 s and −0.33 ± 0.02 μA, respectively; and 1.25 ± 0.05 s and −0.41 ± 0.03 μA, respectively, for channels without isoflurane pre-incubation (green line). The fitting parameters are expressed as mean ± SEM (n = 15).
Mentions: Since isoflurane binds to the pore of the resting state of ELIC (Fig. 2a), pre-incubating the oocytes expressing ELIC with isoflurane for a short period of time is expected to enhance the inhibitory effect. As shown in Fig. 4, pre-incubation with isoflurane not only generated greater inhibition, but also decreased the apparent onset rate of agonist-induced current and slowed channel opening. Data indicate that isoflurane binding stabilizes the closed-channel conformation.

Bottom Line: Electrophysiology measurements with a single-point mutation at position 6' or 13' support the notion that binding at these sites renders isoflurane inhibition.Molecular dynamics simulations suggested that isoflurane binding was more stable in the resting than in a desensitized pore conformation.This study presents compelling evidence for a direct pore-binding mechanism of isoflurane inhibition, which has a general implication for inhibitory action of general anesthetics on pLGICs.

View Article: PubMed Central - PubMed

Affiliation: Department of Anesthesiology, University of Pittsburgh School of Medicine, Pittsburgh, Pennsylvania, 15260, USA.

ABSTRACT
Pentameric ligand-gated ion channels (pLGICs) are targets of general anesthetics, but molecular mechanisms underlying anesthetic action remain debatable. We found that ELIC, a pLGIC from Erwinia chrysanthemi, can be functionally inhibited by isoflurane and other anesthetics. Structures of ELIC co-crystallized with isoflurane in the absence or presence of an agonist revealed double isoflurane occupancies inside the pore near T237(6') and A244(13'). A pore-radius contraction near the extracellular entrance was observed upon isoflurane binding. Electrophysiology measurements with a single-point mutation at position 6' or 13' support the notion that binding at these sites renders isoflurane inhibition. Molecular dynamics simulations suggested that isoflurane binding was more stable in the resting than in a desensitized pore conformation. This study presents compelling evidence for a direct pore-binding mechanism of isoflurane inhibition, which has a general implication for inhibitory action of general anesthetics on pLGICs.

No MeSH data available.


Related in: MedlinePlus