Norfluoxetine inhibits TREK-2 K2P channels by multiple mechanisms including state-independent effects on the selectivity filter gate

Proks, Peter; Schewe, Marcus; Conrad, Linus J.; Rao, Shanlin; Rathje, Kristin; Rodstrom, K.E.J.; Carpenter, E.P.; Baukrowitz, Thomas; Tucker, Stephen J.

doi:10.1085/jgp.202012812

Cited by 26 publications

(38 citation statements)

References 45 publications

(74 reference statements)

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“…Interestingly, in TREK-2 channels, both BL1249 and high ionic-strength resulted in open state peaks in the amplitude histogram that were substantially more symmetric. We have not observed this in other conditions for TREK-2 [ 23 ] and it might therefore be explained by a stabilizing effect of BL1249 and K + on the open state of the filter gate that results in reduced fluctuations of either brief openings, or of subconductance states.…”

Section: Discussionmentioning

confidence: 62%

“…In all previous cases where we have recorded TREK-2 single-channels, the amplitude histograms exhibited skewed distributions, especially in the inward direction [ 17 , 22 , 23 ]. This distortion is likely produced either by very short openings that are not fully resolved, or by a smaller subconductance state, or a combination thereof.…”

Section: Introductionmentioning

confidence: 93%

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Effects of ionic strength on gating and permeation of TREK-2 K2P channels

2021

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In addition to the classical voltage-dependent behavior mediated by the voltage-sensing-domains (VSD) of ion channels, a growing number of voltage-dependent gating behaviors are being described in channels that lack canonical VSDs. A common thread in their mechanism of action is the contribution of the permeating ion to this voltage sensing process. The polymodal K2P K+ channel, TREK2 responds to membrane voltage through a gating process mediated by the interaction of K+ with its selectivity filter. Recently, we found that this action can be modulated by small molecule agonists (e.g. BL1249) which appear to have an electrostatic influence on K+ binding within the inner cavity and produce an increase in the single-channel conductance of TREK-2 channels. Here, we directly probed this K+-dependent gating process by recording both macroscopic and single-channel currents of TREK-2 in the presence of high concentrations of internal K+. Surprisingly we found TREK-2 is inhibited by high internal K+ concentrations and that this is mediated by the concomitant increase in ionic-strength. However, we were still able to determine that the increase in single channel conductance in the presence of BL1249 was blunted in high ionic-strength, whilst its activatory effect (on channel open probability) persisted. These effects are consistent with an electrostatic mechanism of action of negatively charged activators such as BL1249 on permeation, but also suggest that their influence on channel gating is complex.

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Section: Discussionmentioning

confidence: 62%

Section: Introductionmentioning

confidence: 93%

Effects of ionic strength on gating and permeation of TREK-2 K2P channels

2021

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“…Background currents activated by muscimol in B35 cells were blocked in the presence of bicuculline and the TREK-2 inhibitor norfluoxetine [14]. These results strongly indicate that the background K + channel activated by muscimol is TREK-2.…”

Section: Activation Of Trek-2 By Gaba a Receptor Agonistsmentioning

confidence: 62%

“…In B35 cells, muscimol significantly increased the background current, and the muscimolactivated current was restored by washout and blocked in the presence of bicuculline (p < 0.05, Figure 4A). Norfluoxetine, a TREK-2 inhibitor [14], blocked muscimol-induced increase in the background current in B35 cells (Figure 4A). The currents were recorded in the presence of 4-aminopyridine (4-AP, 1 mM), BaCl 2 (1 mM), and TEA (1 mM) to rule out the involvement of other K + channels.…”

Section: Activation Of Trek-2 By Muscimolmentioning

confidence: 93%

Muscimol Directly Activates the TREK-2 Channel Expressed in GABAergic Neurons through Its N-Terminus

Kim

Kwon

Hur³

et al. 2021

IJMS

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The two-pore domain K+ (K2P) channel, which is involved in setting the resting membrane potential in neurons, is an essential target for receptor agonists. Activation of the γ-aminobutyric acid (GABA) receptors (GABAAR and GABABR) reduces cellular excitability through Cl- influx and K+ efflux in neurons. Relatively little is known about the link between GABAAR and the K+ channel. The present study was performed to identify the effect of GABAR agonists on K2P channel expression and activity in the neuroblastic B35 cells that maintain glutamic acid decarboxylase (GAD) activity and express GABA. TASK and TREK/TRAAK mRNA were expressed in B35 cells with a high level of TREK-2 and TRAAK. In addition, TREK/TRAAK proteins were detected in the GABAergic neurons obtained from GABA transgenic mice. Furthermore, TREK-2 mRNA and protein expression levels were markedly upregulated in B35 cells by GABAAR and GABABR agonists. In particular, muscimol, a GABAAR agonist, significantly increased TREK-2 expression and activity, but the effect was reduced in the presence of the GABAAR antagonist bicuculine or TREK-2 inhibitor norfluoxetine. In the whole-cell and single-channel patch configurations, muscimol increased TREK-2 activity, but the muscimol effect disappeared in the N-terminal deletion mutant. These results indicate that muscimol directly induces TREK-2 activation through the N-terminus and suggest that muscimol can reduce cellular excitability by activating the TREK-2 channel and by inducing Cl- influx in GABAergic neurons.

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“…TREK channels are also robustly regulated by protein interaction partners binding to the Ct. A-Kinase Anchoring Protein 150 (AKAP150), substantially increases channel activity, and prevents further activation by mechanical and other regulatory mechanisms [40]. The TREK channels are also targeted by pharmacological agents, volatile anesthetics increase the K + current [29,[41][42][43], and the antidepressant fluoxetine is an inhibitor of TREK-1 and TREK-2 [44][45][46][47][48][49]. The above wide range of factors are not subtle modulators, but one by one evokes a robust effect on TREK/TRAAK channel activity.…”

Section: Introductionmentioning

confidence: 99%

Negative Influence by the Force: Mechanically Induced Hyperpolarization via K2P Background Potassium Channels

Lengyel

Enyedi

Czirják

2021

IJMS

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The two-pore domain K2P subunits form background (leak) potassium channels, which are characterized by constitutive, although not necessarily constant activity, at all membrane potential values. Among the fifteen pore-forming K2P subunits encoded by the KCNK genes, the three members of the TREK subfamily, TREK-1, TREK-2, and TRAAK are mechanosensitive ion channels. Mechanically induced opening of these channels generally results in outward K+ current under physiological conditions, with consequent hyperpolarization and inhibition of membrane potential-dependent cellular functions. In the past decade, great advances have been made in the investigation of the molecular determinants of mechanosensation, and members of the TREK subfamily have emerged among the best-understood examples of mammalian ion channels directly influenced by the tension of the phospholipid bilayer. In parallel, the crucial contribution of mechano-gated TREK channels to the regulation of membrane potential in several cell types has been reported. In this review, we summarize the general principles underlying the mechanical activation of K2P channels, and focus on the physiological roles of mechanically induced hyperpolarization.

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Norfluoxetine inhibits TREK-2 K2P channels by multiple mechanisms including state-independent effects on the selectivity filter gate

Cited by 26 publications

References 45 publications

Effects of ionic strength on gating and permeation of TREK-2 K2P channels

Effects of ionic strength on gating and permeation of TREK-2 K2P channels

Muscimol Directly Activates the TREK-2 Channel Expressed in GABAergic Neurons through Its N-Terminus

Negative Influence by the Force: Mechanically Induced Hyperpolarization via K2P Background Potassium Channels

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