Trichloroethanol enhances the activity of recombinant human TREK-1 and TRAAK channels

Harinath, S.; Sikdar, SK

doi:10.1016/j.neuropharm.2003.11.023

Cited by 23 publications

(28 citation statements)

References 24 publications

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“…hTREK1, when expressed in infected HEK 293 cells, formed functional channels with kinetics similar to that described previously (Harinath and Sikdar, 2004). In physiological K ϩ gradient, an outwardly rectifying, noninactivating, time-dependent current progressively developed upon depolarization, which was absent in uninfected cells and Ad-GFP-infected cells (Supplemental Fig.…”

Section: Resultssupporting

confidence: 72%

“…The AdEasy-1 adenoviral vector system, including the pAdTrack-cytomegalovirus (shuttle vector), pAdEasy-1, and the electrocompetent cells of BJ5183 strain of Escherichia coli, was a generous gift from Prof. Bert Vogelstein (John Hopkins Oncology Center, Baltimore, MD). The Cterminal deletion mutants of hTREK1 (⌬89 and ⌬119 hTREK1) were constructed by PCR mutagenesis as described previously (Harinath and Sikdar, 2004), and the S348A mutant of hTREK1 was a generous gift from Prof. Steve A. N. Goldstein (Yale University Medical School). The other site-directed mutageneses such as the S348D, S366A, and Y352A/F355A were performed with a QuikChange kit (Stratagene, La Jolla, CA), and mutations were verified by DNA sequencing that is illustrated in Supplemental Fig.…”

Section: Methodsmentioning

confidence: 99%

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Inhibition of Human Two-Pore Domain K⁺ Channel TREK1 by Local Anesthetic Lidocaine: Negative Cooperativity and Half-of-Sites Saturation Kinetics

Nayak

Harinath²,

Nama³

et al. 2009

Mol Pharmacol

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TWIK-related Kϩ channel TREK1, a background leak K ϩ channel, has been strongly implicated as the target of several general and local anesthetics. Here, using the whole-cell and single-channel patch-clamp technique, we investigated the effect of lidocaine, a local anesthetic, on the human (h)TREK1 channel heterologously expressed in human embryonic kidney 293 cells by an adenoviral-mediated expression system. Lidocaine, at clinical concentrations, produced reversible, concentration-dependent inhibition of hTREK1 current, with IC 50 value of 180 M, by reducing the single-channel open probability and stabilizing the closed state. We have identified a strategically placed unique aromatic couplet (Tyr352 and Phe355) in the vicinity of the protein kinase A phosphorylation site, Ser348, in the C-terminal domain (CTD) of hTREK1, that is critical for the action of lidocaine. Furthermore, the phosphorylation state of Ser348 was found to have a regulatory role in lidocaine-mediated inhibition of hTREK1. It is interesting that we observed strong intersubunit negative cooperativity (Hill coefficient ϭ 0.49) and half-of-sites saturation binding stoichiometry (halfreaction order) for the binding of lidocaine to hTREK1. Studies with the heterodimer of wild-type (wt)-hTREK1 and ⌬119 Cterminal deletion mutant (hTREK1 wt -⌬119) revealed that single CTD of hTREK1 was capable of mediating partial inhibition by lidocaine, but complete inhibition necessitates the cooperative interaction between both the CTDs upon binding of lidocaine. Based on our observations, we propose a model that explains the unique kinetics and provides a plausible paradigm for the inhibitory action of lidocaine on hTREK1.

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Section: Resultssupporting

confidence: 72%

Section: Methodsmentioning

confidence: 99%

Inhibition of Human Two-Pore Domain K⁺ Channel TREK1 by Local Anesthetic Lidocaine: Negative Cooperativity and Half-of-Sites Saturation Kinetics

Nayak

Harinath²,

Nama³

et al. 2009

Mol Pharmacol

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“…Because of the localization of TREK-1 in the CNS, and its high sensitivity to clinically relevant concentrations of the volatile anesthetics, it has been proposed that TREK-1 is the best candidate to have an important role in anesthesia (268). In support of this assumption, further anesthetics, which do not influence GABA A receptors, chloral hydrate (127), N 2 O, xenon, and cyclopropane (121) were also found to activate TREK-1, but not TASK-1. Even more convincing evidence for the involvement of TREK-1 in the anesthetic action was provided by experiments on a knockout model.…”

Section: General Anesthesiamentioning

confidence: 99%

Molecular Background of Leak K⁺Currents: Two-Pore Domain Potassium Channels

Enyedi

Czirják

2010

Physiological Reviews

779

769

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Two-pore domain K+ (K2P) channels give rise to leak (also called background) K+ currents. The well-known role of background K+ currents is to stabilize the negative resting membrane potential and counterbalance depolarization. However, it has become apparent in the past decade (during the detailed examination of the cloned and corresponding native K2P channel types) that this primary hyperpolarizing action is not performed passively. The K2P channels are regulated by a wide variety of voltage-independent factors. Basic physicochemical parameters (e.g., pH, temperature, membrane stretch) and also several intracellular signaling pathways substantially and specifically modulate the different members of the six K2P channel subfamilies (TWIK, TREK, TASK, TALK, THIK, and TRESK). The deep implication in diverse physiological processes, the circumscribed expression pattern of the different channels, and the interesting pharmacological profile brought the K2P channel family into the spotlight. In this review, we focus on the physiological roles of K2P channels in the most extensively investigated cell types, with special emphasis on the molecular mechanisms of channel regulation.

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“…In our previous report we described the stable expression of human TREK-1 in CHO cells (Harinath and Sikdar, 2004). We utilized the same cell-line for experiments in the present study.…”

Section: Resultsmentioning

confidence: 99%

“…A stable cell-line of CHO cells expressing human TREK-1 channels (CHO TREK-1 ; Harinath and Sikdar, 2004) was maintained in DMEM-F-12 HAM supplemented with 10% fetal bovine serum (v/v), 1% antibiotic-antimycotic (Sigma) and gentamicin (50 g) (Sigma) in a humidified incubator with an atmosphere of 5% CO 2 . When the cells were confluent, they were split and plated onto 35 mm culture dishes.…”

Section: Cell Culture and Transfectionmentioning

confidence: 99%

Inhibition of human TREK-1 channels by caffeine and theophylline

Harinath

Sikdar

2005

Epilepsy Research

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Caffeine (1,3,7-trimethylxanthine) and theophylline (1,3-dimethylxanthine) are used for therapeutic purposes and can cause life-threatening convulsive seizures due to systemic toxicity. The mechanisms for the epileptogenicity of caffeine and theophylline are not clear. TWIK-related K + channels (TREK-1) are highly expressed in the human central nervous system and have a major role in the control of neuronal excitability by regulating the resting membrane potential. In view of their physiological significance, inhibition of TREK-1 channels may be implicated in caffeine-and theophylline-induced seizures. We thus investigated, using whole-cell patch-clamp technique, modulation of hTREK-1 channels expressed in Chinese hamster ovary (CHO) cells by caffeine and theophylline. Caffeine and theophylline produced reversible inhibition of TREK-1 channels in a concentration-dependent manner. The half-maximal inhibitory concentrations (IC 50 ) for caffeine and theophylline were 377 ± 54 M and 486 ± 76 M, respectively. Caffeine and theophylline depolarized the membrane potential of CHO TREK-1 cells in a reversible and concentrationdependent manner. Inhibition by caffeine (5 mM) and theophylline (2 mM) was attenuated in TREK-1 channels with mutation of the PKA consensus sequence at serine 348, suggesting the involvement of cAMP/PKA pathway in the inhibitory process. Inhibition of TREK-1 channels and consequent membrane depolarization may contribute to the convulsive seizures induced by toxic levels of caffeine and theophylline.

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Trichloroethanol enhances the activity of recombinant human TREK-1 and TRAAK channels

Cited by 23 publications

References 24 publications

Inhibition of Human Two-Pore Domain K⁺ Channel TREK1 by Local Anesthetic Lidocaine: Negative Cooperativity and Half-of-Sites Saturation Kinetics

Inhibition of Human Two-Pore Domain K⁺ Channel TREK1 by Local Anesthetic Lidocaine: Negative Cooperativity and Half-of-Sites Saturation Kinetics

Molecular Background of Leak K⁺Currents: Two-Pore Domain Potassium Channels

Inhibition of human TREK-1 channels by caffeine and theophylline

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Trichloroethanol enhances the activity of recombinant human TREK-1 and TRAAK channels

Cited by 23 publications

References 24 publications

Inhibition of Human Two-Pore Domain K+ Channel TREK1 by Local Anesthetic Lidocaine: Negative Cooperativity and Half-of-Sites Saturation Kinetics

Inhibition of Human Two-Pore Domain K+ Channel TREK1 by Local Anesthetic Lidocaine: Negative Cooperativity and Half-of-Sites Saturation Kinetics

Molecular Background of Leak K+Currents: Two-Pore Domain Potassium Channels

Inhibition of human TREK-1 channels by caffeine and theophylline

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Inhibition of Human Two-Pore Domain K⁺ Channel TREK1 by Local Anesthetic Lidocaine: Negative Cooperativity and Half-of-Sites Saturation Kinetics

Inhibition of Human Two-Pore Domain K⁺ Channel TREK1 by Local Anesthetic Lidocaine: Negative Cooperativity and Half-of-Sites Saturation Kinetics

Molecular Background of Leak K⁺Currents: Two-Pore Domain Potassium Channels