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Characterization of TWIK-2, a two-pore domain K+ channel, cloned from the rat middle cerebral artery.

ABSTRACT: TWIK-2, a member of the Two-Pore Domain K channel family, is expressed in a number of mammalian tissues including the vascular system. The function of TWIK-2 is not known. The purpose of this study was to clone the TWIK-2 channel from the rat middle cerebral artery, express it in CHO cells, and characterize the channel's electrical properties. In light of the fact that there are no specific TWIK-2 inhibitors or activators, a better characterization of the channel should enhance our understanding of its role in the vascular system. TWIK-2 was cloned from the rat middle cerebral artery and expressed with an N-terminal green fluorescence protein (GFP) in CHO cells. We report that rTWIK-2-GFP currents were relatively linear at physiological K(+) concentrations but become slightly inwardly rectifying in symmetrical K(+). rTWIK-2-GFP was insensitive to 10 mM TEA, 3 mM 4-aminopyridine, and 10 microM glibenclamide. However, rTWIK-2-GFP was inhibited by Ba(2+) with 50% of the current being blocked at 80 microM. rTWIK-2-GFP activity was enhanced 60% by 100 microM arachidonic acid. The electrophysiological characteristics of TWIK-2 indicate that it could serve an important role in ion homeostasis and regulation of the membrane potential in arteries and arterioles.

SUBMITTER: Lloyd EE

PROVIDER: S-EPMC2847578 | biostudies-literature | 2009 Dec

REPOSITORIES: biostudies-literature

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Characterization of TWIK-2, a two-pore domain K+ channel, cloned from the rat middle cerebral artery.

Lloyd Eric E EE Marrelli Sean P SP Namiranian Khodadad K Bryan Robert M RM

Experimental biology and medicine (Maywood, N.J.) 20091201 12

TWIK-2, a member of the Two-Pore Domain K channel family, is expressed in a number of mammalian tissues including the vascular system. The function of TWIK-2 is not known. The purpose of this study was to clone the TWIK-2 channel from the rat middle cerebral artery, express it in CHO cells, and characterize the channel's electrical properties. In light of the fact that there are no specific TWIK-2 inhibitors or activators, a better characterization of the channel should enhance our understanding ...[more]

PMID: 19934370

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Project description:BACKGROUND AND PURPOSE:TREK two-pore-domain potassium (K2P ) channels play a critical role in regulating the excitability of somatosensory nociceptive neurons and are important mediators of pain perception. An understanding of the roles of TREK channels in pain perception and, indeed, in other pathophysiological conditions, has been severely hampered by the lack of potent and/or selective activators and inhibitors. In this study, we describe a new, selective opener of TREK channels, GI-530159. EXPERIMENTAL APPROACH:The effect of GI-530159 on TREK channels was demonstrated using 86 Rb efflux assays, whole-cell and single-channel patch-clamp recordings from recombinant TREK channels. The expression of K2P 2.1 (TREK1), K2P 10.1 (TREK2) and K2P 4.1 (TRAAK) channels was determined using transcriptome analysis from single dorsal root ganglion (DRG) cells. Current-clamp recordings from cultured rat DRG neurons were used to measure the effect of GI-530159 on neuronal excitability. KEY RESULTS:For recombinant human TREK1 channels, GI-530159 had similar low EC50 values in Rb efflux experiments and electrophysiological recordings. It activated TREK2 channels, but it had no detectable action on TRAAK channels nor any significant effect on other K channels tested. Current-clamp recordings from cultured rat DRG neurones showed that application of GI-530159 at 1 ?M resulted in a significant reduction in firing frequency and a small hyperpolarization of resting membrane potential. CONCLUSIONS AND IMPLICATIONS:This study provides pharmacological evidence for the presence of mechanosensitive TREK K2P channels in sensory neurones and suggests that development of selective K2P channel openers like GI-530159 could aid in the development of novel analgesic agents. LINKED ARTICLES:This article is part of a themed section on Recent Advances in Targeting Ion Channels to Treat Chronic Pain. To view the other articles in this section visit http://onlinelibrary.wiley.com/doi/10.1111/bph.v175.12/issuetoc.

Dataset Information

Characterization of TWIK-2, a two-pore domain K+ channel, cloned from the rat middle cerebral artery.

Publications

Characterization of TWIK-2, a two-pore domain K+ channel, cloned from the rat middle cerebral artery.

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