• The Korean Journal of Physiology and Pharmacology
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• v.20 no.5
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• pp.525-531
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• 2016

The analgesic mechanism of opioids is known to decrease the excitability of substantia gelatinosa (SG) neurons receiving the synaptic inputs from primary nociceptive afferent fiber by increasing inwardly rectifying $K^+$ current. In this study, we examined whether a ${\mu}$-opioid agonist, [D-Ala2,N-Me-Phe4, Gly5-ol]-enkephalin (DAMGO), affects the two-pore domain $K^+$ channel (K2P) current in rat SG neurons using a slice whole-cell patch clamp technique. Also we confirmed which subtypes of K2P channels were associated with DAMGO-induced currents, measuring the expression of K2P channel in whole spinal cord and SG region. DAMGO caused a robust hyperpolarization and outward current in the SG neurons, which developed almost instantaneously and did not show any time-dependent inactivation. Half of the SG neurons exhibited a linear I~V relationship of the DAMGO-induced current, whereas rest of the neurons displayed inward rectification. In SG neurons with a linear I~V relationship of DAMGO-induced current, the reversal potential was close to the $K^+$ equilibrium potentials. The mRNA expression of TWIK (tandem of pore domains in a weak inwardly rectifying $K^+$ channel) related acid-sensitive $K^+$ channel (TASK) 1 and 3 was found in the SG region and a low pH (6.4) significantly blocked the DAMGO-induced $K^+$ current. Taken together, the DAMGO-induced hyperpolarization at resting membrane potential and subsequent decrease in excitability of SG neurons can be carried by the two-pore domain $K^+$ channel (TASK1 and 3) in addition to inwardly rectifying $K^+$ channel.

• Journal of Embryo Transfer
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• v.26 no.3
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• pp.209-214
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• 2011

Endometrium undergoing hormonal change plays important roles in preparation for implantation, fetal growth, and well-being. During pregnancy, cellular remodeling and hormonal changes in endometrium could change two-pore domain K$^+$ channel (K$_{2P}$) expression. This study was performed to identify whether K$_{2P}$ channel expression is changed in endometrium of pregnant Korean cattle, and whether the expression level is modulated by progesterone treatment. We investigated changes in the mRNA and protein expressions of K$_{2P}$ channel in pregnant endometrium using RT-PCR and Western blot analyses. The expression levels of all K$_{2P}$ channel mRNAs tested in this study, except that of TREK-1, were changed in the pregnant endometrium. mRNA levels of TASK-3 and TRAAK were significantly down-regulated, whereas those of TREK-2 and TRESK were up-regulated in the pregnant endometrium. In parallel with the RT-PCR results, Western blot analysis revealed up-regulations of TREK-2 (7.9-fold) and TRESK (2-fold) proteins levels in the pregnant endometrium. In addition, TREK-2 and TRESK protein levels were up-regulated in bovine endometrial cells by progesterone treatment (10 ${\mu}g$/ml). From these results, we suggest that the up-regulation of TREK-2 and TRESK by progesterone may contribute to the regulation of physiological changes during pregnancy.

• Journal of Embryo Transfer
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• v.24 no.3
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• pp.189-197
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• 2009

Two-pore domain $K^+(K_{2P})$ channels contribute to setting the resting membrane potential in excitable and nonexcitable cells. However, the cellular or tissue distribution and function of $K_{2P}$ channels expressed in mammalian germ cells and reproductive organs have not yet been reviewed by researchers. In this review, we focus on expression, localization and expected properties of $K_{2P}$ channels in germ cells and reproductive organs. The $K_{2P}$ channels are expressed in human cytotrophoblast cells, myometrium, placental vascular system, uterine smooth muscle, and pregnant term tissue, suggesting that $K_{2P}$ channels might be involved in the processes of pregnance. The $K_{2P}$ channels are also expressed in mouse zygotes, monkey sperm, ovary, testis, germ cells, and embryos of Korean cattle. Interestingly, $K_{2P}$ channels are modulated by changes in temperature and oxygen concentration which play an important role in embryonic development. Also, $K_{2P}$ channels are responsible for $K^+$ efflux during apoptotic volume decreases in mouse zygotes. These expression patterns and properties of the $K_{2P}$ channels in reproductive organs and germ cells are likely to help the understanding of ion channel-related function in reproductive physiology.

• ALGAE
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• v.36 no.4
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• pp.315-326
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• 2021

Ion channels are membrane protein complexes mediating passive ion flux across the cell membranes. Every organism has a certain set of ion channels that define its physiology. Dinoflagellates are ecologically important microorganisms characterized by effective physiological adaptability, which backs up their massive proliferations that often result in harmful blooms (red tides). In this study, we used a bioinformatics approach to identify homologs of known ion channels that belong to 36 ion channel families. We demonstrated that the versatility of the dinoflagellate physiology is underpinned by a high diversity of ion channels including homologs of animal and plant proteins, as well as channels unique to protists. The analysis of 27 transcriptomes allowed reconstructing a consensus ion channel repertoire (channelome) of dinoflagellates including the members of 31 ion channel families: inwardly-rectifying potassium channels, two-pore domain potassium channels, voltage-gated potassium channels (K_v), tandem K_v, cyclic nucleotide-binding domain-containing channels (CNBD), tandem CNBD, eukaryotic ionotropic glutamate receptors, large-conductance calcium-activated potassium channels, intermediate/small-conductance calcium-activated potassium channels, eukaryotic single-domain voltage-gated cation channels, transient receptor potential channels, two-pore domain calcium channels, four-domain voltage-gated cation channels, cation and anion Cys-loop receptors, small-conductivity mechanosensitive channels, large-conductivity mechanosensitive channels, voltage-gated proton channels, inositole-1,4,5-trisphosphate receptors, slow anion channels, aluminum-activated malate transporters and quick anion channels, mitochondrial calcium uniporters, voltage-dependent anion channels, vesicular chloride channels, ionotropic purinergic receptors, animal volage-insensitive cation channels, channelrhodopsins, bestrophins, voltage-gated chloride channels H⁺/Cl^- exchangers, plant calcium-permeable mechanosensitive channels, and trimeric intracellular cation channels. Overall, dinoflagellates represent cells able to respond to physical and chemical stimuli utilizing a wide range of G-protein coupled receptors- and Ca²⁺-dependent signaling pathways. The applied approach not only shed light on the ion channel set in dinoflagellates, but also provided the information on possible molecular mechanisms underlying vital cellular processes dependent on the ion transport.

• The Korean Journal of Physiology and Pharmacology
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• v.20 no.5
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• pp.547-556
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• 2016

Myometrial relaxation of mouse via expression of two-pore domain acid sensitive (TASK) channels was studied. In our previous report, we suggested that two-pore domain acid-sensing $K^+$ channels (TASK-2) might be one of the candidates for the regulation of uterine circular smooth muscles in mice. In this study, we tried to show the mechanisms of relaxation via TASK-2 channels in marine myometrium. Isometric contraction measurements and patch clamp technique were used to verify TASK conductance in murine myometrium. Western blot and immunehistochemical study under confocal microscopy were used to investigate molecular identity of TASK channel. In this study, we showed that TEA and 4-AP insensitive non-inactivating outward $K^+$ current (NIOK) may be responsible for the quiescence of murine pregnant longitudinal myometrium. The characteristics of NIOK coincided with two-pore domain acid-sensing $K^+$ channels (TASK-2). NIOK in the presence of $K^+$ channel blockers was inhibited further by TASK inhibitors such as quinidine, bupivacaine, lidocaine, and extracellular acidosis. Furthermore, oxytocin and estrogen inhibited NIOK in pregnant myometrium. When compared to non-pregnant myometrium, pregnant myometrium showed stronger inhibition of NIOK by quinidine and increased immunohistochemical expression of TASK-2. Finally, TASK-2 inhibitors induced strong myometrial contraction even in the presence of L-methionine, a known inhibitor of stretch-activated channels in the longitudinal myometrium of mouse. Activation of TASK-2 channels seems to play an essential role for relaxing uterus during pregnancy and it might be one of the alternatives for preventing preterm delivery.

• Journal of Embryo Transfer
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• v.22 no.3
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• pp.149-154
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• 2007

Endometrial cells play important roles in implantation and during pregnancy. This study was carried out to identify whether two-pore domain $K^+\;(K_{2P})$ channels are expressed in endometrial cells of Korean cattle. $K_{2P}$ channels set the resting membrane potential of many kinds of neuronal cells in the central and peripheral nervous systems. RT-PCR data showed that TASK-1, TASK-3, TREK-1, TREK-2, and TRAAK were expressed in bovine endometrial cells, and the mRNA expression levels were similar between endometrial cells with or without endometritis. The protein expression was confirmed by using commercially available polyclonal antibodies (TASK-3, TREK-1, TREK-2, and TRAAK). TASK-3 and TREK-1 were expressed in all area of endometrial cells including nuclei, while TREK-2 and TRAAK were expressed in all area of cells except nuclei. These results demonstrate for the first time the presence of $K_{2P}$ channel in endometrial cells of Korean cattle.

• Journal of the Korea Academia-Industrial cooperation Society
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• v.17 no.3
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• pp.127-135
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• 2016

Two-pore domain potassium (K2P) channels are the targets of physiological stimuli, such as intracellular pH, bioactive lipids, and neurotransmitters, and they set the resting membrane potential. Some types of K2P channels play a critical role in both apoptosis and tumoriogenesis. Among the K2P channels, no antagonists of the TREK2 channel have been reported. The aim of the present study was to determine if the TREK2 channel is blocked and whether cell proliferation is influenced by flavonoids in the TREK2 overexpressing HEK293 cells (HEKT2). The electrophysiological current was recorded using single channel patch clamp techniques and cell proliferation was measured using a XTT assay. The electrophysiological results showed that the TREK2 channel activity was reduced to $91.5{\pm}13.1%$ (n=5) and $82.2{\pm}13.7%$ (n=5) by flavonoids, such as epigallocatechin-3-gallate (EGCG) and quercetin in HEKT2 cells, respectively. In contrast, the EGCG analogue, epicatechin (EC), had no significant inhibitory effects on the TREK2 single channel activity. In addition, cell proliferation was reduced to $69.4{\pm}14.0%$ (n=4) by ECGG in the HEKT2 cells. From these results, EGCG and quercetin represent the first known TREK2 channel inhibitors and only EGCG reduced HEKT2 cell proliferation. This suggests that the flavonoids may work primarily by inhibiting the TREK2 channel, leading to a change in the resting membrane potential, and triggering the initiation of a change in intracellular signaling for cell proliferation. TREK2 channel may, at least in part, contribute to cell proliferation.

• The Korean Journal of Physiology and Pharmacology
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• v.12 no.5
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• pp.245-251
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• 2008

Single-channel recordings of TASK-1 and TASK-3, members of two-pore domain $K^+$ channel family, have not yet been reported in dorsal root ganglion (DRG) neurons, even though their mRNA and activity in whole-cell currents have been detected in these neurons. Here, we report single-channel kinetics of the TASK-3-like $K^+$ channel in DRG neurons and up-regulation of TASK-3 mRNA expression in tissues isolated from animals with spinal cord injury (SCI). In DRG neurons, the single-channel conductance of TASK-3-like $K^+$ channel was $33.0{\pm}0.1$ pS at - 60 mV, and TASK-3 activity fell by $65{\pm}5%$ when the extracellular pH was changed from 7.3 to 6.3, indicating that the DRG $K^+$ channel is similar to cloned TASK-3 channel. TASK-3 mRNA and protein levels in brain, spinal cord, and DRG were significantly higher in injured animals than in sham-operated ones. These results indicate that TASK-3 channels are expressed and functional in DRG neurons and the expression level is up-regulated following SCI, and suggest that TASK-3 channel could act as a potential background $K^+$ channel under SCI-induced acidic condition.

• Journal of Embryo Transfer
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• v.30 no.4
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• pp.277-282
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• 2015

Cryopreservation affects osmotic tolerance and intracellular ion concentration through changes in expression levels of water and ion channels. Control of these changes is important for cell survival after cryopreservation. Relatively little is known about changes in $K^+$ channel expression compared to water channel expression. This study was performed to investigate changes in TASK-2 channel (KCNK5: potassium channel, subfamily K, member 5), a member of two-pore domain $K^+$ channel family, in cryopreserved mouse ovaries. Cryopreservation increased TASK-2 mRNA expression in mouse ovaries. In addition, TASK-2 protein expression was upregulated in vitrified and slowly frozen ovaries. TASK-2 protein was expressed in all area of granulosa cells that surround the oocyte within the follicle, except nucleus. Viability of cells overexpressed with TASK-2 was higher than that of vector-transfected cells. Our results found that TASK-2 expression was increased by cryopreservation and overexpression of TASK-2 decreased cryopreservation-induced cell death. These results suggest that TASK-2 upregulation might reduce cryodamage.

• Journal of the Korea Academia-Industrial cooperation Society
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• v.12 no.6
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• pp.2660-2667
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• 2011

TREK-1 channel is a member of the two-pore domain potassium (K2P) channel family that is regulated by intracellular pH, membrane stretch, polyunsaturated fatty acids, temperature, and some neuroprotectant agents. TREK-1 channel can influence neuronal excitability by regulating leakage of potassium ions and resting membrane potential. TREK-1 channel has been shown to be overexpressed in prostate cancer cells. Although the importance of these properties, relatively little is known about flavonoid effects in the regulations of TREK-1 channel. The purpose of the study was to screening of flavonoids as the TREK-1 channel modulator using one of electrophysiological techniques such as excised inside-out patch configuration. We demonstrated blocking effect on TREK-1 channel by flavonoids such as epigallocatechin-3-gallate (EGCG), curcumin and quercetin in CHO cells transiently expressing TREK-1 channel. The inhibition of TREK-1 channel by quercetin and curcumin was reversible, whereas EGCG was little reversible. Quercetin, EGCG and curcumin decreased the relative channel activity to 73%, 91% and 94%, respectively. The half-inhibitory concentration (IC50) of curcumin, quercetin and EGCG was $1.04{\pm}0.19\;{\mu}M$, $1.13{\pm}0.26\;{\mu}M$ and $13.5{\pm}2.20\;{\mu}M$ in CHO cells expressing TREK-1 channel, respectively. These results indicate that flavonoids might regulate TREK-1 and this regulation might be one of the pharmacological actions of flavonoid in nervous systems and cancer cells.