The Effect of Trifluoroacetic Acid, a Metabolite of Isoflurane on the ATP-sensitive Potassium Channel in Rabbit Ventricular Myocytes

Lee, Dong Jun,Kim, Sung Joo,Ha, Kyung Ho,Kim, Mun Cheol,Cho, Kang Hee 대한마취과학회 2002 Korean Journal of Anesthesiology Vol.43 No.6

Background: Activation of ATP-sensitive K^+ channels (K_ATP channels) in the cardiac muscle produces cardioprotective effects during myocardial ischemia. Previous experimental evidence indicates that volatile anesthetics exert beneficial actions in ischemic myocardium and enhance functional recovery of stunned myocardium. More recently, volatile anesthetics have been demonstrated to produce cardioprotective effects in sunned myocardium in vivo, and these effects are blocked by a K_ATP channel antagonist. This finding suggests that K_ATP channel activation by isoflurane may mediate amtiischemic effects. However, it was demonstrated that isoflurane inhibited K_ATP channel activity in rabbit ventricular myocytes. To explain the discrepancy, the present investigation tested the hypothesis that isoflurane and its metabolite, trifluoroacetic acid, contributes to the activation of K_ATP channel in rabbit ventricular myocytes. Methods: Single ventricular myocytes were isolated from rabbit heats by an enzymatic dissociation procedure. Single-channel currents were measured in inside-out patch configuration of the patch-clamp technique. The perfusing liquid was equilibrated with isoflurane by passing 100% O_2 through a vaporizer. Results: Isoflurane inhibited K_ATP channel activity without a change in the single-channel conductivity. Isoflurane decreased the burst duration and increased the interburst duration. In addition, isoflurane diminished the ATP sensitivity of K_ATP channels. Trifluoroacetic acid increased the burst duration and decreased the interburst duration without a change in the single-channel conductivity. Isoflurane and trifluoroacetic acid diminished the ATP sensitivity of _ATP channels. Conclusions: These results imply that isoflurane and its metabolite could mediate cardioprotective effects via K_ATP channel activation during myocardial ischemia. (Korean J Anesthesiol 2002; 43: S1~S12)

평지방막에 융합된 골격근의 single ATP-sensitive K⁺ channel의 이온투과성에 대한 연구

류판동,Ryu, Pan-dong 대한수의학회 1992 大韓獸醫學會誌 Vol.32 No.4

Properties of unitary ATP-sensitive $K^+$ channels were studied using planar lipid bilayer technique. Vesicles were prepared from bullfrog (Rana catesbeiana) skeletal muscle. ATP-sensitive $K^+$ (K (ATP)) channels were identified by their unitary conductance and sensitivity to ATP. In the symmetrical solution containing 200mM KCI, 10mM Hepes, 1mM EGTA and pH 7.2, single K (ATP) channels showed a linear current-voltage relations with slight inward rectification. Slope conductance at reversal potential was $60.1{\pm}0.43$ pS(n=3)). Micromolar ATP reversibly inhibited the channel activity when applied to the cytoplasmic side. In the range of -50~+50 mV, the channel activity was not voltage-dependent, but the channel gating within a burst was more frequent at negative voltage range. Varying the concentrations of external/internal KCl(mM) to 40/200, 200/200, 200/100 and 200/40 shifted reversal potentials to $-30.8{\pm}2.9$(n=3), $-1.1{\pm}2.7$(n=3), 10.5 and 30.6(mV), respecrivety. These reversal potentials were close to the expected values by the Nernst equation, indicating nearly ideal selectivity for $K^+$ over $Cl^-$. Under bi-ionic conditions of 200mM external test ions and 200mM internal $K^+$, the reversal potentials for each test ion/K pair were measured. The measured reversal potentials were used for the calculation of the releative permeability of alkali cations to $K^+$ ions using the Goldman-Hodgkin-Katz equation. The permeability sequence of 5 cations relative to $K^+$ was $K^+$(1), $Rb^+$(0.49), $Cs^+$(0.27), $Na^+$(0.027) and $Li^+$(0.021). This sequence was recognized as Eisenman's selectivity sequence IV. In addition, modelling the permeation of $K^+$ ion through ATP-sensitive $K^+$ channel revealed that a 3-barrier 2-site multiple occupancy model can reasonably predict the observed current-voltage relations.

Effects of Adenosine on the Ionic Channel Activated by Metabolic Inhibition in Rabbit Ventricular Myocytes

Han. Jin,Kim. Eui-Yong,Ho. Won-Kyung,Earm. Yung-E 대한생리학회 1996 대한생리학회지 Vol.30 No.1

The objective of the present study was to characterize the role of adenosine in regulation of ATP-sensitive K⁺ channel (K_ATP channel) activity in isolated rabbit ventricular myocytes using the patch clamp technique. Internal adenosine had little effects on KaTr channel activity. In an outside-out patch with intrapipette GTP and ATP, external adenosine stimulated K_ATP channel activity. In an inside-out Patch with intrapipette adenosine, ATP reduced K_ATP channel activity, and GTP stimulated K_ATP channel activity. Adenosine receptor activation shifted the half-maximal inhibition Of K_ATP channel from 70 to 241 μm. These results Suggest that activation of adenosine receptors stimulates K_ATP channels in rabbit ventricular myocytes by reducing the apparent affinity of the channel for ATP. The effect may be important for activating K_ATP channels during early phase of myocardial ischemia.

랫드 흑질 신경세포의 전기적 특성과 ATP-sensitive K⁺채널의 전류밀도

한성규,박진봉,류판동,Han, Seong-kyu,Park, Jin-bong,Ryu, Pan-dong 대한수의학회 2000 大韓獸醫學會誌 Vol.40 No.2

Substantia nigra is known to highly express glibenclamide binding site, a protein associated to ATP-sensitive $K^{+}$ ($K_{ATP}$) channel in the brain. However, the functional expression of $K_{ATP}$ channels in the area is not yet known. In this work, we attempted to estimate the functional expression of $K_{ATP}$ channels in neurons of the substantia nigra pars compacta (SNC) in young rats using slice patch clamp technique. Membrane properties and whole cell currents attributable to $K_{ATP}$ channel were examined by the current and voltage clamp method, respectively. In SNC, two sub-populations of neurons were identified. Type I (rhythmic) neurons had low frequency rebound action potentials ($4.5{\pm}0.25Hz$, n=75) with rhythmic pattern. Type II (phasic) neurons were characterized by faster firing ($22.7{\pm}3.16Hz$, n=12). Both time constants and membrane capacitance in rhythmic neurons ($34.0{\pm}1.27$ ms, $270.0{\pm}11.83$ pF) and phasic neurons ($23.7{\pm}4.16$ ms, $184{\pm}35.2$ pF) were also significantly different. The current density of $K_{ATP}$ channels was $6.1{\pm}1.47$ pA/pF (2.44~15.43 pA/pF, n=8) at rhythmic neurons of young rats. Our data show that in SNC there are two types of neurons with different electrical properties and the density of $K_{ATP}$, channel of rhythmic neuron is about 600 channels per neuron.

Identification of ATP-sensitive K<SUP>⁢</SUP> Conductances in Male Rat Major Pelvic Ganglion Neurons

Kyu-Sang Park,Seung-Kyu Cha,Keon-Il Lee,Jae Yeoul Jun,Seong-Woo Jeong,In Deok Kong,Joong Woo Lee 대한생리학회-대한약리학회 2002 The Korean Journal of Physiology & Pharmacology Vol.6 No.5

<P> Major pelvic ganglia (MPG) neurons are classified into sympathetic and parasympathetic neurons according to the electrophysiological properties; membrane capacitance (Cm), expression of T-type Ca<SUP>2⁢</SUP> channels, and the firing patterns during depolarization. In the present study, function and molecular expression of ATP-sensitive K<SUP>⁢</SUP> (K<SUB>ATP</SUB>) channels was investigated in MPG neurons of male rats. Only in parasympathetic MPG neurons showing phasic firing patterns, hyperpolarizing changes were elicited by the application of diazoxide, an activator of K<SUB>ATP</SUB> channels. Glibenclamide (10μM), a K<SUB>ATP</SUB> channel blocker, completely abolished the diazoxide-induced hyperpolarization. Diazoxide increased inward currents at high K<SUP>⁢</SUP> (90 mM) external solution, which was also blocked by glibenclamide. The metabolic inhibition by the treatment with mitochondrial respiratory chain inhibitors (rotenone and antimycin) hyperpolarized the resting membrane potential of parasympathetic neurons, which was not observed in sympathetic neurons. The hyperpolarizing response to metabolic inhibition was partially blocked by glibenclamide. RT-PCR analysis revealed that MPG neurons mainly expressed the K<SUB>ATP</SUB> channel subunits of Kir6.2 and SUR1. Our results suggest that MPG neurons have K<SUB>ATP</SUB> channels, mainly formed by Kir6.2 and SUR1, with phenotype-specificity, and that the conductance through this channel in parasympathetic neurons may contribute to the changes in excitability during hypoxia and/or metabolic inhibition.

Increased Expression of ATP-sensitive K<SUP>+</SUP> Channels Improves the Right Ventricular Tolerance to Hypoxia in Rabbit Hearts

Seong Woo Choi,Jun Seok Ahn,Hyoung Kyu Kim,Nari Kim,Tae-Hoon Choi,Sung-Woo Park,En A Ko,Won Sun Park,Dae-Kyu Song,Jin Han 대한생리학회-대한약리학회 2011 The Korean Journal of Physiology & Pharmacology Vol.15 No.5

ATP-sensitive K⁺ channels (K_ATP) are major component of preventing ischemia-reperfusion injury. However, there is little information regarding to the expressional difference of K_ATP and its function between left and right ventricles. In this study, we measured the lactate dehydrogenase release of rabbit heart slices in vitro and determined the difference of the K_ATP expression at the both ventricles by measuring the level of K_ATP-forming Kir6.2 (OcKir6.2) mRNA using in situ hybridization. The hearts were preconditioned with 15 min hypoxia and reoxygenated for 15 min before a hypoxic period of 60 min, followed by reoxygenation for 180 min. With hypoxic preconditioning (100% N₂) with 15 min, left ventricles (LV) showed higher release of LDH comparing with right ventricles (RV). Adding K_ATP blocker glibenclamide (10ՌM) prior to a hypoxic period of 60 min, hypoxic preconditioning effect of RV was more abolished than LV. With in situ hybridization, the optical density of OcKir6.2 was higher in RV. Therefore, we suggest that different K_ATP expression between LV and RV is responsible for the different response to hypoxia and hypoxic preconditioning of rabbit hearts.

ATP-sensitive K⁺ channels maintain resting membrane potential in interstitial cells of Cajal from the mouse colon

Na, Ji Sun,Hong, Chansik,Kim, Man Woo,Park, Chan Guk,Kang, Hyun Goo,Wu, Mei Jin,Jiao, Han Yi,Choi, Seok,Jun, Jae Yeoul Elsevier 2017 european journal of pharmacology Vol.809 No.-

<P>To investigate the role of ATP-sensitive K+(K-ATP) channels on pacemaker activity in interstitial cells of Cajal (ICC), whole-cell patch clamping, RT-PCR, and intracellular Ca2+([Ca2+](i)) imaging were performed in cultured colonic ICC. Pinacidil (a K+ channel opener) hyperpolarized the membrane and inhibited the generation of pacemaker potential, and this effect was, reversed by glibenclamide (a K-ATP channel blocker). RT-PCR showed that K-ir 6.1 and SUR2B were expressed in Ano-1 positive colonic ICC. Glibenclamide depolarized the membrane and increased pacemaker potential frequency. However, 5-hydroxydecanoic acid (a mitochondrial KATP channel blocker) had no effects on pacemaker potentials. Phorbol 12-myristate 13-acetate (PMA; a protein kinase C activator) blocked the pinacidil-induced effects, and PMA alone depolarized the membrane and increased pacemaker potential frequency. Cell-permeable 8-bromo-cyclic AMP also increased pacemaker potential frequency. Recordings of spontaneous intracellular Ca2+([Ca2+](i)) oscillations showed that glibenclamide increased the frequency of [Ca2+]; oscillations. In small intestinal ICC, glibenclamide alone did not alter the generation of pacemaker potentials, and K-ir 6.2 and SUR2B were expressed in Ano-1 positive ICC. Therefore, K-ATP channels in colonic ICC are activated in resting state and play an important role in maintaining resting membrane potential.</P>

Increased Expression of ATP-sensitive $K^+$ Channels Improves the Right Ventricular Tolerance to Hypoxia in Rabbit Hearts

Choi, Seong-Woo,Ahn, Jun-Seok,Kim, Hyoung-Kyu,Kim, Na-Ri,Choi, Tae-Hoon,Park, Sung-Woo,Ko, En-A,Park, Won-Sun,Song, Dae-Kyu,Han, Jin The Korean Society of Pharmacology 2011 The Korean Journal of Physiology & Pharmacology Vol.15 No.4

ATP-sensitive $K^+$ channels ($K_{ATP}$) are major component of preventing ischemia-reperfusion injury. However, there is little information regarding to the expressional difference of $K_{ATP}$ and its function between left and right ventricles. In this study, we measured the lactate dehydrogenase release of rabbit heart slices in vitro and determined the difference of the $K_{ATP}$ expression at the both ventricles by measuring the level of $K_{ATP}$-forming Kir6.2 (OcKir6.2) mRNA using in situ hybridization. The hearts were preconditioned with 15 min hypoxia and reoxygenated for 15 min before a hypoxic period of 60 min, followed by reoxygenation for 180 min. With hypoxic preconditioning (100% $N_2$) with 15 min, left ventricles (LV) showed higher release of LDH comparing with right ventricles (RV). Adding $K_{ATP}$ blocker glibenclamide ($10{\mu}M$) prior to a hypoxic period of 60 min, hypoxic preconditioning effect of RV was more abolished than LV. With in situ hybridization, the optical density of OcKir6.2 was higher in RV. Therefore, we suggest that different $K_{ATP}$ expression between LV and RV is responsible for the different response to hypoxia and hypoxic preconditioning of rabbit hearts.

Identification of ATP-sensitive $K^+$ Conductances in Male Rat Major Pelvic Ganglion Neurons

Park, Kyu-Sang,Cha, Seung-Kyu,Lee, Keon-Il,Jun, Jae-Yeoul,Jeong, Seong-Woo,Kong, In-Deok,Lee, Joong-Woo The Korean Society of Pharmacology 2002 The Korean Journal of Physiology & Pharmacology Vol.6 No.5

Major pelvic ganglia (MPG) neurons are classified into sympathetic and parasympathetic neurons according to the electrophysiological properties; membrane capacitance (Cm), expression of T-type $Ca^{2+}$ channels, and the firing patterns during depolarization. In the present study, function and molecular expression of ATP-sensitive $K^+\;(K_{ATP})$ channels was investigated in MPG neurons of male rats. Only in parasympathetic MPG neurons showing phasic firing patterns, hyperpolarizing changes were elicited by the application of diazoxide, an activator of $K_{ATP}$ channels. Glibenclamide $(10{\mu}M),$ a $K_{ATP}$ channel blocker, completely abolished the diazoxide-induced hyperpolarization. Diazoxide increased inward currents at high $K^+$ (90 mM) external solution, which was also blocked by glibenclamide. The metabolic inhibition by the treatment with mitochondrial respiratory chain inhibitors (rotenone and antimycin) hyperpolarized the resting membrane potential of parasympathetic neurons, which was not observed in sympathetic neurons. The hyperpolarizing response to metabolic inhibition was partially blocked by glibenclamide. RT-PCR analysis revealed that MPG neurons mainly expressed the $K_{ATP}$ channel subunits of Kir6.2 and SUR1. Our results suggest that MPG neurons have $K_{ATP}$ channels, mainly formed by Kir6.2 and SUR1, with phenotype-specificity, and that the conductance through this channel in parasympathetic neurons may contribute to the changes in excitability during hypoxia and/or metabolic inhibition.

Identification and Characterization of a Novel Bacterial ATP-Sensitive K+ Channel

최승범,Jong-Uk Kim,Hyun Joo,Churl K. Min 한국미생물학회 2010 The journal of microbiology Vol.48 No.3

Five bacterial species that are most likely to have putative prokaryotic inward rectifier K+ (Kir) channels were selected by in silico sequence homology and membrane topology analyses with respect to the number of transmembrane domains (TMs) and the presence of K+ selectivity filter and/or ATP binding sites in reference to rabbit heart inward rectifier K+ channel (Kir6.2). A dot blot assay with genomic DNAs when probed with whole rabbit Kir6.2 cDNA further supported the in silico analysis by exhibiting a stronger hybridization in species with putative Kir’s compared to one without a Kir. Among them, Chromobacterium violaceum gave rise to a putative Kir channel gene, which was PCR-cloned into the bacterial expression vector pET30b(+), and its expression was induced in Escherichia coli and confirmed by gel purification and immunoblotting. On the other hand, this putative bacterial Kir channel was functionally expressed in Xenopus oocytes and its channel activity was measured electrophysiologically by using two electrode voltage clamping (TEVC). Results revealed a K+ current with characteristics similar to those of the ATP-sensitive K+ (K-ATP) channel. Collectively, cloning and functional characterization of bacterial ion channels could be greatly facilitated by combining the in silico analysis and heterologous expression in Xenopus oocytes.