trifluoroacetic acid activates atp-sensitive k+ channels in rabbit ventricular myocytes

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Biochemical and Biophysical Research Communications 285, 1136–1142 (2001)

doi:10.1006/bbrc.2001.5291, available online at http://www.idealibrary.com on

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rifluoroacetic Acid Activates ATP-Sensitive K Channelsn Rabbit Ventricular Myocytes

in Han, Nari Kim, and Euiyong Kim1

epartment of Physiology and Biophysics, College of Medicine, Inje University,33-165 Gaegeum-Dong, Busanjin-Ku, Busan, 614-735 Korea

eceived June 27, 2001

K channel antagonists preclude the cardioprotectionc

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Recent in vivo experimental evidence suggests thatsoflurane-induced cardioprotection may involve KATP

hannel activation during myocardial ischemia. Thectual effect of isoflurane on cardioprotective ion con-uctance, however, such as that mediated by the open-

ng of KATP channels, has been the subject of someontroversy in the past. The investigation reportedere used a patch-clamp technique to test the hypoth-sis that a metabolite of isoflurane, trifluoroaceticcid (TFA), contributes to isoflurane-induced cardio-rotection via KATP channel activation. TFA enhancedhannel activity in a concentration-dependent fash-on, exhibiting half-maximal activation at 0.03 mM.FA increased the number of openings of the channel,ut did not affect the single channel conductance ofATP channels. Analysis of open and closed time distri-utions showed that TFA increased the burst durationnd decreased the interburst interval without elicit-ng changes of less than 5 ms in open and closed timeistributions. TFA diminished the ATP sensitivity ofATP channels in a concentration–response relation-

hip for ATP. These results imply that TFA could me-iate isoflurane-induced cardioprotection via KATP

hannel activation during myocardial ischemia andeperfusion. © 2001 Academic Press

Key Words: KATP channels; isoflurane; trifluoroaceticcid; patch clamp techniques; rabbit ventricularyocytes.

A brief period of ischemic preconditioning has beenroposed to mediate a protective effect against subse-uent, more severe episodes of ischemia (1). ATP-ensitive potassium (KATP) channels are thought to playrole in the phenomenon of ischemic preconditioning

n the heart (2). It has been reported that KATP channelpeners mediate cardioprotective effects (3), whereas

1 To whom correspondence should be addressed. Fax: 182 51 894500. E-mail: [email protected].

1136006-291X/01 $35.00opyright © 2001 by Academic Pressll rights of reproduction in any form reserved.

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onferred by ischemic preconditioning (4).The volatile anesthetic isoflurane also exerts car-

ioprotective effects during ischemia and reperfusion5–7). Numerous mechanisms have been proposed toxplain the cardioprotective action of isoflurane,ncluding reductions in myocardial oxygen consump-ion, beneficial alterations in intracellular calcium ho-eostasis, and the activation of ion channels (8). Re-

ently, both isolated whole-heart experiments (9, 10)nd a clinical study (5) have revealed that KATP channelctivation contributes to isoflurane-induced cardiopro-ection. However, controversy exists regarding the rolehat isoflurane plays in mediating cardioprotection viactivation of KATP channels. It seems that isofluranenhibits KATP channels directly (11), suggesting thatsoflurane may attenuate the cardioprotective effects of

ATP channels during ischemia and reperfusion in theyocardium. To date, possible explanations for the

iscrepancy between these two observations have noteen suggested.The objective of this study was to test the hypothesis

hat the major metabolite of isoflurane, trifluoroaceticcid (TFA), contributes to isoflurane-induced cardio-rotection via the activation of KATP channels. In thistudy, using isolated rabbit ventricular myocytes, wenvestigated the role of TFA in modulating KATP chan-els. The results of this study support the hypothesishat TFA enhances KATP channel activity and therebyontributes to the cardioprotective effects mediate byhe volatile anesthetic isoflurane.

ATERIALS AND METHODS

Cell isolation. Single ventricular myocytes were isolated fromabbit hearts by an enzymatic dissociation procedure, as discussedreviously (11). Briefly, in accordance with national animal-careuidelines, rabbits weighing 150–280 g were anesthetized with so-ium pentobarbital (50 mg/ml, 1 ml/kg body weight) and concomi-antly injected with heparin (300 IU/ml). After adequate anesthesiaas achieved, sternostomy was performed and the heart exposed.rtificial perfusion of the heart was established by cannulation of theorta. The heart was then removed and placed in a Langendorff

perfusion apparatus. Thereafter, an enzymatic method was used toi

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Vol. 285, No. 5, 2001 BIOCHEMICAL AND BIOPHYSICAL RESEARCH COMMUNICATIONS

solate single ventricular cells for electrophysiological experiments.

Electrophysiological recording and data analysis. Single-channelurrents were measured in the cell-attached and inside-out patchonfigurations of the patch-clamp technique (12). Channel activityas measured using a patch-clamp amplifier (EPC-7, LIST, Darm-

tadt, Germany; Axopatch-1D, Axon Instruments, Foster City, CA).ata were stored in digitized format (48 kHz digitization rate) onigital audiotapes using a DTR-1200 recorder (Biologic, Grenoble,rance), and evaluated off-line. They were subsequently filtered at.1–10 kHz, digitized at a sampling rate of 0.4–40 kHz and analyzedith a personal computer (IBM-PC, Pentium-III 450, Busan, Korea)sing pCLAMP (version 6.3 software, Axon Instruments, Inc., Bur-

ingame, CA) through an analogue-to-digital converter interfaceDigidata-1200, Axon Instruments, Inc., Burlingame, CA). The open-ime histogram was formed from continuous recordings of more than0 s. The open probability (P o) was calculated using the formula

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here t j is the time spent at current levels corresponding to j 5 0, 1,, . . . , N (superscript) channels in the open state, T d is the durationf the recording, and N is the number of channels active in the patch.he number of channels in a patch was estimated by dividing theaximum current observed, during an extended period at zero ATP,

y the mean unitary current amplitude. P o was calculated over 30-secords.

Solutions and drugs. The normal Tyrode solution contained: 143M NaCl, 5.4 mM KCl, 1.8 mM CaCl2, 0.5 mM MgCl2, 5.5 mM

lucose, and 5 mM N-2-hydroxyethylpiperazine-N9-2-ethanesulfoniccid (HEPES), adjusted to pH 7.4 with NaOH. The solutions facinghe outside of the cell membrane in the excised patch recordingsontained: 140 mM KCl, 2 mM CaCl2, 1 mM MgCl2, 10 mM glucose,nd 10 mM Hepes, adjusted to pH 7.4 with KOH. The solutionsacing the inside of the cell membrane in the excised patch recordingsontained: 127 mM KCl, 13 mM KOH, 1 mM MgCl2, 5 mM ethylenelycol-bis(-aminoethyl ether)-N,N,N9,N9-tetraacetic acid (EGTA), 10M glucose, and 10 mM Hepes, adjusted to pH 7.4 with KOH. Theodified KB solution had the following composition: 70 mM KOH, 50M L-glutamic acid, 40 mM KCl, 20 mM KH2PO4, 20 mM taurine, 3M MgCl2, 10 mM Hepes, 0.5 mM EGTA, and 10 mM glucose,

djusted to pH 7.4 with KOH. The change of pH that resulted fromddition of TFA was corrected by addition of KOH to take back to 7.4.he DAD-12 Superfusion System (Adams & List Associates, Nework, NY) was used to rapidly exchange (within 100 ms) the batholution and drugs in most experiments.All reagents used in this study were obtained from Sigma (St.

ouis, MO). Experiments were done at a room temperature of 25 6°C.

Statistical analysis. The data were statistically analyzed usingither Student’s unpaired t test when two treatment groups wereompared, or one-way analysis of variance (ANOVA) followed by aost hoc Student–Newman–Keuls test when all pairwise compari-ons among the different treatment groups were made. Tests wereonsidered significant when P , 0.05. All data are presented aseans 6 SE.

ESULTS AND DISCUSSION

To test the hypothesis that TFA is involved in thectivation of KATP channels, the effects of TFA werenvestigated with an inside-out patch configuration.FA (0.1 mM) reversibly enhanced KATP channel activ-

1137

ty, while the addition of 1 mM ATP or 0.5 mM gliben-lamide suppressed the TFA-induced KATP channel ac-ivities, confirming that observed channel activitiesere due to KATP channels (data not shown).Figure 1 shows the concentration-dependent effects of

FA on KATP channel activity. TFA increased KATP chan-el activity at concentrations as low as 0.001 mM andxhibited further activation effects in a concentration-ependent manner (Figs. 1A and 1B). The extent of acti-ation was normalized by the maximum response to 1M TFA, and elicited the concentration–response curve

hown in Fig. 1C. The plot of relative channel activities asfunction of the concentration of TFA ([TFA]i) was fitted

FIG. 1. (A) Representative current trace of concentration-ependent effects of TFA on KATP channel activity in an inside-outatch. The membrane potential was held at 240 mV. The solution-xchange protocol for each concentration of TFA is shown above theurrent trace. Data were sampled at 20 kHz and filtered at 1 kHz.he dashed line indicates the zero current level. (B) Changes in theo of KATP channels produced by different concentrations of TFA.alues are the mean 6 SE of P o. The line was drawn according to theill equation by the least-squares fit (n 5 8). (C) The relationshipetween the concentration of TFA and the relative channel activity ofhe KATP channel, which was obtained with reference to the valuenduced by 1 mM TFA in each patch. Values are the mean 6 SE from5 other experiments similar to that illustrated in A. The solid linen the graph was drawn from calculations that are described in theext.

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o the Hill equation using the least-squares method: Rel-tive channel activity 5 1/{1 1 (Kd/[TFA]i)

n}, where Kd 5TFA]i at the half-maximal activation of the channel, and

is the Hill coefficient. The Kd and n were 0.03 6 0.01M and 1.2 6 0.2, respectively (n 5 16 patches).The traces (Fig. 2A) and all-point amplitude histo-

rams (Fig. 2B) of the unit current are shown for theontrol (left) and 0.05 mM TFA (right) at 260 mV toemonstrate that single channel current amplitudesere not affected by TFA. Figure 2C shows the

urrent–voltage (I–V) relationships obtained from 15atches before and after the application of 0.05 mMFA. The I–V relationships before and after the appli-ation of TFA were linear in the negative membraneotential range, with slope conductance of 77.1 6 4.1nd 76.9 6 3.4 pS before and after 0.05 mM TFA,espectively. There were no statistical differences be-

FIG. 2. (A) The traces of the unit current before (left) and afterright) application of 0.05 mM TFA. The membrane potential waseld at 260 mV. Data were sampled at 20 kHz and filtered at 1 kHz.he dashed line indicates the zero current level. (B) Amplitudeistograms corresponding to the current traces in A. Arrowheads

ndicate peaks corresponding to the unit amplitude, which was 4.9efore (left) and 48 after (right) TFA application. (C) Current–oltage relationships of the unit current of KATP channel before (E)nd after (F) application of TFA. Values are means 6 SE. The lineas drawn by the linear least-squares fit to give a slope conductancef 77.1 pS before and 76.9 pS after the application of TFA.

1138

FA does not affect the conductance of single-channelurrents of KATP channels.

To examine the effect of TFA on the gating kinetics ofATP channels, the open-time and closed-time histo-

rams were calculated at a membrane potential of 250V negative to the reversal potential. The open time

istogram, which was analyzed from the currentecord filtered at a cutoff frequency of 10 kHz, revealedsingle exponential distribution with a time constant

to) of 1.5 ms for the control condition (Fig. 3A, left). Inhe presence of 0.05 mM TFA (Fig. 3A, right), the openime constant did not differ form that in the controlto 5 1.6 ms). The lifetime of a burst was defined as anpening period observed in the records filtered at autoff frequency of 0.1 kHz. The histogram of bursturation consisted of a single exponential distributionFig. 3C). Its time constant, designed as tb, was mark-dly prolonged by TFA (from 10 to 35 ms).The histogram of closed time within bursts was best

tted to a single exponential function (Fig. 3B). Thisnalysis was performed after closed times longer thanms were discarded, and filtered at a cutoff frequency

f 10 kHz. The time constant of the closed time withinursts was designed as tc. The value of tc was nothanged markedly by 0.05 mM TFA (from 0.31 to 0.30s). The closed time between bursts was analyzed by

sing records filtered at a cutoff frequency of 0.1 kHzFig. 3D). The histogram was fitted using a biexponen-ial function, with time constants of a fast (tc1) and alow component (tc2). The tc1 was equivalent to tc fil-ered at a cutoff frequency of 10 kHz, which was dis-orted by heavy filtering. The value of tc1 was notnfluenced by 0.05 mM TFA (from 18 to 14 ms). Thealue of tc2 was 808 ms for the control condition (Fig.D, left). This value was markedly decreased to 105 msy TFA (Fig. 3D, right). A quantitative analysis of thisnd five other patches is shown in Fig. 3E. Thus iteems that TFA does not influence the rapid, open andlosed times within the burst. Rather, it may increasehe number of openings within the burst (Fig. 5A) anday produce an increase in burst durations and a

ecrease in interburst intervals as shown in Fig. 3,esulting in the an increase in the channel activity.Interestingly, we found that 0.5 mM ATP produceduch less block in the presence of TFA than in its

bsence (data not shown), suggesting that the ATPensitivity of the channel may have been changed byFA. Therefore, to determine the effects of TFA on theTP sensitivity of KATP channels, different concentra-

ions of ATP were subsequently applied (Fig. 4). In theoncentration range 1 mM to 10 mM, ATP exerted areater inhibition of the channels in the absence ofFA (Fig. 4A) than in its presence (Fig. 4B). In Fig. 4C,he graph shows the dose–response relationship fornhibition of the P o by ATP before (E) and after (F) thepplication of TFA (Fig. 4C). From 25 patches, the

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nhibitory effects of ATP were quantified by measuringhe P o during ATP application, expressed relative to itsalue in ATP-free solution (Fig. 4D). The continuousines in the graph were fitted curves to the Hill equa-ion using the least-squares method: Relative channel

FIG. 3. Effects of TFA on the kinetic properties of KATP channel.ingle-channel currents were recorded at 250 mV in inside-outatches. Histograms of open (A) and closed (B) times within burstsere analyzed from current records filtered at a cutoff frequency of0 kHz. Histograms of burst (C) and interburst (D) durations werenalyzed from current records filtered at cutoff frequency of 0.1 kHz.n A–D, right panels are the histograms in the presence of 0.05 mMFA. Bin width is 0.05 ms. to, Open-time constant; tc, closed-timeonstant; tb, burst time constant; tc1, tc2, fast and slow components ofnterburst time constant, respectively. (E) Histogram showing theooled data (mean 6 SE) for tb and tc2. *Significant (P , 0.05)ifference from control value (n 5 6 patches).

1139

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TP concentration in the internal solution, K i is theTP concentration evoking the half-maximal inhibi-

ion, and n is the Hill coefficient. In control conditions,i and n were 71.5 6 7.6 and 1.9 6 0.6. In the presence

f TFA, the dose–response relationship for ATP hadhe same slope (1.7 6 0.7, P . 0.05) as in its absence,ut was shifted to higher ATP concentrations (K i 533.2 6 28.8 mM, P , 0.05).

FIG. 4. Representative current traces of the concentration-ependent effects of ATP on KATP channel activity in controls (A) inhe presence of 0.05 mM TFA (B). The membrane potential was heldt 240 mV. The solution-exchange protocol for each concentration ofTP is shown above the current traces. Data were sampled at 20 kHznd filtered at 1 kHz. The dashed line indicates the zero currentevel. (C, D) The relationship between the concentration of ATP andhe channel activity of the KATP channel in controls (E) and in theresence of 0.05 mM TFA (F). In C, the channel activity expressed inn absolute scale (P o). Values are the mean 6 SE of P o. The line wasrawn according to the Hill equation by the least-squares fit (n 50). In D, the relative channel activity was obtained with referenceo the value in ATP-free solution in each patch. Values are theean 6 SE from 24 other experiments similar to that illustrated inand B. The solid line in the graph was drawn from calculations

hat are described in the text. *Significantly different from controlalue (P , 0.05).

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To determine the interaction between TFA and glib-nclamide, the effects of TFA on the dose–responseelation for the channel inhibition for glibenclamideere examined (Fig. 5). In Fig. 5A, two types of channelpenings are represented; one is a KATP channel with aarge conductance and the other the inwardly rectify-ng K1 channel with a small conductance and longerurst-like openings. Application of 0.05 mM TFA pro-uced a significant increase in the channel activity andhe number of openings of the channel. TFA-inducedhannel activity as readily blocked by glibenclamideespite the continuous presence of TFA (Fig. 5A). Inhe absence of TFA, KATP channel activity was abol-shed by glibenclamide at concentrations in excess of0 mM, with an estimated inhibitory K i value and Hill

FIG. 5. Comparison of the effect of glibenclamide on KATP chan-el activity, in the absence and presence of TFA. (A) Representa-ive current traces of the concentration-dependent effects of glib-nclamide on KATP channel activity in the presence of 0.05 mMFA. The membrane potential was held at 240 mV. The solution-xchange protocol for each concentration of ATP is shown abovehe current traces. Data were sampled at 20 kHz and filtered at 1Hz. The dashed line indicates the zero current level. (B) Theelationship between the concentration of gliberclamide andhe relative channel activity of the KATP channel in controls (E)nd in the presence of 0.05 mM TFA (F). Relative effect of glib-nclamide was calculated in each patch as a ratio of P o measuredn the presence of glibenclamide to the value obtained in thebsence of glibenclamide. Solid line was drawn according to theollowing equation: Relative channel activity 5 1/{1 1[glibenclamide]/K i) n}, where [glibenclamide] is the glibenclamideoncentration in the internal solution, K i is the glibenclamideoncentration evoking the half-maximal inhibition, and n is theill coefficient.

1140

atches), respectively (Fig. 5B, E). In the presence of.05 mM TFA (Fig. 5B, F), the channel activity wasnhibited by glibenclamide with an apparent efficacyimilar to that obtained for the channel activity in thebsence of TFA. The K i and Hill coefficient were 0.13 6.04 mM and 0.98 6 0.03 (n 5 9 patches). Thus, it isikely that TFA and glibenclamide do not compete atlibenclamide binding sites. The Hill coefficients werelose to unity for both conditions, which suggests thatnly a single glibenclamide molecule need interactith its binding site to cause channel inhibition.Recent experimental and clinical evidence has indi-

ated that the volatile anesthetic isoflurane can pre-ondition the ischemic heart (7, 8). Although the un-erlying mechanism of this phenomenon has not beenompletely elucidated, much interest has focused on

ATP channels as a potential mechanism. This interests mainly supported by the fact that, in most experi-

ental and clinical studies, isoflurane-induced precon-itioning is abolished by glibenclamide. Thus, it haseen theorized that cardiac KATP channels would bectivated by isoflurane. However, few studies to dateave specifically determined whether isoflurane exertsdirect effect on KATP channels. Roscoe et al. (7) re-

orted that, in human atrial myocytes, isoflurane failso produce any significant changes in either inward orutward KATP currents in whole-cell voltage-clamp con-gurations when administered in concentrations asigh as 3%. Moreover, earlier results derived fromtudy of inside-out patches demonstrated that isoflu-ane inhibited KATP channels (11). The differences inxperimental conditions and animal species betweenur study and that of Roscoe et al. may be responsibleor the discrepancy in results, but these findings that

ATP channel activities are inhibited or are not affectedy isoflurane in experiments on isolated myocytes con-radict the majority of previous studies, both in vivond in vitro, which have demonstrated a cardioprotec-ion by isoflurane-induced KATP channel activation.

There are two possible explanations for the discrep-ncies in the data for isoflurane and its effect on KATP

hannel activity. First, isoflurane diminishes the ATPensitivity of KATP channels, indicating an increasedikelihood of KATP channel activity for a given concen-ration of ATP after isoflurane anesthesia (11). Second,soflurane activates KATP channels by its action on anpstream intermediate, such as an adenosine receptor7). In our previous study, we demonstrated that acti-ation of the adenosine A1 receptor activates KATP

hannels by reducing their sensitivity to ATP (13),upporting the second possibility.In this study, we suggest another possibility for the

bserved effect of isoflurane, namely that its metabo-ites modulate KATP channels. In humans, the principal

etabolite of isoflurane is TFA, which predominates,n addition to fluoride ion and small quantities of other

unidentified organic fluorides (14). All of the identifiedmtt

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etabolites of isoflurane are non-toxic and administra-ion of isoflurane has not been linked with anestheticoxicity.

We have shown here that TFA activated KATP chan-els by reducing the sensitivity of the KATP channel toTP. In this study, TFA was tested at concentrations

anging from 0.0001 to 1 mM. Another study has beenonducted demonstrating serum concentrations of TFAith known MAC hours of isoflurane, indicating thatnder these conditions (patients treated with isoniazidesulting in enzyme induction) the TFA concentrationsre greater than 0.01 mM (15). Furthermore, the peakoncentration of excreted TFA found in human urineas 1–1.3 mM (14), when patients were exposed to

soflurane at a concentration of 0.9%. Theoretically,hese amounts could precondition the heart. Combin-ng our observations and these data, it is clear that theoncentrations of TFA used in this study are of physi-logical relevance, and may be sufficient to activate

ATP channels in clinical situations.KATP channels have been identified in both cardiacyocytes (16) and in coronary vascular smooth muscle

ells (17). Activation of these channels in vascularmooth muscle by ischemia, hypoxia, or KATP channelgonists causes vasodilation. Isoflurane in vivo haslso been shown to produce coronary vasodilationhrough KATP channel activation (10, 17). Such find-ngs, and the results of the current investigation, sug-est that isoflurane and its metabolites should be stud-ed further with regard to their cardioprotectiveffects, which may be specifically related to their mod-lation of the KATP channels in coronary vascularmooth muscle cells. Therefore, to determine the na-ure of the interaction between isoflurane, its metabo-ites, and the KATP channels in coronary arterialmooth muscle cells, it will be necessary to recordingle-channels using patch clamp techniques on theseissues as well as to conduct isometric contraction ex-eriments in arterial rings.In conclusion, these data show that TFA, the pre-

ominant metabolite of isoflurane, activates KATP chan-els by an increase in burst durations and the numberf openings within burst, a decrease in interburst in-ervals and reducing the apparent affinity of the chan-el for ATP, suggesting that these mechanisms mayontribute, at least in part, to the isoflurane-inducedardioprotection during myocardial ischemia andeperfusion. To our knowledge, these are first databtained which relate to the direct effects of TFA on the

ATP channels.

CKNOWLEDGMENTS

This work was supported by Korea Research Foundation GrantKRF-2001-041-F00036). The authors wish especially to thankrofessor K. H. Cho, in the Department of Anesthesiology, Pro-

1141

logy, Professor J. Y. Jung, in the Institute of Malaria, Injeniversity, Professor S. H. Ko, in the Department of Anesthesi-

logy, Chonbuk National University, Professor W. G. Park, in theepartment of Anesthesiology, Yonsei University, and Professor. E. Earm and W. K. Ho, in the Department of Physiology,eoul National University, for their valuable comments to thisroject.

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2. Parratt, J. R., and Kane, K. A. (1994) KATP channels in ischaemicpreconditioning. Cardiovasc. Res. 28, 783–787.

3. Kersten, J. R., Montgomery, M. W., Pagel, P. S., and Warltier,D. C. (2000) Levosimendan, a new positive inotropic drug, de-creases myocardial infarct size via activation of KATP channels.Anesth. Analg. 90, 5–11.

4. Speechly-Dick, M. E., Grover, G. J., and Yellon, D. M. (1995)Does ischemic preconditioning in the human involve proteinkinase C and the ATP-dependent K1 channel? Studies of con-tractile function after simulated ischemia in an atrial in vitromodel. Circ. Res. 77, 1030–1035.

5. Belhomme, D., Peynet, J., Louzy, M., Launay, J. M., Kitakaze,M., and Menasche, P. (1999) Evidence for preconditioning byisoflurane in coronary artery bypass graft surgery. Circulation100, 340–344.

6. Ismaeil, M. S., Tkachenko, I., Gamperl, A. K., Hickey, R. F.,and Cason, B. A. (1999) Mechanisms of isoflurane-inducedmyocardial preconditioning in rabbits. Anesthesiology 90,812– 821.

7. Roscoe, A. K., Christensen, J. D., and Lynch, C. (2000) Isoflu-rane, but not halothane, induces protection of human myo-cardium via adenosine A 1 receptors and adenosinetriphosphate-sensitive potassium channels. Anesthesiology92, 1692–1701.

8. Piriou, V., Chiari, P., Knezynski, S., Bastien, O., Loufoua, J.,Lehot, J. J., Foex, P., Annat, G., and Ovize, M. (2000) Preventionof isoflurane-induced preconditioning by 5-hydroxydecanoateand gadolinium: Possible involvement of mitochondrial adeno-sine triphosphate-sensitive potassium and stretch-activatedchannels. Anesthesiology 93, 756–764.

9. Boutros, A., Wang, J., and Capuano, C. (1997) Isoflurane andhalothane increase adenosine triphosphate preservation, but donot provide additive recovery of function after ischemia, in pre-conditioned rat hearts. Anesthesiology 86, 109–117.

0. Crystal, G. J., Gurevicius, J., Salem, M. R., and Zhou, X. (1997)Role of adenosine triphosphate-sensitive potassium channels incoronary vasodilation by halothane, isoflurane, and enflurane.Anesthesiology 86, 448–458.

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2. Hamill, O. P., Marty, A., Neher, E., Sakmann, B., and Sigworth,F. J. (1981) Improved patch-clamp techniques for high-resolution current recording from cells and cell-free membranepatches. Pflugers Arch. 391, 85–100.

3. Kim, E., Han, J., Ho, W., and Earm, Y. E. (1997) Modulation ofATP-sensitive K1 channels in rabbit ventricular myocytes byadenosine A1 receptor activation. Am. J. Physiol. 272, H325–H333.

4. Hitt, B. A., Mazze, R. I., Cousins, M. J., Edmunds, H. N., Barr,

G. A., and Trudell, J. R. (1974) Metabolism of isoflurane in

1

16. Noma, A. (1983) ATP-regulated K1 channels in cardiac muscle.

1


Fischer 344 rats and man. Anesthesiology 40, 62–67.5. Gauntlett, I. S., Koblin, D. D., Fahey, M. R., Konopka, K.,

Gruenke, L. D., Waskell, L., and Eger, E. I. (1989) Metabolism ofisoflurane in patients receiving isoniazid. Anesth. Analg. 69,245–249.

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Gunther, K., and Goedel-Meinen, L. (1990) Hypoxic dilation ofcoronary arteries is mediated by ATP-sensitive potassium chan-nels. Science 247, 1341–1344.

trifluoroacetic acid activates atp-sensitive k+ channels in rabbit ventricular myocytes

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