Author + information
- Received March 29, 1999
- Revision received July 23, 1999
- Accepted September 13, 1999
- Published online January 1, 2000.
- Tetsuji Miura, MD, PhD∗,†,* (, )
- Yongge Liu, PhD†,
- Hiroyuki Kita, MD, PhD∗,†,
- Takashi Ogawa, MD, PhD∗,† and
- Kazuaki Shimamoto, MD, PhD∗,†
- ↵*Reprint requests and correspondence: Dr. Tetsuji Miura, Second Department of Internal Medicine, Sapporo Medical University School of Medicine, South-1, West-16, Chuo-ku, Sapporo 060-0061, Japan
This study intended to assess the role of mitochondrial ATP-sensitive potassium (mitoKatp) channels and the sequence of signal transduction with protein kinase C (PKC) and adenosine A1receptors in rabbits.
To our knowledge, the link between trigger receptors of preconditioning, PKC and mitoKatpchannels has not been examined in a whole heart model of infarction.
In the first series of experiments, myocardial infarction was induced in isolated buffer-perfused rabbit hearts by 30-min global ischemia and 2-h reperfusion. Infarct size in the left ventricle was determined by tetrazolium staining and expressed as a percentage of area at risk (i.e., the whole left ventricle) (%IS/AR). In the second series of experiments, mitochondria were isolated from the heart, and their respiratory function was examined using glutamate as a substrate.
Pretreatment with r-phenylisopropyladenosine (r-PIA, 1 μmol/liter), an A1-receptor agonist, reduced %IS/AR from 49.8 ± 6.5% to 13.4 ± 2.9%. This protection was abolished by calphostin C, a PKC inhibitor, and by 5-hydroxydecanoate (5-HD), a selective inhibitor of mitoKatpchannels. A selective mitoKatpchannel opener, diazoxide (100 μmol/liter), mimicked the effect of r-PIA on infarct size (%IS/AR = 11.6 ± 4.0%), and this protective effect was also abolished by 5-HD. However, calphostin C failed to block the infarct size–limiting effect of diazoxide. Neither calphostin C nor 5-HD alone modified %IS/AR. State III respiration (Qo2) and respiratory control index (RCI) were reduced after 30 min of ischemia (Qo2= 147.3 ± 5.3 vs. 108.5 ± 12.3, RCI = 22.3 ± 1.1 vs. 12.1 ± 1.8, p < 0.05). This mitochondrial dysfunction was persistent after 10 min of reperfusion (Qo2= 96.1 ± 15.5, RCI = 9.5 ± 1.9). Diazoxide significantly attenuated the respiratory dysfunction after 30 min of ischemia (Qo2= 142.8 ± 9.7, RCI = 16.2 ± 0.8) and subsequent 10-min reperfusion (Qo2= 135.3 ± 7.2, RCI = 19.1 ± 0.8).
These results suggest that mitoKatpchannels are downstream of PKC in the mechanism of infarct-size limitation by A1-receptor activation and that the anti-infarct tolerance afforded by opening of mitoKatpchannels is associated with preservation of mitochondrial function during ischemia/reperfusion.
Exposing the myocardium to brief transient ischemia paradoxically enhances its tolerance against subsequent ischemia insult. The mechanism of this phenomenon, termed “ischemic preconditioning,” has been a subject of intense investigation for a decade (1,2). Studies to date indicate that the preconditioning mechanism is triggered by activation of several classes of receptors, including adenosine and bradykinin receptors, which transmit signals to protein kinase C (PKC) and perhaps subsequent kinase cascades (1–3). However, the effector at the end of the signal transduction that is responsible for myocardial tolerance against ischemic cell death still has not been clarified. One possibility is atp-sensitive potassium channel (Katp) channels. Gross and Auchampach (4)first found that preconditioning protection is abolished by Katpchannel inhibitors and mimicked by Katpchannel openers, suggesting the involvement of this channel in preconditioning. This possibility was further supported by patch-clamp experiments showing that activation of this channel in sarcolemma is induced by adenosine A1receptor (5)and PKC (6). However, shortening of the action potential duration by either preconditioning (7)or a Katpchannel opener (8)did not correlate with the cardioprotection afforded by these manipulations, arguing against the possibility that Katpchannels in the sarcolemma are relevant to preconditioning protection. However, mitochondrial ATP-sensitive potassium channels (mitoKatps) channels, which are in the inner membrane of the mitochondria (9), have received attention as a possible effector of preconditioning, since the opening of mitoKatpchannels appears to be PKC-regulated (10)and cardioprotective against ischemic insult (11,12). In a recent study by Liu et al. (12), diazoxide, a Katpchannel opener, reversibly oxidized flavoproteins in the mitochondria of isolated cardiomyoytes, and this oxidation was inhibited by a selective mitoKatpchannel blocker, 5-hydroxydecanoate (5-HD) (10,13), indicating that the flavoprotein oxidation is due to opening of mitoKatpchannels. Furthermore, diazoxide significantly delayed cardiomyocyte death during 120 min of simulated ischemia, suggesting anti-infarct tolerance afforded by mitoKatpchannel opening (12).
However, to our knowledge, the link between trigger receptors of preconditioning (such as A1receptors), PKC and mitoKatpchannels has not been examined in a whole-heart model of infarction. It is also unknown whether changes in mitochondrial function by opening of mitoKatpchannels during ischemia/reperfusion contribute to infarct-size limitation by preconditioning. Accordingly, in the present study, we first compared an A1-receptor agonist (r-phenylisopropyladenosine [r-PIA]) with diazoxide in regard to infarct size–limiting effects and alteration of their cardioprotective effects by a PKC inhibitor (calphostin C) and a mitoKatpchannel blocker (5-HD). If the mitoKatpchannel is an effector downstream of PKC in the anti-infarct mechanism triggered by A1-receptor stimulation, both r-PIA and diazoxide should similarly limit infarct size, and their effects should be abolished by 5-HD. Furthermore, the effect of r-PIA, but not that of diazoxide, should be prevented by calphostin C. In the second series of experiments, the effects of diazoxide on mitochondrial dysfunction after ischemia and reperfusion were examined to get an insight into the role of mitochondrial function in mitoKatpchannel-mediated cardioprotection.
Experiment 1: role of mitoKatpand PKC in infarct size limitation by preischemic A1-receptor stimulation
Isolated rabbit heart preparation
Male albino rabbits (Japanese White), weighing 2.0 to 2.5 kg, were anesthetized by pentobarbital (30 mg/kg IV), underwent tracheostomies and were ventilated by a respirator (Harvard model 683). The chest was opened by left thoracotomy, and the heart was quickly excised and mounted on a nonrecirculating Langendorff apparatus with a water jacket. Before the heart excision, 2,000 U of heparin was injected intravenously to prevent thrombosis in the coronary arteries. The heart was perfused with a modified Krebs-Henseleit buffer at 37°C (NaCl, 118.5 mmol/liter; KCl, 4.7 mmol/liter; NaHCO3, 24.8 mmol/liter; KH2CO3, 1.2 mmol/liter; MgSO4, 1.2 mmol/liter; CaCl2, 2.5 mmol/liter; glucose, 10.0 mmol/liter), which was continuously oxygenated by 95% O2and 5% CO2. A 3F catheter tip manometer (SPR-524, Millar Instruments Inc., Houston, Texas) was inserted into the left ventricle via the left atrium to monitor ventricular pressure, and coronary flow was measured by collection of coronary effluent in a graduated cylinder. The heart was excluded from the study if the left ventricular systolic pressure was <70 mm Hg or arrhythmias were observed after a 15-min stabilization period.
After stabilization, all of the hearts underwent 30-min of global ischemia and 2-h reperfusion. To maintain myocardial temperature at 37°C during global ischemia, the temperature of the water circulating in the water jacket of the heart chamber was raised to 38°C, and the chamber was sealed with a thick plastic film during the ischemic period. Prior to global ischemia, each heart was subjected to one of the following nine treatments: no drug (untreated controls), diazoxide, 5-HD, diazoxide plus 5-HD, calphostin C, diazoxide plus calphostin C, r-PIA, r-PIA plus calphostin C and r-PIA plus 5-HD. Diazoxide (100 μmol/liter), 5-HD (30 μmol/liter) and r-PIA (1 μmol/liter) were infused for 10 min before the onset of ischemia. Calphostin C (200 nmol) was administered for 8 min before ischemia. The Krebs-Henseleit buffer containing these pharmacologic agents was kept in a reservoir separate from the main reservoir containing normal buffer. Since global no-flow ischemia was induced by clamping a perfusion line to the aorta, the pharmacologic agents stayed in the myocardium during the ischemic period. After 2 h of reperfusion, the heart was removed from the perfusion apparatus, weighed after removal of the atria, and frozen at −20°C.
Determination of infarct size
Infarct size was determined by the same method as that used in our previous studies (3,14). In brief, frozen hearts were sliced in parallel to the AV groove into approximately 2-mm sections and then incubated in 100 mmol/liter sodium phosphate buffer (pH = 7.4) containing 1% triphenyltetrazolium for 20 min at −37°C to visualize infarcts. The heart slices were mounted on a glass press, which compressed the slices uniformly to exactly 2 mm in thickness. A clear acetate sheet was overlaid on the glass press, and the outlines of the LV and infarcted area were traced on the acetate sheet. The traces were read by a computer (Macintosh Quadra 840AV) using a scanner (Hewlett Packard ScanJetIIC), and their areas were measured by NIH image, an image analysis software. The areas of infarct and the LV were multiplied by the thickness of the heart slice to obtain their volumes.
Experiment 2: effect of diazoxide on mitochondrial dysfunction after ischemia and reperfusion
Isolated rabbit heart preparation and experimental protocol
Isolated rabbit hearts were prepared and perfused as in experiment 1. Each heart was subjected either to no drug treatment or to 100 μmol/liter diazoxide for 10 min and rendered ischemic by clamping the perfusion line above the aorta. The left ventricular myocardium was sampled for mitochondria isolation at the end of 30-min ischemia or after 10 min of reperfusion following the 30-min ischemia. Myocardial samples from nonischemic hearts served as controls.
Isolation of mitochondria
Mitochondria were isolated from the ventricular sample according to the method of Vercesi et al. (15)with some modification. The sample (about 1.5 g) was weighed, washed and quickly minced with scissors in 5 ml of isolation buffer (180 mmol/liter KCl, 10 mmol/liter EDTA.2K, 0.5% bovine serum albumin in 10 mmol/liter Tris-HCl buffer, pH = 7.4) containing 0.1% nargarse at 4°C. The minced sample was resuspended in a 10-part volume of isolation buffer and homogenized by a Potter-type homogenizer. The homogenate was centrifuged at 600 gfor 5 min to obtain the supernatant. The supernatant was centrifuged at 10,000 gfor 10 min, and the sediment obtained was washed and suspended in isolation buffer without nargarse. The suspension was recentrifuged at 10,000 gfor 5 min to obtain the final mitochondrial pellet. The pellet was suspended in 150 μl of isolation buffer and used immediately for experiments to determine the respiratory function of mitochondria. Protein concentration in the mitochondria sample was determined using a BCA Protein Assay Kit (PIERCE, Rockford, Illinois).
Determination of mitochondrial respiration
Mitochondrial respiration was measured at 25°C by an oxygen monitor (UC-12 Digital DO/O2/Temp Meter, Central Science, Japan), which polarographically determines oxygen concentration in a water-jacketed sample chamber (volume = 1.2 ml). The assay medium (3 mmol/liter K2HPO4, 1 mmol/liter EDTA.2K, 250 mmol/liter sucrose in 10 mmol/liter Tris-HCl buffer, pH = 7.4) was pre-equilibrated with air at 25°C before being transferred into the sample chamber. A mitochondrial sample (30 or 50 μl) was added to the assay medium in the chamber, and then 10 mmol/liter glutamate was added as a substrate. After state II respiration was recorded for 2 min, state III respiration was initiated by addition of 500 nmol of ADP. The respiratory control index (RCI) and ADP/O ratio were calculated by the method of Chance and Williams (16).
Experiment 3: effect of diazoxide on isolated mitochondria
Rabbit hearts were perfused without ischemia, and mitochondria were isolated from the ventricular myocardium as in Experiment 2. The mitochondria suspension was divided into two parts: one for experiments using an assay medium containing 100 μmol/liter diazoxide and the other for the experiments using a medium containing vehicle (0.1% dimethylsulfoxide) alone. Diazoxide or the vehicle was added to the assay medium immediately before use. Mitochondrial respiratory function was determined as in Experiment 2.
Diazoxide, calphostin C, ADP and bovine albumin were purchased from (SIGMA), St. Louis, Missouri), and 5-HD and r-PIA were obtained from another company (Research Biochemical Institute, Natick, Massachusetts). Nargase was also purchased from another company (Boehringer Ingelheim Bioproducts, Heidelberg, Germany).
Differences in infarct size, hemodynamic parameters and mitochondrial function among the study groups were tested by one-way analysis of variance. When the overall difference was significant in analysis of variance, an intergroup comparison was performed by the Student-Newman-Keul post-hoc test. The unpaired ttest was used to compare mitochondrial function in the diazoxide medium with that in vehicle medium (Experiment 3). The difference was considered significant at p < 0.05. All data are expressed as mean value ± standard error of the mean.
Experiment 1. effects of r-PIA, diazoxide, calphostin C and 5-HD on infarct size in isolated rabbit hearts
Heart rate, left ventricular developed pressure (LVDP) and coronary flow are summarized in Table 1. Under baseline conditions, there were no significant differences in the hemodynamic parameters among all of the study groups. r-PIA reduced the heart rate by 30% and increased coronary flow by approximately 20%, while LVDP was not significantly changed. This coronary vasodilation by r-PIA, an A1agonist, can be explained by the large A2receptor reserve in the coronary artery (17). Diazoxide increased coronary flow by 40% without significant change in heart rate and LVDP. Calphostin C slightly increased the coronary flow, but 5-HD had no effect on the hemodynamic parameters. The effects of diazoxide and r-PIA on coronary flow or heart rate were not abolished by coadministration of either 5-HD or calphostin C. At 2 h after reperfusion, LVDP in the diazoxide-treated hearts was significantly higher than that in the controls; however, all other differences in heart rate, LVDP, and coronary flow between controls and any of the drug-treated groups did not reach statistical significance.
Infarct size data
Infarct size and size of area at risk (i.e., LV mass) are summarized in Table 2. There was no significant difference in the size of area at risk among the study groups. r-PIA reduced infarct size as a percentage of area at risk (%IS/AR) from 49.8 ± 6.5 to 13.4 ± 2.9. Neither 5-HD nor calphostin C alone modified infarct size. However, this infarct size–limiting effect of r-PIA was not observed when calphostin C or 5-HD was added to the pretreatment regimen. Diazoxide achieved infarct-size limitation (%IS/AR = 11.6 ± 4.0), the extent of which was comparable to that by r-PIA. In contrast with the effect of r-PIA, the effect of diazoxide on infarct size was not abolished by calphostin C, although 5-HD blocked diazoxide-induced infarct size–limitation.
Experiment 2: mitochondrial respiratory function after ischemia and reperfusion: diazoxide-treated hearts versus untreated hearts
Table 3summarizes the data of mitochondrial function after ischemia/reperfusion. The ADP/O ratio was not changed after 30-min ischemia and after 10-min reperfusion. However, the rate of oxygen consumption in state III respiration (Qo2) and RCI were reduced after ischemia (from 147.3 ± 5.3 to 108.5 ± 12.3 and from 22.3 ± 1.1 to 12.1 ± 1.8, respectively, both p < 0.05). Both Qo2and RCI remained suppressed at 10 min after reperfusion. However, when the heart was pretreated with 100 μmol/liter diazoxide, the reduction of Qo2and RCI by ischemia was significantly attenuated, and RCI recovered almost to the control level after 10 min of reperfusion.
Experiment 3. effects of diazoxide on respiratory functions of isolated mitochondria
As shown in Table 4, when 100 μmol/liter diazoxide was included in the assay buffer, the ADP/O ratio and Qo2were similar, but RCI was significantly lower compared with RCI in the normal assay buffer (19.1 ± 0.9 vs. 15.3 ± 1.3, p < 0.05). Since RCI is the ratio of Qo2to state IV respiration, this reduction of RCI indicates acceleration of state IV respiration by diazoxide in vitro.
The present study demonstrated that 100 μmol/liter of diazoxide preserved mitochondrial functions after ischemia/reperfusion and reduced infarct size by 75%. This infarct size–limiting effect of diazoxide was comparable to that of 1 μmol/liter of r-PIA, an A1-receptor agonist, and the cardioprotective effects of both agents were abolished by 5-HD. However, calphostin C blocked the infarct-size limitation by r-PIA but not that by diazoxide. These results support the hypothesis that signals triggered by an A1receptor are transmitted via a PKC-mediated pathway to mitoKatpchannels and that the opening of these channels plays a crucial role in preservation of mitochondrial function and cell viability during ischemia.
Doses of diazoxide and 5-HD to examine the function of mitoKatpchannels
A recent study by Garlid et al. (11)showed that mitoKatpchannels have a unique pharmacologic profile. The EC50for diazoxide to activate mitoKatpchannels in reconstituted mitoKatpchannels is estimated to be approximately 0.8 μmol/liter, while >1 mmol/liter of diazoxide is necessary to open sarcolemmal Katpchannels (11). Our results show that 100 μmol/liter diazoxide significantly reduced RCI with insignificant change in Qo2(Table 4), which is consistent with reduction of inner membrane potential by opening of mitoKatpchannels. The dose of diazoxide used in the present study was sufficient to activate smooth muscle type Katpchannels (18), causing a 40% increase in coronary flow (Table 1). However, this vasodilatory effect of diazoxide is unlikely to be essential to the cardioprotection, because the increase in coronary flow during diazoxide infusion was not antagonized by 5-HD, which, however, abolished the infarct size–limiting effect of diazoxide.
The dose of 5-HD used in the present study (30 μmol/liter) is rather low compared with the doses employed for blocking mitoKatpchannels in earlier studies (11,12). The EC50of 5-HD to block mitoKatpchannels, which were opened by diazoxide, in the reconstituted rat heart mitochondria was around 83 μmol/liter (11). Thus, 30 μmol/liter of 5-HD appears rather low, but this dose of 5-HD blocked the infarct size–limiting effects of diazoxide and r-PIA in isolated rabbit hearts (Table 2). Actually, 100 μmol/liter of 5-HD alone appeared to enlarge infarct size in our preliminary experiments. The reason for this apparent contradiction is not clear, but a possible explanation is the difference in temperature during experiments. Although the EC50of 5-HD was obtained by experiments performed at 25°C in the earlier study (11), our infarct-size experiments were conducted at 37°C, at which the EC50of 5-HD could be lower. Most recently, Liu et al. (13)found that 100 μmol/liter of 5-HD selectively blocked the opening of mitoKatpchannels, without inhibiting the response of sarcolemmal Katpchannels, to pinacidil in the rabbit cardiomyocytes. Thus, the present dose of 5-HD appears appropriate to selectively assess the functions of mitoKatpchannels in isolated heart preparations.
Relationship among adenosine receptors, PKC and mitoKatpchannels
A number of earlier studies (19–22)have demonstrated that activation of A1receptors markedly enhances myocardial tolerance against ischemic injury. Although most of the evidence for PKC activation by adenosine receptor stimulation in cardiomyocytes is rather circumstantial, it was demonstrated that A1receptor stimulation activates phospholipase C-β and phospholipase D (20,23,24), both of which produce diacylglycerol, an activator of PKC. In cultured human pediatric myocytes, a relatively high dose of adenosine (i.e., 50 μmol/liter) increased PKC activity by threefold (22). Furthermore, several pharmacologic findings have indicated that PKC has an important role in adenosine-induced cardioprotection. In a previous study, we found that PKC inhibitors (staurosporine and polymyxin B) completely abolished infarct size limitation by r-PIA in the rabbit heart in situ (14). The present study confirmed this previous observation in isolated hearts using a more selective PKC inhibitor, calphostin C (25). The inhibitory effect of PKC inhibitors on anti-infarct tolerance was recently characterized also in isolated cardiomyocyte preparations. Although the optimal dose of adenosine receptor agonists varies depending on the preparations (i.e., human pediatric cardiomyocytes, chick embryo ventricular myocytes and adult rabbit ventricular myocytes), adenosine and A1agonists prevented ischemic myocyte death, and such protection was abolished by chelerythrine and calphostin C (19,22,26). Furthermore, two lines of evidence indicate a link between PKC and Katpchannels in infarct size–limitation. First, PKC-activating phorbol esters facilitate the opening of sarcolemmal Katpchannels (6)and mitoKatpchannels (10). Second, delay of ischemic myocyte death by A1-receptor agonists and a PKC-activating phorbol was prevented by glibenclamide (21,26). Poor selectivity of glibenclamide (11)does not allow differentiation of the roles of sarcolemmal Katpand mitoKatpchannels in these earlier observations. However, a finding that 5-HD at a mitoKatpchannel selective dose completely inhibited cardioprotection by r-PIA (Table 2)and a report that ischemic preconditioning (27)was abolished by 5-HD suggest that the mitoKatpchannel is a major effector in anti-infarct tolerance.
The present study demonstrated that infarct size was limited to 25% of the control value by diazoxide, which is consistent with recent findings by Baines et al. (28)in the in situ rabbit heart. As shown in Table 2, the extent of infarct-size reduction by diazoxide is comparable with that achieved by 1 μmol/liter of r-PIA, and the cardioprotective effects of both agents were similarly abolished by 5-HD. These results further support the role of mitoKatpchannels in infarct size–limitation by A1-receptor activation. Furthermore, considering the failure of calphostin C to block diazoxide-induced cardioprotection, it is likely that opening of mitoKatpchannels occurs downstream of PKC activation in the mechanism of infarct size–limitation by preischemic A1-receptor activation.
In contrast, activation of PKC subsequent to mitoKatpchannel opening is suggested as a mechanism of cardioprotection against calcium paradox in a study by Wang and Ashraf (29). In their study, diazoxide reduced myocardial necrosis by calcium paradox, and this protective effect of diazoxide was abolished by 5-HD and also by chelerythrine, a PKC inhibitor. However, these findings are not necessarily contraditory to the present result of calphostin C, because mechanisms of myocyte destruction are quite different for calcium paradox and ischemia/reperfusion. Separation of the fascia and macula adherens junctions is a crucial initial step for myocyte destruction by calcium paradox (30), while Na+-H+exchange is unlikely to be an important mechanism of calcium overload in this phenomenon (31). In contrast, activation of Na+-H+exchange significantly contributes to myocardial necrosis during ischemia/reperfusion (32), presumably through acceleration of Na+-Ca++exchange, and the change in fascia adherens junctions in ischemic myocytes appears after irreversible changes in the mitochondria (33,34). Thus, although diazoxide is protective against both ischemia/reperfusion and calcium paradox, the sequence of signals and cell protective effectors may be different in these two types of insults.
Protection of mitochondria from ischemic injury by diazoxide
It is not clear how the opening of mitoKatpchannels enhances the tolerance of myocytes to lethal ischemic injury. The function of mitoKatpchannels is thought to regulate mitochondrial volume and thermogenesis (35,36), both of which are apparently unrelated to ischemic injury. The opening of mitoKatpchannels would lower transmembrane potential and thus reduce efficiency of ATP production. Indeed, Portenhauser et al. (37)observed that 400 μmol/liter diazoxide suppressed state III respiration by 22%, and RCI by 44%, in isolated rat mitochondria. In the present experiments, probably because of the lower concentration (i.e., 100 μmol/liter), reduction in RCI by diazoxide was modest, and the change in state III respiration was statistically insignificant. This reduction in RCI per se does not appear to be clearly beneficial for ischemic cardiomyocytes. Furthermore, a recent study by Holmuhamedov et al. (38)suggests that a high dose of Katpchannel openers can even be detrimental to mitochondrial function and structure. They showed that incubation of isolated mitochondrial with 250 μmol/liter pinacidil or 100 μmol/liter cromakalim caused release of cytochrome c and adenylate kinase, two important proteins in the intermembrane space. Thus, the effect of mitoKatpchannel opening on mitochondrial injury during ischemia is probably dependent on the degree of their opening activity. Nevertheless, the present study showed that pretreatment with 100 μmol/liter diazoxide was effective in preserving mitochondrial function during and after 30 min of ischemia (Table 3)and in limiting myocardial necrosis (Table 2).
The effects of diazoxide on mitochondria during ischemia/perfusion (Table 3)are quite compatible with the reported effects of ischemic preconditioning (39–41). Three earlier studies (39–41)consistently showed that ischemic preconditioning preserves mitochondrial function and prevents loss of adenine nucleotide translocase activity during ischemia/reperfusion. However, whether the preservation of mitochondrial function is a part of cardioprotective mechanisms or a result of myocyte protection remains unclear. The contribution of mitochondrial damage to myocardial infarction has been studied for more than a decade, but there is no strong evidence supporting the possibility that a certain degree of reduced mitochondrial function determines the transition from reversible to irreversible myocyte injury (34). Rather, ATP production by mitochondria during ischemia would be theoretically the same, and negligible, regardless of the functional status of mitochondria, because lack of oxygen limits oxidative phosphorylation. However, there is a possibility that mitochondrial function preserved by diazoxide and preconditioning may support energy production in critically injured myocytes at the time of reperfusion, which otherwise fail to survive the subsequent reperfusion period.
The present data indicate that the opening of mitoKatpchannels is downstream of PKC activation in the mechanism of infarct size limitation by adenosine A1-receptor stimulation. How mitoKatpchannel opening enhances anti-infarct tolerance of the cardiomyocytes remains unclear, but preservation of the functional capacity of mitochondria during ischemia/reperfusion may be involved.
☆ This study was supported in part by a grant-in-aid for Scientific Research (No. 08670812) from the Ministry of Education, Science, Sports and Culture of Japan.
This study was presented in part at the 71st Scientific Sessions of the American Heart Association, Dallas, Texas, November 1998.
- infarct size as a percentage of area at risk
- ATP-sensitive potassium channel
- left ventricular developed pressure
- mitochondrial ATP-sensitive potassium channel
- protein kinase C
- the rate of oxygen consumption in state III respiration
- respiratory control index
- Received March 29, 1999.
- Revision received July 23, 1999.
- Accepted September 13, 1999.
- American College of Cardiology
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