Importance of glycolytically derived ATP for Na+ loading via Na+/H+ exchange during metabolic inhibition in guinea pig ventricular myocytes

2001 ◽  
Vol 101 (3) ◽  
pp. 243-251 ◽  
Author(s):  
Hiroshi SATOH ◽  
Shiho SUGIYAMA ◽  
Noriyuki NOMURA ◽  
Hajime TERADA ◽  
Hideharu HAYASHI
Keyword(s):  
Guinea Pig ◽  
Myocardial Ischaemia ◽  
Cell Injury ◽  
Ventricular Myocytes ◽  
Metabolic Inhibition ◽  
Intracellular Acidosis ◽  
Major Pathway ◽  
Intracellular Atp ◽  
Rate Of Increase ◽  
Inhibition Of Glycolysis

The increase in the intracellular Na+ concentration ([Na+]i) during myocardial ischaemia is crucial for ischaemia/reperfusion cell injury, and the cardiac subtype of the Na+/H+ exchanger (NHE-1) has been shown to be a major pathway for Na+ loading. While the importance of glycolytically derived ATP for the optimal functioning of membrane transporters and channels has been suggested, whether NHE-1 is actually activated during myocardial ischaemia remains controversial. Here we examined whether the activity of NHE-1 is predominantly dependent on intracellular ATP generated by glycolysis, and whether the additional inhibition of glycolysis can affect the increase in [Na+]i during the inhibition of oxidative phosphorylation in intact guinea pig ventricular myocytes. The selective inhibition of glycolysis by 2-deoxyglucose prevented the recovery of intracellular pH and the transient increase in [Na+]i following intracellular acidosis induced by a NH4Cl pre-pulse. During severe metabolic inhibition (SMI; induced by amobarbital and carbonyl cyanide m-chlorophenylhydrazone in a glucose-free perfusate), most myocytes changed from rod-shaped to contracted forms by ~ 15 min. [Na+]i increased linearly until rigor contracture occurred, but after rigor contracture the rate of increase was blunted. The increase in [Na+]i during SMI was suppressed significantly by an inhibitor of NHE-1, hexamethylene amiloride. The increase in the intracellular Mg2+ concentration, which can reciprocally indicate depletion of intracellular ATP, was small during the initial 10 min of SMI, but became larger from just a few minutes before rigor contracture. In the presence of 2-deoxyglucose, the time to rigor during SMI was shortened, but the increase in [Na+]i before rigor contracture was not significant, and was much less than that in the absence of 2-deoxyglucose. It is concluded that ATP generated by glycolysis is essential to activate NHE-1, and that the dependence of NHE-1 on glycolysis might affect the increase in [Na+]i observed during myocardial ischaemia.

Survival of metabolically inhibited ventricular myocytes is enhanced by inhibition of rigor and SR Ca2+ cycling

1996 ◽  
Vol 271 (2) ◽  
pp. H643-H650 ◽  
Author(s):  
T. Koyama ◽  
D. Boston ◽  
H. Ikenouchi ◽  
W. H. Barry
Keyword(s):  
Cell Injury ◽  
Ventricular Myocytes ◽  
Metabolic Inhibition ◽  
Atp Depletion ◽  
Resting Membrane Potential ◽  
Adult Rabbit ◽  
Force Development ◽  
Relative Contribution ◽  
Butanedione Monoxime ◽  
Rabbit Ventricular Myocytes

During severe ATP depletion, sarcolemmal rupture resulting from rigor- and/or Ca(2+)-induced myofilament force development is considered to be an important cause of irreversible cell injury. Recent experiments in our laboratory demonstrated that during prolonged metabolic inhibition (MI) in adult rabbit ventricular myocytes, in which rigor was prevented by exposure to 30 mM 2,3-butanedione monoxime (BDM), cyclic uptake and release of cystolic Ca2+ occurred and was associated with strong phasic contractions. To investigate the relative contribution of this sarcoplasmic reticulum Ca2+ cycling and associated force development to energy depletion injury, the effects of BDM together with 7 mM caffeine were examined in isolated rabbit ventricular myocytes subjected to MI with 2 mM NaCN and 20 mM 2-deoxyglucose (2-DG). During 90 min of MI with CN and 2-DG, no cells retained a rod shape in the absence of BDM or caffeine. In the presence of both 30 mM BDM and 7 mM caffeine during MI, preservation of rod morphology was enhanced, and 52 +/- 6.2% of cells retained a rod shape 48 h after metabolic inhibition and had normal ATP content and resting membrane potential. Both systolic and diastolic functions of cells that survived MI, however, were impaired. We conclude that exposure to caffeine together with BDM markedly enhances survival of myocytes during severe prolonged ATP depletion. After recovery, these isolated myocytes show some characteristics of stunning.

Intracellular Ca2+ concentration and pHi during metabolic inhibition

1992 ◽  
Vol 262 (3) ◽  
pp. C628-C634 ◽  
Author(s):  
H. Hayashi ◽  
H. Miyata ◽  
N. Noda ◽  
A. Kobayashi ◽  
M. Hirano ◽  
...  
Keyword(s):  
Cell Shape ◽  
Positive Relationship ◽  
Acidic Solution ◽  
Ventricular Myocytes ◽  
Sodium Cyanide ◽  
Metabolic Inhibition ◽  
Metabolic Energy ◽  
Intracellular Acidosis ◽  
Fura 2 ◽  
Induced Changes

To study the changes in intracellular Ca2+ concentration ([Ca2+]i) and pH (pHi) during metabolic inhibition, rat ventricular myocytes were dual loaded with the acetoxymethyl esters of fura-2 (fura-2/AM) and 2',7'-bis(carboxyethyl)-5,6-carboxyfluorescein (BCECF/AM) and perfused with 2 mM sodium cyanide (NaCN). The percent of rod-shaped cells was 30% of the control after 30 min NaCN in the absence of glucose. [Ca2+]i increased from 82 +/- 8 to 151 +/- 25 (SE) nM (P less than 0.05) when cells were shortened, and to 421 +/- 106 nM (P less than 0.05) when cells were rounded. There was a positive relationship between pCai and pHi (r = 0.425, P less than 0.01). When 50 mM glucose was added during NaCN, there were no significant changes in [Ca2+]i and the percent of rod-shaped cells after 30 min. The pHi of rod-shaped cells, however, decreased to 6.95 +/- 0.03 (P less than 0.01). Although the acidic solution (70% O2-30% CO2) decreased pHi to 6.90 +/- 0.05 (P less than 0.01), there were no changes in the cell shape or [Ca2+]i. The addition of NaCN for 30 min decreased the rod-shaped cells to 18% of the control. Mild acidosis did not cause the changes in cell shape or [Ca2+]i. There was also no protection of the NaCN-induced changes in cell shape by intracellular acidosis. It is likely that the changes in cell shape during metabolic inhibition were related to the depletion of metabolic energy and the increase in [Ca2+]i.

Intramitochondrial [Ca2+] and membrane potential in ventricular myocytes exposed to anoxia-reoxygenation

1998 ◽  
Vol 275 (2) ◽  
pp. H484-H494 ◽  
Author(s):  
T. J. Delcamp ◽  
C. Dales ◽  
L. Ralenkotter ◽  
P. S. Cole ◽  
R. W. Hadley
Keyword(s):  
Membrane Potential ◽  
Guinea Pig ◽  
Mitochondrial Membrane Potential ◽  
Mitochondrial Membrane ◽  
Cell Injury ◽  
Ventricular Myocytes ◽  
Confocal Imaging ◽  
Mitochondrial Depolarization ◽  
Intracellular Ca ◽  
A Cell

The aim of this study was to investigate the role of mitochondrial ionic homeostasis in promoting reoxygenation-induced hypercontracture in cardiac muscle. Mitochondrial membrane potential and intramitochondrial Ca2+ concentration ([Ca2+]) were measured using confocal imaging in guinea pig ventricular myocytes exposed to anoxia and reoxygenation. Anoxia produced a variable, but often profound, mitochondrial depolarization. Some cells mounted a recovery of their mitochondrial membrane potential during reoxygenation; the depolarization was sustained in other cells. Recovery of the mitochondrial membrane potential seemed essential to avoid reoxygenation-induced hypercontracture. Reoxygenation also caused a sizable elevation in intramitochondrial [Ca2+], the amplitude of which was correlated with the likelihood of a cell undergoing hypercontracture. A sustained Ca2+load analogous to that seen during reoxygenation was imposed on cardiac mitochondria through permeabilization of the plasma membrane. Elevation of intracellular [Ca2+] to 800 nM caused a substantial mitochondrial depolarization. We propose that the conditions seen in guinea pig ventricular myocytes during reoxygenation are well suited to produce Ca2+-dependent mitochondrial depolarization, which may play a significant role in promoting irreversible cell injury.

Na+/H+ and Na+/Ca2+ exchange in regulation of [Na+]i and [Ca2+]i during metabolic inhibition

1995 ◽  
Vol 268 (3) ◽  
pp. H1239-H1248 ◽  
Author(s):  
H. Satoh ◽  
H. Hayashi ◽  
H. Katoh ◽  
H. Terada ◽  
A. Kobayashi
Keyword(s):  
Guinea Pig ◽  
Intracellular Ph ◽  
Cell Morphology ◽  
Metabolic Inhibition ◽  
Free Solution ◽  
Intracellular Acidosis ◽  
Energy Depletion ◽  
Fluorescent Indicators ◽  
Low Level ◽  
Percent Change

The relationships among intracellular Na+ and Ca2+ concentrations ([Na+]i and [Ca2+]i, respectively) and cell morphology were investigated during metabolic inhibition (MI) in isolated guinea pig myocytes. [Na+]i and [Ca2+]i were measured using the fluorescent indicators, Na(+)-binding benzofuran isophthalate and fluo 3. During the initial 20 min of MI, [Na+]i increased from 6.2 +/- 0.5 to 18.6 +/- 1.6 mM (n = 31), whereas [Ca2+]i, expressed as the percent change of fluo 3 fluorescence, remained at the low level. In the following 30 min, 94% of the cells developed contracture, and [Ca2+]i began to increase after cells had contracted (167 +/- 14% at 50 min). The level of [Ca2+]i during MI was lower than that during 500 microM strophanthidin perfusion. The increase in [Na+]i was not affected by 10 microM tetrodotoxin but was suppressed by 1 microM hexamethylene amiloride (HMA). The application of 10 mM glucose from the start of MI prevented both the increase in [Na+]i and cell contracture. However, the addition of glucose after 20 min of MI [energy repletion (ER)] led to a dramatic increase in [Ca2+]i (442 +/- 72% at 50 min, n = 31), and 84% of the cells developed contracture. The increase in [Ca2+]i and the cell contracture were suppressed by HMA or Ca(2+)-free solution. Intracellular pH decreased from 7.23 +/- 0.07 to 6.95 +/- 0.09 during MI but did not change after ER (6.90 +/- 0.11 at 35 min, n = 9). These findings suggested that during MI 1) [Na+]i increased by both the activated Na+ influx via Na+/H+ exchange and the suppressed Na+ extrusion via the Na+/K+ pump, 2) Na+/Ca2+ exchange was inhibited by energy depletion and intracellular acidosis, and 3) cell contracture was not related to Ca2+ overload but was related to rigor due to energy depletion.

Effects of anoxia on intracellular Ca2+ and contraction in isolated guinea pig cardiac myocytes

1995 ◽  
Vol 268 (3) ◽  
pp. H1045-H1052 ◽  
Author(s):  
S. Seki ◽  
K. T. MacLeod
Keyword(s):  
Guinea Pig ◽  
Cardiac Myocytes ◽  
Ventricular Myocytes ◽  
Sodium Dithionite ◽  
Metabolic Inhibition ◽  
Cell Length ◽  
Tyrode Solution ◽  
Time To Peak ◽  
Simultaneous Measurements

Single, enzymatically isolated guinea pig ventricular myocytes were exposed to 3-min periods of anoxia with glucose-free Tyrode solution containing 1 mM sodium dithionite (Na2S2O4) and were then reoxygenated for 10 min. The myocytes were exposed to rapid applications of 10 mM caffeine during the control, anoxic, and reoxygenation periods. Intracellular Ca2+ concentration ([Ca2+]i) was measured ratiometrically using indo 1 with simultaneous measurements of cell length. The effects of anoxia on Ca2+ were compared with those of hypoxia and metabolic inhibition. The amplitude of the electrically stimulated (Ca transient) and caffeine-evoked Ca2+ (Caff-Ca) transients decreased during anoxia and recovered after reoxygenation. Diastolic [Ca2+]i did not change during 3 min of anoxia but rose progressively after prolonged anoxia and remained at this higher level on reoxygenation. During metabolic inhibition the Ca transients decreased, while the Caff-Ca transients showed no change in amplitude. During hypoxia the Ca transients decreased. Anoxia slowed the time to peak of the Ca transient, the time to 50% relaxation, and the time to 90% relaxation. The decline of indo 1 fluorescence on rapid caffeine application was slowed during anoxia, metabolic inhibition, and hypoxia and partially recovered after reoxygenation.

Intracellular acidosis decreases the outward Na+-Ca2+exchange current in guinea pig ventricular myocytes

1995 ◽  
Vol 36 (2) ◽  
pp. 146 ◽  
Author(s):  
Ek Ho Lee ◽  
So Ra Park ◽  
Kwang Se Paik ◽  
Chang Kook Suh
Keyword(s):  
Guinea Pig ◽  
Ventricular Myocytes ◽  
Exchange Current ◽  
Intracellular Acidosis

scholarly journals Mitochondrial BKCa channels contribute to protection of cardiomyocytes isolated from chronically hypoxic rats

2011 ◽  
Vol 300 (2) ◽  
pp. H507-H513 ◽  
Author(s):  
Gudrun H. Borchert ◽  
Chengtao Yang ◽  
František Kolář
Keyword(s):  
Protective Effect ◽  
Chronic Hypoxia ◽  
Cell Injury ◽  
Ventricular Myocytes ◽  
Metabolic Inhibition ◽  
Male Rats ◽  
Bkca Channels ◽  
Trypan Blue Exclusion ◽  
Salutary Effect ◽  
Ldh Release

Chronic hypoxia protects the heart against injury caused by acute oxygen deprivation, but its salutary mechanism is poorly understood. The aim was to find out whether cardiomyocytes isolated from chronically hypoxic hearts retain the improved resistance to injury and whether the mitochondrial large-conductance Ca2+-activated K+ (BKCa) channels contribute to the protective effect. Adult male rats were adapted to continuous normobaric hypoxia (inspired O2 fraction 0.10) for 3 wk or kept at room air (normoxic controls). Myocytes, isolated separately from the left ventricle (LVM), septum (SEPM), and right ventricle, were exposed to 25-min metabolic inhibition with sodium cyanide, followed by 30-min reenergization (MI/R). Some LVM were treated with either 30 μM NS-1619 (BKCa opener), or 2 μM paxilline (BKCa blocker), starting 25 min before metabolic inhibition. Cell injury was detected by Trypan blue exclusion and lactate dehydrogenase (LDH) release. Chronic hypoxia doubled the number of rod-shaped LVM and SEPM surviving the MI/R insult and reduced LDH release. While NS-1619 protected cells from normoxic rats, it had no additive salutary effect in the hypoxic group. Paxilline attenuated the improved resistance of cells from hypoxic animals without affecting normoxic controls; it also abolished the protective effect of NS-1619 on LDH release in the normoxic group. While chronic hypoxia did not affect protein abundance of the BKCa channel regulatory β1-subunit, it markedly decreased its glycosylation level. It is concluded that ventricular myocytes isolated from chronically hypoxic rats retain the improved resistance against injury caused by MI/R. Activation of the mitochondrial BKCa channel likely contributes to this protective effect.

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