© 1999 American Heart Association, Inc.
Original Contribution |
Propagation of Cardiomyocyte Hypercontracture by Passage of Na+ Through Gap Junctions
From the Department of Cardiology (M.R-M., D.G-D., J.S-S.), Hospital General Universitario Vall d'Hebron, Barcelona, Spain, and Justus-Liebig-Universität (B.H., H.M.P.), Physiologisches Institut, Giessen, Germany.
Correspondence to David Garcia-Dorado, MD, PhD, Department of Cardiology, Hospital General Universitario Vall d'Hebron, Pg. Vall d'Hebron 119-129, Barcelona 08035, Spain.
 |
Abstract |
|---|
 Top Abstract IntroductionMaterials and MethodsResultsDiscussionReferences |
|---|
Abstract—Prolonged ischemia increases cytosolic Ca2+ concentration in cardiomyocytes. Cells with severely elevated cytosolic Ca2+ may respond to reperfusion, developing hypercontracture, sarcolemmal disruption, and death. Cardiomyocytes are efficiently connected through gap junctions (GJs) to form a functional syncytium, and it has been shown that hypercontracture can be propagated to adjacent myocytes through a GJ-mediated mechanism. This study investigated the mechanism of propagation of cell injury associated with sarcolemmal rupture in end-to-end connected pairs of isolated rat cardiomyocytes. Microinjection of extracellular medium into one of the cells to simulate sarcolemmal disruption induced a marked increase in cytosolic Ca2+ (fura-2) and Na+ (SBFI) in the adjacent cell and its hypercontracture in <30 seconds (22 of 22 cell pairs). This process was not modified when Ca2+ release from the sarcoplasmic reticulum was blocked with 10 µmol/L ryanodine (5 of 5 cell pairs), but it was fully dependent on the presence of Ca2+ in the extracellular buffer. Blockade of L-type Ca2+ channels with 10 µmol/L nifedipine did not alter propagation of hypercontracture. However, the presence of 15 to 20 µmol/L KB-R7943, a highly selective blocker of reverse Na+/Ca2+ exchange, prevented propagation of hypercontracture in 16 of 20 cell pairs (P<0.01) without interfering with GJ permeability, as assessed by the Lucifer Yellow transfer method. Addition of the Ca2+ chelator EGTA (2 mmol/L) to the injection solution prevented hypercontracture in the injected cell but not in the adjacent one (n=5). These results indicate that passage of Na+ through GJ from hypercontracting myocytes with ruptured sarcolemma to adjacent cells, and secondary entry of [Ca2+]o via reverse Na+/Ca2+ exchange, can contribute to cell-to-cell propagation of hypercontracture. This previously unrecognized mechanism could increase myocardial necrosis during ischemia-reperfusion in vivo and be the target of new treatments aimed to limit it.
Key Words: hypercontracture • gap junction • propagation • ischemia • reperfusion
|
Introduction |
|---|
|
TopAbstractIntroductionMaterials and MethodsResultsDiscussionReferences |
|---|
Gap junctions (GJs) play an essential role in the normal function of the cardiovascular system; they mediate the spread of electrical impulses that stimulate synchronized contraction of the cardiac muscle and coordinate the function of endothelial and smooth muscle cells in arterial walls.1 2 GJ-mediated communication is also essential for normal embryonic development as well as for the function of many other adult organs, including the lungs, retina, lens, liver, and central nervous system.3 4 5 Recent studies have demonstrated that neuron GJs remain open during brain ischemia, therefore linking dying cells with potentially viable ones, and that cerebral infarcts may expand over time by a GJ-mediated mechanism.6 The extent of ischemic injury can be significantly decreased when GJ permeability is reduced with octanol.7
However, the role of GJs in chemical communication of cardiac muscle cells, and in particular their possible involvement in cell death secondary to ischemia-reperfusion, is poorly understood. After an ischemic episode, cardiomyocytes may respond to reenergization by developing an abrupt and extreme distortion of their architecture, resulting from development of excessive contractile force.8 This phenomenon, named hypercontracture, causes extreme cell shortening and may occur during the first minutes of reflow as the consequence of ATP availability in the presence of abnormally high [Ca2+]i concentration and cytoskeletal fragility caused by ischemia.9 10
Reperfused myocardial infarcts consist almost exclusively of areas of contraction band necrosis formed by hypercontracted dead myocytes.11 A striking feature of these areas of necrosis is that they usually contain hypercontracted cardiomyocytes in compact clusters, with highly indented and irregular borders or even a patchy appearance, whereas hypercontracted cardiomyocytes in scattered distribution are virtually absent.12 Gradients in wall tension, collateral flow and concentration of catabolites, and diffusion of O2 from endocardial and epicardial surfaces or nonischemic myocardium may influence the regional distribution of the areas of necrosis but cannot explain the continuity of these areas and the absence of scattered hypercontracted myocytes. Studies using computer simulation of infarct morphology demonstrate that some kind of cell-to-cell interaction must be taken into account to reproduce these features of infarct geometry.13 In a recent study, we found that the GJ blocker heptanol, applied only at the time of reenergization, was able to reduce the final extent of myocardial necrosis and to alter infarct geometry in intact rat and pig hearts. At the cellular level, we observed that hypercontracture of a myocyte was consistently propagated to the adjacent cell and that chemical uncoupling with heptanol prevented this spreading of cell injury. Moreover, these effects of heptanol were observed at concentrations that did not afford any protection against reoxygenation-induced hypercontracture in single cells.14 These studies strongly support the hypothesis that the fate of a cardiomyocyte after an ischemic period is not only determined by its own biochemical derangements but may also be influenced by the survival or death of adjacent cells.
There is ample evidence that, in contrast to what happens in isolated cardiomyocytes, in myocardial tissue hypercontracture is associated with sarcolemmal rupture and enzyme release.15 Sarcolemmal disruption causes dramatic changes in cytosolic composition that can be propagated to adjacent cells through GJs and result in cell-to-cell propagation of hypercontracture. The present study was aimed at identifying the chemical messenger responsible for this cell-to-cell propagation of hypercontracture through GJs. Hypercontracture of freshly isolated adult rat cardiomyocytes was induced by microinjection of extracellular media containing 1 mmol/L Ca2+ to simulate sarcolemmal disruption, and its propagation between end-to-end connected cell pairs was analyzed under different interventions.
|
Materials and Methods |
|---|
|
TopAbstractIntroductionMaterials and MethodsResultsDiscussionReferences |
|---|
Cardiomyocyte Isolation Procedure
Animals were handled according to the Declaration of Helsinki, and the experimental procedures were approved by the Research Commission of the Hospital Vall d'Hebron. Ventricular heart muscle cells were isolated from adult male Sprague-Dawley rats (300 g) as previously described.16 Briefly, whole hearts were retrogradely perfused in a Langendorff system with a modified Krebs buffer (in mmol/L, NaCl 110, KCl 2.6, KH2PO4 1.2, MgSO4 1.2, NaHCO3 25, and glucose 11) containing 45 µmol/L CaCl2 and 0.03% collagenase (type II, Serva). Cells from dissociated tissue were subjected to a progressive normalization of Ca2+ levels to a final concentration of 1 mmol/L. Rod-shaped cells were selected by gravity sedimentation in a 4% BSA gradient (fraction V, Boehringer Mannheim) and plated in glass-bottomed dishes with medium 199/HEPES (Sigma) and 4% FCS (Gibco). Two hours after plating, broken nonattached cells were discarded by changing the medium. Five to ten end-to-end–connected myocytes were found in each culture dish, and propagation of hypercontracture between adjacent cells, GJ permeability, and changes in intracellular Na+ and Ca2+ concentrations were studied in these cell pairs.
Microinjection and Experimental Conditions
One of the cells of each pair was microinjected with
extracellular medium to simulate sarcolemmal disruption occurring
during reoxygenation-induced hypercontracture. A
hydraulic micromanipulator with digital control (digital
micromanipulator model 5171 and transjector model 5246, Eppendorf) was
used. Microinjection was performed through a 0.2- to 0.5-µm–tip
sterile micropipette (Sterile Femtotips, Eppendorf) by a pulse pressure
of 400 hPa of 0.1-second duration. The axial and depth limits were set
to allow penetration of the micropipette 3 to 5 µm below cell
surface at an angle of 45°. Previous studies using the same
micropipettes and similar solutions showed that these microinjection
parameters allow effective injection without detectable
mechanical injury.17 In control experiments, the
extracellular medium was a buffer containing (in mmol/L) NaCl 140,
KCl 3.6, MgSO4 1.2, CaCl2
1, and HEPES 20. The modifications of the extracellular buffer included
the addition of (1) 10 µmol/L ryanodine, (2) 10 µmol/L
nifedipine, or (3) 15 µmol/L and 20 µmol/L
KB-R7943, or the replacement of Ca2+ by 2
mmol/L EGTA. The microinjected solution was modified in some
experiments by replacing Ca2+ by 2 mmol/L
EGTA. To assess GJ permeability, 2% Lucifer Yellow was added to the
microinjection solution in a series of experiments. All experiments
were performed at 37°C (Digital Warm Stage Controller, Linkam)
on the stage of an inverted microscope (Olympus IMT-2). Cell images
were continuously videorecorded at x400 magnifications. Relative
changes in cell length were measured on the videorecorded images.
Measurements were performed in both cells every 2 seconds during the
first 30 seconds after microinjection, except when indicated otherwise.
Transfer dye through GJs was monitored under fluorescent
microscopy using a 420-nm excitation light with a bandwidth of 15
nm.
Measurement of [Ca2+]i and
Na+ Concentrations
Ratiofluorescence studies were performed in a
separate series of experiments under the previously described
conditions. Changes in
[Ca2+]i and
Na+ concentrations were analyzed in
myocytes loaded with fura-2 or SBFI (Molecular Probes), respectively.
For loading, cells were incubated for 30 minutes at 37°C in medium
199 with the acetoxymethyl ester of fura-2 (5 µmol/L) or SBFI
(10 µmol/L). Myocytes were then washed twice and postincubated
for 20 minutes in medium 199 to allow hydrolysis of the acetoxymethyl
esters within the cells. Petri dishes with fura-2 or SBFI-loaded
myocytes were positioned on the temperature-controlled stage of an
inverted fluorescent microscope (Olympus IX70) with a fluorite
objective (UplaFL x200 or x400, Olympus). Ratiofluorescence
measurements were performed by a commercially available imaging system
(QuantiCell 900, Applied Imaging). Cells were alternatively excited at
340 and 380 nm, with a bandwidth of 15 nm, by means of a fast-speed
monochromator. Exposure time was set for each excitation wavelength
according to fluorescence intensity and was typically 40 to 80
ms. Emitted light was collected by an air-cooled intensified digital
camera with a resolution of 640x640 pixels. Counts from clusters of
2x2 or 4x4 pixels were pooled to use shorter exposure times and to
improve signal/noise ratio. Ratios of 340/380 were calculated for each
pixel cluster from signal intensities in that cluster in pairs of
images consecutively obtained at the 2 wavelengths, and color-coded
340/380 ratio images were generated. The average ratio was calculated
for regions of interest defined in these images, and the changes in
these average ratio values through time were analyzed.
Field Stimulation of Cardiomyocytes
To assess the potential effects of KB-R7943 on cytosolic
Ca2+ transients and
contractility, changes in cell length and
ratiofluorescence were analyzed in
cardiomyocytes submitted to field stimulation at 1 Hz, as
previously described.18 Biphasic electrical pacing
composed of 2 equal but opposed rectangular 40-V stimuli of 0.5-ms
duration were applied between 2 silver electrodes immersed in the
fluid. Measurements were performed before addition of any drug to the
extracellular fluid and after addition of KB-R7943 at 15
µmol/L.
|
Results |
|---|
|
TopAbstractIntroductionMaterials and MethodsResultsDiscussionReferences |
|---|
Role of [Ca2+]o in the Propagation of Hypercontracture
In control conditions (extracellular and microinjected solutions composed of HEPES buffer with ionic composition mimicking physiological extracellular media), microinjection of extracellular buffer consistently induced hypercontracture of the injected myocyte, and the rapid rise in cytosolic Ca2+ concentration in the microinjected cell was followed within a few seconds by a similar rise in the adjacent cell and by its hypercontracture (Figure 1
).
However, when Ca2+ was replaced in the
extracellular medium by the Ca2+ chelator EGTA,
but not in the injected buffer, the rise in Ca2+
concentration and hypercontracture observed in the injected myocyte
were not propagated to the adjacent cell in 16 of 18 cell pairs (Figure 2). Failure of propagation of
hypercontracture in the absence of
[Ca2+]o was not due to
reduced GJ permeability, because transfer of the GJ-permeant dye
Lucifer Yellow from the microinjected to the adjacent cell was not
modified in the absence of
[Ca2+]o (Figure 2).
|
) and the adjacent one (•). In <30 seconds, both cells
were completely hypercontracted. C, Average changes in fura-2 ratio in
4 experiments, demonstrating the rapid increase in
[Ca2+]i induced by microinjection in both
cells. Change in ratio (
) at a given time is defined as the value
measured at that time divided by the initial value measured before
microinjection. Data are mean±SEM.
|
On the basis of the observed role of
[Ca2+]o in propagation of
hypercontracture, we investigated L-type Ca2+
channels as a likely route of Ca2+ entry into the
adjacent cell. However, this possibility was ruled out by repeating the
microinjection in the presence of 10 µmol/L of the channel
blocker nifedipine. Blockade of L-type
Ca2+ channels did not modify either
Ca2+ rise in the adjacent cell or propagation of
hypercontracture (Figure 2
).
Ca2+ release from the sarcoplasmic reticulum (SR)
had no effect on cell propagation of hypercontracture. Addition of
ryanodine to the extracellular medium (10 µmol/L for 15 minutes)
did not prevent Ca2+ rise or hypercontracture in
the adjacent cell (Figure 2).
Passage of Na+ and Role of Reverse
Na+/Ca2+ Exchange
We used ratiofluorescence imaging of cells loaded with the
Na+ indicator SBFI to demonstrate a rapid and
almost simultaneous increase in cytosolic
Na+ concentration in the microinjected and the
adjacent cell (Figure 3). To determine
the role of reverse
Na+/Ca2+ exchange in the
increase of [Ca2+]i
concentration and hypercontracture of the adjacent cell, microinjection
of extracellular medium was performed in the presence of the novel
isothiourea derivative KB-R7943, a highly selective blocker of
Na+/Ca2+ exchange in its
reverse mode.19 20 Addition of 20 µmol/L of
KB-R7943 to the extracellular medium prevented propagation of
hypercontracture in 12 of 15 cell pairs and markedly delayed it in the
remaining 3 (Figure 3). A similar effect was observed at 15
µmol/L (propagation was observed only in 1 of 5 cell pairs). However,
at 10 µmol/L of KB-R7943, propagation was observed in 12 of 13
cell pairs, although it was significantly delayed (after 30 seconds of
microinjection, hypercontracture was propagated in only 6 of 13 cell
pairs). KB-R7943 at 15 µmol/L had no influence on either cell
shortening (10.3±0.4% and 10.0±0.5%, respectively, in the presence
and the absence of the drug) or Ca2+ transients
induced by field stimulation (Figure 4).
Despite the absence of propagation of hypercontracture, KB-R7943 at
20 µmol/L did not alter GJ permeability, as assessed by
cell-to-cell diffusion of Lucifer Yellow (Figure 3).
|
|
The role of passage of Na+ through GJ and
subsequent exchange with
[Ca2+]o in cell-to-cell
propagation of hypercontracture was further tested in a series of
experiments in which Ca2+ was replaced by 2
mmol/L EGTA in the microinjection buffer. This consistently
resulted in hypercontracture of the adjacent cell despite the absence
of any change in length in the microinjected cell (Figure 3) and
clearly ruled out the mechanical distortion of GJs as the main cause of
propagation of hypercontracture. The fact that the hypercontracture in
the adjacent cell is delayed in these experiments may indicate that
passage of Ca2+ from the primarily injured cell
can contribute to hypercontracture of the adjacent cell.
|
Discussion |
|---|
|
TopAbstractIntroductionMaterials and MethodsResultsDiscussionReferences |
|---|
This study investigated the chemical messenger responsible for cell-to-cell propagation of hypercontracture in pairs of isolated rat cardiomyocytes, in which sarcolemmal rupture of one of the cells was mimicked by injection of extracellular medium. First, we investigated whether the propagation of hypercontracture is due to direct passage of Ca2+ through GJs or to passage of another messenger that induces either Ca2+ entry from the extracellular space or Ca2+ release from intracellular stores. Our results demonstrate that propagation of microinjection-induced hypercontracture to adjacent cells is fully dependent on [Ca2+]o and is caused by the passage of Na+ through GJs that, in turn, is able to induce Ca2+ entry by the reverse mode of Na+/Ca2+ exchanger. L-type Ca2+ channels do not represent an additive route of Ca2+ entry from the extracellular medium to the adjacent cell, given that blockade of the channels with 10 µmol/L of nifedipine did not modify Ca2+ rise in the adjacent cell nor propagation of hypercontracture (Figure 2
). Similar results were obtained with
the inorganic Ca2+ channel blocker
Ni+ (data not shown). The potential role of
Ca2+ release from the SR in the rise in
[Ca2+]i concentration and
hypercontracture of the adjacent cell was ruled out in experiments in
which hypercontracture was induced in control extracellular buffer
containing 10 µmol/L ryanodine. This blocker of
Ca2+ release from the SR had no effect on
Ca2+ rise or hypercontracture (Figure 2).
These results also demonstrate that rapid and important increases in
cytosolic Ca2+ concentration associated with
sarcolemmal rupture do not result in GJ closure rapid enough to prevent
propagation of cell injury. Transmission of cAMP, IP3, or Ca2+ has been involved in cell-to-cell signaling in many cell types, including cardiomyocytes.21 22 23 24 25 It could be expected that direct passage of Ca2+ through GJs without the need of Ca2+ entry from the extracellular space could suffice for cell-to-cell propagation of hypercontracture. Our results demonstrate that this is not the case. A possible explanation is that GJ permeability is closely regulated by [Ca2+]i concentration.26 Previous studies have shown that GJs have a low ability to transmit Ca2+,27 28 and GJ closure caused by exposure to extracellular levels of Ca2+ could prevent passage of sufficient Ca2+ ions as to induce hypercontracture in the adjacent cell. Transfer of Na+ ions, because of the large gradient in cytosolic Na+ concentration between the microinjected and the adjacent cell, may allow Ca2+ influx into the second cell through the reverse Na+/Ca2+ exchange after GJ closure. GJs have been shown to allow efficient equalization of cytosolic Na+ concentration in other cell types.29
The observation that passage of Na+ through GJs can propagate hypercontracture to adjacent cardiomyocytes may have important implications. There is strong evidence that during myocardial reperfusion, hypercontracture is often associated with sarcolemmal disruption8 because of the effect of excessive mechanical stress on a membrane with reduced mechanical resistance.16 Membrane disruption allows influx of extracellular medium with high Na+ concentration into the hypercontracting cell, a situation mimicked by microinjection of extracellular medium. Although GJs may be closed during ischemia, they are expected to reopen rapidly in response to oxidative metabolism. The proposed mechanism of cell-to-cell propagation of hypercontracture may be thus operative during myocardial reperfusion in vivo. The fact that cell-to-cell propagation of hypercontracture depends on sarcolemmal rupture could impose a limit and explain why it does not progress to the whole heart. Cells exposed to less severe ischemia, as those receiving collateral flow, could withstand hypercontracture without developing sarcolemmal rupture and Na+ overload. If the amount of Na+ is not enough to promote exchange in the reverse mode, propagation would be interrupted.
The sarcolemmal Na+/Ca2+ exchanger is a major regulator of [Ca2+]i homeostasis in excitable cells.30 In most mammalian cardiac muscle cells, the exchanger works mainly in the forward mode, ie, as rapid extruder for Ca2+ that has entered the cardiomyocyte via the sarcolemmal L-type Ca2+ channels to trigger the release of Ca2+ from the SR. Available evidence suggests that in heart, the exchanger competes for Ca2+ with the SR Ca-ATPase to bring about relaxation to a degree dependent on species, developmental stage, and/or physiological state of the heart. However, under pathological conditions such as ischemia-reperfusion injury, the direction of its electrogenic exchange can be changed, and the exchanger is thought to cause Ca2+ overload because of an ischemia-dependent increase in Na+.31 Ca2+ influx through reverse Na+/Ca2+ exchange may suffice to induce contraction when intentionally driven into that mode of action.32 In our study, addition of the reverse Na+/Ca2+ exchange inhibitor KB-R7943 prevented propagation of hypercontracture in most cases at concentrations of 15 µmol/L or greater. KB-R7943 has been shown to act as a potent blocker of reverse Na+/Ca2+ exchange in rat cardiomyocytes (90% inhibition at 10 µmol/L) without significant effects on other ion transporters or channels at concentrations <30 µmol/L. At 30 µmol/L, KB-R7943 inhibited the dihydropyridine-sensitive Ca2+ uptake by 35% and forward Na+/Ca2+ exchange by 38% without significant influence on Na+/H+ exchange, sarcolemmal or SR Ca2+ ATPase, or Na+/K+ ATPase activities.19 20 In our study, the concentration of KB-R7943 that effectively prevented propagation of hypercontracture had no significant effect on Ca2+ transients or systolic cell shortening of isolated myocytes submitted to field stimulation. KB-R7943 also slowed hypercontracture of the microinjected cell, which suggests that the reverse mode of Na+/Ca2+ exchange represents one of the routes of Ca2+ entry in myocytes primarily injured with microinjection.
The role of passage of Na+ through GJs and
subsequent exchange with
[Ca2+]o in cell-to-cell
propagation of hypercontracture was further confirmed when
Ca2+ was replaced by EGTA in the microinjection
buffer. This consistently resulted in hypercontracture of the
adjacent cell, at a slower rate than in control conditions, despite the
absence of any change in length in the microinjected cell (Figure 3). This result constitutes direct evidence that propagation of
hypercontracture may occur independently of any physical distortion of
GJs secondary to mechanical forces imposed by hypercontracture on
intercalated discs. However, a possible role of mechanical interaction
in sarcolemmal disruption occurring during hypercontracture cannot be
excluded, as was previously suggested.33 34 In previous
studies injection of EGTA in one cell has not been found to reduce
cytosolic Ca2+ concentration in adjacent,
GJ-connected cells, which indicates that EGTA does not significantly
diffuse through GJs.27 The slower rate of hypercontracture
in the second cell in these experiments could suggest that direct
passage of Ca2+ through GJs, although not
sufficient to induce hypercontracture of the second cell, may
contribute to its development.
The results presented here encourage the study of other potential deleterious effects of GJ-mediated chemical communication in other multicellular systems. Harmful chemical exchange in tissues with well-developed cell-to-cell communication, such as liver or central nervous system, could represent a more generalized mechanism associated with ischemia or hypoxia. In myocardial tissue, cell-to-cell propagation of hypercontracture due to the passage of Na+ through GJs could represent a previously unrecognized mechanism of cell death secondary to ischemia-reperfusion. This mechanism could allow propagation of hypercontracture of severely injured cells to other cells with milder ischemic injury that could have otherwise survived. Considering that myocytes are connected to multiple cells through GJs, even a small probability of cell-to-cell progression of hypercontracture could have a major impact in infarct size. Under experimental conditions, drugs decreasing GJ permeability, such as heptanol or octanol, have successfully reduced transmission of potentially harmful metabolic signals in different cell types.7 14 35 Their clinical application is, however, limited by their effects on myocardial contractile function and their arrhythmogenic properties. This study describes a mechanism of cell injury propagation through GJs that could be the target of a very selective intervention without major effects on GJ permeability or ion homeostasis in normal cells.
|
Acknowledgments |
|---|
This work was supported by the PL-951254 BIOMED-2 program of the European Union and by Grant 97/0948 from the Fondo de Investigaciones Sanitarias (Spain). We thank Y. Puigfel for her technical assistance and A. Garcia-Dorado, J. Inserte, L. Agulló, J. Sagristà, and J. Cinca for their helpful discussions and suggestions.
Received December 7, 1998; accepted May 28, 1999.
|
References |
|---|
|
TopAbstractIntroductionMaterials and MethodsResultsDiscussionReferences |
|---|
1. Dhein S. Gap junction channels in the cardiovascular system: pharmacological and physiological modulation. Trends Pharmacol Sci. 1998;19:229–241.[Medline] [Order article via Infotrieve]
2.
Peters NS, Wit AL. Myocardial architecture and
ventricular arrhythmogenesis. Circulation. 1998;97:1746–1754.
3. Loewenstein WR. Junctional intercellular communication: the cell-to-cell membrane channel. Physiol Rev. 981;61:829–913.
4. Kumar NM, Gilula NB. The gap junction communication channel. Cell. 1996;84:381–388.[Medline] [Order article via Infotrieve]
5. Severs NJ. Cardiac muscle cell interaction: from microanatomy to the molecular make-up of the gap junction. J Histopathol. 1995;10:481–501.
6.
Cotrina ML Astrocytic gap junctions remain open during
ischemic conditions. J Neurosci. 1998;18:2520–2537.
7. Rawanduzy A, Hansen A, Hansen TW, Nedergaard M. Effective reduction of infarct volume by gap junction blockade in a rodent model of stroke. J Neurosurg. 1997;87:916–920.[Medline] [Order article via Infotrieve]
8. Ganote CE. Contraction band necrosis and irreversible myocardial injury. J Mol Cell Cardiol. 1983;15:67–73.[Medline] [Order article via Infotrieve]
9.
Ladilov YV, Siegmund B, Piper HM. Simulated
ischemia increases the susceptibility of rat
cardiomyocytes to hypercontracture. Circ Res. 1997;80:69–75.
10.
Stern MD, Chien AM, Capogrossi MC, Pelto DJ, Lakatta
EG. Direct observation of the "oxygen paradox" in single rat
ventricular myocytes. Circ Res. 1985;56:899–903.
11. Solares J, Garcia-Dorado D, Oliveras J, González MA, Ruiz-Meana M, Barrabés JA, González-Bravo C, Soler-Soler J. Contraction band necrosis at the lateral borders of the area at risk in reperfused infarcts. Virchows Arch. 1995;426:393–399.[Medline] [Order article via Infotrieve]
12.
Garcia-Dorado D, Théroux P, Duran JM, Solares J,
Alonso J, Sanz E, Muñoz R, Elízaga J, Botas J,
Fernández-Avilés F, Soriano J, Esteban E. Selective
inhibition of contractile apparatus: a new approach to
modification of infarct size, infarct composition and infarct geometry
during coronary artery occlusion and reperfusion.
Circulation. 1992;85:1160–1175.
13.
Garcia-Dorado D, Théroux P, Desco M, Solares J,
Elízaga J, Fernández-Avilés F, Alonso J, Soriano
J. Cell-to-cell interaction: a mechanism to explain wave front
progression of myocardial necrosis. Am J Physiol. 1989;256:H1266–H1273.
14.
Garcia-Dorado D, Inserte J, Ruiz-Meana M,
González MA, Solares J, Juliá M, Barrabés JA,
Soler-Soler J. Gap junction uncoupler heptanol prevents cell-to-cell
progression of hypercontracture and limits necrosis during myocardial
reperfusion. Circulation. 1997;96:3579–3586.
15. Hearse DJ, Humphrey SM, Chain EB. Abrupt reoxygenation of the anoxic potassium-arrested perfused rat heart: a study of myocardial enzyme release. J Mol Cell Cardiol. 1973;5:395–407.[Medline] [Order article via Infotrieve]
16. Ruiz-Meana M, Garcia-Dorado D, González MA, Barrabés JA, Soler-Soler J. Effect of osmotic stress on sarcolemmal integrity of isolated cardiomyocytes following transient metabolic inhibition. Cardiovasc Res. 1995;30:64–69.[Medline] [Order article via Infotrieve]
17.
Shubeita HE, Thorburn J, Chien K. Microinjection of
antibodies and expression vectors into living myocardial cells.
Circulation. 1992;85:2236–2246.
18.
Schlüter KD, Weber M, Schraven E, Piper HM. NO
donor SIN-1 protects against reoxygenation-induced
cardiomyocyte injury by a dual action. Am J
Physiol. 1994;267:H1461–H1466.
19.
Iwamoto T, Watano T, Shigekawa M. A novel isothiourea
derivative selectively inhibits the reverse mode of
Na+/Ca2+ exchange in cells
expressing NCX1. J Biol Chem. 1996;271:22391–22397.
20. Watano T, Kimura J, Morita T, Nakanishi H. A novel antagonist, No. 7943, of the Na+/Ca2+ exchange current in guinea-pig cardiac ventricular cells. Br J Pharmacol. 1996;119:555–563.[Medline] [Order article via Infotrieve]
21. Churchill CG, Atkinson MM, Louis F. Mechanical stimulation initiates cell-to-cell calcium signaling in ovine lens epithelial cells. J Cell Sci. 1996;109:355–365.[Abstract]
22. Venance L, Piomelli D, Glowinski J, Giaume C. Inhibition by anandamide of gap junctions and intercellular calcium signaling in striatal astrocytes. Nature. 1995;376:590–594.[Medline] [Order article via Infotrieve]
23.
Christ GJ, Moreno AP, Melman A, Spray DC. Gap
junction-mediated intercellular diffusion of Ca2+
in cultured human corporal smooth muscle cells. Am J
Physiol. 1992;263:C373–C383.
24.
Boitano S, Dirksen ER, Sanderson MJ. Intercellular
propagation of calcium waves mediated by inositol trisphosphate.
Science. 1992;258:292–295.
25.
Sáez JC, Connor JA, Spray DC, Bennett MVL.
Hepatocyte gap junctions are permeable to the second
messenger inositol 1,4,5-trisphosphate, and to calcium ions. Proc
Natl Acad Sci U S A. 1989;86:2708–2712.
26.
Dekker RLC, Fiolet JWT, Van Bavel DE, Coronel R, Opthof
T, Spaan JAE, Janse MJ. Intracellular Ca2+,
intracellular electrical coupling, and mechanical activity in
ischemic rabbit papillary muscle: effects of preconditioning
and metabolic blockade. Circ Res. 1996;79:237–246.
27. Churchill G, Louis C. Roles of Ca2+, inositol trisphosphate and cyclic ADP-ribose in mediating intercellular Ca2+ signaling in sheep lens cells. J Cell Sci. 1998;111:1217–1225.[Abstract]
28. Rose CR, Ransom BR. Gap junctions equalize intracellular Na+ concentration in astrocytes. GLIA. 1997;20:299–307.[Medline] [Order article via Infotrieve]
29.
Schulze D, Kofuji P, Hadley R, Kirby MS, Kieval RS,
Doering A, Niggli E, Lederer WJ. Sodium/calcium exchanger in heart
muscle: molecular biology, cellular function, and its special role in
excitation-contraction coupling. Cardiovasc Res. 1993;27:1726–1734.
30. Tani M. Mechanism of Ca2+ overload in reperfused ischemic myocardium. Annu Rev Physiol. 1990;52:543–559.[Medline] [Order article via Infotrieve]
31. Bers DM, Bassani JW, Bassani RA. Na-Ca exchange and calcium fluxes during contraction and relaxation in mammalian ventricular muscle. Ann N Y Acad Sci. 1996;779:430–442.[Medline] [Order article via Infotrieve]
32.
Siegmund B, Koop A, Klietz T, Schwartz P, Piper HM.
Sarcolemmal integrity and metabolic competence of
cardiomyocytes under anoxia-reoxygenation.
Am J Physiol. 1990;258:H285–H291.
33. Ganote CE. Cell-to-cell interactions contributing to the oxygen paradox. Basic Res Cardiol. 1985;80:141–146.
34. Kimura H, Oyamada Y, Ohshika H, Mori M, Oyamada M. Reversible inhibition of gap junctional intercellular communication synchronous contraction, and synchronism of intracellular Ca2+ fluctuation in cultured neonatal rat cardiac myocytes by heptanol. Exp Cell Res. 1995;220:348–356.[Medline] [Order article via Infotrieve]
35. Diederichs F. Protection of isolated rat heart against the Ca2+ paradox: are gap junctions channels involved? J Mol Cell Cardiol. 1995;27:1301–1309.[Medline] [Order article via Infotrieve]
This article has been cited by other articles:
 |
|
 |
|
  M. Ruiz-Meana, A. Abellan, E. Miro-Casas, E. Agullo, and D. Garcia-Dorado Role of sarcoplasmic reticulum in mitochondrial permeability transition and cardiomyocyte death during reperfusion Am J Physiol Heart Circ Physiol, October 1, 2009; 297(4): H1281 - H1289. [Abstract] [Full Text] [PDF]
|
|
|
|
 |
|
 J. Inserte, J. A. Barrabes, V. Hernando, and D. Garcia-Dorado Orphan targets for reperfusion injury Cardiovasc Res, July 15, 2009; 83(2): 169 - 178. [Abstract] [Full Text] [PDF]
|
|
|
|
|
|
K. Naitoh, T. Yano, T. Miura, T. Itoh, T. Miki, M. Tanno, T. Sato, H. Hotta, Y. Terashima, and K. Shimamoto Roles of Cx43-associated protein kinases in suppression of gap junction-mediated chemical coupling by ischemic preconditioning Am J Physiol Heart Circ Physiol, February 1, 2009; 296(2): H396 - H403. [Abstract] [Full Text] [PDF]
|
|
|
|
 |
|
 J. H.-C. Lin, N. Lou, N. Kang, T. Takano, F. Hu, X. Han, Q. Xu, D. Lovatt, A. Torres, K. Willecke, et al. A Central Role of Connexin 43 in Hypoxic Preconditioning J. Neurosci., January 16, 2008; 28(3): 681 - 695. [Abstract] [Full Text] [PDF]
|
|
|
|
|
|
M. Ruiz-Meana, A. Rodriguez-Sinovas, A. Cabestrero, K. Boengler, G. Heusch, and D. Garcia-Dorado Mitochondrial connexin43 as a new player in the pathophysiology of myocardial ischaemia-reperfusion injury Cardiovasc Res, January 15, 2008; 77(2): 325 - 333. [Abstract] [Full Text] [PDF]
|
|
|
|
|
|
T. Miura, T. Yano, K. Naitoh, M. Nishihara, T. Miki, M. Tanno, and K. Shimamoto {delta}-Opioid receptor activation before ischemia reduces gap junction permeability in ischemic myocardium by PKC-{varepsilon}-mediated phosphorylation of connexin 43 Am J Physiol Heart Circ Physiol, September 1, 2007; 293(3): H1425 - H1431. [Abstract] [Full Text] [PDF]
|
|
|
|
|
|
W.-H. Zimmermann, M. Didie, S. Doker, I. Melnychenko, H. Naito, C. Rogge, M. Tiburcy, and T. Eschenhagen Heart muscle engineering: An update on cardiac muscle replacement therapy Cardiovasc Res, August 1, 2006; 71(3): 419 - 429. [Abstract] [Full Text] [PDF]
|
|
|
|
|
|
D. Garcia-Dorado, A. Rodriguez-Sinovas, M. Ruiz-Meana, J. Inserte, L. Agullo, and A. Cabestrero The end-effectors of preconditioning protection against myocardial cell death secondary to ischemia-reperfusion Cardiovasc Res, May 1, 2006; 70(2): 274 - 285. [Abstract] [Full Text] [PDF]
|
|
|
|
|
|
K. Naitoh, Y. Ichikawa, T. Miura, Y. Nakamura, T. Miki, Y. Ikeda, H. Kobayashi, M. Nishihara, K. Ohori, and K. Shimamoto MitoKATP channel activation suppresses gap junction permeability in the ischemic myocardium by an ERK-dependent mechanism Cardiovasc Res, May 1, 2006; 70(2): 374 - 383. [Abstract] [Full Text] [PDF]
|
|
|
|
|
|
A. Rodriguez-Sinovas, D. Garcia-Dorado, M. Ruiz-Meana, and J. Soler-Soler Protective effect of gap junction uncouplers given during hypoxia against reoxygenation injury in isolated rat hearts Am J Physiol Heart Circ Physiol, February 1, 2006; 290(2): H648 - H656. [Abstract] [Full Text] [PDF]
|
|
|
|
 |
|
 J. Inserte, D. Garcia-Dorado, V. Hernando, and J. Soler-Soler Calpain-Mediated Impairment of Na+/K+-ATPase Activity During Early Reperfusion Contributes to Cell Death After Myocardial Ischemia Circ. Res., September 2, 2005; 97(5): 465 - 473. [Abstract] [Full Text] [PDF]
|
|
|
|
|
|
K. Boengler, G. Dodoni, A. Rodriguez-Sinovas, A. Cabestrero, M. Ruiz-Meana, P. Gres, I. Konietzka, C. Lopez-Iglesias, D. Garcia-Dorado, F. Di Lisa, et al. Connexin 43 in cardiomyocyte mitochondria and its increase by ischemic preconditioning Cardiovasc Res, August 1, 2005; 67(2): 234 - 244. [Abstract] [Full Text] [PDF]
|
|
|
|
|
|
X. Ai and S. M. Pogwizd Connexin 43 Downregulation and Dephosphorylation in Nonischemic Heart Failure Is Associated With Enhanced Colocalized Protein Phosphatase Type 2A Circ. Res., January 7, 2005; 96(1): 54 - 63. [Abstract] [Full Text] [PDF]
|
|
|
|
|
|
S. J. Canyon and G. P. Dobson Protection against ventricular arrhythmias and cardiac death using adenosine and lidocaine during regional ischemia in the in vivo rat Am J Physiol Heart Circ Physiol, September 1, 2004; 287(3): H1286 - H1295. [Abstract] [Full Text] [PDF]
|
|
|
|
 |
|
 A. Rodriguez-Sinovas, D. Garcia-Dorado, M. Ruiz-Meana, and J. Soler-Soler Enhanced effect of gap junction uncouplers on macroscopic electrical properties of reperfused myocardium J. Physiol., August 15, 2004; 559(1): 245 - 257. [Abstract] [Full Text] [PDF]
|
|
|
|
|
|
J. R de Groot and R. Coronel Acute ischemia-induced gap junctional uncoupling and arrhythmogenesis Cardiovasc Res, May 1, 2004; 62(2): 323 - 334. [Abstract] [Full Text] [PDF]
|
|
|
|
|
|
R. Schulz and G. Heusch Connexin 43 and ischemic preconditioning Cardiovasc Res, May 1, 2004; 62(2): 335 - 344. [Abstract] [Full Text] [PDF]
|
|
|
|
|
|
H.M Piper, Y Abdallah, and C Schafer The first minutes of reperfusion: a window of opportunity for cardioprotection Cardiovasc Res, February 15, 2004; 61(3): 365 - 371. [Abstract] [Full Text] [PDF]
|
|
|
|
|
|
D. Garcia-Dorado, A. Rodriguez-Sinovas, and M. Ruiz-Meana Gap junction-mediated spread of cell injury and death during myocardial ischemia-reperfusion Cardiovasc Res, February 15, 2004; 61(3): 386 - 401. [Abstract] [Full Text] [PDF]
|
|
|
|
|
|
T. Miura, Y. Ohnuma, A. Kuno, M. Tanno, Y. Ichikawa, Y. Nakamura, T. Yano, T. Miki, J. Sakamoto, and K. Shimamoto Protective role of gap junctions in preconditioning against myocardial infarction Am J Physiol Heart Circ Physiol, January 1, 2004; 286(1): H214 - H221. [Abstract] [Full Text] [PDF]
|
|
|
|
|
|
F. Padilla, D. Garcia-Dorado, A. Rodriguez-Sinovas, M. Ruiz-Meana, J. Inserte, and J. Soler-Soler Protection afforded by ischemic preconditioning is not mediated by effects on cell-to-cell electrical coupling during myocardial ischemia-reperfusion Am J Physiol Heart Circ Physiol, November 1, 2003; 285(5): H1909 - H1916. [Abstract] [Full Text] [PDF]
|
|
|
|
|
|
J. R. de Groot, T. Veenstra, A. O. Verkerk, R. Wilders, J. P.P. Smits, F. J.G. Wilms-Schopman, R. F. Wiegerinck, J. Bourier, C. N.W. Belterman, R. Coronel, et al. Conduction slowing by the gap junctional uncoupler carbenoxolone Cardiovasc Res, November 1, 2003; 60(2): 288 - 297. [Abstract] [Full Text] [PDF]
|
|
|
|
|
|
F. Vetterlein, C. Schrader, R. Volkmann, M. Neckel, M. Ochs, G. Schmidt, and G. Hellige Extent of damage in ischemic, nonreperfused, and reperfused myocardium of anesthetized rats Am J Physiol Heart Circ Physiol, July 11, 2003; 285(2): H755 - H765. [Abstract] [Full Text] [PDF]
|
|
|
|
 |
|
 J. L. P. Velazquez, M. V. Frantseva, and C. C. Naus Gap Junctions and Neuronal Injury: Protectants or Executioners? Neuroscientist, February 1, 2003; 9(1): 5 - 9. [Abstract] [PDF]
|
|
|
|
 |
|
 H. M. Piper, K. Meuter, and C. Schafer Cellular mechanisms of ischemia-reperfusion injury Ann. Thorac. Surg., February 1, 2003; 75(2): S644 - 648. [Abstract] [Full Text] [PDF]
|
|
|
|
|
|
K. A. Detillieux, F. Sheikh, E. Kardami, and P. A. Cattini Biological activities of fibroblast growth factor-2 in the adult myocardium Cardiovasc Res, January 1, 2003; 57(1): 8 - 19. [Abstract] [Full Text] [PDF]
|
|
|
|
|
|
U. Schwanke, I. Konietzka, A. Duschin, X. Li, R. Schulz, and G. Heusch No ischemic preconditioning in heterozygous connexin43-deficient mice Am J Physiol Heart Circ Physiol, October 1, 2002; 283 (4): H1740 - H1742. [Abstract] [Full Text] [PDF]
|
|
|
|
|
|
J. Inserte, D. Garcia-Dorado, M. Ruiz-Meana, F. Padilla, J. A Barrabes, P. Pina, L. Agullo, H. M. Piper, and J. Soler-Soler Effect of inhibition of Na+/Ca2+ exchanger at the time of myocardial reperfusion on hypercontracture and cell death Cardiovasc Res, September 1, 2002; 55(4): 739 - 748. [Abstract] [Full Text] [PDF]
|
|
|
|
|
|
D. Garcia-Dorado, M. Ruiz-Meana, F. Padilla, A. Rodriguez-Sinovas, and M. Mirabet Gap junction-mediated intercellular communication in ischemic preconditioning Cardiovasc Res, August 15, 2002; 55(3): 456 - 465. [Abstract] [Full Text] [PDF]
|
|
|
|
|
|
M. V. Frantseva, L. Kokarovtseva, C. G. Naus, P. L. Carlen, D. MacFabe, and J. L. Perez Velazquez Specific Gap Junctions Enhance the Neuronal Vulnerability to Brain Traumatic Injury J. Neurosci., February 1, 2002; 22(3): 644 - 653. [Abstract] [Full Text] [PDF]
|
|
|
|
|
|
M. Ruiz-Meana, D. Garcia-Dorado, S. Lane, P. Pina, J. Inserte, M. Mirabet, and J. Soler-Soler Persistence of gap junction communication during myocardial ischemia Am J Physiol Heart Circ Physiol, June 1, 2001; 280(6): H2563 - H2571. [Abstract] [Full Text] [PDF]
|
|
|
|
 |
|
 D. Garcia-Dorado and M. Ruiz-Meana Propagation of Cell Death During Myocardial Reperfusion Physiology, December 1, 2000; 15(6): 326 - 330. [Abstract] [Full Text] [PDF]
|
|
|
|
|
|
G. Taimor Cardiac gap junctions: good or bad? Cardiovasc Res, October 1, 2000; 48(1): 8 - 10. [Full Text] [PDF]
|
|
|
|
 |
|
 W. L. Holman, J. L. Skinner, C. R. Killingsworth, J. M. Rogers, S. Melnick, R. E. Ideker, and S. B. Digerness CONTROLLED POSTCARDIOPLEGIA REPERFUSION: MECHANISM FOR ATTENUATION OF REPERFUSION INJURY J. Thorac. Cardiovasc. Surg., June 1, 2000; 119(6): 1093 - 1101. [Abstract] [Full Text] [PDF]
|
|
|
|
|
|
Z.-S. Jiang, R. R. Padua, H. Ju, B. W. Doble, Y. Jin, J. Hao, P. A. Cattini, I. M. C. Dixon, and E. Kardami Acute protection of ischemic heart by FGF-2: involvement of FGF-2 receptors and protein kinase C Am J Physiol Heart Circ Physiol, March 1, 2002; 282(3): H1071 - H1080. [Abstract] [Full Text] [PDF]
|
|
| |||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||