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Gap junction remodelling is involved in the susceptibility of diabetic rats to hypokalemia-induced ventricular fibrillation
acta histochem. 104(4) 387–391 (2002) © Urban & Fischer Verlag
http://www.urbanfischer.de/journals/actahist
Gap junction remodelling is involved in the susceptibility of diabetic rats to hypokalemia-induced ventricular fibrillation Ludmila Okruhlicova1*, Narcis Tribulova1, Melania Misˇejkova2, Marek Kucˇka2, Radovan Sˇtetka1, Jan Slezak1, and Mordechai Manoach3 1
Institute for Heart Research, Slovak Academy of Sciences, Faculty of Natural Sciences Commenius University, Department of Animal Physiology and Etology, Bratislava, Slovak Republic, and 3 Department of Physiology, Tel Aviv University, Tel Aviv, Israel 2
Received 19 February 2002 and in revised form 3 June 2002 and 12 July 2002; accepted 13 August 2002
Summary The objective of the present study was to examine the susceptibility of diabetic rats with cardiomyopathy to hypokalemia-induced ventricular fibrillation and to localize gap junction protein connexin-43 as well as subcellular changes that may be involved in the development of severe arrhythmia. Our results showed a significantly higher incidence of sustained ventricular fibrillation in diabetic hearts as compared with control hearts, 80% vs 20%, respectively. Diabetic cardiomyopathy itself was accompanied by a distinct decrease in connexin-43-immunopositive gap junctions. Moreover, interstitial fibrosis and subcellular alterations to various degrees were observed in diabetic hearts, and a further deterioration of the ultrastructure and impairment of intercellular junctions, and a stronger local decrease in connexin43 levels due to hypokalemia were found. These changes were heterogeneously distributed throughout the myocardium and occurred earlier and were more pronounced in diabetic hearts than control hearts. In conclusion, our results indicate that diabetic cardiomyopathy is associated with down-regulation of gap junction proteins and may account for the higher vulnerability of diabetic rats to ventricular fibrillation in combination with impairment of intercellular communication due to hypokalemia. Key words: diabetic heart – connexin-43 – ventricular fibrillation
Introduction Experimental studies have demonstrated either increased or decreased susceptibility of the diabetic heart to ischemic injury (Ravingerova et al., 2001). Likewise, there are conflicting data with respect to susceptibility to arrhythmias (Beatch and McNeill, 1988; Ravingerova et al., 2001). Development of cardiomyopathy leads to deterioration of heart function as a result of changes in metabolism (Rodrigues at al., 1995),
alterations in ion transport systems as well as subcellular changes (Dhalla et al., 1998) and extracellular matrix remodelling (Tribulova et al., 1996; Okruhlicova et al., 2002). Recently, it was shown that ageing and hypertension associated with myocardial remodelling result in weakening of cell-cell communication via gap junctions and elevated susceptibility to arrhythmias, such as atrial
*Correspondence to: Dr. Ludmila Okruhlicova, Institute for Heart Research, Slovak Academy of Sciences, Dubravska cesta 9, 842 33 Bratislava, Slovak Republic; tel: +421 2 54774405; fax: +421 2 54776637; e-mail: [email protected]
0065-1281/02/104/04-387 $ 15.00/0
388
Okruhlicova et al.
and/or ventricular fibrillation (Tribulova et al., 1999, 2002). Arrhythmias are known be linked to both impairment of intercellular communication via gap junctions (Peters et al., 1997; Saffitz et al., 1999; Tribulova et al., 2001) and an increase in cytosolic free Ca2+ concentrations (Tribulova et al., 2002). The objective of the present study was the examination of susceptibility of diabetic rats to hypokalemiainduced ventricular fibrillation and to detect changes in gap junctions that may be involved in the genesis of this fatal arrhythmia.
Material and methods Diabetes was induced by a single iv injection of streptozotocin (45 mg/kg) and after 10 weeks, hearts of anaesthetised diabetic rats (n = 12) and age-matched control rats (n = 12) were rapidly excised. Hearts were cannu-
lated via the aorta and perfused in the Langendorff mode at a constant pressure of 70 mm Hg and 37 °C with oxygenated (95% O2 and 5% CO2) Krebs-Henseleit solution (4.4 mmol/l K+). Epicardial electrograms were registered continuously and 20 min of stabilisation was followed by perfusion of the heart with a solution low in K+ ions (1 mmol/l) for a period of 30 min, unless sustained ventricular fibrillation occurred earlier. Ventricular tissue sampling for immunohistochemical localization of gap junction-specific connexin-43 and ultrastructural examination was performed either during the stabilisation period, during low K+ perfusion and/or at the onset of fibrillation. For the examination of ultrastructure, hearts were fixed by perfusion with buffered 2.5% glutaraldehyde and small ventricular blocks of epi-, mid- and endocardial zones were routinely processed and embedded in Epon 812 for cutting ultrathin sections. Immunolabelling of gap junction protein connexin-43 was per-
Fig. 1. Immunodetection of gap junctional connexin-43 in myocardium of control (A, C) and diabetic (B, D) rat hearts. Note the lower number and nonuniform distribution of immunopositive gap junctions in diabetic rat heart (B) as compared with control heart (A) as well as a further focal decrease in immunofluorescence due to hypokalemia in both control (C) and diabetic (D) rats. Bar, 10 µm.
acta histochemica 104, 4 (2002)
Connexin-43 and arrhythmias
Fig. 2. Quantitative evaluation of connexin-43 staining showing a distinct decrease in immunopositivity in diabetic rat hearts (Dia) as compared with control (C) heart and a significant decrease in both groups after perfusion with a K+-deficient solution as compared with normal perfusion. All groups are significantly different (P < 0.05).
389
formed on cryostat sections (10 µm thick). Sections were fixed in 1% paraformaldehyde in PBS for 10 min and incubated with a monoclonal mouse anti-connexin43 antibody (dilution 1:100) for 2 h at room temp and subsequently with goat anti-mouse IgG antibodies conjugated with fluorescein isothiocyanate (dilution 1:50) for 1 h at room temp. Antibodies were purchased from Zymed (San Francisco CA, USA). Non-immune goat serum and incubation without primary antibodies served as negative controls. Sections were examined with a fluorescence microscope (Nikon, Tokyo, Japan) and evaluated with an image analysis system (LUCIA; Laboratory Imaging Ltd, Prague, Czech Republic). The mean of the fluorescence signal was obtained from 15 objects for each group (6 rats). Data were expressed as mean ± SEM and comparison of the groups was performed using ANOVA followed by a Student-test. The level of significance was defined as P < 0.05.
Fig. 3. Subcellular alterations of cardiomyocytes and their junctions in control (A, C) and diabetic (B, D) rat hearts during low K+ perfusion followed by transient arrhythmias. Representative images show distinct (B, C) or slight (A, D) impairment of cell-cell junctions (arrows) as well as hypercontractions induced by Ca2+ overload (A, D) and mitochondria alterations (A, D). Desynchronisation of contraction (A, C, D) between neighbouring cardiomyocytes indicate dysfunction of cell-cell communication via gap junctions.
acta histochemica 104, 4 (2002)
390
Okruhlicova et al.
Results Diabetes-related cardiomyopathy was characterised by various myocardial alterations, such as interstitial fibrosis, microangiopathy and populations of ischemia-like altered cardiomyocytes. These changes were accompanied by a decrease in the activity of oxidative enzymes, ATPases and 5´-nucleotidase and a decrease in density of alkaline phosphatase-positive capillaries (Tribulova et al., 1996; Okruhlicova et al., 2002). Immunolabelling of connexin-43 and ultrastructural examination showed heterogeneous distribution patterns and decreased amounts of gap junctions (Figs. 1–3). Exposure of the heart to hypokalemia that was followed by transient arrhythmias caused heterogeneous and for the larger part reversible but in some cases irreversible injury of cardiomyocytes and intercellular junctions in normal and diabetic rats (Fig. 3). Subcellular alterations indicated calcium overload and disturbances in intercellular synchronisation as occur in cardiomyopathic hearts. Moreover, a significant local decrease in connexin-43 immunostaining (P < 0.05; Fig. 1) preceded ventricular fibrillation. It was more pronounced in diabetic rat hearts as was shown by quantitative evaluation (Fig. 2). Sustained ventricular fibrillation occurred within 30 min perfusion of isolated heart with K+-deficient Krebs-Henseleit solution in 80% of the diabetic hearts versus 20% of control hearts. Premature beats appeared earlier and transient ventricular tachycardia occurred more frequently in diabetic rats (data not shown).
Discussion The major findings of the present study are: 1) changes in distribution and expression patterns of the major ventricular gap junction protein connexin-43 may be involved in hypokalemia-induced ventricular fibrillation, and 2) the higher susceptibility of diabetic rats to fatal arrhythmia. Like hypertension-related myocardial changes, chronic diabetes was manifested by a decrease in the activity of oxidative enzymes that have been monitored with the use of histochemistry (Tribulova et al., 1996, 2000; Okruhlicova et al., 2002) or biochemistry (Chen et al., 1984). Increased alkaline phosphatase activity in the insterstitium delineated clearly areas of early fibrosis. On the other hand, focal depletion of its endothelial activity may indicate abnormal capillary blood supply. Subcellular alterations correlated with histochemical changes in both models of chronic heart failure and revealed ischemia-like injury and extracellular matrix alterations with concomitant fibrosis. These changes were accompanied by decreased immunostaining of connexin-43 that
acta histochemica 104, 4 (2002)
was more heterogeneous in ventricles of diabetic rats, like in hypertensive rats (Tribulova et al., 2001). The closest contact between cardiomyocytes is at the site of gap junctions that are known to be composed of connexin-43 proteins. They form intercellular channels ensuring fast electrical and metabolic signal transduction. Therefore, they are responsible for synchronous and co-ordinated functioning of cardiac muscle. Impairment and/or abnormal cell-cell communication in diabetic hearts (De Mello and Gonzales, 2001; Imanaga et al., 1999) may cause a higher susceptibility to arrhythmias (Manoach et al., 1996; Peters et al., 1997; Saffitz et al., 1999; Tribulova et al., 2001). The present study and previous studies support the assumption that overall myocardial structural remodelling involves gap junction remodelling and as a result decreased cell-cell communication (Tribulova et al., 1999, 2002). Further impairment of cell-cell communication due to low external K+ levels, and transient arrhythmias may contribute to the electrical instability and degeneration of these arrhythmias to ventricular fibrillation. Acknowledgments This study was supported by VEGA grant numbers 2/7155/21 and 2/2064/22. The authors are grateful to Dr. K. Volkovova for providing diabetic rats and Mrs. A. Brichtova and A. Macsaliova for their technical assistance.
References Beatch GN, and McNeill JH (1988) Ventricular arrhythmias following coronary artery occlusion in the streptozotocin diabetic rat. Can J Physiol Pharmacol 66: 312–317 Chen V, Ianuzzo CD, Fong BC, and Spitzer JJ (1984) The effect of acute and chronic diabetes on myocardial metabolism in rats. Diabetes 33: 1078–1084 DeMello W, and Gonzalez M (2001) Cell communication and impulse propagation are impaired in the diabetic heart. J Mol Cell Cardiol 33: A27 (abstract) Dhalla NS, Liu X, Panagia V, and Takeda N (1998) Subcellular remodeling and heart dysfunction in chronic diabetes. Cardiovasc Res 40: 239–247 Imanaga I, Inai T, Hirosawa N, and Shibata Y (1999) Abnormalities of intercellular impulse propagation in diabetic cardiac muscle. Proceedings of the diabetic heart conference. Winnipeg ON, Canada, p 41 (abstract) Manoach M, Tribulova N, and Imanaga I (1996) The protective effect of d-sotalol against hypoxia-induced myocardial uncoupling. Heart Vessels 11: 281–288 Okruhlicova L, Sotnikova R, Sˇ tefek M, Tribulova N, Weismann P, Gajdosˇik A, and Gajdosikova A (2002) L-Arginine reduces structural remodeling in the diabetic rat myocardium. Meth Find Exp Clin Pharmacol, 24: 201–207 Peters NS, Coromilas J, Severs NJ, and Wit AL (1997) Disturbed connexin-43 gap junction distribution correlates with location of re-entrant circuits in the epicardial border zone of healing infarcts that cause ventricular tachycardia. Circulation 95: 988–996
Connexin-43 and arrhythmias Ravingerova T, Neckar J, Kolar F, Stetka R, Volkovova K, Ziegelhoffer A, and Styk J (2001) Ventricular arrhythmias following coronary artery occlusion in rats: is the diabetic heart less or more sensitive to ischemia? Basic Res Cardiol 96: 160–168 Rodrigues B, Cam CM, and McNeill JH (1995) Myocardial substrate metabolism: Implication for diabetic cardiomyopathy. J Mol Cell Cardiol 27: 169–179 Saffitz J, Schuessler RB, and Yamada KA (1999) Mechanisms of remodeling of gap junction distribution and the development of anatomic substrates of arrhythmias. Cardiovasc Res 42: 309–317 Tribulova N, Ravingerova T, Volkovova K, Ziegelhoffer A, Okruhlicova L, Ziegelhoffer B, Styk J, and Slezak J (1996) Resistance of diabetic rat hearts to Ca overload-related injury. Histochemical and ultrastructural study. Diabetes Res Clin Practice 31: 113–122
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Tribulova N, Varon D, Polack-Charcon S, Buscemi P, Slezak J, and Manoach M (1999) Aged heart as a model for prolonged atrial fibrilloflutter. Exp Clin Cardiol 4: 64–72 Tribulova N, Okruhlicova L, Bernatova I, and Pechanova O, (2000) Chronic disturbances in NO production results in histochemical and subcellular alterations of the rat heart. Physiol Res 49: 77–88 Tribulova N, Okruhlicova L, Novakova S, Pancza D, Bernatova I, Pechanova O, Weismann P, Manoach M, Seki S, and Mochizuki M (2002) Hypertension-related intermyocyte junctions remodeling is associated with higher incidence of low K+-induced lethal arrhythmias, in isolated rat heart. Exp Physiol 82: 195–205 Tribulova N, Manoach M, Varon D, Okruhlicova L, Zinman T, and Shainber A (2001) Dispersion of cell-to-cell uncoupling precedes low K+-induced ventricular fibrillation. Physiol Res 50: 247–259
acta histochemica 104, 4 (2002)
http://www.urbanfischer.de/journals/actahist
Gap junction remodelling is involved in the susceptibility of diabetic rats to hypokalemia-induced ventricular fibrillation Ludmila Okruhlicova1*, Narcis Tribulova1, Melania Misˇejkova2, Marek Kucˇka2, Radovan Sˇtetka1, Jan Slezak1, and Mordechai Manoach3 1
Institute for Heart Research, Slovak Academy of Sciences, Faculty of Natural Sciences Commenius University, Department of Animal Physiology and Etology, Bratislava, Slovak Republic, and 3 Department of Physiology, Tel Aviv University, Tel Aviv, Israel 2
Received 19 February 2002 and in revised form 3 June 2002 and 12 July 2002; accepted 13 August 2002
Summary The objective of the present study was to examine the susceptibility of diabetic rats with cardiomyopathy to hypokalemia-induced ventricular fibrillation and to localize gap junction protein connexin-43 as well as subcellular changes that may be involved in the development of severe arrhythmia. Our results showed a significantly higher incidence of sustained ventricular fibrillation in diabetic hearts as compared with control hearts, 80% vs 20%, respectively. Diabetic cardiomyopathy itself was accompanied by a distinct decrease in connexin-43-immunopositive gap junctions. Moreover, interstitial fibrosis and subcellular alterations to various degrees were observed in diabetic hearts, and a further deterioration of the ultrastructure and impairment of intercellular junctions, and a stronger local decrease in connexin43 levels due to hypokalemia were found. These changes were heterogeneously distributed throughout the myocardium and occurred earlier and were more pronounced in diabetic hearts than control hearts. In conclusion, our results indicate that diabetic cardiomyopathy is associated with down-regulation of gap junction proteins and may account for the higher vulnerability of diabetic rats to ventricular fibrillation in combination with impairment of intercellular communication due to hypokalemia. Key words: diabetic heart – connexin-43 – ventricular fibrillation
Introduction Experimental studies have demonstrated either increased or decreased susceptibility of the diabetic heart to ischemic injury (Ravingerova et al., 2001). Likewise, there are conflicting data with respect to susceptibility to arrhythmias (Beatch and McNeill, 1988; Ravingerova et al., 2001). Development of cardiomyopathy leads to deterioration of heart function as a result of changes in metabolism (Rodrigues at al., 1995),
alterations in ion transport systems as well as subcellular changes (Dhalla et al., 1998) and extracellular matrix remodelling (Tribulova et al., 1996; Okruhlicova et al., 2002). Recently, it was shown that ageing and hypertension associated with myocardial remodelling result in weakening of cell-cell communication via gap junctions and elevated susceptibility to arrhythmias, such as atrial
*Correspondence to: Dr. Ludmila Okruhlicova, Institute for Heart Research, Slovak Academy of Sciences, Dubravska cesta 9, 842 33 Bratislava, Slovak Republic; tel: +421 2 54774405; fax: +421 2 54776637; e-mail: [email protected]
0065-1281/02/104/04-387 $ 15.00/0
388
Okruhlicova et al.
and/or ventricular fibrillation (Tribulova et al., 1999, 2002). Arrhythmias are known be linked to both impairment of intercellular communication via gap junctions (Peters et al., 1997; Saffitz et al., 1999; Tribulova et al., 2001) and an increase in cytosolic free Ca2+ concentrations (Tribulova et al., 2002). The objective of the present study was the examination of susceptibility of diabetic rats to hypokalemiainduced ventricular fibrillation and to detect changes in gap junctions that may be involved in the genesis of this fatal arrhythmia.
Material and methods Diabetes was induced by a single iv injection of streptozotocin (45 mg/kg) and after 10 weeks, hearts of anaesthetised diabetic rats (n = 12) and age-matched control rats (n = 12) were rapidly excised. Hearts were cannu-
lated via the aorta and perfused in the Langendorff mode at a constant pressure of 70 mm Hg and 37 °C with oxygenated (95% O2 and 5% CO2) Krebs-Henseleit solution (4.4 mmol/l K+). Epicardial electrograms were registered continuously and 20 min of stabilisation was followed by perfusion of the heart with a solution low in K+ ions (1 mmol/l) for a period of 30 min, unless sustained ventricular fibrillation occurred earlier. Ventricular tissue sampling for immunohistochemical localization of gap junction-specific connexin-43 and ultrastructural examination was performed either during the stabilisation period, during low K+ perfusion and/or at the onset of fibrillation. For the examination of ultrastructure, hearts were fixed by perfusion with buffered 2.5% glutaraldehyde and small ventricular blocks of epi-, mid- and endocardial zones were routinely processed and embedded in Epon 812 for cutting ultrathin sections. Immunolabelling of gap junction protein connexin-43 was per-
Fig. 1. Immunodetection of gap junctional connexin-43 in myocardium of control (A, C) and diabetic (B, D) rat hearts. Note the lower number and nonuniform distribution of immunopositive gap junctions in diabetic rat heart (B) as compared with control heart (A) as well as a further focal decrease in immunofluorescence due to hypokalemia in both control (C) and diabetic (D) rats. Bar, 10 µm.
acta histochemica 104, 4 (2002)
Connexin-43 and arrhythmias
Fig. 2. Quantitative evaluation of connexin-43 staining showing a distinct decrease in immunopositivity in diabetic rat hearts (Dia) as compared with control (C) heart and a significant decrease in both groups after perfusion with a K+-deficient solution as compared with normal perfusion. All groups are significantly different (P < 0.05).
389
formed on cryostat sections (10 µm thick). Sections were fixed in 1% paraformaldehyde in PBS for 10 min and incubated with a monoclonal mouse anti-connexin43 antibody (dilution 1:100) for 2 h at room temp and subsequently with goat anti-mouse IgG antibodies conjugated with fluorescein isothiocyanate (dilution 1:50) for 1 h at room temp. Antibodies were purchased from Zymed (San Francisco CA, USA). Non-immune goat serum and incubation without primary antibodies served as negative controls. Sections were examined with a fluorescence microscope (Nikon, Tokyo, Japan) and evaluated with an image analysis system (LUCIA; Laboratory Imaging Ltd, Prague, Czech Republic). The mean of the fluorescence signal was obtained from 15 objects for each group (6 rats). Data were expressed as mean ± SEM and comparison of the groups was performed using ANOVA followed by a Student-test. The level of significance was defined as P < 0.05.
Fig. 3. Subcellular alterations of cardiomyocytes and their junctions in control (A, C) and diabetic (B, D) rat hearts during low K+ perfusion followed by transient arrhythmias. Representative images show distinct (B, C) or slight (A, D) impairment of cell-cell junctions (arrows) as well as hypercontractions induced by Ca2+ overload (A, D) and mitochondria alterations (A, D). Desynchronisation of contraction (A, C, D) between neighbouring cardiomyocytes indicate dysfunction of cell-cell communication via gap junctions.
acta histochemica 104, 4 (2002)
390
Okruhlicova et al.
Results Diabetes-related cardiomyopathy was characterised by various myocardial alterations, such as interstitial fibrosis, microangiopathy and populations of ischemia-like altered cardiomyocytes. These changes were accompanied by a decrease in the activity of oxidative enzymes, ATPases and 5´-nucleotidase and a decrease in density of alkaline phosphatase-positive capillaries (Tribulova et al., 1996; Okruhlicova et al., 2002). Immunolabelling of connexin-43 and ultrastructural examination showed heterogeneous distribution patterns and decreased amounts of gap junctions (Figs. 1–3). Exposure of the heart to hypokalemia that was followed by transient arrhythmias caused heterogeneous and for the larger part reversible but in some cases irreversible injury of cardiomyocytes and intercellular junctions in normal and diabetic rats (Fig. 3). Subcellular alterations indicated calcium overload and disturbances in intercellular synchronisation as occur in cardiomyopathic hearts. Moreover, a significant local decrease in connexin-43 immunostaining (P < 0.05; Fig. 1) preceded ventricular fibrillation. It was more pronounced in diabetic rat hearts as was shown by quantitative evaluation (Fig. 2). Sustained ventricular fibrillation occurred within 30 min perfusion of isolated heart with K+-deficient Krebs-Henseleit solution in 80% of the diabetic hearts versus 20% of control hearts. Premature beats appeared earlier and transient ventricular tachycardia occurred more frequently in diabetic rats (data not shown).
Discussion The major findings of the present study are: 1) changes in distribution and expression patterns of the major ventricular gap junction protein connexin-43 may be involved in hypokalemia-induced ventricular fibrillation, and 2) the higher susceptibility of diabetic rats to fatal arrhythmia. Like hypertension-related myocardial changes, chronic diabetes was manifested by a decrease in the activity of oxidative enzymes that have been monitored with the use of histochemistry (Tribulova et al., 1996, 2000; Okruhlicova et al., 2002) or biochemistry (Chen et al., 1984). Increased alkaline phosphatase activity in the insterstitium delineated clearly areas of early fibrosis. On the other hand, focal depletion of its endothelial activity may indicate abnormal capillary blood supply. Subcellular alterations correlated with histochemical changes in both models of chronic heart failure and revealed ischemia-like injury and extracellular matrix alterations with concomitant fibrosis. These changes were accompanied by decreased immunostaining of connexin-43 that
acta histochemica 104, 4 (2002)
was more heterogeneous in ventricles of diabetic rats, like in hypertensive rats (Tribulova et al., 2001). The closest contact between cardiomyocytes is at the site of gap junctions that are known to be composed of connexin-43 proteins. They form intercellular channels ensuring fast electrical and metabolic signal transduction. Therefore, they are responsible for synchronous and co-ordinated functioning of cardiac muscle. Impairment and/or abnormal cell-cell communication in diabetic hearts (De Mello and Gonzales, 2001; Imanaga et al., 1999) may cause a higher susceptibility to arrhythmias (Manoach et al., 1996; Peters et al., 1997; Saffitz et al., 1999; Tribulova et al., 2001). The present study and previous studies support the assumption that overall myocardial structural remodelling involves gap junction remodelling and as a result decreased cell-cell communication (Tribulova et al., 1999, 2002). Further impairment of cell-cell communication due to low external K+ levels, and transient arrhythmias may contribute to the electrical instability and degeneration of these arrhythmias to ventricular fibrillation. Acknowledgments This study was supported by VEGA grant numbers 2/7155/21 and 2/2064/22. The authors are grateful to Dr. K. Volkovova for providing diabetic rats and Mrs. A. Brichtova and A. Macsaliova for their technical assistance.
References Beatch GN, and McNeill JH (1988) Ventricular arrhythmias following coronary artery occlusion in the streptozotocin diabetic rat. Can J Physiol Pharmacol 66: 312–317 Chen V, Ianuzzo CD, Fong BC, and Spitzer JJ (1984) The effect of acute and chronic diabetes on myocardial metabolism in rats. Diabetes 33: 1078–1084 DeMello W, and Gonzalez M (2001) Cell communication and impulse propagation are impaired in the diabetic heart. J Mol Cell Cardiol 33: A27 (abstract) Dhalla NS, Liu X, Panagia V, and Takeda N (1998) Subcellular remodeling and heart dysfunction in chronic diabetes. Cardiovasc Res 40: 239–247 Imanaga I, Inai T, Hirosawa N, and Shibata Y (1999) Abnormalities of intercellular impulse propagation in diabetic cardiac muscle. Proceedings of the diabetic heart conference. Winnipeg ON, Canada, p 41 (abstract) Manoach M, Tribulova N, and Imanaga I (1996) The protective effect of d-sotalol against hypoxia-induced myocardial uncoupling. Heart Vessels 11: 281–288 Okruhlicova L, Sotnikova R, Sˇ tefek M, Tribulova N, Weismann P, Gajdosˇik A, and Gajdosikova A (2002) L-Arginine reduces structural remodeling in the diabetic rat myocardium. Meth Find Exp Clin Pharmacol, 24: 201–207 Peters NS, Coromilas J, Severs NJ, and Wit AL (1997) Disturbed connexin-43 gap junction distribution correlates with location of re-entrant circuits in the epicardial border zone of healing infarcts that cause ventricular tachycardia. Circulation 95: 988–996
Connexin-43 and arrhythmias Ravingerova T, Neckar J, Kolar F, Stetka R, Volkovova K, Ziegelhoffer A, and Styk J (2001) Ventricular arrhythmias following coronary artery occlusion in rats: is the diabetic heart less or more sensitive to ischemia? Basic Res Cardiol 96: 160–168 Rodrigues B, Cam CM, and McNeill JH (1995) Myocardial substrate metabolism: Implication for diabetic cardiomyopathy. J Mol Cell Cardiol 27: 169–179 Saffitz J, Schuessler RB, and Yamada KA (1999) Mechanisms of remodeling of gap junction distribution and the development of anatomic substrates of arrhythmias. Cardiovasc Res 42: 309–317 Tribulova N, Ravingerova T, Volkovova K, Ziegelhoffer A, Okruhlicova L, Ziegelhoffer B, Styk J, and Slezak J (1996) Resistance of diabetic rat hearts to Ca overload-related injury. Histochemical and ultrastructural study. Diabetes Res Clin Practice 31: 113–122
391
Tribulova N, Varon D, Polack-Charcon S, Buscemi P, Slezak J, and Manoach M (1999) Aged heart as a model for prolonged atrial fibrilloflutter. Exp Clin Cardiol 4: 64–72 Tribulova N, Okruhlicova L, Bernatova I, and Pechanova O, (2000) Chronic disturbances in NO production results in histochemical and subcellular alterations of the rat heart. Physiol Res 49: 77–88 Tribulova N, Okruhlicova L, Novakova S, Pancza D, Bernatova I, Pechanova O, Weismann P, Manoach M, Seki S, and Mochizuki M (2002) Hypertension-related intermyocyte junctions remodeling is associated with higher incidence of low K+-induced lethal arrhythmias, in isolated rat heart. Exp Physiol 82: 195–205 Tribulova N, Manoach M, Varon D, Okruhlicova L, Zinman T, and Shainber A (2001) Dispersion of cell-to-cell uncoupling precedes low K+-induced ventricular fibrillation. Physiol Res 50: 247–259
acta histochemica 104, 4 (2002)