AIMS: Although voltage-sensitive dye di-4-ANEPPS is a common tool for mapping cardiac electrical activity, reported effects on electrophysiological parameters are rather. The main goals of the study were to reveal effects of the dye on rabbit isolated heart and to verify, whether rabbit isolated heart stained with di-4-ANEPPS is a suitable tool for myocardial ischemia investigation. METHODS AND RESULTS: Study involved experiments on stained (n = 9) and non-stained (n = 11) Langendorff perfused rabbit isolated hearts. Electrophysiological effects of the dye were evaluated by analysis of various electrogram (EG) parameters using common paired and unpaired statistical tests. It was shown that staining the hearts with di-4-ANEPPS leads to only short-term sporadic prolongation of impulse conduction through atria and atrioventricular node. On the other hand, significant irreversible slowing of heart rate and ventricular conduction were found in stained hearts as compared to controls. In patch clamp experiments, significant inhibition of sodium current density was observed in differentiated NG108-15 cells stained by the dye. Although no significant differences in mean number of ventricular premature beats were found between the stained and the non-stained hearts in ischemia as well as in reperfusion, all abovementioned results indicate increased arrhythmogenicity. In isolated hearts during ischemia, prominent ischemic patterns appeared in the stained hearts with 3-4 min delay as compared to the non-stained ones. Moreover, the ischemic changes did not achieve the same magnitude as in controls even after 10 min of ischemia. It resulted in poor performance of ischemia detection by proposed EG parameters, as was quantified by receiver operating characteristics analysis. CONCLUSION: Our results demonstrate significant direct irreversible effect of di-4-ANEPPS on spontaneous heart rate and ventricular impulse conduction in rabbit isolated heart model. Particularly, this should be considered when di-4-ANEPPS is used in ischemia studies in rabbit. Delayed attenuated response of such hearts to ischemia might lead to misinterpretation of obtained results.

• Keywords
• di-4-ANEPPS, electrogram analysis, myocardial ischemia, patch-clamp, rabbit isolated heart, voltage-sensitive dye,
• Publication type
• Journal Article MeSH

BACKGROUND: Detailed quantitative analysis of the effect of left ventricle (LV) hypertrophy on myocardial ischemia manifestation in ECG is still missing. The associations between both phenomena can be studied in animal models. In this study, rabbit isolated hearts with spontaneously increased LV mass were used to evaluate the effect of such LV alteration on ischemia detection criteria and performance. METHODS: Electrophysiological effects of increased LV mass were evaluated on sixteen New Zealand rabbit isolated hearts under non-ischemic and ischemic conditions by analysis of various electrogram (EG) parameters. To reveal hearts with increased LV mass, LV weight/heart weight ratio was proposed. Standard paired and unpaired statistical tests and receiver operating characteristics analysis were used to compare data derived from different groups of animals, monitor EG parameters during global ischemia and evaluate their ability to discriminate between unchanged and increased LV as well as non-ischemic and ischemic state. RESULTS: Successful evaluation of both increased LV mass and ischemia is lead-dependent. Particularly, maximal deviation of QRS and area under QRS associated with anterolateral heart wall respond significantly to even early phase (the 1st-3rd min) of ischemia. Besides ischemia, these parameters reflect increased LV mass as well (with sensitivity reaching approx. 80%). However, the sensitivity of the parameters to both phenomena may lead to misinterpretations, when inappropriate criteria for ischemia detection are selected. Particularly, use of cut-off-based criteria defined from control group for ischemia detection in hearts with increased LV mass may result in dramatic reduction (approx. 15%) of detection specificity due to increased number of false positives. Nevertheless, criteria adjusted to particular experimental group allow achieving ischemia detection sensitivity of 89-100% and specificity of 94-100%, respectively. CONCLUSIONS: It was shown that response of the heart to myocardial ischemia can be successfully evaluated only when taking into account heart-related factors (such as LV mass) and other methodological aspects (such as recording electrodes position, selected EG parameters, cut-off criteria, etc.). Results of this study might be helpful for developing new clinical diagnostic strategies in order to improve myocardial ischemia detection in patients with LV hypertrophy.

• Keywords
• Electrogram, Increased left ventricular mass, Isolated heart, Myocardial ischemia detection, ROC analysis, Rabbit,
• MeSH
• Electrophysiologic Techniques, Cardiac * MeSH
• Electrocardiography * MeSH
• Ventricular Function, Left * MeSH
• Hypertrophy, Left Ventricular complications   diagnosis   physiopathology   MeSH
• Myocardial Ischemia complications   diagnosis   physiopathology   MeSH
• Rabbits MeSH
• Disease Models, Animal MeSH
• Area Under Curve MeSH
• Signal Processing, Computer-Assisted MeSH
• Predictive Value of Tests MeSH
• Isolated Heart Preparation MeSH
• Ventricular Remodeling * MeSH
• Risk Factors MeSH
• ROC Curve MeSH
• Animals MeSH
• Check Tag
• Rabbits MeSH
• Male MeSH
• Female MeSH
• Animals MeSH
• Publication type
• Journal Article MeSH
• Research Support, Non-U.S. Gov't MeSH

Isolated heart model is a relevant tool for cardiovascular system studies. It represents a highly reproducible model for studying broad spectrum of biochemical, physiological, morphological, and pharmaceutical parameters, including analysis of intrinsic heart mechanics, metabolism, and coronary vascular response. Results obtained in this model are under no influence of other organ systems, plasma concentration of hormones or ions and influence of autonomic nervous system. The review describes various isolated heart models, the modes of heart perfusion, and advantages and limitations of various experimental setups. It reports the improvements of perfusion setup according to Langendorff introduced by the authors.

• MeSH
• Biomedical Research * history   methods   MeSH
• History, 19th Century MeSH
• History, 20th Century MeSH
• History, 21st Century MeSH
• Rats MeSH
• Humans MeSH
• Models, Cardiovascular * MeSH
• Mice MeSH
• Perfusion MeSH
• Heart * anatomy & histology   physiology   MeSH
• Anura MeSH
• Animals MeSH
• Check Tag
• History, 19th Century MeSH
• History, 20th Century MeSH
• History, 21st Century MeSH
• Rats MeSH
• Humans MeSH
• Mice MeSH
• Animals MeSH
• Publication type
• Journal Article MeSH
• Historical Article MeSH
• Research Support, Non-U.S. Gov't MeSH
• Review MeSH