Protective Effects Of Cardioplegia With Sodium Channel Inhibitor Tetrodotoxin (TTX) On Ischemia/Reperfusion Myocardium Of Isolated Rat Hearts

Objective: To investigate the protective effects of cardioplegia with sodium channel inhibitor tetrodotoxin(TTX) on ischemia/reperfusion models of isolated rat hearts and isolated cardiomyocytes so as to explore a new myocardial protective method. Methods: Test 1. Langendorff perfusion models and Neely working left ventricular perfusion models of isolated rat hearts were sequentially created in Group TTX and Group STH-2. The hearts in each group were arrested by corresponding cardioplegia in Group TTX(study group, TTX ) and Group STH-2 (control group, only STH-2). The arrested hearts were preserved in the corresponding cardioplegia solution respectively at 7℃ for 5 hours, and then reperfused. Pre-ischemic and post-ischemic indexes were studied including hemodynamic parameters, myocardial enzymology, ATP content and ultrastructural changes. Test 2. Beating ventricular cardiomyocytes were isolated from adult rat hearts by enzymatic dissociation, and randomly created in Group Base, Group STH-2 and Group TTX. Group Base was only treated with K-H buffer solution. Group STH-2 and Group TTX were treated with STH-2 and TTX cardioplegia respectively, and the arrest/re-beating cell models imitating ischemia-reperfusion injury were established. All three groups were imaged by laser scanning confocal microscope with fluorescent probe for measuring [Na+]i and [Ca2+]i of cardiomyocytes in different stages. The beating status and morphology of cardiomyocytes were observed under the inverted microscope. Results: There were no differences in two groups of Langendorff rat hrart models before myocardial ischemia. But the recovery of hemodynamic parameters during reperfusion in Group TTX were better than those in Group STH-2(P<0.01); the releases of CPK and LDH in Group TTX were less than those in Group STH-2(P < 0.05); the activities of Na⁺-K⁺-ATPase, Ca²⁺-ATPase, Ca²⁺-Mg²⁺-ATPase , and contents of ATP in Group TTX kept relative higher level after reperfusion. Compared with Group STH-2, myocardial ultrastructure in Group TTX was better protected . The elevation of [Na⁺]i and [Ca²⁺]i of cardiomyocyte models during arrest in Group TTX was significantly lower than that in Group STH-2. [Na⁺]i and [Ca²⁺]i in both Group STH-2 and Group TTX after re-beating were higher than those in Group Base (P<0.01), but [Na⁺]i and [Ca²⁺]i in Group TTX were significantly lower than those in Group STH-2(P<0.01). There was no difference in beating rate between Group STH-2 and Group TTX(P>0.05). Active cardiomyocytes with normal morphology in Group TTX after re-beating were more counted than those in Group STH-2 (P<0.01). Conclusion: Compared with conventional hyperkalemic depolarizing cardioplegia, polarizing cardioplegia with sodium channel inhibitor TTX could alleviate intracellular Na+ -overload and Ca2+-overload in isolated cardiomyocytes of rat; reduce the consumption of energy; decrease the releases of CPK andLDH after reperfusion; avoid ultrastructural damage and prevent ischemia-reperfusion injury by arresting heart under polarizing condition. Therefore, polarizing cardioplegia with sodium channel inhibitor TTX might be a new and better cardioplegia in myocardial protection and preservation .