An Experimental Study That The Sphingosine 1-phosphate May Be Involved In Occurrence Of Arrhythmia During Acute Myocardial Infarction

Sphingosine 1-phosphate (S1P) is a intermediary metabolite of sphingolipid and an important member of lysophospholipid family, formed by phosphorylation of sphingosine catalyzed by sphingosine kinase. It produces a wide range of biological effects as an extracellular ligand for specific G protein-coupled receptors EDG.s and plays a very important role in physiological and pathological processes. The people more and more pay attention to the relationship between S1P and diseases. The rate of arrhythmia occurrence during acute myocardial infarction is very high. Its mechanism is probably relative to the change of cardiac myocytes metabolism and mechanical and electrophysiological property induced by ischemia and anoxia. The detailed mechanism in this process is still not very clear. The concentration of S1P may increase in the local region of ischemia during acute myocardial infarction because the phospholipase and platelets are activated. It was reported that the level of S1P in coronary heart disease increases with the degree of coronary artery narrow. Moreover, there are EDG receptors sensitive to S1P distributed in cardiac myocytes. All these suggest that S1P may play an important role in arrhythmia occurrence complicated by acute myocardial infarction. To date, the relationship between S1P and arrhythmia complicated by acute myocardial infarction have not been reported yet. The effects of S1P on cardiomyocytes ion channel are very few. In order to further find the relationship between S1P and arrhythmia complicated by acute myocardial infarction, we investigated the effects of S1P on ECG, parameters of cardiac action potential and membrane currents of calcium, potassium and sodium. We expected to obtain a theoretical explanation and supplement of the mechanism of arrhythmia complicated by acute myocardial infarction. Moreover, we also excepted to find some specific antagonist as drugs to block the EDG receptors and the signaling transduction link of S1P for prevention and therapy of this kind of arrhythmia. Ⅰ. Effects of S1P on the arrhythmia during acute myocardial infarction. ECG of rats was used as an index of present study. After increasing the concentration of S1P in the local region of myocardial infarction, the times of extrasystole occurrence (8.13±4.67) times/2h were obviously higher than those of model control group (1.14±1.07) times/2h (p <0.01). But if pretreated with the antagonist of G protein PTX before administrated with S1P, there was no significant difference compared with model control group (p>0.05). All these suggested that S1P could induce the arrhythmia occurrence after acute myocardial infarction via a G-protein-coupled receptors EDG pathway. Ⅱ. Effects of S1P on the action potential parameters of ventricular myocytes in guinea-pig. Current patch clamp technique was used to record the action potential of guinea pig isolated ventricular myocytes. S1P 10.0 μmol·L-1 reduced the membrane potential from (-68.5±7.65) mV to (-60.9±3.69) mV (p<0.05). Therate of reduction was 11.1%,which was blocked by PTX. S1P 10.0 μmol·L-1 reduced the APA from (105.8±9.6) mV to (98.4±8.2) mV and Vmax from (63.4±7.1) V/s to (55.9±6.8) V/s(p<0.05). Compared with control group, S1P 1.0 μmol·L-1 and S1P 10.0 μmol·L-1 could prolong the action potential duration in a dose-dependence manner. APD50 was prolonged from (296.2±47.0) ms to (343.2±49.4) ms and (357.3±52.6) ms (p<0.05) respectively. APD90 was prolonged from (326.1±46.8) ms to (381.3±49.1) ms and (399.2±48.7) ms (p<0.05) respectively. The effects of S1P prolonging the action potential duration were blocked by PTX. All the results suggested that the effects of S1P on membrane potential and action potential duration were via a EDG receptor-G protein-coupled receptors EDG pathway. Ⅲ. Effects of S1P on the calcium current of ventricular myocytes in guinea-pig. The whole cell patch clamp technique was used to record the calcium current of ventricular myocytes, which was from holding potential -50 mV stepping to +70 mV. The calcium current of 32 ventricular myocytes isolated from guinea-pigs were recorded. After application of S1P 1.0 μmol·L-1 and 10.0 μmol·L-1, the amplitude of calcium current was enhanced from (0.256±0.03) mV and (0.242±0.03) mV to (0.367±0.09) mV (p<0.05) and (0.364±0.06) mV (p<0.01) respectively. The percentage of enhancement compared with control group were 43.36% and 50.41% respectively which suggested that S1P could increase the calcium current in a dose-dependence manner. If pretreated with PTX 0.1 mg·L-1 or PKC antagonist Calphostin C 0.1 μmol·L-1, the effects of S1P increasing the calcium current were significantly blocked which suggested that S1P activating L-type calcium channel may be modulated by G-proteincoupled EDG receptor and PKC pathway. Ⅳ. Effects of S1P on the potassium current of ventricular myocytes in guinea-pig. The whole cell patch clamp technique was used to record the potassium current of ventricular myocytes, which was from holding potential -40 mV stepping to +80 mV. The potassium current of 43 ventricular myocytes isolated from guinea-pigs were recorded. After application of S1P 0.1 μmol·L-1, 1.0 μmol·L-1 and 10.0 μmol·L-1, the amplitude of potassium current was decreased from (1.444±0.142) nA ,(1.445±0.121) nA and (1.493±0.140) nA to (1.414±0.142) nA, (1.237±0.120) nA (p <0.01) and (1.175±0.119) nA (p <0.001) respectively. The percentage of decrease compared with control group was 2.08%, 14.39% and 21.30% respectively, which suggested that S1P could decrease the potassium current in a dose-dependence manner. If pretreated with PTX 0.1 mg·L-1or PKC antagonist Calphostin C 0.1 μmol·L-1, the effects of S1P decreasing the potassium were significantly blocked, which suggested that S1P inhibiting potassium channel may be modulated by EDG receptor-G protein-PKC pathway. Ⅴ. Effects of S1P on the sodium current of ventricular myocytes in guinea-pig. The whole cell patch clamp technique was used to record the sodium current of ventricular myocytes, which was from holding potential -80 mV stepping to +70 mV. The sodium current of 38 ventricular myocytes isolated from guinea-pigs was recorded. After application of S1P 0.1 μmol·L-1, 1.0 μmol·L-1 and 10.0 μmol·L-1, the amplitude of sodium current was decreased from (7.086±0.803) nA, (7.164±1.119) nA and (7.099±1.011) nA to...