Positive Inotropic Effect On Myocardium Of Enhancing Na～+/Ca～(2+) Exchange And Protective Effect Of Inhibiting Na～+/Ca～(2+) Exchange On Cardiac Ischemia/Reperfusion Injury In Rats

The cardiac Na+/Ca2+ exchange (NCE) is a double-edged sword. On one hand, the NCE elicits Ca2+ influx through the reverse mode and triggers Ca2+ release from the sarcoplasmic reticulum (SR) to mediate the myocardium contraction. The forward mode of NCE extrudes Ca2+ during aftercontraction and helps the cardiomycytes to diastole. On the other hand, the NCE plays a crucial role in myocardium damage during cardiac ischemia/reperfusion injury. With regard to make good use of the cardiac NCE, we supposed to investigate the positive inotropic effect of a Class III antiarrhythmic agent, dofetilide, by activating the NCE in isolated rat cardiomyocytes and study the protective effect of a NCE inhibitor, KB-R7943 in Langendorff perfused rat heartssuffering cardiac ischemia/reperfusion injury.Whole-cell patch clamp experiment results showed that dofetilide increased INa/ca in a concentration-dependent manner from 0.03 to 1.0 mol/L on both inward and outward transport direction in freshly isolated rat or guinea pig ventricular cells. The EC 50 of outward INa/ca in rat ventricular myocytes was 0.183 fimol/L (95% confidence interval(CI) was 0.058- 0.520 mol/L), and the EC50 of inward Ixa/ca in rat ventricular cells was 0.178 umol/L (95% CI was 0.024-1.296 umol/L). The EC50 of outward and inward INa/ca in guinea pig ventricular myocytes was 0.178 Hmol/L(95% CI was 0.040-0.787 umol/L), and 0.178 umol/L (95% CI was 0.038-0.842 umol/L), respectively. When loaded with Fura-2, the Ca2+ transient and cell shortening images were real-time recorded using a TILL Photonics Imaging System. Results showed that when stimulated by field electrical stimulation, 0.2 mol/L dofetilide significantly enhanced Ca2+ transient and cellshortening in rat ventricular myocytes by 57.24 20.67 (P<0.01) and 3.56 1.24 um(P<0.01), respectively. These effects of dofetilide were inhibited by 0.6u.mol/L KB-R7943, a selective NCE inhibitor, markedly. The calcium sensitivity calculated from cell length devided by Ca2+ transient (cell shortening/(F340/F380)) increased greatly from 0.030 0.014 to 0.036 0.016 (P<0.01) when 0.2 mol/L dofetilide was added to the Tyrode's solution. KB-R7943 at 0.6 mol/L had no significant influence on calcium sensitivity. Dofetilide at 0.2 u,mol/L also shortened the Ca2+ transient duration and myocyte diastolic duration. R75% and R90% was depressed 242.17 77.78 ms (P<0.01) and 385.10 156.33 ms (P<0.05), respectively. The Tdia decreased from 619.00 179.79 ms of control to 289. 17 1 14.04 ms after 0.2 jimol/L dofetilide administration. 0.6 umol/L KB-R7943 significantly prolonged Tdia to 472. 50 1 34.50 ms (P<0.05). When connected the patch clamp system to the Tillvision Ionic Imaging system through the A/D converter of Digidata1322A, the calcium current (ICa) , calcium transient and cell shortening were recorded simultaneously. Nicardipine at 1 umol/L abolished ICa and depressed Ca2+ transient and cell shortening greatly. Dofetilide at 0.2 umol/L had no active effect on Icabut enlarged Ca2+ transient and cell shortening significantly by 87.26 38.10 (P<0.01) and 2.10 0.55 urn (P<0.01), respectively. 0.6umol/L KB-R7943 abolished theeffects of dofetilide on Ca transient and cell shortening without any influence on ICa.The inhibition of KB-R7943 on iNa/ca was examined with whole-cell patch clamp technique. The results showed that KB-R7943 at 0.6 umol/L inhibited NCE equally in both forward and reverse transport modes. A Langendorff perfusion method was used to study the protective effect of KB-R7943 on cardiac regional ischemia/reperfusion injury by inhibiting NCE. When the isolated rat heart was equilibrated with Tyrode's solution for 1 h, the left anterior descending (LAD) coronary artery was ligated for 30 min to produce regional ischemia. After that, the LAD wasreleased and the heart was reperfused for 120 min. When administered during the early period of the reperfusion from Imin before the start of reperfusion to 14 min after LAD release, KB-R7943 at 1 umol/L reduced the depression of left ventricular func...