Effects Of Levosimendan On Calcium Transient In Norepinephrine-cultured Neonatal Rat Ventricular Myocytes

Objective To explore the alterations of the calcium transient in the development of heart failure,and the impact of levosimendan on intracellular calcium dynamics.Methods The cultured neonatal rat ventricular myocytes were divided into 4 groups,normal,norepinephrine(NE)alone(10 umol/L),NE+ levosimendan1(0.1 umol/L),NE+levosimendan2(1umol/L).Calcium transients of myocytes loaded by Fluo-3/AM were observed under Laser scanning confocal microscope,the L-type Ca2+ channel currents were detected with the whole cell patch clamp,Western blots were used to measure the expression of SERCA2 and NCX1.Results(1)Compared with control,calcium wave dispersed,conducted slowly,and exhibited dyssynchrony of Ca2+ release in NE group,NE accelerated the cultured myocytes beating rate(26.7±4.3 vs.11.6±3.6,P<0.01),decreased peak systolic Ca2+(128.37±65.44 vs.155.33±61.77,P<0.05),increased time to peak(Ttp)(0.413±0.324 vs.0.212±0.050,P<0.01)and decay time(1.162±0.524 vs.0.722±0.169,P<0.01)of calcium transient.(2)Levosimendan increased synchrony of calcium transient,reduced Ttp(0.212±0.044 vs.0.413±0.324,NE+LeV1 vs.NE,P<0.01 and 0.205±0.062 vs.0.413±0.324,NE +LeV2 vs.NE,P<0.01,respectively)and decay time(0.735±0.269 vs.1.162±0.524,NE +LeV1 vs.NE,P<0.01 and 0.753±0.152 vs.1.162±0.524,NE +LeV2 vs.NE,P<0.01),had no effect on myocytes beating rate,peak systolic or resting Ca2+ and transient amplitude.(3)There were no concentration differences in the presence of levosimendan.(4)No differences of the L-type calcium current were found among the groups.Both NE and levosimendan shortened the duration of the action potention(APD),the latter was more serious(187.65±11.53 vs.245.40±18.92,NE vs.control,P<0.01),(7.62±0.72 ?8.5±1.11 vs.187.65±11.53,NE+LeV1? NE+LeV2 vs.NE,P<0.01)(5)Western blots showed that the expression of SERCA2 decreased in the NE group,while increased in the presence of levosimendan.The levels of NCX1 displayed the opposite results.Conclusion Our results showed that long-time exposure to NE decreased myocardial contractility by reducing the amplitude of calcium transient,shortening and disrupting the conduction of calcium wave.Levosimendan have no influence on the intracellular calcium homestasis,however,it can exert positive inotropic effect by accelerating the velocity of calcium signaling propagation and synchronzing the calcium release,without increasing the L-type calcium current and intracelluar calcium concentration.Levosimendan significantly shortened APD,improved calcium channel proteins expression and myocardial remodeling.