This study was designed to evaluate the effects of the Na+/Ca2+ exchange (NCX) inhibitor SEA0400 on Ca2+ handling in isolated canine ventricular myocytes.Methods and results
Intracellular Ca2+ ([Ca2+]i) transients, induced by either field stimulation or caffeine flush, were monitored using Ca2+ indicator dyes. [Ca2+]i-dependent modulation of the inhibitory effect of SEA0400 on NCX was characterized by the changes in Ni2+-sensitive current in voltage-clamped myocytes. Sarcoplasmic reticulum (SR) Ca2+ release and uptake were studied in SR membrane vesicles. Gating properties of single-ryanodine receptors were analysed in lipid bilayers. Ca2+ sensitivity of the contractile machinery was evaluated in chemically skinned myocytes. In myocytes paced at 1 Hz, neither diastolic [Ca2+]i nor the amplitude of [Ca2+]i transients was significantly altered by SEA0400 up to the concentration of 1 µM, which was shown to inhibit the exchange current. The blocking effect of SEA0400 on NCX decreased with increasing [Ca2+]i, and it was more pronounced in reverse than in forward mode operation at every [Ca2+]i examined. The rate of decay of the caffeine-induced [Ca2+]i transients was decreased significantly by 1 µM SEA0400; however, this effect was only a fraction of that observed with 10 mM NiCl2. Neither SR Ca2+ release and uptake nor cell shortening and Ca2+ sensitivity of the contractile proteins were influenced by SEA0400.Conclusion
The lack of any major SEA0400-induced shift in Ca2+ transients or contractility of myocytes can well be explained by its limited inhibitory effect on NCX (further attenuated by elevated [Ca2+]i levels) and a concomitant reduction in Ca2+ influx due to the predominantly reverse mode blockade of NCX and suppression of L-type Ca2+ current.