Two types of voltage-dependent Ca2+ channels have been identified in heart: high (ICaL) and low (ICaT) voltage-activated Ca2+ channels. In guinea pig ventricular myocytes, low voltage-activated inward current consists of ICaT and a tetrodotoxin (TTX)-sensitive ICa component (ICa(TTX)). In this study, we reexamined the nature of low-threshold ICa in dog atrium, as well as whether it is affected by Na+ channel toxins. Ca2+ currents were recorded using the whole-cell patch clamp technique. In the absence of external Na+, a transient inward current activated near -50 mV, peaked at -30 mV, and reversed around +40 mV (HP = -90 mV). It was unaffected by 30 μM TTX or micromolar concentrations of external Na+, but was inhibited by 50 μM Ni2+ (by ∼90%) or 5 μM mibefradil (by ∼50%), consistent with the reported properties of ICaT. Addition of 30 μM TTX in the presence of Ni2+ increased the current approximately fourfold (41% of control), and shifted the dose-response curve of Ni2+ block to the right (IC50 from 7.6 to 30 μM). Saxitoxin (STX) at 1 μM abolished the current left in 50 μM Ni2+. In the absence of Ni2+, STX potently blocked ICaT (EC50 = 185 nM) and modestly reduced ICaL (EC50 = 1.6 μM). While TTX produced no direct effect on I CaT elicited by expression of hCaV3.1 and hCa V3.2 in HEK-293 cells, it significantly attenuated the block of this current by Ni2+ (IC50 increased to 550 μM Ni 2+ for CaV3.1 and 15 μM Ni2+ for Ca V3.2); in contrast, 30 μM TTX directly inhibited hCa V3.3-induced ICaT and the addition of 750 μM Ni 2+ to the TTX-containing medium led to greater block of the current that was not significantly different than that produced by Ni2+ alone. 1 μM STX directly inhibited CaV3.1-, CaV3.2-, and CaV3.3-mediated ICaT but did not enhance the ability of Ni2+ to block these currents. These findings provide important new implications for our understanding of structure-function relationships of ICaT in heart, and further extend the hypothesis of a parallel evolution of Na+ and Ca2+ channels from an ancestor with common structural motifs.
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