We compared detailed efficacy of efonidipine and nifedipine, dihydropyridine analogues, and mibefradil using recombinant T- and L-type Ca²⁺ channels expressed separately in mammalian cells. All these Ca²⁺ channel antagonists blocked T-type Ca²⁺ channel currents (I_Ca(T)) with distinct blocking manners: I_Ca(T) was blocked mainly by a tonic manner by nifedipine, by a use-dependent manner by mibefradil, and by a combination of both manners by efonidipine. IC₅₀s of these Ca²⁺ channel antagonists to I_Ca(T) and L-type Ca²⁺ channel current (I_Ca(L)) were 1.2 µmol/l and 0.14 nmol/l for nifedipine; 0.87 and 1.4 µmol/l for mibefradil, and 0.35 µmol/l and 1.8 nmol/l for efonidipine, respectively. Efonidipine, a dihydropyridine analogue, showed high affinity to T-type Ca²⁺ channel.

Effects of bepridil on the low voltage-activated T-type Ca²⁺ channel (Ca_V3.2) current stably expressed in human embryonic kidney (HEK)-293 cells were examined using patch-clamp techniques. Bepridil potently inhibited I_Ca,T with a markedly voltage-dependent manner; the IC₅₀ of bepridil was 0.4 µmol/l at the holding potential of –70 mV, which was 26 times as potent as that at –100 mV (10.6 µmol/l). Steady-state inactivation curve (8.4 ± 1.7 mV) and conductance curve (5.9 ± 1.9 mV) were shifted to the hyperpolarized potential by 10 µmol/l bepridil. Bepridil exerted the tonic blocking action but not the use-dependent block. Bepridil had no effect on the recovery from inactivation of T-type Ca²⁺ channels. Thus, high efficacy of bepridil for terminating atrial fibrillation and atrial flutter may be considered to be attributed, at least in a part, to the T-type Ca²⁺ channel-blocking actions.

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The L-type Ca2+ channel has a unique kinetic property known as voltage-dependent facilitation. Many researchers have repeatedly investigated the mechanism in response to the voltage-dependent facilitation since the first observation by Fenwick et al. in 1982. Electrophysiological evaluations of voltage-dependent facilitation, however, remain inconsistent, partially because of its unclear definition. Some scientists understand it as a current augmentation by a conditioning prepulse prior to the test pulse, and others understand it as a result of the U-shape steady-state inactivation curve. We therefore investigated to identify the distinction between the voltage-dependent facilitation and the steady-state inactivation, by use of Ba2+ as the charge in order to avoid the other inactivation mechanism or the Ca(2+)-dependent inactivation upon this analysis. Conventional whole-cell mode patch clamp technique was applied to chinese hamster fibroblast (CHW) cells that express the alpha1c subunit alone or the alpha1c subunit with the beta subunit (alpha1c/beta) derived from rabbit heart to investigate the voltage-dependent facilitation depending on the composition of the subunits. Coexpression of the beta subunit augmented alpha1 subunit channel current and shifted current-voltage relation towards hyperpolarized direction. In the experiment using conventional double pulse protocol to investigate steady-state inactivation, alpha1c subunit channel current and alpha1c/beta subunit channel current were not fully inactivated. Subtraction of the steady-state inactivation component from whole recovered current enabled us to identify the voltage-dependent facilitation component of the L-type Ca2+ channel. The voltage-dependent facilitation of the alpha1 subunit current and the alpha1c/beta subunit current were identical in kinetics, and could be generated at 0 mV or depolarized potentials partially overlapped with the potential range for the steady-state inactivation of the current. These results suggest that the voltage-dependent facilitation of the L-type Ca2+ channel could be formed by the alpha1c subunit without interaction with the beta subunit, and that the range for the voltage-dependent facilitation and the steady-state inactivation overlap each other at 0 mV or more depolarized potentials up to approximately + 100 mV.