Halothane inhibition of recombinant cardiac L-type Ca²⁺ channels expressed in HEK-293 cells

Background: Volatile anesthetics depress cardiac contractility, which involves inhibition of cardiac L-type calcium channels. To explore the role of voltage-dependent inactivation, the authors analyzed halothane effects on recombinant cardiac L-type calcium channels (α_1Cβ _2a and α_1Cβ_2aα₂/ δ₁), which differ by the α₂/δ ₁ subunit and consequently voltage-dependent inactivation. Methods: HEK-293 cells were transiently cotransfected with complementary DNAs encoding alc tagged with green fluorescent protein and β_2a, with and without α₂/δ₁. Halothane effects on macroscopic barium currents were recorded using patch clamp methodology from cells expressing α_1Cβ_2a and α_1Cβ_2aα₂/δ₁ as identified by fluorescence microscopy. Results: Halothane inhibited peak current (I_peak) and enhanced apparent inactivation (reported by end pulse current amplitude of 300-ms depolarizations [I₃₀₀]) in a concentration-dependent manner in both channel types. α₂/ δ₁ coexpression shifted relations leftward as reported by the 50% inhibitory concentration of I_peak and I₃₀₀/I _peak for α_1Cβ_2a (1.8 and 14.5 mM, respectively) and α_1Cβ_2aα₂/ δ₁ (0.74 and 1.36 mM, respectively). Halothane reduced transmembrane charge transfer primarily through I_peak depression and not by enhancement of macroscopic inactivation for both channels. Conclusions: The results indicate that phenotypic features arising from α₂/ δ₁ coexpression play a key role in halothane inhibition of cardiac L-type calcium channels. These features included marked effects on I_peak inhibition, which is the principal determinant of charge transfer reductions. I_peak depression arises primarily from transitions to nonactivatable states at resting membrane potentials. The findings point to the importance of halothane interactions with states present at resting membrane potential and discount the role of inactivation apparent in current time courses in determining transmembrane charge transfer.