Evaluation of a Two-Site, Three-Barrier Model for Permeation in Ca_V3.1 (α1G) T-Type Calcium Channels: Ca²⁺, Ba²⁺, Mg²⁺, and Na⁺

Abstract

We explored the ability of a two-site, three-barrier (2S3B) Eyring model to describe recently reported data on current flow through open Ca_V3.1 T-type calcium channels, varying Ca²⁺ and Ba²⁺ over a wide range (100 nm–110 mm) while recording whole-cell currents over a wide voltage range (−150 mV to +100 mV) from channels stably expressed in HEK 293 cells. Effects on permeation were isolated using instantaneous current–voltage relationships (IIV) after strong, brief depolarizations to activate channels with minimal inactivation. Most experimental results were reproduced by a 2S3B model. The model described the IIV relationships, apparent affinities for permeation and block for Ca²⁺ and Ba²⁺, and shifts in reversal potential between Ca²⁺ and Ba²⁺. The fit to block by 1 mm ${\text{Mg}}^{2+}_{\text{i}}$ was reasonable, but block by ${\text{Mg}}^{2+}_{\text{o}}$ was described less well. Surprisingly, fits were comparable with strong ion–ion repulsion, with no repulsion, or with intermediate values. With weak repulsion, there was a single high-affinity site, with a low-affinity site near the cytoplasmic side of the pore. With strong repulsion, the net charge of ions in the pore was near +2 over a relatively wide range of concentration and voltage, suggesting a knockoff mechanism. With strong repulsion, Ba²⁺ preferred the inner site, while Ca²⁺ preferred the outer site, potentially explaining faster entry of Ni²⁺ and other pore blockers when Ba²⁺ is the charge carrier.