Evaluation of a two-site, three-barrier model for permeation in Ca(V)3.1 (alpha1G) T-type calcium channels: Ca (2+), Ba (2+), Mg (2+), and Na (+).
We explored the ability of a two-site, three-barrier (2S3B) Eyring model to describe recently reported data on current flow through open Ca(V)3.1 T-type calcium channels, varying Ca(2+) and Ba(2+) 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 Ca(2+) and Ba(2+), and shifts in reversal potential between Ca(2+) and Ba(2+). The fit to block by 1 mM Mg(2+)(i) was reasonable, but block by Mg(2+)(0) 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, Ba(2+) preferred the inner site, while Ca(2+) preferred the outer site, potentially explaining faster entry of Ni(2+) and other pore blockers when Ba(2+) is the charge carrier.
Department of Physiology and Biophysics, Case Western Reserve University, Cleveland, OH 44106, USA.
Kyle V
KV
Lopin
Carlos A
CA
Obejero-Paz
Stephen W
SW
Jones
D001464:Barium / Q000378:metabolism*; D002118:Calcium / Q000378:metabolism*; D020747:Calcium Channels, T-Type / Q000378:metabolism*; D002460:Cell Line; D006801:Humans; D008274:Magnesium / Q000378:metabolism*; D008954:Models, Biological; D012964:Sodium / Q000378:metabolism*
C500994:Cacna1g protein, rat; D020747:Calcium Channels, T-Type; D001464:Barium; D012964:Sodium; D008274:Magnesium; D002118:Calcium