Ni2+ block of Cav3.1 (α1G) T-type calcium channels

Abstract

Ni2+ inhibits current through calcium channels, in part by blocking the pore, but Ni2+ may also allosterically affect channel activity via sites outside the permeation pathway. As a test for pore blockade, we examined whether the effect of Ni2+ on Cav3.1 is affected by permeant ions. We find two components to block by Ni2+, a rapid block with little voltage dependence, and a slow block most visible as accelerated tail currents. Rapid block is weaker for outward vs. inward currents (apparent Kd = 3 vs. 1 mM Ni2+, with 2 mM Ca2+ or Ba2+) and is reduced at high permeant ion concentration (110 vs. 2 mM Ca2+ or Ba2+). Slow block depends both on the concentration and on the identity of the permeant ion (Ca2+vs. Ba2+ vs. Na+). Slow block is 2-3× faster in Ba 2+ than in Ca2+ (2 or 110 mM), and is ∼10× faster with 2 vs. 110 mM Ca2+ or Ba2+. Slow block is orders of magnitude slower than the diffusion limit, except in the nominal absence of divalent cations (∼3 μM Ca2+). We conclude that both fast and slow block of Cav3.1 by Ni2+ are most consistent with occlusion of the pore. The exit rate of Ni2+ for slow block is reduced at high Ni2+ concentrations, suggesting that the site responsible for fast block can "lock in" slow block by Ni 2+, at a site located deeper within the pore. In contrast to the complex pore block observed for Cav3.1, inhibition of Ca v3.2 by Ni2+ was essentially independent of voltage, and was similar in 2 mM Ca2+ vs. Ba2+, consistent with inhibition by a different mechanism, at a site outside the pore. ©2008 Hirn et al.