Complex modulation of Cav3.1 T-type calcium channel by nickel

Nickel is considered to be a selective blocker of low-voltage-activated T-type calcium channel. Recently, the Ni²⁺-binding site with critical histidine-191 (H191) within the extracellular IS3–IS4 domain of the most Ni²⁺-sensitive Ca_v3.2 T-channel isoform has been identified. All calcium channels are postulated to also have intrapore-binding site limiting maximal current carried by permeating divalent cations (PDC) and determining the blockade by non-permeating ones. However, the contribution of the two sites to the overall Ni²⁺ effect and its dependence on PDC remain uncertain. Here we compared Ni²⁺ action on the wild-type “Ni²⁺-insensitive” Ca_v3.1_w/t channel and Ca_v3.1_Q172H mutant having glutamine (Q) equivalent to H191 of Ca_v3.2 replaced by histidine. Each channel was expressed in Xenopus oocytes, and Ni²⁺ blockade of Ca²⁺, Sr²⁺, or Ba²⁺ currents was assessed by electrophysiology. Inhibition of Ca_v3.1_w/t by Ni²⁺ conformed to two sites binding. Ni²⁺ binding with high-affinity site (IC₅₀ = 0.03–3 μM depending on PDC) produced maximal inhibition of 20–30 % and was voltage-dependent, consistent with its location within the channel’s pore. Most of the inhibition (70–80 %) was produced by Ni²⁺ binding with low-affinity site (IC₅₀ = 240–700 μM). Q172H-mutation mainly affected low-affinity binding (IC₅₀ = 120–160 μM). The IC₅₀ of Ni²⁺ binding with both sites in the Ca_v3.1_w/t and Ca_v3.1_Q172H was differentially modulated by PDC, suggesting a varying degree of competition of Ca²⁺, Sr²⁺, or Ba²⁺ with Ni²⁺. We conclude that differential Ni²⁺-sensitivity of T-channel subtypes is determined only by H-containing external binding sites, which, in the absence of Ni²⁺, may be occupied by PDC, influencing in turn the channel’s permeation.