Complex modulation of Ca v 3.1 T-type calcium channel by nickel
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Cellular and Molecular Life Sciences
Research Article
Complex modulation of Cav3.1 T-type calcium channel by nickel Olena V. Nosal · Olga P. Lyubanova · Valeri G. Naidenov · Yaroslav M. Shuba
Received: 24 June 2012 / Revised: 20 November 2012 / Accepted: 27 November 2012 © Springer Basel 2012
Abstract 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 Cav3.2 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 nonpermeating 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” Cav3.1w/t channel and Cav3.1Q172H mutant having glutamine (Q) equivalent to H191 of Cav3.2 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 Cav3.1w/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 voltagedependent, 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 O. V. Nosal · O. P. Lyubanova · Y. M. Shuba International Center of Molecular Physiology of the National Academy of Sciences of Ukraine, Kiev, Ukraine V. G. Naidenov · Y. M. Shuba (*) Bogomoletz Institute of Physiology of the National Academy of Sciences of Ukraine, Bogomoletz Str., 4, Kiev 01024, Ukraine e-mail: [email protected] O. V. Nosal · O. P. Lyubanova · Y. M. Shuba State Key Laboratory of Molecular and Cellular Physiology, Kiev, Ukraine
sites in the Cav3.1w/t and Cav3.1Q172H 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. Keywords T-type calcium channels · Cav3.1 · Xenopus oocytes · Ni2+
Introduction Low-voltage-activated (LVA or T-type) calcium channels constitute a separate group of plasma membrane voltagegated calcium channels (VGCC) with biophysical properties enabling them to pass transient inward calcium current in response to low levels of depolarization (activation threshold around −60 mV) taking place from the relatively hyperpolarized cell’s resting membrane potential (Vr below −60 mV), which is required for the channels’ recuperation from steady-state in
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