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Information × Registration Number 0216U007301, 0110U006269 , R & D reports Title Ion channels and calcium homeostasis as the targets of modulatory influence of endo- and exogenous biological active substances. popup.stage_title Head Shuba Yaroslav M., Доктор біологічних наук Registration Date 04-07-2016 Organization International center of molecular physiology popup.description2 Nickel is considered as selective blocker of low voltage-activated T-type calcium channels. 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 have also 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" 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 ?M 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 ?M). Q172H-mutation mainly affected low-affinity binding (IC50=120-160 ?M). The IC50 of Ni2+ binding with both sites in the Cav3.1w/t and Cav3.1Q172H was differentially modulated by PDC, suggesting various 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 site, which in the absence of Ni2+ may be occupied by PDC, influencing in turn channel's permeation. Product Description popup.authors Любанова О.П. Носаль О.В. Шуба Я.М. popup.nrat_date 2020-04-02 Close