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To calibrate acid\u2010induced intracelluar [Zn<jats:sup>2+<\/jats:sup>] ([Zn<jats:sup>2+<\/jats:sup>]<jats:sub>i<\/jats:sub>) elevations, a low affinity (<jats:italic>K<\/jats:italic><jats:sub>d<\/jats:sub>\u00a0=\u00a039\u00a0\u03bcM at <jats:styled-content style=\"fixed-case\">pH<\/jats:styled-content> 6.1) ratiometric Zn<jats:sup>2+<\/jats:sup> probe, FuraZin\u20101, was used. A <jats:styled-content style=\"fixed-case\">pH<\/jats:styled-content><jats:sub>i<\/jats:sub> drop from 7.2 to 6.1 caused [Zn<jats:sup>2+<\/jats:sup>]<jats:sub>i<\/jats:sub> elevations reaching 2\u00a0\u03bcM; when the thiol\u2010reactive agent <jats:italic>N<\/jats:italic>\u2010ethylmaleimide (<jats:styled-content style=\"fixed-case\">NEM<\/jats:styled-content>) was subsequently applied, [Zn<jats:sup>2+<\/jats:sup>]<jats:sub>i<\/jats:sub> increased further to 5.6\u00a0\u03bcM; analogous acid\u2010 and <jats:styled-content style=\"fixed-case\">NEM<\/jats:styled-content>\u2010induced [Zn<jats:sup>2+<\/jats:sup>]<jats:sub>i<\/jats:sub> elevations could also be detected but not calibrated, using the high affinity Zn<jats:sup>2+<\/jats:sup> probe FluoZin\u20103. The data indicate that <jats:styled-content style=\"fixed-case\">NEM<\/jats:styled-content> causes Zn<jats:sup>2+<\/jats:sup> release from ligands that chelate Zn<jats:sup>2+<\/jats:sup> at <jats:styled-content style=\"fixed-case\">pH<\/jats:styled-content> 6.1. <jats:styled-content style=\"fixed-case\">ATP<\/jats:styled-content> could also chelate Zn<jats:sup>2+<\/jats:sup> at <jats:styled-content style=\"fixed-case\">pH<\/jats:styled-content> 6.1 because its <jats:styled-content style=\"fixed-case\">pK<\/jats:styled-content><jats:sub>a<\/jats:sub> is about 6.8. Therefore, it was tested whether an <jats:styled-content style=\"fixed-case\">ATP<\/jats:styled-content> depletion affects the acid\u2010induced [Zn<jats:sup>2+<\/jats:sup>]<jats:sub>i<\/jats:sub> elevations. The <jats:styled-content style=\"fixed-case\">ATP<\/jats:styled-content> depletion was induced by inhibiting mitochondrial and glycolytic <jats:styled-content style=\"fixed-case\">ATP<\/jats:styled-content> production. Interestingly, an almost complete <jats:styled-content style=\"fixed-case\">ATP<\/jats:styled-content> depletion (confirmed using a luciferin\/luciferase assay) failed to affect the acid\u2010induced [Zn<jats:sup>2+<\/jats:sup>]<jats:sub>i<\/jats:sub> increases. These data suggest that the total amount of Zn<jats:sup>2+<\/jats:sup> accumulated in intracellular <jats:styled-content style=\"fixed-case\">ATP<\/jats:styled-content>\u2010dependent stores (Zn<jats:sup>2+<\/jats:sup>\u2010<jats:styled-content style=\"fixed-case\">ATP<\/jats:styled-content> complexes and organelles that accumulate Zn<jats:sup>2+<\/jats:sup> in an <jats:styled-content style=\"fixed-case\">ATP<\/jats:styled-content>\u2010dependent manner) is negligible compared to the amount of Zn<jats:sup>2+<\/jats:sup> accumulated in the acid\u2010sensitive intracellular ligands.<\/jats:p><\/jats:sec><jats:sec><jats:label \/><jats:p>\n<jats:boxed-text content-type=\"graphic\" position=\"anchor\"><jats:graphic xmlns:xlink=\"http:\/\/www.w3.org\/1999\/xlink\" mimetype=\"image\/png\" position=\"anchor\" specific-use=\"enlarged-web-image\" xlink:href=\"graphic\/jnc13282-fig-0007-m.png\"><jats:alt-text>image<\/jats:alt-text><\/jats:graphic><\/jats:boxed-text>\n<jats:italic>In vitro,<\/jats:italic> upon acidification, Zn<jats:sup>2+<\/jats:sup>\u2010cysteine complexes release Zn<jats:sup>2+<\/jats:sup> and ATP chelates the released Zn<jats:sup>2+<\/jats:sup>. However, <jats:italic>in\u00a0vivo<\/jats:italic> (cultured neurons), an ATP depletion failed to enhance acid\u2010induced [Zn<jats:sup>2+<\/jats:sup>]<jats:sub>i<\/jats:sub> elevations. These [Zn<jats:sup>2+<\/jats:sup>]<jats:sub>i<\/jats:sub> elevations were calibrated using a low affinity ratiometric probe FuraZin\u20101; they reached 2\u00a0\u00b5M levels and increased to 5\u00a0\u00b5M when a thiol\u2010reactive agent, <jats:italic>N<\/jats:italic>\u2010ethylmaleimide, compromised Zn<jats:sup>2+<\/jats:sup> binding by cysteines.<\/jats:p><\/jats:sec>","DOI":"10.1111\/jnc.13282","type":"journal-article","created":{"date-parts":[[2015,8,11]],"date-time":"2015-08-11T15:38:56Z","timestamp":1439307536000},"page":"777-786","source":"Crossref","is-referenced-by-count":3,"title":["Neuronal acid\u2010induced [Zn<sup>2+<\/sup>]<sub>i<\/sub> elevations calibrated using the low\u2010affinity ratiometric probe FuraZin\u20101"],"prefix":"10.1111","volume":"135","author":[{"given":"Lech","family":"Kiedrowski","sequence":"first","affiliation":[{"name":"The Psychiatric Institute Departments of Psychiatry and Pharmacology The University of Illinois at Chicago Chicago Illinois USA"}],"role":[{"role":"author","vocabulary":"crossref"}]}],"member":"311","published-online":{"date-parts":[[2015,9,10]]},"reference":[{"key":"e_1_2_7_2_1","doi-asserted-by":"publisher","DOI":"10.1046\/j.1471-4159.2000.0751878.x"},{"key":"e_1_2_7_3_1","doi-asserted-by":"publisher","DOI":"10.1016\/S0079-6107(96)00013-2"},{"key":"e_1_2_7_4_1","doi-asserted-by":"publisher","DOI":"10.1016\/S0006-3495(91)82034-3"},{"key":"e_1_2_7_5_1","doi-asserted-by":"publisher","DOI":"10.1371\/journal.pone.0125185"},{"key":"e_1_2_7_6_1","doi-asserted-by":"publisher","DOI":"10.1016\/0167-4838(89)90245-8"},{"key":"e_1_2_7_7_1","doi-asserted-by":"crossref","first-page":"8281","DOI":"10.1016\/S0021-9258(18)83180-5","article-title":"Characterization of a H+\u2010ATPase in rat brain synaptic vesicles. 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