{"status":"ok","message-type":"work","message-version":"1.0.0","message":{"indexed":{"date-parts":[[2026,7,2]],"date-time":"2026-07-02T07:51:24Z","timestamp":1782978684039,"version":"3.54.5"},"reference-count":34,"publisher":"Wiley","issue":"24","license":[{"start":{"date-parts":[[2021,9,26]],"date-time":"2021-09-26T00:00:00Z","timestamp":1632614400000},"content-version":"vor","delay-in-days":0,"URL":"http:\/\/onlinelibrary.wiley.com\/termsAndConditions#vor"}],"content-domain":{"domain":["bpspubs.onlinelibrary.wiley.com"],"crossmark-restriction":true},"short-container-title":["British J Pharmacology"],"published-print":{"date-parts":[[2021,12]]},"abstract":"<jats:sec><jats:title>Background and Purpose<\/jats:title><jats:p>In addition to binding to the classical high\u2010affinity extracellular benzodiazepine binding site of the GABA<jats:sub>A<\/jats:sub> receptor, some benzodiazepines occupy transmembrane inter\u2010subunit anaesthetic sites that bind etomidate (\u03b2<jats:sup>+<\/jats:sup>\/\u03b1<jats:sup>\u2212<\/jats:sup> sites) or the barbiturate derivative R\u2010<jats:italic>m<\/jats:italic>TFD\u2010MPAB (\u03b1<jats:sup>+<\/jats:sup>\/\u03b2<jats:sup>\u2212<\/jats:sup> and \u03b3<jats:sup>+<\/jats:sup>\/\u03b2<jats:sup>\u2212<\/jats:sup> sites). We aimed to define the functional effects of these interactions on GABA<jats:sub>A<\/jats:sub> receptor activity and animal behaviour.<\/jats:p><\/jats:sec><jats:sec><jats:title>Experimental Approach<\/jats:title><jats:p>With flumazenil blocking classical high\u2010affinity extracellular benzodiazepine site effects, modulation of GABA\u2010activated currents by diazepam, midazolam and flurazepam was measured electrophysiologically in wildtype and M2\u201015\u2032 mutant \u03b1<jats:sub>1<\/jats:sub>\u03b2<jats:sub>3<\/jats:sub>\u03b3<jats:sub>2L<\/jats:sub> GABA<jats:sub>A<\/jats:sub> receptors. Zebrafish locomotive activity was also assessed in the presence of each benzodiazepine plus flumazenil.<\/jats:p><\/jats:sec><jats:sec><jats:title>Key Results<\/jats:title><jats:p>In the presence of flumazenil, micromolar concentrations of diazepam and midazolam both potentiated and inhibited wildtype GABA<jats:sub>A<\/jats:sub> receptor currents. \u03b2<jats:sub>3<\/jats:sub>N265M (M2\u201015\u2032 in the \u03b2<jats:sup>+<\/jats:sup>\/\u03b1<jats:sup>\u2212<\/jats:sup> sites) and \u03b1<jats:sub>1<\/jats:sub>S270I (M2\u201015\u2032 in the \u03b1<jats:sup>+<\/jats:sup>\/\u03b2<jats:sup>\u2212<\/jats:sup> site) mutations reduced or abolished potentiation by these drugs. In contrast, the \u03b3<jats:sub>2<\/jats:sub>S280W mutation (M2\u201015\u2032 in the \u03b3<jats:sup>+<\/jats:sup>\/\u03b2<jats:sup>\u2212<\/jats:sup> site) abolished inhibition. Flurazepam plus flumazenil only inhibited wildtype receptor currents, an effect unaltered by M2\u201015\u2032 mutations. In the presence of flumazenil, zebrafish locomotion was enhanced by diazepam at concentrations up to 30\u2009\u03bcM and suppressed at 100\u2009\u03bcM, suppressed by midazolam and enhanced by flurazepam.<\/jats:p><\/jats:sec><jats:sec><jats:title>Conclusions and Implications<\/jats:title><jats:p>Benzodiazepine binding to transmembrane anaesthetic binding sites of the GABA<jats:sub>A<\/jats:sub> receptor can produce positive or negative modulation manifesting as decreases or increases in locomotion, respectively. Selectivity for these sites may contribute to the distinct GABA<jats:sub>A<\/jats:sub> receptor and behavioural actions of different benzodiazepines, particularly at high (i.e. anaesthetic) concentrations.<\/jats:p><\/jats:sec>","DOI":"10.1111\/bph.15662","type":"journal-article","created":{"date-parts":[[2021,8,13]],"date-time":"2021-08-13T03:30:12Z","timestamp":1628825412000},"page":"4842-4858","update-policy":"https:\/\/doi.org\/10.1002\/crossmark_policy","source":"Crossref","is-referenced-by-count":11,"title":["Selective actions of benzodiazepines at the transmembrane anaesthetic binding sites of the GABA<sub>A<\/sub> receptor: <i>In vitro<\/i> and <i>in vivo<\/i> studies"],"prefix":"10.1111","volume":"178","author":[{"given":"Megan","family":"McGrath","sequence":"first","affiliation":[{"name":"Department of Anesthesia, Critical Care, and Pain Medicine Massachusetts General Hospital  Boston Massachusetts USA"}],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"Helen","family":"Hoyt","sequence":"additional","affiliation":[{"name":"Department of Anesthesia, Critical Care, and Pain Medicine Massachusetts General Hospital  Boston Massachusetts USA"}],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"Andrea","family":"Pence","sequence":"additional","affiliation":[{"name":"Department of Anesthesia, Critical Care, and Pain Medicine Massachusetts General Hospital  Boston Massachusetts USA"}],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"Stuart A.","family":"Forman","sequence":"additional","affiliation":[{"name":"Department of Anesthesia, Critical Care, and Pain Medicine Massachusetts General Hospital  Boston Massachusetts USA"}],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"Douglas E.","family":"Raines","sequence":"additional","affiliation":[{"name":"Department of Anesthesia, Critical Care, and Pain Medicine Massachusetts General Hospital  Boston Massachusetts USA"}],"role":[{"vocabulary":"crossref","role":"author"}]}],"member":"311","published-online":{"date-parts":[[2021,9,26]]},"reference":[{"key":"e_1_2_12_2_1","doi-asserted-by":"publisher","DOI":"10.1111\/j.1476-5381.1995.tb15903.x"},{"issue":"1","key":"e_1_2_12_3_1","first-page":"S142","article-title":"The concise guide to PHARMACOLOGY 2019\/20: Ion channels","volume":"176","author":"Alexander S. 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