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However, its pharmacological potential in the central nervous system (CNS) remains largely unexplored. In this study, we evaluated the acute toxicity and behavioral responses of adult zebrafish (\n                    <jats:italic>Danio rerio<\/jats:italic>\n                    ) to intraperitoneal (IP) injection of HUM. The behavioral parameters examined were number of line crossings (open\u2010field test) and time spent in the light zone of the tank (light\/dark test). Then were performed experiments with standard antagonists to investigate the possible mechanism of action. Humulene exhibited low acute toxicity (LD\n                    <jats:sub>50<\/jats:sub>\n                    &gt; 40\u00a0mg kg\n                    <jats:sup>\u22121<\/jats:sup>\n                    , IP) and produced a maximal anxiolytic\u2010like effect at an intermediate dose of 20\u00a0mg kg\n                    <jats:sup>\u22121<\/jats:sup>\n                    IP. The pharmacological dissection revealed that this effect was independent of the benzodiazepine (B\n                    <jats:sub>ZD<\/jats:sub>\n                    ) allosteric binding site on the gamma\u2010aminobutyric acid (GABA) type A receptor (GABA\n                    <jats:sub>A<\/jats:sub>\n                    R), but it was completely reversed by granisetron (G\n                    <jats:sub>RAN<\/jats:sub>\n                    ), a serotonin (5\u2010HT) type 3 receptor (5\u2010HT\n                    <jats:sub>3<\/jats:sub>\n                    R) antagonist. Further in silico analysis revealed a low\u2010affinity binding and an allosteric interaction of \u03b1\u2010humulene, respectively, with GABA\n                    <jats:sub>A<\/jats:sub>\n                    R and 5\u2010HT\n                    <jats:sub>3<\/jats:sub>\n                    R channels. Together, these results suggest a noncanonical mechanism involving both GABAergic and serotonergic systems.\n                  <\/jats:p>","DOI":"10.1002\/cbdv.71534","type":"journal-article","created":{"date-parts":[[2026,7,26]],"date-time":"2026-07-26T18:10:02Z","timestamp":1785089402000},"update-policy":"https:\/\/doi.org\/10.1002\/crossmark_policy","source":"Crossref","is-referenced-by-count":1,"title":["Assessment of the Acute Toxicity and Anxiolytic\u2010Like Effect of\n                    <i>\u03b1\u2010<\/i>\n                    Humulene in Adult Zebrafish (\n                    <i>Danio rerio<\/i>\n                    )"],"prefix":"10.1002","volume":"23","author":[{"ORCID":"https:\/\/orcid.org\/0000-0002-4162-8860","authenticated-orcid":false,"given":"Alex de Souza","family":"Borges","sequence":"first","affiliation":[{"name":"Department of Chemical\u2010Biology, Post\u2010Graduate Program in Chemical\u2010Biology Regional University of Cariri, Pimenta Campus  Crato Cear\u00e1 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