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Electroanalytical methodologies have been able to monitor steady state basal extracellular 5-HT levels but are unable to provide insight into how these levels are influenced by key regulatory processes such as release and uptake. We established a new measurement approach, amperometry approach curve profiling, which monitors the extracellular 5-HT level at different electrode\u2013tissue (E\u2013T) distances. Analysis of the current profile can provide information on contributions of regulatory components on the observed extracellular 5-HT level. Measurements were conducted from ex vivo murine ileum and colon using a boron-doped diamond (BDD) microelectrode. Amperometry approach curve profiling coupled with classical pharmacology demonstrated that extracellular 5-HT levels were significantly lower in the colon when compared to the ileum. This difference was due to a greater degree of activity of the 5-HT transporter (SERT) and a reduced amount of 5-HT released from colonic EC cells. The presence of an inhibitory 5-HT<jats:sub>4<\/jats:sub>autoreceptor was observed in the colon, where a 40% increase in extracellular 5-HT was the half maximal inhibitory concentration for activation of the autoreceptor. This novel electroanalytical approach allows estimates of release and re-uptake and their contribution to 5-HT extracellular concentration from intestinal tissue be obtained from a single series of measurements.<\/jats:p>","DOI":"10.1038\/s41598-024-61296-9","type":"journal-article","created":{"date-parts":[[2024,5,7]],"date-time":"2024-05-07T13:01:47Z","timestamp":1715086907000},"update-policy":"https:\/\/doi.org\/10.1007\/springer_crossmark_policy","source":"Crossref","is-referenced-by-count":1,"title":["Amperometry approach curve profiling to understand the regulatory mechanisms governing the concentration of intestinal extracellular serotonin"],"prefix":"10.1038","volume":"14","author":[{"ORCID":"https:\/\/orcid.org\/0000-0003-2113-8650","authenticated-orcid":false,"given":"Mark S.","family":"Yeoman","sequence":"first","affiliation":[],"role":[{"role":"author","vocabulary":"crossref"}]},{"given":"Sara","family":"Fidalgo","sequence":"additional","affiliation":[],"role":[{"role":"author","vocabulary":"crossref"}]},{"ORCID":"https:\/\/orcid.org\/0000-0002-7475-7327","authenticated-orcid":false,"given":"Gianluca","family":"Marcelli","sequence":"additional","affiliation":[],"role":[{"role":"author","vocabulary":"crossref"}]},{"ORCID":"https:\/\/orcid.org\/0000-0002-8773-3850","authenticated-orcid":false,"given":"Bhavik Anil","family":"Patel","sequence":"additional","affiliation":[],"role":[{"role":"author","vocabulary":"crossref"}]}],"member":"297","published-online":{"date-parts":[[2024,5,7]]},"reference":[{"key":"61296_CR1","doi-asserted-by":"publisher","first-page":"83","DOI":"10.1016\/0166-4328(96)00075-7","volume":"73","author":"K Racke","year":"1995","unstructured":"Racke, K., Reimann, A., Schworer, H. & Kilbinger, H. 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