A study of the dopamine transporter using the TRACT assay, a novel in vitro tool for solute carrier drug discovery

Abstract Members of the solute carrier (SLC) transporter protein family are increasingly recognized as therapeutic drug targets. The majority of drug screening assays for SLCs are based on the uptake of radiolabeled or fluorescent substrates. Thus, these approaches often have limitations that compro...

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Autores principales:	Hubert J. Sijben, Julie J. E. van den Berg, Jeremy D. Broekhuis, Adriaan P. IJzerman, Laura H. Heitman
Formato:	article
Lenguaje:	EN
Publicado:	Nature Portfolio 2021
Materias:	Medicine R Science Q
Acceso en línea:	https://doaj.org/article/6dfa7bddb1ad49129d7c2d0b60f32013
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spelling	oai:doaj.org-article:6dfa7bddb1ad49129d7c2d0b60f320132021-12-02T14:01:33ZA study of the dopamine transporter using the TRACT assay, a novel in vitro tool for solute carrier drug discovery10.1038/s41598-020-79218-w2045-2322https://doaj.org/article/6dfa7bddb1ad49129d7c2d0b60f320132021-01-01T00:00:00Zhttps://doi.org/10.1038/s41598-020-79218-whttps://doaj.org/toc/2045-2322Abstract Members of the solute carrier (SLC) transporter protein family are increasingly recognized as therapeutic drug targets. The majority of drug screening assays for SLCs are based on the uptake of radiolabeled or fluorescent substrates. Thus, these approaches often have limitations that compromise on throughput or the physiological environment of the SLC. In this study, we report a novel application of an impedance-based biosensor, xCELLigence, to investigate dopamine transporter (DAT) activity via substrate-induced activation of G protein-coupled receptors (GPCRs). The resulting assay, which is coined the ‘transporter activity through receptor activation’ (TRACT) assay, is based on the hypothesis that DAT-mediated removal of extracellular dopamine directly affects the ability of dopamine to activate cognate membrane-bound GPCRs. In two human cell lines with heterologous DAT expression, dopamine-induced GPCR signaling was attenuated. Pharmacological inhibition or the absence of DAT restored the apparent potency of dopamine for GPCR activation. The inhibitory potencies for DAT inhibitors GBR12909 (pIC50 = 6.2, 6.6) and cocaine (pIC50 = 6.3) were in line with values from reported orthogonal transport assays. Conclusively, this study demonstrates the novel use of label-free whole-cell biosensors to investigate DAT activity using GPCR activation as a readout. This holds promise for other SLCs that share their substrate with a GPCR.Hubert J. SijbenJulie J. E. van den BergJeremy D. BroekhuisAdriaan P. IJzermanLaura H. HeitmanNature PortfolioarticleMedicineRScienceQENScientific Reports, Vol 11, Iss 1, Pp 1-14 (2021)
institution	DOAJ
collection	DOAJ
language	EN
topic	Medicine R Science Q
spellingShingle	Medicine R Science Q Hubert J. Sijben Julie J. E. van den Berg Jeremy D. Broekhuis Adriaan P. IJzerman Laura H. Heitman A study of the dopamine transporter using the TRACT assay, a novel in vitro tool for solute carrier drug discovery
description	Abstract Members of the solute carrier (SLC) transporter protein family are increasingly recognized as therapeutic drug targets. The majority of drug screening assays for SLCs are based on the uptake of radiolabeled or fluorescent substrates. Thus, these approaches often have limitations that compromise on throughput or the physiological environment of the SLC. In this study, we report a novel application of an impedance-based biosensor, xCELLigence, to investigate dopamine transporter (DAT) activity via substrate-induced activation of G protein-coupled receptors (GPCRs). The resulting assay, which is coined the ‘transporter activity through receptor activation’ (TRACT) assay, is based on the hypothesis that DAT-mediated removal of extracellular dopamine directly affects the ability of dopamine to activate cognate membrane-bound GPCRs. In two human cell lines with heterologous DAT expression, dopamine-induced GPCR signaling was attenuated. Pharmacological inhibition or the absence of DAT restored the apparent potency of dopamine for GPCR activation. The inhibitory potencies for DAT inhibitors GBR12909 (pIC50 = 6.2, 6.6) and cocaine (pIC50 = 6.3) were in line with values from reported orthogonal transport assays. Conclusively, this study demonstrates the novel use of label-free whole-cell biosensors to investigate DAT activity using GPCR activation as a readout. This holds promise for other SLCs that share their substrate with a GPCR.
format	article
author	Hubert J. Sijben Julie J. E. van den Berg Jeremy D. Broekhuis Adriaan P. IJzerman Laura H. Heitman
author_facet	Hubert J. Sijben Julie J. E. van den Berg Jeremy D. Broekhuis Adriaan P. IJzerman Laura H. Heitman
author_sort	Hubert J. Sijben
title	A study of the dopamine transporter using the TRACT assay, a novel in vitro tool for solute carrier drug discovery
title_short	A study of the dopamine transporter using the TRACT assay, a novel in vitro tool for solute carrier drug discovery
title_full	A study of the dopamine transporter using the TRACT assay, a novel in vitro tool for solute carrier drug discovery
title_fullStr	A study of the dopamine transporter using the TRACT assay, a novel in vitro tool for solute carrier drug discovery
title_full_unstemmed	A study of the dopamine transporter using the TRACT assay, a novel in vitro tool for solute carrier drug discovery
title_sort	study of the dopamine transporter using the tract assay, a novel in vitro tool for solute carrier drug discovery
publisher	Nature Portfolio
publishDate	2021
url	https://doaj.org/article/6dfa7bddb1ad49129d7c2d0b60f32013
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A study of the dopamine transporter using the TRACT assay, a novel in vitro tool for solute carrier drug discovery

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