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rdf:type |
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lifeskim:mentions |
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pubmed:issue |
1
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pubmed:dateCreated |
2003-9-9
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pubmed:abstractText |
Human placental choriocarcinoma (JAR) cells endogenously expressing glycine transporter type 1a (GlyT1a) have been cultured in 96-well scintillating microplates to develop a homogenous screening assay for the detection of GlyT1 antagonists. In these microplates uptake of [14C]glycine was time dependent and saturable with a Michaelis-Menten constant (Km) of 27+/-3 microM. The GlyT1 transport inhibitors sarcosine, ALX-5407, and Org-24598 were tested and shown to block [14C]glycine uptake with expected IC50 values of 37.5+/-4.6 microM, 2.8+/-0.6 nM, and 6.9+/-0.9 nM, respectively. The [14C]glycine uptake process was sensitive to membrane Na+ gradient as blockade of membrane Na+/K+-ATPase by ouabain or Na+ exchanger by benzamil-disrupted glycine accumulation in JAR cells. Glycine influx was not affected by concentration of dimethyl sulfoxide up to 2%. The versatility of this technological approach was further confirmed by the characterization of a saturable [14C]taurine uptake in JAR cells. Taurine transport was of high affinity with a Km of 10.2+/-1.7 microM and fully inhibited by ALX-5407 (IC50=522 +/-83 nM). The developed assay is homogenous, rapid, versatile and amenable to automation for the discovery of new neurotransmitter transporter inhibitors.
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pubmed:language |
eng
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pubmed:journal |
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pubmed:citationSubset |
IM
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pubmed:chemical |
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pubmed:status |
MEDLINE
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pubmed:month |
Oct
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pubmed:issn |
0003-2697
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pubmed:author |
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pubmed:issnType |
Print
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pubmed:day |
1
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pubmed:volume |
321
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pubmed:owner |
NLM
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pubmed:authorsComplete |
Y
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pubmed:pagination |
31-7
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pubmed:dateRevised |
2005-11-17
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pubmed:meshHeading |
pubmed-meshheading:12963052-Amino Acid Transport Systems, Neutral,
pubmed-meshheading:12963052-Carbon Radioisotopes,
pubmed-meshheading:12963052-Cell Count,
pubmed-meshheading:12963052-Cell Line, Tumor,
pubmed-meshheading:12963052-Chlorides,
pubmed-meshheading:12963052-Dose-Response Relationship, Drug,
pubmed-meshheading:12963052-Female,
pubmed-meshheading:12963052-Gene Expression Regulation, Neoplastic,
pubmed-meshheading:12963052-Glycine,
pubmed-meshheading:12963052-Glycine Plasma Membrane Transport Proteins,
pubmed-meshheading:12963052-Humans,
pubmed-meshheading:12963052-Pregnancy,
pubmed-meshheading:12963052-Scintillation Counting,
pubmed-meshheading:12963052-Sodium,
pubmed-meshheading:12963052-Taurine,
pubmed-meshheading:12963052-Time Factors
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pubmed:year |
2003
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pubmed:articleTitle |
Development of a scintillation proximity assay for analysis of Na+/Cl- -dependent neurotransmitter transporter activity.
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pubmed:affiliation |
Department of Neuroscience West Point, Merck & Co. Inc., West Point, PA 19486, USA.
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pubmed:publicationType |
Journal Article
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