Characterization of a genetically reconstituted high-affinity system for serotonin transport.
By transfecting mouse fibroblast L-M cells with human genomic DNA, we have established and identified several clonal cell lines that stably express a high-affinity serotonin (5-HT)-uptake mechanism absent in untransfected host cells. One such cell line, L-S1, possesses features of 5-[3H]HT uptake similar to those previously characterized in the central nervous system and blood platelets: (i) specificity for 5-HT; (ii) antagonism by imipramine, a known inhibitor of high-affinity 5-HT uptake; (iii) both Na+ and temperature dependences; (iv) kinetic saturability; and (v) high affinity for 5-HT (Km = 0.39 +/- 0.10 microM; Vmax = 2.14 +/- 0.55 pmol/min per mg of protein). This cell line can be used to compare the relative efficacies of known blockers of 5-HT uptake and thereby offers a rapid and reliable assay system for testing novel inhibitors of this system. Since L-S1 contains stably integrated human DNA in its genome, we postulate that the observed 5-HT-uptake system resulted from the expression of human gene(s) coding for the 5-HT transporter. Thus, cell lines such as L-S1 may represent novel means for screening and developing therapeutic agents specific for neurotransmitter-uptake systems as well as substrates for the cloning and elucidation of the genes encoding the various neurotransmitter transporters.
Chang
A S
AS
Laboratory of Cellular and Molecular Neurobiology, Baylor College of Medicine, Woodlands, TX 77381.
Frnka
J V
JV
Chen
D N
DN
Lam
D M
DM
D000818:Animals; D001692:Biological Transport; D002352:Carrier Proteins / Q000235:genetics* / Q000378:metabolism; D004247:DNA / Q000235:genetics*; D006801:Humans; D007700:Kinetics; D007739:L Cells / Q000378:metabolism; D051379:Mice; D012701:Serotonin / Q000378:metabolism*; D013816:Thermodynamics; D014162:Transfection
D002352:Carrier Proteins; D012701:Serotonin; D004247:DNA