Identification of cultured cells expressing ligand-gated cationic channels.
We have identified cultured cells that express ligand-gated cation channels using a simple method which may also be applied to the screening of chemical agents for their use as agonists or antagonists. This assay is based upon the observation that many ligand-gated cation channels are permeable to lithium and agonists induce the flux of lithium into the cells which contain them. Since the accumulation of intracellular lithium can alter the cell cycle, the measurement of [3H]thymidine ([3H]thy) incorporation should reflect this occurrence. This expectation was realized using the PC12 cell line which expresses neuronal-like nicotinic acetylcholine receptor (nAChR). When cholinergic agonists are applied to PC12 cells in the presence of lithium-containing buffer and cells are subsequently pulsed with [3H]thy, the radiolabel incorporation into these cells relative to controls is reduced. If cholinergic antagonists are included or if the concentration of agonist either rapidly desensitizes receptors or is insufficient to induce channel opening, the reduction in [3H]thy incorporation is not observed. This method also provides a rapid way to screen cultured cell lines for those that express ligand-gated cation channels. This assay offers the potential to be automated for the low cost screening of drugs which act upon ligand-gated ion channels.
Rogers
S W
SW
Molecular Neurobiology Laboratory, Salk Institute for Biological Studies, La Jolla, California 92037.
Gahring
L C
LC
Papke
R L
RL
Heinemann
S
S
D000818:Animals; D002412:Cations; D002478:Cells, Cultured; D004247:DNA / Q000235:genetics; D015640:Ion Channel Gating; D007473:Ion Channels / Q000187:drug effects / Q000235:genetics / Q000378:metabolism*; D008024:Ligands; D008094:Lithium / Q000378:metabolism; D010276:Parasympatholytics / Q000494:pharmacology; D010277:Parasympathomimetics / Q000494:pharmacology; D011978:Receptors, Nicotinic / Q000187:drug effects / Q000235:genetics / Q000378:metabolism; D013936:Thymidine / Q000378:metabolism; D014162:Transfection
D002412:Cations; D007473:Ion Channels; D008024:Ligands; D010276:Parasympatholytics; D010277:Parasympathomimetics; D011978:Receptors, Nicotinic; D004247:DNA; D008094:Lithium; D013936:Thymidine