Inducible, heterologous expression of human alpha7-nicotinic acetylcholine receptors in a native nicotinic receptor-null human clonal line.

Abstract

Tetracycline-regulated expression of recombinant nicotinic acetylcholine receptors (nAChR) composed of human alpha7 subunits is achieved in native nAChR-null SH-EP1 human epithelial cells. alpha7 subunits are heterologously expressed as messenger RNA and as components of 125I-labeled alpha-bungarotoxin (I-Bgt)-binding nAChR ( approximately 10 pmol per milligram of membrane protein) at levels sensitive to the amount of tetracycline in cell growth medium. I-Bgt-binding alpha7-nAChR appear on the cell surface pool and in intracellular pools. The pharmacological profile for drug competition toward I-Bgt binding to these recombinant alpha7-nAChR matches that of human native alpha7-nAChR naturally expressed in SH-SY5Y human neuroblastoma cells (rank order potency methyllycaconitine>1, 1-dimethyl-4-phenylpiperazinium>(-)nicotine>cytisine>carbamylch oli ne> /=d-tubocurarine). Chronic exposure to nicotine induces up-regulation of human recombinant alpha7-nAChR (80% up-regulation at 10 microM nicotine) just as it does native alpha7-nAChR in other human cell lines. These studies confirm expression of nAChR as homooligomers of human alpha7 subunits from transgenes, establish a native nAChR-null background for such expression, and demonstrate that this expression can be regulated to facilitate studies of human alpha7-nAChR.

• Authors: J H Peng, L Lucero, J Fryer, J Herl, S S Leonard, R J Lukas
• Journal: Brain research (Brain Res) Vol. 825 Issue 1-2 Pg. 172-9 (Apr 17 1999) ISSN: 0006-8993 [Print] Netherlands
• PMID: 10216184 (Publication Type: Journal Article, Research Support, Non-U.S. Gov't, Research Support, U.S. Gov't, P.H.S.)
• Copyright: Copyright 1999 Elsevier Science B.V.

Chemical References

• Alkaloids
• Azocines
• Bungarotoxins
• Chrna7 protein, human
• DNA, Complementary
• Insecticides
• Iodine Radioisotopes
• Nicotinic Agonists
• Protein Synthesis Inhibitors
• Quinolizines
• RNA, Messenger
• Receptors, Nicotinic
• Recombinant Proteins
• alpha7 Nicotinic Acetylcholine Receptor
• methyllycaconitine
• cytisine
• Dimethylphenylpiperazinium Iodide
• Nicotine
• Carbachol
• Tetracycline
• Acetylcholine
• Tubocurarine
• Aconitine

Topics

• Acetylcholine (physiology)
• Aconitine (analogs & derivatives, pharmacology)
• Alkaloids (pharmacology)
• Azocines
• Blotting, Northern
• Bungarotoxins (pharmacology)
• Carbachol (pharmacology)
• DNA, Complementary
• Dimethylphenylpiperazinium Iodide (pharmacology)
• Gene Expression (drug effects, physiology)
• Humans
• Insecticides (pharmacology)
• Iodine Radioisotopes
• Neuroblastoma
• Neurons (chemistry, physiology)
• Nicotine (pharmacology)
• Nicotinic Agonists (pharmacology)
• Plasmids
• Protein Synthesis Inhibitors (pharmacology)
• Quinolizines
• RNA, Messenger (analysis)
• Receptors, Nicotinic (genetics, metabolism)
• Recombinant Proteins (genetics, metabolism)
• Tetracycline (pharmacology)
• Transfection
• Tubocurarine (pharmacology)
• Tumor Cells, Cultured (chemistry, cytology)
• Up-Regulation (genetics)
• alpha7 Nicotinic Acetylcholine Receptor