Characterization of the recombinant human neuronal nicotinic acetylcholine receptors α3β2 and α4β2 stably expressed in HEK293 cells
| Autor: | Kathryn J. Elliott, Kenneth A. Stauderman, Gonul Velicelebi, Alison Gillespie, Tadimeti S. Rao, Brian O. Claeps, Michael M. Harpold, Laura E. Chavez-Noriega, Edwin C. Johnson, R. T. Reid, Janis Corey-Naeve, James H. Crona |
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| Rok vydání: | 2000 |
| Předmět: |
medicine.medical_specialty
Patch-Clamp Techniques Blotting Western Receptors Nicotinic Biology Kidney Ligands Cell Line Radioligand Assay Cellular and Molecular Neuroscience Cytisine chemistry.chemical_compound Internal medicine Mecamylamine medicine Humans Nicotinic Agonists Patch clamp Acetylcholine receptor Pharmacology Membranes Blotting Northern Molecular biology Electric Stimulation Recombinant Proteins Electrophysiology Nicotinic acetylcholine receptor Endocrinology Nicotinic agonist chemistry Epibatidine RNA Calcium Acetylcholine medicine.drug |
| Zdroj: | Neuropharmacology. 39:2543-2560 |
| ISSN: | 0028-3908 |
| DOI: | 10.1016/s0028-3908(00)00134-9 |
| Popis: | HEK293 cells were stably transfected with the cDNAs encoding full-length human neuronal nicotinic acetylcholine receptor (nAChR) subunit combinations alpha3beta2 or alpha4beta2. [(3)H]-(+/-)Epibatidine ([(3)H]-(+/-)EPI) bound to membranes from A3B2 (alpha3beta2) and A4B2.2 (alpha4beta2) cells with K(d) values of 7.5 and 33.4 pM and B(max) values of 497 and 1564 fmol/mg protein, respectively. Concentration-dependent increases in intracellular free Ca(2+) concentration were elicited by nAChR agonists with a rank order of potency of EPI1,1-dimethyl-4-phenylpiperazinium (DMPP)nicotine (NIC)=suberyldicholine (SUB)cytisine (CYT)=acetylcholine (ACh) for A3B2 cells and EPICYT=SUB=NIC=DMPPACh for A4B2.2 cells. Antagonists of nAChRs blocked NIC-induced responses with a rank order of potency of d-tubocurarine (d-Tubo)=mecamylamine (MEC)dihydro-beta-erythroidine (DHbetaE) in A3B2 cells and MEC=DHbetaEd-Tubo in A4B2.2 cells. Whole-cell patch clamp recordings indicate that the decay rate of macroscopic ACh-induced currents is faster in A3B2 than in A4B2.2 cells and that A3B2 cells are less sensitive to ACh than A4B2.2 cells. ACh currents elicited in alpha3beta2 and alpha4beta2 human nAChRs are maximally potentiated at 20 and 2 mM external Ca(2+), respectively. Our results indicate that stably expressed alpha3beta2 and alpha4beta2 human nAChRs are pharmacologically and functionally distinct. |
| Databáze: | OpenAIRE |
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