The ion channel properties of a rat recombinant neuronal nicotinic receptor are dependent on the host cell type.
1. A stable mammalian cell line (L-alpha 3 beta 4) has been established which expresses the cloned rat neuronal nicotinic acetylcholine receptor (nAChR) subunits alpha 3 and beta 4, which are the most abundant in autonomic ganglia. Ion channel properties of nAChRs expressed in L-alpha 3 beta 4 cells were investigated by single-channel and whole-cell recording techniques, and compared with both rat alpha 3 beta 4 nAChRs expressed in Xenopus oocytes, and endogenous nicotinic receptors in rat superior cervical ganglion (SCG) neurones, using identical solutions for all cell types. 2. Acetylcholine (ACh) caused activation of single ion channel currents with a range of amplitudes. Some channels had high conductances (30-40 pS), and relatively brief lifetimes; these resembled the predominant native channel from SCG. Other channels had low conductances (20-26 pS) and long bursts of openings which were quite unlike native channels, but which were similar to channels formed by alpha 3 beta 4 in oocytes. Both types often occurred in the same patch. 3. Cytisine was about 3 times more potent than ACh (low-concentration potency ratio) in L-alpha 3 beta 4 cells, which is not dissimilar to the 5-fold potency ratio found in both SCG and oocytes, whereas 1,1-dimethyl-4-phenylpiperazinium (DMPP) was less potent than ACh in some cells (as in the oocyte), but more potent in others (as in SCG). 4. While the channels expressed in L-alpha 3 beta 4 cells do not mimic exactly those expressed in rat SCG, they differ considerably from the same subunit combination expressed in oocytes. Larger conductance, SCG-like channels were detected frequently in L-alpha 3 beta 4, but were rarely, if ever, seen in oocytes injected with alpha 3 and beta 4 mRNA. Our results indicate that ion channel properties such as single-channel conductance can be influenced by the choice of heterologous expression system.
Lewis
T M
TM
Wellcome Laboratory for Molecular Pharmacology, Department of Pharmacology, University College London, UK.
Harkness
P C
PC
Sivilotti
L G
LG
Colquhoun
D
D
Millar
N S
NS
D000109:Acetylcholine / Q000494:pharmacology*; D000818:Animals; D002118:Calcium / Q000378:metabolism; D002460:Cell Line; D003001:Cloning, Molecular; D004246:Dimethylphenylpiperazinium Iodide / Q000494:pharmacology; D005260:Female; D005725:Ganglia, Autonomic / Q000502:physiology; D007473:Ion Channels / Q000187:drug effects / Q000502:physiology*; D007739:L Cells; D008564:Membrane Potentials / Q000187:drug effects; D051379:Mice; D009474:Neurons / Q000502:physiology*; D018722:Nicotinic Agonists / Q000494:pharmacology; D009865:Oocytes / Q000502:physiology; D018408:Patch-Clamp Techniques; D012333:RNA, Messenger / Q000378:metabolism; D051381:Rats; D011978:Receptors, Nicotinic / Q000096:biosynthesis / Q000502:physiology*; D011994:Recombinant Proteins / Q000096:biosynthesis; D014162:Transfection; D014981:Xenopus
D007473:Ion Channels; D018722:Nicotinic Agonists; D012333:RNA, Messenger; D011978:Receptors, Nicotinic; D011994:Recombinant Proteins; D004246:Dimethylphenylpiperazinium Iodide; D000109:Acetylcholine; D002118:Calcium