Abstract
Nicotinic acetylcholine receptors (nAChR) of the TE671 cell line were investigated using whole-cell and membrane patch recording techniques. At negative holding potentials (VH), pulses of acetylcholine (ACh) elicited whole-cell inward currents that rapidly desensitized. The EC50 value for ACh at VH = −60 mV was 7.8 μM. The ACh-induced current reversed at ∼0 mV. Desensitization of nAChR by ACh was biphasic and reversible within ∼20 sec. Spermine (1–100 μM) potentiated responses to ACh (10 μM − 1 mM) by reducing the rate of onset of desensitization; potentiation was inhibited by arcaine (10–100 μM). Spermine (1 mM) noncompetitively antagonized the AChinduced current. Antagonism by 1 to 5 mM spermine was voltage-dependent, increasing with negative VH. In 100 μM arcaine, this antagonism was shown to contain a voltage-independent component. Spermine (10 mM) increased the EC50 values for ACh, suggesting that at this concentration the polyamine is also a competitive antagonist. Single channel openings elicited during application of ACh to outside-out patches had a conductance of 47 pS at VH = −60 mV. At 10 and 100 μM, spermine increased channel open probability (po), but at 1 mM spermine, po was not significantly different from controls. The single channel conductance for ACh was unaffected by 10 and 100 μM spermine, but was decreased by 1 mM spermine. Spermine promoted the occurrence of ∼27 pS openings. It is proposed that spermine acts at an excitatory modulatory site similar to that present on N-methyl-d-aspartate receptors and at least three inhibitory sites on nAChR of TE671 cells.
Footnotes
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Send reprint requests to: Dr. I. R. Mellor, Division of Molecular Toxicology, School of Biological Sciences, University of Nottingham, University Park, Nottingham NG7 2RD, UK.
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↵1 This work was financially supported by a NATO Research Grant to P.N.R.U. and by the EC BIOMED-2 program contract PL 962395.
- Abbreviations:
- ACh
- acetylcholine
- nAChR
- nicotinic acetylcholine receptor
- NMDAR
- N-methyl-d-aspartate receptor
- AMPA
- α-amino-3-hydroxy-5-methyl-4-isoxazolepropionic acid
- mAChR
- muscarinic acetylcholine receptor
- HEPES
- N-[2-hydroxyethyl]piperazine-N′-[2-ethanesulphonic acid]
- Received February 5, 1998.
- Accepted April 22, 1998.
- The American Society for Pharmacology and Experimental Therapeutics
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