PHYSOSTIGMINE AND GALANTHAMINE - PROBES FOR A NOVEL BINDING-SITE ON THE ALPHA-4-BETA-2 SUBTYPE OF NEURONAL NICOTINIC ACETYLCHOLINE-RECEPTORS STABLY EXPRESSED IN FIBROBLAST CELLS

In the present study, we demonstrated that the chicken alpha 4 beta 2 neuronal nicotinic receptor stably expressed in transfected mouse fibr oblasts (M10 cells) can be activated via the acetylcholine-binding sit e or via a site that is distinct from that for acetylcholine and recog nizes physostigmine and galanthamine as agonists. In outside-out patch es excised from dexamethasone-induced M10 cells, (+)-anatoxin-a, physo stigmine and galanthamine (each at 1 mu M) activated single channels w ith conductances of 18 and 30 pS. Dihydro-beta-erythroidine (1-30 nM), but not the nicotinic receptor-specific monoclonal antibody FK1, redu ced the frequency of channels activated by anatoxin(l mu M). On the ot her hand, the frequency of channel activity induced by physostigmine ( 1 mu M) was unaffected by dihydro-beta-erythroidine and was markedly d ecreased by FK1. In uninduced M10 cells and in dexamethasone-treated u ntransfected fibroblasts, we observed that physostigmine, galanthamine and nicotinic agonists did not evoke whole-cell or single-channel cur rents. Also, neither [H-3]L-nicotine nor FK1 was able to bind to unind uced M10 cells. In dexamethasone-induced M10 cells, the nicotinic agon ists acetylcholine, anatoxin, 1,1-dimethyl-4-phenylpiperazinium, (-)-n icotine, and cytisine (each at 100 mu M) activated whole-cell currents that showed a marked inward rectification and were sensitive to block ade by dihydro-beta-erythroidine (100 nM). However, neither galanthami ne nor physostigmine could evoke whole-cell currents in cells that wer e responsive to nicotinic agonists. Other effects of physostigmine and galanthamine on the nicotinic receptor that outweigh the agonist prop erties of these compounds could account for their inability to evoke w hole-cell currents.