A functional alpha-bungarotoxin receptor is present in chick cerebellum: purification and characterization.
It has recently been demonstrated that alpha-bungarotoxin receptors, which behave as functional nicotinic receptors, are present in chick CNS. In this paper, we report the purification and characterization of a functional alpha-bungarotoxin receptor from chick cerebellum, a nervous tissue in which a clear inhibition of induced nicotine effects has been reported in vivo. This receptor contains at least three subunits of apparent mol. wt 52,000, 57,000 and 67,000. The use of monoclonal antibodies specific for the alpha 7 subunit demonstrated that 75% of the molecules present in our purified preparation belong to the alpha 7 subtype and that this antibody labels the 57,000 band in western blot, thus indicating that this is the toxin binding subunit. Reconstruction experiments in planar lipid bilayers show that this alpha-bungarotoxin receptor forms a cation selective channel whose opening is blocked by d-tubocurarine. Binding experiments on immobilized receptors over an alpha-bungarotoxin-Sepharose affinity column show that the ligand binding subunit is present in vivo in two copies per receptor. Immunological, pharmacological and functional experiments show that this purified receptor is very similar, but not identical, to the previously characterized chick optic lobe receptor, thus indicating the heterogeneity of these alpha-bungarotoxin receptors in the CNS.
CNR Center of Cytopharmacology, Department of Medical Pharmacology, University of Milan, Italy.
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C
Gotti
W
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Hanke
W R
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Schlue
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Briscini
M
M
Moretti
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F
Clementi
D000818:Animals; D000911:Antibodies, Monoclonal; D001667:Binding, Competitive; D002038:Bungarotoxins / Q000378:metabolism*; D002217:Carbachol / Q000494:pharmacology; D002462:Cell Membrane / Q000378:metabolism; D002499:Centrifugation, Density Gradient; D002531:Cerebellum / Q000378:metabolism*; D002645:Chickens; D002846:Chromatography, Affinity; D004591:Electrophoresis, Polyacrylamide Gel; D018738:Hexamethonium; D006584:Hexamethonium Compounds / Q000494:pharmacology; D007106:Immune Sera; D015640:Ion Channel Gating / Q000187:drug effects; D007700:Kinetics; D008051:Lipid Bilayers; D008970:Molecular Weight; D011978:Receptors, Nicotinic / Q000187:drug effects / Q000302:isolation & purification / Q000378:metabolism / Q000502:physiology*; D014403:Tubocurarine / Q000494:pharmacology; D064569:alpha7 Nicotinic Acetylcholine Receptor
D000911:Antibodies, Monoclonal; D002038:Bungarotoxins; D006584:Hexamethonium Compounds; D007106:Immune Sera; D008051:Lipid Bilayers; D011978:Receptors, Nicotinic; D064569:alpha7 Nicotinic Acetylcholine Receptor; D018738:Hexamethonium; D002217:Carbachol; D014403:Tubocurarine