Abstract
We have characterized the alpha-bungarotoxin receptors (BgtRs) found on the cell surface of undifferentiated pheochromocytoma (PC12) cells. The PC12 cells express a homogeneous population of alpha7-containing receptors that bind alpha-Bgt with high affinity (Kd = 94 pM). The BgtRs mediate most of the response elicited by nicotine, because the BgtR-specific antagonists methyllycaconitine and alpha-Bgt block approximately 90% of the whole-cell current. The binding of nicotinic agonists to cell-surface BgtRs was highly cooperative with four different agonists showing Hill coefficients in the range of 2.3-2.4. A similar agonist binding cooperativity was observed for BgtR homomers formed from chimeric alpha7/5HT3 subunits expressed in tsA 201 cells. Two classes of agonist binding sites, in the ratio of 4:1 for PC12 cell BgtRs and 3:1 for alpha7/5HT3 BgtRs, were revealed by bromoacetylcholine alkylation of the reduced sites on both PC12 BgtRs and alpha7/5HT3 BgtRs. We conclude from this data that PC12 BgtRs and alpha7/5HT3 homomers contain at least three distinguishable agonist binding sites and thus are different from other nicotinic receptors.
Publication types
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Comparative Study
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Research Support, Non-U.S. Gov't
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Research Support, U.S. Gov't, P.H.S.
MeSH terms
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Acetylcholine / analogs & derivatives
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Acetylcholine / metabolism
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Aconitine / analogs & derivatives
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Aconitine / pharmacology
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Alkylation
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Animals
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Binding Sites
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Bungarotoxins / pharmacology
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Centrifugation, Density Gradient
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Cholinergic Agents / pharmacology
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Cross-Linking Reagents / pharmacology
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Neoplasm Proteins / chemistry
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Nerve Tissue Proteins / chemistry*
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Nicotine / pharmacology
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PC12 Cells / chemistry
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Patch-Clamp Techniques
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Rats
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Receptors, Nicotinic / chemistry*
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Receptors, Serotonin / chemistry
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Receptors, Serotonin / genetics
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Receptors, Serotonin, 5-HT3
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Recombinant Fusion Proteins / drug effects
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Recombinant Fusion Proteins / metabolism
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Structure-Activity Relationship
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Succinimides / pharmacology
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alpha7 Nicotinic Acetylcholine Receptor
Substances
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Bungarotoxins
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Cholinergic Agents
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Chrna7 protein, rat
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Cross-Linking Reagents
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Neoplasm Proteins
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Nerve Tissue Proteins
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Receptors, Nicotinic
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Receptors, Serotonin
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Receptors, Serotonin, 5-HT3
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Recombinant Fusion Proteins
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Succinimides
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alpha7 Nicotinic Acetylcholine Receptor
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methyllycaconitine
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Nicotine
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3,3'-dithiobis(sulfosuccinimidyl propionate)
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Acetylcholine
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Aconitine