Sulfhydryl modification of two nicotinic binding sites in mouse brain

Area of Science:

• Neuroscience
• Pharmacology
• Molecular Biology

Background:

• Brain tissue contains distinct binding sites for nicotine and alpha-bungarotoxin, characteristic of nicotinic cholinergic receptors.
• Disulfide bonds play a role in the structure and function of these receptors.

Purpose of the Study:

• To investigate the effects of disulfide reduction on nicotinic receptor binding sites in mouse brain.
• To compare the response of brain nicotinic receptors to disulfide reduction with those in electric tissue.

Main Methods:

• Treatment of DBA mouse brain tissue with dithiothreitol (a disulfide-reducing agent).
• Measurement of nicotine and alpha-bungarotoxin binding before and after reduction and reoxidation.
• Assessment of ligand association and dissociation rates.
• Selective alkylation of reduced binding sites with bromoacetylcholine.

Main Results:

• Dithiothreitol treatment reduced overall binding for both ligands.
• Nicotine binding showed reduced maximal binding, while alpha-bungarotoxin binding showed decreased affinity.
• Agonist affinity for the alpha-bungarotoxin site was reduced at low dithiothreitol concentrations.
• Reduced nicotine sites maintained similar ligand association/dissociation rates, but dissociation was accelerated.
• Both reduced receptor types could be selectively alkylated with bromoacetylcholine.

Conclusions:

• Putative nicotinic receptors in the brain exhibit similar responses to disulfide reduction.
• The observed responses are comparable to those of nicotinic receptors found in electric tissue.
• Disulfide bonds are critical for maintaining the functional integrity of brain nicotinic receptors.