Anti-Peptide Specific Antibodies for the Characterization of Different α Subunits of α-Bungarotoxin Binding Acetylcholine Receptors Present in Chick Optic Lobe

Abstract

Chick optic lobe express α-Bungarotoxin receptors. We have recently purified these receptors which, when reconstituted in a lipid bilayer, behave as functional acetylcholine gated channels. In order to characterize this purified preparation, we raised polyclonal antibodies against peptides obtained from the putative cytoplasmic domain between the hydrophobic sequence M3 and M4 of two previously cloned α-Bungarotoxin receptor subunits, α7 and α8. Both antibodies recognized the receptors present in the membrane extract and in the purified preparation, although the amount of the α-Bungarotoxin receptors precipitated by the two antibodies was quantitatively different. In Western blots of both purified and membrane-bound receptors, these antibodies specifically reacted with an Mr 57000–55000 band. A study was also undertaken to quantify the receptors containing these subunits in different chick brain areas; it was found that the number of these subunits, as well as their ratio, was similar in all the tested areas. Furthermore, the α-Bungarotoxin receptors were present in at least two subtypes, one containing only the α7 subunit and the other both α7 and α8 subunits.