An α4β4 Nicotinic Receptor Subtype Is Present in Chick Retina: Identification, Characterization and Pharmacological Comparison with the Transfected α4β4 and α6β4 Subtypes

Retina from 1-day-old chicks is a valuable tissue model for studying neuronal nicotinic receptors because it expresses a large number of the developmentally regulated high affinity [ 3 H]epibatidine labeled nicotinic receptors. Most of these receptors contain the β4 subunit associated with different α subunits. Using a sequential immunodepletion procedure with anti-α6, anti-β3, anti-β2, and anti-β4 antibodies, we purified an α4β4 nicotinic receptor subtype that accounts for approximately 20 to 25% of the high affinity [ 3 H]epibatidine labeled receptors present in retina at that developmental time. Immunoprecipitation and Western blotting experiments confirmed that the purified subtype contains only the α4 and β4 subunits. This receptor binds a number of agonists and the antagonist dihydro-β-erythroidine with nanomolar affinity, whereas it has micromolar affinity for the α-conotoxin MII and methyllycaconitine toxins and other nicotinic antagonists. Comparison of the pharmacological profile of this purified native subtype with that of the same subtype transiently expressed in human BOSC23 cells showed that they have very similar rank orders and absolute K i values for several nicotinic drugs. Finally, because chick retina expresses an α6β4-containing subtype with a high affinity for the α-conotoxin MII, we used native and transfected α4β4 and α6β4 subtypes to investigate the relative contributions of the α and β subunits to this binding, and found that the α6 subunit determines the high affinity for this toxin.