Drosophila nicotinic receptors: evidence for imidacloprid insecticide and alpha-bungarotoxin binding to distinct sites.

摘要

The principal mammalian brain nicotinic acetylcholine receptors (nAChRs) are the alpha-bungarotoxin (alpha-BGT)-insensitive alpha4beta2 and the alpha-BGT-sensitive alpha7 subtypes assayed with radiolabeled nicotinoids and alpha-BGT, respectively. Drosophila head membranes bind the insecticide radioligand [(3)H]imidacloprid ([(3)H]IMI) and [(3)H]alpha-BGT with K(D) 5.7 and 2.7nM and B(max) 980 and 1400fmol/mg protein, respectively. The hypothesis that [(3)H]IMI at 2.5 or 20nM and [(3)H]alpha-BGT at 1 or 10nM bind to distinct sites or subtypes is tested by using these radioligands alone and together in simultaneous dual binding experiments. These studies show no interference by one radioligand in the binding of the other one, i.e., independent binding, and that both unlabeled IMI and alpha-BGT give biphasic displacement curves. The pharmacological profiles of [(3)H]IMI and [(3)H]alpha-BGT suggest distinct binding sites for the two radioligands. These findings are consistent with those obtained with hybrid receptors assembled from Drosophila alpha subunits and a vertebrate beta subunit and with immunological and protein biochemical approaches. This study, therefore, provides direct evidence for distinct IMI- and alpha-BGT-sensitive sites or subtypes in Drosophila brain.