Nicotinic acetylcholine receptors (nAChRs) are pentameric cation channels that mediate excitatory neurotransmission. It is reasonable to assume that insects have several nAChR subtypes composed of different subunits, but little is known regarding the subunit composition of nAChRs in insects. Insect nAChRs contain multiple target sites for various insecticides, such as neonicotinoids and spinosyns. Our previous studies demonstrated that the neonicotinoids dinotefuran and imidacloprid may bind to distinct nAChR subtypes in the nerve cords of the American cockroach (Periplaneta americana). To gain more insight into this finding, we examined the insecticidal effects of the two neonicotinoids in brown planthoppers (Nilaparvata lugens), in which the expression of an individual subunit gene (Nlα1−3, Nlα6, Nlα8, Nlα10, Nlβ1, and Nlβ2) is specifically reduced by RNA interference. The microinjection of double-stranded RNA (dsRNA) fragments, except for a fragment corresponding to Nlα6, resulted in a decrease in the sensitivity of the planthoppers to imidacloprid. The sensitivity to dinotefuran was decreased only when the dsRNA fragments corresponding to Nlα2 and Nlβ1 were injected. These findings imply that the Nlα1−3, Nlα8, Nlα10, Nlβ1, and Nlβ2 subunits are associated with the imidacloprid-targeting nAChR subtypes and that a major dinotefuran-targeting subtype may contain the Nlα2 and Nlβ1 subunits. We next investigated the effects of the Drosophila α1 subunit (Dα1) gene knockout on the insecticidal effects of dinotefuran and imidacloprid. This knockout line (Dα1KO) demonstrated decreased sensitivity to imidacloprid but the same sensitivity to dinotefuran as the wild type. We also examined the specific binding of [3H]dinotefuran and [3H]imidacloprid to the head membranes of Dα1KO flies. Consequently, the maximum number of [3H]imidacloprid binding sites was diminished by Dα1 knockout, whereas the binding of [3H]dinotefuran was not significantly affected. These findings suggest that the Dα1 subunit constitutes the imidacloprid-targeting subtype but not the major dinotefuran-targeting subtype. Overall, our findings suggest that dinotefuran acts on a narrower range of nAChR subtype(s) than does imidacloprid. The subunit composition of the dinotefuran-targeting subtype(s) remains to be determined.

Loss-of-function of nicotinic acetylcholine receptor subunits reveals distinct insecticidal effects of dinotefuran

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