Structure of ligand-binding site of mutated GABA receptors of insects with resistance for antagonists

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OCR houseflies with an A299S mutation in the binding site of noncompetitive GABA antagonists displayed 1 or 2 orders lower resistance to the insecticide fipronil and EBOB than to the insecticide dieldrin (several thousand-fold resistance). Scatchard analysis showed that OCR receptors had 4-fold lower affinity for [^3H]EBOB and 1.3-fold less binding sites than receptors of susceptible WHO files. Dieldrin was 45-fold less potent in inhibiting [^3H]EBOB binding in OCR receptors than in WHO receptors, while fipronil and EBOB were 2-and 6-fold less potent in OCR receptors than in WHO receptors, respectively. 3, 4, 5,-substituted 1-(2,6-dichioro-4-trifluoromethylphenyl)pyrazoles, fipronil analogues, exhibited different structure-activity relationships in OCR receptors from those in WHO receptors. Although most of the analogues had remarkably low affinities for OCR receptors, several analogues with an electron-withdrawing group in the 4-position of the pyrazole ring showed relatively high affinity. However, the affinities were several hundred-fold lower than the affinities for WHO receptors; IC_<50s> were in the micromolar order. An analogue with a bromine atom in the 4-position had >1600-fold lower affinity for OCR receptors than that for WHO receptors. Although the introduction of an amino group into the 5-position led to a decrease in affinity for OCR receptors in one case, three compounds with an amino group in the 5-position, compounds structurally closely related to fipronil, showed only <4-fold decrease in affinity for OCR receptors, compared to that for WHO receptors. The results indicate that only limited compounds with special structures seem to fit into the GABA antagonist-binding site of dieldrin-resistant houseflies. Factor(s) apart from decrease in affinity might be involved in the mechanism underlying the resistance of OCR files for noncompetitive GABA antagonists.