Thiamethoxam, a new neonicotinoid insecticide acting at nicotinic acetylcholine receptors, was characterized in competition binding assays with [3‐H]‐imidacloprid, a specific nicotinic ligand, using membranes from the aphids Aphis craccivora and Myzus persicae, and from the locust Locusta migratoria. In all insects, Scatchard analysis suggested two binding sites for imidacloprid with Kd values in the range of 1 nM and 10 nM, respectively. The Hill values were significantly below 1 (range of 0.63 to 0.85). In contrast to imidacloprid and nicotine, the potency of thiamethoxam to displace [3‐H]‐imidacloprid varied considerably among these insects. Thiamethoxam was more active than nicotine on Aphis receptors but 100‐fold less in Locusta, a nontarget insect. Comparable relations were found to nithiazine. In Myzus, the inhibition curve for thiamethoxam was shallow. This suggested a heterogeneous receptor population displaying a range of binding affinities to thiamethoxam in this aphid. In all three insects, the other neonicotinoid insecticides studied competed with [3‐H]‐imidacloprid in the same order: thiacloprid > imidacloprid ≥ acetamiprid > nitenpyram. N‐Methylation of imidacloprid strongly reduced the affinity to the imidacloprid site, whereas N‐demethylation of thiamethoxam resulted in a comparable increase of affinity. Supplementary assays were performed with (−)‐[3‐H]‐nicotine and [3‐H]‐α‐bungarotoxin on locust membranes. Overall, the data suggested that the outstanding insecticidal properties of thiamethoxam may be due to either a different binding site on nicotinic receptors, or receptor isoforms, or specific pharmakokinetic behavior, rather than to exceptional affinity to one of the examined binding sites. © 2000 John Wiley & Sons, Inc. J Biochem Toxicol 14:221–230, 2000

No abstract

Cyromazine, an insect growth regulator, affects larval and pupal cuticles in dipterans and some other insects. The mode of action of this aminotriazine is not known yet, though it has been shown not to inhibit the synthesis of chitin and cuticular proteins. Cyromazine may, however, act on some step(s) of sclerotization of the cuticle. In the present study, we have analyzed the key enzyme for the production of sclerotization agents, phenylalanine hydroxylase (PAH), using the enzyme from Drosophila, a cyromazine-sensitive insect. PAH was studied in vitro with cyromazine and three biologically less active derivatives at concentrations ranging from 1 microM to 1 mM. None of the compounds did significantly affect PAH activity. Nor did cyromazine, fed to last instar larvae of Musca domestica, change the relative content of phenylalanine and tyrosine, or the total amount and profile of amino acids of puparial cuticles, which showed a larviform shape typical for cyromazine intoxication. Taken together, this study does not support the hypothesis that phenylalanine hydroxylase represents a target site of cyromazine. In additional studies, the conflicting results, as reported by others, on in vitro inhibition of dihydrofolate reductase (DHFR) by cyromazine were re-examined using the enzymes from larvae of the blowfly, Protophormia terraenovae, and from hen liver. There was no significant inhibition of either DHFR at 100 microM by cyromazine as well as by dicylanil, a pyrimidine analog that is biologically more active than cyromazine. In conclusion, the mode of action of cyromazine remains completely open.