Identification of a high-affinity binding site for dinotefuran in the nerve cord of the American cockroach

The binding of the neonicotinoid insecticide dinotefuran to insect nicotinic acetylcholine receptors (nAChRs) was examined by a centrifugation method using the nerve cord membranes of American cockroaches and [3H]dinotefuran (78 Ci mmol-1). The Kd and Bmax values of [3H]dinotefuran binding were estimated to be 13.7 nM and 14.8 fmol 40 microg-1 protein respectively by Scatchard analysis. Epibatidine, an nAChR agonist, showed a rather lower affinity to the dinotefuran binding site (IC50=991 nM) than dinotefuran (IC50=5.02 nM). Imidacloprid and nereistoxin displayed lower potencies than dinotefuran but higher potencies than epibatidine. The potencies of five dinotefuran analogues in inhibiting the specific binding of [3H]dinotefuran to nerve cord membranes were determined. A good correlation (r2=0.970) was observed between the -log IC50 values of the tested compounds and their piperonyl butoxide-synergised insecticidal activities (-log LD50 values) against German cockroaches. The results indicate that a high-affinity binding site for dinotefuran is present in the nerve cord of the American cockroach and that the binding of ligands to the site leads to the manifestation of insecticidal activity.     

Correlations of the electrophysiological activity of neonicotinoids with their binding and insecticidal activities

The electrophysiological actions of various neonicotinoids, including substituted benzyl derivatives, against recombinant Drosophila SAD/chicken beta2 hybrid nicotinic acetylcholine receptor (nAChR) were measured to analyze the relationships between the in vivo (insecticidal) and in vitro (binding and agonist) activities. Most of the neonicotinoids tested were capable of inducing inward currents by activating the hybrid nAChRs expressed in Xenopus laevis oocytes, whereas some compounds had no agonist activity and only blocked the acetylcholine-induced currents. Variations in the agonist activity were well correlated with those in the binding potency evaluated using [3H]imidacloprid as well as insecticidal activities.     

Insecticidal activity and nicotinic acetylcholine receptor binding of dinotefuran and its analogues in the housefly, Musca domestica

The insecticidal activity of dinotefuran and 23 related compounds against the housefly, Musca domestica (L) was measured by injection with metabolic inhibitors. Dinotefuran was less active than imidacloprid and clothianidin by a factor of 10 in molar concentrations. Their binding activities to the fly-head membrane preparation were measured by using [125I]alpha-bungarotoxin ([125I]alpha-BGTX) and [3H]imidacloprid ([3H]IMI) as radioligands. The activity of some selected compounds measured with [3H]IMI was 10(4)-fold higher than that measured with [125I]alpha-BGTX. With [3H]IMI as a radioligand, dinotefuran was 13-fold less active than imidacloprid. The inhibitory effect of dinotefuran on the binding of [3H]IMI to the membrane preparation was in a competitive manner. Quantitative analysis of the insecticidal activity of the test compounds with the binding activity measured with [3H]IMI showed that the higher the binding activity, the higher was the insecticidal activity.     

Actions of quinolizidine alkaloids on Periplaneta americana nicotinic acetylcholine receptors

BACKGROUND: Botanical insecticides do not play a major role as crop protectants, but they are beneficial in some applications. The authors investigated the actions of naturally occurring alkaloids on insect nicotinic acetylcholine (ACh) receptors (nAChRs) by evaluating their abilities to inhibit specific binding of [³H]imidacloprid (IMI) to nerve‐cord membranes from Periplaneta americana L. Two alkaloids were also tested for their actions on nAChRs expressed by cockroach neurons using patch‐clamp electrophysiology. RESULTS: Four natural quinolizidine alkaloids (matrine, sophocarpine, cytisine and aloperine) exhibited more than 50% inhibition of [³H]IMI binding at 10 µM , although other compounds were found to have no or low inhibitory activity. The rank order of potency based on concentration–inhibition curves was cytisine > sophocarpine ≥ aloperine ≥ matrine. Patch‐clamp analysis indicated that sophocarpine and aloperine were not agonists of nAChRs expressed in P. americana neurons, yet, at 10 µM , aloperine, but not sophocarpine, suppressed ACh‐induced inward currents significantly. CONCLUSION: Three of the four natural alkaloids tested possess structural moieties that are necessary for interaction with P. americana nAChRs. Aloperine, which possesses a unique structure and showed a distinctive dose–response curve, was found to act as an antagonist. Appropriate modifications of these alkaloids might result in novel insecticidal nAChR ligands. Copyright © 2008 Society of Chemical Industry     

Toxicity and nicotinic acetylcholine receptor interaction of imidacloprid and its metabolites in Apis mellifera (Hymenoptera: Apidae)

Acute oral and contact toxicity tests of imidacloprid, an insecticide acting agonistically on nicotinic acetylcholine receptors (nAChR), to adult honeybees, Apis mellifera L var carnica, were carried out by seven different European research facilities. Results indicated that the 48-h oral LD50 of imidacloprid is between 41 and > 81 ng per bee, and the contact LD50 between 49 and 102 ng per bee. The ingested amount of imidacloprid-containing sucrose solution decreased with increasing imidacloprid concentrations and may be attributed to dose-related sub-lethal intoxication symptoms or to antifeedant responses. Some previously reported imidacloprid metabolites occurring at low levels in planta after seed dressing, i.e. olefine-, 5-OH- and 4,5-OH-imidacloprid, showed lower oral LD50 values (> 36, > 49 and 159 ng per bee, respectively) compared with the concurrently tested parent molecule (41 ng per bee). The urea metabolite and 6-chloronicotinic acid (6-CNA) exhibited LD50 values of > 99,500 and > 121,500 ng per bee, respectively. The pharmacological profile of the [3H]imidacloprid binding site in honeybee head membrane preparations is consistent with that anticipated for a nAChR. IC50 values for the displacement of [3H]imidacloprid by several metabolites such as olefine, 5-OH-, 4,5-OH-imidacloprid, urea and 6-CNA were 0.45, 24, 6600, > 100,000, and > 100,000 nM, respectively. Displacement of [3H]imidacloprid by imidacloprid revealed an IC50 value of 2.9 nM, thus correlating well with the observed acute oral toxicity of the compounds in honeybees. Neurons isolated from the antennal lobe of A mellifera and subjected to whole-cell voltage clamp electrophysiology responded to the application of 100 microM acetylcholine with a fast inward current of between 30 and 1600 pA at -70 mV clamp potential. Imidacloprid and two of the metabolites (olefine- and 5-OH-imidacloprid) acted agonistically on these neurons, whereas the others did not induce currents at test concentrations up to 3 mM. The electrophysiological data revealed Hill coefficients of approximately 1, indicating a single binding site responsible for an activation of the receptor and no direct cooperativity or allosteric interaction with a second binding site.     

Structural effects of dinotefuran and analogues in insecticidal and neural activities

The insecticidal potencies of dinotefuran and analogues against the adult male American cockroach, Periplaneta americana (L) were measured by injection with or without metabolic inhibitors. The potency of dinotefuran was close to those of clothianidin and imidacloprid under the conditions used. The nerve-excitatory and nerve-blocking activities were measured with central nerve cords of P americana. The nerve-excitatory activity of dinotefuran was lower than that of imidacloprid, but was comparable with that of clothianidin. The nerve-blocking activity of dinotefuran was comparable with that of imidacloprid and slightly higher than that of clothianidin. Quantitative analyses showed that variations in the insecticidal activity were better correlated with variations in the nerve-blocking activity than with those in the nerve-excitatory activity when the contribution of the hydrophobic factor was allowed for.     

Desnitroimidacloprid and Nicotine Binding Site in Rat Recombinant α4β2 Neuronal Nicotinic Acetylcholine Receptor

Desnitroimidacloprid (desnitro-IMI) is proposed to be a bioactivation product of imidacloprid and to bind at the same site as [³H]nicotine in the nicotinic acetylcholine receptor (nAChR) of mouse brain membranes. The α4β2 nAChR subtype accounts for >90% of the binding sites for nicotine in rat brain. This study further characterizes the binding site for [³]desnitro-IMI and [³H]nicotine in rat recombinant α4β2 nAChR using receptor expressed in Sf9 insect cells so that the assays involved no other receptor subtypes or interference from metabolic activation and detoxification systems. The two radioligands gave the same B_max of 7.5 pmol/mg protein and apparent K_d values of 3.3 nM for nicotine and 8.9 nM for desnitro-IMI by Scatchard analysis at 22°C. However, at 4°C, the observed apparent association rate is slower and dissociation rate is faster for [³H]desnitro-IMI than for [³H]nicotine and due to the rapid rate of dissociation of [³H]desnitro-IMI the K_d calculated from the determined association and dissociation rates more closely approximates 1.0 for both ligands. Eight cholinergic agents and nine nicotinoids are equipotent in displacing [³H]desnitro-IMI and [³H]nicotine, with IC₅₀ values (nM) of 0.5 for epibatidine, 1 for cytisine, 4–6 for nicotine and desnitro-IMI, 15 for acetylcholine, and 155 for imidacloprid, with an overall correlation for inhibitor potencies of _r² = 0.99 (n = 17). This correlation of binding site properties extends to [³H]nicotine in the recombinant α4β2 receptor and rat brain membranes (r² = 0.99, n = 12). Thus, desnitro-IMI and nicotine bind with high affinity to the same site in rat recombinant α4β2 neuronal nAChR. This recombinant receptor can be generated in sufficient quantities for high-throughput target site screening and structural analysis of the binding site.     

含稠杂环结构的螺环丁烯内酯类化合物的合成及杀菌活性

为了发现更高杀菌活性的螺环丁烯内酯类化合物并分析该类化合物的构效关系，设计并合成了一系列未见文献报道的含咪唑并噻唑、咪唑并噻嗪和咪唑并噻嗪酮等稠杂环结构的螺环丁烯内酯类化合物，其结构通过核磁共振氢谱 (1H NMR)、碳谱 (13C NMR)及高分辨质谱 (HRMS)确证。离体杀菌活性测试结果表明，化合物 5f 和 6f 对油菜菌核病菌的EC₅₀值分别为33.2和29.8 mg/L，优于对照药剂咪唑菌酮(46.8 mg/L)，化合物 7b 和 7e 对辣椒疫霉的EC₅₀值分别为45.8和43.5 mg/L，优于咪唑菌酮(50.7 mg/L)，与先导化合物相比，其杀菌活性高于2-甲硫基衍生物，低于2-芳氨基衍生物，表明稠杂环的引入可以提高化合物的杀菌活性，而结构中的NH片段对杀菌活性具有关键作用。     

Effects of monoterpenoid insecticides on [H]-TBOB binding in house fly GABA receptor and Cl uptake in American cockroach ventral nerve cord

Monoterpenoids and their derivatives from plant essential oils showed good insecticidal activities in previous studies, but the mechanisms of their action as natural insecticides are not known yet. In the present work, we evaluated the pharmacological action of five monoterpenoids (α-terpineol, carvacrol, linalool, pulegone, and thymol) on native insect GABA receptors from house flies and American cockroaches using radiotracer methods. In the [³H]-TBOB binding assay, carvacrol, pulegone, and thymol all enhanced the [³H]-TBOB binding to membrane preparation of house fly heads with EC₅₀ values of 48 μM, 432 μM, and 6 mM, respectively. Moreover, these three monoterpenoids at concentrations of 500 μM and 1 mM also significantly increased the ³⁶Cl⁻ uptake induced by GABA in membrane microsacs prepared from American cockroach ventral nerve cords. These results revealed that carvacrol, pulegone, and thymol are all positive allosteric modulators at insect GABA receptors. The other two monoterpenoids that were tested, α-terpineol and linalool, showed little or no effect in both the [³H]-TBOB binding and ³⁶Cl⁻ uptake assays.     

Bicyclophosphorothionate antagonists exhibiting selectivity for housefly GABA receptors

2,6,7-Trioxa-1-phosphabicyclo[2.2.2]octane 1-sulfides (bicyclophosphorothionates) with various C_1–4 alkyl groups at the 3- and 4-positions were synthesized and tested for their ability to compete with [³H]4′-ethynyl-4-n-propylbicycloorthobenzoate (EBOB), a non-competitive antagonist of γ-aminobutyric acid (GABA) receptors, for specific binding to rat-brain and housefly-head membranes, and for their insecticidal activity against houseflies. Among the 3,4-substituted analogues, 20 compounds were selectively active for housefly GABA receptors versus rat GABA receptors. The 3-alkyl groups of C₃ length and the 4-alkyl groups of C₄ length were tolerated in housefly receptors, whereas such bulky substituents were deleterious in rat receptors. The 4-isobutyl-3-isopropyl analogue was the most potent in housefly receptors (IC₅₀ = 45.2 nM ), and tert-butylbicyclophosphorothionate (TBPS), with the 4-tert-butyl group and no 3-substituent, was the most potent in rat receptors (IC₅₀ = 62.2 nM ). Their receptor selectivities (rat IC₅₀/housefly IC₅₀) were 52 and 0.038, respectively. The insecticidal activity (LD₅₀) of 20 active analogues was well correlated with their potency (IC₅₀) in inhibiting [³H]EBOB binding to housefly-head membranes (r = 0.93). The results obtained in the present study indicate that the introduction of appropriate alkyl groups into the 3- and 4-positions of bicyclophosphorothionate leads to non-competitive antagonists with increased affinity and selectivity for housefly ionotropic GABA receptors versus rat GABA_A receptors. © 1999 Society of Chemical Industry     

香豆素型呋虫胺光控化合物的设计、合成、光学特性及杀虫活性

为了提高新烟碱类杀虫剂呋虫胺的利用率,基于光控释放技术,将呋虫胺(dinotefuran,以下简称DIN)与香豆素类光敏保护基团7-二乙基氨基香豆素(7-diethylamino coumarin,简称COU)相连,设计合成了以COU为"笼"的呋虫胺(coumarin-caged dinotefuran,简称COU-D...     

House fly head GABA-gated chloride channel: [3 H] α-Endosulfan binding in relation to polychlorocycloalkane insecticide action

This study attempts to use [³H] α-endosulfan to examine directly the binding site(s) for cyclodienes, lindane and toxaphene (collectively referred to as the polychlorocycloalkane or PCCA insecticides) in the 4-aminobutyric acid (GABA)-gated chloride channel. [³H] α-Endosulfan was prepared by reduction of hexachloronorbornenedicarboxylic anhydride with sodium borotritide, then coupling the labelled alcohol with thionyl chloride followed by HPLC purification (35 Ci mmol^?1, > 99% radiochemical purity). This new candidate radioligand readily partitions into lipid membranes and undergoes indiscriminate adsorption to surfaces resulting in high levels of non-specific binding. This makes it very difficult to differentiate the small portion of specific binding at the site relevant to toxic action. This problem was partially circumvented by incubating [³H] α-endosulfan (0.1 nM) with house-fly head membranes (0.2 mg protein) for 70 min at 22°C giving 23 (±4)% specific binding (40 fmol mg^?1 protein) determined as the difference between the radioligand alone and on preincubation for 15 min with unlabelled α-endosulfan (final concentration 100 nM). This procedure is not appropriate for determination of saturation isotherms and standard binding kinetics. However, the effectiveness of 16 PCCAs (also at 100 nM final concentration) in blocking the specific binding of [³H] α-endosulfan is generally consistent with their relative potencies as inhibitors of 4-[³H] propyl-1-(4-ethynylphenyl)-2,6,7-trioxabicyclo[2.2.2] octane ([³H]EBOB) binding suggesting that the binding site for both [³H]α-endosulfan and the PCCAs is part of the GABA-gated chloride channel. Insecticidal channel blockers of other types (e.g. picrotoxinin, trioxabicyclooctanes, a dithiane, and phenylpyrazoles) and GABA are poor inhibitors of [³H] α-endosulfan binding relative to their potencies as inhibitors of [³H] EBOB binding. It therefore appears that the PCCAs compete directly for the [³H] α-endosulfan site, whereas the other channel blockers bind with different inhibition kinetics or at a site more closely coupled to the EBOB than the α-endosulfan binding domain.     

Sulfoxaflor and the sulfoximine insecticides: Chemistry,mode of action and basis for efficacy on resistant insects

The sulfoximines, as exemplified by sulfoxaflor ([N-[methyloxido[1-[6-(trifluoromethyl)-3-pyridinyl]ethyl]-λ⁴-sulfanylidene] cyanamide] represent a new class of insecticides. Sulfoxaflor exhibits a high degree of efficacy against a wide range of sap-feeding insects, including those resistant to neonicotinoids and other insecticides. Sulfoxaflor is an agonist at insect nicotinic acetylcholine receptors (nAChRs) and functions in a manner distinct from other insecticides acting at nAChRs. The sulfoximines also exhibit structure activity relationships (SAR) that are different from other nAChR agonists such as the neonicotinoids. This review summarizes the sulfoximine SAR, mode of action and the biochemistry underlying the observed efficacy on resistant insect pests, with a particular focus on sulfoxaflor.     

Characterisation of the relationship between binding sites for imidacloprid and other nicotinic ligands in insects

Radioligand binding studies using the neurotoxins α-bungarotoxin, epibatidine, imidacloprid (IMI) and methyllycaconitine reveal heterogeneity at the level of the nicotinic acetylcholine receptor (nAChR) in membranes from the peach potato aphid Myzus persicae (Sulzer) and further suggest the presence of more than one ligand binding site per nAChR. These sites are able to interact allosterically with each other. Of particular interest, [³H]IMI has over an order of magnitude higher affinity in membranes of hemipteran pest species than in non-hemipteran insects, which may help explain why IMI is particularly effective for the control of sucking pests.     

Actions of azadirachtin,a plant allelochemical,against insects

Investigations of the antifeedant mode of action of azadirachtin and four synthetic analogues, 22,23-dihydroazadirachtin, 3-tigloylazadirachtol, 11-methoxydihydroazadirachtin and 22,23-bromoethoxydihydroazadirachtin have revealed that both polyphagous and oligophagous insects are behaviourally responsive to azadirachtin, with the most responsive species being able to differentiate extremely small changes in the parent molecule. In Lepidoptera the antifeedant response is correlated also with increased neural activity of the chemoreceptors. When locusts are treated on crop plants, the antifeedant and physiological actions of azadirachtin and analogues work in concert and result in feeding deterrence, growth and moulting aberrations and mortality with the same order of potency as for antifeedancy. Specific binding studies using [³H]dihydroazadirachtin carried out on locust testes and Spodoptera Sf9 cells have shown that the competitive binding of the different analogues of azadirachtin to these binding sites occurs in a similar order of potency to that found with antifeedant and IGR bioassays. This suggests a causal link between specific binding to membrane proteins and the ability of the molecule to exert biological effects. © 1998 Society of Chemical Industry     

Pyrethroid action on the nicotinic acetylcholine receptor/channel

The interactions of natural pyrethrins and nine pyrethroids with the nicotinic acetylcholine (ACh) receptor/channel complex of Torpedo electric organ membranes were studied. None caused significant reduction in [³H]ACh binding to the receptor sites, but all inhibited [³H]perhydrohistrionicotoxin ([³H]H₁₂-HTX) binding to the channel sites in presence of carbamylcholine. Allethrin inhibited [³H]H₁₂-HTX binding noncompetitively, but [³H]imipramine binding competitively, suggesting that allethrin binds to the receptor's channel sites that bind imipramine. The pyrethroids were divided into two types according to their actions: type I, which included pyrethrins, allethrin, bioallethrin, resmethrin, and tetramethrin, was more potent in inhibiting [³H]H₁₂-HTX binding and acted more rapidly (i.e., in <30 sec). Type II, which included permethrin, fluvalinate, cypermethrin and fenvalerate, was less potent and their potency increased slowly with time. Also, inhibition of the initial rate of [³H]H₁₂-HTX binding by type I compounds increased greatly by the presence of the agonist carbamylcholine, but this was not so with type II compounds. The receptor-regulated ⁴⁵Ca²⁺ flux into Torpedo microsacs was inhibited by pyrethrins and pyrethroids, suggesting that their action on this receptor function is inhibitory. There was very poor correlation between the potencies of pyrethrins and pyrethroids in inhibiting [³H]H₁₂-HTX binding and their toxicities to house flies, mosquitoes, and the American cockroach. However, the high affinities that several pyrethroids have for this nicotinic ACh receptor suggest that pyrethroids may have a synaptic site of action in addition to their well known effects on the axonal channels.     

Investigating nicotinic acetylcholine receptor expression in neonicotinoid resistant Myzus persicae FRC

The peach–potato aphid Myzus persicae is a pest of many commercial crops due to its polyphagous nature of feeding and has a well-documented history of acquiring resistance to insecticides. In 2009 a strain (M. persicae FRC) emerged in southern France with a point mutation (R81T) at the nicotinic acetylcholine receptor (nAChR), the target site for neonicotinoids such as imidacloprid. This point mutation was associated with the loss of the high affinity imidacloprid binding site (pM Kd), with the single remaining binding site (low nM Kd) highly overexpressed compared to laboratory controls (Bass et al., 2011 [1]). Here we report that after 2 years of continuous selection in the glass house environment with neonicotinoids, the total level of IMD-sensitive nAChRs (low nM Kd) in M. persicae FRC is now comparable to laboratory controls (pM and low nM Kd). Interestingly, despite this large reduction in IMD-sensitive nAChRs, this was not associated with any significant alteration in NNIC-lethality. Additionally, sustained absence of neonicotinoid-selection did not alter nAChR protein levels. We suggest that alterations in nAChR protein expression level described in the original characterisation of the field-isolated M. persicae FRC is unlikely to have been a direct consequence of the R81T mutation. Rather, we speculate that nAChR expression in aphids is likely influenced by as yet unknown conditions in the natural field environment that are absent in the laboratory setting.     

Thiamethoxam is a neonicotinoid precursor converted to clothianidin in insects and plants

Neonicotinoid insecticides are compounds acting agonistically on insect nicotinic acetylcholine receptors (nAChR). They are especially active on hemipteran pest species such as aphids, whiteflies, and planthoppers, but also commercialized to control many coleopteran and some lepidopteran pest species. The most prominent member of this class of insecticides is imidacloprid. All neonicotinoid insecticides bind with high affinity (I₅₀-values around 1 nM) to [³H]imidacloprid binding sites on insect nAChRs. One notable ommission is the neonicotinoid thiamethoxam, showing binding affinities up to 10,000-fold less potent than the others, using housefly head membrane preparations. Electrophysiological whole cell voltage clamp studies using neurons isolated from Heliothis virescens ventral nerve cord showed no response to thiamethoxam when applied at concentrations of 0.3 mM, although the symptomology of poisoning in orally and topically treated noctuid larvae suggested strong neurotoxicity. Other neonicotinoids, such as clothianidin, exhibited high activity as agonists on isolated neurons at concentrations as low as 30 nM. There was no obvious correlation between biological efficacy of thiamethoxam against aphids and lepidopterans and receptor affinity in electrophysiological and binding assays. Pharmacokinetic studies using an LC-MS/MS approach to analyze haemolymph samples taken from lepidopteran larvae revealed that thiamethoxam orally applied to 5th instar Spodoptera frugiperda larvae was rapidly metabolized to clothianidin, an open-chain neonicotinoid. Clothianidin shows high affinity to nAChRs in both binding assays and whole cell voltage clamp studies. When applied to cotton plants, thiamethoxam was also quickly metabolized, with clothianidin being the predominant neonicotinoid in planta briefly after application, as indicated by LC-MS/MS analyses. Interestingly, the N-desmethylated derivative of thiamethoxam, N-desmethyl thiamethoxam, was not significantly produced in either lepidopteran larvae or in cotton plants, although it was often mentioned as a possible metabolite, being nearly as active as imidacloprid. In conclusion, our investigations show that thiamethoxam is likely to be a neonicotinoid precursor for clothianidin.     

Structural modifications increase the insecticidal activity of ryanodine

The toxicity of ryanodine ( 1 ) and 9,21-didehydroryanodine ( 2 ) (the principal active ingredients of the botanical insecticide ryania) to adult female house flies (Musca domestica L.) is attributable to binding to the ryanodine receptor (ryr) and thereby disrupting the Ca²⁺-release channel. These ryanoids, assayed in house flies with piperonyl butoxide (PBO) to suppress cytochrome P450-dependent detoxification, give injected KD₅₀ values of 0·07–0·11 μg g^-1, injected LD₅₀ values of 0·39–0·45 μg g^-1 and topical LD₅₀ values of 12– 50 μg g^-1. They inhibit the [³H]ryanodine binding site of house fly and rabbit muscle with IC₅₀ values of 3–10 nM . This study examines the effect of structure on potency, with 15 variants of the cyclohexane substituents, two 4,6-cyclic boron and two methylated derivatives, and four modifications of the isopropyl and ester substituents. The most effective compound examined was 10-deoxy- 2 ( 3 ) which was more potent than 2 by 2–4-fold on injection and 29-fold applied topically following PBO (LD₅₀ 0·41 μg g^-1). Additional high-potency compounds were 10-oxo- 1 and the cyclohexane variants with lactam, 21-nor-9-oxo and 21-nor-10-deoxy substituents. Other modifications usually reduced toxicity. The injected knockdown potency of the ester ryanoids was generally related to their effectiveness in competing with [³H]ryanodine at the ryr of rabbit skeletal muscle. Two non-ester ryanoids, ryanodol and 9,21-didehydroryanodol, were found to be more toxic than predicted from their potency at the ryr and may therefore act in a different manner such as at a K⁺ channel, as suggested by Usherwood and Vais. Clearly ryanoids are challenging prototypes for a potential new generation of insecticides. © 1997 SCI.     

Cross-Resistance to Imidacloprid in Strains of German Cockroach (Blattella germanica) and House Fly (Musca domestica)

The toxicity of a promising new insecticide, imidacloprid, was evaluated against several susceptible and resistant strains of German cockroach and house fly. Imidacloprid rapidly immobilized German cockroaches followed by a period of about 72 h during which some cockroaches recovered. After 72 h there was no further recovery. Imidacloprid-treated houseflies were immobilized more slowly than treated cockroaches, with the maximum effect observed after 72 h, and there was no recovery. Based upon 72-h LD₅₀ values imidacloprid was moderately toxic to German cockroaches (LD₅₀ values were 6–8 ng mg^-1) and had only low toxicity to house flies (LD₅₀ 140 ng mg^-1). Piperonyl butoxide (PBO) blocked the observed recovery in German cockroaches. PBO also greatly enhanced the 72-h LD₅₀ of imidacloprid from 43- to 59-fold in cockroaches and 86-fold in house flies. Two strains of German cockroach (Baygon-R and Pyr-R) showed >4-fold cross-resistance to imidacloprid. This cross-resistance could not be suppressed by PBO, suggesting that P450 monooxygenase-mediated detoxication is not responsible for this cross-resistance. Variation in the level of synergism observed with PBO (between strains) suggests the ‘basal’ level of monooxygenase-mediated detoxication of imidacloprid is quite variable between strains of German cockroach. The AVER and LPR strains of house fly showed significant cross-resistance to imidacloprid. PBO reduced the level of cross-resistance in AVER from >4·2-fold to 0·5-fold (i.e. the AVER strain LD₅₀ was half that of the susceptible strain when both were treated with PBO), but PBO did not suppress the cross-resistance in LPR. These data suggest monooxygenases are the mechanism responsible for cross-resistance to imidacloprid in AVER, but not in the LPR strain. © of SCI.