主要农业害虫对茚虫威的抗性现状及其治理策略

茚虫威属于噁二嗪类杀虫剂,与大多数杀虫剂不同的是其进入害虫体内需要经活化代谢转变成N-去甲氧羰基代谢物（decarbomethoxylated metabolite,DCJW）后不可逆地阻断钠通道,进而发挥杀虫活性。茚虫威由于其作用机制不同于常见的使钠离子通道延迟关闭的菊酯类药剂而被广泛用于鳞翅目和一些同翅目、鞘翅目害虫的防治。抗药性是任何杀虫药剂使用后面临的问题,茚虫威也不例外,许多害虫对其产生了不同程度的抗性。昆虫对茚虫威产生抗性的机制包括酯酶活性、谷胱甘肽S-转移酶（glutathione S-transferase,GST）和P450活性的增加以及分子靶标F1845Y、V1848I、L1014P的突变,这些对茚虫威抗性机制的研究基本都是基于抗性种群和敏感种群开展的,需要进一步验证其对抗性研究的贡献度。针对我国田间害虫种群对茚虫威的抗性现状,及时实施对茚虫威有效的抗性治理是迫切的。对于茚虫威的抗性治理除了传统的杀虫药剂轮用、混用外,需要利用其作用机制特点开展抗性治理策略研究。一是充分利用其活化代谢的特点,开展组合药剂的研究应用;二是菊酯类药剂和茚虫威的作用机制均与钠离子通道有关,但是前者是使钠离子通道关闭延迟,而后者是阻断钠离子通道,开展相关基础研究,使菊酯类药剂与茚虫威合理地用于抗性治理中。本文综述了茚虫威的抗性现状、抗性机制与交互抗性、茚虫威的抗性风险评价,针对茚虫威的抗性特点提出了抗性治理策略。     

Indoxacarb resistance in the house fly, Musca domestica

Indoxacarb (DPX-MP062) is a recently introduced oxadiazine insecticide with activity against a wide range of pests, including house flies. It is metabolically decarbomethoxylated to DCJW. Selection of field collected house flies with indoxacarb produced a New York indoxacarb-resistant (NYINDR) strain with >118-fold resistance after three generations. Resistance in NYINDR could be partially overcome with the P450 inhibitor piperonyl butoxide (PBO), but the synergists diethyl maleate and S,S,S-tributyl phosphorothioate did not alter expression of the resistance, suggesting P450 monooxygenases, but not esterases or glutathione S-transferases are involved in the indoxacarb resistance. Conversely, the NYINDR strain showed only 3.2-fold resistance to DCJW, and this resistance could be suppressed with PBO. Only limited levels of cross-resistance were detected to pyrethroid, organophosphate, carbamate or chlorinated hydrocarbon insecticides in NYINDR. Indoxacarb resistance in the NYINDR strain was inherited primarily as a completely recessive trait. Analysis of the phenotypes vs. mortality data revealed that the major factor for indoxacarb resistance is located on autosome 4 with a minor factor on autosome 3. It appears these genes have not previously been associated with insecticide resistance.     

Effects of pyrethroids on neuronal excitability of adult honeybees Apis mellifera

Pyrethroids act on the nervous system as a primary target organ and exert their neurotoxic effects primarily by altering the conductance of sodium channel, leading to hyperexcitation. However, few studies investigated the effects of pyrethroids on neuronal excitability of honeybee brain neurons. In this study, a whole-cell patch-clamp technique was used to record current threshold, the minimum current to induce an action potential, and peak sodium current in the dissociated honeybee brain neurons treated with bifenthrin, deltamethrin and fluvalinate in vitro & in vivo. The study showed that these pyrethroids greatly suppressed the neuronal excitability as revealed by increasing current injected and inhibited the peak sodium current in honeybees. The three pyrethroids also inhibited steady-state inactivation in addition to reduction of sodium peak current.     

Broad-spectrum insecticide resistance in obliquebanded leafroller Choristoneura rosaceana (Lepidoptera: Tortricidae) from Michigan

Nineteen insecticides, belonging to nine chemical classes, were bioassayed by dietary exposure against two strains of obliquebanded leafroller, Choristoneura rosaceana, collected from Michigan apple orchards. Berrien is a putatively organophosphate-resistant strain from a commercial orchard with a history of insecticide use, and Kalamazoo a susceptible strain from an isolated and unsprayed orchard. The Berrien strain was moderately resistant (about 25-fold) to organophosphates such as azinphos-methyl and chlorpyrifos. Very low resistance (< 10-fold) was also observed to pyrethroids such as cypermethrin, zeta-cypermethrin, bifenthrin, deltamethrin and esfenvalerate, to the ecdysone agonists tebufenozide and methoxyfenozide, and to the chlorinated pyrrole chlorfenapyr. Endosulfan and carbamates such as thiodicarb, methomyl and carbaryl had low intrinsic toxicities against both strains, with little difference in sensitivity between them. There was no resistance to spinosad. Emamectin benzoate was found to be the most toxic insecticide against C rosaceana, with slightly higher lethal doses required for the Berrien strain. Unexpectedly, Berrien exhibited a very high level of resistance (> 700-fold) to indoxacarb, which has never been used in Michigan to control this insect pest. The active metabolite of indoxacarb, DCJW, was considerably more toxic than the parent compound, but the resistance against DCJW was comparable to that seen with indoxacarb. This indicates that a failure to activate indoxacarb was not the mechanism of resistance in Berrien. The low level of resistance to several chemistries recorded in Michigan C rosaceana can be managed at this stage by adopting a rotation of chemistries having different modes of action.     

Voltage‐sensitive sodium channel mutations S989P + V1016G in Aedes aegypti confer variable resistance to pyrethroids,DDT and oxadiazines

BACKGROUND

Aedes aegypti is a vector of several important human pathogens. Control efforts rely primarily on pyrethroid insecticides for adult mosquito control, especially during disease outbreaks. A. aegypti has developed resistance nearly everywhere it occurs and insecticides are used. An important mechanism of resistance is due to mutations in the voltage‐sensitive sodium channel (Vssc) gene. Two mutations, in particular, S989P + V1016G, commonly occur together in parts of Asia.

RESULTS

We have created a strain (KDR:ROCK) that contains the Vssc mutations S989P + V1016G as the only mechanism of pyrethroid resistance within the genetic background of Rockefeller (ROCK), a susceptible lab strain. We created KDR:ROCK by crossing the pyrethroid‐resistant strain Singapore with ROCK followed by four backcrosses with ROCK and Vssc S989P + V1016G genotype selections. We determined the levels of resistance conferred to 17 structurally diverse pyrethroids, the organochloride DDT, and oxadiazines (VSSC blockers) indoxacarb (proinsecticide) and DCJW (the active metabolite of indoxacarb). Levels of resistance to the pyrethroids were variable, ranging from 21‐ to 107‐fold, but no clear pattern between resistance and chemical structure was observed. Resistance is inherited as an incompletely recessive trait. KDR:ROCK had a > 2000‐fold resistance to DDT, 37.5‐fold cross‐resistance to indoxacarb and 13.4‐fold cross‐resistance to DCJW.

CONCLUSION

Etofenprox (and DDT) should be avoided in areas where Vssc mutations S989P + V1016G exist at high frequencies. We found that pyrethroid structure cannot be used to predict the level of resistance conferred by kdr. These results provide useful information for resistance management and for better understanding pyrethroid interactions with VSSC. © 2017 The Authors. Pest Management Science published by John Wiley & Sons Ltd on behalf of Society of Chemical Industry.     

拟环纹豹蛛神经细胞的急性分离及其电压门控钠通道的膜片钳研究

对拟环纹豹蛛雌蛛中枢神经细胞进行急性分离,并利用全细胞膜片钳技术对神经细胞的电压门控钠通道基本电生理学特性进行了研究,同时测定了氯氰菊酯及其不同浓度对Na 通道门控过程的影响。结果表明,拟环纹豹蛛头胸部神经团在消化酶和培养液混合作用下能急性分离出单个神经细胞,在4～6h内适合用于膜片钳研究。在全细胞电压钳条件下,神经细胞Na 通道电流(INa)在-60mV左右激活, 10mV左右达峰值,对河豚毒素敏感。用氯氰菊酯处理后,Na 通道的激活电压向负电位方向移动约10mV,为-70mV左右,峰值提前在 5mV左右出现。INa先增大然后迅速减小。用不同浓度的氯氰菊酯处理神经细胞后,INa在各电压下的差异不大,但达到最大INa所需时间随氯氰菊酯浓度的增加而缩短。     

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.     

Actions of Tefluthrin on Rat Na(v)1.7 Voltage-Gated Sodium Channels Expressed in Xenopus Oocytes

In rats expression of the Na_v1.7 voltage-gated sodium channel isoform is restricted to the peripheral nervous system and is abundant in the sensory neurons of the dorsal root ganglion. We expressed the rat Na_v1.7 sodium channel α subunit together with the rat auxiliary β1 and β2 subunits in Xenopus laevis oocytes and assessed the effects of the pyrethroid insecticide tefluthrin on the expressed currents using the two-electrode voltage clamp method. Tefluthrin at 100 μM modified of Na_v1.7 channels to prolong inactivation of the peak current during a depolarizing pulse, resulting in a marked “late current” at the end of a 40 ms depolarization, and induced a sodium tail current following repolarization. Tefluthrin modification was enhanced up to twofold by the application of a train of up to 100 5 ms depolarizing prepulses. These effects of tefluthrin on Na_v1.7 channels were qualitatively similar to its effects on rat Na_v1.2, Na_v1.3 and Na_v1.6 channels assayed previously under identical conditions. However, Na_v1.7 sodium channels were distinguished by their low sensitivity to modification by tefluthrin, especially compared to Na_v1.3 and Na_v1.6 channels. It is likely that Na_v1.7 channels contribute significantly to the tetrodotoxin-sensitive, pyrethroid-resistant current found in cultured dorsal root ganglion neurons. We aligned the complete amino acid sequences of four pyrethroid-sensitive isoforms (house fly Vssc1; rat Na_v1.3, Na_v1.6 and Na_v1.8) and two pyrethroid-resistant isoforms (rat Na_v1.2 and Na_v1.7) and found only a single site, located in transmembrane segment 6 of homology domain I, at which the amino acid sequence was conserved among all four sensitive isoform sequences but differed in the two resistant isoform sequences. This position, corresponding to Val410 of the house fly Vssc1 sequence, also aligns with sites of multiple amino acid substitutions identified in the sodium channel sequences of pyrethroid-resistant insect populations. These results implicate this single amino acid polymorphism in transmembrane segment 6 of sodium channel homology domain I as a determinant of the differential pyrethroid sensitivity of rat sodium channel isoforms.     

Mode of action of nereistoxin on the neuromuscular transmission in the frog

The effects of nereistoxin on the neuromuscular transmission of the frog have been studied by means of intracellular microelectrode and voltage clamp techniques. Nereistoxin blocked the neuromuscular transmission without causing depolarization of end-plate membranes. A small end-plate potential was elicited upon nerve stimulation. The frequency and amplitude of miniature end-plate potentials decreased after exposure to nereistoxin, and the quantum content of the end-plate potential tended to decrease also. Chemical sensitivity of the end-plate membrane as examined by iontophoretic application of acetylcholine was markedly reduced. Both sodium and potassium components of the end-plate current were suppressed almost equally by application of nereistoxin. It is concluded that inhibition of the mechanisms whereby the end-plate membrane conductances to sodium and potassium ions are increased is the major action of nereistoxin responsible for neuromuscular blockade.     

Lack of cross-resistance to indoxacarb in insecticide-resistant Spodoptera frugiperda (Lepidoptera: Noctuidae) and Plutella xylostella (Lepidoptera: Yponomeutidae)

Two field strains of the fall armyworm, Spodoptera frugiperda (JE Smith), collected from corn in north Florida showed high resistance to carbaryl (626- and 1159-fold) and moderate resistance to parathion-methyl (30- and 39-fold) as compared with a laboratory susceptible strain. A field strain of the diamondback moth, Plutella xylostella (L.) collected from cabbage in north Florida and selected for 20 generations with permethrin showed high resistance to permethrin (987-fold) as compared with a susceptible strain. However, in all instances, no cross-resistance to indoxacarb, a novel oxidiazine insecticide, was observed in these two species. Biochemical studies revealed that, in S. frugiperda, activities of detoxification enzymes (microsomal oxidase, glutathione S-transferase and general esterase) were significantly higher in the field strains than in the susceptible strain, indicating that these detoxification enzymes were not actively involved in the resistance to indoxacarb. The lack of cross-resistance between indoxacarb and permethrin in P. xylostella further supports the notion that the mode of action of these insecticides on the insect sodium channel is different.     

Mechanism of resistance to spinosyn in the tobacco budworm, Heliothis virescens

Topical laboratory selection of tobacco budworm larvae, Heliothis virescens, with technical spinosad for multiple generations resulted in larvae 1068-fold resistant to topical applications of the insecticide and 316.6-fold resistant to insecticide treated diet as compared to the parental strain. The penetration of 2′-O-methyl[¹⁴C]spinosyn A across the cuticle of the susceptible (parental) and selected (resistant) tobacco budworms increased with time 3-12 h after application. A trend of reduced penetration in the resistant strain was found but the differences were not statistically significant. 2′-O-methyl[¹⁴C]spinosyn A when injected into the hemocoel was not metabolized 96 h after treatment in both the susceptible and resistant strain, suggesting that a change in metabolism was not the mechanism of resistance. Electrophysiological studies indicated that dose-dependent spinosyn A-induced currents occurred in neurons from spinosyn resistant and susceptible (adult) tobacco budworms. At both 10 and 100 nM spinosyn A, however, the amplitude of these currents in the resistant insects was significantly smaller than the amplitude of currents observed from neurons from susceptible tobacco budworm adults. This suggests that neurons from resistant insects have decreased sensitivity to spinosyn A. However, the reduced inward currents in the resistant strain may or may not be related to the mode of action of the spinosyns. No statistically significant cross-resistance was noted for the spinosad resistant tobacco budworms for topical applications of permethrin (Pounce^®), profenofos (Curacron^®), emamectin benzoate (Denim^®), or indoxacarb (Steward^®). A statistically significant reduction in susceptibility to acetamiprid (Mospilan^®) in artificial diet as determined from a resistance ratio of 0.482 was found.     

新型高效杀虫剂茚虫威毒理学研究进展

新型噁二嗪类杀虫剂茚虫威对鳞翅目昆虫具有卓越的杀虫活性。神经毒理学研究表明,茚虫威在昆虫体内活化为N 去甲氧羰基代谢物（DCJW）,并作用于失活态钠离子通道,破坏神经冲动传递。近年来对茚虫威的研究表明,昆虫具有对茚虫威产生高水平抗性的风险。     

Effects of alpha- and theta-cypermethrin insecticide on transient outward potassium current in rat hippocampal CA3 neurons

The effects of two cypermethrin insecticides, alpha-cypermethrin and theta-cypermethrin on the transient outward potassium (I_A) channels in freshly dissociated hippocampal CA3 neurons of rat, were studied using whole-cell patch clamp technique. The results indicate that alpha-cypermethrin increased the amplitude of I_A and theta-cypermethrin decreased the amplitude of I_A, and both of them shifted steady activation curve of I_A towards negative potential. Theta-cypermethrin contributed in the inactivation of I_A. Alpha-cypermethrin prolonged the time constants for the recovery from inactivation kinetics of I_A. These results imply that transient outward potassium (I_A) channels are the target of cypermethrin, which may explain the mechanism of toxical effects of cypermethrin on mammalian animals.     

Toxicity,persistence and efficacy of indoxacarb and two other insecticides on Plutella xylostella (Lepidoptera: Plutellidae) immatures in cabbage

Toxicities of indoxacarb on eggs and 5-day-old larvae of diamondback moth, Plutella xylostella L., on cabbage and those of field-aged leaf residues on 5-day-old larvae were determined in the laboratory. The persistence and efficacies of indoxacarb and two other newer insecticides (spinosad and emamectin benzoate) to P. xylostella were tested under field conditions. Results from laboratory bioassays indicate that indoxacarb was highly toxic to P. xylostella larvae through food ingestion, with LC₅₀ and LC₉₀ values of 24.1 and 90.1 mg AI l ^{- 1}, respectively. However, indoxacarb had no significant effects on eggs and larvae through direct contact compared with water control. The toxicity of field-aged leaf residues of indoxacarb (0-, 3-, 5-, 7-, 10-, 14-, 17- and 21-day-old residues) declined slowly and gradually under the field conditions in South Texas. Almost all larvae died on day 5 after feeding on the leaves with 0 - 14-day residue, and the mortalities were as high as 94 and 78% for the 14- and 17-day-old leaf residues. With one application, indoxacarb suppressed P. xylostella larvae below the economic threshold for 14 - 21 days. Two field trials showed that indoxacarb at 0.05 - 0.07 kg AI ha ^{- 1} was effective against P. xylostella, providing marketable cabbage with three applications per season. In addition, indoxacarb was as effective as spinosad, and significantly more effective than emamectin benzoate.     

茚虫威在金银花中的残留动态研究及安全性评价

为了探明茚虫威在金银花中的残留特性和安全性,对茚虫威在金银花中的消解动态及最终残留进行了研究。样品经乙腈提取,超高效液相色谱-串联质谱(UPLC-MS/MS)检测,外标法定量。运用所建立的方法对茚虫威在山东泰安、河北邢台、浙江杭州、湖南邵阳、广东湛江和重庆九龙坡6个试验点的金银花中的残留消解动态和最终残留进行了研究。试验结果表明:茚虫威残留量在金银花中随时间延长而递减,符合一级反应动力学方程,消解速度较快,半衰期为1.5～2.2 d,属易消解型农药。参照中国制定的茶叶中茚虫威的最大残留限量(MRL)5 mg/kg,15%茚虫威悬浮剂按推荐使用剂量90 g/hm~2,施药1次,施药后7 d,所采收的金银花中的茚虫威残留量低于5 mg/kg。研究结果可为茚虫威在金银花上的合理使用及其最大残留限量制定提供参考。     

Toxicity of a formulation of the insecticide indoxacarb to the tarnished plant bug, Lygus lineolaris (Hemiptera: Miridae), and the big-eyed bug, Geocoris punctipes (Hemiptera: Lygaeidae)

Indoxacarb is a new oxadiazine insecticide that has shown outstanding field insecticidal activity. The toxicity of a 145 g litre-1 indoxacarb SC formulation (Steward) was studied on the tarnished plant bug Lygus lineolaris and the big-eyed bug Geocoris punctipes. Both insect species responded very similarly to indoxacarb in topical, tarsal contact and plant feeding toxicity studies. The topical LD50 of the formulation was c 35 ng AI per insect for both species. Prolonged tarsal contact with dry indoxacarb residues did not result in mortality for either insect species. However, both species were susceptible to feeding through dried residues of indoxacarb after spraying on young cotton plants. Feeding on water-washed plants resulted in lower mortality than that observed with unwashed plants, and toxicity declined even more dramatically after a, detergent rinse, indicating that much of the indoxacarb probably resides on the cotton leaf surface or in the waxy cuticle. These results were corroborated by HPLC-mass spectrometry measurements of indoxacarb residues on the plants. Greater mortality for both species was observed in a higher relative humidity environment. Higher levels of accumulated indoxacarb and its active metabolite were detected in dead G punctipes than in L lineolaris after feeding on sprayed, unwashed plants. When female G punctipes ate indoxacarb-treated Heliothis zea eggs, there was significant toxicity. However, only c 15% of the females consumed indoxacarb-treated eggs, and the rest of the females showed a significant diminution of feeding in response to the insecticide. Cotton field studies have shown that indoxacarb treatments at labelled rates lead to a dramatic decline in L lineolaris, with negligible declines in beneficial populations. A major route of intoxication of L lineolaris in indoxacarb-treated cotton fields thus appears to be via oral, and not cuticular, uptake of residues from treated cotton plants. The mechanisms for selectivity/safety for G punctipes are currently under investigation and may be a combination of differential feeding behavior and diminution of feeding by females exposed to indoxacarb-treated eggs.     

A monitoring study to assess the acute mortality effects of indoxacarb on honey bees (Apis mellifera L.) in flowering apple orchards

To evaluate the effect of the indoxacarb 300 g kg(-1) WG, Steward 30WDG, on the honey bee (Apis mellifera L.) in apple orchards, a monitoring study was conducted in Dutch apple orchards in April/May 2004. Before apple flowering began, two honey bee colonies were placed in each orchard to investigate honey bee mortality. Each hive was provided with a Münster dead bee trap to collect dead honey bees. The numbers of dead bees found in these Münster dead traps were counted every 3-4 days for about 2 weeks before and after the period of the insecticide treatment. In nine flowering orchards no indoxacarb was applied during the flowering period, which served as control sites. In 30 flowering orchards indoxacarb was sprayed by the fruit growers according to local practice at 170-260 g formulated product ha(-1) (51-78 g AI ha(-1)). In the control orchards the average mortality was 8 honey bees colony(-1) day(-1). The average daily honey bee mortality before and after indoxacarb application was 8 and 10 honey bees colony(-1) day(-1) respectively. At one test site, indoxacarb was mixed with other plant protection products plus plant nutrients, and in this orchard a slight but biologically non-significant increase in acute honey bee mortality was recorded. It was concluded that the application of indoxacarb caused no effects on honey bee mortality, and that the number of dead honey bees counted in the Münster traps in the orchard treated with indoxacarb was comparable with those determined in control orchards.     

施用茚虫威饵剂后红火蚁的蚁巢迁移及弃尸行为

红火蚁Solenopsis invicta是典型的社会性昆虫, 在防控过程中容易引起红火蚁巢体迁移和弃尸等现象, 影响防控效果。本研究对标记好的蚁巢施用茚虫威饵剂, 然后定时观察新增加的蚁巢数量和工蚁弃尸量。结果表明, 施用0.08%和0.1%茚虫威饵剂后, 所有处理均有新蚁巢出现, 施药后5~10 d, 新蚁巢大量出现, 随后蚁群趋向稳定, 新增蚁巢减少, 至处理后25~30 d, 再次出现新蚁巢。0.1%茚虫威饵剂15 g/巢处理在施药后5 d出现8个新蚁巢, 药后10 d和30 d分别又新增5个蚁巢, 其他时间新增蚁巢数较少。按处理剂量, 药后30 d, 2种药剂15 g/巢处理出现的新巢数量均显著高于25 g/巢处理。0.1%茚虫威15 g/巢处理, 校正蚁巢累计增加率高达124.98%, 0.08%茚虫威饵剂15 g/巢处理为89.05%; 0.08%和0.1%茚虫威饵剂25 g/巢处理, 校正蚁巢累计增加率分别为12.00%和51.34%。施用15 g/巢饵剂后2 d开始有大量工蚁尸体被丢弃, 至药后4 d弃尸量达最大值, 0.08%和0.1%茚虫威饵剂处理弃尸量分别为330.5头/巢和300.2头/巢, 随后呈逐步下降趋势, 至14 d, 弃尸量降至最低。施药后8 d内弃尸量达总弃尸量75.26%以上, 至药后10 d, 已达总弃尸量的90.08%以上。施用0.08%和0.1%茚虫威饵剂25 g/巢的处理新蚁巢出现较少, 表明施用相对低剂量的饵剂, 更容易引起蚁巢迁移。红火蚁的弃尸行为主要发生在饵剂施用后的前8 d。     

Depolarization of motor nerve terminals by pyrethroids in susceptible and kdr-resistant house flies

When applied at concentrations of one nM or higher to a house fly larval neuromuscular preparation, deltamethrin (DM) and fenvalerate (FV) greatly increased miniature excitatory postsynaptic potential (mepsp) rate and blocked neuromuscular transmission. The DM-induced mepsp discharge was abolished by tetrodotoxin (TTX), removal of Ca²⁺ from the saline, or by application of hyperpolarizing stimuli to the nerve, indicating that it was due to depolarization of the presynaptic terminals. Also, in the presence of TTX, K⁺ depolarization increased mepsp rate at the same external K⁺ concentration before and after DM treatment, confirming that DM released transmitter by depolarizing the nerve terminals rather than by altering the voltage dependence of transmitter release. The potassium channel blocker tetraethylammonium (TEA) increased mepsp rate somewhat, while aconitine (20 μM), which keeps sodium channels open, increased mepsp rate consistently. Pretreatment of nerves with a subthreshold dose of TEA greatly increased the mepsp rate-increasing activity of DM and aconitine, while a subthreshold level of aconitine did not synergize DM. These observations suggest that DM, like aconitine, depolarized nerves by modifying the sodium channels. Knockdown resistant (kdr) larvae were resistant to the depolarizing action of DM and aconitine but not to that of TEA, indicating that the kdr gene produced a modified sodium channel which was less sensitive to the action of pyrethroids and aconitine. During sustained transmitter release by DM, evoked release gradually declined, resulting in a condition called early block in which spontaneous release was high and release could be evoked by electrotonic depolarization of the nerve terminals, but not by a nerve action potential. Early block was probably due to conduction block in the nerve terminals. Early block eventually gave way to late block, characterized by the decline of spontaneous release to subnormal levels and complete failure of evoked release. After late block, the calcium ionophore X-537A could not release transmitter, suggesting that late block was due to depletion of available transmitter. DM did not have a direct effect upon extrasynaptic muscle membrane. However, after late block, muscles were left insensitive to the putative transmitters glutamate and aspartate when these were bath or iontophoretically applied. A low rate of mepsps persisted after late block, indicating that the muscles were still sensitive to the natural transmitters.     

Kinetics of sodium channel modification as the basis for the variation in the nerve membrane effects of pyrethroids and DDT analogs

The effects of a wide range of pyrethroids and DDT analogs on the membrane potential and membrane sodium currents were studied in crayfish giant axons. Compounds differed greatly in their ability to produce depolarizing afterpotentials, repetitive firing, and membrane depolarization. The differences observed at the membrane potential level could be explained by differences in the kinetics with which the insecticides interact with the nerve membrane sodium channel. The compounds containing a cyano group at the α position retain sodium channels in a modified open state persistently, depolarize the membrane, and block the action potential without causing repetitive firing. The pyrethroids without an α-cyano group and DDT analogs retain sodium channels in a modified open state only transiently, cause large depolarizing afterpotentials, and evoke repetitive firing with minimal effect on the resting potential. The effects of the phenoxybenzyl pyrethroids were found to be intermediate between these two extremes suggesting that a continuous variation exists in kinetics with which pyrethroids and DDT analogs modify sodium channels. It was not necessary to assume a second site of action to account for the variability observed. The implications of these results to the construction of quantitative structure-activity relationships is discussed.