[논문]피레스로이드계 살충제 퍼메트린이 Heliothis virescens 중추신경세포에 있는 나트륨채널에 작용하는 기작을 전기생리학적으로 연구

이대우; 마이클 아담스

doi:10.5656/ksae.2022.02.1.066

피레스로이드계 살충제 퍼메트린이 Heliothis virescens 중추신경세포에 있는 나트륨채널에 작용하는 기작을 전기생리학적으로 연구
Modification of Insect Sodium Currents by a Pyrethroid Permethrin and Positive Cooperativity with Scorpion Toxins 원문보기

한국응용곤충학회지 = Korean journal of applied entomology, v.61 no.1, 2022년, pp.117 - 128

이대우 (오하이오 대학교, 생물학과, 신경과학프로그램) , 마이클 아담스 (캘리포니아 주립대학교, 곤충 및 세포신경생물학과)

초록
AI-Helper

본 연구는 피레스로이드계 살충제인 퍼메트린이 Heliothis virescens의 중추신경세포의 나트륨채널에 어떻게 작용하는 가를 전기생리학적으로 관찰하였다. 퍼메트린은 나트륨채널의 꼬리전류(I_Na-tail)를 지속적으로 증가시켰으며 이러한 비정상적인 나트륨 전류증가가 나방류의 신경계에 과도한 흥분을 일으겨 살충작용을 하는 것으로 생각된다. 이러한 살충작용은 전갈독과 함께 사용했을때 약 8배의 증가가 있었음을 확인하였다. 전갈독이 살충제의 독성을 강화하는 분자생리학적 기전연구가 계속되면 해충방제에 많은 기여를 할 것으로 생각된다.

Abstract ▼ AI-Helper

In this study, we have examined pyrethroid actions on sodium channels in the pest insect Heliothis virescens. The synthetic pyrethroid permethrin increased steady-state sodium current in H. virescens central neurons and prolonged tail currents (INa-tail) due to extreme slowing of sodium channel deactivation. Prolongation of INa-tail was evident at permethrin concentrations as low as 60 nM, which modified ~1.7% of sodium channels and 10 μM permethrin modified about 30% of channels. The average time constant (τ1) of tail current decay was ~335 ms for permethrin-modified channels. These modified channels activated at more negative potentials and showed slower activation kinetics, and failed to inactivate. Permethrin modification of sodium channels was dramatically potentiated by the α scorpion toxin LqhαIT, showing positive cooperativity between two binding sites. The amplitude of the tail current induced by 0.3 μM permethrin was enhanced ~8-fold by LqhαIT (200 pM). Positive cooperativity was also observed between permethrin and the insect-specific scorpion toxin AaIT as 10 nM permethrin potentiated the shift of voltage dependence caused by AaIT (~2-fold).

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표/그림 (9)

그림 Fig. 1. Modification of sodium channels in Heliothis virescens central neurons by permethrin. (A) 1 μM permethrin increased steady-state current during the test pulse and dramatically slowed deactivation, producing a prolonged tail current. (B) All current except for capacitive transients was completely abolished following application of 100 nM tetrodotoxin (TTX). (C) Extreme prolongation of sodium tail current (I_Na-tail) in the presence of 1 μM permethrin. Double-exponential fit yielded a time constant (τ₁) of ~162 ms. The holding potential (V_H) and test potential (V_T) were -18 mV and -108 mV respectively. Each recording shown here was from a different neuron.
그림 Fig. 2. Concentration-dependent elevation of sodium tail current by permethrin. (A) Example of prolonged tail currents caused by sequential applications of 0.6, 1, and 3 μM permethrin. Depolarizing pulses (-18 mV) were applied with 10 second intervals and the holding potential (V_H) was -108 mV. Each application of progressively higher permethrin concentrations produced rapid increase of tail currents. (B) A plot showing relationship between tail current amplitude and time lapse after application of three successive concentrations. (C) Percentage of modified channels plotted as a function of permethrin concentration. Closed circle represents the mean of four individual experiments. Each open symbol indicates %M (see text for details) from one experiment. (Inset) Superimposed sodium currents recorded 3 min after application of various permethrin concentrations as indicated in the right.
그림 Fig. 3. Voltage-dependent activation of sodium channels before and after exposure to 1 μM permethrin. (A) Typical I/V curves before and after exposure to permethrin. (B) Sodium conductance- potential (gNa-V) curves plotted using an equation gNa = I_Na / (V_T - E_rev), where I_Na is a peak sodium current evoked by the test potential (V_T), and E_rev is a reversal potential of sodium channels. Voltage-dependent activation of channels was expressed by normalized sodium conductance and plotted as a function of test potentials. Each point is an average of 5 experiments and Bars are SEMs in (B).
그림 Fig. 4. Permethrin (1 μM) effects on fast rise and decay of H. virescens sodium currents. (A) Peak sodium current was reduced in the presence of permethrin, while steady-state current was increased. (B) Normalized sodium currents before and after 1 μM permethrin exposure showing that fast rise and decay were not modified by permethrin. Fast decay was fitted with double-exponentials. (C) Time to peak was measured as a latency between peak and onset of depolarizing step, and plotted as a function of test potentials. (D) Fast decay was measured as a tau (τ₁) determined from a double-exponential fit shown in (B). τ₁ was plotted as a function of test potentials. Bars are SEMs.
그림 Fig. 5. Voltage-dependent activation of permethrin-modified sodium channels. Superimposed sodium currents evoked by V_T from -63 to -28 mV with 5 mV increment before (A) and after (B) 1 μM permethrin exposure. Sodium currents in the late phase were taken at the point indicated by upward arrows (Late I_Na at 50 ms from the onset of test potential) before (A) and after (B) app- lication of permethrin and then, plotted as a function of test potential (C). Peak sodium currents (I_Na) were also plotted in (C) for comparison. Voltage dependence of the peak I_Na was not modified by permethrin (see Fig. 3) while peak I_Na was decreased. However, Late I_Na-voltage curve shifted to more negative potentials. There was little or no Late I_Na in control. Conductance-voltage curves (D) clearly show that voltage-dependent activation of permethrin-modified sodium channels was shifted to more negative potentials.
그림 Fig.6. Delayed activation of permethrin-modified sodium channels. (A) At a low membrane potential (-48 mV), permethrin modified sodium channels activated very slowly, while unmodified channels were not opened (control). (B) At a more depolarized potential (-18 mV), activation of modified channels is much faster compared to that at -48 mV, but it is still much delayed compared to that of unmodified channels. Because early part of current is masked by large unmodified currents, a sodiumcurrent recorded after permethrin exposure was subtracted from control current, yielding slowly increasing sodium current (Subtracted I_Na). 1 μM permethrin was applied in this experiment.
그림 Fig. 7. Potentiation of permethrin action by 200 pM LqhαIT. (A) In the presence of 200 pM LqhαIT, permethrin action was dramatically potentiated. Numbers in the right are concentrations of permethrin applied. I_Na-tail was measured at the time point indicated by asterisk. (B) Concentration-response curve in the absence and presence of 200 pM LqhαIT. I_Na-tail was translated into % modified channels (%M). % modified channels were plotted as a function of permethrin concentration. LqhαIT dramatically enhanced permethrin action in terms of increased %M. Each point in the presence of 200 pM LqhαIT is an average of 3 experiments.
그림 Fig. 8. Potentiation of AaIT action by permethrin. (A) Superimposed sodium currents evoked by V_T of -43 mV (refer to a down-arrow in D) before and after exposure to the toxins (AaIT or AaIT + permethrin). Applied at 100 nM AaIT, peak and steady-state sodium currents were increased. Additional application of 10 nM permethrin doubled the peak current. (B) Normalized currents in the presence of AaIT before and after permethrin exposure. There were no changes in rising and decaying phases of sodium currents, indicating permethrin does not directly alter actions of AaIT on the channels. I/V (C) and sodium conductance (D) curves in the absence and presence of AaIT, which caused the shift of I/V curve to negative potentials. 10 nM permethrin shifts the curve further to negative direction.
표 Table 1. Comparison of pyrethroid selectivity on insect versus mammalian sodium channels

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