Cytochrome P450 inhibition impedes pyrethroid effects on insects through Nav channel regulation

Insecticides used in various formulations are inevitable in agricultural pest control and prevention of vector-borne diseases. Piperonyl butoxide (PBO) is a synergist widely used to enhance the effectiveness of insecticides, notably pyrethroids, by inhibiting the detoxifying cytochrome P450 enzymes, thus reducing the capacity of insects to metabolize and resist insecticides. Recent studies, however, reveal an unexpectedly weak restoration of pyrethroid efficacy by PBO, but the underlying mechanism is unknown. Here, we demonstrate that the PBO-induced inhibition of cytochrome P450 impedes the effect of deltamethrin by mainly affecting its interaction with the inactivated state of voltage-gated sodium channels (Nav). We describe a new octopamine-dependent regulatory mechanism involving Gs, PKA, DARPP-32, and PP1-2A, which affect the cytochrome P450 conformation, thus limiting the effect of PBO and modulating the Nav gating. As a result, deltamethrin cannot reach its final binding site in the fenestration to exert its full effect. We confirmed in vivo that under chemical stressor, the level of octopamine is elevated, which decreases the deltamethrin efficacy. Our findings reveal a novel adaptation mechanism that increases insect survival by reducing insecticide efficacy, thus emphasizing the necessity of developing more effective formulations and technologies for pest and vector control.