Molecular Insights into Pharmacological Mechanism of Insect Kir Channels and the Toxicity of Kir Inhibitors on Hemipteran Insects

Inwardly rectifying potassium channels (Kir) play a key role in regulating various physiological processes. However, the structural and pharmacological mechanisms of insect Kir channels remain unclear. In this study, we show that coexpression of different Kir subunits in the same tissue did not affect the rectification properties of strongly rectifying Kir. The Kir inhibitor VU041, along with the insecticide flonicamid and its metabolite flumetnicam, were tested for their inhibitory effects on the homotetrameric Kir1 and Kir2 channels. Both Kir1 and Kir2 channels from the two insect species showed similar pharmacological responses to VU041, flonicamid, and flumetnicam. However, VU041 demonstrated significantly higher inhibitory activity than both insecticides across all four Kir channels, while flumetnicam exhibited the weakest inhibition. Molecular docking analyses indicate that the binding site of VU041 is not the same as that of flonicamid, and flumetnicam. flonicamid, and flumetnicam have binding sites similar to the ATP binding sites in cytoplasmic region of human Kir6.2, whereas VU041 is located in the pore of the ion channel, and serves as a pore blocker that inhibits Kir channels. Mutation analysis confirmed the essential roles of these residues in channel function and binding affinity. Finally, the toxicities of the three inhibitors were evaluated in N. lugens and M. persicae. VU041, a potent inhibitor of the insect Kir channel, showed lower toxicity compared to the other two inhibitors, whereas flumethoxan, which is less active on the Kir1 channel, showed higher toxicity, probably related to the different bioavailability of the different compounds. These findings suggest that the potential of targeting Kir channels as insecticidal strategies requires further evaluation.