Chlorantraniliprobe (Chlo), a potent insecticide, demolishes intracellular Ca2+ homeostasis of insects by inducing uncontrolled Ca2+ release through ryanodine receptors (RyRs). Chlo is lethal to insects but has low toxicity to mammals. In this study, we investigated the effects of Chlo on RyR1 from mammalian skeletal muscle. Ca2+ release assay indicated that Chlo at high concentrations promoted Ca2+ release from sarcoplasmic reticulum through RyR1 channels. Single channel recording of purified RyR1 showed that Chlo activated RyR1 channel, increased channel open probability P-o, reduced channel mean close time T-c, but did not change the channel mean open time T-o, suggesting that Chlo destabilized the closed RyR1 channel, rendered the channel easy to open. The dissociation constant K-d values of Chlo for RyR1 were of micromolar level, approximately 100-fold larger than that for insect RyR. The K-d values were smaller for open states than for closed/blocked states of the RyR1 channel. The maximal binding capacity B-max did not change in the presence of either channel activators or inhibitors/blockers. Our results demonstrate that the insecticide Chlo is a weak activator of mammalian RyR1. It can interact with mammalian RyR1 and activate RyR1 channel but with much lower affinity compared with insect RyR; Chlo has a binding site distinct from all known RyR channel modulators and represents a novel type of RyR channel modulator. Our data provide biochemical and pharmacological insights into its high specificity to insect RyR and high selectivity of poisoning to insects over mammals. (C) 2018 Elsevier Inc. All rights reserved.