The neurotoxic insecticidal mechanism of the nonsteroidal ecdysone agonist RH-5849: K+ channel block in nerve and muscle

RH-5849 (1,2-dibenzoyl-1-tert-butylhydrazine) is the prototype of a novel class of insect growth regulators that induce a lethal premature molt in larval lepidoptera by direct stimulation of ecdysone receptors [K. D. Wing, R. A. Slawecki, and G. R. Carlson, Science 241, 470 (1988); K. D. Wing, Science 241, 467 (1988)]. RH-5849 has also been found to produce, in insects and mammals, acute neurotoxic symptoms that are clearly not due to activation of ecdysone receptors. This paper describes studies leading to a proposed neurotoxic mechanism of RH-5849. The injection of RH-5849 into cockroaches and other insects at 50 microgram/g caused a rapid onset of hyperactivity, leading to prostration and constant movement of all appendages within a few minutes. Paralysis ensued after several hours. RH-5849 at 100 micromolar induced strong contractions in muscles of larval house flies, Musca domestica. Muscle action potentials in response to nerve stimulation or injection of depolarizing current were greatly prolonged. Maintained voltage-activated K+ current (I(K)) in voltage-clamped ventral longitudinal muscles was reduced 70% by 100 micromolar RH-5849, accounting for most of the action potential prolongation and muscular contractions. The transient K+ current (I(A)) and Ca2+-dependent maintained K+ current (I(CS)) were not affected by this treatment. RH-5849 and other diacylhydrazines blocked K+ channels in neurons as well as in muscle. The excitatory postsynaptic current at the house fly larval neuromuscular junction was reversibly enhanced by RH-5849 and another diacylhydrazine, consistent with the block of presynaptic K+ channels. Also, at cercal nerve-giant axon synapses of the cockroach, Periplaneta americana, RH-5849 greatly prolonged excitatory postsynaptic potentials, and after extensive exposure blocked the presynaptic action potential. RH-5849 caused substantial block of (I(K)) at concentrations that would be achieved following injection of doses that cause neurotoxic symptoms, indicating that the K+ channel block is an important neurotoxic mechanism of this class of compounds. Furthermore, a threefold synergism was observed between an analog of RH-5849 and the pyrethroid fenvalerate, when the two compounds were mixed in the ratio of their LD50s, consistent with K+ channel block in vivo. The neurotoxic effects of diacylhydrazines are mediated by a direct action on K+ channels and are not mimicked by beta-ecdysone. The dual actions of diacylhydrazines may help delay or prevent resistance development in certain pest species.